Objective:To analyze the epidemiological characteristics and genetic evolution of human coronavirus OC43（HCoV-OC43）in Hangzhou from 2018 to 2023，and provide molecular epidemiological basis for the prevention and control of HCoV-OC43 infection.Methods:A total of 11 904 samples of cases with acute respiratory tract infection were collected from two sentinel hospitals in Hangzhou. The collection period was from January 2018 to December 2023. HCoV-OC43 was detected by real-time fluorescent quantitative RT-PCR. The positive samples were amplified by PCR using specific primers，and the amplified products were sequenced using third-generation nanopore technology for the whole genome. The genetic evolution features were analyzed.Results:A total of 265 HCoV-OC43 positive samples were detected，with a positive detection rate of 2.23%（265/11 904）. There was no significant difference in detection rate between males and females（χ 2=0.042， P>0.05）；the detection rate varied significantly among different age groups（χ 2=60.104， P<0.01），with higher detection rates observed in children under five years old and individuals over 65 years old. The infection peaks in 2018-2019 and 2023 occurred in spring and summer，while the infection peak in 2020-2022 was in summer and autumn. A total of 85 complete HCoV-OC43 whole genome sequences were obtained through high-throughput sequencing. The evolutionary analysis indicated that the K genotype was predominant in Hangzhou from 2018 to 2023. In 2019，both J and K genotypes co-existed. In 2023，a new genotype，temporarily named genotype L，was discovered，which originated from genotypes J and K. The K genotype had 22 specific amino acid variation sites，most of which were located in the important domain of S protein. The J genotype had seven specific amino acid variation sites，most of which were located in non-structural proteins. The new genotype L had 10 specific amino acid mutation sites，which were distributed across six proteins，and these mutation sites had the most extensive coverage. Conclusions:HCoV-OC43 is an important pathogen causing respiratory diseases in children and the elderly in the Hangzhou area，and there are multiple genotypes in the Hangzhou area，with a new recombinant genotype L discovered in 2023.

Objective:To analyze the epidemiological characteristics and genetic evolution of human coronavirus OC43（HCoV-OC43）in Hangzhou from 2018 to 2023，and provide molecular epidemiological basis for the prevention and control of HCoV-OC43 infection.Methods:A total of 11 904 samples of cases with acute respiratory tract infection were collected from two sentinel hospitals in Hangzhou. The collection period was from January 2018 to December 2023. HCoV-OC43 was detected by real-time fluorescent quantitative RT-PCR. The positive samples were amplified by PCR using specific primers，and the amplified products were sequenced using third-generation nanopore technology for the whole genome. The genetic evolution features were analyzed.Results:A total of 265 HCoV-OC43 positive samples were detected，with a positive detection rate of 2.23%（265/11 904）. There was no significant difference in detection rate between males and females（χ 2=0.042， P>0.05）；the detection rate varied significantly among different age groups（χ 2=60.104， P<0.01），with higher detection rates observed in children under five years old and individuals over 65 years old. The infection peaks in 2018-2019 and 2023 occurred in spring and summer，while the infection peak in 2020-2022 was in summer and autumn. A total of 85 complete HCoV-OC43 whole genome sequences were obtained through high-throughput sequencing. The evolutionary analysis indicated that the K genotype was predominant in Hangzhou from 2018 to 2023. In 2019，both J and K genotypes co-existed. In 2023，a new genotype，temporarily named genotype L，was discovered，which originated from genotypes J and K. The K genotype had 22 specific amino acid variation sites，most of which were located in the important domain of S protein. The J genotype had seven specific amino acid variation sites，most of which were located in non-structural proteins. The new genotype L had 10 specific amino acid mutation sites，which were distributed across six proteins，and these mutation sites had the most extensive coverage. Conclusions:HCoV-OC43 is an important pathogen causing respiratory diseases in children and the elderly in the Hangzhou area，and there are multiple genotypes in the Hangzhou area，with a new recombinant genotype L discovered in 2023.

