Publications 2013

Turner C, Turner P, Hoogenboom G, Aye Mya Thein N, McGready R, Phakaudom K, De Zoysa A, Efstratiou A, Heath PT, Nosten F. 2013. A three year descriptive study of early onset neonatal sepsis in a refugee population on the Thailand Myanmar border. BMC Infect Dis, 13 (1), pp. 601. | Citations: 3 (Scopus) | Show Abstract | Read more

BACKGROUND: Each year an estimated four million neonates die, the majority in the first week of life. One of the major causes of death is sepsis. Proving the incidence and aetiology of neonatal sepsis is difficult, particularly in resource poor settings where the majority of the deaths occur. METHODS: We conducted a three year observational study of clinically diagnosed early onset (<7 days of age) neonatal sepsis (EONS) in infants born to mothers following antenatal care at the Shoklo Malaria Research Unit clinic in Maela camp for displaced persons on the Thailand-Myanmar border. Episodes of EONS were identified using a clinical case definition. Conventional and molecular microbiological techniques were employed in order to determine underlying aetiology. RESULTS: From April 2009 until April 2012, 187 infants had clinical signs of EONS, giving an incidence rate of 44.8 per 1000 live births (95% CI 38.7-51.5). One blood culture was positive for Escherichia coli, E. coli was detected in the cerebrospinal fluid specimen in this infant, and in an additional two infants, by PCR. Therefore, the incidence of bacteriologically proven EONS was 0.7 per 1000 live births (95% CI 0.1-2.1). No infants enrolled in study died as a direct result of EONS. CONCLUSION: A low incidence of bacteriologically proven EONS was seen in this study, despite a high incidence of clinically diagnosed EONS. The use of molecular diagnostics and nonspecific markers of infection need to be studied in resource poor settings to improve the diagnosis of EONS and rationalise antibiotic use.

Abdullah NR, Barber BE, William T, Norahmad NA, Satsu UR, Muniandy PK, Ismail Z, Grigg MJ, Jelip J, Piera K et al. 2013. Plasmodium vivax population structure and transmission dynamics in Sabah Malaysia. PLoS One, 8 (12), pp. e82553. | Citations: 30 (Web of Science Lite) | Show Abstract | Read more

Despite significant progress in the control of malaria in Malaysia, the complex transmission dynamics of P. vivax continue to challenge national efforts to achieve elimination. To assess the impact of ongoing interventions on P. vivax transmission dynamics in Sabah, we genotyped 9 short tandem repeat markers in a total of 97 isolates (8 recurrences) from across Sabah, with a focus on two districts, Kota Marudu (KM, n = 24) and Kota Kinabalu (KK, n = 21), over a 2 year period. STRUCTURE analysis on the Sabah-wide dataset demonstrated multiple sub-populations. Significant differentiation (F ST  = 0.243) was observed between KM and KK, located just 130 Km apart. Consistent with low endemic transmission, infection complexity was modest in both KM (mean MOI  = 1.38) and KK (mean MOI  = 1.19). However, population diversity remained moderate (H E  = 0.583 in KM and H E  = 0.667 in KK). Temporal trends revealed clonal expansions reflecting epidemic transmission dynamics. The haplotypes of these isolates declined in frequency over time, but persisted at low frequency throughout the study duration. A diverse array of low frequency isolates were detected in both KM and KK, some likely reflecting remnants of previous expansions. In accordance with clonal expansions, high levels of Linkage Disequilibrium (I A (S) >0.5 [P<0.0001] in KK and KM) declined sharply when identical haplotypes were represented once (I A (S)  = 0.07 [P = 0.0076] in KM, and I A (S) = -0.003 [P = 0.606] in KK). All 8 recurrences, likely to be relapses, were homologous to the prior infection. These recurrences may promote the persistence of parasite lineages, sustaining local diversity. In summary, Sabah's shrinking P. vivax population appears to have rendered this low endemic setting vulnerable to epidemic expansions. Migration may play an important role in the introduction of new parasite strains leading to epidemic expansions, with important implications for malaria elimination.

Satzke C, Turner P, Virolainen-Julkunen A, Adrian PV, Antonio M, Hare KM, Henao-Restrepo AM, Leach AJ, Klugman KP, Porter BD et al. 2013. Standard method for detecting upper respiratory carriage of Streptococcus pneumoniae: updated recommendations from the World Health Organization Pneumococcal Carriage Working Group. Vaccine, 32 (1), pp. 165-179. | Citations: 128 (Scopus) | Show Abstract | Read more

In 2003 the World Health Organization (WHO) convened a working group and published a set of standard methods for studies measuring nasopharyngeal carriage of Streptococcus pneumoniae (the pneumococcus). The working group recently reconvened under the auspices of the WHO and updated the consensus standard methods. These methods describe the collection, transport and storage of nasopharyngeal samples, as well as provide recommendations for the identification and serotyping of pneumococci using culture and non-culture based approaches. We outline the consensus position of the working group, the evidence supporting this position, areas worthy of future research, and the epidemiological role of carriage studies. Adherence to these methods will reduce variability in the conduct of pneumococcal carriage studies undertaken in the context of pneumococcal vaccine trials, implementation studies, and epidemiology studies more generally so variability in methodology does not confound the interpretation of study findings.

Sriboonvorakul N, Leepipatpiboon N, Dondorp AM, Pouplin T, White NJ, Tarning J, Lindegardh N. 2013. Liquid chromatographic-mass spectrometric method for simultaneous determination of small organic acids potentially contributing to acidosis in severe malaria. J Chromatogr B Analyt Technol Biomed Life Sci, 941 pp. 116-122. | Citations: 18 (Web of Science Lite) | Show Abstract | Read more

Acidosis is an important cause of mortality in severe falciparum malaria. Lactic acid is a major contributor to metabolic acidosis, but accounts for only one-quarter of the strong anion gap. Other unidentified organic acids have an independent strong prognostic significance for a fatal outcome. In this study, a simultaneous bio-analytical method for qualitative and quantitative assessment in plasma and urine of eight small organic acids potentially contributing to acidosis in severe malaria was developed and validated. High-throughput strong anion exchange solid-phase extraction in a 96-well plate format was used for sample preparation. Hydrophilic interaction liquid chromatography (HILIC) coupled to negative mass spectroscopy was utilized for separation and detection. Eight possible small organic acids; l-lactic acid (LA), α-hydroxybutyric acid (aHBA), β-hydroxybutyric acid (bHBA), p-hydroxyphenyllactic acid (pHPLA), malonic acid (MA), methylmalonic acid (MMA), ethylmalonic acid (EMA) and α-ketoglutaric acid (aKGA) were analyzed simultaneously using a ZIC-HILIC column with an isocratic elution containing acetonitrile and ammonium acetate buffer. This method was validated according to U.S. Food and Drug Administration guidelines with additional validation procedures for endogenous substances. Accuracy for all eight acids ranged from 93.1% to 104.0%, and the within-day and between-day precisions (i.e. relative standard deviations) were lower than 5.5% at all tested concentrations. The calibration ranges were: 2.5-2500μg/mL for LA, 0.125-125μg/mL for aHBA, 7.5-375μg/mL for bHBA, 0.1-100μg/mL for pHPLA, 1-1000μg/mL for MA, 0.25-250μg/mL for MMA, 0.25-100μg/mL for EMA, and 30-1500μg/mL for aKGA. Clinical applicability was demonstrated by analyzing plasma and urine samples from five patients with severe falciparum malaria; five acids had increased concentrations in plasma (range LA=177-1169μg/mL, aHBA=4.70-38.4μg/mL, bHBA=7.70-38.0μg/mL, pHPLA=0.900-4.30μg/mL and aKGA=30.2-32.0) and seven in urine samples (range LA=11.2-513μg/mL, aHBA=1.50-69.5μg/mL, bHBA=8.10-111μg/mL, pHPLA=4.30-27.7μg/mL, MMA=0.300-13.3μg/mL, EMA=0.300-48.1μg/mL and aKGA=30.4-107μg/mL). In conclusion, a novel bioanalytical method was developed and validated which allows for simultaneous quantification of eight small organic acids in plasma and urine. This new method may be a useful tool for the assessment of acidosis in patients with severe malaria, and other conditions complicated by acidosis.

WorldWide Antimalarial Resistance Network (WWARN) DP Study Group. 2013. The effect of dosing regimens on the antimalarial efficacy of dihydroartemisinin-piperaquine: a pooled analysis of individual patient data. PLoS Med, 10 (12), pp. e1001564. | Citations: 48 (Scopus) | Show Abstract | Read more

BACKGROUND: Dihydroartemisinin-piperaquine (DP) is increasingly recommended for antimalarial treatment in many endemic countries; however, concerns have been raised over its potential under dosing in young children. We investigated the influence of different dosing schedules on DP's clinical efficacy. METHODS AND FINDINGS: A systematic search of the literature was conducted to identify all studies published between 1960 and February 2013, in which patients were enrolled and treated with DP. Principal investigators were approached and invited to share individual patient data with the WorldWide Antimalarial Resistance Network (WWARN). Data were pooled using a standardised methodology. Univariable and multivariable risk factors for parasite recrudescence were identified using a Cox's regression model with shared frailty across the study sites. Twenty-four published and two unpublished studies (n = 7,072 patients) were included in the analysis. After correcting for reinfection by parasite genotyping, Kaplan-Meier survival estimates were 97.7% (95% CI 97.3%-98.1%) at day 42 and 97.2% (95% CI 96.7%-97.7%) at day 63. Overall 28.6% (979/3,429) of children aged 1 to 5 years received a total dose of piperaquine below 48 mg/kg (the lower limit recommended by WHO); this risk was 2.3-2.9-fold greater compared to that in the other age groups and was associated with reduced efficacy at day 63 (94.4% [95% CI 92.6%-96.2%], p<0.001). After adjusting for confounding factors, the mg/kg dose of piperaquine was found to be a significant predictor for recrudescence, the risk increasing by 13% (95% CI 5.0%-21%) for every 5 mg/kg decrease in dose; p = 0.002. In a multivariable model increasing the target minimum total dose of piperaquine in children aged 1 to 5 years old from 48 mg/kg to 59 mg/kg would halve the risk of treatment failure and cure at least 95% of patients; such an increment was not associated with gastrointestinal toxicity in the ten studies in which this could be assessed. CONCLUSIONS: DP demonstrates excellent efficacy in a wide range of transmission settings; however, treatment failure is associated with a lower dose of piperaquine, particularly in young children, suggesting potential for further dose optimisation.

Maude RR, Hossain MA, Hassan MU, Osbourne S, Sayeed KLA, Karim MR, Samad R, Borooah S, Dhillon B, Day NPJ et al. 2013. Transorbital sonographic evaluation of normal optic nerve sheath diameter in healthy volunteers in Bangladesh. PLoS One, 8 (12), pp. e81013. | Citations: 24 (Scopus) | Show Abstract | Read more

INTRODUCTION: Measurement of optic nerve sheath diameter (ONSD) by ultrasound is increasingly used as a marker to detect raised intracranial pressure (ICP). ONSD varies with age and there is no clear consensus between studies for an upper limit of normal. Knowledge of normal ONSD in a healthy population is essential to interpret this measurement. METHODS: In a prospective observational study, ONSD was measured using a 15 MHz ultrasound probe in healthy volunteers in Chittagong, Bangladesh. The aims were to determine the normal range of ONSD in healthy Bangladeshi adults and children, compare measurements in males and females, horizontal and vertical beam orientations and left and right eyes in the same individual and to determine whether ONSD varies with head circumference independent of age. RESULTS: 136 subjects were enrolled, 12.5% of whom were age 16 or under. Median ONSD was 4.41 mm with 95% of subjects in the range 4.25-4.75 mm. ONSD was bimodally distributed. There was no relationship between ONSD and age (≥4 years), gender, head circumference, and no difference in left vs right eye or horizontal vs vertical beam. CONCLUSIONS: Ultrasonographic ONSD in Bangladeshi healthy volunteers has a narrow bimodal distribution independent of age (≥4 years), gender and head circumference. ONSD >4.75 mm in this population should be considered abnormal.

Turner P, Turner C, Green N, Ashton L, Lwe E, Jankhot A, Day NP, White NJ, Nosten F, Goldblatt D. 2013. Serum antibody responses to pneumococcal colonization in the first 2 years of life: Results from an SE Asian longitudinal cohort study Clinical Microbiology and Infection, 19 (12), | Citations: 25 (Scopus) | Show Abstract | Read more

Assessment of antibody responses to pneumococcal colonization in early childhood may aid our understanding of protection and inform vaccine antigen selection. Serum samples were collected from mother-infant pairs during a longitudinal pneumococcal colonization study in Burmese refugees. Maternal and cord sera were collected at birth and infants were bled monthly (1-24 months of age). Nasopharyngeal swabs were taken monthly to detect colonization. Serum IgG titres to 27 pneumococcal protein antigens were measured in 2624 sera and IgG to dominant serotypes (6B, 14, 19F, 19A and 23F) were quantified in 864 infant sera. Antibodies to all protein antigens were detectable in maternal sera. Titres to four proteins (LytB, PcpA, PhtD and PhtE) were significantly higher in mothers colonized by pneumococci at delivery. Maternally-derived antibodies to PiuA and Spr0096 were associated with delayed pneumococcal acquisition in infants in univariate, but not multivariate models. Controlling for infant age and previous homologous serotype exposure, nasopharyngeal acquisition of serotypes 19A, 23F, 14 or 19F was associated significantly with a ≥2-fold antibody response to the homologous capsule (OR 12.84, 7.52, 6.52, 5.33; p <0.05). Acquisition of pneumococcal serotypes in the nasopharynx of infants was not significantly associated with a ≥2-fold rise in antibodies to any of the protein antigens studied. In conclusion, nasopharyngeal colonization in young children resulted in demonstrable serum IgG responses to pneumococcal capsules and surface/virulence proteins. However, the relationship between serum IgG and the prevention of, or response to, pneumococcal nasopharyngeal colonization remains complex. Mechanisms other than serum IgG are likely to have a role but are currently poorly understood. ©2013 The Authors. Clinical Microbiology and Infection published by John Wiley & Sons Ltd on behalf of the European Society of Clinical Microbiology and Infectious Diseases.

Corbel V, Nosten F, Thanispong K, Luxemburger C, Kongmee M, Chareonviriyaphap T. 2013. Challenges and prospects for dengue and malaria control in Thailand, Southeast Asia Trends in Parasitology, 29 (12), pp. 623-633. | Citations: 27 (Scopus) | Show Abstract | Read more

Despite significant advances in the search for potential dengue vaccines and new therapeutic schemes for malaria, the control of these diseases remains difficult. In Thailand, malaria incidence is falling whereas that of dengue is rising, with an increase in the proportion of reported severe cases. In the absence of antiviral therapeutic options for acute dengue, appropriate case management reduces mortality. However, the interruption of transmission still relies on vector control measures that are currently insufficient to curtail the cycle of epidemics. Drug resistance in malaria parasites is increasing, compromising malaria control and elimination. Deficiencies in our knowledge of vector biology and vectorial capacity also hinder public health efforts for vector control. Challenges to dengue and malaria control are discussed, and research priorities identified. © 2013.

Nic Fhogartaigh C, Dance DAB. 2013. Bacterial gastroenteritis Medicine, 41 (12), pp. 693-699. | Citations: 9 (Scopus) | Show Abstract | Read more

Infectious diarrhoea is a major public health concern worldwide. Bacteria, the focus of this review, are responsible for 20-40% of diarrhoeal episodes, contributing to high rates of childhood mortality in developing regions, and substantial morbidity and economic losses in developed regions. The epidemiology is changing with salmonellosis decreasing in industrialized countries and diarrhoeagenic Escherichia coli contributing to an increasing burden of disease worldwide. Molecular diagnostics have improved our understanding of the epidemiology, aetiology and pathogenesis of bacterial gastroenteritis, and have revealed new pathogenic agents, although widespread introduction of such diagnostics into clinical practice will require careful cost-benefit analyses. The development of antimicrobial resistance in gastrointestinal pathogens has implications for treatment options. We review the epidemiology of infectious diarrhoea, the principal aetiological agents and their clinical features, and the diagnosis, treatment and prevention of bacterial gastroenteritis; we also propose an investigation and management algorithm. © 2013 Elsevier Ltd. All rights reserved.

Douglas NM, Lampah DA, Kenangalem E, Simpson JA, Poespoprodjo JR, Sugiarto P, Anstey NM, Price RN. 2013. Major burden of severe anemia from non-falciparum malaria species in Southern Papua: a hospital-based surveillance study. PLoS Med, 10 (12), pp. e1001575. | Citations: 52 (Web of Science Lite) | Show Abstract | Read more

BACKGROUND: The burden of anemia attributable to non-falciparum malarias in regions with Plasmodium co-endemicity is poorly documented. We compared the hematological profile of patients with and without malaria in southern Papua, Indonesia. METHODS AND FINDINGS: Clinical and laboratory data were linked for all patients presenting to a referral hospital between April 2004 and December 2012. Data were available on patient demographics, malaria diagnosis, hemoglobin concentration, and clinical outcome, but other potential causes of anemia could not be identified reliably. Of 922,120 patient episodes (837,989 as outpatients and 84,131 as inpatients), a total of 219,845 (23.8%) were associated with a hemoglobin measurement, of whom 67,696 (30.8%) had malaria. Patients with P. malariae infection had the lowest hemoglobin concentration (n = 1,608, mean = 8.93 [95% CI 8.81-9.06]), followed by those with mixed species infections (n = 8,645, mean = 9.22 [95% CI 9.16-9.28]), P. falciparum (n = 37,554, mean = 9.47 [95% CI 9.44-9.50]), and P. vivax (n = 19,858, mean = 9.53 [95% CI 9.49-9.57]); p-value for all comparisons <0.001. Severe anemia (hemoglobin <5 g/dl) was present in 8,151 (3.7%) patients. Compared to patients without malaria, those with mixed Plasmodium infection were at greatest risk of severe anemia (adjusted odds ratio [AOR] 3.25 [95% CI 2.99-3.54]); AORs for severe anaemia associated with P. falciparum, P. vivax, and P. malariae were 2.11 (95% CI 2.00-2.23), 1.87 (95% CI 1.74-2.01), and 2.18 (95% CI 1.76-2.67), respectively, p<0.001. Overall, 12.2% (95% CI 11.2%-13.3%) of severe anemia was attributable to non-falciparum infections compared with 15.1% (95% CI 13.9%-16.3%) for P. falciparum monoinfections. Patients with severe anemia had an increased risk of death (AOR = 5.80 [95% CI 5.17-6.50]; p<0.001). Not all patients had a hemoglobin measurement, thus limitations of the study include the potential for selection bias, and possible residual confounding in multivariable analyses. CONCLUSIONS: In Papua P. vivax is the dominant cause of severe anemia in early infancy, mixed P. vivax/P. falciparum infections are associated with a greater hematological impairment than either species alone, and in adulthood P. malariae, although rare, is associated with the lowest hemoglobin concentration. These findings highlight the public health importance of integrated genus-wide malaria control strategies in areas of Plasmodium co-endemicity.

Turner P, Turner C, Green N, Ashton L, Lwe E, Jankhot A, Day NP, White NJ, Nosten F, Goldblatt D. 2013. Serum antibody responses to pneumococcal colonization in the first 2 years of life: results from an SE Asian longitudinal cohort study. Clin Microbiol Infect, 19 (12), pp. E551-E558. | Citations: 20 (European Pubmed Central) | Show Abstract | Read more

Assessment of antibody responses to pneumococcal colonization in early childhood may aid our understanding of protection and inform vaccine antigen selection. Serum samples were collected from mother-infant pairs during a longitudinal pneumococcal colonization study in Burmese refugees. Maternal and cord sera were collected at birth and infants were bled monthly (1–24 months of age). Nasopharyngeal swabs were taken monthly to detect colonization. Serum IgG titres to 27 pneumococcal protein antigens were measured in 2624 sera and IgG to dominant serotypes (6B, 14, 19F, 19A and 23F) were quantified in 864 infant sera. Antibodies to all protein antigens were detect ablein maternal sera. Titres to four proteins (LytB, PcpA, PhtD and PhtE) were significantly higher in mothers colonized by pneumococci at delivery. Maternally-derived antibodies to PiuA and Spr0096 were associated with delayed pneumococcal acquisition in infants in univariate,but not multivariate models. Controlling for infant age and previous homologous serotype exposure, nasopharyngeal acquisition of serotypes 19A, 23F, 14 or 19F was associated significantly with a ≥2-fold antibody response to the homologous capsule (OR 12.84, 7.52,6.52, 5.33; p <0.05). Acquisition of pneumococcal serotypes in the nasopharynx of infants was not significantly associated with a ≥2-fold rise in antibodies to any of the protein antigens studied. In conclusion, nasopharyngeal colonization in young children resulted in demonstrable serum IgG responses to pneumococcal capsules and surface/virulence proteins. However, the relationship between serum IgG and the prevention of, or response to, pneumococcal nasopharyngeal colonization remains complex. Mechanisms other than serum IgG are likely to have a role but are currently poorly understood.

McNamara CW, Lee MCS, Lim CS, Lim SH, Roland J, Nagle A, Simon O, Yeung BKS, Chatterjee AK, McCormack SL et al. 2013. Targeting Plasmodium PI(4)K to eliminate malaria Nature, 504 (7479), pp. 248-253. | Citations: 171 (Scopus) | Show Abstract | Read more

Achieving the goal of malaria elimination will depend on targeting Plasmodium pathways essential across all life stages. Here we identify a lipid kinase, phosphatidylinositol-4-OH kinase (PI(4)K), as the target of imidazopyrazines, a new antimalarial compound class that inhibits the intracellular development of multiple Plasmodium species at each stage of infection in the vertebrate host. Imidazopyrazines demonstrate potent preventive, therapeutic, and transmission-blocking activity in rodent malaria models, are active against blood-stage field isolates of the major human pathogens P. falciparum and P. vivax, and inhibit liver-stage hypnozoites in the simian parasite P. cynomolgi. We show that imidazopyrazines exert their effect through inhibitory interaction with the ATP-binding pocket of PI(4)K, altering the intracellular distribution of phosphatidylinositol-4-phosphate. Collectively, our data define PI(4)K as a key Plasmodium vulnerability, opening up new avenues of target-based discovery to identify drugs with an ideal activity profile for the prevention, treatment and elimination of malaria. © 2013 Macmillan Publishers Limited. All rights reserved.

McNamara CW, Lee MC, Lim CS, Lim SH, Roland J, Simon O, Yeung BK, Chatterjee AK, McCormack SL, Manary MJ et al. 2013. Targeting Plasmodium PI(4)K to eliminate malaria. Nature, 504 (7479), pp. 248-253. | Citations: 157 (Web of Science Lite) | Show Abstract | Read more

Achieving the goal of malaria elimination will depend on targeting Plasmodium pathways essential across all life stages. Here we identify a lipid kinase, phosphatidylinositol-4-OH kinase (PI(4)K), as the target of imidazopyrazines, a new antimalarial compound class that inhibits the intracellular development of multiple Plasmodium species at each stage of infection in the vertebrate host. Imidazopyrazines demonstrate potent preventive, therapeutic, and transmission-blocking activity in rodent malaria models, are active against blood-stage field isolates of the major human pathogens P. falciparum and P. vivax, and inhibit liver-stage hypnozoites in the simian parasite P. cynomolgi. We show that imidazopyrazines exert their effect through inhibitory interaction with the ATP-binding pocket of PI(4)K, altering the intracellular distribution of phosphatidylinositol-4-phosphate. Collectively, our data define PI(4)K as a key Plasmodium vulnerability, opening up new avenues of target-based discovery to identify drugs with an ideal activity profile for the prevention, treatment and elimination of malaria.

Chantratita N, Tandhavanant S, Myers ND, Seal S, Arayawichanont A, Kliangsa-Ad A, Hittle LE, Ernst RK, Emond MJ, Wurfel MM et al. 2013. Survey of innate immune responses to Burkholderia pseudomallei in human blood identifies a central role for lipopolysaccharide. PLoS One, 8 (11), pp. e81617. | Citations: 19 (Scopus) | Show Abstract | Read more

B. pseudomallei is a gram-negative bacterium that causes the tropical infection melioidosis. In northeast Thailand, mortality from melioidosis approaches 40%. As exemplified by the lipopolysaccharide-Toll-like receptor 4 interaction, innate immune responses to invading bacteria are precipitated by activation of host pathogen recognition receptors by pathogen associated molecular patterns. Human melioidosis is characterized by up-regulation of pathogen recognition receptors and pro-inflammatory cytokine release. In contrast to many gram-negative pathogens, however, the lipopolysaccharide of B. pseudomallei is considered only weakly inflammatory. We conducted a study in 300 healthy Thai subjects to investigate the ex vivo human blood response to various bacterial pathogen associated molecular patterns, including lipopolysaccharide from several bacteria, and to two heat-killed B. pseudomallei isolates. We measured cytokine levels after stimulation of fresh whole blood with a panel of stimuli. We found that age, sex, and white blood cell count modulate the innate immune response to B. pseudomallei. We further observed that, in comparison to other stimuli, the innate immune response to B. pseudomallei is most highly correlated with the response to lipopolysaccharide. The magnitude of cytokine responses induced by B. pseudomallei lipopolysaccharide was significantly greater than those induced by lipopolysaccharide from Escherichia coli and comparable to many responses induced by lipopolysaccharide from Salmonella minnesota despite lower amounts of lipid A in the B. pseudomallei lipopolysaccharide preparation. In human monocytes stimulated with B. pseudomallei, addition of polymyxin B or a TLR4/MD-2 neutralizing antibody inhibited the majority of TNF-α production. Challenging existing views, our data indicate that the innate immune response to B. pseudomallei in human blood is largely driven by lipopolysaccharide, and that the response to B. pseudomallei lipopolysaccharide in blood is greater than the response to other lipopolysaccharide expressing isolates. Our findings suggest that B. pseudomallei lipopolysaccharide may play a central role in stimulating the host response in melioidosis.

Flegg JA, Guérin PJ, Nosten F, Ashley EA, Phyo AP, Dondorp AM, Fairhurst RM, Socheat D, Borrmann S, Björkman A et al. 2013. Optimal sampling designs for estimation of Plasmodium falciparum clearance rates in patients treated with artemisinin derivatives. Malar J, 12 (1), pp. 411. | Citations: 15 (Scopus) | Show Abstract | Read more

BACKGROUND: The emergence of Plasmodium falciparum resistance to artemisinins in Southeast Asia threatens the control of malaria worldwide. The pharmacodynamic hallmark of artemisinin derivatives is rapid parasite clearance (a short parasite half-life), therefore, the in vivo phenotype of slow clearance defines the reduced susceptibility to the drug. Measurement of parasite counts every six hours during the first three days after treatment have been recommended to measure the parasite clearance half-life, but it remains unclear whether simpler sampling intervals and frequencies might also be sufficient to reliably estimate this parameter. METHODS: A total of 2,746 parasite density-time profiles were selected from 13 clinical trials in Thailand, Cambodia, Mali, Vietnam, and Kenya. In these studies, parasite densities were measured every six hours until negative after treatment with an artemisinin derivative (alone or in combination with a partner drug). The WWARN Parasite Clearance Estimator (PCE) tool was used to estimate "reference" half-lives from these six-hourly measurements. The effect of four alternative sampling schedules on half-life estimation was investigated, and compared to the reference half-life (time zero, 6, 12, 24 (A1); zero, 6, 18, 24 (A2); zero, 12, 18, 24 (A3) or zero, 12, 24 (A4) hours and then every 12 hours). Statistical bootstrap methods were used to estimate the sampling distribution of half-lives for parasite populations with different geometric mean half-lives. A simulation study was performed to investigate a suite of 16 potential alternative schedules and half-life estimates generated by each of the schedules were compared to the "true" half-life. The candidate schedules in the simulation study included (among others) six-hourly sampling, schedule A1, schedule A4, and a convenience sampling schedule at six, seven, 24, 25, 48 and 49 hours. RESULTS: The median (range) parasite half-life for all clinical studies combined was 3.1 (0.7-12.9) hours. Schedule A1 consistently performed the best, and schedule A4 the worst, both for the individual patient estimates and for the populations generated with the bootstrapping algorithm. In both cases, the differences between the reference and alternative schedules decreased as half-life increased. In the simulation study, 24-hourly sampling performed the worst, and six-hourly sampling the best. The simulation study confirmed that more dense parasite sampling schedules are required to accurately estimate half-life for profiles with short half-life (≤ three hours) and/or low initial parasite density (≤ 10,000 per μL). Among schedules in the simulation study with six or fewer measurements in the first 48 hours, a schedule with measurements at times (time windows) of 0 (0-2), 6 (4-8), 12 (10-14), 24 (22-26), 36 (34-36) and 48 (46-50) hours, or at times 6, 7 (two samples in time window 5-8), 24, 25 (two samples during time 23-26), and 48, 49 (two samples during time 47-50) hours, until negative most accurately estimated the "true" half-life. For a given schedule, continuing sampling after two days had little effect on the estimation of half-life, provided that adequate sampling was performed in the first two days and the half-life was less than three hours. If the measured parasitaemia at two days exceeded 1,000 per μL, continued sampling for at least once a day was needed for accurate half-life estimates. CONCLUSIONS: This study has revealed important insights on sampling schedules for accurate and reliable estimation of Plasmodium falciparum half-life following treatment with an artemisinin derivative (alone or in combination with a partner drug). Accurate measurement of short half-lives (rapid clearance) requires more dense sampling schedules (with more than twice daily sampling). A more intensive sampling schedule is, therefore, recommended in locations where P. falciparum susceptibility to artemisinins is not known and the necessary resources are available. Counting parasite density at six hours is important, and less frequent sampling is satisfactory for estimating long parasite half-lives in areas where artemisinin resistance is present.

Jamsen KM, Duffull SB, Tarning J, Price RN, Simpson JA. 2013. A robust design for identification of the Parasite Clearance Estimator. Malar J, 12 (1), pp. 410. | Citations: 5 (Scopus) | Show Abstract | Read more

BACKGROUND: Anti-malarial efficacy needs to be monitored continually to ensure optimal dosing in the face of emerging anti-malarial drug resistance. The efficacy of artemisinin based combination therapies (ACT) is assessed by repeated measurements of parasite density in the blood of patients following treatment. Parasite density is measured from a capillary or venous blood sample, but this can be logistically and ethically challenging if multiple samples are required within a short time period. The aim of this work was to apply optimal design theory to derive clinically feasible blood sampling schedules from which parasite clearance could be defined using the Parasite Clearance Estimator (PCE), a recently developed tool to identify and quantify artemisinin resistance. METHODS: Robust T-optimal design methodology was applied to offer a sampling schedule that allows for discrimination across models that best describe an individual patient's parasite-time profile. The design was based on typical parasite-time profiles derived from the literature combined with key sampling constraints of no more than six samples per patient within 48 hours of initial treatment. The design was evaluated with a simulation-estimation procedure that implemented the PCE. RESULTS: The optimal sampling times (sampling windows) were: 0 (0 to 1.1), 5.8 (4.0 to 6.0), 9.9 (8.4 to 11.5), 24.8 (24.0 to 24.9), 36.3 (34.8 to 37.2) and 48 (47.3, 48.0) hours post initial treatment. The simulation-estimation procedure showed that the design supported identification of the appropriate method by the PCE to determine an individual's parasite clearance rate constant (the main output calculation from the PCE). CONCLUSIONS: The proposed sampling design requires six samples per patient within the first 48 hours. The derived design requires validation in a real world setting, but should be considered for future studies that intend to employ the PCE.

Lim C, Wannapinij P, White L, Day NPJ, Cooper BS, Peacock SJ, Limmathurotsakul D. 2013. Using a web-based application to define the accuracy of diagnostic tests when the gold standard is imperfect. PLoS One, 8 (11), pp. e79489. | Citations: 8 (European Pubmed Central) | Show Abstract | Read more

BACKGROUND: Estimates of the sensitivity and specificity for new diagnostic tests based on evaluation against a known gold standard are imprecise when the accuracy of the gold standard is imperfect. Bayesian latent class models (LCMs) can be helpful under these circumstances, but the necessary analysis requires expertise in computational programming. Here, we describe open-access web-based applications that allow non-experts to apply Bayesian LCMs to their own data sets via a user-friendly interface. METHODS/PRINCIPAL FINDINGS: Applications for Bayesian LCMs were constructed on a web server using R and WinBUGS programs. The models provided (http://mice.tropmedres.ac) include two Bayesian LCMs: the two-tests in two-population model (Hui and Walter model) and the three-tests in one-population model (Walter and Irwig model). Both models are available with simplified and advanced interfaces. In the former, all settings for Bayesian statistics are fixed as defaults. Users input their data set into a table provided on the webpage. Disease prevalence and accuracy of diagnostic tests are then estimated using the Bayesian LCM, and provided on the web page within a few minutes. With the advanced interfaces, experienced researchers can modify all settings in the models as needed. These settings include correlation among diagnostic test results and prior distributions for all unknown parameters. The web pages provide worked examples with both models using the original data sets presented by Hui and Walter in 1980, and by Walter and Irwig in 1988. We also illustrate the utility of the advanced interface using the Walter and Irwig model on a data set from a recent melioidosis study. The results obtained from the web-based applications were comparable to those published previously. CONCLUSIONS: The newly developed web-based applications are open-access and provide an important new resource for researchers worldwide to evaluate new diagnostic tests.

