We used microarrays and transcriptional profiling of peripheral blood to investigate the host response of 29 individuals who contracted typhoid fever in the Mekong Delta region of Vietnam. Samples were taken over a nine month period encompassing acute disease, convalescence, and recovery. We found that typhoid fever induced a distinct and highly reproducible signature in the peripheral blood that changed during treatment and convalescence, returning in the majority of cases to the "normal" profile as measured in healthy uninfected controls. Unexpectedly, there was a strong, distinct signature of convalescence present at day 9 after infection that remained virtually unchanged one month after acute infection and in some cases persisted as long as nine months despite a complete clinical recovery in all patients. Patients who retain the convalescent signature may be genetically or temporarily incapable of developing an effective immune response and may be more susceptible to reinfection, relapse, or the establishment of a carrier state.
The PCR primers commonly used to detect Plasmodium knowlesi infections in humans were found to cross-react stochastically with P. vivax genomic DNA. A nested primer set that targets one of the P. knowlesi small-subunit rRNA genes was validated for specificity and for sensitivity of detection of <10 parasite genomes.
Clin Microbiol Infect, 15 Suppl 2 (SUPPL. 2), pp. 95-97. | Citations: 17 (European Pubmed Central) | Read more2009. Molecular detection of Bartonella species in rodents from the Lao PDR.
The PCR primers commonly used to detect Plasmodium knowlesi infections in humans were found to cross-react stochastically with P. vivax genomic DNA. A nested primer set that targets one of the P. knowlesi small-subunit rRNA genes was validated for specificity and for sensitivity of detection of <10 parasite genomes. Copyright © 2009, American Society for Microbiology. All Rights Reserved.
Clin Infect Dis, 49 (11), pp. 1638-1640. | Citations: 14 (Scopus) | Read more2009. Artemisinin combination therapy for malaria: beyond good efficacy.
PLOS MEDICINE, 6 (12), pp. e1000207-e1000207. | Citations: 209 (Web of Science Lite) | Read more2009. The Severity of Pandemic H1N1 Influenza in the United States, from April to July 2009: A Bayesian Analysis
Staphylococcal and streptococcal infections are common infectious diseases and can range from mild, superficial skin infections to severe, life-threatening systemic infections. Staphylococcus aureus, group A streptococcus, and Streptococcus pneumoniae are the three major pathogens. The prevalence of invasive infections caused by community-associated meticillin-resistant S. aureus and group A streptococcus appears to be increasing. The emergence of drug resistance (e.g. meticillin and glycopeptide resistance in S. aureus, macrolide resistance in group A streptococci and penicillin resistance in S. pneumoniae), is concerning and a potential threat to successful treatment. Streptococcus suis has recently emerged as an important human pathogen. © 2009 Elsevier Ltd. All rights reserved.
BACKGROUND: Hemolysis causes anemia in falciparum malaria, but its contribution to microvascular pathology in severe malaria (SM) is not well characterized. In other hemolytic diseases, release of cell-free hemoglobin causes nitric oxide (NO) quenching, endothelial activation, and vascular complications. We examined the relationship of plasma hemoglobin and myoglobin to endothelial dysfunction and disease severity in malaria. METHODS: Cell-free hemoglobin (a potent NO quencher), reactive hyperemia peripheral arterial tonometry (RH-PAT) (a measure of endothelial NO bioavailability), and measures of perfusion and endothelial activation were quantified in adults with moderately severe (n = 78) or severe (n = 49) malaria and control subjects (n = 16) from Papua, Indonesia. RESULTS: Cell-free hemoglobin concentrations in patients with SM (median, 5.4 micromol/L; interquartile range [IQR], 3.2-7.4 micromol/L) were significantly higher than in those with moderately severe malaria (2.6 micromol/L; IQR, 1.3-4.5 micromol/L) or controls (1.2 micromol/L; IQR, 0.9-2.4 micromol/L; P < .001). Multivariable regression analysis revealed that cell-free hemoglobin remained inversely associated with RH-PAT, and in patients with SM, there was a significant longitudinal association between improvement in RH-PAT index and decreasing levels of cell-free hemoglobin (P = .047). Cell-free hemoglobin levels were also independently associated with lactate, endothelial activation, and proinflammatory cytokinemia. CONCLUSIONS: Hemolysis in falciparum malaria results in NO quenching by cell-free hemoglobin, and may exacerbate endothelial dysfunction, adhesion receptor expression and impaired tissue perfusion. Treatments that increase NO bioavailability may have potential as adjunctive therapies in SM.
Malaria pigment is an intracellular inclusion body that appears in blood and tissue specimens on microscopic examination and can help in establishing the diagnosis of malaria. In simple light microscopy, it can be difficult to discern from cellular background and artifacts. It has long been known that if polarized light microscopy is used, malaria pigment can be much easier to distinguish. However, this technique is rarely used because of the need for a relatively costly polarization microscope. We describe a simple and economical technique to convert any standard light microscope suitable for examination of malaria films into a polarization microscope.
Health-care-associated methicillin-resistant Staphylococcus aureus (MRSA) infection may cause increased hospital stay, or sometimes death. Quantifying this effect is complicated because the exposure is time dependent: infection may prolong hospital stay, while longer stays increase infection risk. In this paper, the authors overcome these problems by using a multinomial longitudinal model to estimate the daily probability of death and discharge. They then extend the basic model to estimate how the effect of MRSA infection varies over time and to quantify number of excess days in the intensive care unit due to infection. They found that infection decreased the relative risk of discharge (relative risk ratio = 0.68, 95% credible interval: 0.54, 0.82). Infection on the first day of admission resulted in a mean extra stay of 0.3 days (95% credible interval: 0.1, 0.5) for a patient with an Acute Physiology and Chronic Health Evaluation II score of 10 and 1.2 days (95% credible interval: 0.5, 2.0) for a patient with a score of 30. The decrease in the relative risk of discharge remained fairly constant with day of MRSA infection but was slightly stronger closer to the start of infection. Results confirm the importance of MRSA infection in increasing stay in an intensive care unit but suggest that previous work may have systematically overestimated the effect size.
We tracked the effective reproductive number (Rt) over time to assess the impact of important public health control measures in the five most SARS-affected geographic areas in mainland China. As soon as the Chinese authorities gained full control of all activities to combat SARS, Rt decreased dramatically and consistently below one. Many control measures that seriously affected public life were implemented afterwards, i.e., when the epidemic was already dying down.
OBJECTIVE: To quantify the transmissibility of severe acute respiratory syndrome (SARS) in hospitals in mainland China and to assess the effectiveness of control measures. METHODS: We report key epidemiological details of three major hospital outbreaks of SARS in mainland China, and estimate the evolution of the effective reproduction number in each of the three hospitals during the course of the outbreaks. RESULTS: The three successive hospital outbreaks infected 41, 99 and 91 people of whom 37%, 60% and 70% were hospital staff. These cases resulted in 33 deaths, five of which occurred in hospital staff. In a multivariate logistic regression, age and whether or not the case was a healthcare worker (HCW) were found to be significant predictors of mortality. The estimated effective reproduction numbers (95% CI) for the three epidemics peaked at 8 (5, 11), 9 (4, 14) and 12 (7, 17). In all three hospitals the epidemics were rapidly controlled, bringing the reproduction number below one within 25, 10 and 5 days respectively. CONCLUSIONS: This work shows that in three major hospital epidemics in Beijing and Tianjin substantially higher rates of transmission were initially observed than those seen in the community. In all three cases the hospital epidemics were rapidly brought under control, with the time to successful control becoming shorter in each successive outbreak.
Lancet, 374 (9700), pp. 1478-1480. | Citations: 6 (European Pubmed Central) | Read more2009. Intermittent preventive treatment of malaria in infancy.
NEW ENGLAND JOURNAL OF MEDICINE, 361 (18), pp. 1808-1808. | Citations: 4 (Web of Science Lite)2009. Artemisinin Resistance in Plasmodium falciparum Malaria REPLY
NEW ENGLAND JOURNAL OF MEDICINE, 361 (17), pp. 1714-1714. | Citations: 10 (Web of Science Lite) | Read more2009. Artemisinin Resistance in Plasmodium falciparum Malaria (vol 361, pg 455, 2009)
Melioidosis is an infectious disease with a propensity for relapse, despite prolonged antibiotic eradication therapy for 12 to 20 weeks. A pharmacokinetic (PK) simulation study was performed to determine the optimal dosing of cotrimoxazole (trimethoprim-sulfamethoxazole [TMP-SMX]) used in current eradication regimens in Thailand and Australia. Data for bioavailability, protein binding, and coefficients of absorption and elimination were taken from published literature. Apparent volumes of distribution were correlated with body mass and were estimated separately for Thai and Australian populations. In vitro experiments demonstrated concentration-dependent killing. In Australia, the currently used eradication regimen (320 [TMP]/1,600 [SMX] mg every 12 h [q12h]) was predicted to achieve the PK-pharmacodynamic (PD) target (an area under the concentration-time curve from 0 to 24 h/MIC ratio of >25 for both TMP and SMX) for strains with the MIC90 of Australian strains (< or = 1/19 mg/liter). In Thailand, the former regimen of 160/800 mg q12h would not be expected to attain the target for strains with an MIC of > or = 1/19 mg/liter, but the recently implemented weight-based regimen (<40 kg [body weight], 160/800 mg q12h; 40 to 60 kg, 240/1,200 mg q12h; >60 kg, 320/1,600 mg q12h) would be expected to achieve adequate concentrations for strains with an MIC of < or = 1/19 mg/liter. The results were sensitive to the variance of the PK parameters. Prospective PK-PD studies of Asian populations are needed to optimize TMP-SMX dosing in melioidosis.
A prospective study in Thailand identified 106 patients with culture-proven leptospirosis. The accuracy of the microscopic agglutination test (MAT) in predicting the infecting serovar was evaluated in 78/106 (74%) patients with a diagnostic titer. MAT correctly determined the infecting serovar in 26 cases (33%), indicating that this assay is a poor predictor of infecting serovar in our setting.
BACKGROUND: Castor oil is one of the most popular drugs for induction of labour in a non-medical setting; however, published data on safety and effectiveness of this compound to induce labour remain sparse. AIM: To assess the safety and effectiveness of castor oil for induction of labour in pregnancies with an ultrasound estimated gestational at birth of more than 40 weeks. METHODS: Data were extracted from hospital-based records of all pregnant women who attended antenatal clinics on the Thai-Burmese border and who were more than 40 weeks pregnant. The effectiveness of castor oil to induce labour was expressed as time to birth and analysed with a Cox proportional hazards regression model. Measures associated with safety were fetal distress, meconium-stained amniotic fluid, tachysystole of the uterus, uterine rupture, abnormal maternal blood pressure during labour, Apgar scores, neonatal resuscitation, stillbirth, post-partum haemorrhage, severe diarrhoea and maternal death. Proportions were compared using Fisher's exact test. RESULTS: Of 612 women with a gestation of more than 40 weeks, 205 received castor oil for induction and 407 did not. The time to birth was not significantly different between the two groups (hazard ratio 0.99 (95% confidence interval: 0.81 to 1.20; n = 509)). Castor oil use was not associated with any harmful effects on the mother or fetus. CONCLUSIONS: Castor oil for induction of labour had no effect on time to birth nor were there any harmful effects observed in this large series. Our findings leave no justification for recommending castor oil for this purpose.
OBJECTIVES: Ultrasound examination of the fetus is a powerful tool for assessing gestational age and detecting obstetric problems but is rarely available in developing countries. The aim of this study was to assess the intraobserver and interobserver agreement of fetal biometry by locally trained health workers in a refugee camp on the Thai-Burmese border. METHODS: One expatriate doctor and four local health workers participated in the study, which included examinations performed on every fifth pregnant woman with a singleton pregnancy between 16 and 40 weeks' gestation, and who had undergone an early dating ultrasound scan, attending the antenatal clinic in Maela refugee camp. At each examination, two examiners independently measured biparietal diameter (BPD), head circumference (HC), abdominal circumference (AC) and femur length (FL), with one of the examiners obtaining duplicate measurements of each parameter. Intraobserver measurement error was assessed using the intraclass correlation coefficient (ICC) and interobserver error was assessed by the Bland and Altman 95% limits of agreement method. RESULTS: A total of 4188 ultrasound measurements (12 per woman) were obtained in 349 pregnancies at a median gestational age of 27 (range, 16-40) weeks in 2008. The ICC for BPD, HC, AC and FL was greater than 0.99 for all four trainees and the doctor (range, 0.996-0.998). For gestational ages between 18 and 24 weeks, interobserver 95% limits of agreement corresponding to differences in estimated gestational age of less than +/- 1 week were calculated for BPD, HC, AC and FL. Measurements by local health workers showed high levels of agreement with those of the expatriate doctor. CONCLUSIONS: Locally trained health workers working in a well organized unit with ongoing quality control can obtain accurate fetal biometry measurements for gestational age estimation. This experience suggests that training of local health workers in developing countries is possible and could allow effective use of obstetric ultrasound imaging.
Med Sci (Paris), 25 (10), pp. 867-869. | Citations: 2 (European Pubmed Central) | Read more2009. [Malaria in pregnancy: a therapeutic dilemma].
PURPOSE OF REVIEW: Unlike Plasmodium falciparum, Plasmodium vivax rarely causes severe disease in healthy travellers or in temperate endemic regions and has been regarded as readily treatable with chloroquine. However, in tropical areas, recent reports have highlighted severe and fatal disease associated with P. vivax infection. We review the evidence for severe disease and the spread of drug-resistant P. vivax and speculate how these maybe related. RECENT FINDINGS: Studies from Indonesia, Papua New Guinea, Thailand and India have shown that 21-27% of patients with severe malaria have P. vivax monoinfection. The clinical spectrum of these cases is broad with an overall mortality of 0.8-1.6%. Major manifestations include severe anaemia and respiratory distress, with infants being particularly vulnerable. Most reports of severe and fatal vivax malaria come from endemic regions where populations have limited access to healthcare, a high prevalence of comorbidity and where drug-resistant P. vivax strains and partially effective primaquine regimens significantly undermine the radical cure and control of this relapsing infection. The mechanisms underlying severe disease in vivax malaria remain poorly defined. SUMMARY: Severe, fatal and multidrug-resistant vivax malaria challenge our perception of P. vivax as a benign disease. Strategies to understand and address these phenomena are needed urgently if the global elimination of malaria is to succeed.