Objective:To investigate the co-infection and phylogenetic analysis of human rhinovirus (HRV) in 2019 novel coronavirus (2019-nCoV) positive samples.Methods:Ten common respiratory viruses, including HRV were detected by real-time fluorescence quantitative polymerase chain reaction (qPCR) in 7 213 samples of 2019-nCoV positive cases and the co-infection characteristics were analyzed. The VP4/VP2 gene fragment of HRV was amplified and sequenced.Phylogenetic trees were constructed.Results:HRV accounted for 1.34% of the 2019-nCoV positive samples (97/7 213), followed by common coronavirus (0.50%, 36/7 213). The co-infection rate of HRV in 2019-nCoV positive samples was significantly different from that of other viruses ( χ2=318.09, P<0.001). There was significant difference in HRV co-infection rate among different age groups ( χ2=36.77, P<0.001), the peak was in<18 years age group. The co-infection rate of HRV had no significant difference in different seasons. The VP4/VP2 gene fragments of 39 HRV strains (40.21%, 39/97) were successfully sequenced and made phylogenetic analysis. There were 10 strains of HRV-A, 9 strains of HRV-B and 20 strains of HRV-C. Seventeen subtypes were identified, of which B6 (66.67%, 6/9) and C15 (70%, 14/20) were the most prevalent and other subtypes were scattered. Conclusions:The co-infection rate of HRV in patients with 2019-nCoV infection was the highest. The highest co-infection rate was in<18 years age group. Group A, B, and C of HRV were found in 2019-nCoV positive samples, and serotypes present diversity.

Descriptive epidemiological methods were used to analyze the epidemiological characteristics of the local cluster of COVID-19 in the logistic park of Yuhang District in Hangzhou in March 2022. The cluster epidemic was detected by a case who actively visited the fever clinic. The epidemic lasted for 8 days, and a total of 58 cases (53 workers, 2 students, 1 farmer, 1 teacher and 1 unemployed) were found, including 40 males and 18 females. The age was (33.29±12.22) years. There cases were mainly in Yuhang District (48 cases, 82.77%) and Shangcheng District (7 cases, 12.07%) of Hangzhou. The real-time regeneration number peaked at 2.31 on March 10 th and decreased to 0.37 on March 15 th. The sequencing result of the indicated case was 100% homologous with the sequence uploaded from South Korea on March 4 th, 2022.

Objective:To analyze the prevalence of influenza B virus in Hangzhou from 2014 to 2020 and the genetic evolution of seven reassortant strains of influenza B virus.Methods:Influenza viruses were isolated from throat swabs collected from 16 943 patients with influenza-like illness in Hangzhou from January 2014 to December 2020. The subtypes of influenza viruses were identified by real-time RT-PCR. Eight genes ( PB2, PB1, PA, HA, NP, NA, MP and NS) of influenza B viruses were amplified with specific primers and then analyzed with nanopore sequencing and phylogenetic analysis. Results:From January 2014 to December 2020, there were 1 090 influenza B virus-positive samples, including 474 samples of Yamagata lineage and 616 samples of Victoria lineage, were identified in Hangzhou with an overall positive rate of 6.43% (1 090/16 943). Whole genomes of 228 strains of influenza B virus were obtained by nanopore sequencing and seven reassortant strains of influenza B virus were found. There were four reassortant influenza B viruses of Yamagata lineage with NA gene fragments from viruses of Victoria lineage, two strains of Yamagata lineage (H644_BY and H648_BY) with NP and NA gene fragments from Victoria lineage and one strain of Victoria lineage with PB2, PB1, PA and NS gene fragments from Yamagata lineage. Meanwhile, these seven strains possessed several mutations in the antigenic sites of HA and NA genes. Conclusions:Several rare reassortant strains of influenza B virus with epidemic potential were detected in Hangzhou from 2014 to 2020, which indicated that the traditional detection methods should be improved and more attention should be paid to the reassortant influenza B viruses and the match between epidemic and vaccine strains.