Lim C, Wannapinij P, White L, Day NPJ, Cooper BS, Peacock SJ, Limmathurotsakul D. 2013. Using a web-based application to define the accuracy of diagnostic tests when the gold standard is imperfect PLoS ONE, 8 (11), | Citations: 14 (Scopus) | Show Abstract | Read more

Background: Estimates of the sensitivity and specificity for new diagnostic tests based on evaluation against a known gold standard are imprecise when the accuracy of the gold standard is imperfect. Bayesian latent class models (LCMs) can be helpful under these circumstances, but the necessary analysis requires expertise in computational programming. Here, we describe open-access web-based applications that allow non-experts to apply Bayesian LCMs to their own data sets via a user-friendly interface. Methods/Principal Findings: Applications for Bayesian LCMs were constructed on a web server using R and WinBUGS programs. The models provided (http://mice.tropmedres.ac) include two Bayesian LCMs: the two-tests in two-population model (Hui and Walter model) and the three-tests in one-population model (Walter and Irwig model). Both models are available with simplified and advanced interfaces. In the former, all settings for Bayesian statistics are fixed as defaults. Users input their data set into a table provided on the webpage. Disease prevalence and accuracy of diagnostic tests are then estimated using the Bayesian LCM, and provided on the web page within a few minutes. With the advanced interfaces, experienced researchers can modify all settings in the models as needed. These settings include correlation among diagnostic test results and prior distributions for all unknown parameters. The web pages provide worked examples with both models using the original data sets presented by Hui and Walter in 1980, and by Walter and Irwig in 1988. We also illustrate the utility of the advanced interface using the Walter and Irwig model on a data set from a recent melioidosis study. The results obtained from the web-based applications were comparable to those published previously. Conclusions: The newly developed web-based applications are open-access and provide an important new resource for researchers worldwide to evaluate new diagnostic tests. © 2013 Lim et al.

Corbel V, Nosten F, Thanispong K, Luxemburger C, Kongmee M, Chareonviriyaphap T. 2013. Challenges and prospects for dengue and malaria control in Thailand, Southeast Asia. Trends Parasitol, 29 (12), pp. 623-633. | Citations: 22 (Web of Science Lite) | Show Abstract | Read more

Despite significant advances in the search for potential dengue vaccines and new therapeutic schemes for malaria, the control of these diseases remains difficult. In Thailand, malaria incidence is falling whereas that of dengue is rising, with an increase in the proportion of reported severe cases. In the absence of antiviral therapeutic options for acute dengue, appropriate case management reduces mortality. However, the interruption of transmission still relies on vector control measures that are currently insufficient to curtail the cycle of epidemics. Drug resistance in malaria parasites is increasing, compromising malaria control and elimination. Deficiencies in our knowledge of vector biology and vectorial capacity also hinder public health efforts for vector control. Challenges to dengue and malaria control are discussed, and research priorities identified.

Domingo GJ, Satyagraha AW, Anvikar A, Baird K, Bancone G, Bansil P, Carter N, Cheng Q, Culpepper J, Eziefula C et al. 2013. G6PD testing in support of treatment and elimination of malaria: recommendations for evaluation of G6PD tests. Malar J, 12 (1), pp. 391. | Citations: 59 (Scopus) | Show Abstract | Read more

Malaria elimination will be possible only with serious attempts to address asymptomatic infection and chronic infection by both Plasmodium falciparum and Plasmodium vivax. Currently available drugs that can completely clear a human of P. vivax (known as "radical cure"), and that can reduce transmission of malaria parasites, are those in the 8-aminoquinoline drug family, such as primaquine. Unfortunately, people with glucose-6-phosphate dehydrogenase (G6PD) deficiency risk having severe adverse reactions if exposed to these drugs at certain doses. G6PD deficiency is the most common human enzyme defect, affecting approximately 400 million people worldwide.Scaling up radical cure regimens will require testing for G6PD deficiency, at two levels: 1) the individual level to ensure safe case management, and 2) the population level to understand the risk in the local population to guide Plasmodium vivax treatment policy. Several technical and operational knowledge gaps must be addressed to expand access to G6PD deficiency testing and to ensure that a patient's G6PD status is known before deciding to administer an 8-aminoquinoline-based drug.In this report from a stakeholder meeting held in Thailand on October 4 and 5, 2012, G6PD testing in support of radical cure is discussed in detail. The focus is on challenges to the development and evaluation of G6PD diagnostic tests, and on challenges related to the operational aspects of implementing G6PD testing in support of radical cure. The report also describes recommendations for evaluation of diagnostic tests for G6PD deficiency in support of radical cure.

Kloprogge F, Piola P, Dhorda M, Muwanga S, Turyakira E, Apinan S, Lindegårdh N, Nosten F, Day NPJ, White NJ et al. 2013. Population Pharmacokinetics of Lumefantrine in Pregnant and Nonpregnant Women With Uncomplicated Plasmodium falciparum Malaria in Uganda. CPT Pharmacometrics Syst Pharmacol, 2 (11), pp. e83. | Citations: 22 (Scopus) | Show Abstract | Read more

Pregnancy alters the pharmacokinetic properties of many antimalarial compounds. The objective of this study was to evaluate the pharmacokinetic properties of lumefantrine in pregnant and nonpregnant women with uncomplicated Plasmodium falciparum malaria in Uganda after a standard fixed oral artemether-lumefantrine treatment. Dense venous (n = 26) and sparse capillary (n = 90) lumefantrine samples were drawn from pregnant patients. A total of 17 nonpregnant women contributed with dense venous lumefantrine samples. Lumefantrine pharmacokinetics was best described by a flexible absorption model with multiphasic disposition. Pregnancy and body temperature had a significant impact on the pharmacokinetic properties of lumefantrine. Simulations from the final model indicated 27% lower day 7 concentrations in pregnant women compared with nonpregnant women and a decreased median time of 0.92 and 0.42 days above previously defined critical concentration cutoff values (280 and 175 ng/ml, respectively). The standard artemether-lumefantrine dose regimen in P. falciparum malaria may need reevaluation in nonimmune pregnant women.CPT: Pharmacometrics & Systems Pharmacology (2013) 2, e83; doi:10.1038/psp.2013.59; advance online publication 13 November 2013.

Kanoksil M, Jatapai A, Peacock SJ, Limmathurotsakul D. 2013. Correction: Epidemiology, Microbiology and Mortality Associated with Community-Acquired Bacteremia in Northeast Thailand: A Multicenter Surveillance Study. PLoS One, 8 (10), | Show Abstract | Read more

[This corrects the article on p. e54714 in vol. 8.].

Kager LM, Weehuizen TA, Wiersinga WJ, Roelofs JJTH, Meijers JCM, Dondorp AM, van 't Veer C, van der Poll T. 2013. Endogenous α2-antiplasmin is protective during severe gram-negative sepsis (melioidosis). Am J Respir Crit Care Med, 188 (8), pp. 967-975. | Citations: 19 (Scopus) | Show Abstract | Read more

RATIONALE: α2-Antiplasmin (A2AP) is a major inhibitor of fibrinolysis by virtue of its capacity to inhibit plasmin. Although the fibrinolytic system is strongly affected by infection, the functional role of A2AP in the host response to sepsis is unknown. OBJECTIVES: To study the role of A2AP in melioidosis, a common form of community-acquired sepsis in Southeast Asia and Northern Australia caused by the gram-negative bacterium Burkholderia pseudomallei. METHODS: In a single-center observational study A2AP was measured in patients with culture-proven septic melioidosis. Wild-type and A2AP-deficient (A2AP(-/-)) mice were intranasally infected with B. pseudomallei to induce severe pneumosepsis (melioidosis). Parameters of inflammation and coagulation were measured, and survival studies were performed. MEASUREMENTS AND MAIN RESULTS: Patients with melioidosis showed elevated A2AP plasma levels. Likewise, A2AP levels in plasma and lung homogenates were elevated in mice infected with B. pseudomallei. A2AP-deficient (A2AP(-/-)) mice had a strongly disturbed host response during experimental melioidosis as reflected by enhanced bacterial growth at the primary site of infection accompanied by increased dissemination to distant organs. In addition, A2AP(-/-) mice showed more severe lung pathology and injury together with an increased accumulation of neutrophils and higher cytokine levels in lung tissue. A2AP deficiency further was associated with exaggerated systemic inflammation and coagulation, increased distant organ injury, and enhanced lethality. CONCLUSIONS: This study is the first to identify A2AP as a protective mediator during gram-negative (pneumo)sepsis by limiting bacterial growth, inflammation, tissue injury, and coagulation.

Smithuis FM, Kyaw MK, Phe UO, van der Broek I, Katterman N, Rogers C, Almeida P, Kager PA, Stepniewska K, Lubell Y et al. 2013. Entomological determinants of insecticide-treated bed net effectiveness in Western Myanmar. Malar J, 12 (1), pp. 364. | Citations: 7 (European Pubmed Central) | Show Abstract | Read more

BACKGROUND: In a large cluster randomized control trial of insecticide-treated bed nets (ITN) in Western Myanmar the malaria protective effect of ITN was found to be highly variable and, in aggregate, the effect was not statistically significant. A coincident entomological investigation measured malaria vector abundance and biting behaviour and the human population sleeping habits, factors relevant to ITN effectiveness. METHODS: Entomological surveys were carried out using different catching methods to identify potential malaria vector species and characterise their biting habits. The salivary glands were dissected from all female anophelines caught to identify sporozoites by microscopy. FINDINGS: Between 1995 and 2000 a total of 4,824 female anopheline mosquitoes were caught with various catching methods. A total of 916 person nights yielded 3,009 female anopheline mosquitoes between 6 pm and 6 am. Except for Anopheles annularis, which showed no apparent preference (51% outdoor biting), all major species showed a strong preference for outdoor biting; Anopheles epiroticus (79%), Anopheles subpictus (72%), Anopheles maculatus (92%), Anopheles aconitus (85%) and Anopheles vagus (72%). Most human biting occurred in the early evening with the peak biting time between 6 pm and 7 pm (35%). Overall 51% (1447/2837) of all bites recorded were between 6 pm and 8 pm. A large proportion of children were not sleeping under an ITN during peak biting times. Only one An. annularis mosquito (0.02%) had malaria sporozoites identified in the salivary glands. CONCLUSIONS: Peak vector biting occurred early in the evening and mainly occurred outdoors. The limited efficacy of ITN in this area of Western Myanmar may be explained by the biting behaviour of the prevalent Anopheles mosquito vectors in this area.

Smithuis FM, Kyaw MK, Phe UO, van der Broek I, Katterman N, Rogers C, Almeida P, Kager PA, Stepniewska K, Lubell Y et al. 2013. The effect of insecticide-treated bed nets on the incidence and prevalence of malaria in children in an area of unstable seasonal transmission in western Myanmar. Malar J, 12 (1), pp. 363. | Citations: 32 (Scopus) | Show Abstract | Read more

BACKGROUND: Insecticide-treated bed nets (ITN) reduce malaria morbidity and mortality consistently in Africa, but their benefits have been less consistent in Asia. This study's objective was to evaluate the malaria protective efficacy of village-wide usage of ITN in Western Myanmar and estimate the cost-effectiveness of ITN compared with extending early diagnosis and treatment services. METHODS: A cluster-randomized controlled trial was conducted in Rakhine State to assess the efficacy of ITNs in preventing malaria and anaemia in children and their secondary effects on nutrition and development. The data were aggregated for each village to obtain cluster-level infection rates. In total 8,175 children under 10 years of age were followed up for 10 months, which included the main malaria transmission period. The incidence and prevalence of Plasmodium falciparum and Plasmodium vivax infections, and the biting behaviour of Anopheles mosquitoes in the area were studied concurrently. The trial data along with costs for current recommended treatment practices were modelled to estimate the cost-effectiveness of ITNs compared with, or in addition to extending the coverage of early diagnosis and treatment services. RESULTS: In aggregate, malaria infections, spleen rates, haemoglobin concentrations, and weight for height, did not differ significantly during the study period between villages with and without ITNs, with a weighted mean difference of -2.6 P. falciparum episodes per 1,000 weeks at risk (95% Confidence Interval -7 to 1.8). In areas with a higher incidence of malaria there was some evidence ITN protective efficacy. The economic analysis indicated that, despite the uncertainty and variability in their protective efficacy in the different study sites, ITN could still be cost-effective, but not if they displaced funding for early diagnosis and effective treatment which is substantially more cost-effective. CONCLUSION: In Western Myanmar deployment of ITNs did not provide consistent protection against malaria in children living in malaria endemic villages. Early diagnosis and effective treatment is a more cost effective malaria control strategy than deployment of ITNs in this area where the main vector bites early in the evening, often before people are protected by an ITN.

Malleret B, Xu F, Mohandas N, Suwanarusk R, Chu C, Leite JA, Low K, Turner C, Sriprawat K, Zhang R et al. 2013. Significant biochemical, biophysical and metabolic diversity in circulating human cord blood reticulocytes. PLoS One, 8 (10), pp. e76062. | Citations: 40 (Scopus) | Show Abstract | Read more

BACKGROUND: The transition from enucleated reticulocytes to mature normocytes is marked by substantial remodeling of the erythrocytic cytoplasm and membrane. Despite conspicuous changes, most studies describe the maturing reticulocyte as a homogenous erythropoietic cell type. While reticulocyte staging based on fluorescent RNA stains such as thiazole orange have been useful in a clinical setting; these 'sub-vital' stains may confound delicate studies on reticulocyte biology and may preclude their use in heamoparasite invasion studies. DESIGN AND METHODS: Here we use highly purified populations of reticulocytes isolated from cord blood, sorted by flow cytometry into four sequential subpopulations based on transferrin receptor (CD71) expression: CD71high, CD71medium, CD71low and CD71negative. Each of these subgroups was phenotyped in terms of their, morphology, membrane antigens, biomechanical properties and metabolomic profile. RESULTS: Superficially CD71high and CD71medium reticulocytes share a similar gross morphology (large and multilobular) when compared to the smaller, smooth and increasingly concave reticulocytes as seen in the in the CD71low and CD71negativesamples. However, between each of the four sample sets we observe significant decreases in shear modulus, cytoadhesive capacity, erythroid receptor expression (CD44, CD55, CD147, CD235R, and CD242) and metabolite concentrations. Interestingly increasing amounts of boric acid was found in the mature reticulocytes. CONCLUSIONS: Reticulocyte maturation is a dynamic and continuous process, confounding efforts to rigidly classify them. Certainly this study does not offer an alternative classification strategy; instead we used a nondestructive sampling method to examine key phenotypic changes of in reticulocytes. Our study emphasizes a need to focus greater attention on reticulocyte biology.

Luangasanatip N, Hongsuwan M, Lubell Y, Limmathurotsakul D, Teparrukkul P, Chaowarat S, Day NPJ, Graves N, Cooper BS. 2013. Long-term survival after intensive care unit discharge in Thailand: a retrospective study. Crit Care, 17 (5), pp. R219. | Citations: 10 (Scopus) | Show Abstract | Read more

INTRODUCTION: Economic evaluations of interventions in the hospital setting often rely on the estimated long-term impact on patient survival. Estimates of mortality rates and long-term outcomes among patients discharged alive from the intensive care unit (ICU) are lacking from lower- and middle-income countries. This study aimed to assess the long-term survival and life expectancy (LE) amongst post-ICU patients in Thailand, a middle-income country. METHODS: In this retrospective cohort study, data from a regional tertiary hospital in northeast Thailand and the regional death registry were linked and used to assess patient survival time after ICU discharge. Adult ICU patients aged at least 15 years who had been discharged alive from an ICU between 1 January 2004 and 31 December 2005 were included in the study, and the death registry was used to determine deaths occurring in this cohort up to 31st December 2010. These data were used in conjunction with standard mortality life tables to estimate annual mortality and life expectancy. RESULTS: This analysis included 10,321 ICU patients. During ICU admission, 3,251 patients (31.5%) died. Of 7,070 patients discharged alive, 2,527 (35.7%) were known to have died within the five-year follow-up period, a mortality rate 2.5 times higher than that in the Thai general population (age and sex matched). The mean LE was estimated as 18.3 years compared with 25.2 years in the general population. CONCLUSIONS: Post-ICU patients experienced much higher rates of mortality than members of the general population over the five-year follow-up period, particularly in the first year after discharge. Further work assessing Health Related Quality of Life (HRQOL) in both post-ICU patients and in the general population in developing countries is needed.

Shanks GD, White NJ. 2013. The activation of vivax malaria hypnozoites by infectious diseases. Lancet Infect Dis, 13 (10), pp. 900-906. | Citations: 46 (Web of Science Lite) | Show Abstract | Read more

The periodicity of vivax malaria relapses may be explained by the activation of latent hypnozoites acquired from a previous malarial infection. The activation stimulus could be the febrile illness associated with acute malaria or a different febrile infection. We review historical records to examine the association between relapses of Plasmodium vivax and febrile infectious diseases. In data from British soldiers in Palestine, epidemic falciparum malaria triggered a smaller epidemic of P vivax relapses only in those who had been extensively exposed to malaria previously. Relapses did not follow pandemic influenza infection. Evidence from three simultaneous typhoid and malaria epidemics suggest that typhoid fever might activate P vivax hypnozoites. Some data lend support to the notion that vivax malaria relapse followed febrile illness caused by relapsing fever, trench fever, epidemic typhus, and Malta fever (brucellosis). These observations suggest that systemic parasitic and bacterial infections, but not viral infections, can activate P vivax hypnozoites. Specific components of the host's acute febrile inflammatory response, and not fever alone, are probably important factors in the provocation of a relapse of vivax malaria.

Elliott I, Dittrich S, Paris D, Sengduanphachanh A, Phoumin P, Newton PN. 2013. The use of dried cerebrospinal fluid filter paper spots as a substrate for PCR diagnosis of the aetiology of bacterial meningitis in the Lao PDR Clinical Microbiology and Infection, 19 (10), | Citations: 10 (Scopus) | Show Abstract | Read more

We investigated whether dried cerebrospinal fluid (CSF) conserved on filter paper can be used as a substrate for accurate PCR diagnosis of important causes of bacterial meningitis in the Lao PDR. Using mock CSF, we investigated and optimized filter paper varieties, paper punch sizes, elution volumes and quantities of DNA template to achieve sensitive and reliable detection of bacterial DNA from filter paper specimens. FTA Elute Micro Card™ (Whatman, Maidstone, UK) was the most sensitive, consistent and practical variety of filter paper. Following optimization, the lower limit of detection for Streptococcus pneumoniae from dried mock CSF spots was 14 genomic equivalents (GE)/μL (interquartile range 5.5 GE/μL) or 230 (IQR 65) colony forming units/mL. A prospective clinical evaluation for S. pneumoniae, S. suis and Neisseria meningitidis was performed. Culture and PCR performed on fresh liquid CSF from patients admitted with a clinical diagnosis of meningitis (n = 73) were compared with results derived from dried CSF spots. Four of five fresh PCR-positive CSF samples also tested PCR positive from dried CSF spots, with one patient under the limit of detection. In a retrospective study of S. pneumoniae samples (n = 20), the median (IQR; range) CSF S. pneumoniae bacterial load was 1.1 × 104 GE/μL (1.2 × 105; 1 to 6.1 × 106 DNA GE/μL). Utilizing the optimized methodology, we estimate an extrapolated sensitivity of 90%, based on the range of CSF genome counts found in Laos. Dried CSF filter paper spots could potentially help us to better understand the epidemiology of bacterial meningitis in resource-poor settings and guide empirical treatments and vaccination policies. © 2013 The Authors. Clinical Microbiology and Infection published by John Wiley & Sons Ltd on behalf of the European Society of Clinical Microbiology and Infectious Disease.

Hanson J, Lam SWK, Alam S, Pattnaik R, Mahanta KC, Uddin Hasan M, Mohanty S, Mishra S, Cohen S, Day N et al. 2013. The reliability of the physical examination to guide fluid therapy in adults with severe falciparum malaria: an observational study. Malar J, 12 (1), pp. 348. | Citations: 11 (Scopus) | Show Abstract | Read more

BACKGROUND: Adults with severe malaria frequently require intravenous fluid therapy to restore their circulating volume. However, fluid must be delivered judiciously as both under- and over-hydration increase the risk of complications and, potentially, death. As most patients will be cared for in a resource-poor setting, management guidelines necessarily recommend that physical examination should guide fluid resuscitation. However, the reliability of this strategy is uncertain. METHODS: To determine the ability of physical examination to identify hypovolaemia, volume responsiveness, and pulmonary oedema, clinical signs and invasive measures of volume status were collected independently during an observational study of 28 adults with severe malaria. RESULTS: The physical examination defined volume status poorly. Jugular venous pressure (JVP) did not correlate with intravascular volume as determined by global end diastolic volume index (GEDVI; r(s) = 0.07, p = 0.19), neither did dry mucous membranes (p = 0.85), or dry axillae (p = 0.09). GEDVI was actually higher in patients with decreased tissue turgor (p < 0.001). Poor capillary return correlated with GEDVI, but was present infrequently (7% of observations) and, therefore, insensitive. Mean arterial pressure (MAP) correlated with GEDVI (rs = 0.16, p = 0.002), but even before resuscitation patients with a low GEDVI had a preserved MAP. Anuria on admission was unrelated to GEDVI and although liberal fluid resuscitation led to a median hourly urine output of 100 ml in 19 patients who were not anuric on admission, four (21%) developed clinical pulmonary oedema subsequently. MAP was unrelated to volume responsiveness (p = 0.71), while a low JVP, dry mucous membranes, dry axillae, increased tissue turgor, prolonged capillary refill, and tachycardia all had a positive predictive value for volume responsiveness of ≤50%. Extravascular lung water ≥11 ml/kg indicating pulmonary oedema was present on 99 of the 353 times that it was assessed during the study, but was identified on less than half these occasions by tachypnoea, chest auscultation, or an elevated JVP. A clear chest on auscultation and a respiratory rate <30 breaths/minute could exclude pulmonary oedema on 82% and 72% of occasions respectively. CONCLUSIONS: Findings on physical examination correlate poorly with true volume status in adults with severe malaria and must be used with caution to guide fluid therapy. TRIAL REGISTRATION: Clinicaltrials.gov identifier: NCT00692627.

Koirala S, Basnyat B, Arjyal A, Shilpakar O, Shrestha K, Shrestha R, Shrestha UM, Agrawal K, Koirala KD, Thapa SD et al. 2013. Gatifloxacin versus ofloxacin for the treatment of uncomplicated enteric fever in Nepal: an open-label, randomized, controlled trial. PLoS Negl Trop Dis, 7 (10), pp. e2523. | Citations: 17 (Scopus) | Show Abstract | Read more

BACKGROUND: Fluoroquinolones are the most commonly used group of antimicrobials for the treatment of enteric fever, but no direct comparison between two fluoroquinolones has been performed in a large randomised trial. An open-label randomized trial was conducted to investigate whether gatifloxacin is more effective than ofloxacin in the treatment of uncomplicated enteric fever caused by nalidixic acid-resistant Salmonella enterica serovars Typhi and Paratyphi A. METHODOLOGY AND PRINCIPAL FINDINGS: Adults and children clinically diagnosed with uncomplicated enteric fever were enrolled in the study to receive gatifloxacin (10 mg/kg/day) in a single dose or ofloxacin (20 mg/kg/day) in two divided doses for 7 days. Patients were followed for six months. The primary outcome was treatment failure in patients infected with nalidixic acid resistant isolates. 627 patients with a median age of 17 (IQR 9-23) years were randomised. Of the 218 patients with culture confirmed enteric fever, 170 patients were infected with nalidixic acid-resistant isolates. In the ofloxacin group, 6 out of 83 patients had treatment failure compared to 5 out of 87 in the gatifloxacin group (hazard ratio [HR] of time to failure 0.81, 95% CI 0.25 to 2.65, p = 0.73). The median time to fever clearance was 4.70 days (IQR 2.98-5.90) in the ofloxacin group versus 3.31 days (IQR 2.29-4.75) in the gatifloxacin group (HR = 1.59, 95% CI 1.16 to 2.18, p = 0.004). The results in all blood culture-confirmed patients and all randomized patients were comparable. CONCLUSION: Gatifloxacin was not superior to ofloxacin in preventing failure, but use of gatifloxacin did result in more prompt fever clearance time compared to ofloxacin. TRIAL REGISTRATION: ISRCTN 63006567 (www.controlled-trials.com).

Participants in the Community Engagement and Consent Workshop , Kilifi, Kenya , March 2011. 2013. Consent and community engagement in diverse research contexts. J Empir Res Hum Res Ethics, 8 (4), pp. 1-18. | Citations: 34 (Scopus) | Show Abstract | Read more

Consent and community engagement (CE) in health research are two aspects of a single concern-that research is carried out in a respectful manner where social value is maximized. There are important overlaps and interdependencies between consent and CE; for example, CE can provide insights into how best to tailor consent to context and can be an important component of consent processes. Engaging communities can also have intrinsic and instrumental value beyond consent; for example, as a means of showing respect and identifying appropriate ways of working respectfully. In this paper we critically examine how CE and consent processes are characterized, conducted, and evaluated in diverse health research contexts, and propose a preliminary research agenda to support future learning in these critical areas.

Aguas R, Ferguson NM. 2013. Feature selection methods for identifying genetic determinants of host species in RNA viruses. PLoS Comput Biol, 9 (10), pp. e1003254. | Citations: 7 (Scopus) | Show Abstract | Read more

Despite environmental, social and ecological dependencies, emergence of zoonotic viruses in human populations is clearly also affected by genetic factors which determine cross-species transmission potential. RNA viruses pose an interesting case study given their mutation rates are orders of magnitude higher than any other pathogen--as reflected by the recent emergence of SARS and Influenza for example. Here, we show how feature selection techniques can be used to reliably classify viral sequences by host species, and to identify the crucial minority of host-specific sites in pathogen genomic data. The variability in alleles at those sites can be translated into prediction probabilities that a particular pathogen isolate is adapted to a given host. We illustrate the power of these methods by: 1) identifying the sites explaining SARS coronavirus differences between human, bat and palm civet samples; 2) showing how cross species jumps of rabies virus among bat populations can be readily identified; and 3) de novo identification of likely functional influenza host discriminant markers.

Tanomsing N, Imwong M, Sutherland CJ, Dolecek C, Hien TT, Nosten F, Day NPJ, White NJ, Snounou G. 2013. Genetic marker suitable for identification and genotyping of Plasmodium ovale curtisi and Plasmodium ovale wallikeri. J Clin Microbiol, 51 (12), pp. 4213-4216. | Citations: 12 (Scopus) | Show Abstract | Read more

We present a seminested PCR method that specifically discriminates between Plasmodium ovale curtisi and P. ovale wallikeri with high sensitivity. The test is based on species-specific amplification of a size-polymorphic fragment of the tryptophan-rich antigen gene, potra, which also permits discrimination of intraspecific sequence variants at this locus.

Suntornsut P, Kasemsupat K, Silairatana S, Wongsuvan G, Jutrakul Y, Wuthiekanun V, Day NPJ, Peacock SJ, Limmathurotsakul D. 2013. Prevalence of melioidosis in patients with suspected pulmonary tuberculosis and sputum smear negative for acid-fast bacilli in northeast Thailand. Am J Trop Med Hyg, 89 (5), pp. 983-985. | Citations: 10 (Scopus) | Show Abstract | Read more

The clinical and radiological features of pulmonary melioidosis can mimic tuberculosis. We prospectively evaluated 118 patients with suspected pulmonary tuberculosis who were acid-fast bacilli (AFB) smear negative at Udon Thani Hospital, northeast Thailand. Culture of residual sputum from AFB testing was positive for Burkholderia pseudomallei in three patients (2.5%; 95% confidence interval [CI] 0.5-7.3%). We propose that in melioidosis-endemic areas, residual sputum from AFB testing should be routinely cultured for B. pseudomallei.

Goodyear A, Strange L, Rholl DA, Silisouk J, Dance DAB, Schweizer HP, Dow S. 2013. An improved selective culture medium enhances the isolation of Burkholderia pseudomallei from contaminated specimens. Am J Trop Med Hyg, 89 (5), pp. 973-982. | Citations: 5 (Scopus) | Show Abstract | Read more

Burkholderia pseudomallei is a Gram-negative environmental bacterium found in tropical climates that causes melioidosis. Culture remains the diagnostic gold standard, but isolation of B. pseudomallei from heavily contaminated sites, such as fecal specimens, can be difficult. We recently reported that B. pseudomallei is capable of infecting the gastrointestinal tract of mice and suggested that the same may be true in humans. Thus, there is a strong need for new culture techniques to allow for efficient detection of B. pseudomallei in fecal and other specimens. We found that the addition of norfloxacin, ampicillin, and polymyxin B to Ashdown's medium (NAP-A) resulted in increased specificity without affecting the growth of 25 B. pseudomallei strains. Furthermore, recovery of B. pseudomallei from human clinical specimens was not affected by the three additional antibiotics. Therefore, we conclude that NAP-A medium provides a new tool for more sensitive isolation of B. pseudomallei from heavily contaminated sites.

Turner P, Turner C, Watthanaworawit W, Carrara V, Cicelia N, Deglise C, Phares C, Ortega L, Nosten F. 2013. Respiratory virus surveillance in hospitalised pneumonia patients on the Thailand-Myanmar border. BMC Infect Dis, 13 (1), pp. 434. | Citations: 11 (European Pubmed Central) | Show Abstract | Read more

BACKGROUND: Pneumonia is a significant cause of morbidity and mortality in the developing world. Viruses contribute significantly to pneumonia burden, although data for low-income and tropical countries are scarce. The aim of this laboratory-enhanced, hospital-based surveillance was to characterise the epidemiology of respiratory virus infections among refugees living on the Thailand-Myanmar border. METHODS: Maela camp provides shelter for ~45,000 refugees. Inside the camp, a humanitarian organisation provides free hospital care in a 158-bed inpatient department (IPD). Between 1st April 2009 and 30th September 2011, all patients admitted to the IPD with a clinical diagnosis of pneumonia were invited to participate. Clinical symptoms and signs were recorded and a nasopharyngeal aspirate (NPA) collected. NPAs were tested for adenoviruses, human metapneumovirus (hMPV), influenza A & B, and RSV by PCR. RESULTS: Seven hundred eight patient episodes (698 patients) diagnosed as pneumonia during the enhanced surveillance period were included in this analysis. The median patient age was 1 year (range: < 1-70), and 90.4% were aged < 5 years. At least one virus was detected in 53.7% (380/708) of episodes. Virus detection was more common in children aged < 5 years old (<1 year: OR 2.0, 95% CI 1.2-3.4, p = 0.01; 1-4 years: OR 1.4, 95% CI 0.8-2.3, p = 0.2). RSV was detected in 176/708 (24.9%); an adenovirus in 133/708 (18.8%); an influenza virus in 68/708 (9.6%); and hMPV in 33/708 (4.7%). Twenty-eight episodes of multiple viral infections were identified, most commonly adenovirus plus another virus. RSV was more likely to be detected in children <5 years (OR 12.3, 95% CI 3.0-50.8, p = 0.001) and influenza viruses in patients ≥5 years (OR 2.8, 95% CI 1.5-5.4, p = 0.002). IPD treatment was documented in 702/708 cases; all but one patient received antimicrobials, most commonly a beta-lactam (amoxicillin/ampicillin +/-gentamicin in 664/701, 94.7%). CONCLUSIONS: Viral nucleic acid was identified in the nasopharynx in half the patients admitted with clinically diagnosed pneumonia. Development of immunisations targeting common respiratory viruses is likely to reduce the incidence of pneumonia in children living refugee camps and similar settings.

Chantratita N, Tandhavanant S, Wongsuvan G, Wuthiekanun V, Teerawattanasook N, Day NPJ, Limmathurotsakul D, Peacock SJ. 2013. Rapid detection of Burkholderia pseudomallei in blood cultures using a monoclonal antibody-based immunofluorescent assay. Am J Trop Med Hyg, 89 (5), pp. 971-972. | Citations: 13 (Scopus) | Show Abstract | Read more

Melioidosis is a severe bacterial infection caused by Burkholderia pseudomallei. Rapid antimicrobial therapy is necessary to improve patient outcome, which is aided by direct detection of B. pseudomallei in clinical samples. A drawback for all antigen assays is that the number of B. pseudomallei in blood usually falls below the achievable level of detection. We performed a prospective cohort study of 461 patients with 541 blood cultures to evaluate the utility of a pre-incubation step prior to detection of B. pseudomallei using a monoclonal antibody-based immunofluorescent assay (Mab-IFA). The Mab-IFA was positive in 74 of 76 patients with melioidosis (sensitivity = 97.4%), and negative in 385 patients who did not have blood cultures containing B. pseudomallei (specificity = 100%). The Mab-IFA could be a valuable supplementary tool for rapid detection. We recommend the use of the Mab-IFA to test blood cultures that flag positive in regions where melioidosis is endemic.

Sonthayanon P, Chierakul W, Wuthiekanun V, Limmathurotsakul D, Amornchai P, Smythe LD, Day NP, Peacock SJ. 2013. Molecular confirmation of co-infection by pathogenic Leptospira spp. and Orientia tsutsugamushi in patients with acute febrile illness in Thailand. Am J Trop Med Hyg, 89 (4), pp. 797-799. | Citations: 7 (Scopus) | Show Abstract | Read more

Leptospirosis and scrub typhus are major causes of acute febrile illness in rural Asia, where co-infection is reported to occur based on serologic evidence. We re-examined whether co-infection occurs by using a molecular approach. A duplex real-time polymerase chain reaction was developed that targeted a specific 16S ribosomal RNA gene of pathogenic Leptospira spp. and Orientia tsutsugamushi. Of 82 patients with an acute febrile illness who had dual infection on the basis of serologic tests, 5 (6%) had polymerase chain reaction results positive for both pathogens. We conclude that dual infection occurs, but that serologic tests may overestimate the frequency of co-infections.