ASIAN PACIFIC JOURNAL OF TROPICAL MEDICINE, 2 (5), pp. 14-19.2009. SDS-PAGE analysis of whole cell protein and outer membrane protein patterns of clinical isolates of Burkholderia pseudomallei
Artemether-lumefantrine has become one of the most widely used antimalarial drugs in the world. The objective of this study was to determine the population pharmacokinetic properties of lumefantrine in pregnant women with uncomplicated multidrug-resistant Plasmodium falciparum malaria on the northwestern border of Thailand. Burmese and Karen women (n = 103) with P. falciparum malaria and in the second and third trimesters of pregnancy were treated with artemether-lumefantrine (80/480 mg) twice daily for 3 days. All patients provided five capillary plasma samples for drug quantification, and the collection times were randomly distributed over 14 days. The concentration-time profiles of lumefantrine were assessed by nonlinear mixed-effects modeling. The treatment failure rate (PCR-confirmed recrudescent infections at delivery) was high; 16.5% (95% confidence interval, 9.9 to 25.1). The population pharmacokinetics of lumefantrine were described well by a two-compartment open model with first-order absorption and elimination. The final model included interindividual variability in all pharmacokinetic parameters and a linear covariate relationship between the estimated gestational age and the central volume of distribution. A high proportion of all women (40%, 41/103) had day 7 capillary plasma concentrations of <355 ng/ml (which corresponds to approximately <280 ng/ml in venous plasma), a threshold previously associated with an increased risk of therapeutic failure in nonpregnant patients in this area. Predictive modeling suggests that a twice-daily regimen given for 5 days would be preferable in later pregnancy. In conclusion, altered pharmacokinetic properties of lumefantrine contribute to the high rates of failure of artemether-lumefantrine treatment in later pregnancy. Dose optimization is urgently needed.
It is well understood that sociocultural practices strongly influence Taenia solium transmission; however, the extent to which interspecific parasite competition moderates Taenia transmission has yet to be determined. This is certainly the case in Southeast Asia where T. solium faces competition in both the definitive host (people) and the intermediate host (pigs). In people, adult worms of T. solium, T. saginata and T. asiatica compete through density-dependent crowding mechanisms. In pigs, metacestodes of T. solium, T. hydatigena and T. asiatica compete through density-dependent immune-mediated interactions. Here, we describe the biological and epidemiological implications of Taenia competition and propose that interspecific competition has a moderating effect on the transmission dynamics of T. solium in the region. Furthermore, we argue that this competitive ecological scenario should be considered in future research and surveillance activities examining T. solium cysticercosis and taeniasis in Southeast Asia.
BACKGROUND: Progress in therapy for cryptococcal meningitis has been slow because of the lack of a suitable marker of treatment response. Previously, we demonstrated the statistical power of a novel endpoint, the rate of clearance of infection, based on serial quantitative cultures of cerebrospinal fluid, to differentiate the fungicidal activity of alternative antifungal drug regimens. We hypothesized that the rate of clearance of infection should also be a clinically meaningful endpoint. METHODS: We combined data from cohorts of patients with human immunodeficiency virus-associated cryptococcal meningitis from Thailand, South Africa, and Uganda, for whom the rate of clearance of infection was determined, and clinical and laboratory data prospectively collected, and explored the association between the rate of clearance of infection and mortality by Cox survival analyses. RESULTS: The combined cohort comprised 262 subjects. Altered mental status at presentation, a high baseline organism load, and a slow rate of clearance of infection were independently associated with increased mortality at 2 and 10 weeks. Rate of clearance of infection was associated with antifungal drug regimen and baseline cerebrospinal fluid interferon-gamma levels. CONCLUSIONS: The results support the use of the rate of clearance of infection or early fungicidal activity as a means to explore antifungal drug dosages and combinations in phase II studies. An increased understanding of how the factors determining outcome interrelate may help clarify opportunities for intervention.
J Infect, 59 (3), pp. 219-222. | Citations: 3 (Web of Science Lite) | Read more2009. Apolipoprotein E-epsilon2 confers risk of pulmonary tuberculosis in women from the Indian subcontinent--a preliminary study.
Ultrasound in Obstetrics & Gynecology, 34 (S1), pp. 13-13. | Read more2009. OC08.01: Quality assessment of fetal biometry in locally trained sonographers in a developing country setting
BACKGROUND: The incidence of community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) infection is rising in the developed world but appears to be rare in developing countries. One explanation for this difference is that resource poor countries lack the diagnostic microbiology facilities necessary to detect the presence of CA-MRSA carriage and infection. METHODOLOGY AND PRINCIPAL FINDINGS: We developed diagnostic microbiology capabilities at the Angkor Hospital for Children, Siem Reap, western Cambodia in January 2006 and in the same month identified a child with severe community-acquired impetigo caused by CA-MRSA. A study was undertaken to identify and describe additional cases presenting between January 2006 and December 2007. Bacterial isolates underwent molecular characterization using multilocus sequence typing, staphylococcal cassette chromosome mec (SCCmec) typing, and PCR for the presence of the genes encoding Panton-Valentine Leukocidin (PVL). Seventeen children were identified with CA-MRSA infection, of which 11 had skin and soft tissue infection and 6 had invasive disease. The majority of cases were unrelated in time or place. Molecular characterization identified two independent MRSA clones; fifteen isolates were sequence type (ST) 834, SCCmec type IV, PVL gene-negative, and two isolates were ST 121, SCCmec type V, PVL gene-positive. CONCLUSIONS: This represents the first ever report of MRSA in Cambodia, spread of which would pose a significant threat to public health. The finding that cases were mostly unrelated in time or place suggests that these were sporadic infections in persons who were CA-MRSA carriers or contacts of carriers, rather than arising in the context of an outbreak.
BACKGROUND: Clinical management of malaria is a major health issue in sub-Saharan Africa. New strategies based on intermittent preventive treatment (IPT) can tackle disease burden by simultaneously reducing frequency of infections and life-threatening illness in infants (IPTi) and children (IPTc), while allowing for immunity to build up. However, concerns as to whether immunity develops efficiently in treated individuals, and whether there is a rebound effect after treatment is halted, have made it imperative to define the effects that IPTi and IPTc exert on the clinical malaria scenario. METHODS AND FINDINGS: Here, we simulate several schemes of intervention under different transmission settings, while varying immunity build up assumptions. Our model predicts that infection risk and effectiveness of acquisition of clinical immunity under prophylactic effect are associated to intervention impact during treatment and follow-up periods. These effects vary across regions of different endemicity and are highly correlated with the interplay between the timing of interventions in age and the age dependent risk of acquiring an infection. However, even when significant rebound effects are predicted to occur, the overall intervention impact is positive. CONCLUSIONS: IPTi is predicted to have minimal impact on the acquisition of clinical immunity, since it does not interfere with the occurrence of mild infections, thus failing to reduce the underlying force of infection. On the contrary, IPTc has a significant potential to reduce transmission, specifically in areas where it is already low to moderate.
Recurrent melioidosis can be caused by two different mechanisms: relapse or re-infection. We examined the pattern of organ involvement in the first and second episodes in individual patients. Evaluation of 140 patients with recurrence showed that similar patterns of disease occurred during the first and second episode, independent of whether this was caused by relapse or re-infection. Copyright © 2009 by The American Society of Tropical Medicine and Hygiene.
Eur J Clin Pharmacol, 65 (8), pp. 847-847. | Read more2009. The pharmacokinetics of artemether and lumefantrine in pregnant women with uncomplicated falciparum malaria.
Neorickettsia sennetsu has been described from Japan and Malaysia, causing a largely forgotten infectious mononucleosis-like disease. Because it is believed to be contracted from eating raw fish, frequently consumed in the Lao PDR, we looked for evidence of N. sennetsu among Lao patients and fish. A buffy coat from 1 of 91 patients with undifferentiated fever was positive by 16S rRNA amplification and sequencing and real-time polymerase chain reactions (PCR) targeting two N. sennetsu genes. Lao blood donors and patients with fever, hepatitis, or jaundice (N = 1,132) had a high prevalence (17%) of immunofluorescence assay IgG anti-N. sennetsu antibodies compared with 4% and 0% from febrile patients (N = 848) in Thailand and Malaysia, respectively. We found N. sennetsu DNA by PCR, for the first time, in a fish (Anabas testudineus). These data suggest that sennetsu may be an under-recognized cause of fever and are consistent with the hypothesis that it may be contracted from eating raw fish.
Recurrent melioidosis can be caused by two different mechanisms: relapse or re-infection. We examined the pattern of organ involvement in the first and second episodes in individual patients. Evaluation of 140 patients with recurrence showed that similar patterns of disease occurred during the first and second episode, independent of whether this was caused by relapse or re-infection.
Classical swine fever (CSF) is a highly contagious and severe viral disease of swine resulting in substantial production losses in different farming systems in many regions of the world. The accurate and rapid detection of CSF outbreaks is reliant on sensitive and specific laboratory testing and is a key component of disease control. Specific detection of CSF virus can be achieved by virus isolation in tissue culture, antigen capture or the detection of viral RNA using molecular techniques. In order to reduce the time taken to achieve a diagnostic result and simplify testing methods, an antigen capture ELISA using immunomagnetic beads (IMB) as the solid phase was developed and compared to a microplate-based antigen capture (AC)-ELISA. The IMB-ELISA has up to 64-fold greater analytical sensitivity than the AC-ELISA and initial estimates of diagnostic sensitivity and specificity are 100%. The IMB-ELISA has a highly robust, rapid and stable test format and is simpler to perform than the AC-ELISA. The IMB-ELISA has the added advantage that a result can be sensitively and specifically determined by eye, lending it to the possibility of adaptation to a near-to-field test with minimal equipment or expertise needed.
Trends Parasitol, 25 (8), pp. 350-351. | Citations: 3 (European Pubmed Central) | Read more2009. Fluorescein angiography findings strengthen the theoretical basis for trialling neuroprotective agents in cerebral malaria.
BACKGROUND: Artemisinin-based combination therapies are the recommended first-line treatments of falciparum malaria in all countries with endemic disease. There are recent concerns that the efficacy of such therapies has declined on the Thai-Cambodian border, historically a site of emerging antimalarial-drug resistance. METHODS: In two open-label, randomized trials, we compared the efficacies of two treatments for uncomplicated falciparum malaria in Pailin, western Cambodia, and Wang Pha, northwestern Thailand: oral artesunate given at a dose of 2 mg per kilogram of body weight per day, for 7 days, and artesunate given at a dose of 4 mg per kilogram per day, for 3 days, followed by mefloquine at two doses totaling 25 mg per kilogram. We assessed in vitro and in vivo Plasmodium falciparum susceptibility, artesunate pharmacokinetics, and molecular markers of resistance. RESULTS: We studied 40 patients in each of the two locations. The overall median parasite clearance times were 84 hours (interquartile range, 60 to 96) in Pailin and 48 hours (interquartile range, 36 to 66) in Wang Pha (P<0.001). Recrudescence confirmed by means of polymerase-chain-reaction assay occurred in 6 of 20 patients (30%) receiving artesunate monotherapy and 1 of 20 (5%) receiving artesunate-mefloquine therapy in Pailin, as compared with 2 of 20 (10%) and 1 of 20 (5%), respectively, in Wang Pha (P=0.31). These markedly different parasitologic responses were not explained by differences in age, artesunate or dihydroartemisinin pharmacokinetics, results of isotopic in vitro sensitivity tests, or putative molecular correlates of P. falciparum drug resistance (mutations or amplifications of the gene encoding a multidrug resistance protein [PfMDR1] or mutations in the gene encoding sarco-endoplasmic reticulum calcium ATPase6 [PfSERCA]). Adverse events were mild and did not differ significantly between the two treatment groups. CONCLUSIONS: P. falciparum has reduced in vivo susceptibility to artesunate in western Cambodia as compared with northwestern Thailand. Resistance is characterized by slow parasite clearance in vivo without corresponding reductions on conventional in vitro susceptibility testing. Containment measures are urgently needed. (ClinicalTrials.gov number, NCT00493363, and Current Controlled Trials number, ISRCTN64835265.)
BACKGROUND: Counterfeit oral artesunate has been a major public health problem in mainland SE Asia, impeding malaria control. A countrywide stratified random survey was performed to determine the availability and quality of oral artesunate in pharmacies and outlets (shops selling medicines) in the Lao PDR (Laos). METHODS: In 2003, 'mystery' shoppers were asked to buy artesunate tablets from 180 outlets in 12 of the 18 Lao provinces. Outlets were selected using stratified random sampling by investigators not involved in sampling. Samples were analysed for packaging characteristics, by the Fast Red Dye test, high-performance liquid chromatography (HPLC), mass spectrometry (MS), X-ray diffractometry and pollen analysis. RESULTS: Of 180 outlets sampled, 25 (13.9%) sold oral artesunate. Outlets selling artesunate were more commonly found in the more malarious southern Laos. Of the 25 outlets, 22 (88%; 95%CI 68-97%) sold counterfeit artesunate, as defined by packaging and chemistry. No artesunate was detected in the counterfeits by any of the chemical analysis techniques and analysis of the packaging demonstrated seven different counterfeit types. There was complete agreement between the Fast Red dye test, HPLC and MS analysis. A wide variety of wrong active ingredients were found by MS. Of great concern, 4/27 (14.8%) fakes contained detectable amounts of artemisinin (0.26-115.7 mg/tablet). CONCLUSION: This random survey confirms results from previous convenience surveys that counterfeit artesunate is a severe public health problem. The presence of artemisinin in counterfeits may encourage malaria resistance to artemisinin derivatives. With increasing accessibility of artemisinin-derivative combination therapy (ACT) in Laos, the removal of artesunate monotherapy from pharmacies may be an effective intervention.
A specific retinopathy has been described in African children with cerebral malaria, but in adults this has not been extensively studied. Since the structure and function of the retinal vasculature greatly resembles the cerebral vasculature, study of retinal changes can reveal insights into the pathophysiology of cerebral malaria. A detailed observational study of malarial retinopathy in Bangladeshi adults was performed using high-definition portable retinal photography. Retinopathy was present in 17/27 adults (63%) with severe malaria and 14/20 adults (70%) with cerebral malaria. Moderate or severe retinopathy was more frequent in cerebral malaria (11/20, 55%) than in uncomplicated malaria (3/15, 20%; P=0.039), bacterial sepsis (0/5, 0%; P=0.038) or healthy controls (0/18, 0%; P<0.001). The spectrum of malarial retinopathy was similar to that previously described in African children, but no vessel discolouration was observed. The severity of retinal whitening correlated with admission venous plasma lactate (P=0.046), suggesting that retinal ischaemia represents systemic ischaemia. In conclusion, retinal changes related to microvascular obstruction were common in adults with severe falciparum malaria and correlated with disease severity and coma, suggesting that a compromised microcirculation has important pathophysiological significance in severe and cerebral malaria. Portable retinal photography has potential as a valuable tool to study malarial retinopathy.
The pathophysiology of coma in cerebral malaria (CM) is not well understood. Obstruction of microcirculatory flow is thought to play a central role, but other hypotheses include roles for parasite- and host-derived factors such as immune mediators, and for increased blood-brain barrier permeability leading to raised intracranial pressure. The retinal vasculature is a direct extension of the cerebral vasculature. It is the only vascular bed easily accessible for visualisation and provides a unique opportunity to observe vascular pathology and its effect on neurological tissue. A specific retinopathy has been well described in African children with CM and its severity correlates with outcome. This retinopathy has been less well described in adults. The central mechanism causing malarial retinopathy appears to be microvascular obstruction, which has been demonstrated in affected retinas by fluorescein angiography. The presence in a central nervous system tissue of microvascular obstruction strongly supports the hypothesis that the sequestration of erythrocytes in small blood vessels and consequent obstruction of microcirculatory flow is an important mechanism causing coma and death in CM. Despite advances in the antimalarial treatment of severe malaria, its mortality remains approximately 15-20%. Adjunctive treatment targeting sequestration is a promising strategy to further lower mortality.
