Genetic diversity of circumsporozoite protein in Plasmodium knowlesi isolates from Malaysian Borneo and Peninsular Malaysia

Abstract

Background: Understanding the genetic diversity of candidate genes for malaria vaccines such as circumsporozoite protein (csp) may enhance the development of vaccines for treating Plasmodium knowlesi. Hence, the aim of this study is to investigate the genetic diversity of non-repeat regions of csp in P. knowlesi from Malaysian Borneo and Peninsular Malaysia. Methods: A total of 46 csp genes were subjected to polymerase chain reaction amplification. The genes were obtained from P. knowlesi isolates collected from different divisions of Sabah, Malaysian Borneo, and Peninsular Malaysia. The targeted gene fragments were cloned into a commercial vector and sequenced, and a phylogenetic tree was constructed while incorporating 168 csp sequences retrieved from the GenBank database. The genetic diversity and natural evolution of the csp sequences were analysed using MEGA6 and DnaSP ver. 5.10.01. A genealogical network of the csp haplotypes was generated using NETWORK ver. 4.6.1.3. Results: The phylogenetic analysis revealed indistinguishable clusters of P. knowlesi isolates across different geographic regions, including Malaysian Borneo and Peninsular Malaysia. Nucleotide analysis showed that the csp non-repeat regions of zoonotic P. knowlesi isolates obtained in this study underwent purifying selection with population expansion, which was supported by extensive haplotype sharing observed between humans and macaques. Novel variations were observed in the C-terminal non-repeat region of csp. Conclusions: The csp non-repeat regions are relatively conserved and there is no distinct cluster of P. knowlesi isolates from Malaysian Borneo and Peninsular Malaysia. Distinctive variation data obtained in the C-terminal non-repeat region of csp could be beneficial for the design and development of vaccines to treat P. knowlesi.

Genetic polymorphism of the thrombospondin-related apical merozoite protein (TRAMP) of Plasmodium knowlesi in Malaysia

“…The haplotype network revealed no distinct geographical separation of PkTRAMP between Peninsular Malaysia and Malaysian Borneo. This is similar to P. knowlesi proteins such as the PkCSP (Chong et al 2020) and PkMSP7D . However, some P. knowlesi genetic markers such as PkDBPαII (Fong et al 2015), PkAMA-1 (Ng et al 2021), PkCOXI, and PkA-type 18S rRNA (Yusof et al 2016) showed distinct geographical separation.…”

“…In the current study, the nucleotide diversity of PkTRAMP (π = 0.009) was found to be lower than that of other P. knowlesi genes such as merozoite surface protein 7D (MSP7D) (π = 0.052) , merozoite surface protein 3 (MSP3) (π = 0.046) (De Silva et al 2017), and circumsporozoite protein (csp) (π = 0.020) (Chong et al 2020), but almost similar to other low polymorphism genes such as AMA-1 (π = 0.006) (Ng et al 2021) and merozoite surface protein 4 (MSP4) (π = 0.007) . The haplotype diversity of PkTRAMP (Hd = 0.999) was similar to other P. knowlesi genes such as AMA-1 (Hd = 1.000) (Ng et al 2021) and MSP3 (Hd = 0.999) (De Silva et al 2017).…”