The plasticity of Plasmodium falciparum gametocytaemia in relation to age in Burkina Faso

Malaria Journal The plasticity of Plasmodium falciparum gametocytaemia in relation to age in Burkina Faso Andr Lin Oudraogo 0 2 Teun Bousema 1 Sake J de Vlas 3 Nadine Cuzin-Ouattara 0 Jan-Peter Verhave 2 Chris Drakeley 1 Adrian JF Luty 2 Robert Sauerwein 2 0 Centre National de Recherche et de Formation sur le Paludisme , BP 2208, Ouagadougou 01 , Burkina Faso 1 Department of Infectious & Tropical Diseases, London School of Hygiene & Tropical Medicine , London , UK 2 Department of Medical Microbiology, Radboud University Nijmegen Medical Centre , Nijmegen , The Netherlands 3 Department of Public Health, Erasmus MC, University Medical Center Rotterdam , P.O. Box 2040, 3000 CA Rotterdam , The Netherlands Background: Malaria transmission depends on the presence of gametocytes in the peripheral blood. In this study, the age-dependency of gametocytaemia was examined by microscopy and molecular tools. Methods: A total of 5,383 blood samples from individuals of all ages were collected over six cross sectional surveys in Burkina Faso. One cross-sectional study used quantitative nucleic acid sequence based amplification (QTNASBA) for parasite quantification (n = 412). The proportion of infections with concurrent gametocytaemia and median proportion of gametocytes among all parasites were calculated. Results: Asexual parasite prevalence and gametocyte prevalence decreased with age. Gametocytes made up 1.8% of the total parasite population detected by microscopy in the youngest age group. This proportion gradually increased to 18.2% in adults (p < 0.001). Similarly, gametocytes made up 0.2% of the total parasite population detected by QT-NASBA in the youngest age group, increasing to 5.7% in adults (p < 0.001). This age pattern in gametocytaemia was also evident in the proportion of gametocyte positive slides without concomitant asexual parasites which increased from 13.4% (17/127) in children to 45.6% (52/114) in adults (OR 1.55, 95% CI 1.38-1.74, p < 0.001). Conclusions: The findings of this study suggest that although gametocytes are most commonly detected in children, the proportion of asexual parasites that is committed to develop into gametocytes may increase with age. These findings underscore the importance of adults for the human infectious reservoir for malaria. - Background The malaria parasites life- cycle is composed of several developmental stages, one of which is the transmissible sexual stage comprising male and female gametocytes. Mature gametocytes are apparently benign, causing no overt disease symptoms. They appear to be developmentally arrested at the G0 phase of the cell cycle and circulate within erythrocytes in the peripheral blood of the human host until they are taken up by a feeding female mosquito. In the mosquito midgut, gametocyte activation and fertilization take place. The subsequent formation of sporogonic stages results in the development of thousands of sporozoites that migrate to and invade the salivary glands, rendering the mosquito infectious to humans. During the course of an infection with Plasmodium falciparum, gametocytes are generated from asexual stage parasites. Only a small fraction of the asexual parasites of P. falciparum commit to form gametocytes [1] and as a result only a fraction of infected individuals also harbour gametocytes [1,2]. It is now understood that this apparently low occurrence of gametocytes is partly a reflection of the low sensitivity of microscopy for the detection of gametocytes [2,3]. However, the fact remains that asexual parasitaemia is not always accompanied by gametocyte carriage [1-3], and that the relationship between asexual parasite density and gametocyte prevalence or density is not straightforward. Some studies report a positive association between asexual parasite densities and gametocyte prevalence [4-6] and density [4] while others observe inverse associations [7,8] or report that the association may be modified by age [2]. Factors that trigger and regulate the commitment of asexual stage parasites to gametocytes are largely unknown but are thought to include intrinsic parasite factors [9], anti-malarial treatment [4,6,10] and treatment outcome [4-6,11] , fever [7,8], haematological disruptions [6,12,13] and the presence of competing parasite strains [14,15] or species [6,16]. In general the mechanism of sexual commitment appears to be highly plastic and environment sensitive [17,18]. The flexible gametocyte production can be interpreted as a response mechanism of the parasite to stressful situations: if the survival of the asexual stage parasite is challenged, the investment in transmission stages increases. Here, we explore age-dependent variation in gametocytaemia in a series of cross-sectional surveys in an area of seasonal malaria transmission in Burkina Faso, using both microscopy and quantitative nucleic acid sequence based amplification (QT-NASBA). Methods Study site and population The study was carried out in the vicinity of Ouagadougou, the capital of Burkina Faso. The area has the ecological characteristics of Sudan savannah. Participating populations from six villages (longitude: 146- 179; latitude: 1252-1261) were of the same ethnic group (Mossi) and had similar age distributions. Transmission intensity is intense and seasonal in this region. Study subjects were given detailed explanations of the procedures, risk and benefits involved in the study and their consent was obtained. The study protocol was viewed and approved by the Ministry of Health of Burkina Faso (Researchs Authorization number 2000/3174/MS/SG/ DEP). Blood sample collection Cross-sectional surveys were performed in January, May, August and December 2002 and in April and December 2003. Participants were randomly selected from previously determined age groups (0.5-4, 5-9, 10-14, 15-24 and 25+ years) based on census lists and computer generated randomization tables. Thick and thin blood smears were made from finger-prick blood. The body temperature was measured and febrile individuals who were parasitaemic were treated with chloroquine according to the national policy in 2002. In the cross-sectional survey of December 2003, a single finger prick sample was used for blood smears and the collection of nucleic acids for quantitative-nucleic acid sequence based amplification (QT-NASBA); 100 L blood samples were collected from 412 volunteers of all ages from the six villages. The first part of the RNA extraction was done in the field following the original guanidinium isothiocyanate (GuSCN) RNA extraction method [19] until the nucleic acids were bound to silica dioxide particles. At this point, samples were stored at -20C and transferred Microscopical detection of P. falciparum parasites Samples were considered negative if no parasites were detected in 100 high-power fields of Giemsa-stained thick blood smears. Both asexual stage and gametocyte densities were assessed in the thick smear by counting against 500 and (...truncated)