Wild bird surveillance for highly pathogenic avian influenza (HPAI) H5N1 virus from 2004 to 2007 in Thailand indicated that the prevalence of infection with avian influenza H5N1 virus in wild birds was low (1.0%, 95% confidence interval [CI]: 0.7-1.2, 60/6,263 pooled samples). However, the annual prevalence varied considerably over this period, with a peak of 2.7% (95% CI: 1.4, 4.1) in 2004. Prevalence dropped to 0.5% (95% CI: 0.3, 0.8]) and 0.6% (95% CI: 0.3, 1.0) in 2005 and 2006, respectively, and then increased to 1.8% (95% CI: 1.0, 2.6) in 2007. During this period, 16 species from 12 families of wild birds tested positive for H5N1 virus infection. All samples from juvenile birds were negative for H5N1 virus, whereas 0.6% (95% CI: 0.4, 0.9) of pooled samples from adult birds were positive. Most positive samples originated from peridomestic resident species. Infected wild bird samples were only found in provinces where poultry outbreaks had occurred. Detection of H5N1 virus infection in wild birds was reported up to 3 yr after eradication of the poultry outbreaks in those provinces. As observed with outbreaks in poultry, the frequencies of H5N1 outbreaks in wild birds were significantly higher in winter. Further understanding of the mechanisms of persistence and ongoing HPAI H5N1 transmission between wild birds and domestic poultry is needed.
Neth J Med, 67 (7), pp. 291-294. | Citations: 1 (European Pubmed Central)2009. Turning green with shock.
Southeast Asian J Trop Med Public Health, 40 (4), pp. 781-784. | Citations: 6 (European Pubmed Central) | Show Abstract2009. Lack of correlation of Burkholderia pseudomallei quantities in blood, urine, sputum and pus.
We evaluated the correlation of Burkholderia pseudomallei quantities in blood versus urine, sputum or pus. Correlations between bacterial counts in blood and other samples were not found. It is likely that an initial seeding event to extracellular organs is followed by independent growth of B. pseudomallei, and that bacteria in the urine were not passively filtered from the bloodstream.
A deterministic state-transition model for mastitis transmission was developed to explore population level effects of antibiotic treatment regimens targeting chronic subclinical mastitis caused by major gram-positive pathogens in lactating dairy cows. Behavior and sensitivity of model outputs to changes in key parameters were explored. Outcomes included the size of the state variables describing proportions of infected quarters and basic and effective reproductive numbers. Treatment effects were estimated by calculating proportional reductions in state variables at equilibrium for populations implementing a treatment program relative to populations with no intervention. In general the relationships between parameters were complex and non-linear, although the model outputs were especially sensitive to changes in the value of the transmission rate parameter. Interaction between the parameters resulted in large variations in treatment effect estimates. Effect estimates calculated from model outputs showed a quadratic curve with a clear optimum at low, but not the lowest, transmission rates. These results indicated that overall positive population level effects of lactation therapy would be realized for herds that have successfully implemented practices that reduce the transmission rate of pathogens. A key finding is that in herds with high transmission rates, treatment of chronically infected quarters was predicted to have little impact on the proportion of infected quarters and no positive population level effect in reducing the force of infection and new infection rates. Results of this study suggest that field trials to evaluate efficacy of antimicrobial treatment should include estimates of indirect treatment effects.
NEUROTOXICOLOGY AND TERATOLOGY, 31 (4), pp. 246-246. | Read more2009. Treating severe malaria with pre-referral artesunate saves fives and prevents CNS damage
BACKGROUND TO THE DEBATE: In a 2007 article in PLoS Medicine, Holger J. Schünemann and colleagues described a new process used by the World Health Organization for rapidly developing clinical management guidelines in emergency situations. These situations include outbreaks of emerging infectious diseases. The authors discussed how they developed such a "rapid advice" guideline for the pharmacological management of avian influenza A (H5N1) virus infection. The guideline recommends giving the antiviral drug oseltamivir at a dose of 75 mg twice daily for five days. In this Debate, Nicholas White argues that such dosing is inadequate, Robert Webster and Elena Govorkova say that combination antiviral therapy should be used, and Tim Uyeki reminds us that clinical care of patients with H5N1 entails much more than antiviral treatment. These issues may also apply to therapy of patients hospitalized with severe disease due to novel swine-origin influenza A (H1N1) virus infection.
BACKGROUND: In areas where malaria is endemic, infants aged <3 months appear to be relatively protected from symptomatic and severe Plasmodium falciparum malaria, but less is known about the effect of Plasmodium vivax infection in this age group. METHODS: To define malaria morbidity in the first year of life in an area where both multidrug-resistant P. falciparum and P. vivax are highly prevalent, data were gathered on all infants attending a referral hospital in Papua, Indonesia, using systematic data forms and hospital computerized records. Additional clinical and laboratory data were prospectively collected from inpatients aged <3 months. RESULTS: From April 2004 through April 2008, 4976 infants were admitted to the hospital, of whom 1560 (31%) had malaria, with infection equally attributable to P. falciparum and P. vivax. The case-fatality rate was similar for inpatients with P. falciparum malaria (13 [2.2%] of 599 inpatients died) and P. vivax malaria (6 [1.0%] of 603 died; P= .161), whereas severe malarial anemia was more prevalent among those with P. vivax malaria (193 [32%] of 605 vs. 144 [24%] of 601; P= .025). Of the 187 infants aged <3 months, 102 (56%) had P. vivax malaria, and 55 (30%) had P. falciparum malaria. In these young infants, infection with P. vivax was associated with a greater risk of severe anemia (odds ratio, 2.4; 95% confidence interval, 1.03-5.91; P= .041) and severe thrombocytopenia (odds ratio, 3.3; 95% confidence interval, 1.07-10.6; P= .036) compared with those who have P. falciparum infection. CONCLUSIONS: P. vivax malaria is a major cause of morbidity in early infancy. Preventive strategies, early diagnosis, and prompt treatment should be initiated in the perinatal period.
BACKGROUND: The Plasmodium purine salvage enzyme, hypoxanthine guanine xanthine phosphoribosyl transferase (HGXPRT) can protect mice against Plasmodium yoelii pRBC challenge in a T cell-dependent manner and has, therefore, been proposed as a novel vaccine candidate. It is not known whether natural exposure to Plasmodium falciparum stimulates HGXPRT T cell reactivity in humans. METHODS: PBMC and plasma collected from malaria-exposed Indonesians during infection and 7-28 days after anti-malarial therapy, were assessed for HGXPRT recognition using CFSE proliferation, IFNgamma ELISPOT assay and ELISA. RESULTS: HGXPRT-specific T cell proliferation was found in 44% of patients during acute infection; in 80% of responders both CD4+ and CD8+ T cell subsets proliferated. Antigen-specific T cell proliferation was largely lost within 28 days of parasite clearance. HGXPRT-specific IFN-gamma production was more frequent 28 days after treatment than during acute infection. HGXPRT-specific plasma IgG was undetectable even in individuals exposed to malaria for at least two years. CONCLUSION: The prevalence of acute proliferative and convalescent IFNgamma responses to HGXPRT demonstrates cellular immunogenicity in humans. Further studies to determine minimal HGXPRT epitopes, the specificity of responses for Plasmodia and associations with protection are required. Frequent and robust T cell proliferation, high sequence conservation among Plasmodium species and absent IgG responses distinguish HGXPRT from other malaria antigens.
Trans R Soc Trop Med Hyg, 103 (6), pp. 643-644. | Citations: 5 (European Pubmed Central) | Read more2009. The role of mathematical modelling in malaria elimination and eradication (Comment on: Can malaria be eliminated?).
The antimalarial susceptibility of ring stage (> 80%) Plasmodium vivax from the Republic of Korea, where long incubation-period strains are prevalent, was evaluated using the schizont maturation inhibition technique. During 2005-2007, susceptibility to seven antimalarial drugs was evaluated with 24 fresh isolates. The geometric mean (95% confidence interval) 50% inhibition concentration (IC(50)) were quinine 60 (54-75) ng/mL, chloroquine 39 (22-282) ng/mL, piperaquine 27 (17-58) ng/mL, mefloquine 39 (35-67) ng/mL, pyrimethamine 138 (89-280) ng/mL, artesunate 0.6 (0.5-0.8) ng/mL, and primaquine 122 (98-232) ng/mL. Positive correlations were found between quinine and mefloquine (r = 0.6, P = 0.004), piperaquine and chloroquine (r = 0.6, P = 0.008), and piperaquine and primaquine IC(50) values (r = 0.5, P = 0.01). Compared with P. vivax in Thailand, P. vivax in the Republic of Korea was more sensitive to quinine and mefloquine, but equally sensitive to chloroquine and artesunate.
Am J Trop Med Hyg, 80 (6), pp. 881. | Citations: 4 (Web of Science Lite) | Read more2009. Severe retinal whitening in an adult with cerebral malaria.
Foot-and-mouth disease (FMD) causes sporadic disease outbreaks in the Lao People's Democratic Republic (Lao PDR) and appears to be endemic within a livestock population largely susceptible to infection. As Lao PDR is a major thoroughfare for transboundary animal movement, regular FMD outbreaks occur causing economic hardship for farmers and their families. The dominant serotype causing outbreaks between 1998 and 2006 was type O. Using phylogenetic analysis, type O isolated viruses were divided into two topotypes: South East Asia (SEA) and the Middle East-South Asia (ME-SA). Type A virus was reported only in 2003 and 2006 and type Asia 1 only in 1996 and 1998.
BACKGROUND: An assessment of the correlation between anti-malarial treatment outcome and molecular markers would improve the early detection and monitoring of drug resistance by Plasmodium falciparum. The purpose of this systematic review was to determine the risk of treatment failure associated with specific polymorphisms in the parasite genome or gene copy number. METHODS: Clinical studies of non-severe malaria reporting on target genetic markers (SNPs for pfmdr1, pfcrt, dhfr, dhps, gene copy number for pfmdr1) providing complete information on inclusion criteria, outcome, follow up and genotyping, were included. Three investigators independently extracted data from articles. Results were stratified by gene, codon, drug and duration of follow-up. For each study and aggregate data the random effect odds ratio (OR) with 95%CIs was estimated and presented as Forest plots. An OR with a lower 95th confidence interval > 1 was considered consistent with a failure being associated to a given gene mutation. RESULTS: 92 studies were eligible among the selection from computerized search, with information on pfcrt (25/159 studies), pfmdr1 (29/236 studies), dhfr (18/373 studies), dhps (20/195 studies). The risk of therapeutic failure after chloroquine was increased by the presence of pfcrt K76T (Day 28, OR = 7.2 [95%CI: 4.5-11.5]), pfmdr1 N86Y was associated with both chloroquine (Day 28, OR = 1.8 [95%CI: 1.3-2.4]) and amodiaquine failures (OR = 5.4 [95%CI: 2.6-11.3, p < 0.001]). For sulphadoxine-pyrimethamine the dhfr single (S108N) (Day 28, OR = 3.5 [95%CI: 1.9-6.3]) and triple mutants (S108N, N51I, C59R) (Day 28, OR = 3.1 [95%CI: 2.0-4.9]) and dhfr-dhps quintuple mutants (Day 28, OR = 5.2 [95%CI: 3.2-8.8]) also increased the risk of treatment failure. Increased pfmdr1 copy number was correlated with treatment failure following mefloquine (OR = 8.6 [95%CI: 3.3-22.9]). CONCLUSION: When applying the selection procedure for comparative analysis, few studies fulfilled all inclusion criteria compared to the large number of papers identified, but heterogeneity was limited. Genetic molecular markers were related to an increased risk of therapeutic failure. Guidelines are discussed and a checklist for further studies is proposed.
We performed indirect immunofluorescence assays (IFAs) to compare levels of IgM and IgG antibodies to Orientia tsutsugamushi and Rickettsia typhi in admission-phase serum samples and filter paper blood spots (assayed immediately and stored at 5.4 degrees C and 29 degrees C for 30 days) collected on the same day from 53 adults with suspected scrub typhus and murine typhus admitted to Mahosot Hospital Vientiane, Lao People's Democratic Republic. The sensitivities and specificities of admission-phase filter paper blood spots in comparison to paired sera were between 91% and 95% and 87% and 100%, respectively, for the diagnosis of scrub typhus and murine typhus. The classification of patients as having or not having typhus did not significantly differ after storage of the blood spots for 30 days (P > 0.4) at 5.4 degrees C and 29 degrees C. Because filter paper blood samples do not require sophisticated and expensive storage and transport, they may be an appropriate specimen collection technique for the diagnosis of rickettsial disease in the rural tropics.
Vaccine research is a combinatorial science requiring computational analysis of vaccine components, formulations and optimization. We have developed a framework that combines computational tools for the study of immune function and vaccine development. This framework, named ImmunoGrid combines conceptual models of the immune system, models of antigen processing and presentation, system-level models of the immune system, Grid computing, and database technology to facilitate discovery, formulation and optimization of vaccines. ImmunoGrid modules share common conceptual models and ontologies. The ImmunoGrid portal offers access to educational simulators where previously defined cases can be displayed, and to research simulators that allow the development of new, or tuning of existing, computational models. The portal is accessible at <igrid-ext.cryst.bbk.ac.uk/immunogrid>.
Partial nucleotide sequences (459 bp) of the groEL gene (encoding the 60-kDa heat shock protein, HSP60) from 23 contemporary isolates of Orientia tsutsugamushi isolated from patients with acute scrub typhus in Thailand were compared with 16 reference strain sequences to evaluate the potential of groEL as a conserved and representative target for molecular diagnostics.. Overall nucleotide identity within all available O. tsutsugamushi isolates (n = 39) was 98.8% (range: 95.0-100), reflecting a high degree of conservation; nucleotide identities were 67.5% and 65.6%, respectively, when typhus and spotted fever group rickettsiae were included.. A highly sensitive and quantitative real-time PCR assay was designed and evaluated using 61 samples, including buffy coats from patients in Thailand and Laos. Reliable and accurate quantitation of bacterial loads allows further investigation of other diagnostic methods and may lead to an improved understanding of the pathophysiology of acute scrub typhus, an important but under-recognized disease.
Long considered a benign infection, Plasmodium vivax is now recognized as a cause of severe and fatal malaria, despite its low parasite biomass, the increased deformability of vivax-infected red blood cells and an apparent paucity of parasite sequestration. Severe anemia is associated with recurrent bouts of hemolysis of predominantly uninfected erythrocytes with increased fragility, and lung injury is associated with inflammatory increases in alveolar-capillary membrane permeability. Although rare, vivax-associated coma challenges our understanding of pathobiology caused by Plasmodium spp. Host and parasite factors contribute to the risk of severe disease, and comorbidities might contribute to vivax mortality. In this review, we discuss potential mechanisms underlying the syndromes of uncomplicated and severe vivax malaria, identifying key areas for future research.
