RAPID ONSET OF TRANSMISSION-REDUCING ANTIBODIES IN JAVANESE MIGRANTS EXPOSED TO MALARIA IN PAPUA, INDONESIA

J. TEUN BOUSEMA 0 1 WILL ROEFFEN 0 1 MIKE VAN DER KOLK 0 1 SAKE J. DE VLAS 0 1 MARGA VAN DE VEGTE-BOLMER 0 1 MICHAEL J. BANGS 0 1 KARINA TEELEN 0 1 LILIANA KURNIAWAN 0 1 JASON D. MAGUIRE 0 1 J. KEVIN BAIRD 0 1 ROBERT W. SAUERWEIN 0 1 0 Medical Microbiology 268, Radboud University Nijmegen Medical Centre , P.O. Box 9101, 6500 HB Nijmegen , The Netherlands 1 Department of Medical Microbiology, Radboud University, Nijmegen Medical Centre , Nijmegen , The Netherlands ; Department of Public Health, Erasmus Medical Centre, University Medical Center Rotterdam, The Netherlands; U.S. Naval Medical Research Unit No. 2, Jakarta , Indonesia; National Institute of Health Research and Development , Jakarta , Indonesia Transmission of Plasmodium falciparum malaria is initiated by sexual stages in the mosquito. Anti-Pfs48/ 45 and anti-Pfs230 sexual stage antibodies that are ingested together with parasites can reduce parasite development and subsequently malaria transmission. Acquisition of sexual stage immunity was studied in a cohort of 102 non-immune Javanese individuals migrating to hyperendemic Papua Indonesia. Seroprevalence of antibodies against Pfs48/45 and Pfs230 and functional transmission-reducing activity (TRA) were measured upon arrival and at 6, 12, and 24 months. Asexual parasitemia and gametocytemia were assessed every two weeks. The TRA and seroreactivity increased with the number of P. falciparum infections. The longitudinally sustained association between TRA and antibodies against Pfs48/45 (odds ratio [OR] 3.74, 95% confidence interval [CI] 1.51-9.29) and Pfs230 (OR 3.72, 95% CI 1.36-10.17) suggests that functional transmission reducing immunity is acquired after limited exposure to infection. - Successful transmission of Plasmodium falciparum parasites from humans to mosquitoes depends on the presence of infectious gametocytes in the circulation, which are derived from a small fraction of asexual parasites. After ingestion of the blood meal by mosquitoes, fertilization takes place in the midgut, which will ultimately result in the generation of infectious sporozoites in the salivary glands. Specific antibodies against sexual stages can interfere with fertilization and subsequent sporogonic development when co-ingested with the blood meal, as shown in natural and experimental infections.14 This transmission-reducing activity (TRA) is associated with antibodies directed against the sexual stage-specific antigens Pfs48/45 and Pfs230.58 The rate of development and persistence of TRA is largely unknown. Although it was suggested to be low grade and rapidly transient,9 persistent TRA has also been observed.10 Acquisition of immunity directed against asexual blood stage parasites has been intensively studied people migrating from a non-endemic to a malariaendemic area.1115 In these transmigrants, clinical and parasitologic immunity developed quickly after exposure13 with inconsistent evidence for an independent role of age.1116 The objective of this study was to examine the development of sexual stage-specific immunity in a longitudinal cohort of Javanese children and adults from non-endemic Java after migration to malaria endemic Papua. Antibody reactivity against sexual stage-specific antigens Pfs230 and Pfs48/45 was determined and compared with antibodies against asexual stage antigen glutamate-rich protein (GLURP) and whole parasite extracts. The GLURP-specific antibodies have previously been associated with protection against high levels of parasitemia17 and clinical disease.1820 The development of sexual stage-specific immunity was evaluated by detecting circulating antibodies against Pfs230 and Pfs48/45 and by assessing functional TRA in the standard membrane-feeding assay. MATERIALS AND METHODS Study population. The study site was a newly created transmigration village (designated SP2) located near the northeastern coast of Papua, Indonesia. After informed consent was obtained, healthy volunteer adults between 20 and 40 years of age and children between 6 and 12 years of age were included in the study. The study was reviewed and approved by the institutional ethical review boards of the United States Navy and the Indonesian Ministry of Health under U.S. Department of Defense Protocol #30820. The included human subjects were treated according to relevant regulations of the Indonesian Ministry of Health and the United States government (code 32 of Federal Regulation, Part 219, Protection of Human Subjects; U.S. Navy, SECNAVINST 3900.39B). Subjects were excluded if admitting a history of residence in a malaria-endemic area in the previous five years. Details on recruitment, follow-up, and the epidemiology of malaria in this cohort have been described elsewhere.16 Enrollment started in 1996 and follow-up ended in 1999. The subjects originated from Java and were enrolled in the study within 48 hours of arrival in Papua, Indonesia. Malaria transmission in Java has been very low for many decades, typically around 0.01 malaria cases/1,000 person-years after the exclusion of the few foci of hypoendemic transmission.21 In contrast, malaria transmission in Papua is perennial and often hyperendemic to holoendemic, with incidence rates from 500 to 5,000 infections/1,000 person-years.22 Follow-up. Malaria episodes were actively detected over a period of 24 months by periodic clinical assessments and blood smears. Blood smears were collected at two-week intervals or at any time a subject presented to the clinic with symptoms (chills, fever, etc.) suggesting malaria infection. Blood was microscopically examined for P. falciparum and P. vivax asexual parasites and gametocytes by Giemsa-stained blood films. Parasites were counted against the number of white blood cells. A slide was considered negative after examination of 200 microscopic fields, corresponding to 1,600 2,400 white blood cells and an estimated diagnostic threshold of five parasites per microliter. Parasite densities were calculated using a normal density of 8,000 white blood cells per microliter of blood. The first episode of symptomatic or asymptomatic P. falciparum malaria was recorded as first infection. Each subsequent P. falciparum parasitemia was classified as a new infection if it occurred more than 28 days after directly observed chemotherapy of the previous infection. Sera were collected on the day of arrival and after 6, 12, and 24 months at SP2.12,14,16 To study the acquisition of immunity in time, we selected samples from subjects for whom at least three samples were available. We selected 51 subjects 612 years of age and 51 subjects 2040 years of age. The incidence of malaria infection in these individuals was not different from that of the general population. Enzyme-linked imunosorbent assays (ELISAs). Antigen preparation. Mature gametocytes of P. falciparum (NF54 strain) were produced in an automated static culture system as described by Ponnudurai and others,23 isolated,4 and stored at 7 (...truncated)