ABSTRACT
Objective To analyze the polymorphism of Plasmodium lactate dehydrogenase (pLDH) gene and predict B-cell epitopes in pLDH peptides in four species of human malaria parasites. Methods The blood samples and epidemiological characteristics were collected from malaria cases in Yunnan Province registered in the National Notifiable Disease Report System. The pLDH genes of four human Plasmodium species were amplified using nested PCR assay and sequenced. The polymorphisms of pLDH genes was analyzed using the software MEGA version 7.0.26 and DnaSP version 5.10, and the B-cell epitopes were predicted in pLDH peptides using the Immune Epitope Database (IEDB). Results The sequences of P. vivax LDH (PvLDH), P. falciparum LDH (PfLDH), P. ovale LDH (PoLDH) and P. malariae LDH (PmLDH) genes were obtained from 153, 29, 17 and 11 blood samples from patients with P. vivax, P. falciparum, P. ovale and P. malariae malaria, respectively, which included 15, 2, 4 and 2 haplotypes and had a nucleotide diversity (π) of 0.104. A high level of intra-species differentiation was seen in the PoLDH gene (π = 0.012), and the π values were all < 0.001 for PvLDH, PfLDH and PmLDH genes. Active regions of B-cell antigen were predicted in the pLDH peptide chain of four human malaria parasites, of 4 to 5 in each chain, and the activity score was approximately 0.430. Among these peptide chains, the “86-PGKSDKEWNRD-96” short-peptide was a B-cell epitope shared by all four species of human malaria parasites, and the “266-GQYGHS (T)-271” short-peptide was present in PvLDH and PoLDH peptide chains, while “212-EEVEGIFDR-220” was only found in the PvLDH peptide chain, and “208-LISDAE-213” was only seen in the PfLDH peptide chain. Conclusions The PoLDH gene polymorphism may be derived from the weak negative purification selection, while PvLDH, PfLDH and PmLDH genes may maintain a relatively conservative state. There may be two B-cell epitopes “212-EEVEGIFDR-220” and “208-LISDAE-213” in the proximal region of the C terminal in the pLDH peptide chain, which is feasible to differentiate between P. vivax and P. falciparum infections.
ABSTRACT
Objective To analyze the genetic diversity of Pomacea canaliculata based on the mitochondria DNA cytochrome c oxidase subunitⅠ(mtDNA COⅠ)gene as a molecular marker in Lincang City of Yunnan Province,so as to provide the scien-tific data for monitoring Angiostrongylus cantonensis in local areas. Methods The genotypes and polymorphisms of 38 speci-mens of P.canaliculata collected from Mengding Town of Lincang City were analyzed by sequencing COⅠgene.The phylogenet-ic tree and genetic distances were produced based on the haplotypes from GenBank and the present study by using the neighbour-joining method with the software MEGA version 6.06. Results Totally 31 sequences were acquired in the present study and they produced 3 unique haplotypes.Haplotype 1 showed a higher frequency compared to the others and it accounted for 83.9 % (26/31).The data showed that the least genetic distances ranged from 0 to 0.052 between P.canaliculata and 3 haplotypes,as well as the largest genetic distances ranged from 0.021 to 0.239 between Pila conica and 3 haplotypes.Otherwise,the analysis of the phylogenetic trees based on COⅠgene sequences of P.canaliculata indicated that all of 3 haplotypes clustered into one big clade with that from Japan(GenBank accession number: AB433769),China(GenBank accession number: KT313034)and USA(GenBank accession number:EU523129),which owned the closet relationship amongst them.Their genetic relationships were distantly related to the GenBank's reference sequences of P.insularum(GenBank accession number:EF514942),P.cam-ena(GenBank accession number: EF515059)and so on. Conclusion There is a P. canaliculata species in Lincang City of Yunnan Province as well as a high genetic diversity amongst the acquired 3 haplotypes in this study.