Epidemic And Genetic Characteristics Of Influenza A(H1N1) In Shandong Province At The Late Stage Of Pandemic

ObjectiveThis year is the 10 th year of the 2009 influenza a(H1N1)pandemic.In August 2010,the WHO declared that the new influenza a(H1N1)virus became the new seasonal H1N1 influenza virus in the post-pandemic influenza.Describe and analyse the epidemiological characteristics of A Influenza virus(H1N1)in Shandong province since the H1N1 Influenza Pandemic,analyse the risk factors of severe cases and HA and NA genes characterristic,building genetic evolutionary tree and discussing its genetic variation regularity,providing the theoretical basis for predicting the development trend of influenza and the scientific basis for the future influenza surveillance and prevention and control.Methods1.Download influenza surveillance data from Chinese influenza surveillance information system and Infectious disease reporting information system and select the A influenza virus(H1N1)cases,describing the time,age and regional epidemiological characteristics.Select the severe death cases and analyse its risk factors.2.Select the A(H1N1)influenza strains from April 2014 to March 2018,extract RNA of A(H1N1)virus,design the primers of HA and NA genes influenza by Primer5.0software.Using the relevant primers of HA and NA genes for RT-PCR amplification,sending the products to the relevant company for sequencing,Collating and analysing the genes sequence with Edit Seq and MEGA5.05 software.According to the determined gene sequence,it is translated into amino acid sequence and compared with vaccine strains to find out the corresponding mutation site,and find out whether it is on the antigen-determining cluster or whether it is a key marker gene cluster.The evolutionary tree constructed is used to determine the geographical location of common ancestor of each branch node.Results1.The annual strain distribution results from 2014 to 2018 and risk factors of death in severe casesDuring the monitoring year of 2014-2018,a total of 8004 influenza virus strains(positive separation rate 13.37%)were detected in Shandong Provincial national influenza network laboratory,and the number of samples significantly increased from October to March of the next year,A(H1N1)influenza virus,seasonal H3N2 virus and type B virus circulated alternately.Positive cases were mainly concentrated in the age group of 5-14 years old,accounting for 16.68%.Weihai,Heze and Dezhou were the top three in terms of the positive isolation rate.A total of 63 severe cases were collected,including 40 males(63.49%)and 23females(36.51%),the single factor analysis show that gender,age,tachypnea,platelet toxic shock,respiratory failure,infection and multi-function viscera not congruent factors have an effect on influenza a(H1N1)severe death,multiariable logistic regression analysis of abnormal platelet content and respiratory failure is the a/H1N1 influenza a risk factor for severe cases of death.2.Genetic characteristics of HA and NA in A(H1N1)influenza strains from April2014 to March 2018The homology analysis of HA and NA gene sequences of H1N1 virus isolated from shandong province were 94.3～99.9% and 97.8～99.6%.HA gene phylogenetic tree analysis showed that the strains of 2015-2016 monitoring year were clustered,which was closely related to A/California/07/2009(H1N1),and most of the H1N1 influenza virus in 2016-2018 monitoring year were clustered,which was closely related to A/Michigan/45/2015-(H1N1).NA gene phylogenetic tree analysis showed that the most strains of 2015-2016 monitoring year gather into a gene cluster,with A/California/07/2009(H1N1)the genetic distance of the shortest,the 2016-2018 monitoring annual A(H1N1)influenza virus gather in a large branch,with A/Michigan/45/2015(H1N1)the genetic distance of the shortest,and the A/California/07/2009(H1N1)the genetic distance is farther.Evolutionary tree showed that the strains of influenza a(H1N1)virus had obvious temporal characteristics,but no obvious geographical characteristics.Compared with vaccine strains,the majority of influenza a(H1N1)strains in the 2014-2018 surveillance year had 18(3.17%)amino acid sites that were mutated,and 4 of them were located in antigenic site.The glycosylation sites analysis showed that a new glycosylation sites NQS/T162-164 was added.18(3.83%)amino acid sites of NA protein showed significant variation,and the glycosylation site NFS386-388 was lost.There’s no H275 Y mutation at the drug site.Conclusion1.The epidemic peak of influenza virus is from October to March of the next year,A(H1N1)influenza virus,seasonal H3N2 virus and influenza B virus circulate alternately.2.Thrombocytopenia and respiratory failure are risk factors for death in severe cases of influenza.3.The HA and NA genes of the H1N1 influenza strains isolated from Shandong province from April 2014 to March 2018 showed high homology.Amino acid variation occurred in both HA and NA proteins,partial amino acid substitution of HA protein occurred in the antigen-determining cluster area,and antigenic drift of HA gene was generally possible.The vaccine strain A/California/07/2009(H1N1)recommended by WHO has A certain protective effect on the H1N1 influenza strain monitored in 2015-2016,while A/Michigan/45/2015(H1N1)has A certain protective effect on the H1N1 influenza strain monitored in 2016-2018.4.From April 2014 to March 2018,the newly added glycosylation site of HA protein of influenza a(H1N1)strain isolated and the loss of a glycosylation site of NA protein may both affect the function of HA and NA genes.