IgM in human immunity to Plasmodium falciparum malaria.

Most studies on human immunity to malaria have focused on the roles of immunoglobulin G (IgG), whereas the roles of IgM remain undefined. Analyzing multiple human cohorts to assess the dynamics of malaria-specific IgM during experimentally induced and naturally acquired malaria, we identified IgM activity against blood-stage parasites. We found that merozoite-specific IgM appears rapidly in Plasmodium falciparum infection and is prominent during malaria in children and adults with lifetime exposure, together with IgG. Unexpectedly, IgM persisted for extended periods of time; we found no difference in decay of merozoite-specific IgM over time compared to that of IgG. IgM blocked merozoite invasion of red blood cells in a complement-dependent manner. IgM was also associated with significantly reduced risk of clinical malaria in a longitudinal cohort of children. These findings suggest that merozoite-specific IgM is an important functional and long-lived antibody response targeting blood-stage malaria parasites that contributes to malaria immunity.

A study protocol for a randomised open-label clinical trial of artesunate-mefloquine versus chloroquine in patients with non-severe Plasmodium knowlesi malaria in Sabah, Malaysia (ACT KNOW trial).

INTRODUCTION: Malaria due to Plasmodium knowlesi is reported throughout South-East Asia, and is the commonest cause of it in Malaysia. P. knowlesi replicates every 24 h and can cause severe disease and death. Current 2010 WHO Malaria Treatment Guidelines have no recommendations for the optimal treatment of non-severe knowlesi malaria. Artemisinin-combination therapies (ACT) and chloroquine have each been successfully used to treat knowlesi malaria; however, the rapidity of parasite clearance has not been prospectively compared. Malaysia's national policy for malaria pre-elimination involves mandatory hospital admission for confirmed malaria cases with discharge only after two negative blood films; use of a more rapidly acting antimalarial agent would have health cost benefits. P. knowlesi is commonly microscopically misreported as P. malariae, P. falciparum or P. vivax, with a high proportion of the latter two species being chloroquine-resistant in Malaysia. A unified ACT-treatment protocol would provide effective blood stage malaria treatment for all Plasmodium species. METHODS AND ANALYSIS: ACT KNOW, the first randomised controlled trial ever performed in knowlesi malaria, is a two-arm open-label trial with enrolments over a 2-year period at three district sites in Sabah, powered to show a difference in proportion of patients negative for malaria by microscopy at 24 h between treatment arms (clinicaltrials.gov #NCT01708876). Enrolments started in December 2012, with completion expected by September 2014. A total sample size of 228 is required to give 90% power (α 0.05) to determine the primary end point using intention-to-treat analysis. Secondary end points include parasite clearance time, rates of recurrent infection/treatment failure to day 42, gametocyte carriage throughout follow-up and rates of anaemia at day 28, as determined by survival analysis. ETHICS AND DISSEMINATION: This study has been approved by relevant institutional ethics committees in Malaysia and Australia. Results will be disseminated to inform knowlesi malaria treatment policy in this region through peer-reviewed publications and academic presentations. TRIAL REGISTRATION NUMBER: NCT01708876.

Factors that are associated with the risk of acquiring Plasmodium knowlesi malaria in Sabah, Malaysia: a case-control study protocol.

INTRODUCTION: Plasmodium knowlesi has long been present in Malaysia, and is now an emerging cause of zoonotic human malaria. Cases have been confirmed throughout South-East Asia where the ranges of its natural macaque hosts and Anopheles leucosphyrus group vectors overlap. The majority of cases are from Eastern Malaysia, with increasing total public health notifications despite a concurrent reduction in Plasmodium falciparum and P. vivax malaria. The public health implications are concerning given P. knowlesi has the highest risk of severe and fatal disease of all Plasmodium spp in Malaysia. Current patterns of risk and disease vary based on vector type and competence, with individual exposure risks related to forest and forest-edge activities still poorly defined. Clustering of cases has not yet been systematically evaluated despite reports of peri-domestic transmission and known vector competence for human-to-human transmission. METHODS AND ANALYSIS: A population-based case-control study will be conducted over a 2-year period at two adjacent districts in north-west Sabah, Malaysia. Confirmed malaria cases presenting to the district hospital sites meeting relevant inclusion criteria will be requested to enrol. Three community controls matched to the same village as the case will be selected randomly. Study procedures will include blood sampling and administration of household and individual questionnaires to evaluate potential exposure risks associated with acquisition of P. knowlesi malaria. Secondary outcomes will include differences in exposure variables between P. knowlesi and other Plasmodium spp, risk of severe P. knowlesi malaria, and evaluation of P. knowlesi case clustering. Primary analysis will be per protocol, with adjusted ORs for exposure risks between cases and controls calculated using conditional multiple logistic regression models. ETHICS: This study has been approved by the human research ethics committees of Malaysia, the Menzies School of Health Research, Australia, and the London School of Hygiene and Tropical Medicine, UK.