Thaipadungpanit J, Wuthiekanun V, Chantratita N, Yimsamran S, Amornchai P, Boonsilp S, Maneeboonyang W, Tharnpoophasiam P, Saiprom N, Mahakunkijcharoen Y et al. 2013. Leptospira species in floodwater during the 2011 floods in the Bangkok Metropolitan Region, Thailand. Am J Trop Med Hyg, 89 (4), pp. 794-796. | Citations: 15 (Scopus) | Show Abstract | Read more

Floodwater samples (N = 110) collected during the 2011 Bangkok floods were tested for Leptospira using culture and polymerase chain reaction (PCR); 65 samples were PCR-positive for putatively non-pathogenic Leptospira species, 1 sample contained a putatively pathogenic Leptospira, and 6 samples contained Leptospira clustering phylogenetically with the intermediate group. The low prevalence of pathogenic and intermediate Leptospira in floodwater was consistent with the low number of human leptospirosis cases reported to the Bureau of Epidemiology in Thailand. This study provides baseline information on environmental Leptospira in Bangkok together with a set of laboratory tests that could be readily deployed in the event of future flooding.

White NJ. 2013. Pharmacokinetic and pharmacodynamic considerations in antimalarial dose optimization. Antimicrob Agents Chemother, 57 (12), pp. 5792-5807. | Citations: 39 (Web of Science Lite) | Show Abstract | Read more

Antimalarial drugs have usually been first deployed in areas of malaria endemicity at doses which were too low, particularly for high-risk groups such as young children and pregnant women. This may accelerate the emergence and spread of resistance, thereby shortening the useful life of the drug, but it is an inevitable consequence of the current imprecise method of dose finding. An alternative approach to dose finding is suggested in which phase 2 studies concentrate initially on pharmacokinetic-pharmacodynamic (PK-PD) characterization and in vivo calibration of in vitro susceptibility information. PD assessment is facilitated in malaria because serial parasite densities are readily assessed by microscopy, and at low densities by quantitative PCR, so that initial therapeutic responses can be quantitated accurately. If the in vivo MIC could be characterized early in phase 2 studies, it would provide a sound basis for the choice of dose in all target populations in subsequent combination treatments. Population PK assessments in phase 2b and phase 3 studies which characterize PK differences between different age groups, clinical disease states, and human populations can then be combined with the PK-PD observations to provide a sound evidence base for dose recommendations in different target groups.

Deeny SR, Cooper BS, Cookson B, Hopkins S, Robotham JV. 2013. Targeted versus universal screening and decolonization to reduce healthcare-associated meticillin-resistant Staphylococcus aureus infection. J Hosp Infect, 85 (1), pp. 33-44. | Citations: 18 (Web of Science Lite) | Show Abstract | Read more

BACKGROUND: The benefits of universal meticillin-resistant Staphylococcus aureus (MRSA) admission screening, compared with screening targeted patient groups and the additional impact of discharge screening, are uncertain. AIMS: To quantify the impact of MRSA screening plus decolonization treatment on MRSA infection rates. To compare universal with targeted screening policies, and to evaluate the additional impact of screening and decolonization on discharge. METHODS: A stochastic, individual-based model of MRSA transmission was developed that included patient movements between general medical and intensive care unit (ICU) wards, and between the hospital and community, informed by 18 months of individual patient data from a 900-bed tertiary care hospital. We simulated the impact of universal and targeted [for ICU, acute care of the elderly (ACE) or readmitted patients] MRSA screening and decolonization policies, both on admission and discharge. FINDINGS: Universal admission screening plus decolonization resulted in 77% (95% confidence interval: 76-78) reduction in MRSA infections over 10 years. Screening only ACE specialty or ICU patients yielded 62% (61-63) and 66% (65-67) reductions, respectively. Targeted policies reduced the number of screens by up to 95% and courses of decolonization by 96%. In addition to screening on admission, screening on discharge had little impact, with a maximum 7% additional reduction in infection. CONCLUSIONS: Compared with universal screening, targeted screening substantially reduced the amount of screening and decolonization required to achieve only 12% lower reduction in infection. Targeted screening and decolonization could lower the risk of resistance emerging as well as offer a more efficient use of resources.

Wittekamp BHJ, Oostdijk EAN, Cooper BS, Brun-Buisson C, Bonten MJM. 2013. Studies of selective decontamination. Lancet Infect Dis, 13 (9), pp. 736-737. | Read more

Newton P. 2013. Causes of fever - hoof beats are rarely from zebras TROPICAL MEDICINE & INTERNATIONAL HEALTH, 18 pp. 7-7. | Citations: 1 (Web of Science Lite)

Plancon A, Newton P. 2013. Building bridges between law enforcement and public health communities to combat pharmaceutical crime TROPICAL MEDICINE & INTERNATIONAL HEALTH, 18 pp. 20-20.

Lee AS, Cooper BS, Malhotra-Kumar S, Chalfine A, Daikos GL, Fankhauser C, Carevic B, Lemmen S, Martínez JA, Masuet-Aumatell C et al. 2013. Comparison of strategies to reduce meticillin-resistant Staphylococcus aureus rates in surgical patients: a controlled multicentre intervention trial. BMJ Open, 3 (9), pp. e003126. | Citations: 35 (Scopus) | Show Abstract | Read more

OBJECTIVE: To compare the effect of two strategies (enhanced hand hygiene vs meticillin-resistant Staphylococcus aureus (MRSA) screening and decolonisation) alone and in combination on MRSA rates in surgical wards. DESIGN: Prospective, controlled, interventional cohort study, with 6-month baseline, 12-month intervention and 6-month washout phases. SETTING: 33 surgical wards of 10 hospitals in nine countries in Europe and Israel. PARTICIPANTS: All patients admitted to the enrolled wards for more than 24 h. INTERVENTIONS: The two strategies compared were (1) enhanced hand hygiene promotion and (2) universal MRSA screening with contact precautions and decolonisation (intranasal mupirocin and chlorhexidine bathing) of MRSA carriers. Four hospitals were assigned to each intervention and two hospitals combined both strategies, using targeted MRSA screening. OUTCOME MEASURES: Monthly rates of MRSA clinical cultures per 100 susceptible patients (primary outcome) and MRSA infections per 100 admissions (secondary outcome). Planned subgroup analysis for clean surgery wards was performed. RESULTS: After adjusting for clustering and potential confounders, neither strategy when used alone was associated with significant changes in MRSA rates. Combining both strategies was associated with a reduction in the rate of MRSA clinical cultures of 12% per month (adjusted incidence rate ratios (aIRR) 0.88, 95% CI 0.79 to 0.98). In clean surgery wards, strategy 2 (MRSA screening, contact precautions and decolonisation) was associated with decreasing rates of MRSA clinical cultures (15% monthly decrease, aIRR 0.85, 95% CI 0.74 to 0.97) and MRSA infections (17% monthly decrease, aIRR 0.83, 95% CI 0.69 to 0.99). CONCLUSIONS: In surgical wards with relatively low MRSA prevalence, a combination of enhanced standard and MRSA-specific infection control approaches was required to reduce MRSA rates. Implementation of single interventions was not effective, except in clean surgery wards where MRSA screening coupled with contact precautions and decolonisation was associated with significant reductions in MRSA clinical culture and infection rates. TRIAL REGISTRATION: clinicaltrials.gov identifier: NCT00685867.

Mayxay M, Castonguay-Vanier J, Chansamouth V, Dubot-Pérès A, Paris DH, Phetsouvanh R, Tangkhabuanbutra J, Douangdala P, Inthalath S, Souvannasing P et al. 2013. Causes of non-malarial fever in Laos: a prospective study. Lancet Glob Health, 1 (1), pp. e46-e54. | Citations: 98 (Scopus) | Show Abstract | Read more

BACKGROUND: Because of reductions in the incidence of Plasmodium falciparum malaria in Laos, identification of the causes of fever in people without malaria, and discussion of the best empirical treatment options, are urgently needed. We aimed to identify the causes of non-malarial acute fever in patients in rural Laos. METHODS: For this prospective study, we recruited 1938 febrile patients, between May, 2008, and December, 2010, at Luang Namtha provincial hospital in northwest Laos (n=1390), and between September, 2008, and December, 2010, at Salavan provincial hospital in southern Laos (n=548). Eligible participants were aged 5-49 years with fever (≥38°C) lasting 8 days or less and were eligible for malaria testing by national guidelines. FINDINGS: With conservative definitions of cause, we assigned 799 (41%) patients a diagnosis. With exclusion of influenza, the top five diagnoses when only one aetiological agent per patient was identified were dengue (156 [8%] of 1927 patients), scrub typhus (122 [7%] of 1871), Japanese encephalitis virus (112 [6%] of 1924), leptospirosis (109 [6%] of 1934), and bacteraemia (43 [2%] of 1938). 115 (32%) of 358 patients at Luang Namtha hospital tested influenza PCR-positive between June and December, 2010, of which influenza B was the most frequently detected strain (n=121 [87%]). Disease frequency differed significantly between the two sites: Japanese encephalitis virus infection (p=0·04), typhoid (p=0·006), and leptospirosis (p=0·001) were more common at Luang Namtha, whereas dengue and malaria were more common at Salavan (all p<0·0001). With use of evidence from southeast Asia when possible, we estimated that azithromycin, doxycycline, ceftriaxone, and ofloxacin would have had significant efficacy for 258 (13%), 240 (12%), 154 (8%), and 41 (2%) of patients, respectively. INTERPRETATION: Our findings suggest that a wide range of treatable or preventable pathogens are implicated in non-malarial febrile illness in Laos. Empirical treatment with doxycycline for patients with undifferentiated fever and negative rapid diagnostic tests for malaria and dengue could be an appropriate strategy for rural health workers in Laos. FUNDING: Wellcome Trust, WHO-Western Pacific Region, Foundation for Innovative New Diagnostics, US Centers for Disease Control and Prevention

Goehler A, Trung TT, Hopf V, Kohler A, Wuthiekanun V, Peacock S, Limmathurotsakul D, Steinmetz I. 2013. Environmental presence of Burkholderia pseudomallei in rice fields as determined by direct PCR from soil samples INTERNATIONAL JOURNAL OF MEDICAL MICROBIOLOGY, 303 pp. 75-75.

Turner C, Carrara V, Aye Mya Thein N, Chit Mo Mo Win N, Turner P, Bancone G, White NJ, McGready R, Nosten F. 2013. Neonatal intensive care in a Karen refugee camp: a 4 year descriptive study. PLoS One, 8 (8), pp. e72721. | Citations: 18 (Scopus) | Show Abstract | Read more

BACKGROUND: A third of all deaths in children aged <5 years occur in the neonatal period. Neonatal intensive care is often considered too complex and expensive to be implemented in resource poor settings. Consequently the reductions that have been made in infant mortality in the poorest countries have not been made in the neonatal period. This manuscript describes the activities surrounding the introduction of special care baby unit (SCBU) in a refugee setting and the resulting population impact. METHODS: A SCBU was developed in Maela refugee camp on the Thailand-Myanmar border. This unit comprised of a dedicated area, basic equipment, drugs and staff training. Training was built around neonatal guidelines, comprising six clinical steps: recognition, resuscitation, examination, supportive medical care, specialised medical care, and counselling of parents with sick newborns. RESULTS: From January 2008 until December 2011, 952 infants were admitted to SCBU. The main admission diagnoses were early onset neonatal sepsis, jaundice and prematurity. Early prematurity (<34 weeks) carried the highest risk of mortality (OR 9.5, 95% CI 5.4-16.5, p<0.001). There was a significant decrease in mortality from 19.3% (2008) to 4.8% (2011) among the infants admitted for prematurity (p=0.03). The neonatal mortality in Maela camp as a whole declined by 51% from 21.8 to 10.7 deaths per 1000 live births over the corresponding period (p=0.04). Staff expressed more confidence in their ability to take care of neonates and there was a more positive attitude towards premature infants. CONCLUSION: Neonatal mortality can be reduced in a resource poor setting by introduction of a simple low cost unit specialising in care of sick neonates and run by local health workers following adequate training. Training in recognition and provision of simple interventions at a high standard can increase staff confidence and reduce fatalistic attitudes towards premature neonates.

Nantakomol D, Paul R, Palasuwan A, Day NPJ, White NJ, Imwong M. 2013. Evaluation of the phenotypic test and genetic analysis in the detection of glucose-6-phosphate dehydrogenase deficiency. Malar J, 12 (1), pp. 289. | Citations: 22 (Scopus) | Show Abstract | Read more

BACKGROUND: Glucose-6-phosphate dehydrogenase (G6PD) deficiency is particularly prevalent in historically malaria-endemic countries. Although most individuals with G6PD deficiency are asymptomatic, deficiency can result in acute haemolytic anaemia after exposure to oxidative agents. A reliable test is necessary for diagnosing the deficiency to prevent an acute haemolytic crisis following, for example, anti-malarial treatment. The aim of this study was to investigate which method was the best predictor of this disorder. METHODS: The present study investigated four G6PD activity detections (fluorescence spot (FS), methaemoglobin reduction (MR), biochemical and cytochemical test). These methods accompanied with mutation analysis of blood samples were taken from 295 apparently healthy individuals with unknown G6PD deficiency status. RESULTS: Molecular characterization of 295 Thai adults revealed an overall prevalence of 14.2%. The G6PD Viangchan (871 G>A) was the most common (83.3%), followed by G6PD Mahidol (487G>A) (11.9%), and G6PD Union (1360 C>T) (4.8%). There were two cases of G6PD deficiency carrying the double mutations of Viangchan (871G > A)-Mahidol (487G > A) and Viangchan (871G > A)-Union (1360C > T). In comparison, the prevalence of G6PD deficiency was 6.1% by FS test and 7.1% by MR test. G6PD activity was 11 ± 2.5 IU/gHb in non-deficient females (mean ± SD), and 10.9 ± 0.6 IU/gHb in non-deficient males. The upper and lower limit cut-off points for partial and severe deficiency in adults were 5.7 IU/gHb (60% of the normal mean) and 0.95 IU/gHb (10% of the normal mean), respectively. All hemizygote, homozygote and double mutations were associated with severe enzyme deficiency (the residual enzyme activity <10% of the normal mean), whereas only 14.3% of the heterozygote mutations showed severe enzyme deficiency. Based on the cut-off value <5.7 IU/gHb, the quantitative G6PD assay diagnosed 83% of cases as G6PD-deficient. Using a cut-off number of negative cell >20% in the cytochemical assay to define G6PD deficiency, the prevalence of G6PD deficiency was closest to the molecular analysis (12.9% G6PD-deficient) compared to the others methods. CONCLUSION: The cytochemical method is a significant predictor of this disease, while FS and MR test are recommended for the detection of severe G6PD deficiency in developing countries.

Ralph AP, Yeo TW, Salome CM, Waramori G, Pontororing GJ, Kenangalem E, Sandjaja, Tjitra E, Lumb R, Maguire GP et al. 2013. Impaired pulmonary nitric oxide bioavailability in pulmonary tuberculosis: association with disease severity and delayed mycobacterial clearance with treatment. J Infect Dis, 208 (4), pp. 616-626. | Citations: 13 (Scopus) | Show Abstract | Read more

BACKGROUND: Nitric oxide (NO), a key macrophage antimycobacterial mediator that ameliorates immunopathology, is measurable in exhaled breath in individuals with pulmonary tuberculosis. We investigated relationships between fractional exhale NO (FENO) and initial pulmonary tuberculosis severity, change during treatment, and relationship with conversion of sputum culture to negative at 2 months. METHODS: In Papua, we measured FENO in patients with pulmonary tuberculosis at baseline and serially over 6 months and once in healthy controls. Treatment outcomes were conversion of sputum culture results at 2 months and time to conversion of sputum microscopy results. RESULTS: Among 200 patients with pulmonary tuberculosis and 88 controls, FENO was lower for patients with pulmonary tuberculosis at diagnosis (geometric mean FENO, 12.7 parts per billion [ppb]; 95% confidence interval [CI], 11.6-13.8) than for controls (geometric mean FENO, 16.6 ppb; 95% CI, 14.2-19.5; P = .002), fell further after treatment initiation (nadir at 1 week), and then recovered by 6 months (P = .03). Lower FENO was associated with more-severe tuberculosis disease, with FENO directly proportional to weight (P < .001) and forced vital-capacity (P = .001) and inversely proportional to radiological score (P = .03). People whose FENO increased or remained unchanged by 2 months were 2.7-fold more likely to achieve conversion of sputum culture than those whose FENO decreased (odds ratio, 2.72; 95% CI, 1.05-7.12; P = .04). CONCLUSIONS: Among patients with pulmonary tuberculosis, impaired pulmonary NO bioavailability is associated with more-severe disease and delayed mycobacterial clearance. Measures to increase pulmonary NO warrant investigation as adjunctive tuberculosis treatments.

Ralph AP, Waramori G, Pontororing GJ, Kenangalem E, Wiguna A, Tjitra E, Sandjaja, Lolong DB, Yeo TW, Chatfield MD et al. 2013. L-arginine and vitamin D adjunctive therapies in pulmonary tuberculosis: a randomised, double-blind, placebo-controlled trial. PLoS One, 8 (8), pp. e70032. | Citations: 53 (Scopus) | Show Abstract | Read more

BACKGROUND: Vitamin D (vitD) and L-arginine have important antimycobacterial effects in humans. Adjunctive therapy with these agents has the potential to improve outcomes in active tuberculosis (TB). METHODS: In a 4-arm randomised, double-blind, placebo-controlled factorial trial in adults with smear-positive pulmonary tuberculosis (PTB) in Timika, Indonesia, we tested the effect of oral adjunctive vitD 50,000 IU 4-weekly or matching placebo, and L-arginine 6.0 g daily or matching placebo, for 8 weeks, on proportions of participants with negative 4-week sputum culture, and on an 8-week clinical score (weight, FEV1, cough, sputum, haemoptysis). All participants with available endpoints were included in analyses according to the study arm to which they were originally assigned. Adults with new smear-positive PTB were eligible. The trial was registered at ClinicalTrials.gov NCT00677339. RESULTS: 200 participants were enrolled, less than the intended sample size: 50 received L-arginine + active vitD, 49 received L-arginine + placebo vit D, 51 received placebo L-arginine + active vitD and 50 received placebo L-arginine + placebo vitD. According to the factorial model, 99 people received arginine, 101 placebo arginine, 101 vitamin D, 99 placebo vitamin D. Results for the primary endpoints were available in 155 (4-week culture) and 167 (clinical score) participants. Sputum culture conversion was achieved by week 4 in 48/76 (63%) participants in the active L-arginine versus 48/79 (61%) in placebo L-arginine arms (risk difference -3%, 95% CI -19 to 13%), and in 44/75 (59%) in the active vitD versus 52/80 (65%) in the placebo vitD arms (risk difference 7%, 95% CI -9 to 22%). The mean clinical outcome score also did not differ between study arms. There were no effects of the interventions on adverse event rates including hypercalcaemia, or other secondary outcomes. CONCLUSION: Neither vitD nor L-arginine supplementation, at the doses administered and with the power attained, affected TB outcomes. REGISTRY: ClinicalTrials.gov. Registry number: NCT00677339.

Kulpeng W, Sornsrivichai V, Chongsuvivatwong V, Rattanavipapong W, Leelahavarong P, Cairns J, Lubell Y, Teerawattananon Y. 2013. Variation of health-related quality of life assessed by caregivers and patients affected by severe childhood infections. BMC Pediatr, 13 (1), pp. 122. | Citations: 16 (Scopus) | Show Abstract | Read more

BACKGROUND: The agreement between self-reported and proxy measures of health status in ill children is not well established. This study aimed to quantify the variation in health-related quality of life (HRQOL) derived from young patients and their carers using different instruments. METHODS: A hospital-based cross-sectional survey was conducted between August 2010 and March 2011. Children with meningitis, bacteremia, pneumonia, acute otitis media, hearing loss, chronic lung disease, epilepsy, mild mental retardation, severe mental retardation, and mental retardation combined with epilepsy, aged between five to 14 years in seven tertiary hospitals were selected for participation in this study. The Health Utilities Index Mark 2 (HUI2), and Mark 3 (HUI3), and the EuroQoL Descriptive System (EQ-5D) and Visual Analogue Scale (EQ-VAS) were applied to both paediatric patients (self-assessment) and caregivers (proxy-assessment). RESULTS: The EQ-5D scores were lowest for acute conditions such as meningitis, bacteremia, and pneumonia, whereas the HUI3 scores were lowest for most chronic conditions such as hearing loss and severe mental retardation. Comparing patient and proxy scores (n = 74), the EQ-5D exhibited high correlation (r = 0.77) while in the HUI2 and HUI3 patient and caregiver scores were moderately correlated (r = 0.58 and 0.67 respectively). The mean difference between self and proxy-assessment using the HUI2, HUI3, EQ-5D and EQ-VAS scores were 0.03, 0.05, -0.03 and -0.02, respectively. In hearing-impaired and chronic lung patients the self-rated HRQOL differed significantly from their caregivers. CONCLUSIONS: The use of caregivers as proxies for measuring HRQOL in young patients affected by pneumococcal infection and its sequelae should be employed with caution. Given the high correlation between instruments, each of the HRQOL instruments appears acceptable apart from the EQ-VAS which exhibited low correlation with the others.

Lim P, Dek D, Try V, Eastman RT, Chy S, Sreng S, Suon S, Mao S, Sopha C, Sam B et al. 2013. Ex vivo susceptibility of Plasmodium falciparum to antimalarial drugs in western, northern, and eastern Cambodia, 2011-2012: association with molecular markers. Antimicrob Agents Chemother, 57 (11), pp. 5277-5283. | Citations: 25 (Scopus) | Show Abstract | Read more

In 2008, dihydroartemisinin (DHA)-piperaquine (PPQ) became the first-line treatment for uncomplicated Plasmodium falciparum malaria in western Cambodia. Recent reports of increased treatment failure rates after DHA-PPQ therapy in this region suggest that parasite resistance to DHA, PPQ, or both is now adversely affecting treatment. While artemisinin (ART) resistance is established in western Cambodia, there is no evidence of PPQ resistance. To monitor for resistance to PPQ and other antimalarials, we measured drug susceptibilities for parasites collected in 2011 and 2012 from Pursat, Preah Vihear, and Ratanakiri, in western, northern, and eastern Cambodia, respectively. Using a SYBR green I fluorescence assay, we calculated the ex vivo 50% inhibitory concentrations (IC50s) of 310 parasites to six antimalarials: chloroquine (CQ), mefloquine (MQ), quinine (QN), PPQ, artesunate (ATS), and DHA. Geometric mean IC50s (GMIC50s) for all drugs (except PPQ) were significantly higher in Pursat and Preah Vihear than in Ratanakiri (P ≤ 0.001). An increased copy number of P. falciparum mdr1 (pfmdr1), an MQ resistance marker, was more prevalent in Pursat and Preah Vihear than in Ratanakiri and was associated with higher GMIC50s for MQ, QN, ATS, and DHA. An increased copy number of a chromosome 5 region (X5r), a candidate PPQ resistance marker, was detected in Pursat but was not associated with reduced susceptibility to PPQ. The ex vivo IC50 and pfmdr1 copy number are important tools in the surveillance of multidrug-resistant (MDR) parasites in Cambodia. While MDR P. falciparum is prevalent in western and northern Cambodia, there is no evidence for PPQ resistance, suggesting that DHA-PPQ treatment failures result mainly from ART resistance.

Thanapongpichat S, McGready R, Luxemburger C, Day NPJ, White NJ, Nosten F, Snounou G, Imwong M. 2013. Microsatellite genotyping of Plasmodium vivax infections and their relapses in pregnant and non-pregnant patients on the Thai-Myanmar border. Malar J, 12 (1), pp. 275. | Citations: 13 (Scopus) | Show Abstract | Read more

BACKGROUND: Plasmodium vivax infections in pregnancy are associated with low birth weight and anaemia. This parasites species is also characterised by relapses, erythrocytic infections initiated by the activation of the dormant liver stages, the hypnozoites, to mature. Genotyping of P. vivax using microsatellite markers has opened the way to comparative investigations of parasite populations. The aim of the study was to assess whether there were any differences between the parasites found in pregnant and non-pregnant patients, and/or between the admission infections and recurrent episodes during follow-up. METHODS: Blood samples were collected from 18 pregnant and 18 non-pregnant patients, who had at least two recurrent episodes during follow-up, that were recruited in two previous trials on the efficacy of chloroquine treatment of P. vivax infections on the Thai-Myanmar border. DNA was purified and the P. vivax populations genotyped with respect to eight polymorphic microsatellite markers. Analyses of the genetic diversity, multiplicity of infection (MOI), and a comparison of the genotypes in the samples from each patient were conducted. RESULTS: The P. vivax parasites present in the samples exhibited high genetic diversity (6 to 15 distinct allelic variants found for the 8 loci). Similar expected heterozygosity (He) values were obtained for isolates from pregnant (0.837) and non-pregnant patients (0.852). There were modest differences between the MOI values calculated for both admission and recurrence samples from the pregnant patients (2.00 and 2.05, respectively) and the equivalent samples from the non-pregnant patients (1.67 and 1.64, respectively). Furthermore, the mean number of distinct alleles enumerated in the admission samples from the pregnant (6.88) and non-pregnant (7.63) patients were significantly lower than that found in the corresponding recurrent episodes samples (9.25 and 9.63, respectively). CONCLUSIONS: The P. vivax populations circulating in inhabitants along the Thai-Myanmar border, an area of low malaria transmission, displayed high genetic diversity. A subtle increase in the multiplicity of P. vivax infections in pregnant patients suggests a higher susceptibility to infection. The higher allelic diversity in the relapse as compared to the admission samples in both patient groups is consistent with the hypothesis that a febrile episode promotes the activation of hypnozoites.

Pasaribu AP, Chokejindachai W, Sirivichayakul C, Tanomsing N, Chavez I, Tjitra E, Pasaribu S, Imwong M, White NJ, Dondorp AM. 2013. A randomized comparison of dihydroartemisinin-piperaquine and artesunate-amodiaquine combined with primaquine for radical treatment of vivax malaria in Sumatera, Indonesia. J Infect Dis, 208 (11), pp. 1906-1913. | Citations: 17 (Scopus) | Show Abstract | Read more

BACKGROUND: A high prevalence of chloroquine-resistant Plasmodium vivax in Indonesia has shifted first-line treatment to artemisinin-based combination therapies, combined with primaquine (PQ) for radical cure. Which combination is most effective and safe remains to be established. METHODS: We conducted a prospective open-label randomized comparison of 14 days of PQ (0.25 mg base/kg) plus either artesunate-amodiaquine (AAQ + PQ) or dihydroartemisinin-piperaquine (DHP + PQ) for the treatment of uncomplicated monoinfection P. vivax malaria in North Sumatera, Indonesia. Patients were randomized and treatments were given without prior testing for G6PD status. The primary outcome was parasitological failure at day 42. Patients were followed up to 1 year. RESULTS: Between December 2010 and April 2012, 331 patients were included. After treatment with AAQ + PQ, recurrent infection occurred in 0 of 167 patients within 42 days and in 15 of 130 (11.5%; 95% confidence interval [CI], 6.6%-18.3%) within a year. With DHP + PQ, this was 1 of 164 (0.6%; 95% CI, 0.01%-3.4%) and 13 of 143 (9.1%; 95% CI, 4.9%-15.0%), respectively (P > .2). Intravascular hemolysis occurred in 5 patients, of which 3 males were hemizygous for the G6PD-Mahidol mutation. Minor adverse events were more frequent with AAQ + PQ. CONCLUSIONS: In North Sumatera, Indonesia, AAQ and DHP, both combined with PQ, were effective for blood-stage parasite clearance of uncomplicated P. vivax malaria. Both treatments were safe, but DHP + PQ was better tolerated. CLINICAL TRIALS REGISTRATION: NCT01288820.

Tarning J, Kloprogge F, Dhorda M, Jullien V, Nosten F, White NJ, Guerin PJ, Piola P. 2013. Pharmacokinetic properties of artemether, dihydroartemisinin, lumefantrine, and quinine in pregnant women with uncomplicated plasmodium falciparum malaria in Uganda. Antimicrob Agents Chemother, 57 (10), pp. 5096-5103. | Citations: 26 (Scopus) | Show Abstract | Read more

Pregnancy alters the pharmacokinetic properties of many drugs used in the treatment of malaria, usually resulting in lower drug exposures. This increases the risks of treatment failure, adverse outcomes for the fetus, and the development of resistance. The pharmacokinetic properties of artemether and its principal metabolite dihydroartemisinin (n = 21), quinine (n = 21), and lumefantrine (n = 26) in pregnant Ugandan women were studied. Lumefantrine pharmacokinetics in a nonpregnant control group (n = 17) were also studied. Frequently sampled patient data were evaluated with noncompartmental analysis. No significant correlation was observed between estimated gestational age and artemether, dihydroartemisinin, lumefantrine, or quinine exposures. Artemether/dihydroartemisinin and quinine exposures were generally low in these pregnant women compared to values reported previously for nonpregnant patients. Median day 7 lumefantrine concentrations were 488 (range, 30.7 to 3,550) ng/ml in pregnant women compared to 720 (339 to 2,150) ng/ml in nonpregnant women (P = 0.128). There was no statistical difference in total lumefantrine exposure or maximum concentration. More studies with appropriate control groups in larger series are needed to characterize the degree to which pregnant women are underdosed with current antimalarial dosing regimens.

Dondorp AM, Ringwald P. 2013. Artemisinin resistance is a clear and present danger. Trends Parasitol, 29 (8), pp. 359-360. | Citations: 32 (Web of Science Lite) | Read more

Paris DH, Shelite TR, Day NP, Walker DH. 2013. Unresolved problems related to scrub typhus: a seriously neglected life-threatening disease. Am J Trop Med Hyg, 89 (2), pp. 301-307. | Citations: 90 (Scopus) | Read more

Dondorp AM, Ringwald P. 2013. Artemisinin resistance is a clear and present danger Trends in Parasitology, 29 (8), pp. 359-360. | Citations: 36 (Scopus) | Read more

Dubot-Pérès A, Vongphrachanh P, Denny J, Phetsouvanh R, Linthavong S, Sengkeopraseuth B, Khasing A, Xaythideth V, Moore CE, Vongsouvath M et al. 2013. An epidemic of dengue-1 in a remote village in rural Laos. PLoS Negl Trop Dis, 7 (8), pp. e2360. | Citations: 16 (Web of Science Lite) | Show Abstract | Read more

In the Lao PDR (Laos), urban dengue is an increasingly recognised public health problem. We describe a dengue-1 virus outbreak in a rural northwestern Lao forest village during the cool season of 2008. The isolated strain was genotypically "endemic" and not "sylvatic," belonging to the genotype 1, Asia 3 clade. Phylogenetic analyses of 37 other dengue-1 sequences from diverse areas of Laos between 2007 and 2010 showed that the geographic distribution of some strains remained focal overtime while others were dispersed throughout the country. Evidence that dengue viruses have broad circulation in the region, crossing country borders, was also obtained. Whether the outbreak arose from dengue importation from an urban centre into a dengue-naïve community or crossed into the village from a forest cycle is unknown. More epidemiological and entomological investigations are required to understand dengue epidemiology and the importance of rural and forest dengue dynamics in Laos.

Karkey A, Thompson CN, Tran Vu Thieu N, Dongol S, Le Thi Phuong T, Voong Vinh P, Arjyal A, Martin LB, Rondini S, Farrar JJ et al. 2013. Differential epidemiology of Salmonella Typhi and Paratyphi A in Kathmandu, Nepal: a matched case control investigation in a highly endemic enteric fever setting. PLoS Negl Trop Dis, 7 (8), pp. e2391. | Citations: 31 (European Pubmed Central) | Show Abstract | Read more

BACKGROUND: Enteric fever, a systemic infection caused by the bacteria Salmonella Typhi and Salmonella Paratyphi A, is endemic in Kathmandu, Nepal. Previous work identified proximity to poor quality water sources as a community-level risk for infection. Here, we sought to examine individual-level risk factors related to hygiene and sanitation to improve our understanding of the epidemiology of enteric fever in this setting. METHODOLOGY AND PRINCIPAL FINDINGS: A matched case-control analysis was performed through enrollment of 103 blood culture positive enteric fever patients and 294 afebrile community-based age and gender-matched controls. A detailed questionnaire was administered to both cases and controls and the association between enteric fever infection and potential exposures were examined through conditional logistic regression. Several behavioral practices were identified as protective against infection with enteric fever, including water storage and hygienic habits. Additionally, we found that exposures related to poor water and socioeconomic status are more influential in the risk of infection with S. Typhi, whereas food consumption habits and migration play more of a role in risk of S. Paratyphi A infection. CONCLUSIONS AND SIGNIFICANCE: Our work suggests that S. Typhi and S. Paratyphi A follow different routes of infection in this highly endemic setting and that sustained exposure to both serovars probably leads to the development of passive immunity. In the absence of a polyvalent vaccine against S. Typhi and S. Paratyphi A, we advocate better systems for water treatment and storage, improvements in the quality of street food, and vaccination with currently available S. Typhi vaccines.

