Publications

Background The efficacy of anti-malarial drugs is determined by the level of parasite susceptibility, anti-malarial drug bioavailability and pharmacokinetics, and host factors including immunity. Host immunity improves the in vivo therapeutic efficacy of anti-malarial drugs, but the mechanism and magnitude of this effect has not been characterized. This study characterized the effects of 'immune' plasma to Plasmodium falciparum on the in vitro susceptibility of P. falciparum to anti-malarial drugs. Methods Titres of antibodies against blood stage antigens (mainly the ring-infected erythrocyte surface antigen [RESA]) were measured in plasma samples obtained from Thai patients with acute falciparum malaria. 'Immune' plasma was selected and its effects on in vitro parasite growth and multiplication of the Thai P. falciparum laboratory strain TM267 were assessed by light microscopy. The in vitro susceptibility to quinine and artesunate was then determined in the presence and absence of 'immune' plasma using the 3 H-hypoxanthine uptake inhibition method. Drug susceptibility was expressed as the concentrations causing 50% and 90% inhibition (IC 50 and IC 90 ), of 3 H-hypoxanthine uptake. Results Incubation with 'immune' plasma reduced parasite maturation and decreased parasite multiplication in a dose dependent manner. 3 H-hypoxanthine incorporation after incubation with 'immune' plasma was decreased significantly compared to controls (median [range]; 181.5 [0 to 3,269] cpm versus 1,222.5 [388 to 5,932] cpm) ( p = 0.001). As a result 'immune' plasma reduced apparent susceptibility to quinine substantially; median (range) IC 50 6.4 (0.5 to 23.8) ng/ml versus 221.5 (174.4 to 250.4) ng/ml ( p = 0.02), and also had a borderline effect on artesunate susceptibility; IC 50 0.2 (0.02 to 0.3) ng/ml versus 0.8 (0.2 to 2.3) ng/ml ( p = 0.08). Effects were greatest at low concentrations, changing the shape of the concentration-effect relationship. IC 90 values were not significantly affected; median (range) IC 90 448.0 (65 to > 500) ng/ml versus 368.8 (261 to 501) ng/ml for quinine ( p > 0.05) and 17.0 (0.1 to 29.5) ng/ml versus 7.6 (2.3 to 19.5) ng/ml for artesunate ( p = 0.4). Conclusions 'Immune' plasma containing anti-malarial antibodies inhibits parasite development and multiplication and increases apparent in vitro anti-malarial drug susceptibility of P. falciparum . The IC 90 was much less affected than the IC 50 measurement.

Noninfected erythrocytes form rosettes around those infected with trophozoites and schizonts of Plasmodium falciparum in vitro. These rosettes are thought to contribute to the microvascular obstruction which underlies the pathophysiology of severe falciparum malaria. To determine whether the percentage of infected erythrocytes forming rosettes for a parasite isolates in vitro correlates with the in vivo severity of disease, we studied the rosette formation behavior of 35 isolates of P. falciparum from patients with uncomplicated, severe, and cerebral malaria. There was a wide variation in the degree of rosette formation (0 to 53%). Four parasite isolates formed rosettes well (30 to 53%), and seven isolates formed rosettes poorly or not at all (0 to 5%), while the majority of the isolates formed rosettes to various degrees between these two extremes. In this relatively small sample of patients, we were unable to demonstrate a significant association between in vitro rosette formation and patients with cerebral malaria or conscious patients with significant renal (serum creatinine greater than 200 mumol/liter) or hepatic dysfunction (serum bilirubin greater than 50 mumol/liter and aspartate aminotransferase greater than 50 Reitman-Frankel units). However, there was an inverse relationship between rosette formation and cytoadherence (r = -0.575, P less than 0.01) which could not be explained on the basis of steric hindrance. This finding suggests that cytoadherence and rosette formation properties are intrinsic to the parasites, with isolates having a greater propensity for one or the other but not both. Further studies are required to establish the occurrence and pathophysiological role of rosette formation in vivo.

No abstract is provided for this article.

Imwong, M., Pukrittakayamee, S., Looareesuwan, S., Poirriez, J., Pasvol, G., White, N. J., and Snounou, G. 2001. Plasmodium vivax: Polymerase chain reaction amplification artifacts limit the suitability of pvgam1 as a genetic marker. Experimental Parasitology 99, 175–179.

