Phylogeographic Evidence for 2 Genetically Distinct Zoonotic Plasmodium knowlesi Parasites, Malaysia.

Ruhani Yusof,Frankie Anak Thomas Sitam,Mun Yik Fong,Md Atique Ahmed,Sahlawati Mustakim,Hie Ung Ngian,J Rovie-Ryan Japning,Georges Snounou,Ananias A Escalante,Jenarun Jelip,Rohela Mahmud,Yee Ling Lau,Hani Mat Hussin

doi:10.3201/eid2208.151885

Abstract

Infections of humans with the zoonotic simian malaria parasite Plasmodium knowlesi occur throughout Southeast Asia, although most cases have occurred in Malaysia, where P. knowlesi is now the dominant malaria species. This apparently skewed distribution prompted an investigation of the phylogeography of this parasite in 2 geographically separated regions of Malaysia, Peninsular Malaysia and Malaysian Borneo. We investigated samples collected from humans and macaques in these regions. Haplotype network analyses of sequences from 2 P. knowlesi genes, type A small subunit ribosomal 18S RNA and cytochrome c oxidase subunit I, showed 2 genetically distinct divergent clusters, 1 from each of the 2 regions of Malaysia. We propose that these parasites represent 2 distinct P. knowlesi types that independently became zoonotic. These types would have evolved after the sea-level rise at the end of the last ice age, which separated Malaysian Borneo from Peninsular Malaysia.

Highlights

Infections of humans with the zoonotic simian malaria parasite Plasmodium knowlesi occur throughout Southeast Asia, most cases have occurred in Malaysia, where P. knowlesi is the dominant malaria species
The results of the various analyses conducted on the P. knowlesi parasites collected from Peninsular Malaysia and Malaysian Borneo strongly support the conclusion that the 2 geographically separated regions of this country harbor genetically distinct P. knowlesi populations
A measure of species evenness, was high for both the Peninsular Malaysia (0.906 ± 0. 0.025) and the Malaysian Borneo (0.871 ± 0.031) isolates, which may indicate a sustained transmission of P. knowlesi in both regions of Malaysia over long periods

Summary

Materials and Methods

Sample Collection The Medical Research Ethic Committee of the Ministry of Health Malaysia, Sabah State Director of Health, Kelantan State Director of Health, and directors of state and district hospitals approved this study. Ethical approval was granted by the Medical Research and Ethics Committee of the Malaysian Ministry of Health (Reference Number: KKM/NIHSEC/800/-2/2/2/P13–316), the Medical Ethics Committee of University Malaya Medical Centre, and the Department of Wildlife and National Parks. We used previously collected human blood samples for this study [7]: 78 microscopically confirmed P. knowlesi– positive blood samples from patients in 8 states in Malaysia, including Sabah and Sarawak in Malaysian Borneo (Figure 1). All samples were collected during September 2012–December 2013 (Tables 1, 2) In addition to these samples, we included previously published sequences deposited into GenBank during 2003–2015 in the analyses; these sequences were derived from samples collected from humans and macaques (online Technical Appendix Table, http://wwwnc.cdc.gov/EID/ article/22/8/15-1885-Techapp1.pdf)

Total sequences

Nucleotide diversity

Discussion

Findings

FST values