ABSTRACT
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.
ABSTRACT
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.
ABSTRACT
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.
ABSTRACT
Objective:To analyze the prevalence of influenza B virus in Hangzhou between 2014 and 2019, and the variation in hemagglutinin (HA) and neuraminidase (NA) genes.Methods:Viruses were isolated from throat swabs collected from 10 481 patients with influenza-like illness in Hangzhou from October 2014 to September 2019. The subtypes of influenza viruses were identified by real-time RT-PCR. HA and NA genes of some influenza B virus strains were amplified with specific primers and then analyzed with gene sequencing and phylogenetic analysis.Results:Influenza B virus had circulated in Hangzhou every year since 2014 and caused a much higher morbidity in people aged 5-14 years. The Victoria lineage of influenza B virus circulating in Hangzhou between 2014 and 2019 belonged to the V1A clade, while the Yamagata lineage belonged to the Y3 clade. Homology analysis showed that 124 strains shared 94.67%-100.00% homology in HA gene sequences and 128 strains shared 97.13%-100.00% homology in NA gene sequences. Influenza B virus possessed several mutations in the antigenic sites of HA and NA.Conclusions:Influenza B virus played a critical role in the influenza epidemics in Hangzhou. During 2014 to 2019, genetic mutations in the antigenic sites of HA and NA and reassortant strains were detected, and there was a mismatch between epidemic and vaccine strains. However, no drug-resistant virus was found.
ABSTRACT
Objective:To investigate the characteristics of clinical bacteria and H3N2 influenza A virus coinfection and variations in the hemagglutinin (HA) cleavage site of H3N2 among influenza-like cases.Methods:A total of 12 250 samples were collected from influenza-like cases for real-time PCR detection of H3N2 influenza virus from January 2013 to December 2018. To analyze the characteristics of co-infection, some H3N2-positive samples were selected and analyzed by real-time PCR to detect Staphylococcus aureus, Streptococcus pneumoniae and haemophilus influenzae type B. HA genes of H3N2 isolates were amplified by RT-PCR and sequenced. A phylogenetic tree was constructed based on HA gene sequences. Amino acid variations in cleavage sites were analyzed. Results:H3N2 influenza viruses had been detected every year since 2013, causing 44.69% influenza-positive cases. There were 295 randomly selected H3N2-positive samples, of which 29.2% had clinical bacterial infection. The HA cleavage sites of 210 H3N2 isolates were sequenced and 68 had variations, including 63 carrying K342R (PEKQTR to PERQTR) single-amino acid site variation. The co-infection rate was 31.25% (45/144) in unmutated samples and 23.53% (16/68) in mutated samples (χ 2=1.34, P>0.05). The H3N2 influenza viruses circulating in Hangzhou mainly belonged to two evolutionary clusters of 3c.3a and 3c.2a, and the viruses with K342R mutation at the cleavage site belonged to the evolutionary cluster of 3c.3a. Conclusions:H3N2 influenza virus played an important role in the epidemic of influenza virus in Hangzhou. There were some bacterial co-infections in influenza-positive cases. Cleavage site variations showed regional epidemic characteristics, but had no significant correlation with bacterial co-infection.
ABSTRACT
Objective To analyze the prevalence of influenza A (H3N2) virus in Hangzhou be-tween 2012 and 2017 and to investigate the genetic variations in hemagglutinin ( HA) and neuraminidase ( NA) . -ethods Throat swab samples were collected for viral isolation from 12185 patients with suspected influenza in Hangzhou area from January 2012 to December 2017. Influenza virus subtypes were identified by real-time RT-PCR. HA and NA genes of some isolated Influenza A (H3N2) viruses were amplified with spe-cific primers and then analyzed by sequencing and phylogenetic analysis. Results Influenza A (H3N2) virus was the predominant subtype circulating in Hangzhou during 2012 to 2017. It caused high morbidity in elderly people (Z=81. 039, P<0. 05). Most of the isolated influenza A (H3N2) viruses belonged to the phylogenetic clades of 3C. 3a and 3C. 2a. These viruses shared a homology of 96. 7％-100％ in nucleotide sequences of both HA and NA genes, but possessed several HA and NA mutations in antigenic sites. Con-clusions Influenza A (H3N2) virus was an important pathogen causing influenza epidemics in Hangzhou during 2012 to 2017. HA and NA genes showed many mutations in antigenic sites. No drug resistant virus was reported.
