The Pk41 surface protein diversity, natural selection, sub population and geographical clustering: a 6-cysteine protein family member.

INTRODUCTION

The zoonotic malaria parasite has currently become the most dominant form of infection in humans in Malaysia and is an emerging infectious disease in most Southeast Asian countries. The P41 is a merozoite surface protein belonging to the 6-cysteine family and is a well-characterized vaccine candidate in and ; however, no study has been done in the orthologous gene of . This study investigates the level of polymorphism, haplotypes and natural selection of genes in clinical isolates from Malaysia.

METHOD

Thirty-five full-length sequences from clinical isolates of Malaysia along with four laboratory lines (along with H-strain) were downloaded from public databases. For comparative analysis between species, orthologous genes from , and were also downloaded. Genetic diversity, polymorphism, haplotype and natural selection were determined using DnaSP 5.10 software. Phylogenetic relationships between genes were determined using MEGA 5.0 software.

RESULTS

Analysis of 39 full-length sequences along with the H-strain identified 36 SNPs (20 non-synonymous and 16 synonymous substitutions) resulting in 31 haplotypes. Nucleotide diversity across the full-length gene was low and was similar to its ortholog in ; . Domain-wise amino acid analysis of the two s48/45 domains indicated low level of polymorphisms for both the domains, and the glutamic acid rich region had extensive size variations. In the central domain, upstream to the glutamate rich region, a unique two to six (K-E) repeat region was identified within the clinical isolates. Overall, the genes were indicative of negative/purifying selection due to functional constraints. Domain-wise analysis of the s48/45 domains also indicated purifying selection. However, analysis of Tajima's D across the genes identified non-synonymous SNPs in the s48/45 domain II with high positive values indicating possible epitope binding regions. All the 6-cysteine residues within the s48/45 domains were conserved within the clinical isolates indicating functional conservation of these regions. Phylogenetic analysis of full-length genes indicated geographical clustering and identified three subpopulations of ; one originating in the laboratory lines and two originating from Sarawak, Malaysian Borneo.

CONCLUSION

This is the first study to report on the polymorphism and natural selection of genes from clinical isolates of Malaysia. The results reveal that there is low level of polymorphism in both s48/45 domains, indicating that this antigen could be a potential vaccine target. However, genetic and molecular immunology studies involving higher number of samples from various parts of Malaysia would be necessary to validate this antigen's candidacy as a vaccine target for .