Phommasone K, Paris DH, Anantatat T, Castonguay-Vanier J, Keomany S, Souvannasing P, Blacksell SD, Mayxay M, Newton PN. 2013. Concurrent Infection with murine typhus and scrub typhus in southern Laos--the mixed and the unmixed. PLoS Negl Trop Dis, 7 (8), pp. e2163. | Citations: 18 (Scopus) | Read more

Turner P, Turner C, Watthanaworawit W, Carrara V, Cicelia N, Deglise C, Phares C, Ortega L, Nosten F. 2013. Respiratory virus surveillance in hospitalised pneumonia patients on the Thailand-Myanmar border BMC Infectious Diseases, 13 (1), | Citations: 12 (Scopus) | Show Abstract | Read more

Background: Pneumonia is a significant cause of morbidity and mortality in the developing world. Viruses contribute significantly to pneumonia burden, although data for low-income and tropical countries are scarce. The aim of this laboratory-enhanced, hospital-based surveillance was to characterise the epidemiology of respiratory virus infections among refugees living on the Thailand-Myanmar border.Methods: Maela camp provides shelter for ~45,000 refugees. Inside the camp, a humanitarian organisation provides free hospital care in a 158-bed inpatient department (IPD). Between 1st April 2009 and 30th September 2011, all patients admitted to the IPD with a clinical diagnosis of pneumonia were invited to participate. Clinical symptoms and signs were recorded and a nasopharyngeal aspirate (NPA) collected. NPAs were tested for adenoviruses, human metapneumovirus (hMPV), influenza A & B, and RSV by PCR.Results: Seven hundred eight patient episodes (698 patients) diagnosed as pneumonia during the enhanced surveillance period were included in this analysis. The median patient age was 1 year (range: < 1-70), and 90.4% were aged < 5 years. At least one virus was detected in 53.7% (380/708) of episodes. Virus detection was more common in children aged < 5 years old (<1 year: OR 2.0, 95% CI 1.2-3.4, p = 0.01; 1-4 years: OR 1.4, 95% CI 0.8-2.3, p = 0.2). RSV was detected in 176/708 (24.9%); an adenovirus in 133/708 (18.8%); an influenza virus in 68/708 (9.6%); and hMPV in 33/708 (4.7%). Twenty-eight episodes of multiple viral infections were identified, most commonly adenovirus plus another virus. RSV was more likely to be detected in children <5 years (OR 12.3, 95% CI 3.0-50.8, p = 0.001) and influenza viruses in patients ≥5 years (OR 2.8, 95% CI 1.5-5.4, p = 0.002). IPD treatment was documented in 702/708 cases; all but one patient received antimicrobials, most commonly a beta-lactam (amoxicillin/ampicillin +/-gentamicin in 664/701, 94.7%).Conclusions: Viral nucleic acid was identified in the nasopharynx in half the patients admitted with clinically diagnosed pneumonia. Development of immunisations targeting common respiratory viruses is likely to reduce the incidence of pneumonia in children living refugee camps and similar settings. © 2013 Turner et al.; licensee BioMed Central Ltd.

Harris JP, Lopman BA, Cooper BS. 2013. Correction BMJ Open, 3 (8), pp. e003060corr1-e003060corr1. | Read more

Yeo TW, Lampah DA, Rooslamiati I, Gitawati R, Tjitra E, Kenangalem E, Price RN, Duffull SB, Anstey NM. 2013. A randomized pilot study of L-arginine infusion in severe falciparum malaria: preliminary safety, efficacy and pharmacokinetics. PLoS One, 8 (7), pp. e69587. | Citations: 24 (Web of Science Lite) | Show Abstract | Read more

BACKGROUND: Decreased nitric oxide (NO) and hypoargininemia are associated with severe falciparum malaria and may contribute to severe disease. Intravenous L-arginine increases endothelial NO in moderately-severe malaria (MSM) without adverse effects. The safety, efficacy and pharmacokinetics of L-arginine or other agents to improve NO bioavailability in severe malaria have not been assessed. METHODS: In an open-label pilot study of L-arginine in adults with severe malaria (ARGISM-1 Study), patients were randomized to 12 g L-arginine hydrochloride or saline over 8 hours together with intravenous artesunate. Vital signs, selected biochemical measures (including blood lactate and L-arginine) and endothelial NO bioavailability (using reactive hyperemia peripheral arterial tonometry [RH-PAT]) were assessed serially. Pharmacokinetic analyses of L-arginine concentrations were performed using NONMEM. RESULTS: Six patients received L-arginine and two saline infusions. There were no deaths in either group. There were no changes in mean systolic (SBP) and diastolic blood pressure (DBP) or other vital signs with L-arginine, although a transient but clinically unimportant mean maximal decrease in SBP of 14 mmHg was noted. No significant changes in mean potassium, glucose, bicarbonate, or pH were seen, with transient mean maximal increases in plasma potassium of 0.3 mmol/L, and mean maximal decreases in blood glucose of 0.8 mmol/L and bicarbonate of 2.3 mEq/L following L-arginine administration. There was no effect on lactate clearance or RH-PAT index. Pharmacokinetic modelling (n = 4) showed L-arginine concentrations 40% lower than predicted from models developed in MSM. CONCLUSION: In the first clinical trial of an adjunctive treatment aimed at increasing NO bioavailability in severe malaria, L-arginine infused at 12 g over 8 hours was safe, but did not improve lactate clearance or endothelial NO bioavailability. Future studies may require increased doses of L-arginine. TRIAL REGISTRATION: ClinicalTrials.gov NCT00616304.

Simpson JA, Jamsen KM, Anderson TJC, Zaloumis S, Nair S, Woodrow C, White NJ, Nosten F, Price RN. 2013. Nonlinear mixed-effects modelling of in vitro drug susceptibility and molecular correlates of multidrug resistant Plasmodium falciparum. PLoS One, 8 (7), pp. e69505. | Citations: 4 (Scopus) | Show Abstract | Read more

The analysis of in vitro anti-malarial drug susceptibility testing is vulnerable to the effects of different statistical approaches and selection biases. These confounding factors were assessed with respect to pfmdr1 gene mutation and amplification in 490 clinical isolates. Two statistical approaches for estimating the drug concentration associated with 50% effect (EC50 ) were compared: the commonly used standard two-stage (STS) method, and nonlinear mixed-effects modelling. The in vitro concentration-effect relationships for, chloroquine, mefloquine, lumefantrine and artesunate, were derived from clinical isolates obtained from patients on the western border of Thailand. All isolates were genotyped for polymorphisms in the pfmdr1 gene. The EC50 estimates were similar for the two statistical approaches but 15-28% of isolates in the STS method had a high coefficient of variation (>15%) for individual estimates of EC50 and these isolates had EC50 values that were 32 to 66% higher than isolates derived with more precision. In total 41% (202/490) of isolates had amplification of pfmdr1 and single nucleotide polymorphisms were found in 50 (10%). Pfmdr1 amplification was associated with an increase in EC50 for mefloquine (139% relative increase in EC50 for 2 copies, 188% for 3+ copies), lumefantrine (82% and 75% for 2 and 3+ copies respectively) and artesunate (63% and 127% for 2 and 3+ copies respectively). In contrast pfmdr1 mutation at codons 86 or 1042 were associated with an increase in chloroquine EC50 (44-48%). Sample size calculations showed that to demonstrate an EC50 shift of 50% or more with 80% power if the prevalence was 10% would require 430 isolates and 245 isolates if the prevalence was 20%. In conclusion, although nonlinear mixed-effects modelling did not demonstrate any major advantage for determining estimates of anti-malarial drug susceptibility, the method includes all isolates, thereby, potentially improving confirmation of candidate molecular markers of anti-malarial drug susceptibility.

Russell B, Malleret B, Suwanarusk R, Anthony C, Kanlaya S, Lau YL, Woodrow CJ, Nosten F, Renia L. 2013. Field-based flow cytometry for ex vivo characterization of Plasmodium vivax and P. falciparum antimalarial sensitivity. Antimicrob Agents Chemother, 57 (10), pp. 5170-5174. | Citations: 13 (Web of Science Lite) | Show Abstract | Read more

Ex vivo antimalarial sensitivity testing in human malaria parasites has largely depended on microscopic determination of schizont maturation. While this microscopic method is sensitive, it suffers from poor precision and is laborious. The recent development of portable, low-cost cytometers has allowed us to develop and validate a simple, field-optimized protocol using SYBR green and dihydroethidium for the accurate and objective determination of antimalarial drug sensitivity in freshly isolated Plasmodium vivax and Plasmodium falciparum.

Yeo TW, Lampah DA, Kenangalem E, Tjitra E, Price RN, Anstey NM. 2013. Increased carboxyhemoglobin in adult falciparum malaria is associated with disease severity and mortality. J Infect Dis, 208 (5), pp. 813-817. | Citations: 6 (Scopus) | Show Abstract | Read more

Heme oxygenase 1 expression is increased in pediatric patients with malaria. The carboxyhemoglobin level (a measure of heme oxygenase 1 activity) has not been assessed in adult patients with malaria. Results of pulse co-oximetry revealed that the mean carboxyhemoglobin level was elevated in 29 Indonesian adults with severe falciparum malaria (10%; 95% confidence interval [CI], 8%-13%) and in 20 with severe sepsis (8%; 95% CI, 5%-12%), compared with the mean levels in 32 patients with moderately severe malaria (7%; 95% CI, 5%-8%) and 36 controls (3.6%; 95% CI, 3%-5%; P < .001). An increased carboxyhemoglobin level was associated with an increased odds of death among patients with severe malaria (odds ratio, 1.2 per percentage point increase; 95% CI, 1.02-1.5). While also associated with severity and fatality, methemoglobin was only modestly increased in patients with severe malaria. Increased carboxyhemoglobin levels during severe malaria and sepsis may exacerbate organ dysfunction by reducing oxygen carriage and cautions against the use of adjunctive CO therapy, which was proposed on the basis of mouse models.

Newton PN, Stepniewska K, Dondorp A, Silamut K, Chierakul W, Krishna S, Davis TME, Suputtamongkol Y, Angus B, Pukrittayakamee S et al. 2013. Prognostic indicators in adults hospitalized with falciparum malaria in Western Thailand. Malar J, 12 (1), pp. 229. | Citations: 17 (Scopus) | Show Abstract | Read more

BACKGROUND: Severe malaria remains a major cause of death and morbidity amongst adults in the Asiatic tropics. METHODS: A retrospective analysis of the clinical and laboratory data of 988 adult patients, hospitalized with Plasmodium falciparum malaria and prospectively recruited to malaria studies in western Thailand between 1986 and 2002, was performed to assess the factors associated with a fatal outcome. Different severity scores and classifications for defining severe malaria were compared and, using multiple logistic regression, simple models for predicting mortality developed. RESULTS: The proportion of patients fulfilling the WHO 2000 definition of severe malaria was 78.1%, and their mortality was 10%. Mortality in patients given parenteral artesunate or artemether (16/317, 5%) was lower than in those given parenteral quinine (59/442, 13%) (P < 0.001). Models using parameter sets based on WHO 1990, 2000 and Adapted AQ criteria plus blood smear parasite-stage assessment gave the best mortality prediction. A malaria prognostic index (MPI), derived from the dataset using five clinical or laboratory variables gave similar prognostic accuracy. CONCLUSIONS: The mortality of severe malaria in adults has fallen and the switch from quinine to artesunate has probably been an important contributor. Prognostic indices based on WHO 2000 definitions, and other simpler indices based on fewer variables, provide clinically useful predictions of outcome in Asian adults with severe malaria.

Turner P, Turner C, Jankhot A, Phakaudom K, Nosten F, Goldblatt D. 2013. Field evaluation of culture plus latex sweep serotyping for detection of multiple pneumococcal serotype colonisation in infants and young children. PLoS One, 8 (7), pp. e67933. | Citations: 7 (Scopus) | Show Abstract | Read more

BACKGROUND: Nasopharyngeal swab (NPS) culture by World Health Organisation (WHO) methodology underestimates multiple pneumococcal serotype colonisation compared to a simple culture and latex sweep method. The impacts of this on descriptions of pneumococcal serotype distributions and colonisation dynamics in infancy are not clear. METHODS: 8,736 NPS collected from infants enrolled into a longitudinal study were processed to evaluate the field utility of the latex sweep method. 1,107 had previously been cultured by WHO methodology. Additionally, colonisation results were compared in 100 matched pairs of infants, where swabs from an individual were cultured either by WHO or latex sweep method. RESULTS: In 1,107 swabs cultured by both methods, the latex sweep method was three times more likely to detect colonisation with multiple pneumococcal serotypes than the WHO method (p<0.001). At least one common serotype was identified in 91.2% of swabs from which typeable pneumococci were detected by both methods. Agreement improved with increasing colonisation density (p = 0.03). Estimates of age at first pneumococcal acquisition and colonisation duration were not affected by culture/serotyping method. However, a greater number of serotype carriage episodes were detected in infants cultured by latex sweep (p = 0.03). The overall rate of non-vaccine type pneumococcal acquisition was also greater in infants cultured by latex sweep (p = 0.04). CONCLUSIONS: Latex sweep serotyping was feasible to perform on a large specimen collection. Multiple serotype colonisation detection was significantly improved compared with WHO methodology. However, use of the latex sweep method is unlikely to significantly alter colonisation study serotype distribution or colonisation dynamics results.

Vlek ALM, Cooper BS, Kypraios T, Cox A, Edgeworth JD, Auguet OT. 2013. Clustering of antimicrobial resistance outbreaks across bacterial species in the intensive care unit. Clin Infect Dis, 57 (1), pp. 65-76. | Citations: 11 (Scopus) | Show Abstract | Read more

BACKGROUND: There are frequent reports of intensive care unit (ICU) outbreaks due to transmission of particular antibiotic-resistant bacteria. Less is known about the burden of outbreaks of resistance due to horizontal transfer of mobile genetic elements between species. Moreover, the potential of existing statistical software as a preliminary means for detecting such events has never been assessed. This study uses a software package to determine the burden of species and resistance outbreaks in 2 adjacent ICUs and to look for evidence of clustering of resistance outbreaks consistent with interspecies transmission of resistance elements. METHODS: A retrospective analysis of data from 2 adjacent 15-bed adult ICUs between 2002 and 2009 was undertaken. Detection of bacterial species-groups and resistance outbreaks was conducted using SaTScan and WHONet-SaTScan software. Resampling and permutation methods were applied to investigate temporal clustering of outbreaks. RESULTS: Outbreaks occurred for 69% of bacterial species-groups (18/26), and resistance outbreaks were detected against 63% of antibiotics (10/16). Resistance outbreaks against 7 of 10 antibiotics were observed in multiple species-groups simultaneously and there was evidence of inter-species-group dependence for 4 of 7 antibiotics; background temporal changes in resistance did not explain the temporal aggregation of outbreaks in 3 of 7 antibiotics. CONCLUSIONS: Species outbreaks occurred for the majority of bacteria commonly identified in the ICU. There was evidence for frequent temporal clustering of resistance outbreaks consistent with interspecies transmission of resistance elements. Wider application of outbreak detection software combined with targeted sequencing of bacterial genomes is needed to understand the contribution of interspecies gene transfer to resistance emergence.

Stoesser N, Moore CE, Pocock JM, An KP, Emary K, Carter M, Sona S, Poda S, Day N, Kumar V, Parry CM. 2013. Pediatric bloodstream infections in Cambodia, 2007 to 2011. Pediatr Infect Dis J, 32 (7), pp. e272-e276. | Citations: 19 (Scopus) | Show Abstract | Read more

BACKGROUND: Pediatric bacterial bloodstream infections (BSIs) are a major cause of morbidity and mortality worldwide. Epidemiological data from resource-limited settings in southeast Asia, such as Cambodia, are sparse but have important implications for treatment and public health strategies. METHODS: We retrospectively investigated BSI in children at a pediatric hospital and its satellite clinic in Siem Reap, Cambodia, from January 1, 2007, to July 31, 2011. The range of bacterial pathogens and their antimicrobial susceptibility patterns were analyzed in conjunction with demographic, clinical and outcome data. RESULTS: Of 7682 blood cultures with results (99.9% of cultures taken), 606 (7.9%) episodes of BSI were identified in 588 children. The incidence of BSI increased from 14 to 50/1000 admissions (P < 0.001); this was associated with an increased sampling rate. Most BSI were community acquired (89.1%). Common pathogens included Salmonella Typhi (22.8% of all isolates), Staphylococcus aureus (12.2%), Streptococcus pneumoniae (10.0%), Klebsiella pneumoniae (6.4%) and Escherichia coli (6.3%). 21.5% of BSI were caused by a diverse group of uncommon organisms, the majority of which were environmental Gram-negative species. No Listeria monocytogenes or Group B streptococcal BSI were identified. Antimicrobial resistance, particularly among the Enterobacteriaceae, was common. Overall mortality was substantial (19.0%), higher in neonates (36.9%) and independently associated with meningitis/meningoencephalitis and K. pneumoniae infection. CONCLUSIONS: BSI is a common problem in Cambodian children attending hospital and associated with significant mortality. Further studies are needed to clarify the epidemiology of neonatal sepsis, the contribution of atypical organisms and the epidemiology of pneumococcal disease before the introduction of vaccine.

Anekthananon T, Pukrittayakamee S, Ratanasuwan W, Jittamala P, Werarak P, Charunwatthana P, Suwanagool S, Lawpoolsri S, Stepniewska K, Sapchookul P et al. 2013. Oseltamivir and inhaled zanamivir as influenza prophylaxis in Thai health workers: a randomized, double-blind, placebo-controlled safety trial over 16 weeks (vol 68, pg 697, 2013) JOURNAL OF ANTIMICROBIAL CHEMOTHERAPY, 68 (7), pp. 1695-1695. | Read more

Pratt B, Zion D, Lwin KM, Cheah PY, Nosten F, Loff B. 2013. Ancillary Care: From Theory to Practice in International Clinical Research. Public Health Ethics, 6 (2), pp. 154-169. | Citations: 11 (Web of Science Lite) | Show Abstract | Read more

How international research might contribute to justice in global health has not been substantively addressed by bioethics. This article describes how the provision of ancillary care can link international clinical research to the reduction of global health disparities. It identifies the ancillary care obligations supported by a theory of global justice, showing that Jennifer Ruger's health capability paradigm requires the delivery of ancillary care to trial participants for a limited subset of conditions that cause severe morbidity and mortality. Empirical research on the Shoklo Malaria Research Unit's (SMRU) vivax malaria treatment trial was then undertaken to demonstrate whether and how these obligations might be upheld in a resource-poor setting. Our findings show that fulfilment of the ancillary care obligations is feasible where there is commitment from chief investigators and funders and is strongly facilitated by SMRU's dual role as a research unit and medical non-governmental organization.

Harris JP, Lopman BA, Cooper BS, O'Brien SJ. 2013. Does spatial proximity drive norovirus transmission during outbreaks in hospitals? BMJ Open, 3 (7), pp. e003060-e003060. | Citations: 14 (Scopus) | Show Abstract | Read more

OBJECTIVE: To assess the role of spatial proximity, defined as patients sharing bays, in the spread of norovirus during outbreaks in hospitals. DESIGN: Enhanced surveillance of norovirus outbreaks between November 2009 and November 2011. METHODS: Data were gathered during 149 outbreaks of norovirus in hospital wards from five hospitals in two major cities in England serving a population of two million. We used the time between the first two cases of each outbreak to estimate the serial interval for norovirus in this setting. This distribution and dates of illness onset were used to calculate epidemic trees for each outbreak. We then used a permutation test to assess whether proximity, for all outbreaks, was more extreme than would be expected by chance under the null hypothesis that proximity was not associated with transmission risk. RESULTS: 65 outbreaks contained complete data on both onset dates and ward position. We estimated the serial interval to be 1.86 days (95% CI 1.6 to 2.2 days), and with this value found strong evidence to reject the null hypothesis that proximity was not significant (p<0.001). Sensitivity analysis using different values of the serial interval showed that there was evidence to reject the null hypothesis provided the assumed serial interval was less than 2.5 days. CONCLUSIONS: Our results provide evidence that patients occupying the same bay as patients with symptomatic norovirus infection are at an increased risk of becoming infected by these patients compared with patients elsewhere in the same ward.

Kager LM, Schouten M, Wiersinga WJ, de Boer JD, Lattenist LCW, Roelofs JJTH, Meijers JCM, Levi M, Dondorp AM, Esmon CT et al. 2013. Overexpression of the endothelial protein C receptor is detrimental during pneumonia-derived gram-negative sepsis (Melioidosis). PLoS Negl Trop Dis, 7 (7), pp. e2306. | Citations: 10 (Web of Science Lite) | Show Abstract | Read more

BACKGROUND: The endothelial protein C receptor (EPCR) enhances anticoagulation by accelerating activation of protein C to activated protein C (APC) and mediates anti-inflammatory effects by facilitating APC-mediated signaling via protease activated receptor-1. We studied the role of EPCR in the host response during pneumonia-derived sepsis instigated by Burkholderia (B.) pseudomallei, the causative agent of melioidosis, a common form of community-acquired Gram-negative (pneumo)sepsis in South-East Asia. METHODOLOGY/PRINCIPAL FINDINGS: Soluble EPCR was measured in plasma of patients with septic culture-proven melioidosis and healthy controls. Experimental melioidosis was induced by intranasal inoculation of B. pseudomallei in wild-type (WT) mice and mice with either EPCR-overexpression (Tie2-EPCR) or EPCR-deficiency (EPCR(-/-)). Mice were sacrificed after 24, 48 or 72 hours. Organs and plasma were harvested to measure colony forming units, cellular influxes, cytokine levels and coagulation parameters. Plasma EPCR-levels were higher in melioidosis patients than in healthy controls and associated with an increased mortality. Tie2-EPCR mice demonstrated enhanced bacterial growth and dissemination to distant organs during experimental melioidosis, accompanied by increased lung damage, neutrophil influx and cytokine production, and attenuated coagulation activation. EPCR(-/-) mice had an unremarkable response to B. pseudomallei infection as compared to WT mice, except for a difference in coagulation activation in plasma. CONCLUSION/SIGNIFICANCE: Increased EPCR-levels correlate with accelerated mortality in patients with melioidosis. In mice, transgenic overexpression of EPCR aggravates outcome during Gram-negative pneumonia-derived sepsis caused by B. pseudomallei, while endogenous EPCR does not impact on the host response. These results add to a better understanding of the regulation of coagulation during severe (pneumo)sepsis.

Turner C, Carrara V, Thien NAM, Paw NMK, Rijken M, McGready R, Nosten F. 2013. Changes in the body weight of term infants, born in the tropics, during the first seven days of life. BMC Pediatr, 13 (1), pp. 93. | Citations: 6 (European Pubmed Central) | Show Abstract | Read more

BACKGROUND: Identifying unwell neonates, particularly in the first week of life, is often subjective. If normal values are known, calculating the weight lost or gained from birth weight can be a useful adjunct in the evaluation of the health of a neonate. METHODS: Serial body weights of well, term, breast fed infants who were attending for routine follow up, were recorded at the Shoklo Malaria Research Unit clinic in Maela Camp for displaced persons on the Thailand Myanmar border. Newborn examination was routine. Weight loss, expressed as percent weight lost from birth weight, and weight gain, expressed as a velocity (g/kg/day), was calculated for the first seven days of life. The results from normal birth weight infants, low birth weight infants (<2.5 kg) and small for gestational age infants (SGA) were examined. RESULTS: In the first week of life there were no significant differences in weight gained or lost across the three study groups. The maximum weight lost was 4.4% (95% CI 4.1 - 4.6%), which occurred on day three. Weight gain ranged from 13 g/kg/day [95% CI 10 - 16] on day four to 18 g/kg/day [95% CI 15 - 20] on days six and seven. CONCLUSIONS: Use of these normal values for weight gain and loss, allows infants falling outside of the expected range (95% CI) to be easily identified and subsequently highlighted as needing further medical review.

Douglas NM, Simpson JA, Phyo AP, Siswantoro H, Hasugian AR, Kenangalem E, Poespoprodjo JR, Singhasivanon P, Anstey NM, White NJ et al. 2013. Gametocyte dynamics and the role of drugs in reducing the transmission potential of Plasmodium vivax. J Infect Dis, 208 (5), pp. 801-812. | Citations: 24 (Web of Science Lite) | Show Abstract | Read more

BACKGROUND: Designing interventions that will reduce transmission of vivax malaria requires knowledge of Plasmodium vivax gametocyte dynamics. METHODS: We analyzed data from a randomized controlled trial in northwestern Thailand and 2 trials in Papua, Indonesia, to identify and compare risk factors for vivax gametocytemia at enrollment and following treatment. RESULTS: A total of 492 patients with P. vivax infections from Thailand and 476 patients (162 with concurrent falciparum parasitemia) from Indonesia were evaluable. Also, 84.3% (415/492) and 66.6% (209/314) of patients with monoinfection were gametocytemic at enrollment, respectively. The ratio of gametocytemia to asexual parasitemia did not differ between acute and recurrent infections (P = .48 in Thailand, P = .08 in Indonesia). High asexual parasitemia was associated with an increased risk of gametocytemia during follow-up in both locations. In Thailand, the cumulative incidence of gametocytemia between day 7 and day 42 following dihydroartemisinin + piperaquine (DHA + PIP) was 6.92% vs 29.1% following chloroquine (P < .001). In Indonesia, the incidence of gametocytemia was 33.6% following artesunate + amodiaquine (AS + AQ), 7.42% following artemether + lumefantrine, and 6.80% following DHA + PIP (P < .001 for DHA + PIP vs AS + AQ). CONCLUSIONS: P. vivax gametocyte carriage mirrors asexual-stage infection. Prevention of relapses, particularly in those with high asexual parasitemia, is likely the most important strategy for interrupting P. vivax transmission.

South East Asia Infectious Disease Clinical Research Network. 2013. Effect of double dose oseltamivir on clinical and virological outcomes in children and adults admitted to hospital with severe influenza: double blind randomised controlled trial. BMJ, 346 (may30 2), pp. f3039. | Citations: 58 (Scopus) | Show Abstract | Read more

OBJECTIVE: To investigate the validity of recommendations in treatment guidelines to use higher than approved doses of oseltamivir in patients with severe influenza. DESIGN: Double blind randomised trial. SETTING: Thirteen hospitals in Indonesia, Singapore, Thailand, and Vietnam. PARTICIPANTS: Patients aged ≥ 1 year admitted to hospital with confirmed severe influenza. INTERVENTIONS: Oral oseltamivir at double dose (150 mg twice a day/paediatric equivalent) versus standard dose (75 mg twice a day/paediatric equivalent). MAIN OUTCOME MEASURE: Viral status according to reverse transcriptase polymerase chain reaction (RT-PCR) for influenza RNA in nasal and throat swabs on day five. RESULTS: Of 326 patients (including 246 (75.5%) children aged <15), 165 and 161 were randomised to double or standard dose oseltamivir, respectively. Of these, 260 (79.8%) were infected with influenza virus A (133 (40.8%) with A/H3N2, 72 (22.1%) with A/H1N1-pdm09, 38 (11.7%) with seasonal A/H1N1, 17 (5.2%) with A/H5N1) and 53 (16.2%) with influenza virus B. A further 3.9% (13) were false positive by rapid antigen test (negative by RT-PCR and no rise in convalescent haemagglutination inhibition titers). Similar proportions of patients were negative for RT-PCR on day five of treatment: 115/159 (72.3%, 95% confidence interval 64.9% to 78.7%) double dose recipients versus 105/154 (68.2%, 60.5% to 75.0%) standard dose recipients; difference 4.2% (-5.9 to 14.2); P=0.42. No differences were found in clearance of virus in subgroup analyses by virus type/subtype, age, and duration of illness before randomisation. Mortality was similar: 12/165 (7.3%, 4.2% to 12.3%) in double dose recipients versus 9/161 (5.6%, 3.0% to 10.3%) in standard dose recipients. No differences were found between double and standard dose arms in median days on supplemental oxygen (3 (interquartile range 2-5) v 3.5 (2-7)), in intensive care (4.5 (3-6) v 5 (2-11), and on mechanical ventilation (2.5 (1-16) v 8 (1-16)), respectively. No important differences in tolerability were found. CONCLUSIONS: There were no virological or clinical advantages with double dose oseltamivir compared with standard dose in patients with severe influenza admitted to hospital. REGISTRATION: Clinical Trials NCT00298233.

Watthanaworawit W, Turner P, Turner C, Tanganuchitcharnchai A, Richards AL, Bourzac KM, Blacksell SD, Nosten F. 2013. A prospective evaluation of real-time PCR assays for the detection of Orientia tsutsugamushi and Rickettsia spp. for early diagnosis of rickettsial infections during the acute phase of undifferentiated febrile illness. Am J Trop Med Hyg, 89 (2), pp. 308-310. | Citations: 22 (Scopus) | Show Abstract | Read more

One hundred and eighty febrile patients were analyzed in a prospective evaluation of Orientia tsutsugamushi and Rickettsia spp. real-time polymerase chain reaction (PCR) assays for early diagnosis of rickettsial infections. By paired serology, 3.9% (7 of 180) and 6.1% (11 of 180) of patients were confirmed to have acute scrub or murine typhus, respectively. The PCR assays for the detection of O. tsutsugamushi and Rickettsia spp. had high specificity (99.4% [95% confidence interval (CI): 96.8-100] and 100% [95% CI: 97.8-100], respectively). The PCR results were also compared with immunoglobulin M (IgM) immunofluorescence assay (IFA) on acute sera. For O. tsutsugamushi, PCR sensitivity was twice that of acute specimen IgM IFA (28.6% versus 14.3%; McNemar's P = 0.3). For Rickettsia spp., PCR was four times as sensitive as acute specimen IgM IFA (36.4% versus 9.1%; P = 0.08), although this was not statistically significant. Whole blood and buffy coat, but not serum, were acceptable specimens for these PCRs. Further evaluation of these assays in a larger prospective study is warranted.

Katangwe T, Purcell J, Bar-Zeev N, Denis B, Montgomery J, Alaerts M, Heyderman RS, Dance DAB, Kennedy N, Feasey N, Moxon CA. 2013. Human melioidosis, Malawi, 2011. Emerg Infect Dis, 19 (6), pp. 981-984. | Citations: 19 (Web of Science Lite) | Show Abstract | Read more

A case of human melioidosis caused by a novel sequence type of Burkholderia pseudomallei occurred in a child in Malawi, southern Africa. A literature review showed that human cases reported from the continent have been increasing.

Cooper B, Brun-Buisson C, Bonten M. 2013. Daily chlorhexidine bathing and hospital-acquired infection. N Engl J Med, 368 (24), pp. 2330-2331. | Citations: 16 (Web of Science Lite) | Read more

Worby CJ, Jeyaratnam D, Robotham JV, Kypraios T, O'Neill PD, De Angelis D, French G, Cooper BS. 2013. Estimating the effectiveness of isolation and decolonization measures in reducing transmission of methicillin-resistant Staphylococcus aureus in hospital general wards. Am J Epidemiol, 177 (11), pp. 1306-1313. | Citations: 23 (Scopus) | Show Abstract | Read more

Infection control for hospital pathogens such as methicillin-resistant Staphylococcus aureus (MRSA) often takes the form of a package of interventions, including the use of patient isolation and decolonization treatment. Such interventions, though widely used, have generated controversy because of their significant resource implications and the lack of robust evidence with regard to their effectiveness at reducing transmission. The aim of this study was to estimate the effectiveness of isolation and decolonization measures in reducing MRSA transmission in hospital general wards. Prospectively collected MRSA surveillance data from 10 general wards at Guy's and St. Thomas' hospitals, London, United Kingdom, in 2006-2007 were used, comprising 14,035 patient episodes. Data were analyzed with a Markov chain Monte Carlo algorithm to model transmission dynamics. The combined effect of isolation and decolonization was estimated to reduce transmission by 64% (95% confidence interval: 37, 79). Undetected MRSA-positive patients were estimated to be the source of 75% (95% confidence interval: 67, 86) of total transmission events. Isolation measures combined with decolonization treatment were strongly associated with a reduction in MRSA transmission in hospital general wards. These findings provide support for active methods of MRSA control, but further research is needed to determine the relative importance of isolation and decolonization in preventing transmission.

Choosakulchart P, Kittisopee T, Takdhada S, Lubell Y, Robinson J. 2013. Cost-utility evaluation of influenza vaccination patients with existing coronary heart diseases in Thailand ASIAN BIOMEDICINE, 7 (3), pp. 425-435. | Citations: 1 (Web of Science Lite) | Show Abstract | Read more

Background: Influenza can exacerbate chronic coronary heart diseases (CHD) and health policy recommends influenza vaccination in this population group. However, cost effectiveness of influenza vaccination in protecting CHD population has not been, to our knowledge, well studied before especially in CHD patients with different disease severities. Objectives: To assess life-time cost utility of influenza vaccination in CHD patients either with angina and/or cardiac arrest/myocardial infarction (CA/MI) and to identify the most cost-effective influenza vaccination strategies. Method: The Markov model of CHD progression concurrent with the influenza infection was developed to quantify life-time costs and health effects of the three influenza vaccination strategies compared with no influenza vaccination (base case): (1) influenza vaccination in all CHD patients, (2) influenza vaccination in CA/MI patients-only, and (3) influenza vaccination in angina patients-only. The cost-effectiveness analysis (CEA) was based on the societal perspective. Deterministic and probabilistic sensitivity analyses were performed to identify variables that influence the sensitivity of the results and examine the effects of model parameters uncertainty, respectively. Results: For the base case, the expected value (EV) results of no influenza vaccination, influenza vaccination in all CHD groups, influenza vaccination in angina patients, and influenza vaccination in CA/MI are 346,437 Thai baht (THB) yielded 18.26 Quality adjusted life year (QALYs), 454,664 THB yielded 21.46 QALYs, 360,786 THB yielded 19.96 QALYs, and 437,901 THB yielded 19.72 QALYs; respectively. CEA graph comparing all influenza vaccination strategies shows that vaccination in all CHD patients groups and angina patients are in the costeffectiveness frontier, but not influenza vaccination in CA/MI patients. The cost-effectiveness rankings report shows that the willingness-to-pay (WTP) threshold (100,000 THB) is greater than the incremental cost effectiveness ratio (ICER) of vaccination in all CHD groups (ICER = 33,813 THB per QALY gained) and angina group (8,420 THB per QALY gained) and therefore the vaccination in all CHD groups, which is more expensive, but more effective would be recommended. The deterministic sensitivity analysis shows the most influential parameters driving the cost-effectiveness of vaccination strategies are the effect of influenza vaccination on CHD both for acute myocardial infarction and cardiovascular death, respectively. The probabilistic sensitivity analysis shows the same influenza strategy recommendation (vaccination in all CHD groups) as the base case analysis. Conclusion: From a societal perspective, influenza vaccination in all CHD groups is recommended. The information from economic modeling should be confirmed by primary economic research.