PLOS MEDICINE, 6 (5), pp. e1000085-e1000085. | Citations: 52 (Web of Science Lite) | Read more2009. Hedging against Antiviral Resistance during the Next Influenza Pandemic Using Small Stockpiles of an Alternative Chemotherapy
Ann Intern Med, 150 (8), pp. 567-568. | Citations: 25 (Web of Science Lite) | Read more2009. Gallbladder carriage of Salmonella paratyphi A may be an important factor in the increasing incidence of this infection in South Asia.
A liquid chromatographic tandem mass spectroscopy method for the quantification of artemisinin in human heparinised plasma has been developed and validated. The method uses Oasis HLB mu-elution solid phase extraction 96-well plates to facilitate a high throughput of 192 samples a day. Artesunate (internal standard) in a plasma-water solution was added to plasma (50 microL) before solid phase extraction. Artemisinin and its internal standard artesunate were analysed by liquid chromatography and MS/MS detection on a Hypersil Gold C18 (100 mm x 2.1 mm, 5 microm) column using a mobile phase containing acetonitrile-ammonium acetate 10mM pH 3.5 (50:50, v/v) at a flow rate of 0.5 mL/min. The method has been validated according to published FDA guidelines and showed excellent performance. The within-day, between-day and total precisions expressed as R.S.D., were lower than 8% at all tested quality control levels including the upper and lower limit of quantification. The limit of detection was 0.257 ng/mL for artemisinin and the calibration range was 1.03-762 ng/mL using 50 microL plasma. The method was free from matrix effects as demonstrated both graphically and quantitatively.
BACKGROUND: Infection with the Gram-negative bacterium Burkholderia pseudomallei is an important cause of community-acquired lethal sepsis in endemic regions in southeast Asia and northern Australia and is increasingly reported in other tropical areas. In animal models, production of interferon-gamma (IFN-gamma) is critical for resistance, but in humans the characteristics of IFN-gamma production and the bacterial antigens that are recognized by the cell-mediated immune response have not been defined. METHODS: Peripheral blood from 133 healthy individuals who lived in the endemic area and had no history of melioidosis, 60 patients who had recovered from melioidosis, and 31 other patient control subjects were stimulated by whole bacteria or purified bacterial proteins in vitro, and IFN-gamma responses were analyzed by ELISPOT and flow cytometry. FINDINGS: B. pseudomallei was a potent activator of human peripheral blood NK cells for innate production of IFN-gamma. In addition, healthy individuals with serological evidence of exposure to B. pseudomallei and patients recovered from active melioidosis developed CD4(+) (and CD8(+)) T cells that recognized whole bacteria and purified proteins LolC, OppA, and PotF, members of the B. pseudomallei ABC transporter family. This response was primarily mediated by terminally differentiated T cells of the effector-memory (T(EMRA)) phenotype and correlated with the titer of anti-B. pseudomallei antibodies in the serum. CONCLUSIONS: Individuals living in a melioidosis-endemic region show clear evidence of T cell priming for the ability to make IFN-gamma that correlates with their serological status. The ability to detect T cell responses to defined B. pseudomallei proteins in large numbers of individuals now provides the opportunity to screen candidate antigens for inclusion in protein or polysaccharide-conjugate subunit vaccines against this important but neglected disease.
Mefloquine is widely used in combination with artemisinin derivatives for the treatment of falciparum malaria. Mefloquine resistance in Plasmodium falciparum has been related to increased copy numbers of multidrug-resistant gene 1 (pfmdr1). We studied the ex vivo dynamics of pfmdr1 gene amplification in culture-adapted P. falciparum in relation to mefloquine resistance and parasite fitness. A Thai P. falciparum isolate (isolate TM036) was assessed by the use of multiple genetic markers as a single genotype. Resistance was selected by exposure to stepwise increasing concentrations of mefloquine up to 30 ng/ml in continuous culture. The pfmdr1 gene copy numbers increased as susceptibility to mefloquine declined (P = 0.03). No codon mutations at positions 86, 184, 1034, 1042, and 1246 in the pfmdr1 gene were detected. Two subclones of selected parasites (average copy numbers, 2.3 and 3.1, respectively) showed a fitness disadvantage when they were grown together with the original parasites containing a single pfmdr1 gene copy in the absence of mefloquine; the multiplication rates were 6.3% and 8.7% lower, respectively (P < 0.01). Modeling of the dynamics of the pfmdr1 copy numbers over time in relation to the relative fitness of the parasites suggested that net pfmdr1 gene amplification from one to two copies occurs once in every 10(8) parasites and that amplification from two to three copies occurs once in every 10(3) parasites. pfmdr1 gene amplification in P. falciparum is a frequent event and confers mefloquine resistance. Parasites with multiple copies of the pfmdr1 gene have decreased survival fitness in the absence of drug pressure.
Severe Plasmodium falciparum malaria is a major cause of global mortality, yet the immunological factors underlying progression to severe disease remain unclear. CD4(+)CD25(+) regulatory T cells (Treg cells) are associated with impaired T cell control of Plasmodium spp infection. We investigated the relationship between Treg cells, parasite biomass, and P. falciparum malaria disease severity in adults living in a malaria-endemic region of Indonesia. CD4(+)CD25(+)Foxp3(+)CD127(lo) Treg cells were significantly elevated in patients with uncomplicated (UM; n = 17) and severe malaria (SM; n = 16) relative to exposed asymptomatic controls (AC; n = 10). In patients with SM, Treg cell frequency correlated positively with parasitemia (r = 0.79, p = 0.0003) and total parasite biomass (r = 0.87, p<0.001), both major determinants for the development of severe and fatal malaria, and Treg cells were significantly increased in hyperparasitemia. There was a further significant correlation between Treg cell frequency and plasma concentrations of soluble tumor necrosis factor receptor II (TNFRII) in SM. A subset of TNFRII(+) Treg cells with high expression of Foxp3 was increased in severe relative to uncomplicated malaria. In vitro, P. falciparum-infected red blood cells dose dependently induced TNFRII(+)Foxp3(hi) Treg cells in PBMC from malaria-unexposed donors which showed greater suppressive activity than TNFRII(-) Treg cells. The selective enrichment of the Treg cell compartment for a maximally suppressive TNFRII(+)Foxp3(hi) Treg subset in severe malaria provides a potential link between immune suppression, increased parasite biomass, and malaria disease severity. The findings caution against the induction of TNFRII(+)Foxp3(hi) Treg cells when developing effective malaria vaccines.
BACKGROUND: Artemether-lumefantrine is the most widely recommended artemisinin-based combination treatment for falciparum malaria. Quantification of artemether and its metabolite dihydroartemisinin in biological matrices has traditionally been difficult, with sensitivity being an issue. RESULTS: A high-throughput bioanalytical method for the analysis of artemether and its metabolite dihydroartemisinin in human plasma using solid-phase extraction in the 96-well plate format and liquid chromatography coupled to positive ion mode tandem mass spectroscopy has been developed and validated according to US FDA guidelines. The method uses 50 µl plasma and covers the calibration range 1.43-500 ng/ml with a limit of detection at 0.36 ng/ml. CONCLUSIONS: The developed liquid chromatography-tandem mass spectrometry assay is more sensitive than all previous methods despite using a lower plasma volume (50 µl) and is highly suitable for clinical studies where plasma volumes are limited, such as pediatric trials.
The diagnosis of severe Streptococcus pneumoniae infection relies heavily on insensitive culture techniques. To improve the usefulness of PCR assays, we developed a dual-PCR protocol (targeted at pneumolysin and autolysin) for EDTA blood samples. This was compared to the Binax NOW S. pneumoniae urine antigen test in patients with bacteremic pneumococcal infections. Patients with nonbacteremic community-acquired pneumonia also were tested by these methods to determine what proportion could be confirmed as pneumococcal infections. A direct comparison was made in a group of patients who each had both tests performed. The Binax NOW S. pneumoniae urine antigen test was positive in 51 of 58 bacteremic pneumococcal cases (sensitivity, 88%; 95% confidence interval [CI], 77 to 95%), whereas the dual PCR was positive in 31 cases (sensitivity, 53.5%; 95% CI, 40 to 67%; P < 0.0001), and all of these had detectable urinary antigens. Both tests gave positive results in 2 of 51 control patients (referred to as other-organism septicemia), giving a specificity of 96% (95% CI, 86.5 to 99.5%). In 77 patients with nonbacteremic community-acquired pneumonia, urinary antigen was detected significantly more often (in 21 patients [27%]) than a positive result by the dual-PCR protocol (6 [8%]) (P = 0.002). The development of a dual-PCR protocol enhanced the sensitivity compared to that of the individual assays, but it is still significantly less sensitive than the Binax NOW urine antigen test, as well as being more time-consuming and expensive. Urinary antigen detection is the nonculture diagnostic method of choice for patients with possible severe pneumococcal infection.
OBJECTIVES: To assess impact of serial lumbar punctures on association between cerebrospinal fluid (CSF) opening pressure and prognosis in HIV-associated cryptococcal meningitis; to explore time course and relationship of opening pressure with neurological findings, CSF fungal burden, immune response, and CD4 cell count. DESIGN: Evaluation of 163 HIV-positive ART-naive patients enrolled in three trials of amphotericin B-based therapy for cryptococcal meningitis in Thailand and South Africa. METHODS: Study protocols required four lumbar punctures with measurements of opening pressure over the first 2 weeks of treatment and additional lumbar punctures if opening pressure raised. Fungal burden and clearance, CSF immune parameters, CD4 cell count, neurological symptoms and signs, and outcome at 2 and 10 weeks were compared between groups categorized by opening pressure at cryptococcal meningitis diagnosis. RESULTS: Patients with higher baseline fungal burden had higher baseline opening pressure. High fungal burden appeared necessary but not sufficient for development of high pressure. Baseline opening pressure was not associated with CD4 cell count, CSF pro-inflammatory cytokines, or altered mental status. Day 14 opening pressure was associated with day 14 fungal burden. Overall mortality was 12% (20/162) at 2 weeks and 26% (42/160) at 10 weeks, with no significant differences between opening pressure groups. CONCLUSION: Studies are needed to define factors, in addition to fungal burden, associated with raised opening pressure. Aggressive management of raised opening pressure through repeated CSF drainage appeared to prevent any adverse impact of raised opening pressure on outcome in patients with cryptococcal meningitis. The results support increasing access to manometers in resource-poor settings and routine management of opening pressure in patients with cryptococcal meningitis.
PLoS Med, 6 (3), pp. e52. | Citations: 74 (Web of Science Lite) | Read more2009. Guidelines for field surveys of the quality of medicines: a proposal.
Indications:For the treatment of malaria in children.Patients:Unspecified number of children from Papua New Guinea.TypeofStudy:Letters to the editor.DosageDuration:Dosage and duration not stated.ComparativeDrug:Dihydroartemisinin-piperaquine, dosage and duration not stated.Results:-AdverseEffects:No adverse events were mentioned.FreeText:Comment on Riamet (N Engl J Med 2008;359:2545-57). The study reported that Riamet had a more favorable efficacy than dihydroartemisinin-piperaquine in children with malaria from Papua New Guinea. The comment states that Riamet was not superior to dihydroartemisinin-piperaquine since fat was given with the treatment only in the Riamet group thereby increasing its bioavailability, and no significant difference was noted between groups in the primary end point. Furthermore, the per-protocol analysis with a high dropout rate from a small sample resulted in overestimation of the risk of treatment failure and wide 95% confidence intervals (6.4% to 20.0%). Finally, the comment states that both Riamet and dihydroartemisinin-piperaquine should be given with fat (milk, biscuit, or other food) to increase bioavailability.
EUROPEAN JOURNAL OF CLINICAL NUTRITION, 63 (3), pp. 450-450. | Read more2009. Postpartum traditions and nutrition practices among urban Lao women and their infants in Vientiane, Lao PDR (vol 63, pg 323, 2009)
OBJECTIVE: To explore the cost-effectiveness of artesunate against quinine based principally on the findings of a large multi-centre trial carried out in Southeast Asia. METHODS: Trial data were used to compare mortality of patients with severe malaria, treated with either artesunate or quinine. This was combined with retrospectively collected cost data to estimate the incremental cost per death averted with the use of artesunate instead of quinine. RESULTS: The incremental cost per death averted using artesunate was approximately 140 USD. Artesunate maintained this high level of cost-effectiveness also when allowing for the uncertainty surrounding the cost and effectiveness assessments. CONCLUSION: This analysis confirms the vast superiority of artesunate for treatment of severe malaria from an economic as well as a clinical perspective.
The effects of loading doses and probenecid coadministration on oseltamivir pharmacokinetics at four increasing dose levels in groups of eight healthy adult Thai volunteers (125 individual series) were evaluated. Doses of up to 675 mg were well-tolerated. The pharmacokinetics were dose linear. Oseltamivir phosphate (OS) was rapidly and completely absorbed and converted (median conversion level, 93%) to the active carboxylate metabolite. Median elimination half-lives (and 95% confidence intervals [CI]) were 1.0 h (0.9 to 1.1 h) for OS and 5.1 h (4.7 to 5.7 h) for oseltamivir carboxylate (OC). One subject repeatedly showed markedly reduced OS-to-OC conversion, indicating constitutionally impaired carboxylesterase activity. The coadministration of probenecid resulted in a mean contraction in the apparent volume of distribution of OC of 40% (95% CI, 37 to 44%) and a reduction in the renal elimination of OC of 61% (95% CI, 58 to 62%), thereby increasing the median area under the concentration-time curve (AUC) for OC by 154% (range, 71 to 278%). The AUC increase for OC in saliva was approximately three times less than the AUC increase for OC in plasma. A loading dose 1.25 times the maintenance dose should be given for severe influenza pneumonia. Probenecid coadministration may allow considerable dose saving for oseltamivir, but more information on OC penetration into respiratory secretions is needed to devise appropriate dose regimens.
Amodiaquine retains efficacy against infection by chloroquine-resistant Plasmodium falciparum; however, little information is available on its efficacy against infection by chloroquine-resistant Plasmodium vivax. Patients presenting to a rural clinic with a pure P. vivax infection that recurred after recent antimalarial treatment were retreated, this time with amodiaquine monotherapy, and the risk of further recurrence within 4 weeks was assessed. Of the 87 patients with pure P. vivax infection, 15 patients did not complete a full course of treatment, 4 of whom were intolerant to treatment. In the 72 patients completing treatment, 91% (63 of 69) had cleared their parasitemia within 48 h with no early treatment failure. Follow-up to day 28 or recurrent parasitemia was achieved for 56 patients (78%). The cumulative incidence of treatment failure by day 28 was 22.8% (95% confidence interval, 7.3 to 38%). The in vitro sensitivity profile was determined for a separate set of isolates from outpatients with pure P. vivax infection. The median 50% inhibitory concentration of amodiaquine was 11.3 nM (range, 0.37 to 95.8) and was correlated significantly with that of chloroquine (Spearman rank correlation coefficient, 0.602; P < 0.001). Although amodiaquine results in a rapid clinical response, the risk of recurrence by day 28 is unacceptably high, reducing its suitability as an alternative treatment of infection by chloroquine-resistant P. vivax in this region.
