Etiology analysis of hand, foot and mouth disease cases and genetic characteristics of coxsackievirus A6 in six prefectures/cities of Yunnan Province in 2018

Objective To detect the enterovirus VP4 and VP1 genes in 510 stool samples collected from hand, foot and mouth disease (HFMD) cases and analyze the phylogenetic characteristics of the entire VP1 genes of coxsackievirus A6 (CV-A6) strains in six prefectures/cities of Yunnan Province in 2018. Methods Viral RNA was abstracted from the stool samples. VP4 gene sequences were amplified by RT-PCR and sequenced using the MD91/OL68-1 primer pair to identify viral genotypes. Whole VP1 gene sequences were amplified and sequenced using appropriate primer pairs. The whole VP1 gene sequences of CV-A6 reference strains were downloaded from GenBank. MEGA5.2 software was used to analyze the similarity in nucleotide and amino acid sequences between different strains and phylogenetic tree was constructed for analysis of genetic characteristics and molecular epidemiology. Results VP4 and VP1 gene sequences were obtained from 57 out of 510 stool samples with a positive rate of 11.17% (57/510). There were 43 CV-A6 (8.43%, 43/510), six CV-A10 (1.17%, 6/510), two enterovirus A71 (EV-A71, 0.39%, 2/510) and two CV-A9 (0.39%, 2/510) strains. The other four strains were CV-A4 (0.19%, 1/510), CV-A5 (0.19%, 1/510), CV-B1 (0.19%, 1/510) and E11 (0.19%, 1/510). The phylogenetic analysis showed that all 43 CV-A6 strains belonged to sub-genotype D3. Conclusions In the 510 HFMD samples, CV-A6 strains were mostly detected with a detection rate of 8.43% and accounted for 75.44% (43/57) of all isolates, followed by CV-A10 (1.17%, 6/510) and EV-A71 (0.39%, 2/510). There was a large HFMD outbreak mainly caused by CV-A6 in Yunnan Province in 2018. The outbreak was caused by CV-A6 of sub-genotype D3, as was the case with pervious outbreaks in China. Key words: Hand, foot and mouth disease; Etiology; Coxsackievirus A6; VP4 gene; VP1 gene; Genetic characteristics analysis