[Polymorphism analysis of the block 5 region in merozoite surface protein-1 gene of imported and local Plasmodium vivax in Yunnan Province].

Abstract

Objective: To analyze the sequence of Plasmodium vivax merozoite surface protein-1(PvMSP-1) and allele polymorphism in imported and local vivax malaria parasites in Yunnan Province.

Methods: Blood samples on filter paper were collected from imported and local vivax malaria cases in Yunnan Province during August 2012 and September 2015 and information of epidemiological history was recorded. Plasmodium DNA was extracted by a DNA extraction kit, and the block 5 region in PvMSP-1 gene was amplified by PCR. The PCR products were sequenced and blasted with reference sequences M75674, AAN86237, M60807, ABJ53045, AAN86238 and BAA18977. The sequence polymorphism in block 5 region of PvMSP-1 was analyzed with MEGA 5.04 and Arlequin3.5.1 softwares. The conserved sites, genetic distances among sequences and Shannon-wiener index among alleles were calculated. The clustering tree was drawn according to the genetic distances between the amino-acid sequences.

Results: A total of 847 blood samples were collected from the malaria cases, comprising of 61 samples from local cases, 66 from imported cases from Africa, and 720 from Myanmar. The block 5 region in PvMSP-1 was successfully amplified in 278 samples, and sequencing was successfully made in 206 of them. The peptide coded by the block 5 region had a length of 193 to 222 aa. The amino acid sequence alignment showed that in 206 samples the proportion of genotypes of Sal-1, Belem and Recombine was 59.2%(122/206),23.3%(48/206) and 17.5%(36/206), respectively. The proportion of Sal-1 genotype in imported cases from Myanmar and Africa and in local cases was 58.8%(104/177),73.3%(11/15) and 50%(7/14), respectively. The genotypes Sal-1, Belem and Recombine had 51, 9 and 6 different alleles. The 66 alleles had a Shannon Wiener index (H’) of 0.955 and an expected heterozygosis (He) of 0.567. The 206 DNA sequences had a 665-bp homologous locus, comprising of 75 conserved sites (11.3%,75/665) and 590 variable sites (88.7%, 590/665). The genetic distances between sequences were all less than 0.4. The clustering analysis showed that the 206 sequences were clustered into two categories with three branches. The homology of Recombine with Belem genotype was 91%-92%, higher than with Sal-1 genotype (82%-83%).

Conclusion: The block 5 region in PvMSP-1 gene from local and imported Plasmodium vivax in Yunnan Province has varied forms of alleles, and the Sal-1 genotype is predominant among the three genotypes.