Hygiene + Medizin, 34 (3), pp. 108-109.2009. New study on selective intestinal decontamination shows only marginal benefit
BACKGROUND: We investigated differences in breast cancer mortality between younger (younger than 40 years of age) and older (40 years of age and older) women by stage at diagnosis to identify patient and tumor characteristics accounting for disparities. STUDY DESIGN: We conducted a retrospective study of women diagnosed with breast cancer in the 1988 to 2003 Surveillance, Epidemiology, and End Results Program data. Multivariate Cox regression models calculated adjusted hazard ratios (aHR) and 95% confidence intervals to compare overall and stage-specific breast cancer mortality in women younger than 40 years old and women 40 years and older, controlling for potential confounding variables identified in univariate tests. RESULTS: Of 243,012 breast cancer patients, 6.4% were younger than 40 years old, and 93.6% were 40 years of age or older. Compared with older women, younger women were more likely to be African American, single, diagnosed at later stages, and treated by mastectomy. Younger women had tumors that were more likely to be higher grade, larger size, estrogen receptor/progesterone receptor-negative, and lymph-node positive (p < 0.001). Younger women were more likely to die from breast cancer compared with older women (crude HR = 1.39; CI, 1.34 to 1.45). Controlling for confounders, younger women were more likely to die compared with older women if diagnosed with stage I (aHR = 1.44; CI, 1.27 to 1.64) or stage II (aHR = 1.09; CI, 1.03 to 1.15) disease and less likely to die if diagnosed with stage IV disease (aHR = 0.85; CI, 0.76 to 0.95). CONCLUSIONS: Higher breast cancer mortality in younger women was attributed to poorer outcomes with early-stage disease. Additional studies should focus on specific tumor biology contributing to the increased mortality of younger women with early-stage breast cancer.
BACKGROUND: Most malaria deaths occur in rural areas. Rapid progression from illness to death can be interrupted by prompt, effective medication. Antimalarial treatment cannot rescue terminally ill patients but could be effective if given earlier. If patients who cannot be treated orally are several hours from facilities for injections, rectal artesunate can be given before referral and acts rapidly on parasites. We investigated whether this intervention reduced mortality and permanent disability. METHODS: In Bangladesh, Ghana, and Tanzania, patients with suspected severe malaria who could not be treated orally were allocated randomly to a single artesunate (n=8954) or placebo (n=8872) suppository by taking the next numbered box, then referred to clinics at which injections could be given. Those with antimalarial injections or negative blood smears before randomisation were excluded, leaving 12 068 patients (6072 artesunate, 5996 placebo) for analysis. Primary endpoints were mortality, assessed 7-30 days later, and permanent disability, reassessed periodically. All investigators were masked to group assignment. Analysis was by intention to treat. This study is registered in all three countries, numbers ISRCTN83979018, 46343627, and 76987662. RESULTS: Mortality was 154 of 6072 artesunate versus 177 of 5996 placebo (2.5%vs 3.0%, p=0.1). Two versus 13 (0.03%vs 0.22%, p=0.0020) were permanently disabled; total dead or disabled: 156 versus 190 (2.6%vs 3.2%, p=0.0484). There was no reduction in early mortality (56 vs 51 deaths within 6 h; median 2 h). In patients reaching clinic within 6 h (median 3 h), pre-referral artesunate had no significant effect on death after 6 h or permanent disability (71/4450 [1.6%] vs 82/4426 [1.9%], risk ratio 0.86 [95% CI 0.63-1.18], p=0.35). In patients still not in clinic after more than 6 h, however, half were still not there after more than 15 h, and pre-referral rectal artesunate significantly reduced death or permanent disability (29/1566 [1.9%] vs 57/1519 [3.8%], risk ratio 0.49 [95% CI 0.32-0.77], p=0.0013). INTERPRETATION: If patients with severe malaria cannot be treated orally and access to injections will take several hours, a single inexpensive artesunate suppository at the time of referral substantially reduces the risk of death or permanent disability. FUNDING: UNICEF/UNDP/World Bank Special Programme for Research and Training in Tropical Diseases (WHO/TDR); WHO Global Malaria Programme (WHO/GMP); Sall Family Foundation; the European Union (QLRT-2000-01430); the UK Medical Research Council; USAID; Irish Aid; the Karolinska Institute; and the University of Oxford Clinical Trial Service Unit (CTSU).
Understanding the evolution of drug resistance in malaria is a central area of study at the intersection of evolution and medicine. Antimalarial drug resistance is a major threat to malaria control and directly related to trends in malaria attributable mortality. Artemisinin combination therapies (ACT) are now recommended worldwide as first line treatment for uncomplicated malaria, and losing them to resistance would be a disaster for malaria control. Understanding the emergence and spread of antimalarial drug resistance in the context of different scenarios of antimalarial drug use is essential for the development of strategies protecting ACTs. In this study, we review the basic mechanisms of resistance emergence and describe several simple equations that can be used to estimate the probabilities of de novo resistance mutations at three stages of the parasite life cycle: sporozoite, hepatic merozoite and asexual blood stages; we discuss the factors that affect parasite survival in a single host in the context of different levels of antimalarial drug use, immunity and parasitaemia. We show that in the absence of drug effects, and despite very different parasite numbers, the probability of resistance emerging at each stage is very low and similar in all stages (for example per-infection probability of 10(-10)-10(-9) if the per-parasite chance of mutation is 10(-10) per asexual division). However, under the selective pressure provided by antimalarial treatment and particularly in the presence of hyperparasitaemia, the probability of resistance emerging in the blood stage of the parasite can be approximately five orders of magnitude higher than in the absence of drugs. Detailed models built upon these basic methods should allow us to assess the relative probabilities of resistance emergence in the different phases of the parasite life cycle.
OBJECTIVES: Sepsis is associated with immunosuppression (characterized by a reduced capacity of circulating monocytes to release proinflammatory cytokines), which has been implicated in late mortality. Melioidosis, caused by the Gram-negative bacterium Burkholderia pseudomallei, is an important cause of community-acquired sepsis in Southeast Asia with a mortality of up to 40%. Previous in vitro and murine studies have suggested a key role for the so-called negative regulators of the toll-like receptor (TLR) signaling pathway in immunosuppression. In this study, we investigated the expression of these negative TLR regulators in patients with septic melioidosis in association with the responsiveness of peripheral blood leukocytes of these patients to lipopolysaccharide and B. pseudomallei. DESIGN: Ex vivo study. SETTING: Academic research laboratory. PATIENTS: Thirty-two healthy controls and 34 patients with sepsis caused by B. pseudomallei. INTERVENTIONS: None. MEASUREMENTS: 1) Plasma cytokine levels; 2) ex vivo cytokine production capacity of whole blood; and 3) purified mononuclear cell-derived messenger RNA (mRNA) levels of key inhibitory molecules of the TLR-signaling cascade were investigated. MAIN RESULTS: In accordance with an immunosuppressed state, whole blood of patients demonstrated a strongly decreased capacity to release the proinflammatory cytokines tumor necrosis factor-[alpha], interleukin-1[beta], and the chemokine interleukin-8 after ex vivo stimulation with lipopolysaccharide or B. pseudomallei. Analysis of myeloid-differentiation-88-short, interleukin-1R-associated-kinase (IRAK)-M, IRAK-1, suppressor-of-cytokine signaling-3, Src-homology-2-domain-containing inositol-5-phosphatase-1, single-immunoglobulin-interleukin-1R-related-molecule, and A20 mRNA expression in purified mononuclear cells showed decreased IRAK-1 and elevated IRAK-M expression in patients with septic melioidosis. Immunosuppression was correlated with mortality; furthermore, patients who eventually died had higher IRAK-M mRNA levels on admission than the patients who survived. CONCLUSIONS: Immunosuppression in sepsis caused by B. pseudomallei is associated with an upregulation of IRAK-M and an indicator of poor outcome.
OBJECTIVE: Markers of oxidative stress are reported to be increased in severe malaria. It has been suggested that the antioxidant N-acetylcysteine (NAC) may be beneficial in treatment. We studied the efficacy and safety of parenteral NAC as an adjunct to artesunate treatment of severe falciparum malaria. DESIGN: A randomized, double-blind, placebo-controlled trial on the use of high-dose intravenous NAC as adjunctive treatment to artesunate. SETTING: A provincial hospital in Western Thailand and a tertiary referral hospital in Chittagong, Bangladesh. PATIENTS: One hundred eight adult patients with severe falciparum malaria. INTERVENTIONS: Patients were randomized to receive NAC or placebo as an adjunctive treatment to intravenous artesunate. MEASUREMENTS AND MAIN RESULTS: A total of 56 patients were treated with NAC and 52 received placebo. NAC had no significant effect on mortality, lactate clearance times (p = 0.74), or coma recovery times (p = 0.46). Parasite clearance time was increased from 30 hours (range, 6-144 hours) to 36 hours (range, 6-120 hours) (p = 0.03), but this could be explained by differences in admission parasitemia. Urinary F2-isoprostane metabolites, measured as a marker of oxidative stress, were increased in severe malaria compared with patients with uncomplicated malaria and healthy volunteers. Admission red cell rigidity correlated with mortality, but did not improve with NAC. CONCLUSION: Systemic oxidative stress is increased in severe malaria. Treatment with NAC had no effect on outcome in patients with severe falciparum malaria in this setting.
Orientia tsutsugamushi, the cause of scrub typhus, is a major pathogen in the Asia-Pacific region. The severity of infection ranges from mild features to multiorgan failure and death. The aim of this prospective study was to define the O. tsutsugamushi loads in the blood samples of patients with scrub typhus on the day of hospital admission and to determine whether this was associated with disease severity. Quantitation was performed using a real-time PCR assay targeting the 16S rRNA gene of O. tsutsugamushi. A total of 155 patients with a confirmed diagnosis of scrub typhus had a median (interquartile range [IQR], range) O. tsutsugamushi DNA load in blood of 13 (0 to 334, 0 to 310,253) copies/ml. This included 74 patients who had undetectable bacterial loads. An analysis of bacterial load versus clinical features for all 155 patents demonstrated that duration of illness (P < 0.001), presence of eschar (P = 0.004), and concentrations of aspartate aminotransferase, alanine aminotransferase, and alkaline phosphatase (P < 0.001 for all three) were positively correlated with bacterial load. Patients who died had a significantly higher bacterial load than those who survived (mean [standard deviation] values: 17,154 [12.7] versus 281 [5.2] copies/ml; P < 0.001). This study has demonstrated a relationship between bacterial load and disease severity in adults with scrub typhus.
Multiple displacement amplification (MDA) using Phi29 has proved to be an efficient, high-fidelity method for whole genome amplification in many organisms. This project was designed to evaluate this approach for use with the malaria parasite Plasmodium falciparum. In particular, we were concerned that the AT richness and presence of contaminating human DNA could limit efficiency of MDA in this system. We amplified 60 DNA samples using phi29 and scored 14 microsatellites, 9 single-nucleotide polymorphisms (SNPs), and gene copy number at GTP-cyclohydrolase I both before and after MDA. We observed 100% concordance in 829 microsatellite genotypes and in 499 SNP genotypes. Furthermore, copy number estimates for the GTP-cyclohydrolase I gene were correlated (r(2) = 0.67) in pre- and postamplification samples. These data confirm that MDA permits scoring of a range of different types of polymorphisms in P. falciparum malaria and can be used to extend the life of valuable DNA stocks.
By contrast with high-income countries, Staphylococcus aureus disease ranks low on the public-health agenda in low-income countries. We undertook a literature review of S aureus disease in resource-limited countries in south and east Asia, and found that its neglected status as a developing world pathogen does not equate with low rates of disease. The incidence of the disease seems to be highest in neonates, its range of clinical manifestations is as broad as that seen in other settings, and the mortality rate associated with serious S aureus infection, such as bacteraemia, is as high as 50%. The prevalence of meticillin-resistant S aureus (MRSA) infection across much of resource-limited Asia is largely unknown. Antibiotic drugs are readily and widely available from pharmacists in most parts of Asia, where ease of purchase and frequent self-medication are likely to be major drivers in the emergence of drug resistance. In our global culture, the epidemiology of important drug-resistant pathogens in resource-limited countries is inextricably linked with the health of both developing and developed communities. An initiative is needed to raise the profile of S aureus disease in developing countries, and to define a programme of research to find practical solutions to the health-care challenges posed by this important global pathogen.
Maturation of Plasmodium falciparum decreases the deformability of infected red blood cells (RBCs), increasing their clearance as they attempt to pass through endothelial slits of the splenic sinus. Previous studies of Plasmodium vivax-infected RBCs led to opposite conclusions with respect to cellular deformability. To resolve this controversy, P. vivax-infected RBCs were passed through a 2-microm microfluidic channel. In contrast to P. falciparum-infected RBCs, mature P. vivax-infected RBCs readily became deformed through 2-microm constrictions. After this extreme deformation, 67% of P. vivax-infected RBCs recovered a normal appearance; however, 15% of uninfected RBCs were destroyed. Results suggest mechanisms for both avoidance of splenic clearance and anemia in vivax malaria.
Bipolar elongation of filaments of the bacterial actin homolog ParM drives movement of newly replicated plasmid DNA to opposite poles of a bacterial cell. We used a combination of vitreous sectioning and electron cryotomography to study this DNA partitioning system directly in native, frozen cells. The diffraction patterns from overexpressed ParM bundles in electron cryotomographic reconstructions were used to unambiguously identify ParM filaments in Escherichia coli cells. Using a low-copy number plasmid encoding components required for partitioning, we observed small bundles of three to five intracellular ParM filaments that were situated close to the edge of the nucleoid. We propose that this may indicate the capture of plasmid DNA within the periphery of this loosely defined, chromosome-containing region.
The hepatitis C virus (HCV), which currently infects an estimated 3% of people worldwide, has been present in some human populations for several centuries, notably HCV genotypes 1 and 2 in West Africa and genotype 6 in Southeast Asia. Here we use newly developed methods of sequence analysis to conduct the first comprehensive investigation of the epidemic and evolutionary history of HCV in Asia. Our analysis includes new HCV core (n = 16) and NS5B (n = 14) gene sequences, obtained from serum samples of jaundiced patients from Laos. These exceptionally diverse isolates were analyzed in conjunction with all available reference strains using phylogenetic and Bayesian coalescent methods. We performed statistical tests of phylogeographic structure and applied a recently developed "relaxed molecular clock" approach to HCV for the first time, which indicated an unexpectedly high degree of rate variation. Our results reveal a >1,000-year-long development of genotype 6 in Asia, characterized by substantial phylogeographic structure and two distinct phases of epidemic history, before and during the 20th century. We conclude that HCV lineages representing preexisting and spatially restricted strains were involved in multiple, independent local epidemics during the 20th century. Our analysis explains the generation and maintenance of HCV diversity in Asia and could provide a template for further investigations of HCV spread in other regions.