Objective:To investigate the epidemic characteristics of influenza in children and the features of severe influenza.Methods:From January 2016 to September 2022, 1 600 samples from hospitalized cases of severe acute respiratory tract infection and 7 660 samples from outpatients with influenza-like illness were collected. Influenza virus was detected by real-time RT-PCR. Other respiratory viruses in the samples of severe hospitalized cases and some samples of outpatients were detected. Clinical features of influenza virus infection and co-infection were analyzed.Results:The positive rate of influenza virus in the 1 600 hospitalized cases of severe acute respiratory infection was 6.63% (106 cases). H1N1, H3N2, BV and BY were deteted in 49.06% (52 cases), 17.92% (19 cases), 29.25% (31 cases) and 3.77% (4 cases) of the 106 cases, respectively. The positive rate of influenza virus in the 7 660 out-patient cases was 15.01% (1 150 cases), and H1N1, H3N2, BV and BY were detected in 22.17% (255 cases), 30.96% (356 cases), 41.39% (476 cases) and 5.48% (63 cases) of the infected cases, respectively. Influenza A (H1N1) virus was more likely to cause severe influenza in children (χ 2=37.978, P<0.001), while seasonal H3N2 and BV strains were less likely to cause severe influenza in children (χ 2=7.871, P=0.005; χ 2=5.948, P=0.015). There was no statistically significant difference in the positive rates of BY lineage in the two groups. Severe influenza mainly occured in the peak season of influenza epidemic. There was no significant difference in the clinical manifestations between the children infected with the four different influenza viruses. In the 106 severe cases of influenza, the co-infection rate of influenza virus with other respiratory viruses was 17.92% (19 cases), while the co-infection rate reached 34.81% (47 cases) in 135 outpatient cases of influenza. The difference in the co-infection rates was statistically significant between outpatient and hospitalized cases (χ 2=10.734, P=0.001). Conclusions:Influenza A (H1N1) virus was more likely to cause severe influenza in infants and young children in comparison with seasonal H3N2 and BV. There was no significant difference in the clinical features of influenza caused by H1N1, H3N2, BV and BY. Co-infection of influenza virus with other respiratory viruses is not a major risk factor for severe influenza in infants.

Objective:To study the epidemiological features of local influenza A(H1N1)pdm09 epidemic strains through analyzing the changes in lineages and to analyze how well the vaccine strains were matched to the circulating strains in Hangzhou.Methods:Of 1 112 clinical specimens from laboratory-confirmed A(H1N1)pdm09 infections in Hangzhou in consecutive seasons from 2009 to 2020, 208 (18.7%) with high viral load (Ct value <30) were randomly selected from 10 influenza epidemics for full-length hemagglutinin gene ( HA) gene sequencing. Genetic variation, evolution and lineage changes of these representative local strains were analyzed by comparison with vaccine strains and reference strains. Results:Since the 2009 pandemic, A(H1N1)pdm09 had become one of the predominant viruses causing seasonal influenza and been reported to co-circulate with influenza A(H3N2) and influenza B viruses in Hangzhou in the past decade. It caused 10 local influenza epidemics in the 12 consecutive seasons from 2009 to 2020. HA gene sequencing revealed complex sources and rapid variation of the local A(H1N1)pdm09 strains. The main epidemic strains often genetically drifted from the recommended northern hemisphere vaccine strains due to lineage changes. Conclusions:This study suggested that it was essential to update the recommended vaccine strains year by year. Besides, enhanced periodic monitoring of influenza A(H1N1)pdm09 strains circulating in the region was important for the prevention and control of influenza A(H1N1)pdm09 infection in the next epidemic season.