Highly effective therapy for maternal malaria associated with a lower risk of vertical transmission.

BACKGROUND: The epidemiology of congenital malaria was investigated in a hospital-based malaria surveillance study in Papua, Indonesia. METHODS: From April 2005 to January 2010, 4878 delivering women and their newborns underwent prospective clinical review and malaria screening by peripheral blood microscopy. FINDINGS: Congenital malaria occurred in 8 per 1000 (38/4884) live births, with Plasmodium falciparum accounting for 76.3% (29) and P. vivax for 15.8% (6) of infections. Maternal malaria at delivery (adjusted odds ratio [AOR], 9.5; 95% confidence interval [CI], 4.2-21.5; P < .001), age ≤ 16 years (AOR, 4; 95% CI, 1.4-12.1; P = .011), and prior malaria during pregnancy (AOR, 2.2; 95% CI, 1.1-4.4, P = .022) were independent risk factors for vertical transmission. Of 29 mothers and neonates with contemporaneous peripheral parasitemia, 17% (5) had discordant parasite species, suggesting possible antenatal malaria transmission. Newborns with malaria were at significantly greater risk of low birth weight (AOR, 2.8; 95% CI, 1.2-6.6; P = .002). Following introduction of dihydroartemisinin-piperaquine for uncomplicated malaria in the second and third trimesters of pregnancy, congenital malaria incidence fell from 3.2% to 0.2% (odds ratio, 0.07; 95% CI, .03-.15; P < .001). CONCLUSIONS: Congenital malaria is an important cause of neonatal morbidity in this region co-endemic for P. falciparum and P. vivax malaria. The introduction of artemisinin-combination therapy was associated with a significant risk reduction in the vertical transmission of malaria.

In vivo and in vitro efficacy of chloroquine against Plasmodium malariae and P. ovale in Papua, Indonesia.

Reports of potential drug-resistant strains of Plasmodium malariae in western Indonesia raise concerns that chloroquine resistance may be emerging in P. malariae and P. ovale. In order to assess this, in vivo and in vitro efficacy studies were conducted in patients with monoinfection in Papua, Indonesia. Consecutive patients with uncomplicated malaria due to P. ovale or P. malariae were enrolled in a prospective clinical trial, provided with supervised chloroquine treatment, and followed for 28 days. Blood from patients with P. malariae or P. ovale parasitemia greater than 1,000 per microliter underwent in vitro antimalarial drug susceptibility testing using a modified schizont maturation assay. Of the 57 evaluable patients in the clinical study (P. malariae, n = 46; P. ovale, n = 11), none had recurrence with the same species during follow-up. The mean parasite reduction ratio at 48 h was 86 (95% confidence interval [CI], 57 to 114) for P. malariae and 150 (95% CI, 54 to 245) for P. ovale (P = 0.18). One patient infected with P. malariae, with 93% of parasites at the trophozoite stage, was still parasitemic on day 4. In vitro drug susceptibility assays were carried out successfully for 40 isolates (34 infected with P. malariae and 6 with P. ovale). The P. malariae infections at trophozoite stages had significantly higher chloroquine 50% effective concentrations (EC(50)s) (median, 127.9 nM [range, 7.9 to 2,980]) than those initially exposed at the ring stage (median, 14.0 nM [range, 3.5 to 27.0]; P = 0.01). The EC(50) for chloroquine in P. ovale was also higher in an isolate initially at the trophozoite stage (23.2 nM) than in the three isolates predominantly at ring stage (7.8 nM). Chloroquine retains adequate efficacy against P. ovale and P. malariae, but its marked stage specificity of action may account for reports of delayed parasite clearance times.