Climo MW, Wong ES. 2013. Daily chlorhexidine bathing and hospital-acquired infection. N Engl J Med, 368 (24), pp. 2332. | Citations: 3 (European Pubmed Central) | Read more

Turner P, Turner C, Green N, Ashton L, Lwe E, Jankhot A, Day NP, White NJ, Nosten F, Goldblatt D. 2013. Serum antibody responses to pneumococcal colonisation in the first two years of life: results from a SE Asian longitudinal cohort study. Clin Microbiol Infect, 19 (12), pp. E551-E558. | Citations: 27 (Web of Science Lite) | Show Abstract | Read more

Assessment of antibody responses to pneumococcal colonisation in early childhood may aid our understanding of protection and inform vaccine antigen selection. Serum samples were collected from mother-infant pairs during a longitudinal pneumococcal colonisation study in Burmese refugees. Maternal and cord sera were collected at birth and infants were bled monthly (1-24 months of age). Nasopharyngeal swabs were taken monthly to detect colonisation. Serum IgG titres to 27 pneumococcal protein antigens were measured in 2,624 sera and IgG to dominant serotypes (6B,14,19F,19A,23F) were quantified in 864 infant sera. Antibodies to all protein antigens were detectable in maternal sera. Titres to four proteins (LytB,PcpA,PhtD,PhtE) were significantly higher in mothers colonised by pneumococci at delivery. Maternally-derived antibodies to PiuA and Spr0096 were associated with delayed pneumococcal acquisition in infants in univariate, but not multivariate models. Controlling for infant age and previous homologous serotype exposure, nasopharyngeal acquisition of serotypes 19A, 23F, 14, or 19F were associated significantly with a ≥2-fold antibody response to the homologous capsule (OR 12.84, 7.52, 6.52, 5.33; p<0.05). Acquisition of pneumococcal serotypes in the nasopharynx of infants was not significantly associated with a ≥2-fold rise in antibodies to any of the protein antigens studied. In conclusion, nasopharyngeal colonisation in young children resulted in demonstrable serum IgG responses to pneumococcal capsules and surface/virulence proteins. However, the relationship between serum IgG and the prevention of, or response to, pneumococcal nasopharyngeal colonisation remains complex. Mechanisms other than serum IgG are likely to have a role but are currently poorly understood. This article is protected by copyright. All rights reserved.

Macareo L, Lwin KM, Cheah PY, Yuentrakul P, Miller RS, Nosten F. 2013. Triangular test design to evaluate tinidazole in the prevention of Plasmodium vivax relapse. Malar J, 12 (1), pp. 173. | Citations: 8 (Scopus) | Show Abstract | Read more

BACKGROUND: There are very few drugs that prevent the relapse of Plasmodium vivax malaria in man. Tinidazole is a 5-nitroimidazole approved in the USA for the treatment of indications including amoebiasis and giardiasis. In the non-human primate relapsing Plasmodium cynomolgi/macaque malaria model, tinidazole cured one of six macaques studied with an apparent mild delay to relapse in the other five of 14-28 days compared to 11-12 days in controls. One study has demonstrated activity against P. vivax in man. Presented here are the results of a pilot phase II, randomized, open-label study conducted along the Thai-Myanmar border designed to evaluate the efficacy of tinidazole to prevent relapse of P. vivax when administered with chloroquine. METHODS: This study utilized a modified triangular test sequential analysis which allows repeated statistical evaluation during the course of enrolment while maintaining a specified power and type 1 error and minimizing recruitment of subjects. Enrolment was to be halted when a pre-specified success/failure ratio was surpassed. The study was designed to have a 5% type 1 error and 90% power to show whether tinidazole would produce a relapse rate of less than 20% or greater than 45% through Day 63 of weekly follow-up after initiation of treatment and initial parasite clearance with 3 days of an oral weight based dosing of chloroquine and five days of 2 grams/day of tinidazole. RESULTS: All subjects cleared their parasitaemia by Day 3. Six of the first seven subjects treated with tinidazole relapsed prior to Day 63 (average Day 48.3 (range 42-56)). This exceeded the upper boundary of the triangular test and enrolment to receive tinidazole was halted. A concurrent cohort of five subjects definitively treated with standard doses of primaquine and chloroquine (historically 100% effective) showed no episodes of recurrent P. vivax parasitaemia during the 63-day protocol specified follow-up period. CONCLUSIONS: Tinidazole is ineffective in preventing relapse of P. vivax at the dose used. The macaque relapsing model appeared to correctly predict outcome in humans. Use of the modified triangular test allowed minimal enrolment and limited unnecessary exposure to the study drug and reduced costs. This adds weight to the ethical and economic advantages of this study design to evaluate similarly situated drugs. TRIAL REGISTRATION: ClinicalTrials.gov NCT00811096.

Tandhavanant S, Wongsuvan G, Wuthiekanun V, Teerawattanasook N, Day NPJ, Limmathurotsakul D, Peacock SJ, Chantratita N. 2013. Monoclonal antibody-based immunofluorescence microscopy for the rapid identification of Burkholderia pseudomallei in clinical specimens. Am J Trop Med Hyg, 89 (1), pp. 165-168. | Citations: 14 (European Pubmed Central) | Show Abstract | Read more

The diagnosis of melioidosis depends on the culture of Burkholderia pseudomallei, which takes at least 48 hours. We used a polyclonal-FITC-based immunofluorescence microscopic assay (Pab-IFA) on clinical samples to provide a rapid presumptive diagnosis. This has limitations including photobleaching and batch-to-batch variability. This study evaluated an IFA based on a monoclonal antibody specific to B. pseudomallei (Mab-IFA) and Alexa Fluor 488. A diagnostic evaluation was performed on a prospective cohort of 951 consecutive patients with suspected melioidosis. A total of 1,407 samples were tested. Test accuracy was defined against culture as the gold standard, and was also compared against Pab-IFA. A total of 88 samples from 64 patients were culture positive for B. pseudomallei. The diagnostic sensitivity and specificity of the Mab-IFA was comparable to the Pab-IFA (48.4% versus 45.3% for sensitivity, and 99.8% versus 98.8% for specificity). We have incorporated the Mab-IFA into our routine practice.

Liverani M, Waage J, Barnett T, Pfeiffer DU, Rushton J, Rudge JW, Loevinsohn ME, Scoones I, Smith RD, Cooper BS et al. 2013. Understanding and managing zoonotic risk in the new livestock industries. Environ Health Perspect, 121 (8), pp. 873-877. | Citations: 26 (Scopus) | Show Abstract | Read more

BACKGROUND: In many parts of the world, livestock production is undergoing a process of rapid intensification. The health implications of this development are uncertain. Intensification creates cheaper products, allowing more people to access animal-based foods. However, some practices associated with intensification may contribute to zoonotic disease emergence and spread: for example, the sustained use of antibiotics, concentration of animals in confined units, and long distances and frequent movement of livestock. OBJECTIVES: Here we present the diverse range of ecological, biological, and socioeconomic factors likely to enhance or reduce zoonotic risk, and identify ways in which a comprehensive risk analysis may be conducted by using an interdisciplinary approach. We also offer a conceptual framework to guide systematic research on this problem. DISCUSSION: We recommend that interdisciplinary work on zoonotic risk should take into account the complexity of risk environments, rather than limiting studies to simple linear causal relations between risk drivers and disease emergence and/or spread. In addition, interdisciplinary integration is needed at different levels of analysis, from the study of risk environments to the identification of policy options for risk management. CONCLUSION: Given rapid changes in livestock production systems and their potential health implications at the local and global level, the problem we analyze here is of great importance for environmental health and development. Although we offer a systematic interdisciplinary approach to understand and address these implications, we recognize that further research is needed to clarify methodological and practical questions arising from the integration of the natural and social sciences.

Nacher M, Guérin PJ, Demar-Pierre M, Djossou F, Nosten F, Carme B. 2013. Made in Europe: will artemisinin resistance emerge in French Guiana? Malar J, 12 (1), pp. 152. | Citations: 11 (Web of Science Lite) | Show Abstract | Read more

Resistance to artemisinin casts a shadow on the fight against malaria. The importance of illegal gold miners and of malaria in isolated regions of French Guiana constitutes a threat that endangers the fight against malaria in the Amazon. The hurdles of French laws and the remoteness of the territory from France make it impossible for the system to adapt to the problem of total inaccessibility of an important part of the malaria problem. Transmission is high in these areas and gold miners self-medicate with erratic regimens of artemisinin combinations, thus creating perfect conditions for the emergence of resistance. What needs to be done is being done, but within the limits of national law, with some results. However, facing the same difficult problem, Suriname shows more flexibility and is doing much better than French Guiana despite having lower resources. Local authorities in French Guiana cannot overrule the laws that block appropriate malaria care from reaching a third of malaria-exposed persons. Thus the health authorities in France should take immediate calibrated legislative and financial measures to avoid a predictable disaster.

Elliott I, Dittrich S, Paris D, Sengduanphachanh A, Phoumin P, Newton PN. 2013. The use of dried cerebrospinal fluid filter paper spots as a substrate for PCR diagnosis of the aetiology of bacterial meningitis in the Lao PDR. Clin Microbiol Infect, 19 (10), pp. E466-E472. | Citations: 8 (Web of Science Lite) | Show Abstract | Read more

We investigated whether dried cerebrospinal fluid (CSF) conserved on filter paper can be used as a substrate for accurate PCR diagnosis of important causes of bacterial meningitis in the Lao PDR. Using mock CSF, we investigated and optimized filter paper varieties, paper punch sizes, elution volumes and quantities of DNA template to achieve sensitive and reliable detection of bacterial DNA from filter paper specimens. FTA Elute Micro Card™ (Whatman, Maidstone, UK) was the most sensitive, consistent and practical variety of filter paper. Following optimization, the lower limit of detection for Streptococcus pneumoniae from dried mock CSF spots was 14 genomic equivalents (GE)/μL (interquartile range 5.5 GE/μL) or 230 (IQR 65) colony forming units/mL. A prospective clinical evaluation for S. pneumoniae, S. suis and Neisseria meningitidis was performed. Culture and PCR performed on fresh liquid CSF from patients admitted with a clinical diagnosis of meningitis (n = 73) were compared with results derived from dried CSF spots. Four of five fresh PCR-positive CSF samples also tested PCR positive from dried CSF spots, with one patient under the limit of detection. In a retrospective study of S. pneumoniae samples (n = 20), the median (IQR; range) CSF S. pneumoniae bacterial load was 1.1 × 10(4) GE/μL (1.2 × 10(5) ; 1 to 6.1 × 10(6) DNA GE/μL). Utilizing the optimized methodology, we estimate an extrapolated sensitivity of 90%, based on the range of CSF genome counts found in Laos. Dried CSF filter paper spots could potentially help us to better understand the epidemiology of bacterial meningitis in resource-poor settings and guide empirical treatments and vaccination policies.

Hendriksen ICE, Mtove G, Kent A, Gesase S, Reyburn H, Lemnge MM, Lindegardh N, Day NPJ, von Seidlein L, White NJ et al. 2013. Population pharmacokinetics of intramuscular artesunate in African children with severe malaria: implications for a practical dosing regimen. Clin Pharmacol Ther, 93 (5), pp. 443-450. | Citations: 19 (Scopus) | Show Abstract | Read more

Parenteral artesunate (ARS) is the drug of choice for the treatment of severe malaria. Pharmacokinetics data on intramuscular ARS are limited with respect to the main treatment group that carries the highest mortality, namely, critically ill children with severe malaria. A population pharmacokinetic study of ARS and dihydroartemisinin (DHA) was conducted from sparse sampling in 70 Tanzanian children of ages 6 months to 11 years. All the children had been admitted with severe falciparum malaria and were treated with intramuscular ARS (2.4 mg/kg at 0, 12, and 24 h). Venous plasma concentration-time profiles were characterized using nonlinear mixed-effects modeling (NONMEM). A one-compartment disposition model accurately described first-dose population pharmacokinetics of ARS and DHA. Body weight significantly affected clearance and apparent volume of distribution (P < 0.001), resulting in lower ARS and DHA exposure levels in smaller children. An adapted dosing regimen including a practical dosing table per weight band is proposed for young children based on the pharmacokinetic model.

Bright AT, Alenazi T, Shokoples S, Tarning J, Paganotti GM, White NJ, Houston S, Winzeler EA, Yanow SK. 2013. Genetic analysis of primaquine tolerance in a patient with relapsing vivax malaria. Emerg Infect Dis, 19 (5), pp. 802-805. | Citations: 16 (Scopus) | Show Abstract | Read more

Patients with Plasmodium vivax malaria are treated with primaquine to prevent relapse infections. We report primaquine failure in a patient with 3 relapses without any possibility of re-infection. Using whole genome sequencing of the relapsing parasite isolates, we identified single nucleotide variants as candidate molecular markers of resistance.

Siengsanan-Lamont J, Robertson ID, Blacksell SD, Ellis T, Saengchoowong S, Suwanpukdee S, Yongyuttawichai P, Cheewajorn K, Jangjaras J, Chaichoun K et al. 2013. A Study of Risk Factors for Infection with HPAI H5N1 in Small Poultry Farms in Thailand Using a Questionnaire Survey Zoonoses and Public Health, 60 (3), pp. 209-214. | Citations: 1 (Scopus) | Show Abstract | Read more

A questionnaire was used to collect data on small poultry farm management and wild bird observed in poultry keeping areas to identify putative risk factors for infection with HPAI H5N1. The study was conducted in 2008 in four subdistricts of central Thailand that had experienced outbreaks of HPAI H5N1 in poultry. Descriptive and inferential analyses including univariable analyses and multivariable logistic regression were used to identify putative risk factors. Risk factors included purchasing native chickens/fighting cocks from commercial hatcheries, replacing or restocking birds individually, and observing lesser whistling ducks (Dendrocygna javanica) on the farm daily. Selecting healthy animals when purchasing animals to ensure that they were disease free was a protective factor. To fully understand the epidemiology of infection of small poultry farms with HPAI H5N1, control of movement of domestic poultry and serological and virological testing of the poultry population should be applied. © 2012 Blackwell Verlag GmbH.

Miotto O, Almagro-Garcia J, Manske M, Macinnis B, Campino S, Rockett KA, Amaratunga C, Lim P, Suon S, Sreng S et al. 2013. Multiple populations of artemisinin-resistant Plasmodium falciparum in Cambodia. Nat Genet, 45 (6), pp. 648-655. | Citations: 247 (Scopus) | Show Abstract | Read more

We describe an analysis of genome variation in 825 P. falciparum samples from Asia and Africa that identifies an unusual pattern of parasite population structure at the epicenter of artemisinin resistance in western Cambodia. Within this relatively small geographic area, we have discovered several distinct but apparently sympatric parasite subpopulations with extremely high levels of genetic differentiation. Of particular interest are three subpopulations, all associated with clinical resistance to artemisinin, which have skewed allele frequency spectra and high levels of haplotype homozygosity, indicative of founder effects and recent population expansion. We provide a catalog of SNPs that show high levels of differentiation in the artemisinin-resistant subpopulations, including codon variants in transporter proteins and DNA mismatch repair proteins. These data provide a population-level genetic framework for investigating the biological origins of artemisinin resistance and for defining molecular markers to assist in its elimination.

Ch'ng J-H, Mok S, Bozdech Z, Lear MJ, Boudhar A, Russell B, Nosten F, Tan KS-W. 2013. A whole cell pathway screen reveals seven novel chemosensitizers to combat chloroquine resistant malaria. Sci Rep, 3 (1), pp. 1734. | Citations: 15 (Scopus) | Show Abstract | Read more

Due to the widespread prevalence of resistant parasites, chloroquine (CQ) was removed from front-line antimalarial chemotherapy in the 1990s despite its initial promise of disease eradication. Since then, resistance-conferring mutations have been identified in transporters such as the PfCRT, that allow for the efflux of CQ from its primary site of action, the parasite digestive vacuole. Chemosensitizing/chemoreversing compounds interfere with the function of these transporters thereby sensitizing parasites to CQ once again. However, compounds identified thus far have disappointing in vivo efficacy and screening for alternative candidates is required to revive this strategy. In this study, we propose a simple and direct means to rapidly screen for such compounds using a fluorescent-tagged CQ molecule. When this screen was applied to a small library, seven novel chemosensitizers (octoclothepin, methiothepin, metergoline, loperamide, chlorprothixene, L-703,606 and mibefradil) were quickly elucidated, including two which showed greater potency than the classical chemosensitizers verapamil and desipramine.

De Beaudrap P, Turyakira E, White LJ, Nabasumba C, Tumwebaze B, Muehlenbachs A, Guérin PJ, Boum Y, McGready R, Piola P. 2013. Impact of malaria during pregnancy on pregnancy outcomes in a Ugandan prospective cohort with intensive malaria screening and prompt treatment. Malar J, 12 (1), pp. 139. | Citations: 42 (European Pubmed Central) | Show Abstract | Read more

BACKGROUND: Malaria in pregnancy (MiP) is a major public health problem in endemic areas of sub-Saharan Africa and has important consequences on birth outcome. Because MiP is a complex phenomenon and malaria epidemiology is rapidly changing, additional evidence is still required to understand how best to control malaria. This study followed a prospective cohort of pregnant women who had access to intensive malaria screening and prompt treatment to identify factors associated with increased risk of MiP and to analyse how various characteristics of MiP affect delivery outcomes. METHODS: Between October 2006 and May 2009, 1,218 pregnant women were enrolled in a prospective cohort. After an initial assessment, they were screened weekly for malaria. At delivery, blood smears were obtained from the mother, placenta, cord and newborn. Multivariate analyses were performed to analyse the association between mothers' characteristics and malaria risk, as well as between MiP and birth outcome, length and weight at birth. This study is a secondary analysis of a trial registered with ClinicalTrials.gov, number NCT00495508. RESULTS: Overall, 288/1,069 (27%) mothers had 345 peripheral malaria infections. The risk of peripheral malaria was higher in mothers who were younger, infected with HIV, had less education, lived in rural areas or reported no bed net use, whereas the risk of placental infection was associated with more frequent malaria infections and with infection during late pregnancy. The risk of pre-term delivery and of miscarriage was increased in mothers infected with HIV, living in rural areas and with MiP occurring within two weeks of delivery.In adjusted analysis, birth weight but not length was reduced in babies of mothers exposed to MiP (-60 g, 95%CI: -120 to 0 for at least one infection and -150 g, 95%CI: -280 to -20 for >1 infections). CONCLUSIONS: In this study, the timing, parasitaemia level and number of peripherally-detected malaria infections, but not the presence of fever, were associated with adverse birth outcomes. Hence, prompt malaria detection and treatment should be offered to pregnant women regardless of symptoms or other preventive measures used during pregnancy, and with increased focus on mothers living in remote areas.

Woodrow CJ, Dahlström S, Cooksey R, Flegg JA, Le Nagard H, Mentré F, Murillo C, Ménard D, Nosten F, Sriprawat K et al. 2013. High-throughput analysis of antimalarial susceptibility data by the WorldWide Antimalarial Resistance Network (WWARN) in vitro analysis and reporting tool. Antimicrob Agents Chemother, 57 (7), pp. 3121-3130. | Citations: 22 (Scopus) | Show Abstract | Read more

Assessment of in vitro susceptibility is a fundamental component of antimalarial surveillance studies, but wide variations in the measurement of parasite growth and the calculation of inhibitory constants make comparisons of data from different laboratories difficult. Here we describe a Web-based, high-throughput in vitro analysis and reporting tool (IVART) generating inhibitory constants for large data sets. Fourteen primary data sets examining laboratory-determined susceptibility to artemisinin derivatives and artemisinin combination therapy partner drugs were collated from 11 laboratories. Drug concentrations associated with half-maximal inhibition of growth (IC50s) were determined by a modified sigmoid Emax model-fitting algorithm, allowing standardized analysis of 7,350 concentration-inhibition assays involving 1,592 isolates. Examination of concentration-inhibition data revealed evidence of apparent paradoxical growth at high concentrations of nonartemisinin drugs, supporting amendment of the method for calculating the maximal drug effect in each assay. Criteria for defining more-reliable IC50s based on estimated confidence intervals and growth ratios improved correlation coefficients for the drug pairs mefloquine-quinine and chloroquine-desethylamodiaquine in 9 of 11 and 8 of 8 data sets, respectively. Further analysis showed that maximal drug inhibition was higher for artemisinins than for other drugs, particularly in ELISA (enzyme-linked immunosorbent assay)-based assays, a finding consistent with the earlier onset of action of these drugs in the parasite life cycle. This is the first high-throughput analytical approach to apply consistent constraints and reliability criteria to large, diverse antimalarial susceptibility data sets. The data also illustrate the distinct biological properties of artemisinins and underline the need to apply more sensitive approaches to assessing in vitro susceptibility to these drugs.

Das D, Price RN, Bethell D, Guerin PJ, Stepniewska K. 2013. Early parasitological response following artemisinin-containing regimens: a critical review of the literature. Malar J, 12 (1), pp. 125. | Citations: 22 (Scopus) | Show Abstract | Read more

BACKGROUND: Parasitaemia on Day 3 has been proposed as a useful alert of potential artemisinin resistance, however, the normal variation of parasite clearance observed in artemisinin-based combination therapy clinical trials is poorly documented. METHODS: The trends in early parasitological response following treatment with an artemisinin anti-malarial regimen were reviewed. A PubMed literature search identified all studies using an artemisinin regimen for uncomplicated falciparum malaria published between January 2000 and December 2011. Data from clinical studies were extracted for analysis using a standardized questionnaire. RESULTS: In total 65,078 patients were enrolled into 213 clinical trials with 413 treatment arms containing either an artemisinin derivative alone (n=26) or in combination with a partner drug (n=387). The proportion of patients remaining parasitaemic at 24, 48 and 72 hours was documented in 115 (28%), 167 (40%) and 153 (37%) treatment arms, respectively. Excluding resistance studies in Cambodia, the median proportion of patients still parasitaemic was 53.8% [range 3-95, IQR=30.5-69.2] on Day 1, 6% [range 0-65.9, IQR=2-11.5] on Day 2 and 0 [range 0-12.6, IQR=0-2] on Day 3. Comparing studies from 2000 to 2005 and 2006 to 2011, the median proportion of patients reported to remain parasitaemic at 72 hours decreased in Africa (1.2% vs 0%, p=0.007), but increased in Asia (0.4% vs 3.9%, p=0.076). In 95% of studies the proportion of patients with peripheral parasitaemia was less than 6% at 72 hours. CONCLUSIONS: These results highlight the normal distribution of early parasitological responses following ACT, and the influence that heterogeneity in study design, host and parasite factors have in confounding a surveillance system based on Day 3 parasite positivity. Greater understanding of factors influencing parasite clearance is crucial, but will require analysis of pooled data from individual patient records.

Nkhoma SC, Stepniewska K, Nair S, Phyo AP, McGready R, Nosten F, Anderson TJC. 2013. Genetic evaluation of the performance of malaria parasite clearance rate metrics. J Infect Dis, 208 (2), pp. 346-350. | Citations: 9 (European Pubmed Central) | Show Abstract | Read more

Accurate measurement of malaria parasite clearance rates (CRs) following artemisinin (ART) treatment is critical for resistance surveillance and research, and various CR metrics are currently used. We measured 13 CR metrics in 1472 ART-treated hyperparasitemia infections for which 6-hour parasite counts and parasite genotypes (93 single nucleotide polymorphisms [SNPs]) were available. We used heritability to evaluate the performance of each metric. Heritability ranged from 0.06 ± 0.06 (SD) for 50% parasite clearance times to 0.67 ± 0.04 (SD) for clearance half-lives estimated from 6-hour parasite counts. These results identify the measures that should be avoided and show that reliable clearance measures can be obtained with abbreviated monitoring protocols.

Jiang J, Paris DH, Blacksell SD, Aukkanit N, Newton PN, Phetsouvanh R, Izzard L, Stenos J, Graves SR, Day NPJ, Richards AL. 2013. Diversity of the 47-kD HtrA nucleic acid and translated amino acid sequences from 17 recent human isolates of Orientia. Vector Borne Zoonotic Dis, 13 (6), pp. 367-375. | Citations: 18 (European Pubmed Central) | Show Abstract | Read more

Orientia tsutsugamushi, the etiologic agent of potentially fatal scrub typhus, is characterized by a high antigenic diversity, which complicates the development of a broadly protective vaccine. Efficacy studies in murine and nonhuman primate models demonstrated the DNA vaccine candidate pKarp47, based upon the O. tsutsugamushi Karp 47-kD HtrA protein gene, to be a successful immunoprophylactic against scrub typhus. To characterize 47-kD HtrA protein diversity among human isolates of Orientia, we sequenced the full open reading frame (ORF) of the 47-kD HtrA gene and analyzed the translated amino acid sequences of 17 patient isolates from Thailand (n=13), Laos (n=2), Australia (n=1), and the United Arab Emirates (UAE) (n=1) and 9 reference strains: Karp (New Guinea), Kato (Japan), Ikeda (Japan), Gilliam (Burma), Boryong (Korea), TA763, TH1811 and TH1817 (Thailand), and MAK243 (China). The percentage identity (similarity) of translated amino acid sequences between 16 new isolates and 9 reference strains of O. tsutsugamushi ranged from 96.4% to 100% (97.4% to 100%). However, inclusion of the recently identified Orientia chuto sp. nov. reduced identity (similarity) values to 82.2% to 83.3% (90.4% to 91.4%). These results demonstrate the diversity of Orientia 47-kD HtrA among isolates encountered by humans and therefore provide support for the necessity of developing a broadly protective scrub typhus vaccine that takes this diversity into account.

White NJ, Turner GDH, Day NPJ, Dondorp AM. 2013. Lethal malaria: Marchiafava and Bignami were right. J Infect Dis, 208 (2), pp. 192-198. | Citations: 53 (Scopus) | Show Abstract | Read more

One hundred and twenty years ago, the Italian malariologists Marchiafava and Bignami proposed that the fundamental pathological process underlying lethal falciparum malaria was microvascular obstruction. Since then, several alternative hypotheses have been proposed. These formed the basis for adjunctive interventions, which have either been ineffective or harmful. Recent evidence strongly suggests that Marchiafava and Bignami were right.

Chheng K, Carter MJ, Emary K, Chanpheaktra N, Moore CE, Stoesser N, Putchhat H, Sona S, Reaksmey S, Kitsutani P et al. 2013. A prospective study of the causes of febrile illness requiring hospitalization in children in Cambodia. PLoS One, 8 (4), pp. e60634. | Citations: 46 (Scopus) | Show Abstract | Read more

BACKGROUND: Febrile illnesses are pre-eminent contributors to morbidity and mortality among children in South-East Asia but the causes are poorly understood. We determined the causes of fever in children hospitalised in Siem Reap province, Cambodia. METHODS AND FINDINGS: A one-year prospective study of febrile children admitted to Angkor Hospital for Children, Siem Reap. Demographic, clinical, laboratory and outcome data were comprehensively analysed. Between October 12(th) 2009 and October 12(th) 2010 there were 1225 episodes of febrile illness in 1180 children. Median (IQR) age was 2.0 (0.8-6.4) years, with 850 (69%) episodes in children <5 years. Common microbiological diagnoses were dengue virus (16.2%), scrub typhus (7.8%), and Japanese encephalitis virus (5.8%). 76 (6.3%) episodes had culture-proven bloodstream infection, including Salmonella enterica serovar Typhi (22 isolates, 1.8%), Streptococcus pneumoniae (13, 1.1%), Escherichia coli (8, 0.7%), Haemophilus influenzae (7, 0.6%), Staphylococcus aureus (6, 0.5%) and Burkholderia pseudomallei (6, 0.5%). There were 69 deaths (5.6%), including those due to clinically diagnosed pneumonia (19), dengue virus (5), and melioidosis (4). 10 of 69 (14.5%) deaths were associated with culture-proven bloodstream infection in logistic regression analyses (odds ratio for mortality 3.4, 95% CI 1.6-6.9). Antimicrobial resistance was prevalent, particularly in S. enterica Typhi, (where 90% of isolates were resistant to ciprofloxacin, and 86% were multi-drug resistant). Comorbid undernutrition was present in 44% of episodes and a major risk factor for acute mortality (OR 2.1, 95% CI 1.1-4.2), as were HIV infection and cardiac disease. CONCLUSION: We identified a microbiological cause of fever in almost 50% of episodes in this large study of community-acquired febrile illness in hospitalized children in Cambodia. The range of pathogens, antimicrobial susceptibility, and co-morbidities associated with mortality described will be of use in the development of rational guidelines for infectious disease treatment and control in Cambodia and South-East Asia.

Lee W-C, Russell B, Lau Y-L, Fong M-Y, Chu C, Sriprawat K, Suwanarusk R, Nosten F, Renia L. 2013. Giemsa-stained wet mount based method for reticulocyte quantification: a viable alternative in resource limited or malaria endemic settings. PLoS One, 8 (4), pp. e60303. | Citations: 4 (Web of Science Lite) | Show Abstract | Read more

The quantity of circulating reticulocytes is an important indicator of erythropoietic activity in response to a wide range of haematological pathologies. While most modern laboratories use flow cytometry to quantify reticulocytes, most field laboratories still rely on 'subvital' staining. The specialist 'subvital' stains, New Methylene Blue (NMB) and Brilliant Crésyl Blue are often difficult to procure, toxic, and show inconsistencies between batches. Here we demonstrate the utility of Giemsa's stain (commonly used microbiology and parasitology) in a 'subvital' manner to provide an accurate method to visualize and count reticulocytes in blood samples from normal and malaria-infected individuals.

Vink JP, Laubscher M, Vlutters R, Silamut K, Maude RJ, Hasan MU, DE Haan G. 2013. An automatic vision-based malaria diagnosis system. J Microsc, 250 (3), pp. 166-178. | Citations: 15 (Scopus) | Show Abstract | Read more

Malaria is a worldwide health problem with 225 million infections each year. A fast and easy-to-use method, with high performance is required to differentiate malaria from non-malarial fevers. Manual examination of blood smears is currently the gold standard, but it is time-consuming, labour-intensive, requires skilled microscopists and the sensitivity of the method depends heavily on the skills of the microscopist. We propose an easy-to-use, quantitative cartridge-scanner system for vision-based malaria diagnosis, focusing on low malaria parasite densities. We have used special finger-prick cartridges filled with acridine orange to obtain a thin blood film and a dedicated scanner to image the cartridge. Using supervised learning, we have built a Plasmodium falciparum detector. A two-step approach was used to first segment potentially interesting areas, which are then analysed in more detail. The performance of the detector was validated using 5,420 manually annotated parasite images from malaria parasite culture in medium, as well as using 40 cartridges of 11,780 images containing healthy blood. From finger prick to result, the prototype cartridge-scanner system gave a quantitative diagnosis in 16 min, of which only 1 min required manual interaction of basic operations. It does not require a wet lab or a skilled operator and provides parasite images for manual review and quality control. In healthy samples, the image analysis part of the system achieved an overall specificity of 99.999978% at the level of (infected) red blood cells, resulting in at most seven false positives per microlitre. Furthermore, the system showed a sensitivity of 75% at the cell level, enabling the detection of low parasite densities in a fast and easy-to-use manner. A field trial in Chittagong (Bangladesh) indicated that future work should primarily focus on improving the filling process of the cartridge and the focus control part of the scanner.

Ramachandran S, Singhal M, McKenzie KG, Osborn JL, Arjyal A, Dongol S, Baker SG, Basnyat B, Farrar J, Dolecek C et al. 2013. A Rapid, Multiplexed, High-Throughput Flow-Through Membrane Immunoassay: A Convenient Alternative to ELISA. Diagnostics (Basel), 3 (2), pp. 244-260. | Citations: 1 (European Pubmed Central) | Show Abstract | Read more

This paper describes a rapid, high-throughput flow-through membrane immunoassay (FMIA) platform. A nitrocellulose membrane was spotted in an array format with multiple capture and control reagents for each sample detection area, and assay steps were carried out by sequential aspiration of sample and reagents through each detection area using a 96-well vacuum manifold. The FMIA provides an alternate assay format with several advantages over ELISA. The high surface area of the membrane permits high label concentration using gold labels, and the small pores and vacuum control provide rapid diffusion to reduce total assay time to ~30 min. All reagents used in the FMIA are compatible with dry storage without refrigeration. The results appear as colored spots on the membrane that can be quantified using a flatbed scanner. We demonstrate the platform for detection of IgM specific to lipopolysaccharides (LPS) derived from Salmonella Typhi. The FMIA format provides analytical results comparable to ELISA in less time, provides integrated assay controls, and allows compensation for specimen-to-specimen variability in background, which is a particular challenge for IgM assays.

Turner C, Mya Thein NA, Turner P, Nosten F, White NJ. 2013. Rectal pH in Well and Unwell Infants. J Trop Pediatr, 59 (2), pp. 162. | Read more

Hanson JP, Lam SWK, Mohanty S, Alam S, Pattnaik R, Mahanta KC, Hasan MU, Charunwatthana P, Mishra SK, Day NPJ et al. 2013. Fluid resuscitation of adults with severe falciparum malaria: Effects on acid-base status, renal function, and extravascular lung water Critical Care Medicine, 41 (4), pp. 972-981. | Citations: 38 (Scopus) | Show Abstract | Read more

Objective: To evaluate the efficacy and safety of liberal fluid resuscitation of adults with severe malaria. DESIGN, Setting, Patients, and Methods:: Twenty-eight Bangladeshi and Indian adults with severe falciparum malaria received crystalloid resuscitation guided by transpulmonary thermodilution (PiCCO) in an intensive care setting. Systemic hemodynamics, microvascular indices and measures of acidosis, renal function, and pulmonary edema were followed prospectively. Results: All patients were hypovolemic (global end-diastolic volume index < 680 mL/m) on enrollment. Patients received a median (range) 3230 mL (390-7300) of isotonic saline in the first 6 hours and 5450 mL (710-13,720) in the first 24 hours. With resuscitation, acid-base status deteriorated in 19 of 28 (68%), and there was no significant improvement in renal function. Extravascular lung water increased in 17 of 22 liberally resuscitated patients (77%); eight of these patients developed pulmonary edema, five of whom died. All other patients survived. All patients with pulmonary edema during the study were hypovolemic or euvolemic at the time pulmonary edema developed. Plasma lactate was lower in hypovolemic patients before (rs = 0.38; p = 0.05) and after (rs = 0.49; p = 0.01) resuscitation but was the strongest predictor of mortality before (chi-square = 9.9; p = 0.002) and after resuscitation (chi-square = 11.1; p < 0.001) and correlated with the degree of visualized microvascular sequestration of parasitized erythrocytes at both time points (rs = 0.55; p = 0.003 and rs = 0.43; p = 0.03, respectively). Persisting sequestration was evident in 7 of 15 patients (47%) 48 hours after enrollment. Conclusions: Lactic acidosis-the strongest prognostic indicator in adults with severe falciparum malaria-results from sequestration of parasitized erythrocytes in the microcirculation, not from hypovolemia. Liberal fluid resuscitation has little effect on this sequestration and does not improve acid-base status or renal function. Pulmonary edema-secondary to increased pulmonary vascular permeability-is common, unpredictable, and exacerbated by fluid loading. Liberal fluid replacement of adults with severe malaria should be avoided. © 2013 by the Society of Critical Care Medicine and Lippincott Williams & Wilkins.

