Etiological Investigation Of HFMD In Fuzhou And Study On The Evolution Of Gene

Objective1.To investigate etiology of HFMD comprehensively, and to master and clarifythe pathogenic spectrum and the genetic characteristics of HFMD in Fuzhou, in orderto provide scientific evidence for the prevention and control of HFMD.2.To identify the CVA6positive proportion,and to analyze epidemiologicalcharacteristics of CVA6positive samples.Methods1.1031samples were collected from clinically diagnosed cases with HFMD,real-time quantitative RT-PCR, Nested RT-PCR and BLAST of VP1sequences werecarried out for nucleonic acid identification and typing.2. To identify the other enteroviruses by a pair of CVA6specific primers.Combined with epidemiological data, To analyze the distribution of CVA6positivesamples.3. Specimens identified EV71,CVA16,CVA6was performed for Viral isolationand complete VP1and5’-UTR amplification and sequencing, compared and analyzedby MEGA5.0.1.The Homology and Phylogenetic analysis constructed base on thealignment of the complete VP1and5’-UTR sequences of Fuzhou strains and thecorresponding strains from GenBank were performed by Mega5.0.2.Results1. Among the1031clinical samples878(85.16%)cases were tested EV positive,including426cases of EV71positive with the positive rate of45.52%(426/878),93cases of CVA16positive with the positive rate of10.60%(93/878),and359otherenterovirus positive with the positive rate of40.88%(359/878). Among the otherenterovirus positive samples, we found218cases of CVA6positive with the positiverate of24.83%(218/878).CVA10and Echo3were identified. 2. Epidemiological characteristics.Time: the positive ratio of CVA6changedsignificantly.the cases of September mostly caused by other enteroviruses, with thepositive ratio of CVA6of62.50%. Region: the constituent ratio of Yongtai washigher than other counties (cities), with the constituent ratio of54.55%(95%CI:25.12%-83.97%). Population: Most of CVA6cases were aging from0to5years old(94.04%), gender ratio was1.66:1.3. Fuzhou EV71strain shared the highest VP1similarity with the C4aSubgenotype (92.9%-97.3%), with the similarity of96.7%with Fuyang (Anhui2008).The VP1phylogenetic tree: EV71can be divided into three genotypes and Fuzhoustrains were grouped to subgenotype C4a.For5’-UTR sequence, Fuzhou EV71strainshared the highest VP1similarity with the C4a Subgenotype (96.7%-98.8%). The5’-UTR phylogenetic tree: Fuzhou EV71strain clustered into C4a subgenotype.4. Fuzhou CVA16strain shared the highest VP1similarity with the B1bsubgenotype (90.8%-93.2%). The VP1phylogenetic tree: CVA16can be divided intotwo genotypes and Fuzhou strains were grouped to B1b subgenotype. For5’-UTRsequence, Fuzhou CVA16strain shared the highest VP1similarity with the B1bsubgenotype (94.8%-97%). The5’-UTR phylogenetic tree: Fuzhou EV71strainclustered into B1b subgenotype.5. The vp1nucleotide homologies between Fuzhou strains and prototype ofCVA6were77.4%-79.8%.The VP1phylogenetic tree: CVA6can be divided intothree cluster and Fuzhou strains were grouped to C cluster. Fuzhou CVA6strainshared the highest VP1similarity with the C cluster (86.1%-98%).For5’-UTRsequence, the nucleotide homology between Fuzhou strain and cluster A, C were89.1%-91.3%and91.7%-99.7%, respectively. Fuzhou CVA6strain shared the highestVP1similarity with the cluster C.Fuzhou CVA6strain shared the highest VP1similarity with the B1b subgenotype (94.8%-97%). The5’-UTR phylogenetic tree: theCVA65’-UTR phylogenetic tree illustrated the same classification as the VP1tree. Conclusions:1. EV71, CVA16and CVA6are the main intestinal pathogen of HFMF inFuzhou. CVA6has become the second largest pathogens over CVA16.2. Fuzhou EV71strain belongs to C4a subgenotype, with CVA16strainbelonging to B1b subgenotype, and CVA6strain belonging to C cluster, which isconsistent with the trend of mainland china in recent years.3. The VP1tree phylogenetic tree illustrated the same classification as the5’UTR tree. The combination of VP1and5’-UTR phylogenetic tree will assist withboth individual patient diagnosis and public health measures.