Comparison of allelic frequencies between blood donors and <i>P. knowlesi</i> infected patients.

<p>Comparison of allelic frequencies between blood donors and <i>P. knowlesi</i> infected patients.</p

Proportion of predicted phenotypes of blood donors and <i>P. knowlesi</i> infected patients.

<p>Proportion of predicted phenotypes of blood donors and <i>P. knowlesi</i> infected patients.</p

Median Joining network of PkDBPαRII haplotypes.

<p>The network shows geographical clustering of PkDBPαRII haplotypes from Peninsular Malaysia (yellow) and North Borneo (red). Amino acid sequences used for the construction of this network were from a previous study [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0155627#pone.0155627.ref009" target="_blank">9</a>].</p

Genetic Diversity, Natural Selection and Haplotype Grouping of <i>Plasmodium knowlesi</i> Gamma Protein Region II (PkγRII): Comparison with the Duffy Binding Protein (PkDBPαRII)

<div><p>Background</p><p><i>Plasmodium knowlesi</i> is a simian malaria parasite that has been reported to cause malaria in humans in Southeast Asia. This parasite invades the erythrocytes of humans and of its natural host, the macaque <i>Macaca fascicularis</i>, via interaction between the Duffy binding protein region II (PkDBPαRII) and the Duffy antigen receptor on the host erythrocytes. In contrast, the <i>P</i>. <i>knowlesi</i> gamma protein region II (PkγRII) is not involved in the invasion of <i>P</i>. <i>knowlesi</i> into humans. PkγRII, however, mediates the invasion of <i>P</i>. <i>knowlesi</i> into the erythrocytes of <i>M</i>. <i>mulata</i>, a non-natural host of <i>P</i>. <i>knowlesi</i> via a hitherto unknown receptor. The haplotypes of PkDBPαRII in <i>P</i>. <i>knowlesi</i> isolates from Peninsular Malaysia and North Borneo have been shown to be genetically distinct and geographically clustered. Also, the PkDBPαRII was observed to be undergoing purifying (negative) selection. The present study aimed to determine whether similar phenomena occur in PkγRII.</p><p>Methods</p><p>Blood samples from 78 knowlesi malaria patients were used. Forty-eight of the samples were from Peninsular Malaysia, and 30 were from Malaysia Borneo. The genomic DNA of the samples was extracted and used as template for the PCR amplification of the PkγRII. The PCR product was cloned and sequenced. The sequences obtained were analysed for genetic diversity and natural selection using MEGA6 and DnaSP (version 5.10.00) programmes. Genetic differentiation between the PkγRII of Peninsular Malaysia and North Borneo isolates was estimated using the Wright’s F_ST fixation index in DnaSP (version 5.10.00). Haplotype analysis was carried out using the Median-Joining approach in NETWORK (version 4.6.1.3).</p><p>Results</p><p>A total of 78 PkγRII sequences was obtained. Comparative analysis showed that the PkγRII have similar range of haplotype (Hd) and nucleotide diversity (π) with that of PkDBPαRII. Other similarities between PkγRII and PkDBPαRII include undergoing purifying (negative) selection, geographical clustering of haplotypes, and high inter-population genetic differentiation (F_ST index). The main differences between PkγRII and PkDBPαRII include length polymorphism and no departure from neutrality (as measured by Tajima’s D statistics) in the PkγRII.</p><p>Conclusion</p><p>Despite the biological difference between PkγRII and PkDBPαRII, both generally have similar genetic diversity level, natural selection, geographical haplotype clustering and inter-population genetic differentiation index.</p></div

Median Joining network of PkγRII haplotypes.

<p>The network shows geographical clustering of PkγRII haplotypes from Peninsular Malaysia (yellow) and North Borneo (red). Note that haplotypes from each region are divided into two subgroups.</p

Amino acid sequence polymorphism in the PkγRII haplotypes from Peninsular Malaysia and North Borneo.

<p>Polymorphic amino acid residues are listed for each haplotype. Residues identical to those of haplotype H1 are marked by dots. Total number of sequences for each haplotype is listed in the panel on the right.</p

Genetic diversity and selection pressure of PkγRII of <i>P</i>. <i>knowlesi</i> isolates in Peninsular Malaysia and North Borneo.

<p>Genetic diversity and selection pressure of PkγRII of <i>P</i>. <i>knowlesi</i> isolates in Peninsular Malaysia and North Borneo.</p

Sliding window (size = 10 bp, length = 5 bp) analysis of genetic diversity (π) across the Th2R/Th3R region.

<p>The x-axis represents CSP amino acid positions 278 to 336. The Th2R and Th3R regions are indicated.</p

Most abundant PkCSP Th2R/Th2R haplotypes.

<p>Most abundant PkCSP Th2R/Th2R haplotypes.</p

Median Joining Network of CSP generated using Network program.

<p>The network shows relationship among the 27 haplotypes based on the human Th2R/Th3R sequences from Peninsular Malaysia, Singapore and macaques from Peninsular Malaysia. Numbers in larger circles represent number of haplotypes, unnumbered circles represent single haplotypes. The smaller red circles are median vectors generated by the program.</p