Turner C, Thein NAM, Turner P, Nosten F, White NJ. 2013. Erratum to Rectal pH in well and unwell infants [J Trop Pediatr, 58 (2012) 311-313] Journal of Tropical Pediatrics, 59 (2), pp. 162. | Read more

Rekić D, Röshammar D, Bergstrand M, Tarning J, Calcagno A, D'Avolio A, Ormaasen V, Vigan M, Barrail-Tran A, Ashton M et al. 2013. External validation of the bilirubin-atazanavir nomogram for assessment of atazanavir plasma exposure in HIV-1-infected patients. AAPS J, 15 (2), pp. 308-315. | Citations: 2 (Web of Science Lite) | Show Abstract | Read more

Atazanavir increases plasma bilirubin levels in a concentration-dependent manner. Due to less costly and readily available assays, bilirubin has been proposed as a marker of atazanavir exposure. In this work, a previously developed nomogram for detection of suboptimal atazanavir exposure is validated against external patient populations. The bilirubin nomogram was validated against 311 matching bilirubin and atazanavir samples from 166 HIV-1-infected Norwegian, French, and Italian patients on a ritonavir-boosted regimen. In addition, the nomogram was evaluated in 56 Italian patients on an unboosted regimen. The predictive properties of the nomogram were validated against observed atazanavir plasma concentrations. The use of the nomogram to detect non-adherence was also investigated by simulation. The bilirubin nomogram predicted suboptimal exposure in the patient populations on a ritonavir-boosted regimen with a negative predictive value of 97% (95% CI 95-100). The bilirubin nomogram and monitoring of atazanavir concentrations had similar predictive properties for detecting non-adherence based on simulations. Although both methods performed adequately during a period of non-adherence, they had lower predictive power to detect past non-adherence episodes. Using the bilirubin nomogram for detection of suboptimal atazanavir exposure in patients on a ritonavir-boosted regimen is a rapid and cost-effective alternative to routine measurements of the actual atazanavir exposure in plasma. Its application may be useful in clinical settings if atazanavir concentrations are not available.

Majumder MI, Haque MM, Ahmed MW, Alam MN, Rahman MW, Akter F, Basher A, Maude RJ, Faiz MA. 2013. Melioidosis in an adult male. Mymensingh Med J, 22 (2), pp. 413-416. | Citations: 4 (European Pubmed Central) | Show Abstract

Infection with Burkholderia pseudomallei has been described, albeit rarely, patients in Bangladesh. Infection usually follows percutaneous inoculation or inhalation of the causative bacterium, which is present in soil and surface water in the endemic region. A 35-year-young male farmer presented with prolonged fever and significant weight loss. Patient gradually deteriorated despite getting different antibiotics including intravenous ceftriaxone and metronidazole. Panels of investigations were done which revealed no diagnostic confirmation except uncontrolled diabetes and multiple abscesses in different organs. Melioidosis was suspected and serum samples were positive for Burkholderia pseudomallei antibody. The case illustrates the importance of non-specific nature of the clinical presentation and high index of suspicion of uncommon diseases like melioidosis where the disease has not been considered as an endemic.

von Seidlein L, Auburn S, Espino F, Shanks D, Cheng Q, McCarthy J, Baird K, Moyes C, Howes R, Ménard D et al. 2013. Review of key knowledge gaps in glucose-6-phosphate dehydrogenase deficiency detection with regard to the safe clinical deployment of 8-aminoquinoline treatment regimens: a workshop report. Malar J, 12 (1), pp. 112. | Citations: 73 (Web of Science Lite) | Show Abstract | Read more

The diagnosis and management of glucose-6-phosphate dehydrogenase (G6PD) deficiency is a crucial aspect in the current phases of malaria control and elimination, which will require the wider use of 8-aminoquinolines for both reducing Plasmodium falciparum transmission and achieving the radical cure of Plasmodium vivax. 8-aminoquinolines, such as primaquine, can induce severe haemolysis in G6PD-deficient individuals, potentially creating significant morbidity and undermining confidence in 8-aminoquinoline prescription. On the other hand, erring on the side of safety and excluding large numbers of people with unconfirmed G6PD deficiency from treatment with 8-aminoquinolines will diminish the impact of these drugs. Estimating the remaining G6PD enzyme activity is the most direct, accessible, and reliable assessment of the phenotype and remains the gold standard for the diagnosis of patients who could be harmed by the administration of primaquine. Genotyping seems an unambiguous technique, but its use is limited by cost and the large range of recognized G6PD genotypes. A number of enzyme activity assays diagnose G6PD deficiency, but they require a cold chain, specialized equipment, and laboratory skills. These assays are impractical for care delivery where most patients with malaria live. Improvements to the diagnosis of G6PD deficiency are required for the broader and safer use of 8-aminoquinolines to kill hypnozoites, while lower doses of primaquine may be safely used to kill gametocytes without testing. The discussions and conclusions of a workshop conducted in Incheon, Korea in May 2012 to review key knowledge gaps in G6PD deficiency are reported here.

Eziefula AC, Staedke SG, Yeung S, Webb E, Kamya M, White NJ, Bousema T, Drakeley C. 2013. Study protocol for a randomised controlled double-blinded trial of the dose-dependent efficacy and safety of primaquine for clearance of gametocytes in children with uncomplicated falciparum malaria in Uganda. BMJ Open, 3 (3), pp. e002759-e002759. | Citations: 6 (Scopus) | Show Abstract | Read more

OBJECTIVES: For the purpose of blocking transmission of Plasmodium falciparum malaria from humans to mosquitoes, a single dose of primaquine is recommended by the WHO as an addition to artemisinin combination therapy. Primaquine clears gametocytes but causes dose-dependent haemolysis in individuals with glucose-6-phosphate dehydrogenase (G6PD) deficiency. Evidence is needed to inform the optimal dosing of primaquine for malaria elimination programmes and for the purpose of interrupting the spread of artemisinin-resistant malaria. This study investigates the efficacy and safety of reducing doses of primaquine for clearance of gametocytes in participants with normal G6PD status. METHODS AND ANALYSIS: In this prospective, four-armed randomised placebo-controlled double-blinded trial, children aged 1-10 years, weighing over 10 kg, with haemoglobin ≥8 g/dl and uncomplicated P falciparum malaria are treated with artemether lumefantrine and randomised to receive a dose of primaquine (0.1, 0.4 or 0.75 mg base/kg) or placebo on the third day of treatment. Participants are followed up for 28 days. Gametocytaemia is measured by quantitative nucleic acid sequence-based analysis on days 0, 2, 3, 7, 10 and 14 with a primary endpoint of the number of days to gametocyte clearance in each treatment arm and secondarily the area under the curve of gametocyte density over time. Analysis is for non-inferiority of efficacy compared to the reference dose, 0.75 mg base/kg. Safety is assessed by pair-wise comparisons of the arithmetic mean (±SD) change in haemoglobin concentration per treatment arm and analysed for superiority to placebo and incidence of adverse events. Ethics and dissemination Approval was obtained from the ethical committees of Makerere University School of Medicine, the Ugandan National Council of Science and Technology and the London School of Hygiene and Tropical Medicine. RESULTS: These will be disseminated to inform malaria elimination policy, through peer-reviewed publication and academic presentations.

Meeyai A, Cooper BS, Coker R. 2013. Analysis of 2009 pandemic influenza A/H1N1 outcomes in 19 European countries: association with completeness of national strategic plans. BMJ Open, 3 (3), pp. e002253-e002253. | Citations: 3 (Scopus) | Show Abstract | Read more

OBJECTIVE: To describe changes in reported influenza activity associated with the 2009 H1N1 pandemic in European countries and determine whether there is a correlation between these changes and completeness of national strategic pandemic preparedness. DESIGN: A retrospective correlational study. SETTING: Countries were included if their national strategic plans had previously been analysed and if weekly influenza-like illness (ILI) data from sentinel networks between week 21, 2006 and week 20, 2010 were more than 50% complete. OUTCOME MEASURES: For each country we calculated three outcomes: the percentage change in ILI peak height during the pandemic relative to the prepandemic mean; the timing of the ILI peak and the percentage change in total cases relative to the prepandemic mean. Correlations between these outcomes and completeness of a country's national strategic pandemic preparedness plan were assessed using the Pearson product-moment correlation coefficient. RESULTS: Nineteen countries were included. The ILI peak occurred earlier than the mean seasonal peak in 17 countries. In 14 countries the pandemic peak was higher than the seasonal peak, though the difference was large only in Norway, the UK and Greece. Nine countries experienced more total ILI cases during the pandemic compared with the mean for prepandemic years. Five countries experienced two distinct pandemic peaks. There was no clear pattern of correlation between overall completeness of national strategic plans and pandemic influenza outcome measures and no evidence of association between these outcomes and components of pandemic plans that might plausibly affect influenza outcomes (public health interventions, vaccination, antiviral use, public communication). Amongst the 17 countries with a clear pandemic peak, only the correlation between planning for essential services and change in total ILI cases significantly differed from zero: correlation coefficient (95% CI) 0.50 (0.02, 0.79). CONCLUSIONS: The diversity of pandemic influenza outcomes across Europe is not explained by the marked variation in the completeness of pandemic plans.

Nilsen A, LaCrue AN, White KL, Forquer IP, Cross RM, Marfurt J, Mather MW, Delves MJ, Shackleford DM, Saenz FE et al. 2013. Quinolone-3-diarylethers: a new class of antimalarial drug. Sci Transl Med, 5 (177), pp. 177ra37. | Citations: 107 (Scopus) | Show Abstract | Read more

The goal for developing new antimalarial drugs is to find a molecule that can target multiple stages of the parasite's life cycle, thus impacting prevention, treatment, and transmission of the disease. The 4(1H)-quinolone-3-diarylethers are selective potent inhibitors of the parasite's mitochondrial cytochrome bc1 complex. These compounds are highly active against the human malaria parasites Plasmodium falciparum and Plasmodium vivax. They target both the liver and blood stages of the parasite as well as the forms that are crucial for disease transmission, that is, the gametocytes, the zygote, the ookinete, and the oocyst. Selected as a preclinical candidate, ELQ-300 has good oral bioavailability at efficacious doses in mice, is metabolically stable, and is highly active in blocking transmission in rodent models of malaria. Given its predicted low dose in patients and its predicted long half-life, ELQ-300 has potential as a new drug for the treatment, prevention, and, ultimately, eradication of human malaria.

Instiaty I, Lindegardh N, Jittmala P, Hanpithakpong W, Blessborn D, Pukrittayakamee S, White NJ, Tarning J. 2013. Comparison of oseltamivir and oseltamivir carboxylate concentrations in venous plasma, venous blood, and capillary blood in healthy volunteers. Antimicrob Agents Chemother, 57 (6), pp. 2858-2862. | Citations: 5 (Scopus) | Show Abstract | Read more

Oseltamivir and oseltamivir carboxylate concentrations were measured in venous plasma, venous blood, and capillary blood taken simultaneously from 24 healthy volunteers. Median (range) venous-blood-to-plasma ratios were 1.42 (0.920 to 1.97) for oseltamivir and 0.673 (0.564 to 0.814) for oseltamivir carboxylate. Capillary blood/venous plasma ratios were 1.32 (0.737 to 3.16) for oseltamivir and 0.685 (0.502 to 1.34) for oseltamivir carboxylate. Oseltamivir concentrations in venous and capillary blood were similar. Oseltamivir carboxylate showed a time-dependent distribution between venous and capillary blood.

Awab GR, Pukrittayakamee S, Jamornthanyawat N, Yamin F, Dondorp AM, Day NP, White NJ, Woodrow CJ, Imwong M. 2013. Prevalence of antifolate resistance mutations in Plasmodium falciparum isolates in Afghanistan. Malar J, 12 (1), pp. 96. | Citations: 10 (Scopus) | Show Abstract | Read more

BACKGROUND: Artesunate plus sulphadoxine-pyrimethamine (AS+SP) is now first-line treatment for Plasmodium falciparum infection in several south Asian countries, including Afghanistan. Molecular studies provide a sensitive means to investigate the current state of drug susceptibility to the SP component, and can also provide information on the likely efficacy of other potential forms of artemisinin-combination therapy. METHODS: During the years 2007 to 2010, 120 blood spots from patients with P. falciparum malaria were obtained in four provinces of Afghanistan. PCR-based methods were used to detect drug-resistance mutations in dhfr, dhps, pfcrt and pfmdr1, as well as to determine copy number of pfmdr1. RESULTS: The majority (95.5%) of infections had a double mutation in the dhfr gene (C59R, S108N); no mutations at dhfr positions 16, 51 or 164 were seen. Most isolates were wild type across the dhps gene, but five isolates from the provinces of Kunar and Nangarhar in eastern Afghanistan had the triple mutation A437G / K540E / A581G; all five cases were successfully treated with three receiving AS+SP and two receiving dihydroartemisinin-piperaquine. All isolates showed the pfcrt SVNMT chloroquine resistance haplotype. Five of 79 isolates had the pfmdr1 N86Y mutation, while 52 had pfmdr1 Y184F; positions 1034, 1042 and 1246 were wild type in all isolates. The pfmdr1 gene was not amplified in any sample. CONCLUSIONS: This study indicates that shortly after the adoption of AS+SP as first-line treatment in Afghanistan, most parasites had a double mutation haplotype in dhfr, and a small number of isolates from eastern Afghanistan harboured a triple mutation haplotype in dhps. The impact of these mutations on the efficacy of AS+SP remains to be assessed in significant numbers of patients, but these results are clearly concerning since they suggest a higher degree of SP resistance than previously detected. Further focused molecular and clinical studies in this region are urgently required.

Boel ME, Rijken MJ, Leenstra T, Phyo AP, Pimanpanarak M, Keereecharoen NL, Proux S, Laochan N, Imwong M, Singhasivanon P et al. 2013. Malaria in the post-partum period; a prospective cohort study. PLoS One, 8 (3), pp. e57890. | Citations: 4 (Web of Science Lite) | Show Abstract | Read more

BACKGROUND: Several studies have shown a prolonged or increased susceptibility to malaria in the post-partum period. A matched cohort study was conducted to evaluate prospectively the susceptibility to malaria of post-partum women in an area where P.falciparum and P.vivax are prevalent. METHODS: In an area of low seasonal malaria transmission on the Thai-Myanmar border pregnant women attending antenatal clinics were matched to a non-pregnant, non-post-partum control and followed up prospectively until 12 weeks after delivery. RESULTS: Post-partum women (n = 744) experienced significantly less P.falciparum episodes than controls (hazard ratio (HR) 0.39 (95%CI 0.21-0.72) p = 0.003) but significantly more P.vivax (HR 1.34 (1.05-1.72) p = 0.018). The reduced risk of falciparum malaria was accounted for by reduced exposure, whereas a history of P.vivax infection during pregnancy was a strong risk factor for P.vivax in post-partum women (HR 13.98 (9.13-21.41), p<0.001). After controlling for effect modification by history of P.vivax, post-partum women were not more susceptible to P.vivax than controls (HR: 0.33 (0.21-0.51), p<0.001). Genotyping of pre-and post-partum infections [Formula in text] showed that each post-partum P.falciparum was a newly acquired infection. CONCLUSIONS: In this area of low seasonal malaria transmission post-partum women were less likely to develop falciparum malaria but more likely to develop vivax malaria than controls. This was explained by reduced risk of exposure and increased risk of relapse, respectively. There was no evidence for altered susceptibility to malaria in the post-partum period. The treatment of vivax malaria during and immediately after pregnancy needs to be improved.

Phetsouvanh R, Thojaikong T, Phoumin P, Sibounheuang B, Phommasone K, Chansamouth V, Lee SJ, Newton PN, Blacksell SD. 2013. Inter- and intra-operator variability in the reading of indirect immunofluorescence assays for the serological diagnosis of scrub typhus and murine typhus. Am J Trop Med Hyg, 88 (5), pp. 932-936. | Citations: 14 (Scopus) | Show Abstract | Read more

Inter- and intra-observer variation was examined among six microscopists who read 50 scrub typhus (ST) and murine typhus (MT) indirect immunofluorescence assay (IFA) immunoglobulin M (IgM) slides. Inter-observer agreement was moderate (κ = 0.45) for MT and fair (κ = 0.32) for ST, and was significantly correlated with experience (P = 0.03 and P = 0.004, respectively); κ-scores for intra-observer agreement between morning and afternoon readings (range = 0.35-0.86) were not correlated between years of experience for ST and MT IFAs (Spearman's ρ = 0.31, P = 0.54 and P = 0.14, respectively; P = 0.78). Storage at 4°C for 2 days showed a change from positive to negative in 20-32% of slides. Although the titers did not dramatically change after 14 days of storage, the final interpretation (positive to negative) did change in 36-50% of samples, and it, therefore, recommended that slides should be read as soon as possible after processing.

Kyaw MP, Nyunt MH, Chit K, Aye MM, Aye KH, Aye MM, Lindegardh N, Tarning J, Imwong M, Jacob CG et al. 2013. Reduced susceptibility of Plasmodium falciparum to artesunate in southern Myanmar. PLoS One, 8 (3), pp. e57689. | Citations: 117 (Scopus) | Show Abstract | Read more

BACKGROUND: Plasmodium falciparum resistance to artemisinins, the first line treatment for malaria worldwide, has been reported in western Cambodia. Resistance is characterized by significantly delayed clearance of parasites following artemisinin treatment. Artemisinin resistance has not previously been reported in Myanmar, which has the highest falciparum malaria burden among Southeast Asian countries. METHODS: A non-randomized, single-arm, open-label clinical trial of artesunate monotherapy (4 mg/kg daily for seven days) was conducted in adults with acute blood-smear positive P. falciparum malaria in Kawthaung, southern Myanmar. Parasite density was measured every 12 hours until two consecutive negative smears were obtained. Participants were followed weekly at the study clinic for three additional weeks. Co-primary endpoints included parasite clearance time (the time required for complete clearance of initial parasitemia), parasite clearance half-life (the time required for parasitemia to decrease by 50% based on the linear portion of the parasite clearance slope), and detectable parasitemia 72 hours after commencement of artesunate treatment. Drug pharmacokinetics were measured to rule out delayed clearance due to suboptimal drug levels. RESULTS: The median (range) parasite clearance half-life and time were 4.8 (2.1-9.7) and 60 (24-96) hours, respectively. The frequency distributions of parasite clearance half-life and time were bimodal, with very slow parasite clearance characteristic of the slowest-clearing Cambodian parasites (half-life longer than 6.2 hours) in approximately 1/3 of infections. Fourteen of 52 participants (26.9%) had a measurable parasitemia 72 hours after initiating artesunate treatment. Parasite clearance was not associated with drug pharmacokinetics. CONCLUSIONS: A subset of P. falciparum infections in southern Myanmar displayed markedly delayed clearance following artemisinin treatment, suggesting either emergence of artemisinin resistance in southern Myanmar or spread to this location from its site of origin in western Cambodia. Resistance containment efforts are underway in Myanmar. TRIAL REGISTRATION: Australian New Zealand Clinical Trials Registry ACTRN12610000896077.

Kurt K, Rasigade J-P, Laurent F, Goering RV, Žemličková H, Machova I, Struelens MJ, Zautner AE, Holtfreter S, Bröker B et al. 2013. Subpopulations of Staphylococcus aureus clonal complex 121 are associated with distinct clinical entities. PLoS One, 8 (3), pp. e58155. | Citations: 28 (Scopus) | Show Abstract | Read more

We investigated the population structure of Staphylococcus aureus clonal complex CC121 by mutation discovery at 115 genetic housekeeping loci from each of 154 isolates, sampled on five continents between 1953 and 2009. In addition, we pyro-sequenced the genomes from ten representative isolates. The genome-wide SNPs that were ascertained revealed the evolutionary history of CC121, indicating at least six major clades (A to F) within the clonal complex and dating its most recent common ancestor to the pre-antibiotic era. The toxin gene complement of CC121 isolates was correlated with their SNP-based phylogeny. Moreover, we found a highly significant association of clinical phenotypes with phylogenetic affiliations, which is unusual for S. aureus. All isolates evidently sampled from superficial infections (including staphylococcal scalded skin syndrome, bullous impetigo, exfoliative dermatitis, conjunctivitis) clustered in clade F, which included the European epidemic fusidic-acid resistant impetigo clone (EEFIC). In comparison, isolates from deep-seated infections (abscess, furuncle, pyomyositis, necrotizing pneumonia) were disseminated in several clades, but not in clade F. Our results demonstrate that phylogenetic lineages with distinct clinical properties exist within an S. aureus clonal complex, and that SNPs serve as powerful discriminatory markers, able to identify these lineages. All CC121 genomes harboured a 41-kilobase prophage that was dissimilar to S. aureus phages sequenced previously. Community-associated MRSA and MSSA from Cambodia were extremely closely related, suggesting this MRSA arose in the region.

Price RN. 2013. Potential of artemisinin-based combination therapies to block malaria transmission. J Infect Dis, 207 (11), pp. 1627-1629. | Citations: 10 (Scopus) | Read more

Carrara VI, Lwin KM, Phyo AP, Ashley E, Wiladphaingern J, Sriprawat K, Rijken M, Boel M, McGready R, Proux S et al. 2013. Malaria burden and artemisinin resistance in the mobile and migrant population on the Thai-Myanmar border, 1999-2011: an observational study. PLoS Med, 10 (3), pp. e1001398. | Citations: 100 (Scopus) | Show Abstract | Read more

BACKGROUND: The Shoklo Malaria Research Unit has been working on the Thai-Myanmar border for 25 y providing early diagnosis and treatment (EDT) of malaria. Transmission of Plasmodium falciparum has declined, but resistance to artesunate has emerged. We expanded malaria activities through EDT and evaluated the impact over a 12-y period. METHODS AND FINDINGS: Between 1 October 1999 and 30 September 2011, the Shoklo Malaria Research Unit increased the number of cross-border (Myanmar side) health facilities from two to 11 and recorded the number of malaria consultations. Changes in malaria incidence were estimated from a cohort of pregnant women, and prevalence from cross-sectional surveys. In vivo and in vitro antimalarial drug efficacy were monitored. Over this period, the number of malaria cases detected increased initially, but then declined rapidly. In children under 5 y, the percentage of consultations due to malaria declined from 78% (95% CI 76-80) (1,048/1,344 consultations) to 7% (95% CI 6.2-7.1) (767/11,542 consultations), p<0.001. The ratio of P. falciparum/P. vivax declined from 1.4 (95% CI 1.3-1.4) to 0.7 (95% CI 0.7-0.8). The case fatality rate was low (39/75,126; 0.05% [95% CI 0.04-0.07]). The incidence of malaria declined from 1.1 to 0.1 episodes per pregnant women-year. The cumulative proportion of P. falciparum decreased significantly from 24.3% (95% CI 21.0-28.0) (143/588 pregnant women) to 3.4% (95% CI 2.8-4.3) (76/2,207 pregnant women), p<0.001. The in vivo efficacy of mefloquine-artesunate declined steadily, with a sharp drop in 2011 (day-42 PCR-adjusted cure rate 42% [95% CI 20-62]). The proportion of patients still slide positive for malaria at day 3 rose from 0% in 2000 to reach 28% (95% CI 13-45) (8/29 patients) in 2011. CONCLUSIONS: Despite the emergence of resistance to artesunate in P. falciparum, the strategy of EDT with artemisinin-based combination treatments has been associated with a reduction in malaria in the migrant population living on the Thai-Myanmar border. Although limited by its observational nature, this study provides useful data on malaria burden in a strategically crucial geographical area. Alternative fixed combination treatments are needed urgently to replace the failing first-line regimen of mefloquine and artesunate. Please see later in the article for the Editors' Summary.

Cheng AC, Currie BJ, Dance DAB, Funnell SGP, Limmathurotsakul D, Simpson AJH, Peacock SJ. 2013. Clinical definitions of melioidosis. Am J Trop Med Hyg, 88 (3), pp. 411-413. | Citations: 30 (Scopus) | Show Abstract | Read more

Clinical definitions of melioidosis and inhalation-acquired melioidosis (Burkholderia pseudomallei infection) are described together with the evidence used to develop these definitions. Such definitions support accurate public health reporting, preparedness planning for deliberate B. pseudomallei release, design of experimental models, and categorization of naturally acquired melioidosis.

Malmberg M, Ferreira PE, Tarning J, Ursing J, Ngasala B, Björkman A, Mårtensson A, Gil JP. 2013. Plasmodium falciparum drug resistance phenotype as assessed by patient antimalarial drug levels and its association with pfmdr1 polymorphisms. J Infect Dis, 207 (5), pp. 842-847. | Citations: 44 (Scopus) | Show Abstract | Read more

BACKGROUND: Multidrug-resistant Plasmodium falciparum is a major threat to global malaria control. Parasites develop resistance by gradually acquiring genetic polymorphisms that decrease drug susceptibility. The aim of this study was to investigate the extent to which parasites with different genetic characteristics are able to withstand individual drug blood concentrations. METHODS: We analyzed 2 clinical trials that assessed the efficacy and effectiveness of artemether-lumefantrine. As a proof of concept, we used measured day 7 lumefantrine concentrations to estimate the concentrations at which reinfections multiplied. P. falciparum multidrug resistance gene 1 (pfmdr1) genotypes of these parasites were then correlated to drug susceptibility. RESULTS: Reinfecting parasites with the pfmdr1 N86/184F/D1246 haplotype were able to withstand lumefantrine blood concentrations 15-fold higher than those with the 86Y/Y184/1246Y haplotype. CONCLUSIONS: By estimating drug concentrations, we were able to quantify the contribution of pfmdr1 single-nucleotide polymorphisms to reduced lumefantrine susceptibility. The method can be applied to all long-half-life antimalarial drugs, enables early detection of P. falciparum with reduced drug susceptibility in vivo, and represents a novel way for unveiling molecular markers of antimalarial drug resistance.

Limmathurotsakul D, Dance DAB, Wuthiekanun V, Kaestli M, Mayo M, Warner J, Wagner DM, Tuanyok A, Wertheim H, Yoke Cheng T et al. 2013. Systematic review and consensus guidelines for environmental sampling of Burkholderia pseudomallei. PLoS Negl Trop Dis, 7 (3), pp. e2105. | Citations: 45 (Scopus) | Show Abstract | Read more

BACKGROUND: Burkholderia pseudomallei, a Tier 1 Select Agent and the cause of melioidosis, is a Gram-negative bacillus present in the environment in many tropical countries. Defining the global pattern of B. pseudomallei distribution underpins efforts to prevent infection, and is dependent upon robust environmental sampling methodology. Our objective was to review the literature on the detection of environmental B. pseudomallei, update the risk map for melioidosis, and propose international consensus guidelines for soil sampling. METHODS/PRINCIPAL FINDINGS: An international working party (Detection of Environmental Burkholderia pseudomallei Working Party (DEBWorP)) was formed during the VIth World Melioidosis Congress in 2010. PubMed (January 1912 to December 2011) was searched using the following MeSH terms: pseudomallei or melioidosis. Bibliographies were hand-searched for secondary references. The reported geographical distribution of B. pseudomallei in the environment was mapped and categorized as definite, probable, or possible. The methodology used for detecting environmental B. pseudomallei was extracted and collated. We found that global coverage was patchy, with a lack of studies in many areas where melioidosis is suspected to occur. The sampling strategies and bacterial identification methods used were highly variable, and not all were robust. We developed consensus guidelines with the goals of reducing the probability of false-negative results, and the provision of affordable and 'low-tech' methodology that is applicable in both developed and developing countries. CONCLUSIONS/SIGNIFICANCE: The proposed consensus guidelines provide the basis for the development of an accurate and comprehensive global map of environmental B. pseudomallei.

Choudhury AAK, Conlan JV, Racloz VN, Reid SA, Blacksell SD, Fenwick SG, Thompson ARC, Khamlome B, Vongxay K, Whittaker M. 2013. The economic impact of pig-associated parasitic zoonosis in Northern Lao PDR EcoHealth, 10 (1), pp. 54-62. | Citations: 13 (Scopus) | Show Abstract | Read more

The parasitic zoonoses human cysticercosis (Taenia solium), taeniasis (other Taenia species) and trichinellosis (Trichinella species) are endemic in the Lao People's Democratic Republic (Lao PDR). This study was designed to quantify the economic burden pig-associated zoonotic disease pose in Lao PDR. In particular, the analysis included estimation of the losses in the pork industry as well as losses due to human illness and lost productivity. A Markov-probability based decision-tree model was chosen to form the basis of the calculations to estimate the economic and public health impacts of taeniasis, trichinellosis and cysticercosis. Two different decision trees were run simultaneously on the model's human cohort. A third decision tree simulated the potential impacts on pig production. The human capital method was used to estimate productivity loss. The results found varied significantly depending on the rate of hospitalisation due to neurocysticerosis. This study is the first systematic estimate of the economic impact of pig-associated zoonotic diseases in Lao PDR that demonstrates the significance of the diseases in that country. © 2013 International Association for Ecology and Health.

Anekthananon T, Pukrittayakamee S, Ratanasuwan W, Jittamala P, Werarak P, Charunwatthana P, Suwanagool S, Lawpoolsri S, Stepniewska K, Sapchookul P et al. 2013. Oseltamivir and inhaled zanamivir as influenza prophylaxis in Thai health workers: a randomized, double-blind, placebo-controlled safety trial over 16 weeks. J Antimicrob Chemother, 68 (3), pp. 697-707. | Citations: 7 (Web of Science Lite) | Show Abstract | Read more

OBJECTIVES: Long-term chemoprophylaxis using neuraminidase inhibitors may be needed during influenza epidemics but safety data are limited to several weeks. We sought to assess the tolerability of oseltamivir and zanamivir as primary prophylaxis over 16 weeks. METHODS: We conducted a parallel group, double blind, 2 (active drug) :1 (placebo) randomized trial of oral oseltamivir/placebo or inhaled zanamivir/placebo over 16 weeks in healthy, Thai hospital professionals at two Bangkok hospitals. The primary endpoint was study withdrawal due to drug-related (possibly, probably, definitely) serious or adverse events (AEs) graded ≥ 2. RESULTS: Recruited subjects numbered 129 oseltamivir/65 placebo and 131 zanamivir/65 placebo. A total of 102 grade ≥ 2 AEs were reported or detected in 69 subjects: 23/129 (17.8%) versus 15/65 (23.1%) (P=0.26), and 23/131 (17.6%) versus 8/65 (12.3%) (P=0.28). Intercurrent infections/fevers [26/102 (25.5%)], abnormal biochemistry [25/102 (24.5%)] and gastrointestinal symptoms [18/102 (17.6%)] were the most frequently reported AEs. There were no drug-related study withdrawals. Eight serious AEs were all due to intercurrent illnesses. Laboratory, lung function and ECG parameters were similar between drugs and placebos. CONCLUSIONS: Oseltamivir and zanamivir were well tolerated in healthy hospital professionals. Both drugs can be recommended for primary influenza prophylaxis for up to 16 weeks.

Creek DJ, Bigira V, McCormack S, Arinaitwe E, Wanzira H, Kakuru A, Tappero JW, Sandison TG, Lindegardh N, Nosten F et al. 2013. Pharmacokinetic predictors for recurrent malaria after dihydroartemisinin-piperaquine treatment of uncomplicated malaria in Ugandan infants. J Infect Dis, 207 (11), pp. 1646-1654. | Citations: 15 (Scopus) | Show Abstract | Read more

BACKGROUND: Although dihydroartemisinin-piperaquine (DP) is used primarily in children, pharmacokinetic/pharmacodynamic (PK/PD) data on DP use in young children are lacking. METHODS: We conducted a prospective PK/PD study of piperaquine in 107 young children in Uganda. Samples were collected up to 28 days after 218 episodes of malaria treatment, which occurred during follow-up periods of up to 5 months. Malaria follow-up was conducted actively to day 28 and passively to day 63. RESULTS: The median capillary piperaquine concentration on day 7 after treatment was 41.9 ng/mL. Low piperaquine concentrations were associated with an increased risk of recurrent malaria for up to 42 days, primarily in those receiving trimethoprim-sulfamethoxazole (TMP-SMX) prophylaxis. In children not receiving TMP-SMX, low piperaquine concentrations were only modestly associated with an increased risk of recurrent malaria. However, for children receiving TMP-SMX, associations were strong and evident for all sampling days, with PQ concentrations of ≤ 27.3 ng/mL on day 7 associated with a greatly increased risk of recurrent malaria. Notably, of 132 cases of recurrent malaria, 119 had detectable piperaquine concentrations at the time of presentation with recurrent malaria. CONCLUSIONS: These piperaquine PK/PD data represent the first in children <2 years of age. Piperaquine exposure on day 7 correlated with an increased risk of recurrent malaria after DP treatment in children receiving TMP-SMX prophylaxis. Interestingly, despite strong associations, infants remained at risk for malaria, even if they had residual levels of piperaquine.