ABSTRACT
Objective@#To determine the epidemic characteristics of respiratory viruses, mycoplasma pneumonia(MP) and chlamydia pneumoniae(CP) in outpatients and hospitalized children with acute respiratory tract infections(ARI), to lay a foundation for the prevention and control of ARI.@*Methods@#From 2011 to 2013, children with ARI, including outpatients and inpatients, were involved in this study. One nasopharyngeal aspirate or throat swab specimen was collected from each patient.Real time PCRs were performed to detect common respiratory tract viruses, MP and CP.@*Results@#At least one pathogen was identified in each of 610 out of 908 patients and the overall positive rate was 67.2%. The positive rate in inpatient(76.7%)was higher than that in outpatient(43.0%) (χ2=94.79, P<0.001). Simultaneous detection of two or more viruses was found in 206 cases.Co-infection was more frequent in inpatients than in outpatients(29.0% VS 6.6%, P<0.001). Significant differences of the detection rate were observed in RSV, PIV, HRV, Flu, human bocavirus (hBoV), adenovirus (AdV), saffold virus(SAFV), MP and CP between the inpatient and outpatient group. Respiratory syncytial virus(RSV)(34.5%) was the most prevalent virus detected among hospitalized children, followed by MP(15.0%)and human rhinovirus(HRV)(14.6%). Whereas adenovirus(AdV) (15.2%)was the most frequently identified virus in the outpatient group, followed by influenza virus(Flu)(11.7%))and PIV(7.8%). Except for RSV and Flu, co-infection of the other pathogens was more frequent than its mono-infection in inpatients. Significant differences of the AdV co-infection rate were observed between the inpatient and outpatient group(χ2=18.90, P<0.001). Compared with mono-infection, co-infection has no significant effect on the clinical presentation.@*Conclusions@#The detection rate of respiratory pathogens was higher in inpatients than in outpatients with ARI, and co-infections were more popular in children hospitalized, it may show that co-infection had some correlation with disease severity.
ABSTRACT
Objective To investigate the clinical and epidemiological characteristics of Saffold virus (SAFV)infection in outpatient and hospitalized children with acute respiratory tract infections(ARI). Methods A total of 1060 clinical specimens were collected from children with ARI in the Affiliated Children's Hospital,Zhejiang University School of Medicine from March 2011 to February 2014, including 256 samples of throat swabs from outpatients,and 804 samples of trachea suctions from hospitalized patients. Real time PCR(RT-PCR)was performed to detect 5'UTR segment of SAFV.SPSS 17.0 software was used to analyze the test results and clinical data.Results The positive detection rates of SAFV in outpatients and hospitalized children with ARI were 2.3%(6/256)and 13.2%(106/804), respectively(χ2=24.147, P<0.01).Among the hospitalized children,the positive detection rates of SAFV in children <1 year,1-<3 years,3-<6 years and 6-12 years were 14.0%,11.2%,11.1% and 8.3%, respectively(χ2=1.845,P>0.05).The positive rates of SAFV in males and females were 12.7% and 17.7%(χ2=0.279,P>0.05).The detection rate of SAFV in autumn was highest(21.2%), followed by that in spring (14.6%),winter(9.5%)and summer(8.8%)(χ2=15.625, P<0.01).The co-infection rates with other respiratory pathogens of hospitalized and outpatients children were 76.4%(81/106)and 66.7% (4/6).Among the hospitalized patients, the rate of co-infection with respiratory syncytial virus was the highest(36.8%),followed by rhinovirus(27.4%), metapneumovirus(10.4%)and parainfluenza virus (10.4%).Among children with ARI,the fever rate of SAFV-positive cases was lower than that of SAFV-negative cases(χ2=4.069,P<0.05).Conclusions The detection rate of SAFV in hospitalized children with ARI is significantly higher than that in outpatients,and SAFV infection was dominated by co-infection. The prevalence of SAFV in the Hangzhou area presents a certain local epidemic pattern.