BACKGROUND: Facilitation of endogenous neuroprotective pathways, such as the erythropoietin (Epo) pathway, has been proposed as adjuvant treatment strategies in cerebral malaria. Whether different endogenous protein expression levels of Epo or differences in the abundance of its receptor components could account for the extent of structural neuropathological changes or neurological complications in adults with severe malaria was investigated. METHODS: High sensitivity immunohistochemistry was used to assess the frequency, distribution and concordance of Epo and components of its homodimeric and heteromeric receptors, Epo receptor and CD131, within the brainstem of adults who died of severe malaria. The following relationships with Epo and its receptor components were also defined: (i) sequestration and indicators of hypoxia; (ii) vascular damage in the form of plasma protein leakage and haemorrhage; (iii) clinical complications and neuropathological features of severe malaria disease. Brainstems of patients dying in the UK from unrelated non-infectious causes were examined for comparison. RESULTS: The incidence of endogenous Epo in parenchymal brain cells did not greatly differ between severe malaria and non-neurological UK controls at the time of death. However, EpoR and CD131 labelling of neurons was greater in severe malaria compared with non-neurological controls (P = .009). EpoR labelling of vessels was positively correlated with admission peripheral parasite count (P = .01) and cerebral sequestration (P < .0001). There was a strong negative correlation between arterial oxygen saturation and EpoR labelling of glia (P = .001). There were no significant correlations with indicators of vascular damage, neuronal chromatolysis, axonal swelling or vital organ failure. CONCLUSION: Cells within the brainstem of severe malaria patients showed protein expression of Epo and its receptor components. However, the incidence of endogeneous expression did not reflect protection from vascular or neuronal injury, and/or clinical manifestations, such as coma. These findings do not provide support for Epo as an adjuvant neuroprotective agent in adults with severe malaria.
BACKGROUND: Artemisinin combination treatments (ACT) are recommended as first line treatment for falciparum malaria throughout the malaria affected world. We reviewed the efficacy of a 3-day regimen of mefloquine and artesunate regimen (MAS(3)), over a 13 year period of continuous deployment as first-line treatment in camps for displaced persons and in clinics for migrant population along the Thai-Myanmar border. METHODS AND FINDINGS: 3,264 patients were enrolled in prospective treatment trials between 1995 and 2007 and treated with MAS(3). The proportion of patients with parasitaemia persisting on day-2 increased significantly from 4.5% before 2001 to 21.9% since 2002 (p<0.001). Delayed parasite clearance was associated with increased risk of developing gametocytaemia (AOR = 2.29; 95% CI, 2.00-2.69, p = 0.002). Gametocytaemia on admission and carriage also increased over the years (p = 0.001, test for trend, for both). MAS(3) efficacy has declined slightly but significantly (Hazards ratio 1.13; 95% CI, 1.07-1.19, p<0.001), although efficacy in 2007 remained well within acceptable limits: 96.5% (95% CI, 91.0-98.7). The in vitro susceptibility of P. falciparum to artesunate increased significantly until 2002, but thereafter declined to levels close to those of 13 years ago (geometric mean in 2007: 4.2 nM/l; 95% CI, 3.2-5.5). The proportion of infections caused by parasites with increased pfmdr1 copy number rose from 30% (12/40) in 1996 to 53% (24/45) in 2006 (p = 0.012, test for trend). CONCLUSION: Artesunate-mefloquine remains a highly efficacious antimalarial treatment in this area despite 13 years of widespread intense deployment, but there is evidence of a modest increase in resistance. Of particular concern is the slowing of parasitological response to artesunate and the associated increase in gametocyte carriage.
BACKGROUND: Analytical approaches for the interpretation of anti-malarial clinical trials vary considerably. The aim of this study was to quantify the magnitude of the differences between efficacy estimates derived from these approaches and identify the factors underlying these differences. METHODS: Data from studies conducted in Africa and Thailand were compiled and the risk estimates of treatment failure, adjusted and unadjusted by genotyping, were derived by three methods (intention to treat (ITT), modified intention to treat (mITT) and per protocol (PP)) and then compared. RESULTS: 29 clinical trials (15 from Africa and 14 from Thailand) with a total of 65 treatment arms (38 from Africa and 27 from Thailand) were included in the analysis. Of the 15,409 patients enrolled, 2,637 (17.1%) had incomplete follow up for the unadjusted analysis and 4,489 (33.4%) for the adjusted analysis. Estimates of treatment failure were consistently higher when derived from the ITT or PP analyses compared to the mITT approach. In the unadjusted analyses the median difference between the ITT and mITT estimates was greater in Thai studies (11.4% [range 2.1-31.8]) compared to African Studies (1.8% [range 0-11.7]). In the adjusted analyses the median difference between PP and mITT estimates was 1.7%, but ranged from 0 to 30.9%. The discrepancy between estimates was correlated significantly with the proportion of patients with incomplete follow-up; p < 0.0001. The proportion of studies with a major difference (> 5%) between adjusted PP and mITT was 28% (16/57), with the risk difference greater in African (37% 14/38) compared to Thai studies (11% 2/19). In the African studies, a major difference in the adjusted estimates was significantly more likely in studies in high transmission sites (62% 8/13) compared to studies in moderate transmission sites (24% 6/25); p = 0.035. CONCLUSION: Estimates of anti-malarial clinical efficacy vary significantly depending on the analytical methodology from which they are derived. In order to monitor temporal and spatial trends in anti-malarial efficacy, standardized analytical tools need to be applied in a transparent and systematic manner.
BACKGROUND: Preventing the emergence of anti-malarial drug resistance is critical for the success of current malaria elimination efforts. Prevention strategies have focused predominantly on qualitative factors, such as choice of drugs, use of combinations and deployment of multiple first-line treatments. The importance of anti-malarial treatment dosing has been underappreciated. Treatment recommendations are often for the lowest doses that produce "satisfactory" results. METHODS: The probability of de-novo resistant malaria parasites surviving and transmitting depends on the relationship between their degree of resistance and the blood concentration profiles of the anti-malarial drug to which they are exposed. The conditions required for the in-vivo selection of de-novo emergent resistant malaria parasites were examined and relative probabilities assessed. RESULTS: Recrudescence is essential for the transmission of de-novo resistance. For rapidly eliminated anti-malarials high-grade resistance can arise from a single drug exposure, but low-grade resistance can arise only from repeated inadequate treatments. Resistance to artemisinins is, therefore, unlikely to emerge with single drug exposures. Hyperparasitaemic patients are an important source of de-novo anti-malarial drug resistance. Their parasite populations are larger, their control of the infection insufficient, and their rates of recrudescence following anti-malarial treatment are high. As use of substandard drugs, poor adherence, unusual pharmacokinetics, and inadequate immune responses are host characteristics, likely to pertain to each recurrence of infection, a small subgroup of patients provides the particular circumstances conducive to de-novo resistance selection and transmission. CONCLUSION: Current dosing recommendations provide a resistance selection opportunity in those patients with low drug levels and high parasite burdens (often children or pregnant women). Patients with hyperparasitaemia who receive outpatient treatments provide the greatest risk of selecting de-novo resistant parasites. This emphasizes the importance of ensuring that only quality-assured anti-malarial combinations are used, that treatment doses are optimized on the basis of pharmacodynamic and pharmacokinetic assessments in the target populations, and that patients with heavy parasite burdens are identified and receive sufficient treatment to prevent recrudescence.
Although hyponatremia occurs in most patients with severe malaria, its pathogenesis, prognostic significance, and optimal management have not been established. Clinical and biochemical data were prospectively collected from 171 consecutive Bangladeshi adults with severe malaria. On admission, 57% of patients were hyponatremic. Plasma sodium and Glasgow Coma Score were inversely related (r(s) = -0.36, P < 0.0001). Plasma antidiuretic hormone concentrations were similar in hyponatremic and normonatremic patients (median, range: 6.1, 2.3-85.3 versus 32.7, 3.0-56.4 pmol/L; P = 0.19). Mortality was lower in hyponatremic than normonatremic patients (31.6% versus 51.4%; odds ratio [95% confidence interval]: 0.44 [0.23-0.82]; P = 0.01 by univariate analysis). Plasma sodium normalized with crystalloid rehydration from (median, range) 127 (123-140) mmol/L on admission to 136 (128-149) mmol/L at 24 hours (P = 0.01). Hyponatremia in adults with severe malaria is common and associated with preserved consciousness and decreased mortality. It likely reflects continued oral hypotonic fluid intake in the setting of hypovolemia and requires no therapy beyond rehydration.
BACKGROUND: Characterization of anti-malarial drug concentration profiles is necessary to optimize dosing, and thereby optimize cure rates and reduce both toxicity and the emergence of resistance. Population pharmacokinetic studies determine the drug concentration time profiles in the target patient populations, including children who have limited sampling options. Currently, population pharmacokinetic studies of anti-malarial drugs are designed based on logistical, financial and ethical constraints, and prior knowledge of the drug concentration time profile. Although these factors are important, the proposed design may be unable to determine the desired pharmacokinetic profile because there was no formal consideration of the complex statistical models used to analyse the drug concentration data. METHODS: Optimal design methods incorporate prior knowledge of the pharmacokinetic profile of the drug, the statistical methods used to analyse data from population pharmacokinetic studies, and also the practical constraints of sampling the patient population. The methods determine the statistical efficiency of the design by evaluating the information of the candidate study design prior to the pharmacokinetic study being conducted. RESULTS: In a hypothetical population pharmacokinetic study of intravenous artesunate, where the number of patients and blood samples to be assayed was constrained to be 50 and 200 respectively, an evaluation of varying elementary designs using optimal design methods found that the designs with more patients and less samples per patient improved the precision of the pharmacokinetic parameters and inter-patient variability, and the overall statistical efficiency by at least 50%. CONCLUSION: Optimal design methods ensure that the proposed study designs for population pharmacokinetic studies are robust and efficient. It is unethical to continue conducting population pharmacokinetic studies when the sampling schedule may be insufficient to estimate precisely the pharmacokinetic profile.
BACKGROUND: The soil-dwelling Gram-negative bacterium Burkholderia pseudomallei is the cause of melioidosis. Extreme structuring of genotype and genotypic frequency has been demonstrated for B. pseudomallei in uncultivated land, but its distribution and genetic diversity in agricultural land where most human infections are probably acquired is not well defined. METHODS: Fixed-interval soil sampling was performed in a rice paddy in northeast Thailand in which 100 grams of soil was sampled at a depth of 30 cm from 10x10 sampling points each measuring 2.5 m by 2.5 m. Soil was cultured for the presence of B. pseudomallei and genotyping of colonies present on primary culture plates was performed using a combination of pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST). PRINCIPAL FINDINGS: B. pseudomallei was cultured from 28/100 samples. Genotyping of 630 primary colonies drawn from 11 sampling points demonstrated 10 PFGE banding pattern types, which on MLST were resolved into 7 sequence types (ST). Overlap of genotypes was observed more often between sampling points that were closely positioned. Two sampling points contained mixed B. pseudomallei genotypes, each with a numerically dominant genotype and one or more additional genotypes present as minority populations. CONCLUSIONS: Genetic diversity and structuring of B. pseudomallei exists despite the effects of flooding and the physical and chemical processes associated with farming. These findings form an important baseline for future studies of environmental B. pseudomallei.
Several antimalarials can cause significant prolongation of the electrocardiograph QT interval, which can be associated with an increased risk of potentially lethal ventricular arrhythmias. High doses of artemether and artemotil have been associated with QT prolongation in dogs, raising the possibility of a class effect with the artemisinin derivatives. Serial electrocardiograms were recorded, and QTc interval was calculated before and after administration of artesunate by intravenous injection in patients with severe falciparum malaria in Bangladesh. Of 21 adult patients with severe malaria enrolled, 8 (38%) died. The mean QTc interval was unaffected by bolus intravenous artesunate (2.4 mg/kg). In two patients, the QTc interval exceeded 0.5 seconds, but in both cases, an alternative explanation was plausible. No effect was observed on the JTc or PR interval, QRS width, blood pressure, or heart rate. Intravenous artesunate does not have significant cardiovascular effects in patients with severe falciparum malaria.
BACKGROUND: Invasive Staphylococcus aureus infection is increasingly recognised as an important cause of serious sepsis across the developing world, with mortality rates higher than those in the developed world. The factors determining mortality in developing countries have not been identified. METHODS: A prospective, observational study of invasive S. aureus disease was conducted at a provincial hospital in northeast Thailand over a 1-year period. All-cause and S. aureus-attributable mortality rates were determined, and the relationship was assessed between death and patient characteristics, clinical presentations, antibiotic therapy and resistance, drainage of pus and carriage of genes encoding Panton-Valentine Leukocidin (PVL). PRINCIPAL FINDINGS: A total of 270 patients with invasive S. aureus infection were recruited. The range of clinical manifestations was broad and comparable to that described in developed countries. All-cause and S. aureus-attributable mortality rates were 26% and 20%, respectively. Early antibiotic therapy and drainage of pus were associated with a survival advantage (both p<0.001) on univariate analysis. Patients infected by a PVL gene-positive isolate (122/248 tested, 49%) had a strong survival advantage compared with patients infected by a PVL gene-negative isolate (all-cause mortality 11% versus 39% respectively, p<0.001). Multiple logistic regression analysis using all variables significant on univariate analysis revealed that age, underlying cardiac disease and respiratory infection were risk factors for all-cause and S. aureus-attributable mortality, while one or more abscesses as the presenting clinical feature and procedures for infectious source control were associated with survival. CONCLUSIONS: Drainage of pus and timely antibiotic therapy are key to the successful management of S. aureus infection in the developing world. Defining the presence of genes encoding PVL provides no practical bedside information and draws attention away from identifying verified clinical risk factors and those interventions that save lives.
BACKGROUND: Malaria has recently been identified as a candidate for global eradication. This process will take the form of a series of national eliminations. Key issues must be considered specifically for elimination strategy when compared to the control of disease. Namely the spread of drug resistance, data scarcity and the adverse effects of failed elimination attempts. Mathematical models of various levels of complexity have been produced to consider the control and elimination of malaria infection. If available, detailed data on malaria transmission (such as the vector life cycle and behaviour, human population behaviour, the acquisition and decay of immunity, heterogeneities in transmission intensity, age profiles of clinical and subclinical infection) can be used to populate complex transmission models that can then be used to design control strategy. However, in many malaria countries reliable data are not available and policy must be formed based on information like an estimate of the average parasite prevalence. METHODS: A simple deterministic model, that requires data in the form of a single estimate of parasite prevalence as an input, is developed for the purpose of comparison with other more complex models. The model is designed to include key aspects of malaria transmission and integrated control. RESULTS: The simple model is shown to have similar short-term dynamic behaviour to three complex models. The model is used to demonstrate the potential of alternative methods of delivery of controls. The adverse effects on clinical infection and spread of resistance are predicted for failed elimination attempts. Since elimination strategies present an increased risk of the spread of drug resistance, the model is used to demonstrate the population level protective effect of multiple controls against this very serious threat. CONCLUSION: A simple model structure for the elimination of malaria is suitable for situations where data are sparse yet strategy design requirements are urgent with the caveat that more complex models, populated with new data, would provide more information, especially in the long-term.