In vitro activity of pyronaridine against multidrug-resistant Plasmodium falciparum and Plasmodium vivax.

Pyronaridine, a Mannich base antimalarial, has demonstrated high in vivo and in vitro efficacy against chloroquine-resistant Plasmodium falciparum. Although this drug has the potential to become a prominent artemisinin combination therapy, little is known about its efficacy against drug-resistant Plasmodium vivax. The in vitro antimalarial susceptibility of pyronaridine was assessed in multidrug-resistant P. vivax (n = 99) and P. falciparum (n = 90) isolates from Papua, Indonesia, using a schizont maturation assay. The median 50% inhibitory concentration (IC(50)) of pyronaridine was 1.92 nM (range, 0.24 to 13.8 nM) against P. falciparum and 2.58 nM (range, 0.13 to 43.6 nM) against P. vivax, with in vitro susceptibility correlating significantly with chloroquine, amodiaquine, and piperaquine (r(s) [Spearman's rank correlation coefficient] = 0.45 to 0.62; P < 0.001). P. falciparum parasites initially at trophozoite stage had higher IC(50)s of pyronaridine than those exposed at the ring stage (8.9 nM [range, 0.6 to 8.9 nM] versus 1.6 nM [range, 0.6 to 8.9 nM], respectively; P = 0.015), although this did not reach significance for P. vivax (4.7 nM [range, 1.4 to 18.7 nM] versus 2.5 nM [range, 1.4 to 15.6 nM], respectively; P = 0.085). The excellent in vitro efficacy of pyronaridine against both chloroquine-resistant P. vivax and P. falciparum highlights the suitability of the drug as a novel partner for artemisinin-based combination therapy in regions where the two species are coendemic.

Pulmonary tuberculosis, impaired lung function, disability and quality of life in a high-burden setting.

SETTING: Tuberculosis treatment clinic in Papua, Indonesia. OBJECTIVE: To document the impact of pulmonary tuberculosis (PTB) on lung function, exercise tolerance and quality of life (QOL). DESIGN: A prospective cohort study of 115 patients with smear-positive PTB followed for 6 months. Demographics, disease history, sputum microbiology, spirometry, 6-minute weight.walk distance (6MWWD) and QOL (modified St George's Respiratory Questionnaire) were measured at diagnosis and at 2 and 6 months. Analysis was restricted to the 69/115 (60%) subjects who attended all follow-up visits. RESULTS: Subjects who attended all visits were less likely than the full cohort to be of Papuan ethnicity (P < 0.05), were more likely to be cured (P < 0.001) and had better lung function at diagnosis (P < 0.05). Significant lung function impairment (forced expiratory volume in 1 second [FEV(1)] <60% predicted) was found in 27/69 (39%) at diagnosis. Although this fell during treatment (P < 0.01), 17/69 (24.6%) had persisting significant lung function impairment at treatment completion. As lung function recovered, exercise tolerance (6MWWD) rose by 12.3% (P < 0.001) and QOL improved (P < 0.001). CONCLUSION: In a high-burden setting, PTB causes prolonged, significant impairment of lung function, exercise tolerance and QOL. Current measures of disease burden are likely to underestimate the true impact of disease. Earlier diagnosis and disease-modifying treatments may reduce the long-term impact of PTB.

Cost-effectiveness of artesunate for the treatment of severe malaria.

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.

In vivo and in vitro efficacy of amodiaquine monotherapy for treatment of infection by chloroquine-resistant Plasmodium vivax.

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.

Amplification of pvmdr1 associated with multidrug-resistant Plasmodium vivax.