West TE, Chantratita N, Chierakul W, Limmathurotsakul D, Wuthiekanun V, Myers ND, Emond MJ, Wurfel MM, Hawn TR, Peacock SJ, Skerrett SJ. 2013. Impaired TLR5 functionality is associated with survival in melioidosis. J Immunol, 190 (7), pp. 3373-3379. | Citations: 23 (Web of Science Lite) | Show Abstract | Read more

Melioidosis is infection caused by the flagellated saprophyte Burkholderia pseudomallei. TLR5 is a pathogen recognition receptor activated by bacterial flagellin. We studied a genetic variant that encodes a defective TLR5 protein, TLR5(1174C)>T, to elucidate the role of TLR5 in melioidosis. We measured NF-κB activation induced by B. pseudomallei in human embryonic kidney-293 cells transfected with TLR5 and found that B. pseudomallei induced TLR5(1174C)- but not TLR5(1174T)-dependent activation of NF-κB. We tested the association of TLR5(1174C)>T with outcome in 600 Thai subjects with melioidosis. In a dominant model, TLR5(1174C)>T was associated with protection against in-hospital death (adjusted odds ratio: 0.20; 95% confidence interval: 0.08-0.50; p = 0.001) and organ failure (adjusted odds ratio: 0.37; 95% confidence interval: 0.19-0.71; p = 0.003). We analyzed blood cytokine production induced by flagellin or heat-killed B. pseudomallei by TLR5(1174C)>T genotype in healthy subjects. Flagellin induced lower monocyte-normalized levels of IL-6, IL-8, TNF-α, IL-10, MCP-1, IL-1ra, G-CSF, and IL-1β in carriers of TLR5(1174T) compared with carriers of TLR5(1174C). B. pseudomallei induced lower monocyte-normalized levels of IL-10 in carriers of TLR5(1174T). We conclude that the hypofunctional genetic variant TLR5(1174C)>T is associated with reduced organ failure and improved survival in melioidosis. This conclusion suggests a deleterious immunoregulatory effect of TLR5 that may be mediated by IL-10 and identifies this receptor as a potential therapeutic target in melioidosis.

Day N. 2013. The researcher of the future...takes advantage of international opportunities. Lancet, 381 Suppl 1 (S1), pp. S8-S9. | Read more

Aubry F, Vongsouvath M, Nougairède A, Phetsouvanh R, Sibounheuang B, Charrel R, Rattanavong S, Phommasone K, Sengvilaipraserth O, de Lamballerie X et al. 2013. Complete Genome of a Genotype I Japanese Encephalitis Virus Isolated from a Patient with Encephalitis in Vientiane, Lao PDR. Genome Announc, 1 (1), | Citations: 9 (European Pubmed Central) | Show Abstract | Read more

Japanese encephalitis virus (JEV) (Flaviviridae, Flavivirus) is an arthropod-borne flavivirus transmitted by Culex species mosquitoes. We report here the complete genome of the JEV genotype I strain JEV_CNS769_Laos_2009 isolated from an infected patient in Vientiane, Lao People's Democratic Republic (PDR) (Laos).

Choudhury AAK, Conlan JV, Racloz VN, Reid SA, Blacksell SD, Fenwick SG, Thompson ARC, Khamlome B, Vongxay K, Whittaker M. 2013. The economic impact of pig-associated parasitic zoonosis in Northern Lao PDR. Ecohealth, 10 (1), pp. 54-62. | Citations: 11 (Web of Science Lite) | Show Abstract | Read more

The parasitic zoonoses human cysticercosis (Taenia solium), taeniasis (other Taenia species) and trichinellosis (Trichinella species) are endemic in the Lao People's Democratic Republic (Lao PDR). This study was designed to quantify the economic burden pig-associated zoonotic disease pose in Lao PDR. In particular, the analysis included estimation of the losses in the pork industry as well as losses due to human illness and lost productivity. A Markov-probability based decision-tree model was chosen to form the basis of the calculations to estimate the economic and public health impacts of taeniasis, trichinellosis and cysticercosis. Two different decision trees were run simultaneously on the model's human cohort. A third decision tree simulated the potential impacts on pig production. The human capital method was used to estimate productivity loss. The results found varied significantly depending on the rate of hospitalisation due to neurocysticerosis. This study is the first systematic estimate of the economic impact of pig-associated zoonotic diseases in Lao PDR that demonstrates the significance of the diseases in that country.

Stoesser N, Emary K, Soklin S, Peng An K, Sophal S, Chhomrath S, Day NPJ, Limmathurotsakul D, Nget P, Pangnarith Y et al. 2013. The value of intermittent point-prevalence surveys of healthcare-associated infections for evaluating infection control interventions at Angkor Hospital for Children, Siem Reap, Cambodia. Trans R Soc Trop Med Hyg, 107 (4), pp. 248-253. | Citations: 12 (Scopus) | Show Abstract | Read more

BACKGROUND: There are limited data on the epidemiology of paediatric healthcare-associated infection (HCAI) and infection control in low-income countries. We describe the value of intermittent point-prevalence surveys for monitoring HCAI and evaluating infection control interventions in a Cambodian paediatric hospital. METHODS: Hospital-wide, point-prevalence surveys were performed monthly in 2011. Infection control interventions introduced during this period included a hand hygiene programme and a ventilator-associated pneumonia (VAP) care bundle. RESULTS: Overall HCAI prevalence was 13.8/100 patients at-risk, with a significant decline over time. The highest HCAI rates (50%) were observed in critical care; the majority of HCAIs were respiratory (61%). Klebsiella pneumoniae was most commonly isolated and antimicrobial resistance was widespread. Hand hygiene compliance doubled to 51.6%, and total VAP cases/1000 patient-ventilator days fell from 30 to 10. CONCLUSION: Rates of HCAI were substantial in our institution, and antimicrobial resistance a major concern. Point-prevalence surveys are effective for HCAI surveillance, and in monitoring trends in response to infection control interventions.

Bitencourt AR, Vicentin EC, Jimenez MC, Ricci R, Leite JA, Costa FT, Ferreira LC, Russell B, Nosten F, Rénia L et al. 2013. Antigenicity and immunogenicity of Plasmodium vivax merozoite surface protein-3. PLoS One, 8 (2), pp. e56061. | Citations: 13 (Scopus) | Show Abstract | Read more

A recent clinical trial in African children demonstrated the potential utility of merozoite surface protein (MSP)-3 as a vaccine against Plasmodium falciparum malaria. The present study evaluated the use of Plasmodium vivax MSP-3 (PvMSP-3) as a target antigen in vaccine formulations against malaria caused by P. vivax. Recombinant proteins representing MSP-3α and MSP-3β of P. vivax were expressed as soluble histidine-tagged bacterial fusions. Antigenicity during natural infection was evaluated by detecting specific antibodies using sera from individuals living in endemic areas of Brazil. A large proportion of infected individuals presented IgG antibodies to PvMSP-3α (68.2%) and at least 1 recombinant protein representing PvMSP-3β (79.1%). In spite of the large responder frequency, reactivity to both antigens was significantly lower than was observed for the immunodominant epitope present on the 19-kDa C-terminal region of PvMSP-1. Immunogenicity of the recombinant proteins was studied in mice in the absence or presence of different adjuvant formulations. PvMSP-3β, but not PvMSP-3α, induced a TLR4-independent humoral immune response in the absence of any adjuvant formulation. The immunogenicity of the recombinant antigens were also tested in formulations containing different adjuvants (Alum, Salmonella enterica flagellin, CpG, Quil A,TiterMax® and incomplete Freunds adjuvant) and combinations of two adjuvants (Alum plus flagellin, and CpG plus flagellin). Recombinant PvMSP-3α and PvMSP-3β elicited higher antibody titers capable of recognizing P. vivax-infected erythrocytes harvested from malaria patients. Our results confirm that P. vivax MSP-3 antigens are immunogenic during natural infection, and the corresponding recombinant proteins may be useful in elucidating their vaccine potential.

Green MD, Mayxay M, Beach R, Pongvongsa T, Phompida S, Hongvanthong B, Vanisaveth V, Newton PN, Vizcaino L, Swamidoss I. 2013. Evaluation of a rapid colorimetric field test to assess the effective life of long-lasting insecticide-treated mosquito nets in the Lao PDR. Malar J, 12 (1), pp. 57. | Citations: 4 (European Pubmed Central) | Show Abstract | Read more

BACKGROUND: Malaria morbidity and mortality have been significantly reduced through the proper use of insecticide-treated mosquito nets, but the extra protection afforded by the insecticide diminishes over time. The insecticide depletion rates vary according to location where wash frequency and wear are influenced by cultural habits as well as the availability of water. Monitoring of available insecticides on the net surface is essential for determining the effective life of the net. Therefore, a rapid and inexpensive colorimetric field test for cyanopyrethroids (Cyanopyrethroid Field Test or CFT) was used to measure surface levels of deltamethrin on insecticide-coated polyester nets (PowerNets™) in rural Lao PDR over a two-year period. METHODS: Net surface levels of deltamethrin were measured by wiping the net with filter paper and measuring the adsorbed deltamethrin using the CFT. A relationship between surface levels of deltamethrin and whole net levels was established by comparing results of the CFT with whole levels assayed by high-performance liquid chromatography (HPLC). An effective deltamethrin surface concentration (EC80) was determined by comparing mosquito mortality (WHO Cone Test) with CFT and HPLC results. Five positions (roof to bottom) on each of 23 matched nets were assayed for deltamethrin surface levels at 6, 12, and 24 months. Mosquito mortality assays (WHO Cone Tests) were performed on a subset of eleven 24-month old nets and compared with the proportion of failed nets as predicted by the CFT. RESULTS: At six months, the nets retained about 80% of the baseline (new net) levels of deltamethrin with no significant differences between net positions. At 12 months, ~15-40%, and at 24 months <10% of deltamethrin was retained on the nets, with significant differences appearing between positions. Results from the CFT show that 93% of the nets failed (deltamethrin surface levels </= EC80) at 24 months. This value is in agreement with 91% failure as determined by the WHO Cone Test on a subset of 11 nets. The CFT results show that 50% of the nets from Laos failed at 12 months of normal use. CONCLUSION: The CFT is a useful and accurate indicator of net efficacy and may be substituted for mosquito bioassays.

Prapansilp P, Medana I, Mai NTH, Day NPJ, Phu NH, Yeo TW, Hien TT, White NJ, Anstey NM, Turner GDH. 2013. A clinicopathological correlation of the expression of the angiopoietin-Tie-2 receptor pathway in the brain of adults with Plasmodium falciparum malaria. Malar J, 12 (1), pp. 50. | Citations: 16 (Web of Science Lite) | Show Abstract | Read more

BACKGROUND: Plasma angiopoietin (Ang)-2 is associated with disease severity and mortality in adults and children with falciparum malaria. However the mechanism of action of the angiopoietins in fatal malaria is unclear. This study aimed to determine whether the expression of Ang-1 and Ang-2 and their receptor Tie-2 in cerebral endothelial or parenchymal cells was specific to cerebral malaria (CM), correlated with coma or other severe clinical features, and whether plasma and CSF levels of these markers correlated with the clinical and neuropathological features of severe and fatal malaria in Vietnamese adults. METHODS: Immunohistochemistry was performed for Ang-1, Ang-2 and Tie-2 on post-mortem brain tissue from fatal malaria cases and controls. Quantitative ELISA for plasma and cerebrospinal fluid levels of Ang-1, Ang-2 and Tie-2 was done to compare fatal cases with surviving patients from the same study. RESULTS: Immunohistochemistry revealed significant differences in expression in endothelial and parenchymal cells compared to controls. However there was no significant difference in expression of these markers on endothelial cells, astroglial cells or neurons between CM and non-cerebral malaria cases. Immunostaining of Ang-1, Ang-2 and Tie-2 was also not associated with Plasmodium falciparum-infected erythrocyte sequestration in the brain. However Ang-1 and Ang-2 expression in neurons was significantly correlated with the incidence of microscopic haemorrhages. Plasma levels of Ang-2 and Ang-2/Ang-1 ratio were associated with the number of severe malaria complications and were significant and independent predictors of metabolic acidosis and fatal outcome. CONCLUSIONS: The independent prognostic significance of Ang-2 and the Ang-2/Ang-1 ratio in severe malaria was confirmed, although immunohistochemistry in fatal cases did not reveal increased expression on brain endothelium in cerebral versus non-cerebral cases. Activation of the Ang-Tie-2 pathway in severe malaria is therefore related to acidosis, number of severity criteria and outcome, but is not a specific event in the brain during cerebral malaria.

Hay SI, Price RN, Baird JK. 2013. The epidemiology of Plasmodium vivax. Preface. Adv Parasitol, 81 pp. xi-xii. | Citations: 1 (Web of Science Lite) | Read more

Pan A, Lee A, Cooper B, Chalfine A, Daikos GL, Garilli S, Goossens H, Malhotra-Kumar S, Martínez JA, Patroni A, Harbarth S. 2013. Risk factors for previously unknown meticillin-resistant Staphylococcus aureus carriage on admission to 13 surgical wards in Europe Journal of Hospital Infection, 83 (2), pp. 107-113. | Citations: 12 (Scopus) | Show Abstract | Read more

Background: Early identification of meticillin-resistant Staphylococcus aureus (MRSA) carriers may be helpful for clinical and epidemiological reasons. Aim: To identify and compare risk factors of previously unknown MRSA carriage on admission to 13 surgical wards in France, Greece, Italy, and Spain. Methods: The study was a prospective observational cohort study which enrolled consecutive patients screened for MRSA on admission to surgical wards. Sociodemographic data, comorbidities and possible risk factors for MRSA were recorded. A multivariate logistic regression model was used to predict probabilities of previously unknown MRSA colonization on admission based on patient characteristics. Prediction rules for MRSA carriage were developed and evaluated using the c-statistic. Findings: Of 2901 patients enrolled, admission screening identified 111 (3.8%) new MRSA carriers. Independent risk factors for MRSA carriage were urinary catheterization (odds ratio: 4.4; 95% confidence interval: 2.0-9.9), nursing home residency (3.8; 1.9-7.7), chronic skin disease (2.9; 1.5-5.8), wounds/ulcers (2.4; 1.5-4.0), recent hospitalization (2.2; 1.5-3.3), diabetes (1.6, 1.02-2.5), and age >70 years (1.5; 1.03-2.3). However, risk factors varied between centres. The c-statistic for the common prediction rule for all centres was 0.64, indicating limited predictive power. Conclusions: Risk profiles for MRSA carriers vary between surgical wards in European countries. Identifying local risk factors is important, as a common European prediction rule was found to be of limited clinical value. © 2012 The Healthcare Infection Society.

Limmathurotsakul D, Kanoksil M, Wuthiekanun V, Kitphati R, deStavola B, Day NPJ, Peacock SJ. 2013. Activities of daily living associated with acquisition of melioidosis in northeast Thailand: a matched case-control study. PLoS Negl Trop Dis, 7 (2), pp. e2072. | Citations: 66 (Scopus) | Show Abstract | Read more

BACKGROUND: Melioidosis is a serious infectious disease caused by the Category B select agent and environmental saprophyte, Burkholderia pseudomallei. Most cases of naturally acquired infection are assumed to result from skin inoculation after exposure to soil or water. The aim of this study was to provide evidence for inoculation, inhalation and ingestion as routes of infection, and develop preventive guidelines based on this evidence. METHODS/PRINCIPAL FINDINGS: A prospective hospital-based 1∶2 matched case-control study was conducted in Northeast Thailand. Cases were patients with culture-confirmed melioidosis, and controls were patients admitted with non-infectious conditions during the same period, matched for gender, age, and diabetes mellitus. Activities of daily living were recorded for the 30-day period before onset of symptoms, and home visits were performed to obtain drinking water and culture this for B. pseudomallei. Multivariable conditional logistic regression analysis based on 286 cases and 512 controls showed that activities associated with a risk of melioidosis included working in a rice field (conditional odds ratio [cOR] = 2.1; 95% confidence interval [CI] 1.4-3.3), other activities associated with exposure to soil or water (cOR = 1.4; 95%CI 0.8-2.6), an open wound (cOR = 2.0; 95%CI 1.2-3.3), eating food contaminated with soil or dust (cOR = 1.5; 95%CI 1.0-2.2), drinking untreated water (cOR = 1.7; 95%CI 1.1-2.6), outdoor exposure to rain (cOR = 2.1; 95%CI 1.4-3.2), water inhalation (cOR = 2.4; 95%CI 1.5-3.9), current smoking (cOR = 1.5; 95%CI 1.0-2.3) and steroid intake (cOR = 3.1; 95%CI 1.4-6.9). B. pseudomallei was detected in water source(s) consumed by 7% of cases and 3% of controls (cOR = 2.2; 95%CI 0.8-5.8). CONCLUSIONS/SIGNIFICANCE: We used these findings to develop the first evidence-based guidelines for the prevention of melioidosis. These are suitable for people in melioidosis-endemic areas, travelers and military personnel. Public health campaigns based on our recommendations are under development in Thailand.

Chairat K, Tarning J, White NJ, Lindegardh N. 2013. Pharmacokinetic properties of anti-influenza neuraminidase inhibitors. J Clin Pharmacol, 53 (2), pp. 119-139. | Citations: 13 (European Pubmed Central) | Show Abstract | Read more

Neuraminidase inhibitors are the mainstay of anti-influenza treatment. Oseltamivir is the most widely used drug but is currently available only as an oral formulation. Resistance spreads rapidly in seasonal H1N1 influenza A viruses, which were universally resistant in 2008, because of the H275Y mutation in the neuraminidase (NA) gene. Oseltamivir is a prodrug for the active carboxylate metabolite. Ex vivo conversion in blood samples may have confounded early pharmacokinetic studies. Oseltamivir shows dose linear kinetics, and oseltamivir carboxylate has an elimination half-life (t(1/2) β) after oral administration in healthy individuals of approximately 7.7 hours. Oseltamivir carboxylate is eliminated primarily by tubular secretion, and both clearance and tissue distribution are reduced by probenecid. The H275Y mutation in NA confers high-level oseltamivir resistance and intermediate peramivir resistance but does not alter zanamivir susceptibility. Zanamivir is available as a powder for inhalation, and a parenteral form is under development. Zanamivir distributes in an apparent volume of distribution approximating that of extracellular water and is rapidly eliminated (t(1/2) β of approximately 3.0 hours). Peramivir is slowly eliminated (t(1/2) β of 7.7-20.8 hours) and is prescribed as either a once-daily injection or as a single infusion. Laninamivir is a recently developed slowly eliminated compound for administration by inhalation.

Peacock SJ, Limmathurotsakul D, Lubell Y, Koh GC, White LJ, Day NP, Titball RW. 2013. Correction: Melioidosis Vaccines: A Systematic Review and Appraisal of the Potential to Exploit Biodefense Vaccines for Public Health Purposes. PLoS Negl Trop Dis, 7 (2), | Show Abstract | Read more

[This corrects the article on p. e1488 in vol. 6.].

White NJ. 2013. Primaquine to prevent transmission of falciparum malaria. The Lancet infectious diseases, 13 (2), pp. 175-181. | Citations: 51 (Scopus) | Show Abstract

Falciparum malaria is transmitted by anopheline mosquitoes that have fed on blood containing gametocytes of Plasmodium falciparum. In areas of low malaria transmission, where symptomatic infections contribute substantially to malaria transmission, the use of gametocytocidal drugs reduces the incidence of malaria. Artemisinin-based combination therapies provide high cure rates and substantially reduce gametocyte carriage. Artemisinin resistance in P falciparum lessens overall gametocytocidal activity, which provides a selective pressure to the spread of these resistant parasites. The 8-aminoquinoline compounds possess unique gametocytocidal properties and rapidly sterilise the mature transmissible stages of P falciparum. The addition of one dose of primaquine to artemisinin-based combination regimens could help to counter the spread of artemisinin resistance. Although primaquine is commonly recommended for falciparum and vivax malaria, concerns about drug-related haemolysis frequently prevent its administration. The limited available evidence on transmission-blocking effects of primaquine and its forerunner plasmoquine suggests that doses lower than currently recommended (0.50-0.75 mg base per kg), which would be safer, might still be very effective. Copyright © 2013 Elsevier Ltd. All rights reserved.

Kauss T, Gaubert A, Boyer C, Ba BB, Manse M, Massip S, Léger JM, Fawaz F, Lembege M, Boiron JM et al. 2013. Pharmaceutical development and optimization of azithromycin suppository for paediatric use International Journal of Pharmaceutics, 441 (1-2), pp. 218-226. | Citations: 11 (Scopus) | Show Abstract | Read more

Pharmaceutical development and manufacturing process optimization work was undertaken in order to propose a potential paediatric rectal formulation of azithromycin as an alternative to existing oral or injectable formulations. The target product profile was to be easy-to-use, cheap and stable in tropical conditions, with bioavailability comparable to oral forms, rapidly achieving and maintaining bactericidal concentrations. PEG solid solution suppositories were characterized in vitro using visual, HPLC, DSC, FTIR and XRD analyses. In vitro drug release and in vivo bioavailability were assessed; a study in rabbits compared the bioavailability of the optimized solid solution suppository to rectal solution and intra-venous product (as reference) and to the previous, non-optimized formulation (suspended azithromycin suppository). The bioavailability of azithromycin administered as solid solution suppositories relative to intra-venous was 43%, which compared well to the target of 38% (oral product in humans). The results of 3-month preliminary stability and feasibility studies were consistent with industrial production scale-up. This product has potential both as a classical antibiotic and as a product for use in severely ill children in rural areas. Industrial partners for further development are being sought. © 2012 Elsevier B.V. All rights reserved.

Koh GCKW, Schreiber MF, Bautista R, Maude RR, Dunachie S, Limmathurotsakul D, Day NPJ, Dougan G, Peacock SJ. 2013. Host responses to melioidosis and tuberculosis are both dominated by interferon-mediated signaling. PLoS One, 8 (1), pp. e54961. | Citations: 41 (Web of Science Lite) | Show Abstract | Read more

Melioidosis (Burkholderia pseudomallei infection) is a common cause of community-acquired sepsis in Northeast Thailand and northern Australia. B. pseudomallei is a soil saprophyte endemic to Southeast Asia and northern Australia. The clinical presentation of melioidosis may mimic tuberculosis (both cause chronic suppurative lesions unresponsive to conventional antibiotics and both commonly affect the lungs). The two diseases have overlapping risk profiles (e.g., diabetes, corticosteroid use), and both B. pseudomallei and Mycobacterium tuberculosis are intracellular pathogens. There are however important differences: the majority of melioidosis cases are acute, not chronic, and present with severe sepsis and a mortality rate that approaches 50% despite appropriate antimicrobial therapy. By contrast, tuberculosis is characteristically a chronic illness with mortality <2% with appropriate antimicrobial chemotherapy. We examined the gene expression profiles of total peripheral leukocytes in two cohorts of patients, one with acute melioidosis (30 patients and 30 controls) and another with tuberculosis (20 patients and 24 controls). Interferon-mediated responses dominate the host response to both infections, and both type 1 and type 2 interferon responses are important. An 86-gene signature previously thought to be specific for tuberculosis is also found in melioidosis. We conclude that the host responses to melioidosis and to tuberculosis are similar: both are dominated by interferon-signalling pathways and this similarity means gene expression signatures from whole blood do not distinguish between these two diseases.

Boonsilp S, Thaipadungpanit J, Amornchai P, Wuthiekanun V, Bailey MS, Holden MTG, Zhang C, Jiang X, Koizumi N, Taylor K et al. 2013. A single multilocus sequence typing (MLST) scheme for seven pathogenic Leptospira species. PLoS Negl Trop Dis, 7 (1), pp. e1954. | Citations: 67 (Scopus) | Show Abstract | Read more

BACKGROUND: The available Leptospira multilocus sequence typing (MLST) scheme supported by a MLST website is limited to L. interrogans and L. kirschneri. Our aim was to broaden the utility of this scheme to incorporate a total of seven pathogenic species. METHODOLOGY AND FINDINGS: We modified the existing scheme by replacing one of the seven MLST loci (fadD was changed to caiB), as the former gene did not appear to be present in some pathogenic species. Comparison of the original and modified schemes using data for L. interrogans and L. kirschneri demonstrated that the discriminatory power of the two schemes was not significantly different. The modified scheme was used to further characterize 325 isolates (L. alexanderi [n = 5], L. borgpetersenii [n = 34], L. interrogans [n = 222], L. kirschneri [n = 29], L. noguchii [n = 9], L. santarosai [n = 10], and L. weilii [n = 16]). Phylogenetic analysis using concatenated sequences of the 7 loci demonstrated that each species corresponded to a discrete clade, and that no strains were misclassified at the species level. Comparison between genotype and serovar was possible for 254 isolates. Of the 31 sequence types (STs) represented by at least two isolates, 18 STs included isolates assigned to two or three different serovars. Conversely, 14 serovars were identified that contained between 2 to 10 different STs. New observations were made on the global phylogeography of Leptospira spp., and the utility of MLST in making associations between human disease and specific maintenance hosts was demonstrated. CONCLUSION: The new MLST scheme, supported by an updated MLST website, allows the characterization and species assignment of isolates of the seven major pathogenic species associated with leptospirosis.

Pan-ngum W, Blacksell SD, Lubell Y, Pukrittayakamee S, Bailey MS, de Silva HJ, Lalloo DG, Day NPJ, White LJ, Limmathurotsakul D. 2013. Estimating the true accuracy of diagnostic tests for dengue infection using bayesian latent class models. PLoS One, 8 (1), pp. e50765. | Citations: 23 (Scopus) | Show Abstract | Read more

BACKGROUND: Accuracy of rapid diagnostic tests for dengue infection has been repeatedly estimated by comparing those tests with reference assays. We hypothesized that those estimates might be inaccurate if the accuracy of the reference assays is not perfect. Here, we investigated this using statistical modeling. METHODS/PRINCIPAL FINDINGS: Data from a cohort study of 549 patients suspected of dengue infection presenting at Colombo North Teaching Hospital, Ragama, Sri Lanka, that described the application of our reference assay (a combination of Dengue IgM antibody capture ELISA and IgG antibody capture ELISA) and of three rapid diagnostic tests (Panbio NS1 antigen, IgM antibody and IgG antibody rapid immunochromatographic cassette tests) were re-evaluated using bayesian latent class models (LCMs). The estimated sensitivity and specificity of the reference assay were 62.0% and 99.6%, respectively. Prevalence of dengue infection (24.3%), and sensitivities and specificities of the Panbio NS1 (45.9% and 97.9%), IgM (54.5% and 95.5%) and IgG (62.1% and 84.5%) estimated by bayesian LCMs were significantly different from those estimated by assuming that the reference assay was perfect. Sensitivity, specificity, PPV and NPV for a combination of NS1, IgM and IgG cassette tests on admission samples were 87.0%, 82.8%, 62.0% and 95.2%, respectively. CONCLUSIONS: Our reference assay is an imperfect gold standard. In our setting, the combination of NS1, IgM and IgG rapid diagnostic tests could be used on admission to rule out dengue infection with a high level of accuracy (NPV 95.2%). Further evaluation of rapid diagnostic tests for dengue infection should include the use of appropriate statistical models.

Kanoksil M, Jatapai A, Peacock SJ, Limmathurotsakul D. 2013. Epidemiology, microbiology and mortality associated with community-acquired bacteremia in northeast Thailand: a multicenter surveillance study. PLoS One, 8 (1), pp. e54714. | Citations: 38 (Scopus) | Show Abstract | Read more

BACKGROUND: National statistics in developing countries are likely to underestimate deaths due to bacterial infections. Here, we calculated mortality associated with community-acquired bacteremia (CAB) in a developing country using routinely available databases. METHODS/PRINCIPAL FINDINGS: Information was obtained from the microbiology and hospital database of 10 provincial hospitals in northeast Thailand, and compared with the national death registry from the Ministry of Interior, Thailand for the period between 2004 and 2010. CAB was defined in patients who had pathogenic organisms isolated from blood taken within 2 days of hospital admission without a prior inpatient episode in the preceding 30 days. A total of 15,251 CAB patients identified, of which 5,722 (37.5%) died within 30 days of admission. The incidence rate of CAB between 2004 and 2010 increased from 16.7 to 38.1 per 100,000 people per year, and the mortality rate associated with CAB increased from 6.9 to 13.7 per 100,000 people per year. In 2010, the mortality rate associated with CAB was lower than that from respiratory tract infection, but higher than HIV disease or tuberculosis. The most common causes of CAB were Escherichia coli (23.1%), Burkholderia pseudomallei (19.3%), and Staphylococcus aureus (8.2%). There was an increase in the proportion of Extended-Spectrum Beta-Lactamases (ESBL) producing E. coli and Klebsiella pneumoniae over time. CONCLUSIONS: This study has demonstrated that national statistics on causes of death in developing countries could be improved by integrating information from readily available databases. CAB is neglected as an important cause of death, and specific prevention and intervention is urgently required to reduce its incidence and mortality.

Sunyakumthorn P, Paris DH, Chan T-C, Jones M, Luce-Fedrow A, Chattopadhyay S, Jiang J, Anantatat T, Turner GDH, Day NPJ, Richards AL. 2013. An intradermal inoculation model of scrub typhus in Swiss CD-1 mice demonstrates more rapid dissemination of virulent strains of Orientia tsutsugamushi. PLoS One, 8 (1), pp. e54570. | Citations: 18 (European Pubmed Central) | Show Abstract | Read more

Scrub typhus is an important endemic disease of the Asia-Pacific region caused by Orientia tsutsugamushi. To develop an effective vaccine to prevent scrub typhus infection, a better understanding of the initial host-pathogen interaction is needed. The objective of this study was to investigate early bacterial dissemination in a CD-1 Swiss outbred mouse model after intradermal injection of O. tsutsugamushi. Three human pathogenic strains of O. tsutsugamushi (Karp, Gilliam, and Woods) were chosen to investigate the early infection characteristics associated with bacterial virulence. Tissue biopsies of the intradermal injection site and draining lymph nodes were examined using histology and immunohistochemistry to characterize bacterial dissemination, and correlated with quantitative real-time PCR for O. tsutsugamushi in blood and tissue from major organs. Soluble adhesion molecules were measured to examine cellular activation in response to infection. No eschar formation was seen at the inoculation site and no clinical disease developed within the 7 day period of observation. However, O. tsutsugamushi was localized at the injection site and in the draining lymph nodes by day 7 post inoculation. Evidence of leukocyte and endothelial activation was present by day 7 with significantly raised levels of sL-selectin, sICAM-1 and sVCAM-1. Infection with the Karp strain was associated with earlier and higher bacterial loads and more extensive dissemination in various tissues than the less pathogenic Gilliam and Woods strains. The bacterial loads of O. tsutsugamushi were highest in the lungs and spleens of mice inoculated with Karp and Gilliam, but not Woods strains. Strains of higher virulence resulted in more rapid systemic infection and dissemination in this model. The CD-1 mouse intradermal inoculation model demonstrates features relevant to early scrub typhus infection in humans, including the development of regional lymphadenopathy, leukocyte activation and distant organ dissemination after low-dose intradermal injection with O. tsutsugamushi.

Hanson JP, Lam SWK, Mohanty S, Alam S, Pattnaik R, Mahanta KC, Hasan MU, Charunwatthana P, Mishra SK, Day NPJ et al. 2013. Fluid resuscitation of adults with severe falciparum malaria: effects on Acid-base status, renal function, and extravascular lung water. Crit Care Med, 41 (4), pp. 972-981. | Citations: 35 (Web of Science Lite) | Show Abstract | Read more

OBJECTIVE: To evaluate the efficacy and safety of liberal fluid resuscitation of adults with severe malaria. DESIGN, SETTING, PATIENTS, AND METHODS: Twenty-eight Bangladeshi and Indian adults with severe falciparum malaria received crystalloid resuscitation guided by transpulmonary thermodilution (PiCCO) in an intensive care setting. Systemic hemodynamics, microvascular indices and measures of acidosis, renal function, and pulmonary edema were followed prospectively. RESULTS: All patients were hypovolemic (global end-diastolic volume index<680 mL/m) on enrollment. Patients received a median (range) 3230 mL (390-7300) of isotonic saline in the first 6 hours and 5450 mL (710-13,720) in the first 24 hours. With resuscitation, acid-base status deteriorated in 19 of 28 (68%), and there was no significant improvement in renal function. Extravascular lung water increased in 17 of 22 liberally resuscitated patients (77%); eight of these patients developed pulmonary edema, five of whom died. All other patients survived. All patients with pulmonary edema during the study were hypovolemic or euvolemic at the time pulmonary edema developed. Plasma lactate was lower in hypovolemic patients before (rs=0.38; p=0.05) and after (rs=0.49; p=0.01) resuscitation but was the strongest predictor of mortality before (chi-square=9.9; p=0.002) and after resuscitation (chi-square=11.1; p<0.001) and correlated with the degree of visualized microvascular sequestration of parasitized erythrocytes at both time points (rs=0.55; p=0.003 and rs=0.43; p=0.03, respectively). Persisting sequestration was evident in 7 of 15 patients (47%) 48 hours after enrollment. CONCLUSIONS: Lactic acidosis--the strongest prognostic indicator in adults with severe falciparum malaria--results from sequestration of parasitized erythrocytes in the microcirculation, not from hypovolemia. Liberal fluid resuscitation has little effect on this sequestration and does not improve acid-base status or renal function. Pulmonary edema--secondary to increased pulmonary vascular permeability--is common, unpredictable, and exacerbated by fluid loading. Liberal fluid replacement of adults with severe malaria should be avoided.