BACKGROUND: Investigations of Plasmodium vivax are restricted to samples collected from infected persons or primates, because this parasite cannot be maintained in in vitro cultures. Contamination of P. vivax isolates with host leukocytes and platelets is detrimental to a range of ex vivo and molecular investigations. Easy-to-produce CF11 cellulose filters have recently provided us with an inexpensive method for the removal of leukocytes and platelets. This contrasted with previous reports of unacceptably high levels of infected red blood cell (IRBC) retention by CF11. The aims of this study were to compare the ability of CF11 cellulose filters and the commercial filter Plasmodipur at removing leukocyte and platelet, and to investigate the retention of P. vivax IRBCs by CF11 cellulose filtration. METHODS AND RESULTS: Side-by-side comparison of six leukocyte removal methods using blood samples from five healthy donor showed that CF11 filtration reduced the mean initial leukocyte counts from 9.4 x 103 per microl [95%CI 5.2-13.5] to 0.01 x 103 [95%CI 0.01-0.03]. The CF11 was particularly effective at removing neutrophils. CF11 treatment also reduced initial platelet counts from 211.6 x 103 per microl [95%CI 107.5-315.7] to 0.8 x 103 per microl [95%CI -0.7-2.2]. Analysis of 30 P. vivax blood samples before and after CF11 filtration showed only a minor loss in parasitaemia (<or= 7.1% of initial counts). Stage specific retention of P. vivax IRBCs was not observed. CONCLUSION: CF11 filtration is the most cost and time efficient method for the production of leukocyte- and platelet-free P. vivax-infected erythrocytes from field isolates.
BACKGROUND: Most information on invasive Staphylococcus aureus infections comes from temperate countries. There are considerable knowledge gaps in epidemiology, treatment, drug resistance and outcome of invasive S. aureus infection in the tropics. METHODS: A prospective, observational study of S. aureus bacteraemia was conducted in a 1000-bed regional hospital in northeast Thailand over 1 year. Detailed clinical data were collected and final outcomes determined at 12 weeks, and correlated with antimicrobial susceptibility profiles of infecting isolates. PRINCIPAL FINDINGS: Ninety-eight patients with S. aureus bacteraemia were recruited. The range of clinical manifestations was similar to that reported from temperate countries. The prevalence of endocarditis was 14%. The disease burden was highest at both extremes of age, whilst mortality increased with age. The all-cause mortality rate was 52%, with a mortality attributable to S. aureus of 44%. Methicillin-resistant S. aureus (MRSA) was responsible for 28% of infections, all of which were healthcare-associated. Mortality rates for MRSA and methicillin-susceptible S. aureus (MSSA) were 67% (18/27) and 46% (33/71), respectively (p = 0.11). MRSA isolates were multidrug resistant. Only vancomycin or fusidic acid would be suitable as empirical treatment options for suspected MRSA infection. CONCLUSIONS: S. aureus is a significant pathogen in northeast Thailand, with comparable clinical manifestations and a similar endocarditis prevalence but higher mortality than industrialised countries. S. aureus bacteraemia is frequently associated with exposure to healthcare settings with MRSA causing a considerable burden of disease. Further studies are required to define setting-specific strategies to reduce mortality from S. aureus bacteraemia, prevent MRSA transmission, and to define the burden of S. aureus disease and emergence of drug resistance throughout the developing world.
BACKGROUND: Artemisinin combination therapy (ACT) is now the recommended first-line treatment for falciparum malaria throughout the world. Initiatives to eliminate malaria are critically dependent on its efficacy. There is recent worrying evidence that artemisinin resistance has arisen on the Thai-Cambodian border. Urgent containment interventions are planned and about to be executed. Mathematical modeling approaches to intervention design are now integrated into the field of malaria epidemiology and control. The use of such an approach to investigate the likely effectiveness of different containment measures with the ultimate aim of eliminating artemisinin-resistant malaria is described. METHODS: A population dynamic mathematical modeling framework was developed to explore the relative effectiveness of a variety of containment interventions in eliminating artemisinin-resistant malaria in western Cambodia. RESULTS: The most effective intervention to eliminate artemisinin-resistant malaria was a switch of treatment from artemisinin monotherapy to ACT (mean time to elimination 3.42 years (95% CI 3.32-3.60 years). However, with this approach it is predicted that elimination of artemisinin-resistant malaria using ACT can be achieved only by elimination of all malaria. This is because the various forms of ACT are more effective against infections with artemisinin-sensitive parasites, leaving the more resistant infections as an increasing proportion of the dwindling parasite population. CONCLUSION: Containment of artemisinin-resistant malaria can be achieved by elimination of malaria from western Cambodia using ACT. The "last man standing" is the most resistant and thus this strategy must be sustained until elimination is truly achieved.
West Nile virus (WNV) has emerged globally as an increasingly important pathogen for humans and domestic animals. Studies of the evolutionary diversity of the virus over its known history will help to elucidate conserved sites, and characterize their correspondence to other pathogens and their relevance to the immune system. We describe a large-scale analysis of the entire WNV proteome, aimed at identifying and characterizing evolutionarily conserved amino acid sequences. This study, which used 2,746 WNV protein sequences collected from the NCBI GenPept database, focused on analysis of peptides of length 9 amino acids or more, which are immunologically relevant as potential T-cell epitopes. Entropy-based analysis of the diversity of WNV sequences, revealed the presence of numerous evolutionarily stable nonamer positions across the proteome (entropy value of < or = 1). The representation (frequency) of nonamers variant to the predominant peptide at these stable positions was, generally, low (< or = 10% of the WNV sequences analyzed). Eighty-eight fragments of length 9-29 amino acids, representing approximately 34% of the WNV polyprotein length, were identified to be identical and evolutionarily stable in all analyzed WNV sequences. Of the 88 completely conserved sequences, 67 are also present in other flaviviruses, and several have been associated with the functional and structural properties of viral proteins. Immunoinformatic analysis revealed that the majority (78/88) of conserved sequences are potentially immunogenic, while 44 contained experimentally confirmed human T-cell epitopes. This study identified a comprehensive catalogue of completely conserved WNV sequences, many of which are shared by other flaviviruses, and majority are potential epitopes. The complete conservation of these immunologically relevant sequences through the entire recorded WNV history suggests they will be valuable as components of peptide-specific vaccines or other therapeutic applications, for sequence-specific diagnosis of a wide-range of Flavivirus infections, and for studies of homologous sequences among other flaviviruses.
BACKGROUND: In Vietnam the blackwater fever syndrome (BWF) has been associated with malaria infection, quinine ingestion and G6PD deficiency. The G6PD variants within the Vietnamese Kinh contributing to the disease risk in this population, and more generally to haemoglobinuria, are currently unknown. METHOD: Eighty-two haemoglobinuria patients and 524 healthy controls were screened for G6PD deficiency using either the methylene blue reduction test, the G-6-PDH kit or the micro-methaemoglobin reduction test. The G6PD gene variants were screened using SSCP combined with DNA sequencing in 82 patients with haemoglobinuria, and in 59 healthy controls found to be G6PD deficient. RESULTS: This study confirmed that G6PD deficiency is strongly associated with haemoglobinuria (OR = 15, 95% CI [7.7 to 28.9], P < 0.0001). Six G6PD variants were identified in the Vietnamese population, of which two are novel (Vietnam1 [Glu3Lys] and Vietnam2 [Phe66Cys]). G6PD Viangchan [Val291Met], common throughout south-east Asia, accounted for 77% of the variants detected and was significantly associated with haemoglobinuria within G6PD-deficient ethnic Kinh Vietnamese (OR = 5.8 95% CI [114-55.4], P = 0.022). CONCLUSION: The primary frequency of several G6PD mutations, including novel mutations, in the Vietnamese Kinh population are reported and the contribution of G6PD mutations to the development of haemoglobinuria are investigated.
BACKGROUND: The fixed dose antimalarial combination of dihydroartemisinin-piperaquine (DP) is a promising new artemisinin-based combination therapy (ACT). We present an individual patient data analysis of efficacy and tolerability in acute uncomplicated falciparum malaria, from seven published randomized clinical trials conducted in Africa and South East Asia using a predefined in-vivo protocol. Comparator drugs were mefloquine-artesunate (MAS3) in Thailand, Myanmar, Laos and Cambodia; artemether-lumefantrine in Uganda; and amodiaquine+sulfadoxine-pyrimethamine and artesunate+amodiaquine in Rwanda. METHODS AND FINDINGS: In total 3,547 patients were enrolled: 1,814 patients (32% children under five years) received DP and 1,733 received a comparator antimalarial at 12 different sites and were followed for 28-63 days. There was no significant heterogeneity between trials. DP was well tolerated with 1.7% early vomiting. There were less adverse events with DP in children and adults compared to MAS3 except for diarrhea; ORs (95%CI) 2.74 (2.13 to 3.51) and 3.11 (2.31 to 4.18), respectively. DP treatment resulted in a rapid clearance of fever and parasitaemia. The PCR genotype corrected efficacy at Day 28 of DP assessed by survival analysis was 98.7% (95%CI 97.6-99.8). DP was superior to the comparator drugs in protecting against both P.falciparum recurrence and recrudescence (P = 0.001, weighted by site). There was no difference between DP and MAS3 in treating P. vivax co-infections and in suppressing the first relapse (median interval to P. vivax recurrence: 6 weeks). Children under 5 y were at higher risk of recurrence for both infections. The proportion of patients developing gametocytaemia (P = 0.002, weighted by site) and the subsequent gametocyte carriage rates were higher with DP (11/1000 person gametocyte week, PGW) than MAS3 (6/1000 PGW, P = 0.001, weighted by site). CONCLUSIONS: DP proved a safe, well tolerated, and highly effective treatment of P.falciparum malaria in Asia and Africa, but the effect on gametocyte carriage was inferior to that of MAS3.
BACKGROUND: Pharmacokinetic (PK) data on amodiaquine (AQ) and artesunate (AS) are limited in children, an important risk group for malaria. The aim of this study was to evaluate the PK properties of a newly developed and registered fixed dose combination (FDC) of artesunate and amodiaquine. METHODS: A prospective population pharmacokinetic study of AS and AQ was conducted in children aged six months to five years. Participants were randomized to receive the new artesunate and amodiaquine FDC or the same drugs given in separate tablets. Children were divided into two groups of 70 (35 in each treatment arm) to evaluate the pharmacokinetic properties of AS and AQ, respectively. Population pharmacokinetic models for dihydroartemisinin (DHA) and desethylamodiaquine (DeAq), the principal pharmacologically active metabolites of AS and AQ, respectively, and total artemisinin anti-malarial activity, defined as the sum of the molar equivalent plasma concentrations of DHA and artesunate, were constructed using the non-linear mixed effects approach. Relative bioavailability between products was compared by estimating the ratios (and 95% CI) between the areas under the plasma concentration-time curves (AUC). RESULTS: The two regimens had similar PK properties in young children with acute malaria. The ratio of loose formulation to fixed co-formulation AUCs, was estimated as 1.043 (95% CI: 0.956 to 1.138) for DeAq. For DHA and total anti-malarial activity AUCs were estimated to be the same. Artesunate was rapidly absorbed, hydrolysed to DHA, and eliminated. Plasma concentrations were significantly higher following the first dose, when patients were acutely ill, than after subsequent doses when patients were usually afebrile and clinically improved. Amodiaquine was converted rapidly to DeAq, which was then eliminated with an estimated median (range) elimination half-life of 9 (7 to 12) days. Efficacy was similar in the two treatments groups, with cure rates of 0.946 (95% CI: 0.840-0.982) in the AS+AQ group and 0.892 (95% CI: 0.787 - 0.947) in the AS/AQ group. Four out of five patients with PCR confirmed recrudescences received AQ doses < 10 mg/kg. Both regimens were well tolerated. No child developed severe, post treatment neutropaenia (<1,000/muL). There was no evidence of AQ dose related hepatotoxicity, but one patient developed an asymptomatic rise in liver enzymes that was resolving by Day-28. CONCLUSION: The bioavailability of the co-formulated AS-AQ FDC was similar to that of the separate tablets for desethylamodiaquine, DHA and the total anti-malarial activity. These data support the use this new AS-AQ FDC in children with acute uncomplicated falciparum malaria.
BACKGROUND: Mefloquine and artesunate combination therapy is the recommended first-line treatment for uncomplicated malaria throughout much of south-east Asia. Concerns have been raised about the potential central nervous system (CNS) effects of both drug components and there are no detailed reports in very young children. METHODS: Children, aged between three months and five years, with acute uncomplicated Plasmodium falciparum malaria were randomized to either 7 days of artesunate monotherapy or the same schedule of artesunate plus mefloquine on day 7 and 8. Neurological testing targeting coordination and behaviour was carried out at day 0, 7, 9, 10, 14 and 28. Non-febrile healthy control children from the same population were tested on days 0, 7, 14 and 28. RESULTS: From December 1994 to July 1997, 91 children with uncomplicated P. falciparum, 45 treated with artesunate monotherapy, 46 treated with mefloquine and artesunate combination therapy and 36 non-febrile controls, underwent neurological testing. Malaria and fever had a significant negative impact on testing performance. By contrast, the anti-malarial treatments were not associated with worsening performances in the various components of the test. Artesunate and mefloquine do not appear to have a significant influence on coordination and behaviour. Children treated with mefloquine were significantly less likely to suffer recurrent malaria infection during follow-up compared to those treated with artesunate alone (P = 0.033). CONCLUSION: In keeping with the results of randomized controlled trials in adults, mefloquine was not associated with a decrease in specific items of neurological performance. Likewise, children treated with artesunate did not perform significantly differently to control children. This study does not exclude subtle or rare treatment CNS effects of artesunate or mefloquine. Treatment of acute uncomplicated malaria results in a significant improvement on items of neurological performance.
BACKGROUND: A number of molecular tools have been developed to monitor the emergence and spread of anti-malarial drug resistance to Plasmodium falciparum. One of the major obstacles to the wider implementation of these tools is the absence of practical methods enabling high throughput analysis. Here a new Zip-code array is described, called FlexiChip, linked to a dedicated software program, which largely overcomes this problem. METHODS: Previously published microarray probes detecting single-nucleotide polymorphisms (SNP) associated with parasite resistance to anti-malarial drugs (ResMalChip) were adapted for a universal microarray FlexiChip format. To evaluate the overall sensitivity of the FlexiChip package (microarray + software), the results of FlexiChip were compared to ResMalChip microarray, using the same extension probes and with the same PCR products. In both cases, sequence results were used as gold standard to calculate sensitivity and specificity. FlexiChip results obtained with a set of field isolates were then compared to those assessed in an independent reference laboratory. RESULTS: The FlexiChip package gave results identical to the ResMalChip results in 92.7% of samples (kappa coefficient 0.8491, with a standard error 0.021) and had a sensitivity of 95.88% and a specificity of 97.68% compared to the sequencing as the reference method. Moreover the method performed well compared to the results obtained in the reference laboratories, with 99.7% of identical results (kappa coefficient 0.9923, S.E. 0.0523). CONCLUSION: Microarrays could be employed to monitor P. falciparum drug resistance markers with greater cost effectiveness and the possibility for high throughput analysis. The FlexiChip package is a promising tool for use in poor resource settings of malaria endemic countries.