ABSTRACT
Objective To study the epidemiological characteristics and genetic evolution of human bocavirus ( HBoV ) infection in hospitalized children with severe acute respiratory infection ( SARI ) in Hangzhou.Methods A total of 1388 clinical specimens were collected from children with SARI admitted in Affiliated Children' s Hospital, Zhejiang University School of Medicine from January 2011 to December 2014.HBoV1-4 and other respiratory pathogens were identified by fluorescent real -time polymerase chain reaction (fRT-PCR).The VP1 gene in HBoV1 positive samples was amplified and sequenced for genetic analysis with Clustal X and MEGA 6.0.Chi-square test and Fisher exact probability were used to analyze the data.Results Eighty five HBoV positive samples were detected from 1388 samples (6.12％), among which 83 (97.65％) were HBoV1 positive and 2 (2.35％) were HBoV2 positive.The positive rates of HBoV in males and females were 6.54％and 5.35％(χ2 =0.780, P>0.05).The posititve detection rate of HBoV in all age groups was statistically significant (χ2 =47.446,P <0.01).The detection rate in children aged 6 months-1 year was highest (12.84％), in children aged >3 years was lowest (1.64％), in children aged ≤6 months and aged 1-3 years was 3.04％ and 3.33％, respectively.The detection rate of HBoV in summer was the highest (14.97％), followed by that in autumn (7.14％), spring (3.19％) and winter (1.97％) (χ2 =58.807, P<0.01).The detection rates of HBoV in 2011 to 2014 were 7.39％, 7.31％, 5.58％ and 4.72％ (χ2 =3.447, P >0.05 ).The co-infection rate with other respiratory pathogens was 62.35％.The main pathogens were human rhinovirus (33.96％), parainfluenza virus (28.30％) and respiratory syncytial virus (20.75％).The incidence of anhelation and wheezing in HBoV positive group was higher than that in HBoV negative group (χ2 =15.161 and 13.914, P <0.01). Sequence analysis of VP 1 gene showed that 44 isolates belonged to the same branch ( clade 1 ) as Swedish strain ST2, and 2 isolates HZ12-S32 and HZ12-S199 belonged to a separated branch.Conclusion HBoV is an important causative agent of hospitalized children with SARI in Hangzhou area and has high co -infection with other respiratory pathogens.Most of the strains belong to the same clade as the Swedish strain ST 2, and two strains of HZ12-S32 and HZ12-S199 are identified in a separated clade.
ABSTRACT
Objective To detect adenovirus from children with acute upper/lower respiratory tract infections and to investigate the genetic evolution of virus .Methods A total of 1 178 clinical specimens were collected from the Children ’ s Hospital , Zhejiang University School of Medicine during March 2011 and February 2013, including 513 throat swabs from children with acute upper respiratory tract infection and 665 nasopharyngeal aspirates from children with acute lower respiratory tract infections .Besides, 9 specimens in an outbreak of adenovirus infection during 2011 and 2014 were also collected .Adenovirus was identified by real-time fluorescent polymerase chain reaction (RT-qPCR).The hypervariable region (HVR)-7 region of hexon gene in positive samples was amplified and sequenced for typing and phylogenetic analysis .Other respiratory viruses were also detected with RT-qPCR in adenovirus positive samples .Clinical characteristics of adenovirus infection were analyzed in children with lower respiratory tract infections .Chi-square test and Fisher exact probability were used for data analysis .Results Among 1 178 samples from sporadic cases , 104 samples (8.83%) were adenovirus positive .The rates of adenovirus infection in upper respiratory tract infection group and lower respiratory tract infection group were 13.65%(70/513) and 5.11%(34/665), respectively (χ2 =26.193, P3 years (χ2 =6.575 and 7.334, P0.05), but two children with co-infection died.Conclusions Adenovirus infection is more common in upper respiratory tract infection .Adenovirus type 3 and type 7 are the most prevalent serotypes in sporadic cases , while type 3 is the most prevalent serotype in outbreak cases .