BACKGROUND: Multidrug-resistant strains of Plasmodium vivax are emerging in Southeast Asia. METHODS: In vitro drug susceptibility and pvmdr1 genotype were determined in P. vivax field isolates from Indonesia and Thailand. RESULTS: Increased pvmdr1 copy number was present in 21% of isolates from Thailand (15/71) and none from Indonesia (0/114; P < .001). Compared with Indonesian isolates, the median IC(50) of Thai isolates was lower for chloroquine (36 vs. 114 nmol/L; P < .001) but higher for amodiaquine (34 vs. 13.7 nmol/L; P = .032), artesunate (8.33 vs. 1.58 nmol/L; P < .001), and mefloquine (111 vs. 9.87 nmol/L; P < .001). In 11 cryopreserved Thai isolates, those with increased pvmdr1 copy number had a higher IC(50) for mefloquine (78.6 vs. 38 nmol/L for single-copy isolates; P = .006). Compared with isolates with the wild-type allele, the Y976F mutation of pvmdr1 was associated with reduced susceptibility to chloroquine (154 nmol/L [range, 4.6-3505] vs. 34 nmol/L [range, 6.7-149]; P < .001) but greater susceptibility to artesunate (1.8 vs. 9.5 nmol/L; P = .009) and mefloquine (14 vs. 121 nmol/L; P < .001). CONCLUSIONS: Amplification of pvmdr1 and single-nucleotide polymorphisms are correlated with susceptibility of P. vivax to multiple antimalarial drugs. Chloroquine and mefloquine appear to exert competitive evolutionary pressure on pvmdr1, similar to that observed with pfmdr1 in Plasmodium falciparum.

Stronger activity of human immunodeficiency virus type 1 protease inhibitors against clinical isolates of Plasmodium vivax than against those of P. falciparum.

Recent studies using laboratory clones have demonstrated that several antiretroviral protease inhibitors (PIs) inhibit the growth of Plasmodium falciparum at concentrations that may be of clinical significance, especially during human immunodeficiency virus type 1 (HIV-1) and malaria coinfection. Using clinical isolates, we now demonstrate the in vitro effectiveness of two HIV-1 aspartic PIs, saquinavir (SQV) and ritonavir (RTV), against P. vivax (n = 30) and P. falciparum (n = 20) from populations subjected to high levels of mefloquine and artesunate pressure on the Thailand-Myanmar border. The median 50% inhibitory concentration values of P. vivax to RTV and SQV were 2,233 nM (range, 732 to 7,738 nM) and 4,230 nM (range, 1,326 to 8,452 nM), respectively, both within the therapeutic concentration range commonly found for patients treated with these PIs. RTV was fourfold more effective at inhibiting P. vivax than it was at inhibiting P. falciparum, compared to a twofold difference in SQV sensitivity. An increased P. falciparum mdr1 copy number was present in 33% (3/9) of isolates and that of P. vivax mdr1 was present in 9% of isolates (2/22), but neither was associated with PI sensitivity. The inter-Plasmodium sp. variations in PI sensitivity indicate key differences between P. vivax and P. falciparum. PI-containing antiretroviral regimens may demonstrate prophylactic activity against both vivax and falciparum malaria in HIV-infected patients who reside in areas where multidrug-resistant P. vivax or P. falciparum is found.

Determinants of in vitro drug susceptibility testing of Plasmodium vivax.

In Papua, Indonesia, the antimalarial susceptibility of Plasmodium vivax (n = 216) and P. falciparum (n = 277) was assessed using a modified schizont maturation assay for chloroquine, amodiaquine, artesunate, lumefantrine, mefloquine, and piperaquine. The most effective antimalarial against P. vivax and P. falciparum was artesunate, with geometric mean 50% inhibitory concentrations (IC50s) (95% confidence intervals [CI]) of 1.31 nM (1.07 to 1.59) and 0.64 nM (0.53 to 0.79), respectively. In contrast, the geometric mean chloroquine IC50 for P. vivax was 295 nM (227 to 384) compared to only 47.4 nM (42.2 to 53.3) for P. falciparum. Two factors were found to significantly influence the in vitro drug response of P. vivax: the initial stage of the parasite and the duration of the assay. Isolates of P. vivax initially at the trophozoite stage had significantly higher chloroquine IC50s (478 nM [95% CI, 316 to 722]) than those initially at the ring stage (84.7 nM [95% CI, 45.7 to 157]; P < 0.001). Synchronous isolates of P. vivax and P. falciparum which reached the target of 40% schizonts in the control wells within 30 h had significantly higher geometric mean chloroquine IC50s (435 nM [95% CI, 169 to 1,118] and 55.9 nM [95% CI, 48 to 64.9], respectively) than isolates that took more than 30 h (39.9 nM [14.6 to 110.4] and 36.9 nM [31.2 to 43.7]; P < 0.005). The results demonstrate the marked stage-specific activity of chloroquine with P. vivax and suggest that susceptibility to chloroquine may be associated with variable growth rates. These findings have important implications for the phenotypic and downstream genetic characterization of P. vivax.