BACKGROUND: In areas where non-falciparum malaria is common rapid diagnostic tests (RDTs) capable of distinguishing malaria species reliably are needed. Such tests are often based on the detection of parasite lactate dehydrogenase (pLDH). METHODS: In Dawei, southern Myanmar, three pLDH based RDTs (CareStart Malaria pLDH (Pan), CareStart Malaria pLDH (Pan, Pf) and OptiMAL-IT)were evaluated in patients presenting with clinically suspected malaria. Each RDT was read independently by two readers. A subset of patients with microscopically confirmed malaria had their RDTs repeated on days 2, 7 and then weekly until negative. At the end of the study, samples of study batches were sent for heat stability testing. RESULTS: Between August and November 2007, 1004 patients aged between 1 and 93 years were enrolled in the study. Slide microscopy (the reference standard) diagnosed 213 Plasmodium vivax (Pv) monoinfections, 98 Plasmodium falciparum (Pf) mono-infections and no malaria in 650 cases. The sensitivities (sens) and specificities (spec), of the RDTs for the detection of malaria were- CareStart Malaria pLDH (Pan) test: sens 89.1% [CI95 84.2-92.6], spec 97.6% [CI95 96.5-98.4]. OptiMal-IT: Pf+/- other species detection: sens 95.2% [CI95 87.5-98.2], spec 94.7% [CI95 93.3-95.8]; non-Pf detection alone: sens 89.6% [CI95 83.6-93.6], spec 96.5% [CI95 94.8-97.7]. CareStart Malaria pLDH (Pan, Pf): Pf+/- other species: sens 93.5% [CI95 85.4-97.3], spec 97.4% [95.9-98.3]; non-Pf: sens 78.5% [CI95 71.1-84.4], spec 97.8% [CI95 96.3-98.7]. Inter-observer agreement was excellent for all tests (kappa > 0.9). The median time for the RDTs to become negative was two days for the CareStart Malaria tests and seven days for OptiMAL-IT. Tests were heat stable up to 90 days except for OptiMAL-IT (Pf specific pLDH stable to day 20 at 35 degrees C). CONCLUSION: None of the pLDH-based RDTs evaluated was able to detect non-falciparum malaria with high sensitivity, particularly at low parasitaemias. OptiMAL-IT performed best overall and would perform best in an area of high malaria prevalence among screened fever cases. However, heat stability was unacceptable and the number of steps to perform this test is a significant drawback in the field. A reliable, heat-stable, highly sensitive RDT, capable of diagnosing all Plasmodium species has yet to be identified.
ANNALS OF NUTRITION AND METABOLISM, 55 pp. 215-215.2009. MUAC AND BMI: USEFULNESS IN PREGNANCY AMONG REFUGEE AND MIGRANT POPULATIONS ALONG THE THAI-MYANMAR BORDER
ANNALS OF NUTRITION AND METABOLISM, 55 pp. 299-299.2009. GROWTH VELOCITY OF TERM INFANTS BORN IN MAELA CAMP FOR DISPLACED POPULATION ALONG THE THAI-MYANMAR BORDER
ANNALS OF NUTRITION AND METABOLISM, 55 pp. 440-441.2009. IRON, THIAMINE, AND MICRONUTRIENT STATUS IN PREGNANT AND BREAST-FEEDING WOMEN: ASSESSMENT OF FOOD RATIONS AND SUPPLEMENTATION IN MAELA REFUGEE CAMP, NORTHERN THAILAND
Background: The fixed dose antimalarial combination of dihydroartemisinin-piperaquine (DP) is a promising new artemisinin-based combination therapy (ACT). We present an individual patient data analysis of efficacy and tolerability in acute uncomplicated falciparum malaria, from seven published randomized clinical trials conducted in Africa and South East Asia using a predefined in-vivo protocol. Comparator drugs were mefloquine-artesunate (MAS3) in Thailand, Myanmar, Laos and Cambodia; artemether-lumefantrine in Uganda; and amodiaquine+sulfadoxine-pyrimethamine and artesunate+amodiaquine in Rwanda. Methods and Findings: In total 3,547 patients were enrolled: 1,814 patients (32% children under five years) received DP and 1,733 received a comparator antimalarial at 12 different sites and were followed for 28-63 days. There was no significant heterogeneity between trials. DP was well tolerated with 1.7% early vomiting. There were less adverse events with DP in children and adults compared to MAS3 except for diarrhea; ORs (95%CI) 2.74 (2.13 to 3.51) and 3.11 (2.31 to 4.18), respectively. DP treatment resulted in a rapid clearance of fever and parasitaemia. The PCR genotype corrected efficacy at Day 28 of DP assessed by survival analysis was 98.7% (95%CI 97.6-99.8). DP was superior to the comparator drugs in protecting against both P.falciparum recurrence and recrudescence (P = 0.001, weighted by site). There was no difference between DP and MAS3 in treating P. vivax co-infections and in suppressing the first relapse (median interval to P. vivax recurrence: 6 weeks). Children under 5 y were at higher risk of recurrence for both infections. The proportion of patients developing gametocytaemia (P = 0.002, weighted by site) and the subsequent gametocyte carriage rates were higher with DP (11/1000 person gametocyte week, PGW) than MAS3 (6/ 1000 PGW, P = 0.001, weighted by site). Conclusions: DP proved a safe, well tolerated, and highly effective treatment of P.falciparum malaria in Asia and Africa, but the effect on gametocyte carriage was inferior to that of MAS3. © 2009 Zwang et al.
N Engl J Med, 361 (18), pp. 1808-1809. | Citations: 3 (European Pubmed Central) | Read more2009. Peginterferon alfa-2b or alfa-2a with ribavirin for hepatitis C.
médecine/sciences, 25 (10), pp. 867-869. | Citations: 2 (Scopus) | Read more2009. Paludisme et grossesse : un dilemme thérapeutique
European Journal of Clinical Pharmacology, 65 (8), pp. 847. | Read more2009. Erratum:The pharmacokinetics of artemether and lumefantrine in pregnant women with uncomplicated falciparum malaria(European Journal of Clinical Pharmacology (2006) 62 (1021-1031) DOI: 10.1007/s00228-006-0199-7)
European Journal of Clinical Nutrition, 63 (3), pp. 450. | Read more2009. Erratum: Postpartum traditions and nutrition practices among urban Lao women and their infants in Vientiane, Lao PDR (European Journal of Clinical Nutrition (2009) vol. 63 (323-331) 10.1038/sj.ejcn.1602928)
BACKGROUND: Chromobacterium violaceum is a Gram negative facultative anaerobic bacillus, found in soil and stagnant water, that usually has a violet pigmented appearance on agar culture. It is rarely described as a human pathogen, mostly from tropical and subtropical areas. CASE PRESENTATION: A 53 year-old farmer died with Chromobacterium violaceum septicemia in Laos. A modified oxidase method was used to demonstrate that this violacious organism was oxidase positive. Forensic analysis of the glucose-6-phosphate dehydrogenase genotypes of his family suggest that the deceased patient did not have this possible predisposing condition. CONCLUSION: C. violaceum infection should be included in the differential diagnosis in patients presenting with community-acquired septicaemia in tropical and subtropical areas. The apparently neglected but simple modified oxidase test may be useful in the oxidase assessment of other violet-pigmented organisms or of those growing on violet coloured agar.
BACKGROUND: Little is known about how primary care physicians (PCPs) in Asia diagnose and manage prostatitis-like symptoms. This study investigated the clinical diagnosis of and care provided for prostatitis-like symptoms by PCPs in a Malaysian population, and compared these findings to reports from other areas. METHODS: All members of the Penang Private Medical Practitioners' Society were asked to complete a self-administered survey. Nonresponders were contacted after 3 weeks and received a telephone request after 6 weeks. RESULTS: Of the 786 practitioners contacted, 669 considered themselves to be PCPs, including 279 (42%) who responded to the survey. Adult males with prostatitis-like symptoms typically constitute <1% of the patients seen by PCPs. Most PCPs (72%) believe that prostatitis-like symptoms are caused by bacterial infection. 61% of PCPs base their diagnosis of prostatitis-like symptoms on clinical history, a physical examination and dipstick urinalysis. Standard management was to prescribe 1 or 2 courses of antimicrobials. CONCLUSIONS: Despite the 8.7% prevalence found in a previous survey in this population, prostatitis remains underdiagnosed in Malaysia. In contrast to many other clinical settings, urologists in Malaysia see a large proportion of newly diagnosed and treatment-naive prostatitis patients, providing an opportunity for clinical diagnostic and treatment studies.
Cochrane Database of Systematic Reviews, (4), | Read more2009. Antiviral prophylaxis for varicella zoster in immunocompromised patients (excluding haematological malignancies)
BACKGROUND: Selective digestive tract decontamination (SDD) and selective oropharyngeal decontamination (SOD) are infection-prevention measures used in the treatment of some patients in intensive care, but reported effects on patient outcome are conflicting. METHODS: We evaluated the effectiveness of SDD and SOD in a crossover study using cluster randomization in 13 intensive care units (ICUs), all in The Netherlands. Patients with an expected duration of intubation of more than 48 hours or an expected ICU stay of more than 72 hours were eligible. In each ICU, three regimens (SDD, SOD, and standard care) were applied in random order over the course of 6 months. Mortality at day 28 was the primary end point. SDD consisted of 4 days of intravenous cefotaxime and topical application of tobramycin, colistin, and amphotericin B in the oropharynx and stomach. SOD consisted of oropharyngeal application only of the same antibiotics. Monthly point-prevalence studies were performed to analyze antibiotic resistance. RESULTS: A total of 5939 patients were enrolled in the study, with 1990 assigned to standard care, 1904 to SOD, and 2045 to SDD; crude mortality in the groups at day 28 was 27.5%, 26.6%, and 26.9%, respectively. In a random-effects logistic-regression model with age, sex, Acute Physiology and Chronic Health Evaluation (APACHE II) score, intubation status, and medical specialty used as covariates, odds ratios for death at day 28 in the SOD and SDD groups, as compared with the standard-care group, were 0.86 (95% confidence interval [CI], 0.74 to 0.99) and 0.83 (95% CI, 0.72 to 0.97), respectively. CONCLUSIONS: In an ICU population in which the mortality rate associated with standard care was 27.5% at day 28, the rate was reduced by an estimated 3.5 percentage points with SDD and by 2.9 percentage points with SOD. (Controlled Clinical Trials number, ISRCTN35176830.)
BACKGROUND: Accurate measures of the severity of pandemic (H1N1) 2009 influenza (pH1N1) are needed to assess the likely impact of an anticipated resurgence in the autumn in the Northern Hemisphere. Severity has been difficult to measure because jurisdictions with large numbers of deaths and other severe outcomes have had too many cases to assess the total number with confidence. Also, detection of severe cases may be more likely, resulting in overestimation of the severity of an average case. We sought to estimate the probabilities that symptomatic infection would lead to hospitalization, ICU admission, and death by combining data from multiple sources. METHODS AND FINDINGS: We used complementary data from two US cities: Milwaukee attempted to identify cases of medically attended infection whether or not they required hospitalization, while New York City focused on the identification of hospitalizations, intensive care admission or mechanical ventilation (hereafter, ICU), and deaths. New York data were used to estimate numerators for ICU and death, and two sources of data--medically attended cases in Milwaukee or self-reported influenza-like illness (ILI) in New York--were used to estimate ratios of symptomatic cases to hospitalizations. Combining these data with estimates of the fraction detected for each level of severity, we estimated the proportion of symptomatic patients who died (symptomatic case-fatality ratio, sCFR), required ICU (sCIR), and required hospitalization (sCHR), overall and by age category. Evidence, prior information, and associated uncertainty were analyzed in a Bayesian evidence synthesis framework. Using medically attended cases and estimates of the proportion of symptomatic cases medically attended, we estimated an sCFR of 0.048% (95% credible interval [CI] 0.026%-0.096%), sCIR of 0.239% (0.134%-0.458%), and sCHR of 1.44% (0.83%-2.64%). Using self-reported ILI, we obtained estimates approximately 7-9 x lower. sCFR and sCIR appear to be highest in persons aged 18 y and older, and lowest in children aged 5-17 y. sCHR appears to be lowest in persons aged 5-17; our data were too sparse to allow us to determine the group in which it was the highest. CONCLUSIONS: These estimates suggest that an autumn-winter pandemic wave of pH1N1 with comparable severity per case could lead to a number of deaths in the range from considerably below that associated with seasonal influenza to slightly higher, but with the greatest impact in children aged 0-4 and adults 18-64. These estimates of impact depend on assumptions about total incidence of infection and would be larger if incidence of symptomatic infection were higher or shifted toward adults, if viral virulence increased, or if suboptimal treatment resulted from stress on the health care system; numbers would decrease if the total proportion of the population symptomatically infected were lower than assumed.
BACKGROUND: The effectiveness of single-drug antiviral interventions to reduce morbidity and mortality during the next influenza pandemic will be substantially weakened if transmissible strains emerge which are resistant to the stockpiled antiviral drugs. We developed a mathematical model to test the hypothesis that a small stockpile of a secondary antiviral drug could be used to mitigate the adverse consequences of the emergence of resistant strains. METHODS AND FINDINGS: We used a multistrain stochastic transmission model of influenza to show that the spread of antiviral resistance can be significantly reduced by deploying a small stockpile (1% population coverage) of a secondary drug during the early phase of local epidemics. We considered two strategies for the use of the secondary stockpile: early combination chemotherapy (ECC; individuals are treated with both drugs in combination while both are available); and sequential multidrug chemotherapy (SMC; individuals are treated only with the secondary drug until it is exhausted, then treated with the primary drug). We investigated all potentially important regions of unknown parameter space and found that both ECC and SMC reduced the cumulative attack rate (AR) and the resistant attack rate (RAR) unless the probability of emergence of resistance to the primary drug p(A) was so low (less than 1 in 10,000) that resistance was unlikely to be a problem or so high (more than 1 in 20) that resistance emerged as soon as primary drug monotherapy began. For example, when the basic reproductive number was 1.8 and 40% of symptomatic individuals were treated with antivirals, AR and RAR were 67% and 38% under monotherapy if p(A) = 0.01. If the probability of resistance emergence for the secondary drug was also 0.01, then SMC reduced AR and RAR to 57% and 2%. The effectiveness of ECC was similar if combination chemotherapy reduced the probabilities of resistance emergence by at least ten times. We extended our model using travel data between 105 large cities to investigate the robustness of these resistance-limiting strategies at a global scale. We found that as long as populations that were the main source of resistant strains employed these strategies (SMC or ECC), then those same strategies were also effective for populations far from the source even when some intermediate populations failed to control resistance. In essence, through the existence of many wild-type epidemics, the interconnectedness of the global network dampened the international spread of resistant strains. CONCLUSIONS: Our results indicate that the augmentation of existing stockpiles of a single anti-influenza drug with smaller stockpiles of a second drug could be an effective and inexpensive epidemiological hedge against antiviral resistance if either SMC or ECC were used. Choosing between these strategies will require additional empirical studies. Specifically, the choice will depend on the safety of combination therapy and the synergistic effect of one antiviral in suppressing the emergence of resistance to the other antiviral when both are taken in combination.
PLoS Curr, 1 pp. RRN1130. | Citations: 26 (European Pubmed Central) | Read more2009. The Early Transmission Dynamics of H1N1pdm Influenza in the United Kingdom.
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