Ethnic Conflict and Democratization in Africa edited by Harvey Glickman (Atlanta, GA: The African Studies Association Press, 1995, pp.484, $29). Citizen and Subject. Contemporary Africa and the Legacy of Late Colonialism by Mahmood Mamdani. (Kampala/Cape Town/London: Fountain Publishers/David Philip/James Currey, pp.xii + 353, £14.95). Big Men, Small Boys and Politics in Ghana. Power, Ideology and the Burden of History, 1982–94 by Paul Nugent (London: Pinter, pp.xiv + 306, £55). Tributors, Supports and Merchant Capital: Mining and Underdevelopment in Sierra Leone by Alfred Zack‐Williams (Aldershot, Avebury, 1995, pp.239, £42.50). Limits of Adjustments in Africa edited by Poul Engberg‐Pedersen, Peter Gibbon, Phil Raikes and Lars Udsholt (Oxford: Centre for Development Research, Copenhagen, in association with James Currey, 1996, pp.xi + 430, £45 hb, £16.95 pb). Structurally Adjusted Africa: Poverty, Debt and Basic Needs edited by David Simon, Wim Van Spengen, Chris Dixon and Anders Narman (London: Pluto Press, 1995, pp.ix + 245, £45). The Tanzanian Peasantry: Further Studies edited by Peter Foster and Sam Maghimbi (Aldershot: Avebury, 1995, pp.xxxiv + 329, £42.50). Soldiers in Zimbabwe's Liberation War edited by Ngwabi Bhebe and Terence Ranger (Heinemann, New Hampshire, James Currey, London, and University of Zimbabwe Publications, 1995, Volume 1, pp.203, £12.95 pb). Apartheid's Last Stand: The Rise and Fall of the South African Security State by Chris Alden (London: Macmillan, 1996, pp.333, £45). South Africa's Defence and Security into the 21st Century edited by William Gutteridge (Aldershot, Dartmouth Publishing Company, 1996, pp.185, £37.50). In Search of Cool Ground: War, Flight and Homecoming in Northeast Africa edited by Tim Allen (Geneva: with London: James Currey and Trenton, NJ: Africa World Press, United Nations Research Institute for Social Development (UNRISD), pp.xv + 336, £14.95 pb, £35 hb). Africa in the Contemporary International Disorder: Crises and Possibilities edited by Mulugeta Agonafer (Lanham, MD: University Press of America, 1996, pp.xvi + 405, $58). Kampala Women Getting By: Wellbeing in the Time of AIDS by Sandra Wallman (London: James Currey, 1996, $35 hb, $14.95 pb). Winning Hearts and Minds: British Governments, the Media and Colonial Counter‐Insurgency 1994–1960 by Susan L. Carruthers (London and New York: Leicester University Press, 1995, pp.xi + 307, £45 hb, £6,99 pb). Challenging the State: Crisis and Innovation in Latin America and Africa by Merilee S. Grindle (Cambridge: Cambridge University Press, 1996, pp.243, £14.95). Democracy Development, and the Countryside: Urban‐Rural Struggles in India by Ashutosh Varshney (Cambridge: Cambridge University Press, 1995, pp.214. £35). The Hindu Nationalist Movement and Indian Politics, 1925 to the 1990s. Strategies of Identity‐Building, Implantation and Mobilisation (with special reference to Central india) by Christophe Jaffrelot (London: Hurst & Co., 1996, pp.xxiii + 592, £45 hb, £19.50 pb). Political Oppositions in Industrialising Asia edited by Garry Rodan (London and New York; Routledge, 1996, pp.xvii + 338, £45 hb; £14.99 pb). The 1992 PNG Election. Change anal Continuity in Electoral Politics edited by Yaw Saffu (Canberra: Australian National University; Political and Social Change Monograph 23, 1996, pp.xii + 409, pb). Ethnic Conflict and Development The Case of Fiji by Ralph R. Premdas (Aldershot: Avebury, pp.xi + 198, £45 hb). The Reality of Aid: An Independent Review of International Aid by Judith Randel and Tony German (London: Earthscan, 1996, pp.iii + 244, £14.95). Gender in Third World Politics by Georgina Waylen (Boulder, Lynne Rienner Publishers, 1996, pp.xii + 163, £18.95 pb, £45 hb).