Objective: In order to analyze the characteristics of the outbreak of acute respiratory tract infection in children caused by respiratory syncytial virus(RSV). Methods: The field epidemiological investigations were conducted for the two outbreaks in kindergartens in Hangzhou. Data were analyzed by descriptive method. Samples with positive respiratory syncytial virus nucleic acid were sequenced using PCR. Results: The two outbreaks occurred in kindergartens. There were 21 cases in kindergarten A, lasting 11 days, and 43 cases in kindergarten B, lasting 33 days. The epidemic curve showed a proliferation pattern. The cases were concentrated in nurseries and K1 classes, primarily among children aged 2-4 years. The most common symptoms were fever and cough, mainly upper respiratory tract infection, and no severe cases were found. Upper respiratory tract samples were collected and detected as positive for RSV. Four samples were sequenced and identified as subgroup B. Conclusion During the outbreak of acute respiratory infection in kindergartens, respiratory syncytial virus should be given primary consideration in the process of identification of the outbreak caused by other respiratory infections, and strictly control measures should be taken to reduce the long term impact of the epidemic.

Objective:To analyze the epidemiological and etiological characteristics of dengue fever in Hangzhou in 2018.Methods:RT-PCR was used to detect the nucleic acids and analyze the serotypes of dengue viruses (DENV) in serum samples collected from dengue fever cases. Phylogenetic trees based on the E gene sequences of DENV isolated from the serum samples were then constructed and analyzed. Epidemiological characteristics of these dengue fever cases were analyzed. Results:A total of 80 cases of dengue fever were detected in Hangzhou in 2018 with 55 imported cases and 25 indigenous cases (24 caused by DENV-1 and one by DENV-3). These indigenous cases mainly occurred during late July to early October with people above 50 years old accounting for 68%. Phylogenetic analysis showed that DENV-1 strains isolated from the indigenous cases in Yuhang, Jianggan-Shangcheng and Qiantang districts all belonged to genotype Ⅰ, and were respectively closely related to the strains from Indonesia in 2015, Myanmar in 2017, Ningbo in 2018 and Hangzhou imported cases from Thailand in 2018. The indigenous DENV-3 strain belonged to genotype Ⅲ, and shared 99.5% homology with the Singapore strain in 2013.Conclusions:Imported cases accounted for a large fraction of the dengue fever cases in Hangzhou, which brought a high risk to indigenous outbreak. Due to multiple imported cases, the current epidemic presented a characteristic of multiple small-scale outbreaks.

Objective:To understand the viral genomic characteristics of a 2019-novel coronavirus (2019-nCoV) strain in the first COVID-19 patient found in Hangzhou, China.Methods:Viral RNA was extracted in throat swab and sputum sample of the patient and was performed real-time reverse transcription PCR detection and obtained viral genome by high-throughput sequencing method. Phylogenetic analysis was conducted using 29 2019-nCoV genomes and 30 β-coronavirus genomes deposited in NCBI GenBank. Fifteen genomes from Wuhan were grouped by mutation sites and others were identified by Wuhan's or specific mutation sites.Results:A 29 833 bp length genome of the first 2019-nCoV strain in Hangzhou was obtained, covering full length of the coding regions of coronavirus. Phylogenetic analysis showed that the genome was closest to the genome of a bat SARS-like coronavirus strain RaTG13 with an identity of 96.11% (28 666/29 826). Among the genes between two genomes, E genes were highly conserved (99.56%), while S genes had lowest identity (92.87%). The genome sequence similarities among 29 strains from China (Hangzhou, Wuhan, and Shenzhen), Japan, USA, and Finland, were all more than 99.9%; however, some single nucleotide polymorphisms were identified in some strains.Conclusion:The genome of Hangzhou 2019-nCoV strain was very close to the genomes of strains from other cities in China and overseas collected at early epidemic phase. The 2019-nCoV genome sequencing method used in this paper provides an useful tool for monitoring variation of viral genes.