Short Report: Prevalence and Chloroquine Sensitivity of Plasmodium malariae in Madagascar

Abstract. We report the results of clinical studies carried out at six sites in Madagascar, between January and October 2006. The aims were (i) to update our knowledge of the burden of Plasmodium malariae infection and (ii) to assess the therapeutic efficacy of chloroquine for uncomplicated quartan malaria. Our findings confirm that P. malariae is the third leading cause of malaria, accounting for 1.1% of all malarial infections. They also demonstrate that chloroquine-currently recommended for the home management of presumed malaria in children under the age of five years and commonly used by adults-remains highly effective in patients with uncomplicated P. malariae infection. Plasmodium malariae, one of the four species of Plasmodium affecting humans, is found in tropical and subtropical regions, often in sympatry with other Plasmodium species, as in Madagascar. Its reported prevalence varies from less than 4% to more than 20% in endemic regions. 9 Fever displays quartan (4-day) periodicity, parasite density is usually considerably below 1000 parasites per ml of blood, and infection is rarely life-threatening in the absence of complications, such as nephrotic syndrome. 17 Recent WHO guidelines for the testing of therapeutic efficacy have not yet been applied to P. malariae infection. We therefore report here the results of a clinical study carried out at six sites in 2006. The aims of this study were to: (i) update our knowledge of the burden of Plasmodium malariae infections and (ii) assess the therapeutic efficacy of CQ for uncomplicated quartan malaria. This clinical study, approved by the National Ethics Committee of the Ministry of Health and Family Planning of Madagascar (N°102-SANPF-2006), was conducted in 2006, at six primary health centers and assessed the therapeutic efficacy of the main antimalarial drugs used in Madagascar. All patients with fever or a history of fever in the 48 h before their arrival at the health center were screened with the rapid diagnostic test (RDT) based on the detection of Plasmodiumspecific lactate dehydrogenase (pLDH) (OptiMAL-IT™, DiaMed AG © , Cressier sur Morat, Switzerland). Giemsastained thin and thick blood films were prepared for each patient with a positive RDT result. A microbiologist differentiated between the different Plasmodium species and parasitemia was assessed. Patients with histologically confirmed P. malariae infection were enrolled according to inclusion criteria adapted from the WHO guidelines (2001) 18 for P. vivax therapeutic efficacy tests. Informed consent was obtained from all adults and from at least one parent for minors. Patients were then treated with the standard chloroquine regimen of 25 mg/kg over 3 days and followed for 28 days. Clinical examination, including axillary temperature recording and the screening of thick and thin films, was carried out on days 0, 1, 2, 3, 7, 14, 21, and 28. On these days, capillary blood was spotted onto filter paper for PCR confirmation. Hemoglobin concentration was measured on days 0 and 28. DNA was extracted from blood spots with Instagene® Matrix resin (BioRad©, Marnes la Coquette, France), according to the manufacturer's instructions. The parasite species was confirmed by real-time PCR, using species-specific primers as described by de Monbrison, From January to October 2006, 6,692 patients were screened: 66.8% tested negative for malaria. Of the patients whose test results indicated the presence of malaria, 91.9% tested positive for P. falciparum and 8.1% for other Plasmodium species. Microscopy results confirmed by species-specific real-time PCR showed that 86.9% of the patients infected with nonfalciparum species were infected with P. vivax and 13.1% were infected with P. malariae. None of the patients tested positive for P. ovale. Plasmodium malariae infection accounted for 0% (Ihosy) to 2.0% (Maevatanana) of all malaria cases, depending on the site considered 1039 were six male and 10 female patients (62.5%), aged from 1 to 25 years (median 4.5), with a median weight of 10.5 kg (range 5-63). All patients had had fever during the past 48 hours and none declared having taken antimalarial drugs. Neither microscopy nor real-time PCR showed mixed infections (P. malariae with other species). The geometric mean of asexual parasite count was 947/L (range 250-14,000) at baseline. None of the patients had detectable gametocytes on microscopy at day 0 or during the time of follow-up. Mean hemoglobin concentration was 9.1 g/dL on day 0, and the mean increase in hemoglobin concentration observed on day 28 (mean of individual increases in Hb) was 1.0 g/dL. Fifteen patients successfully cleared P. malariae parasitemia after CQ treatment. One case was excluded on day 28 because P. falciparum parasites were detected on a blood smear (confirmed by real-time PCR). Microscopy showed parasite clearance before day 7 in all patients. However, realtime PCR showed parasites to be present in the blood, as described for diagnosis in one patient, and this patient displayed clearance between days 7 and 14 Our data confirm that P. malariae is the third leading cause of malaria in Madagascar after P. falciparum and P. vivax. No P. ovale parasites were observed due to the very low prevalence of this specie in Madagascar. 1040 Eastern Madagascar. CQ remains the drug most widely available (distribution and financial criteria) and is the first drug used in most of areas in Madagascar, particularly in the prepackaged PaluStop form, sold at an affordable price (US $0.025), or as Ody Tazomoka, distributed free at primary public health facilities. In conclusion, our findings demonstrate that oral CQ, which is commonly used, remains highly effective in patients with uncomplicated P. malariae infection, the third leading cause of malaria in Madagascar. Received April 30, 2007. Accepted for publication July 2, 2007. Acknowledgments: We thank the patients and healthcare workers involved in the national network for the surveillance of malaria resistance in Madagascar (Réseau d'Etude de la Résistance, RER) from which these samples were derived, and the staff of the Ministry of Health of Madagascar for their collaboration