Clinical and pharmacological determinants of the therapeutic response to dihydroartemisinin-piperaquine for drug-resistant malaria.

Dihydroartemisinin-piperaquine (DHP) is an important new treatment for drug-resistant malaria, although pharmacokinetic studies on the combination are limited. In Papua, Indonesia, we assessed determinants of the therapeutic efficacy of DHP for uncomplicated malaria. Plasma piperaquine concentrations were measured on day 7 and day 28, and the cumulative risk of parasitological failure at day 42 was calculated using survival analysis. Of the 598 patients in the evaluable population 342 had infections with Plasmodium falciparum, 83 with Plasmodium vivax, and 173 with a mixture of both species. The unadjusted cumulative risks of recurrence were 7.0% (95% confidence interval [CI]: 4.6 to 9.4%) for P. falciparum and 8.9% (95% CI: 6.0 to 12%) for P. vivax. After correcting for reinfections the risk of recrudescence with P. falciparum was 1.1% (95% CI: 0.1 to 2.1%). The major determinant of parasitological failure was the plasma piperaquine concentration. A concentration below 30 ng/ml on day 7 was observed in 38% (21/56) of children less than 15 years old and 22% (31/140) of adults (P = 0.04), even though the overall dose (mg per kg of body weight) in children was 9% higher than that in adults (P < 0.001). Patients with piperaquine levels below 30 ng/ml were more likely to have a recurrence with P. falciparum (hazard ratio [HR] = 6.6 [95% CI: 1.9 to 23]; P = 0.003) or P. vivax (HR = 9.0 [95% CI: 2.3 to 35]; P = 0.001). The plasma concentration of piperaquine on day 7 was the major determinant of the therapeutic response to DHP. Lower plasma piperaquine concentrations and higher failure rates in children suggest that dose revision may be warranted in this age group.

Two fixed-dose artemisinin combinations for drug-resistant falciparum and vivax malaria in Papua, Indonesia: an open-label randomised comparison.

BACKGROUND: The burden of Plasmodium vivax infections has been underappreciated, especially in southeast Asia where chloroquine resistant strains have emerged. Our aim was to compare the safety and efficacy of dihydroartemisinin-piperaquine with that of artemether-lumefantrine in patients with uncomplicated malaria caused by multidrug-resistant P falciparum and P vivax. METHODS: 774 patients in southern Papua, Indonesia, with slide-confirmed malaria were randomly assigned to receive either artemether-lumefantrine or dihydroartemisinin-piperaquine and followed up for at least 42 days. The primary endpoint was the overall cumulative risk of parasitological failure at day 42 with a modified intention-to-treat analysis. This trial is registered with ClinicalTrials.gov, trial number 00157833. FINDINGS: Of the 754 evaluable patients enrolled, 466 had infections with P falciparum, 175 with P vivax, and 113 with a mixture of both species. The overall risk of failure at day 42 was 43% (95% CI 38-48) for artemether-lumefantrine and 19% (14-23) for dihydroartemisinin-piperaquine (hazard ratio=3.0, 95% CI 2.2-4.1, p<0.0001). After correcting for reinfections, the risk of recrudescence of P falciparum was 4.4% (2.6-6.2) with no difference between regimens. Recurrence of vivax occurred in 38% (33-44) of patients given artemether-lumefantrine compared with 10% (6.9-14.0) given dihydroartemisinin-piperaquine (p<0.0001). At the end of the study, patients receiving dihydroartemisinin-piperaquine were 2.0 times (1.2-3.6) less likely to be anaemic and 6.6 times (2.8-16) less likely to carry vivax gametocytes than were those given artemether-lumefantrine. INTERPRETATION: Both dihydroartemisinin-piperaquine and artemether-lumefantrine were safe and effective for the treatment of multidrug-resistant uncomplicated malaria. However, dihydroartemisinin-piperaquine provided greater post-treatment prophylaxis than did artemether-lumefantrine, reducing P falciparum reinfections and P vivax recurrences, the clinical public-health importance of which should not be ignored.

Dihydroartemisinin-piperaquine versus artesunate-amodiaquine: superior efficacy and posttreatment prophylaxis against multidrug-resistant Plasmodium falciparum and Plasmodium vivax malaria.

