Enterovirus Infection In Hospitalized Children With Hand,Foot And Mouth Disease:Diagnosis,Genetic And Clinical Feature

ObjectiveAnalysis of the pathogenic components of hand,foot and mouth disease(HFMD)in hospitalized children from 2014 to 2016 in Tianjin Children's Hospital,molecular typing of enterovirus causing HFMD epidemic,and study on the association between MDA5 and CTLA4 gene polymorphisms and the susceptibility of enterovirus infections of hand,foot mouth disease,and analysis of the clinical manifestations of various types of infected viruses,will contribute to the prevention,control,diagnosis,and treatment of this disease.MethodAll 499 samples of HFMD cases which were collected from Tianjin Children's Hospital from Jul.2014 to Nov.2016,and subsequently were tested for the nucleotide acid of enterovirus(EV),human enterovirus71(EV71),Coxsackievirus A16(CVA16)by real-time reverse transcription polymerase chain reaction(RT-PCR).Among 170 sample of other EV positive were selected to amplify and sequence the whole VP1 region via RT-PCR,homology was analyzed and phylogenetic tree were constructed by comparison of the sequence with all subgenotype of EV by Chromas1.62 and MEGA6.06.Statistical analysis was used by SPSS20.0.Serum samples from 389 children with VP1 genotype successfully performed by RT-PCR were tested for EV71 IgM antibodies,CVB-IgM and CVB-IgG antibodies by enzyme-linked immunosorbent assay(ELISA).The polymorphisms of MDA5 gene rs1990760 and CTLA4 gene rs231775 in 151 HFMD-infected patients with HFMD and normal controls were analyzed by direct PCR.ResultsAll the 499 HFMD,429 EV positive cases were found,the constituent ratio of EV71,CVA16 and other EV were 54.1%(232/429),6.3%(27/429)and 39.6%(170/429),respectively.In all 170 samples of other EV,47 were CVA10(11.0%,47/429)and 35 were CVA6(8.2%,35/429).Phylogenetic analysis of 28 EV71 VP1,22 CVA10 VP1 and 16 CVA6 VP1 showed that EV71 belonged to genotype C4 a,CVA10 belonged to genotype G and CVA6 belonged to genotype F,respectively.The specificities of EV71-IgM,CVB-IgM,and CVB-IgG were 93.6%,82.6%,and 71.5%,respectively.Compared with the asymptomatic control group,the genotypes and alleles of MDA5 gene rs1990760 and CTLA4 gene rs231775 in patients with enterovirus infected HFMD were not statistically significant(P all> 0.05).Conclusion1.All 21 different EV were detected in this study.Although the positive rate of EV71 was still the highest,the types and proportions of other non-EV71 non-CVA16 enteroviruses increasing.And we found that the EV71 in this study was still subtype C4 a by phylogenetic tree,whereas CVA10 clustered into G clusters and CVA6 clustered into F clusters.2.In our study,30 were found to have common symptoms of the 499 children,402 were severe,and 67 were critically illness.The prevalence of children under 5 was the most common,and 1-year-old children had the highest positive detection rate.We should be guarded against the possibility of young children developing severe illness.EV71 had the longest average hospital stay.The number of HFMD-induced severe HFMD cases was the highest,reaching 49%,and 96.12% of EV71-infected children had neurological complications.The average age of children with HFMD caused by CVA16 infection was the highest.3.The positive predictive value of CVB-IgM/IgG antibody is too low by ELISA test,and there is a cross reaction between EV71-IgM and CVB-IgM CVB-IgG.4.Genetic polymorphisms of MDA5 gene rs1990760 G/A and CTLA4 gene rs231775 A/G in HFMD-infected HFMD patients were not significantly associated with the disease,affecting the susceptibility and severity of HFMD The degree of host immune response mechanism needs further study.