Age-dependent carriage of alleles and haplotypes of Plasmodium falciparum sera5, eba-175, and csp in a region of intense malaria transmission in Uganda

Abstract

Background: The development of malaria vaccines is constrained by genetic polymorphisms exhibited by Plasmodium falciparum antigens. We investigated the age-dependent distribution of alleles or haplotypes of three P. falciparum malaria vaccine candidates, circumsporozoite protein (csp), erythrocyte binding antigen 175 (eba-175) and serine repeat antigen 5 (sera5) in a region of intense malaria transmission in Uganda. Methods & Materials: A cross sectional study was carried out between August and November 2009. Blood samples were collected after informed consent from 250 individuals below 5 years, 5-10 years and above 10 years olds. P. falciparum DNA was extracted from all samples. Alleles of sera5 and eba-175 were determined by polymerase chain reaction (PCR) amplification followed by resolution of PCR products by agarose gel electrophoresis and allele calling using photographs of ethidium bromide-stained gels. Haplotypes of CSP were identified by sequencing 63 PCR products and using P.falciparum 7G8 strain sequence as a reference. Results: Both eba-175 FCR3 (48/178) and CAMP (16/178) alleles were observed with the FCR3 (24/67) allele being predominant among children aged below 5 years old while the CAMP (12/67) allele was predominant among older individuals. Both sera5 alleles ORI (6/204) and ORII (103/204) were observed in the population but ORII was more prevalent. SERA5 ORII allele was significantly associated with age (P values < 0.0001), parasite density (P value < 0.0001) and clinical outcomes (P value = 0.018). There was marked CSP diversity in the Th2/Th3 region. Out of 63 sequences, 16 conformed to the reference strain and one (1/16) was similar with a West African haplotype and the majority (14/16) of the haplotypes were unique to this study region. Conclusion: There was an age-dependent distribution of CSP haplotypes with more haplotypes being harbored by < 5-year olds, (10/16) compared to adults (2/16). Interestingly, the CSP haplotype corresponding to 3D7 whose prototypical sequence is identical to the sequence of the leading malaria vaccine candidate RTS, S was not observed. Our data suggest that eba-175 FCR3 allele, sera5 ORII allele, and CSP haplotypes are targets of host immunity and under immune selection pressure in Apac District.