Phylogenetic Analysis Of HPIV-3 Gene And Clinical Characteristics In Children With Acute Respiratory Tract Infection In Qingdao In 2018-2019

Objective:The purpose of this research is to:(1)discuss human parainfluenza virus type3(HPIV-3)infection in children with acute respiratory tract infection(ARTI)in Qingdao,Shandong Province;(2)analyze the gene characteristics of HPIV-3 hemagglutinin-neuraminidase(HN)protein;(3)explore the clinical characteristics of HPIV-3 infection.It may provide basic data and scientific evidence for the molecular epidemiological study of HPIV-3 in China and the prevention and treatment of acute respiratory tract infection in children.Methods:This study belonged to sentinel monitoring.A total of 1674 respiratory samples were collected from children with ARTI,in three hospitals(Qingdao Women and Children’s Hospital;West Coast Branch of Affiliated Hospital of Qingdao University;Laoshan Branch of Affiliated Hospital of Qingdao University)from January 2018 to December 2019.Furthermore,sixteen common respiratory viruses were screened by Multiplex real-time fluorescence polymerase chain reaction.The infection rate of HPIV-3,gender distribution,age distribution,and seasonal characteristics of onset in patients with ARTI were examined.For HPIV-3 positive specimens,we performed nested PCR to amplify the full length of the HN gene in HPIV-3,and then sequencing.Comparative analysis and homology analysis were performed for the sequence results and the prototype strain of HPIV-3 in Gen Bank,as well as representative strains epidemic in China and other countries.And then we used the Neighbor-Joining(NJ)method to establish a phylogenetic tree.At the same time,the hospital’s electronic medical record systems were used to collect data about HPIV-3positive children,and to analyze and compare the laboratory test indexes of HPIV-3 single infections and mixed infections.Results:In the 1674 respiratory specimens collected from children with ARTI,positive specimens for viruses were 989,with a positive rate of 59.08%.Among the 989 samples,90 samples were HPIV-3 positive,with a rate of 5.37%.The positive rates of HPIV-3 in male and female children were 5.50%and 5.20%,respectively.And the difference was not statistically significant(X~2=0.072,P=0.788).In each age group(under 1 year,1-3 years,3-6 years,and above 6 years),the positive rate for HPIV-3 was 8.73%,6.18%,3.25%,and0.92%,respectively.The difference between the four age groups was statistically significant(X~2=20.928,P<0.001).HPIV-3 infection in Qingdao shows a seasonal distribution,and the positive rates during the four seasons were 9.20%,18.56%,3.70%,and 0.50%,respectively,with a statistically significant difference(X~2=90.536,P<0.001).Children are mainly to be diagnosed with HPIV-3 infection with bronchopneumonia(45.6%),pneumonia(22.2%)and asthmatic bronchitis(17.5%).Clinical symptoms are mainly cough(100.0%),fever(88.6%)and expectoration(60.0%).And abnormal chest CT findings are mainly increased lung markings(78.9%),blurred lung margins(50.0%),and patchy high-density shadows(45.6%).Additionally,in these 90 HPIV-3-positive children,74 were detected single-virus infected,and 16 were detected as multiple-virus infected.And the mixed infection rate was 0.96%.However,no significant difference was observed in laboratory test indexes between the HPIV-3 single infection group and the mixed infection group(P>0.05).44 full-length sequences of HPIV-3 HN gene were obtained by nested PCR.Homology analysis showed that the nucleotide and amino acid homology of 44 Qingdao isolates were 97.0%-100.0%and 98.5%-100.0%,respectively.Additionally,nucleotide homology compared with the representative strain of South Africa and amino acid homology compared with the representative strain of Australia were the lowest,the homologies were 93.1%and 96.1%,respectively.While the nucleotide and amino acid homologies with the representative strain from Malaysia were the highest(99.8%and100.0%,respectively).The results of sequence alignment and evolutionary analysis showed that the HPIV-3HN gene belonged to the C3a and C3b branches of the C3 genotype,with30 strains of subtype C3a and 14 strains of subtype C3b.Conclusions:From January 2018 to December 2019,HPIV-3 was one of the most important viral pathogens causing acute respiratory tract infections of children in Qingdao,Shandong Province.Infection with HPIV-3 is mainly concentrated in spring and summer,and children under 1-year-old are most susceptible.The clinical characteristics of HPIV-3infection are non-specific and the infection of HPIV-3 is commonly accompanied with other respiratory viruses.Two of the most common co-infection viruses with HPIV-3 are Human enterovirus(HEV)and Human rhinovirus(HRV).The C3a and C3b branches of HPIV-3 C3 genotype were circulating prevalent in Qingdao,Shandong Province.The variation rate of the HN gene was low.And the HN gene has certain regional characteristics in evolution.