BACKGROUND: Antimalarial drug resistance is now well established in both Plasmodium falciparum and Plasmodium vivax. In southern Papua, Indonesia, where both strains of plasmodia coexist, we have been conducting a series of studies to optimize treatment strategies. METHODS: We conducted a randomized trial that compared the efficacy and safety of dihydroartemisinin-piperaquine (DHP) with artesunate-amodiaquine (AAQ). The primary end point was the overall cumulative parasitological failure rate at day 42. RESULTS: Of the 334 patients in the evaluable patient population, 185 were infected with P. falciparum, 80 were infected with P. vivax, and 69 were infected with both species. The overall parasitological failure rate at day 42 was 45% (95% confidence interval [CI], 36%-53%) for AAQ and 13% (95% CI, 7.2%-19%) for DHP (hazard ratio [HR], 4.3; 95% CI, 2.5-7.2; P<.001). Rates of both recrudescence of P. falciparum infection and recurrence of P. vivax infection were significantly higher after receipt of AAQ than after receipt of DHP (HR, 3.4 [95% CI, 1.2-9.4] and 4.3 [95% CI, 2.2-8.2], respectively; P<.001). By the end of the study, AAQ recipients were 2.95-fold (95% CI, 1.2- to 4.9-fold) more likely to be anemic and 14.5-fold (95% CI, 3.4- to 61-fold) more likely to have carried P. vivax gametocytes. CONCLUSIONS: DHP was more effective and better tolerated than AAQ against multidrug-resistant P. falciparum and P. vivax infections. The prolonged therapeutic effect of piperaquine delayed the time to P. falciparum reinfection, decreased the rate of recurrence of P. vivax infection, and reduced the risk of P. vivax gametocyte carriage and anemia.

Therapeutic response of multidrug-resistant Plasmodium falciparum and P. vivax to chloroquine and sulfadoxine-pyrimethamine in southern Papua, Indonesia.

To determine the level of antimalarial drug resistance in southern Papua, Indonesia, we assessed the therapeutic efficacy of chloroquine plus sulfadoxine-pyrimethamine (CQ+SP) for Plasmodium falciparum infections as well as CQ monotherapy for P. vivax infections. Patients with P. falciparum failing therapy were re-treated with unsupervised quinine+/-doxycycline therapy and those with P. vivax with either unsupervised quinine+/-doxycycline or amodiaquine. In total, 143 patients were enrolled in the study (103 treated with CQ+SP and 40 with CQ). Early treatment failures occurred in four patients (4%) with P. falciparum and six patients (15%) with P. vivax. The failure rate by Day 28 for P. vivax was 65% (95% CI 49-81). After PCR correction for re-infections, the Day 42 recrudescence rate for P. falciparum infections was 48% (95% CI 31-65). Re-treatment with unsupervised quinine+/-doxycycline resulted in further recurrence of malaria in 48% (95% CI 31-65) of P. falciparum infections and 70% (95% CI 37-100) of P. vivax infections. Eleven patients with recurrent P. vivax were re-treated with amodiaquine; there were no early or late treatment failures. In southern Papua, a high prevalence of drug resistance of P. falciparum and P. vivax exists both to first- and second-line therapies. Preliminary data indicate that amodiaquine retains superior efficacy compared with CQ for CQ-resistant P. vivax.

A community-based TB drug susceptibility study in Mimika District, Papua Province, Indonesia.

SETTING: A district level tuberculosis (TB) control programme in Papua Province, Indonesia. OBJECTIVE: To determine the nature and extent of drug-resistant TB in newly diagnosed sputum smear-positive patients. METHODS: Sputum was collected from previously untreated smear-positive pulmonary TB patients diagnosed in the district over a 10-month period. Sputum specimens were processed and inoculated into a BACTEC MGIT960 tube. Isolates were identified by Ziehl-Neelsen staining, hybridisation with nucleic acid probes and biochemical investigations. Susceptibility testing was performed using the radiometric proportion method. Pyrazinamide testing was performed using the Wayne indirect method. RESULTS: One hundred and seven patients had sputum sent to a reference laboratory; 101 (94.4%) were culture-positive for Mycobacterium tuberculosis, with 87 (86.1%) fully sensitive to first-line anti-tuberculosis drugs. Two per cent were multidrug-resistant (MDR-TB) and 12 (11.9%) had other drug resistance. Each of the MDR-TB isolates was susceptible to amikacin, capreomycin, ciprofloxacin and para-aminosalicylic acid (PAS), but were resistant to rifabutin. One isolate was also resistant to ethionamide. CONCLUSIONS: MDR-TB is present in Indonesia but is not a major problem for TB control in this district. Generalisability to other districts in Indonesia, particularly large urban areas, needs to be confirmed by future studies.