Turner C, Turner P, Carrara V, Burgoine K, Tha Ler Htoo S, Watthanaworawit W, Day NP, White NJ, Goldblatt D, Nosten F. 2013. High rates of pneumonia in children under two years of age in a South East Asian refugee population. PLoS One, 8 (1), pp. e54026. | Citations: 15 (Web of Science Lite) | Show Abstract | Read more

BACKGROUND: There are an estimated 150 million episodes of childhood pneumonia per year, with 11-20 million hospital admissions and 1.575 million deaths. Refugee children are particularly vulnerable, with poorly defined pneumonia epidemiology. METHODS: We followed a birth cohort of 955 refugee infants, born over a one-year period, until two years of age. Clinical and radiographic pneumonia were diagnosed according to WHO criteria. Detailed characteristics were collected to determine risk factors for clinical, radiological and multiple episodes of pneumonia. Investigations were taken during a pneumonia episode to help determine or to infer an aetiological diagnosis. FINDINGS: The incidence of clinical pneumonia was 0.73 (95% CI 0.70-0.75) episodes per child year (/CY) and of radiological primary endpoint pneumonia (PEP) was 0.22/CY (95% CI 0.20-0.24). The incidence of pneumonia without severe signs was 0.50/CY (95% CI 0.48-0.53), severe pneumonia 0.15/CY (95% CI 0.13-0.17) and very severe pneumonia 0.06/CY (0.05-0.07). Virus was detected, from a nasopharyngeal aspirate, in 61.3% of episodes. A reduced volume of living space per person (IRR 0.99, 95% CI 0.99-1.0, p = 0.003) and young maternal age (IRR 1.59, 95% CI 1.12-2.27, p = 0.01) were risk factors for developing pneumonia. The risk of a child having >1 episode of pneumonia was increased by having a shorter distance to the next house (IRR 0.86, 95% CI 0.74-1.00, p = 0.04). Infants were at risk of having an episode of PEP if there was a shorter distance from stove to bed (IRR 0.89, 95% CI 0.80-0.99, p = 0.03). Raised CRP and neutrophil values were associated with PEP. CONCLUSIONS: There was a high incidence of pneumonia in young children in this SE Asian refugee population. Viral infections were important, however CXR and non-specific marker findings suggested that bacteria may be involved in up to a third of cases.

Auburn S, Marfurt J, Maslen G, Campino S, Ruano Rubio V, Manske M, Machunter B, Kenangalem E, Noviyanti R, Trianty L et al. 2013. Effective preparation of Plasmodium vivax field isolates for high-throughput whole genome sequencing. PLoS One, 8 (1), pp. e53160. | Citations: 18 (European Pubmed Central) | Show Abstract | Read more

Whole genome sequencing (WGS) of Plasmodium vivax is problematic due to the reliance on clinical isolates which are generally low in parasitaemia and sample volume. Furthermore, clinical isolates contain a significant contaminating background of host DNA which confounds efforts to map short read sequence of the target P. vivax DNA. Here, we discuss a methodology to significantly improve the success of P. vivax WGS on natural (non-adapted) patient isolates. Using 37 patient isolates from Indonesia, Thailand, and travellers, we assessed the application of CF11-based white blood cell filtration alone and in combination with short term ex vivo schizont maturation. Although CF11 filtration reduced human DNA contamination in 8 Indonesian isolates tested, additional short-term culture increased the P. vivax DNA yield from a median of 0.15 to 6.2 ng µl(-1) packed red blood cells (pRBCs) (p = 0.001) and reduced the human DNA percentage from a median of 33.9% to 6.22% (p = 0.008). Furthermore, post-CF11 and culture samples from Thailand gave a median P. vivax DNA yield of 2.34 ng µl(-1) pRBCs, and 2.65% human DNA. In 22 P. vivax patient isolates prepared with the 2-step method, we demonstrate high depth (median 654X coverage) and breadth (≥89%) of coverage on the Illumina GAII and HiSeq platforms. In contrast to the A+T-rich P. falciparum genome, negligible bias was observed in coverage depth between coding and non-coding regions of the P. vivax genome. This uniform coverage will greatly facilitate the detection of SNPs and copy number variants across the genome, enabling unbiased exploration of the natural diversity in P. vivax populations.

Kolyva C, Kingston H, Tachtsidis I, Mohanty S, Mishra S, Patnaik R, Maude RJ, Dondorp AM, Elwell CE. 2013. Oscillations in cerebral haemodynamics in patients with falciparum malaria. Adv Exp Med Biol, 765 pp. 101-107. | Citations: 11 (Scopus) | Show Abstract | Read more

Spontaneous oscillations in cerebral haemodynamics studied with near-infrared spectroscopy (NIRS), become impaired in several pathological conditions. We assessed the spectral characteristics of these oscillations in 20 patients with falciparum malaria admitted to Ispat General Hospital, Rourkela, India. Monitoring included continuous frontal lobe NIRS recordings within 24 h of admission (Day 0), together with single measurements of a number of clinical and chemical markers recorded on admission. Seven patients returned for follow-up measurements on recovery (FU). A 2,048 sampling-point segment of oxygenated haemoglobin concentration ([ΔHbO(2)]) data was subjected to Fourier analysis per patient, and power spectral density was derived over the very low frequency (VLF: 0.02-0.04 Hz), low frequency (LF: 0.04-0.15 Hz) and high frequency (HF: 0.15-0.4 Hz) bands. At Day 0, VLF spectral power was 21.1 ± 16.4, LF power 7.2 ± 4.6 and HF power 2.6 ± 5.0, with VLF power being statistically significantly higher than LF and HF (P < 0.005). VLF power tended to decrease in the severely ill patients and correlated negatively with heart rate (r = 0.57, P < 0.01), while LF power correlated positively with aural body temperature (r = 0.49, P < 0.05). In all but one of the patients who returned for FU measurements, VLF power increased after recovery. This may be related to autonomic dysfunction in severe malaria, a topic of little research to date. The present study demonstrated that application of NIRS in a resource-poor setting is feasible and has potential as a research tool.

Deen J, White NJ, Lubell Y. 2013. Bloodstream infections in south and southeast Asia - authors' reply. Lancet Infect Dis, 13 (1), pp. 15. | Read more

Day NPJ. 2013. Panton-Valentine leucocidin and staphylococcal disease The Lancet Infectious Diseases, 13 (1), pp. 5-6. | Citations: 1 (Scopus) | Read more

Wang H, Yuan Z, Barnes E, Yuan M, Li C, Fu Y, Xia X, Li G, Newton PN, Vongsouvath M et al. 2013. Eight novel hepatitis c virus genomes reveal the changing taxonomic structure of genotype 6 Journal of General Virology, 94 (PART11), pp. 76-80. | Citations: 11 (Scopus) | Show Abstract | Read more

Analysis of partial hepatitis C virus sequences has revealed many novel genotype 6 variants that cannot be unambiguously classified, which obscure the distinctiveness of pre-existing subtypes. To explore this uncertainty, we obtained genomes of 98.0-98.8% full-length for eight such variants (KM35, QC273, TV257, TV476, TV533, L349, QC271 and DH027) and characterized them using phylogenetic analyses and per cent nucleotide similarities. The former four are closely related phylogenetically to subtype 6k, TV533 and L349 to subtype 6l, QC271 to subtypes 6i and 6j, and DH027 to subtypes 6m and 6n. The former six defined a high-level grouping that comprised subtypes 6k and 6l, plus related strains. The threshold between intra- and intersubtype diversity in this group was indistinct. We propose that similar results would be seen elsewhere if more intermediate variants like QC271 and DH027 were sampled. © 2013 SGM.

Vaid N, Langan KM, Maude RJ. 2013. Post-exposure prophylaxis in resource-poor settings: review and recommendations for pre-departure risk assessment and planning for expatriate healthcare workers. Trop Med Int Health, 18 (5), pp. 588-595. | Citations: 3 (European Pubmed Central) | Show Abstract | Read more

It is estimated that more than 3 million healthcare workers worldwide suffer needlestick and splash injuries whilst at work resulting in the potential transmission of blood-borne pathogens via exposure to bodily fluids. Under-reporting and the subsequent management of occupational injuries is a problem both in the United Kingdom and abroad. Many expatriate health care workers will work in low resource settings where the risk of transmission is greatest but in contrast to wealthier countries such as the United Kingdom, there is often a lack of effective systems for its safe management. This article provides important information about this risk and how to minimise it. The reasons for an increased risk in transmission, its subsequent management and pre-departure planning are discussed, together with the evidence for initiation of post-exposure prophylaxis; current National and International guidelines as well as the urgent need for International standardisation of these is also discussed.

Orth H, Jensen BO, Holtfreter MC, Kocheril SJ, Mallach S, MacKenzie C, Müller-Stöver I, Henrich B, Imwong M, White NJ et al. 2013. Plasmodium knowlesi infection imported to Germany, January 2013. Euro Surveill, 18 (40), pp. 4-6. | Citations: 15 (Scopus) | Show Abstract | Read more

Plasmodium knowlesi was known as a plasmodium of macaques until P. knowlesi transmission to humans was recognised in Borneo and later throughout South-East Asia. We describe here a case of a P. knowlesi infection imported to Germany from Thailand. The patient had not taken antimalarial chemoprophylaxis and suffered from daily fever attacks. Microscopy revealed trophozoites and gametocytes resembling P. malariae. P. knowlesi malaria was confirmed by PCR.

Luangasanatip N, Hongsuwan M, Lubell Y, Limmathurotsakul D, Teparrukkul P, Chaowarat S, Day NPJ, Graves N, Cooper BS. 2013. Long-term survival after intensive care unit discharge in Thailand: a retrospective study CRITICAL CARE, 17 (5), | Citations: 9 (Web of Science Lite) | Read more

Lwin KM, Peto TJ, White NJ, Day NPJ, Nosten F, Parker M, Cheah PY. 2013. The practicality and sustainability of a community advisory board at a large medical research unit on the Thai-Myanmar border Health, 05 (02), pp. 229-236. | Read more

Seo S, Englund JA, Nguyen JT, Pukrittayakamee S, Lindegardh N, Tarning J, Tambyah PA, Renaud C, Went GT, de Jong MD, Boeckh MJ. 2013. Combination therapy with amantadine, oseltamivir and ribavirin for influenza A infection: safety and pharmacokinetics. Antivir Ther, 18 (3), pp. 377-386. | Citations: 33 (Scopus) | Show Abstract | Read more

BACKGROUND: Antiviral resistance among influenza A viruses is associated with high morbidity and mortality in immunocompromised hosts. However, treatment strategies for drug-resistant influenza A are not established. A triple-combination antiviral drug (TCAD) regimen consisting of amantadine (AMT), oseltamivir (OSL) and ribavirin (RBV) demonstrated good efficacy in an animal model. METHODS: We first analysed the pharmacokinetics (PKs) of TCAD therapy in healthy volunteers. We then performed a pilot study of TCAD therapy in patients undergoing chemotherapy or haematopoietic cell transplantation. AMT (75 mg), OSL (50 mg) and RBV (200 mg) were administered three times a day for 10 days. The safety and PKs of TCAD therapy were monitored. RESULTS: The PKs of TCAD therapy in healthy volunteers was shown to be similar to the PKs of each drug individually from a single dose. In the pilot study, six immunocompromised patients received TCAD therapy and one patient received OSL monotherapy. All but one patient completed 10 days of TCAD therapy without side effects; one patient receiving TCAD was withdrawn from the study because of respiratory failure and ultimately recovered. Viral load was decreased after TCAD therapy, despite the presence of either AMT- or OSL-resistant virus in two cases. One patient with 2009 influenza A/H1N1 receiving OSL monotherapy developed confirmed OSL resistance during treatment. CONCLUSIONS: TCAD therapy had similar PKs to each individual antiviral during monotherapy following a single dose and can be administered safely in immunocompromised patients.

Takala-Harrison S, Clark TG, Jacob CG, Cummings MP, Miotto O, Dondorp AM, Fukuda MM, Nosten F, Noedl H, Imwong M et al. 2013. Genetic loci associated with delayed clearance of Plasmodium falciparum following artemisinin treatment in Southeast Asia. Proc Natl Acad Sci U S A, 110 (1), pp. 240-245. | Citations: 167 (Scopus) | Show Abstract | Read more

The recent emergence of artemisinin-resistant Plasmodium falciparum malaria in western Cambodia could threaten prospects for malaria elimination. Identification of the genetic basis of resistance would provide tools for molecular surveillance, aiding efforts to contain resistance. Clinical trials of artesunate efficacy were conducted in Bangladesh, in northwestern Thailand near the Myanmar border, and at two sites in western Cambodia. Parasites collected from trial participants were genotyped at 8,079 single nucleotide polymorphisms (SNPs) using a P. falciparum-specific SNP array. Parasite genotypes were examined for signatures of recent positive selection and association with parasite clearance phenotypes to identify regions of the genome associated with artemisinin resistance. Four SNPs on chromosomes 10 (one), 13 (two), and 14 (one) were significantly associated with delayed parasite clearance. The two SNPs on chromosome 13 are in a region of the genome that appears to be under strong recent positive selection in Cambodia. The SNPs on chromosomes 10 and 13 lie in or near genes involved in postreplication repair, a DNA damage-tolerance pathway. Replication and validation studies are needed to refine the location of loci responsible for artemisinin resistance and to understand the mechanism behind it; however, two SNPs on chromosomes 10 and 13 may be useful markers of delayed parasite clearance in surveillance for artemisinin resistance in Southeast Asia.

Witkowski B, Khim N, Chim P, Kim S, Ke S, Kloeung N, Chy S, Duong S, Leang R, Ringwald P et al. 2013. Reduced artemisinin susceptibility of Plasmodium falciparum ring stages in western Cambodia. Antimicrob Agents Chemother, 57 (2), pp. 914-923. | Citations: 119 (Scopus) | Show Abstract | Read more

The declining efficacy of artemisinin derivatives against Plasmodium falciparum in western Cambodia is a major concern. The knowledge gap in the understanding of the mechanisms involved hampers designing monitoring tools. Here, we culture-adapted 20 isolates from Pailin and Ratanakiri (areas of artemisinin resistance and susceptibility in western and eastern Cambodia, respectively) and studied their in vitro response to dihydroartemisinin. No significant difference between the two sets of isolates was observed in the classical isotopic test. However, a 6-h pulse exposure to 700 nM dihydroartemisinin (ring-stage survival assay -RSA]) revealed a clear-cut geographic dichotomy. The survival rate of exposed ring-stage parasites (ring stages) was 17-fold higher in isolates from Pailin (median, 13.5%) than in those from Ratanakiri (median, 0.8%), while exposed mature stages were equally and highly susceptible (0.6% and 0.7%, respectively). Ring stages survived drug exposure by cell cycle arrest and resumed growth upon drug withdrawal. The reduced susceptibility to artemisinin in Pailin appears to be associated with an altered in vitro phenotype of ring stages from Pailin in the RSA.

Fowkes FJI, McGready R, Johnstone-Robertson S, Nosten F, Beeson JG. 2013. Antibody boosting and longevity following tetanus immunization during pregnancy. Clin Infect Dis, 56 (5), pp. 749-750. | Citations: 6 (European Pubmed Central) | Read more

Hendriksen ICE, Maiga D, Lemnge MM, Mtove G, Gesase S, Reyburn H, Lindegardh N, Day NPJ, von Seidlein L, Dondorp AM et al. 2013. Population pharmacokinetic and pharmacodynamic properties of intramuscular quinine in Tanzanian children with severe Falciparum malaria. Antimicrob Agents Chemother, 57 (2), pp. 775-783. | Citations: 5 (European Pubmed Central) | Show Abstract | Read more

Although artesunate is clearly superior, parenteral quinine is still used widely for the treatment of severe malaria. A loading-dose regimen has been recommended for 30 years but is still often not used. A population pharmacokinetic study was conducted with 75 Tanzanian children aged 4 months to 8 years with severe malaria who received quinine intramuscularly; 69 patients received a loading dose of 20 mg quinine dihydrochloride (salt)/kg of body weight. Twenty-one patients had plasma quinine concentrations detectable at baseline. A zero-order absorption model with one-compartment disposition pharmacokinetics described the data adequately. Body weight was the only significant covariate and was implemented as an allometric function on clearance and volume parameters. Population pharmacokinetic parameter estimates (and percent relative standard errors [%RSE]) of elimination clearance, central volume of distribution, and duration of zero-order absorption were 0.977 liters/h (6.50%), 16.7 liters (6.39%), and 1.42 h (21.5%), respectively, for a typical patient weighing 11 kg. Quinine exposure was reduced at lower body weights after standard weight-based dosing; there was 18% less exposure over 24 h in patients weighing 5 kg than in those weighing 25 kg. Maximum plasma concentrations after the loading dose were unaffected by body weight. There was no evidence of dose-related drug toxicity with the loading dosing regimen. Intramuscular quinine is rapidly and reliably absorbed in children with severe falciparum malaria. Based on these pharmacokinetic data, a loading dose of 20 mg salt/kg is recommended, provided that no loading dose was administered within 24 h and no routine dose was administered within 12 h of admission. (This study has been registered with Current Controlled Trials under registration number ISRCTN 50258054.).

Das D, Tripura R, Phyo AP, Lwin KM, Tarning J, Lee SJ, Hanpithakpong W, Stepniewska K, Menard D, Ringwald P et al. 2013. Effect of high-dose or split-dose artesunate on parasite clearance in artemisinin-resistant falciparum malaria. Clin Infect Dis, 56 (5), pp. e48-e58. | Citations: 35 (Scopus) | Show Abstract | Read more

BACKGROUND: The emergence of Plasmodium falciparum resistance to artemisinins on the Cambodian and Myanmar-Thai borders poses severe threats to malaria control. We investigated whether increasing or splitting the dose of the short-half-life drug artesunate improves parasite clearance in falciparum malaria in the 2 regions. METHODS: In Pailin, western Cambodia (from 2008 to 2010), and Wang Pha, northwestern Thailand (2009-2010), patients with uncomplicated falciparum malaria were randomized to oral artesunate 6 mg/kg/d as a once-daily or twice-daily dose for 7 days, or artesunate 8 mg/kg/d as a once-daily or twice-daily dose for 3 days, followed by mefloquine. Parasite clearance and recrudescence for up to 63 days of follow-up were assessed. RESULTS: A total of 159 patients were enrolled. Overall median (interquartile range [IQR]) parasitemia half-life (half-life) was 6.03 (4.89-7.28) hours in Pailin versus 3.42 (2.20-4.85) hours in Wang Pha (P = .0001). Splitting or increasing the artesunate dose did not shorten half-life in either site. Pharmacokinetic profiles of artesunate and dihydroartemisinin were similar between sites and did not correlate with half-life. Recrudescent infections occurred in 4 of 79 patients in Pailin and 5 of 80 in Wang Pha and was not different between treatment arms (P = .68). CONCLUSIONS: Increasing the artesunate treatment dose up to 8 mg/kg/d or splitting the dose does not improve parasite clearance in either artemisinin resistant or more sensitive infections with P. falciparum. Clinical Trials Registration. ISRCTN15351875.

Dondorp AM. 2013. Editorial commentary: single-dose primaquine as gametocytocidal treatment in patients with uncomplicated falciparum malaria. Clin Infect Dis, 56 (5), pp. 694-696. | Citations: 4 (European Pubmed Central) | Read more

Yeo TW, Lampah DA, Kenangalem E, Tjitra E, Price RN, Anstey NM. 2013. Impaired skeletal muscle microvascular function and increased skeletal muscle oxygen consumption in severe falciparum malaria. J Infect Dis, 207 (3), pp. 528-536. | Citations: 28 (Web of Science Lite) | Show Abstract | Read more

BACKGROUND: Organ dysfunction and tissue hypoxia in severe falciparum malaria result from an imbalance between oxygen delivery and demand. In severe malaria, microvascular obstruction from parasite sequestration decreases oxygen delivery. However, host microvascular function (defined as the capacity to increase oxygen delivery in response to ischemia) and oxygen consumption have not been assessed. METHODS: We used near-infrared resonance spectroscopy to measure thenar muscle microvascular function (StO(2)recov) and oxygen consumption (VO(2)) in 36 adults in Papua, Indonesia, with severe malaria, 33 with moderately severe malaria (MSM), 24 with severe sepsis, and 36 healthy controls. RESULTS: In the severe malaria group, the StO(2)recov of 2.7%/second was 16% and 22% lower than that in the MSM group (3.1%/second) and control group (3.5%/second), respectively (P < .001), and comparable to that in the severe sepsis group (2.5%/second). In the severe malaria group, StO(2)recov was inversely correlated with lactate level (r = -0.63; P < .001) and predicted death (area under the receiver operating characteristic curve, 0.71 [95% confidence interval {CI}, .51-.92]), with each percentage decrease associated with an increased odds of mortality (odds ratio, 2.49 [95% CI, 1.05-6.2]). Conversely, VO(2) increased in the severe malaria group by 18%, compared with levels in the control and severe sepsis groups (P < .001), and was associated with parasite biomass (r = 0.49; P = .04). CONCLUSIONS: Impaired microvascular function is associated with increased mortality among individuals with severe malaria, while oxygen consumption is increased. Tissue hypoxia may result not only from microvascular obstruction, but also from impaired ability of the microvasculature to match oxygen delivery to increased oxygen demand.

Hendriksen ICE, White LJ, Veenemans J, Mtove G, Woodrow C, Amos B, Saiwaew S, Gesase S, Nadjm B, Silamut K et al. 2013. Defining falciparum-malaria-attributable severe febrile illness in moderate-to-high transmission settings on the basis of plasma PfHRP2 concentration. J Infect Dis, 207 (2), pp. 351-361. | Citations: 45 (Web of Science Lite) | Show Abstract | Read more

BACKGROUND: In malaria-endemic settings, asymptomatic parasitemia complicates the diagnosis of malaria. Histidine-rich protein 2 (HRP2) is produced by Plasmodium falciparum, and its plasma concentration reflects the total body parasite burden. We aimed to define the malaria-attributable fraction of severe febrile illness, using the distributions of plasma P. falciparum HRP2 (PfHRP2) concentrations from parasitemic children with different clinical presentations. METHODS: Plasma samples were collected from and peripheral blood slides prepared for 1435 children aged 6-60 months in communities and a nearby hospital in northeastern Tanzania. The study population included children with severe or uncomplicated malaria, asymptomatic carriers, and healthy control subjects who had negative results of rapid diagnostic tests. The distributions of plasma PfHRP2 concentrations among the different groups were used to model severe malaria-attributable disease. RESULTS: The plasma PfHRP2 concentration showed a close correlation with the severity of infection. PfHRP2 concentrations of >1000 ng/mL denoted a malaria-attributable fraction of severe disease of 99% (95% credible interval [CI], 96%-100%), with a sensitivity of 74% (95% CI, 72%-77%), whereas a concentration of <200 ng/mL denoted severe febrile illness of an alternative diagnosis in >10% (95% CI, 3%-27%) of patients. Bacteremia was more common among patients in the lowest and highest PfHRP2 concentration quintiles. CONCLUSIONS: The plasma PfHRP2 concentration defines malaria-attributable disease and distinguishes severe malaria from coincidental parasitemia in African children in a moderate-to-high transmission setting.

Hanson J, Dondorp AM, Day NP, White NJ. 2013. Reply to Cunnington et al. J Infect Dis, 207 (2), pp. 370-371. | Citations: 1 (European Pubmed Central) | Read more

Nkhoma SC, Nair S, Al-Saai S, Ashley E, McGready R, Phyo AP, Nosten F, Anderson TJC. 2013. Population genetic correlates of declining transmission in a human pathogen. Mol Ecol, 22 (2), pp. 273-285. | Citations: 64 (Web of Science Lite) | Show Abstract | Read more

Pathogen control programs provide a valuable, but rarely exploited, opportunity to directly examine the relationship between population decline and population genetics. We investigated the impact of an ~12-fold decline in transmission on the population genetics of Plasmodium falciparum infections (n = 1731) sampled from four clinics on the Thai-Burma border over 10 years and genotyped using 96 genome-wide SNPs. The most striking associated genetic change was a reduction in the frequency of infections containing multiple parasite genotypes from 63% in 2001 to 14% in 2010 (P = 3 × 10(-15)). Two measures of the clonal composition of populations (genotypic richness and the β-parameter of the Pareto distribution) declined over time as more people were infected by parasites with identical multilocus genotypes, consistent with increased selfing and a reduction in the rate at which multilocus genotypes are broken apart by recombination. We predicted that the reduction in transmission, multiple clone carriage and outbreeding would be mirrored by an increased influence of genetic drift. However, geographical differentiation and expected heterozygosity remained stable across the sampling period. Furthermore, N(e) estimates derived from allele frequencies fluctuation between years remained high (582 to ∞) and showed no downward trend. These results demonstrate how genetic data can compliment epidemiological assessments of infectious disease control programs. The temporal changes in a single declining population parallel to those seen in comparisons of parasite genetics in regions of differing endemicity, strongly supporting the notion that reduced opportunity for outbreeding is the key driver of these patterns.

Newton PN, Day NPJ. 2013. Scrub Typhus pp. 542-545. | Read more

White NJ. 2013. Melioidosis and Glanders pp. 580-583. | Citations: 1 (Scopus) | Read more

Wuthiekanun V, Amornchai P, Paris DH, Langla S, Thaipadunpanit J, Chierakul W, Smythe LD, White NJ, Day NPJ, Limmathurotsakul D, Peacock SJ. 2013. Rapid isolation and susceptibility testing of Leptospira spp. using a new solid medium, LVW agar. Antimicrob Agents Chemother, 57 (1), pp. 297-302. | Citations: 17 (Scopus) | Show Abstract | Read more

Pathogenic Leptospira spp., the causative agents of leptospirosis, are slow-growing Gram-negative spirochetes. Isolation of Leptospira from clinical samples and testing of antimicrobial susceptibility are difficult and time-consuming. Here, we describe the development of a new solid medium that facilitates more-rapid growth of Leptospira spp. and the use of this medium to evaluate the Etest's performance in determining antimicrobial MICs to drugs in common use for leptospirosis. The medium was developed by evaluating the effects of numerous factors on the growth rate of Leptospira interrogans strain NR-20157. These included the type of base agar, the concentration of rabbit serum (RS), and the concentration and duration of CO(2) incubation during the initial period of culture. The highest growth rate of NR-20157 was achieved using a Noble agar base supplemented with 10% RS (named LVW agar), with an initial incubation at 30°C in 5% CO(2) for 2 days prior to continuous culture in air at 30°C. These conditions were used to develop the Etest for three species, L. interrogans (NR-20161), L. kirschnerii (NR-20327), and L. borgpetersenii (NR-20151). The MICs were read on day 7 for all samples. The Etest was then performed on 109 isolates of pathogenic Leptospira spp. The MIC(90) values for penicillin G, doxycycline, cefotaxime, ceftriaxone, and chloramphenicol were 0.64 units/ml and 0.19, 0.047, 0.5, and 2 μg/ml, respectively. The use of LVW agar, which enables rapid growth, isolation of single colonies, and simple antimicrobial susceptibility testing for Leptospira spp., provides an opportunity for new areas of fundamental and applied research.

Day NPJ. 2013. Panton-Valentine leucocidin and staphylococcal disease. Lancet Infect Dis, 13 (1), pp. 5-6. | Citations: 1 (European Pubmed Central) | Read more

Castonguay-Vanier J, Davong V, Bouthasavong L, Sengdetka D, Simmalavong M, Seupsavith A, Dance DAB, Baker S, Le Thi Phuong T, Vongsouvath M, Newton PN. 2013. Evaluation of a simple blood culture amplification and antigen detection method for diagnosis of Salmonella enterica serovar typhi bacteremia. J Clin Microbiol, 51 (1), pp. 142-148. | Citations: 13 (Scopus) | Show Abstract | Read more

In most areas where typhoid is endemic, laboratory diagnosis is not possible due to the lack of appropriate facilities. We investigated whether the combination of blood culture amplification of Salmonella enterica serovar Typhi with an S. Typhi antigen rapid diagnostic test (RDT) could be an accurate and inexpensive tool for the accelerated diagnosis of patients with acute typhoid in Laos. For a panel of 23 Gram-negative reference pathogens, the Standard Diagnostics (catalog no. 15FK20; Kyonggi-do, South Korea) RDT gave positive results for S. Typhi NCTC 8385, S. Typhi NCTC 786 (Vi negative), Salmonella enterica serovar Enteritidis (ATCC 13076), and Salmonella enterica serovar Ndolo NCTC 8700 (all group D). In a prospective study of 6,456 blood culture bottles from 3,028 patients over 15 months, 392 blood culture bottles (6.1%) from 221 (7.3%) patients had Gram-negative rods (GNRs) seen in the blood culture fluid. The sensitivity, negative predictive value, specificity, and positive predictive value were 96.7%, 99.5%, 97.9%, and 87.9%, respectively, for patients with proven S. Typhi bacteremia and 91.2%, 98.4%, 98.9%, and 93.9% for patients with group D Salmonella. The median (range) number of days between diagnosis by RDT and reference assays was 1 (-1 to +2) day for those with confirmed S. Typhi. The use of antigen-based pathogen detection in blood culture fluid may be a useful, relatively rapid, inexpensive, and accurate technique for the identification of important causes of bacteremia in the tropics.

Wootton CI, Elliott IAM, Sengdetkha D, Vongsouvath M, Phongmany S, Dance DAB. 2013. Melioidosis: an unusual cause of recurrent buttock abscesses. Clin Exp Dermatol, 38 (4), pp. 427-428. | Read more

Stoesser N, Pocock J, Moore CE, Soeng S, Hor P, Sar P, Limmathurotsakul D, Day N, Kumar V, Khan S et al. 2013. The epidemiology of pediatric bone and joint infections in Cambodia, 2007-11. J Trop Pediatr, 59 (1), pp. 36-42. | Citations: 11 (Web of Science Lite) | Show Abstract | Read more

There are limited data on osteoarticular infections from resource-limited settings in Asia. A retrospective study of patients presenting to the Angkor Hospital for Children, Cambodia, January 2007-July 2011, identified 81 cases (28% monoarticular septic arthritis, 51% single-limb osteomyelitis and 15% multisite infections). The incidence was 13.8/100 000 hospital attendances. The median age was 7.3 years, with a male/female ratio of 1.9:1; 35% presented within 5 days of symptom onset (median 7 days). Staphylococcus aureus was cultured in 29 (36%) cases (52% of culture-positive cases); one isolate was methicillin-resistant (MRSA). Median duration of antimicrobial treatment was 29 days (interquartile range 21-43); rates of surgical intervention were 96%, and 46% of children had sequelae, with one fatality. In this setting osteoarticular infections are relatively common with high rates of surgical intervention and sequelae. Staphylococcus aureus is the commonest culturable cause, but methicillin-resistant S. aureus is not a major problem, unlike in other Asian centers.

Siengsanan-Lamont J, Robertson ID, Blacksell SD, Ellis T, Saengchoowong S, Suwanpukdee S, Yongyuttawichai P, Cheewajorn K, Jangjaras J, Chaichoun K et al. 2013. A study of risk factors for infection with HPAI H5N1 in small poultry farms in Thailand using a questionnaire survey. Zoonoses Public Health, 60 (3), pp. 209-214. | Citations: 1 (European Pubmed Central) | Show Abstract | Read more

A questionnaire was used to collect data on small poultry farm management and wild bird observed in poultry keeping areas to identify putative risk factors for infection with HPAI H5N1. The study was conducted in 2008 in four subdistricts of central Thailand that had experienced outbreaks of HPAI H5N1 in poultry. Descriptive and inferential analyses including univariable analyses and multivariable logistic regression were used to identify putative risk factors. Risk factors included purchasing native chickens/fighting cocks from commercial hatcheries, replacing or restocking birds individually, and observing lesser whistling ducks (Dendrocygna javanica) on the farm daily. Selecting healthy animals when purchasing animals to ensure that they were disease free was a protective factor. To fully understand the epidemiology of infection of small poultry farms with HPAI H5N1, control of movement of domestic poultry and serological and virological testing of the poultry population should be applied.

Pan A, Lee A, Cooper B, Chalfine A, Daikos GL, Garilli S, Goossens H, Malhotra-Kumar S, Martínez JA, Patroni A et al. 2013. Risk factors for previously unknown meticillin-resistant Staphylococcus aureus carriage on admission to 13 surgical wards in Europe. J Hosp Infect, 83 (2), pp. 107-113. | Citations: 12 (Web of Science Lite) | Show Abstract | Read more

BACKGROUND: Early identification of meticillin-resistant Staphylococcus aureus (MRSA) carriers may be helpful for clinical and epidemiological reasons. AIM: To identify and compare risk factors of previously unknown MRSA carriage on admission to 13 surgical wards in France, Greece, Italy, and Spain. METHODS: The study was a prospective observational cohort study which enrolled consecutive patients screened for MRSA on admission to surgical wards. Sociodemographic data, comorbidities and possible risk factors for MRSA were recorded. A multivariate logistic regression model was used to predict probabilities of previously unknown MRSA colonization on admission based on patient characteristics. Prediction rules for MRSA carriage were developed and evaluated using the c-statistic. FINDINGS: Of 2901 patients enrolled, admission screening identified 111 (3.8%) new MRSA carriers. Independent risk factors for MRSA carriage were urinary catheterization (odds ratio: 4.4; 95% confidence interval: 2.0-9.9), nursing home residency (3.8; 1.9-7.7), chronic skin disease (2.9; 1.5-5.8), wounds/ulcers (2.4; 1.5-4.0), recent hospitalization (2.2; 1.5-3.3), diabetes (1.6, 1.02-2.5), and age >70 years (1.5; 1.03-2.3). However, risk factors varied between centres. The c-statistic for the common prediction rule for all centres was 0.64, indicating limited predictive power. CONCLUSIONS: Risk profiles for MRSA carriers vary between surgical wards in European countries. Identifying local risk factors is important, as a common European prediction rule was found to be of limited clinical value.

Harris JP, Lopman BA, Cooper BS, O'Brien SJ. 2013. Correction. BMJ Open, 3 (8), | Citations: 3 (Web of Science Lite) | Read more

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