Analysis Of Virus Community Of Children With Acute Respiratory Diseases With Unknown Etiology And Genetic Characteristics Of Novel Viruses

ObjectiveAcute respiratory infection is a major public health problem and a leading cause of illness and death in children under 5 years of age.Viruses are the main cause of acute respiratory diseases in children.A large number of viruses have been found in the respiratory tract,and the new respiratory virus,SARS-Co V-2,has also caused huge physical and mental health damage and property losses to people all over the world in the past three years.With the development of molecular biology technology,multiple real-time fluorescent polymerase chain amplification technology has been widely used in the screening of respiratory viruses,but for new,mutated or low-titer viruses,this technology cannot detect and may false negatives occur.At the same time,the routine screening for pathogens of respiratory infections in hospitals does not include all respiratory viruses,usually including influenza virus,respiratory syncytial virus and parainfluenza virus.Therefore,it is very important to use viral metagenomics technology to explore the composition of respiratory viruses.It can quickly detect the viruses in the samples and discover the dominant viruses in the composition of respiratory viruses in people in a certain area.MethodsIn this study,445 throat swab samples of children(<5 years old)with unexplained acute respiratory diseases were collected in Xuzhou Children’s Hospital,and bacteria,mycoplasma,influenza virus and respiratory syncytial virus had been excluded by the hospital’s routine screening of respiratory pathogens.The 445 samples were categorised according to the time of collection in spring(March-May),summer(June-August),autumn(September-November)and winter(December-February),and the samples were evenly distributed according to 10 libraries per season,for a total of 40 libraries.The metagenomics method was used to study the composition of the viral community in the respiratory tract of children in this region,and species diversity analysis was used to compare the differences in the composition of the viral community among the four seasons of spring,summer,autumn and winter.The genetic diversity of the viruses found in this study was further explored by constructing a phylogenetic tree for phylogenetic analysis,as well as comparing the high-level structure of conserved region proteins to predict functional changes,and finally an epidemiological investigation of important viruses was carried out using polymerase chain reaction.Results(1)Composition of viral communities in the respiratory tractThrough deep sequencing and data analysis of 40 libraries,the composition of respiratory viruses in this study was described in detail.At the family level,the sequences identified as viruses in the libraries were classified into 17 viral families in total,among which Picornaviridae,Parvoviridae,Anelloviridae and Paramyxoviridae were relatively abundant viral families in the whole.In addition,we ranked the top five relative abundance families,and the abundance of viruses from high to low was Picornaviridae,Parvoviridae,Coronaviridae,Anelloviridae and Paramyxoviridae.The family Picornaviridae dominates the RNA viruses,while the Parvoviridae family occupies the largest proportion among the DNA viruses.At the level of virus genus,a total of 67 virus genera were found to be classified into the above-mentioned 17 virus families,and the highly abundant virus genera were Betatorquevirus,Bocaparvovirus,Respirovirus,Betacoronavirus,Cardiovirus,Enterovirus and Parechovirus,and the abundance of other virus genera were low.Overall,most of the virus genera with high abundance belonged to the two most abundant virus families.(2)Differences in the composition of respiratory viruses among the four seasonsThe virus composition of the four seasons was not the same.At the family level,the abundance of Parvoviridae was the highest in the spring group,followed by Picornaviridae,and the rest of the virus families were all low abundance.The virus community composition of the summer group was dominated by Picornaviridae.The virus composition of the autumn and winter groups was more abundant than that of the spring and summer groups,and the abundance of each virus family in autumn and winter was very balanced.Likewise,comparisons of virus composition at the genus level indicated the same.At the species level,9 species were shared by the four seasons and could be detected in all four seasons.There are 56 species that exist alone in one of the four groups,among which the autumn group has the most individual species,and the winter group has the least.Alpha diversity analysis showed that there were no significant differences in the species abundance and richness of viruses among the four seasons,while Beta diversity analysis showed that there were significant differences in the composition of respiratory virus communities between groups.(3)Phylogenetic analysis of virusesA total of 23 nearly complete viral sequences were spliced in this study.Through phylogenetic analysis,it was found that 9 sequences belonged to the Enterovirus genus of the Picornaviridae family,1 sequence belonged to the Cardiovirus genus of the Picornaviridae family,3 sequences belonged to the Paraenterovirus genus of the Picornaviridae family,5 sequences belonged to the genus Bocavirus in the family Parvoviridae,and 5 sequences belonged to the family Anelloviridae.RNA viruses are prone to mutation and recombination.The phenomenon of viral recombination was also found this time,and recombination analysis was carried out to speculate on the sequence of its primary and secondary parents.(4)Epidemiological survey of important virusesHuman enterovirus(HEV)and human bocavirus(HBo V)had the highest abundance in the virus community,so further epidemiological investigations were carried out on these two important viruses.The results showed that the positivity rates for HEV and HBo V were 2.92% and 8.09% respectively.Also,both viral distributions had seasonal variations,with spring being the high season for HBo V infections,while summer was the peak season for HEV infections.ConclusionsThis study examined the composition of respiratory viruses of unknown etiology in children with pronounced acute respiratory symptoms,identified the prevalent viruses and their genetic characteristics,and investigated the prevalence and seasonal variation of the two viruses in the region.There are obvious differences in the virus composition among the four seasons,and the virus composition in autumn and winter is more abundant than that in spring and summer.In addition,we detected potential recombination events that may indicate the presence and spread of viral infection.This study reveals the composition and diversity of respiratory virus communities in children with unexplained respiratory tract infections in Xuzhou,which will provide useful information for the prevention and treatment of acute respiratory infections in children and help to formulate effective prevention strategies.