An alternative method for sputum storage and transport for mycobacterium tuberculosis drug resistance surveys.

SETTING: A district level tuberculosis (TB) programme in Indonesia. OBJECTIVE: To evaluate whether a single sputum specimen could be stored by refrigeration for an extended period of time, then transported to a reference laboratory and successfully cultured for Mycobacterium tuberculosis. METHODS: Single sputum specimens were collected from newly diagnosed smear-positive pulmonary TB patients, refrigerated at the study site without addition of 1% cetylpyridinium chloride, batched and sent to the reference laboratory, where they were decontaminated and inoculated into BACTEC MGIT 960 liquid media. RESULTS: One hundred and seven patients were enrolled. The median specimen storage time was 12 days (range 1-38) and median transportation time was 4 days (2-12). The median time from specimen collection until processing was 18 days (4-42). Only 4 (3.7%) specimens failed to grow Mycobacterium species and M. tuberculosis was isolated from 101 (94.4%) specimens. Six specimens with breakthrough contamination successfully grew M. tuberculosis after a second decontamination procedure. CONCLUSIONS: Single sputum specimens collected at a remote setting, refrigerated for relatively long periods without preservatives and transported without refrigeration to a reference laboratory can yield a high positive culture rate. These findings offer potential logistic simplification and cost savings for drug resistance surveys in low-resource countries.

Glycosylphosphatidylinositols in malaria pathogenesis and immunity: potential for therapeutic inhibition and vaccination.

Glycosylphosphatidylinositols (GPIs) are found in the outer cell membranes of all eukaryotes. GPIs anchor a diverse range of proteins to the surface of Plasmodium falciparum, but may also exist free of protein attachment. In vitro and in vivo studies have established GPIs as likely candidate toxins in malaria, consistent with the prevailing paradigm that attributes induction of inflammatory cytokines, fever and other pathology to parasite toxins released when schizonts rupture. Although evolutionarily conserved, sufficient structural differences appear to exist that impart upon plasmodial GPIs the ability to activate second messengers in mammalian cells and elicit immune responses. In populations exposed to P. falciparum, the antibody response to purified GPIs is characterised by a predominance of immunoglobulin (Ig)G over IgM and an increase in the prevalence, level and persistence of responses with increasing age. It remains unclear, however, if these antibodies or other cellular responses to GPIs mediate anti-toxic immunity in humans; anti-toxic immunity may comprise either reduction in the severity of disease or maintenance of the malaria-tolerant state (i.e. persistent asymptomatic parasitaemia). P. falciparum GPIs are potentially amenable to specific therapeutic inhibition and vaccination; more needs to be known about their dual roles in malaria pathogenesis and protection for these strategies to succeed.

Seroprevalence of Burkholderia pseudomallei in East Timorese refugees: implications for healthcare in East Timor.

Melioidosis is a disease with protean clinical manifestations caused by the bacterium Burkholderia pseudomallei. It is endemic in countries surrounding the newly independent East Timor, but has yet to be isolated or demonstrated serologically in that country. One illness that can be clinically indistinguishable from melioidosis is pulmonary tuberculosis, a condition with a very high prevalence in East Timor. We used an indirect hemagglutination test (IHA) to measure antibodies to B. pseudomallei in 407 East Timorese evacuated to Darwin, Australia, in September 1999. Assuming a positive IHA titer as > or = 1:40, the overall seroprevalence rate was 17.0%, in keeping with other seroprevalence studies from the region. The IHA titres ranged up to 1:320. After adjusting for age, females were 2.5 times more likely to be seropositive than males (p = 0.0001). There was an inverse relationship between seropositivity and age. This study shows that exposure to B. pseudomallei occurs in East Timor melioidosis is also likely to occur. Due to the lack of laboratory facilities at present, it may be some time before a laboratory-confirmed case proves that melioidosis occurs. In the meantime, clinicians in East Timor should include melioidosis in the differential diagnosis of the many conditions that it may mimic.