| Objective:To investigate the prevalence, epidemiology, genotype, recombination and clinical profiles of human bocavirus (HBoV) and human adenovirus (HAdV) among children with severe acute respiratory tract infection (SARI). Determine the epidemic strains and molecular biological characteristics so that provide reference for the control and prevention of related diseases as well as the molecular epidemiological study of human bocavirus and human adenovirus.Methods:Induced sputum (IS) samples were collected from 293 hospitalized children with severe community acquired pneumonia (CAP) from September 2007 to April 2008 in Wenling Hospital of Zhejiang Province. From May 2008 to March 2010, 259 nasopharyngeal aspirates (NPAs) were collected from hospitalized children with severe acute respiratory tract infection (SARI) in Beijing Children’s Hospital, m addition, from June 2013 to March 2014, a total of 441 NPAs were collected from children with SARI in Shanghai Paediatrics Hospital Affiliated Fundan University. Human adenovirus and human bocavirus were detected by nested PCR for typing followed by sequencing. The complete genome of human bocavirus was amplified to investigate the phylogenetic and mutation characteristics. The human adenovirus was isolated, purification and appraisal. The main coding sequences of structure gene and complete genome were amplified by PCR and next generation sequencing respectively to determine the phylogenetic and recombination profiles. All data were used to investigate the related molecular epidemiology characteristics, phylogenetic, recombination profiles and clinical presentations.Results:Part one. Typing and analysis of Human bocavirus:Typing detection of human bocavirus:All of the samples,200 (200/993,20.1%, 95%CI:17.3%～22.9%) were found to contain HBoV. And the infection rate of HBoV in Beijing, Shanghai and Wenling were 21.6%(56/259,95%CI:16.0%～27.3%), 17.1%(78/441,95%CI:13.8%-21.6%) and 22.5%(66/293,95%CI:17.1%～28.0%), respectively. The difference of region distribution of HBoV infection was not significant. Through sequences alignment and phylogenetic analysis, we found HBoV-positive cases 98.5%(197/200) were positive for HBoV1. The HBoV2 and HBoV3 contained only one and two positive samples, respectively. The HBoV showed a high co-infection rate with other respiratory viruses (63.6%-94.6%).The epidemiological and clinical profiles:No significant difference in gender distribution of HBoV infecion. Most of the SARI children with HBoV infection were younger than two years, mainly were between 6 months and 2 years old (P<0.05). And the peak seasons for HBoV infection were autumn, winter and spring. No significant difference of clinical profiles was shown between patients with HBoV and non-HBoV.The phylogenetic and mutation analysis of HBoV:The phylogenetic analysis both of the complete genome and coding sequences reveals that the virus was relative conserved, and the results of phylogenetic tree of the VP1/VP2 was similar to the complete genome.Part two. Typing and analysis of human adenovirus:Typing detection of human adenovirus:In total,127 (12.8%,127/993,95%CI: 10.6%-15.0%) of 993 SARI patients were positive for HAdV, and the infection rates of HAdV in Beijing, Shanghai and Wenling and were 20.1%(52/259,95%CI: 14.6%～25.5%),11.6%(51/441,95%CI:8.4%～14.7%) and 8.2%(24/293,95%CI: 4.9%～11.5%), respectively. The infection rate of Beijing was higher than the Wenling and Shanghai. We found HAdV-B (10 of HAdV-3,24 of HAdV-7) was 65.4%(34/52, 95%CI:43.4%-87.4%), HAdV-C (6 of HAdV-1,5 of HAdV-2/6,6 of others) was 32.7%(17/52,95%CI:17.2%-48.2%) and HAdV-D contained only one (HAdV-37) in Beijing. And HAdV-37 and HAdV-57 were first detected from the respiratory samples. In Shanghai, typing data shown that HAdV-B (17 of HAdV-3,16 of HAdV-7) was most dominant as 64.7%(33/51,95%CI:42.6%～82.8%), followed by HAdV-C as 27.5%(14/51,95%CI:13.1%～41.8%), either HAdV-D (HAdV-8) or HAdV-E (HAdV～4) contained only one case, and HAdV-F (HAdV-41) found in two cases. In Wenling, we found HAdV-C (8 of HAdV-2,6 of HAdV-5,5 of HAdV-1,1 of HAdV-6) was 83.3%(20/24,95%CI:46.8%～100%), HAdV-B (2 of HAdV-3,1 of HAdV-7) was 12.5%(3/24,95%CI:0～26.6%) and HAdV-E contained only one (HAdV-4) case. The HAdV showed a high co-infection rate with other respiratory viruses (64.7%-90.4%).The epidemiological and clinical profiles:No significant difference in gender distribution. Most of the SARI children with HAdV infection were between 6 months and 2 years old (P<0.05). The peak seasons for HAdV infection were winter and spring. No significant difference of clinical profiles was shown between SARI patients with HAdV and non-HAdV.The phylogenetic and recombination analysis of HAdV:The phylogenetic and recombination analysis reveals the virus occur recombination, which is the recombinant strain of HAdV-1,-2,-5,-6 and HAdV-57 of specie HAdV-C.Conclusion:In this study, we got the molecular epidemiology characteristics of HBoV and HAdV among children with SARI and the epidemic strains in different regions. In addition, the sequence of human bocavirus was conserved and the VP1/VP2 can be the region for genotyping. And we detected HAdV-37 and HAdV-57 from the respiratory samples the first time. The human adenovirus occurred recombination between strains of the species HAdV-C.Which is the recombination strain of HAdV-1,-2,-5 and HAdV-6. |
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