A fixed artesunate-mefloquine combination, comprising three daily doses of 8 mg of mefloquine/kg of body weight and 4 mg of artesunate/kg, has been developed recently. This study was designed to construct a population pharmacokinetic model describing this new dosage regimen of mefloquine given as loose tablets together with artesunate. In two randomized trials in Thailand which evaluated the efficacy, safety, and tolerability of this new regimen, the members of a subgroup of 50 patients were randomized to have capillary blood sampling before treatment and at five randomly assigned time points during the 63-day follow-up period. Mefloquine levels in capillary whole blood were assayed by liquid chromatography with UV detection. A pharmacokinetic model for mefloquine was constructed using mixed-effects modeling. A one-compartment model with first-order absorption and elimination was selected to describe the kinetic properties of mefloquine. For capillary whole-blood mefloquine, the area under the concentration curve (AUC) was 40% higher than previous estimates for patients given the equivalent conventional-dose regimen (mefloquine given as 15 mg/kg and then 10 mg/kg on the second and third days of treatment). The half-life (t1/2) of the carboxylic acid metabolite was estimated as 26 days, and the metabolite was eliminated more slowly than the parent drug (population t1/2 estimate, 10.5 days). Splitting the 25 mg/kg dose of mefloquine into three doses of 8 mg/kg each resulted in improved oral bioavailability compared to the conventional split-dose regimen results. This new regimen is well tolerated and results in an equivalent therapeutic response.

Background Prophylaxis for high-risk populations, such as forest workers, could be one component for malaria elimination in the Greater Mekong Sub-region. A study was conducted to assess the malaria incidence in forest rangers and the feasibility of malaria prophylaxis for rangers sleeping in forest camps. Methods Forest rangers deployed in the Bu Gia Map National Park, Vietnam were invited to participate in the study. Plasmodium infections were cleared using presumptive treatment, irrespective of malaria status, with a 3-day course dihydroartemisinin/piperaquine (DP) and a 14-day course of primaquine. Before returning to the forest, study participants were randomly allocated to a 3-day course of DP or placebo. Fifteen days after returning from their forest deployment the participants were tested for Plasmodium infections using uPCR. Results Prior to treatment, 30 of 150 study participants (20%) were found to be infected with Plasmodium . Seventeen days (median) after enrolment the rangers were randomized to DP or placebo 2 days before returning to forest camps where they stayed between 2 and 20 days (median 9.5 days). One ranger in the DP-prophylaxis arm and one in the placebo arm were found to be infected with Plasmodium falciparum 15 days (median) after returning from the forest. The evaluable P. falciparum isolates had molecular markers indicating resistance to artemisinins (K13-C580Y) and piperaquine (plasmepsin), but none had multiple copies of pfmdr1 associated with mefloquine resistance. Conclusion Anti-malarial prophylaxis in forest rangers is feasible. The findings of the study highlight the threat of multidrug-resistant malaria. Trial registration NCT02788864

Falsified medicines are a major threat to global health. Antimalarial drugs have been particularly targeted by criminals. As DNA analysis has revolutionized forensic criminology, we hypothesized that these techniques could also be used to investigate the origins of falsified medicines. Medicines may contain diverse adventitious biological contamination, and the sealed nature of blister-packages may capture and preserve genetic signals from the manufacturing processes allowing identification of production source(s). We conducted a blinded pilot study to determine if such environmental DNA (eDNA) could be detected in eleven samples of falsified and genuine artesunate antimalarial tablets, collected in SE Asia, which could be indicative of origin. Massively Parallel Sequencing (MPS) was used to characterize microbial and eukaryote diversity. Two mitochondrial DNA analysis approaches were explored to detect the presence of human DNA. Trace eDNA from these low biomass samples demonstrated sample specific signals using two target markers. Significant differences in bacterial and eukaryote DNA community structures were observed between genuine and falsified tablets and between different packaging types of falsified artesunate. Human DNA, which was indicative of likely east Asian ancestry, was found in falsified tablets. This pilot study of the ‘pharmabiome’ shows the potential of environmental DNA as a powerful forensic tool to assist with the identification of the environments, and hence location and timing, of the source and manufacture of falsified medicines, establish links between seizures and complement existing tools to build a more complete picture of criminal trade routes. The finding of human DNA in tablets raises important ethical issues that need to be addressed.

