| Objective:This study in order to kown well the clinical characteristics of our hospital(Kunming children’s hospital)by different pathogens after the infection of hospitalized children with hand,foot and mouth disease,as well as indepth understand of the different clinical charectestistics after infection by different pathogens,and improve to discieminate the different pathogen infectionclinical with diagnosis and treatment,then perform better treatment.In addition,the molecular epidemiological research of HFMD pathogens and the whole gene study of CA5 strains which in order to understand the molecular epidemiological characteristics of HFMD and HFMD pathogens in our hospital and study the whole gene of CA5(Coxsacieverus A)virus strain furtherly,were carried out.This study can help us to understand the evolution of hand,foot and mouth disease pathogens in our hospital futherly,and this study will play a great role in promoting the vaccine development of hand,foot and mouth disease virus(except EV71)in the future.Methods:Firstly,we collect basic information,clinical manifestations and laboratory examinations of the children confirmed with HFMD in the inpatient Haitai case system of Kunming Children’s Hospital from 2019 to 2020.We use SPSS.20.0 software for statistical analysis;Nextly,We collected 155 stool samples of hospitalized children with HFMD from 2019 to 2020.we were deposited fecal samples in the refrigerator,processed simples simply,extracted the viral nucleic acid,in the stool sample to amplify the nucleic acid by using the RT-PCR method.We sent the products obtained to Kunming Kinco Gene Company for sequencing,compared part of the sequenced VP1 sequences on NCBI,and used MEGA7.0 to construct a phylogenetic tree of the top three dominant virus strains detected.Finally,we analyze its evolutionary relationship.Lastly,we analyzed the two detected CA5(Coxsackievirus A5)strains.We obtained two reference strains of CA5 strains at NCBI,and used the CA5 reference strains to design primers on Primer Premier 5.After that,we use primers to perform PCR amplification,subject the PCR products to agarose gel electrophoresis,and then send the product of the target band after electrophoresis to Kunming Kinco Sequencing Company for sequencing.The head and tail ends are sent to the gene company for gene cloning,the different sequences obtained are spliced to obtain the full gene sequence of the two CA5 viruses,and then the evolutionary tree and gene recombination analysis are performed using MEGA 7.0 and Simplot 3.5.1.Results:1.A total of 1,294 inpatient children with HFMD were collected from 2019 to 2020.Among them,the incidence of HFMD in the age group of 1 to 2 years is slightly higher than that of other age groups.The age of onset in EV71,CA16(Coxsackievirus A16),CA6(Coxsackievirus A6),CA10(Coxsackievirus A10),其他(Enteroviruses,here means other types of enteroviruses except the above)is mainly in childhood,and the incidence of boys is more Girl.Among them,EV71 infection is more likely to cause severe and critical hand,foot and mouth disease.There were statistical differences in the clinical features of different pathogens in each group in terms of fever peaks,anorexia,nervous system symptoms,and circulatory system involvement(P<0.05).2.We collected a total of 155 stool samples from inpatient children with HFMD from 2019 to 2020.Among the 155 children with HFMD,108 were boys and 47 were girls,with a male-to-female ratio of 2.29:1.The age is mainly distributed in the age range of 1 to 2 years,99 cases of which are mild and 55 cases are severe.Enteroviruses were detected in 105 stool samples of children with HFMD and a total of 10 pathogens were detected.In this study,155 stool samples from children with HFMD showed no mixed infections.Among them,the top three dominant viruses tested were CA6(65.7%69/105),CA16(11.4% 12/105),and CA10(10.4% 11/105).CA6,CA16,and CA10 belong to D3,B1 a,and D2 subtypes,respectively.CA6 is the main pathogen causing hand,foot and mouth disease.3.The complete gene sequences of the two CA5 strains obtained by PCR amplification have the most homology with the CA5 strain isolated in Xiangyang in2017 There is recombination with other types of enteroviruses.For example,the recombination of CA12 is mainly in 5’UTR,VP2,VP3,and VP1.There is recombination with CA10 in the 5’UTR in the gene structure region,recombination with CA4 in 3D and 3’UTR,and recombination with CA16 in the 2A region.Conclusion:1.The main pathogen spectrum of inpatient children with hand,foot and mouth disease in this area has changed,and CA6 has become the main pathogen causing HFMD.The clinical characteristics of HFMD need to arouse the attention of researchers in the process of diagnosis and treatment in the future.2.It is imminent to strengthen the in-depth study of CA16,CA10,CA2,CA5,CA4 and other enteroviruses that cause hand,foot and mouth disease in children.3.The two CA5 strain of whole gene studied that which have high homology and recombination with Xiangyang isolates in this study. |