Background Cambodia has seen a marked reduction in the incidence of Plasmodium falciparum over the past decade without a corresponding decline in Plasmodium vivax incidence. It is unknown to what extent local transmission is sustained by a chain of clinical and sub-clinical infections or by continued re-introduction via migration. Using an ultrasensitive molecular technique, 20 villages in western Cambodia were surveyed to detect the low season prevalence of P. falciparum and P. vivax and local treatment records were reviewed. Methods During March to May 2015 cross-sectional surveys were conducted in 20 villages in Battambang, western Cambodia. Demographic and epidemiological data and venous blood samples were collected from 50 randomly selected adult volunteers in each village. Blood was tested for Plasmodium infections by rapid diagnostic test (RDT), microscopy and high volume (0.5 ml packed red blood cell) quantitative polymerase chain reaction (uPCR). Positive samples were analysed by nested PCR to determine the Plasmodium species. Malaria case records were collected from the Provincial Health Department and village malaria workers to determine incidence and migration status. Results Among the 1000 participants, 91 (9.1%) were positive for any Plasmodium infection by uPCR, seven (0.7%) by microscopy, and two (0.2%) by RDT. uPCR P. vivax prevalence was 6.6%, P. falciparum 0.7%, and undetermined Plasmodium species 1.8%. Being male (adjusted OR 2.0; 95% CI 1.2-3.4); being a young adult <30 years (aOR 2.1; 95% CI 1.3–3.4); recent forest travel (aOR 2.8; 95% CI 1.6–4.8); and, a history of malaria (aOR 5.2; 95% CI 2.5–10.7) were independent risk factors for parasitaemia. Of the clinical malaria cases diagnosed by village malaria workers, 43.9% (297/634) and 38.4% (201/523) were among migrants in 2013 and in 2014, respectively. Plasmodium vivax prevalence determined by uPCR significantly correlated with vivax malaria incidences in both 2014 and 2015 (p = 0.001 and 0.002, respectively), whereas no relationship was observed in falciparum malaria (p = 0.36 and p = 0.59, respectively). Discussion There was heterogeneity in the malaria parasite reservoir between villages, and Plasmodium prevalence correlated with subsequent malaria incidence. The association was attributable chiefly to P. vivax infections, which were nine-fold more prevalent than P. falciparum infections. In the absence of a radical cure with 8-aminoquinolines, P. vivax transmission will continue even as P. falciparum prevalence declines. Migration was associated with over a third of incident cases of clinical malaria. Trial registration clinicaltrials.gov (NCT01872702). Registered 4 June 2013

Amodiaquine is a 4-aminoquinoline used extensively for the treatment and prevention of malaria. Orally administered amodiaquine is largely converted to the active metabolite desethylamodiaquine. Amodiaquine can cause bradycardia, hypotension, and electrocardiograph QT interval prolongation, but the relationship of these changes to drug concentrations is not well characterized.

Pyronaridine-artesunate is a newly introduced artemisinin-based combination treatment which may be deployed together with primaquine. A single-dose, randomized, three-sequence crossover study was conducted in healthy Thai volunteers to characterize potential pharmacokinetic interactions between these drugs. Seventeen healthy adults received a single oral dose of primaquine alone (30 mg base) and were then randomized to receive pyronaridine-artesunate alone (540-180 mg) or pyronaridine-artesunate plus primaquine in combination, with intervening washout periods between all treatments. The pharmacokinetic properties of primaquine, its metabolite carboxyprimaquine, artesunate, its metabolite dihydroartemisinin, and pyronaridine were assessed in 15 subjects using a noncompartmental approach followed by a bioequivalence evaluation. All drugs were well tolerated. The single oral dose of primaquine did not result in any clinically relevant pharmacokinetic alterations to pyronaridine, artesunate, or dihydroartemisinin exposures. There were significantly higher primaquine maximum plasma drug concentrations (geometric mean ratio, 30%; 90% confidence interval [CI], 17% to 46%) and total exposures (15%; 6.4% to 24%) during coadministration with pyronaridine-artesunate than when primaquine was given alone. Pyronaridine, like chloroquine and piperaquine, increases plasma primaquine concentrations. (This study has been registered at ClinicalTrials.gov under registration no. NCT01552330.).