ABSTRACT
We investigated the genetic diversity and evolution of the M gene of human influenza A viruses in Hangzhou (Zhejiang province, China) from 2009 to 2013, including subtypes of A(H1N1) pdm09 strains and seasonal A(H3N2) strains. Subtypes of analyzed viruses were identified by cell culture and real-time reverse transcription-polymerase chain reaction, followed by cloning, sequencing and phylogenetic analyses of the M gene. Assessment of 5675 throat swabs revealed a positive rate for the influenza virus of 20.46%, and 827 cases were diagnosed as. infections due to influenza A viruses. Seventy-six influenza-A strains were selected randomly from nine stages during six phases of a virus epidemic. Sequences of the M gene showed high homology among six epidemics with identities of amino-acid sequences of 98.98-100%. All strains contained the adamantine-resistant mutation S31N in its M2 protein. Two of the A(H1N1)pdm09 strains had double mutants of V27A/S31N or V271/S31N. One of the seasonal A(H3N2) viruses had another form of double-mutant R45H/S31N. Evolutionary rate of the M gene was much lower than that of the HA gene and NA gene. Compared with A(H3N2) strains, higher positive pressure on the M1 and M2 proteins of A(H1N1) pdm09 viruses was observed. Separate analyses of M1 and M2 proteins revealed very different selection pressures. Knowledge of the genetic diversity and evolution of the M gene of human influenza-A viruses will be valuable for the control and prevention of diseases.
Subject(s)
Humans , Amino Acid Substitution , China , Epidemiology , Evolution, Molecular , Genetic Variation , Influenza A Virus, H1N1 Subtype , Classification , Genetics , Influenza A Virus, H3N2 Subtype , Classification , Genetics , Influenza, Human , Epidemiology , Virology , Phylogeny , Selection, Genetic , Viral Matrix Proteins , Genetics , Viral Proteins , Chemistry , GeneticsABSTRACT
Objective To investigate the genetic variation and molecular evolution of human bo-cavirus 1 (HBoV1) strains isolated during 2009 to 2014 in Hangzhou, China.Methods Throat swab sam-ples were collected from children with acute respiratory tract infections in the Children′s Hospital Affiliated to the Zhejiang University School of Medicine from 2009 to 2014.Real-time PCR was performed for the detec-tion of HBoV1 strains.Fifteen HBoV1 strains with high virus load were screened out for the amplification and sequencing of complete genomes.The complete genomes were submitted to GenBank for further analysis with bioinformatics software.Results A total of 48 nucleotide mutations were detected in the complete genomes of 15 HBoV1 strains, resulting in 11 amino acid mutations with 5 of them located in the active region of phospholipase A2 ( PLA2) .The 15 HBoV1 isolates along with 16 HBoV1 strains in GenBank were classified into three clusters as indicated by the phylogenetic analysis based on their complete coding sequences.All of the 15 strains were belonged to clusterⅠ, the representative strain of which was the Sweden prototype strain ST2.The phylogenetic trees constructed using genes encoding the capsid proteins VP1 and VP2 were highly similar to those based on the complete coding sequences.The estimated mean evolutionary rate of HBoV1 with regard to the complete coding sequence was 3.03×10-4(95%HPD, 2.14×10-4-3.92×10-4 ) substitu-tions per site per year.With regard to each gene, the NS1 gene was considered to the most conserved gene while the NP1 gene showed the highest substitution rate.The dN/dS ratios (ω) of the four genes were all less than 1, indicating that all of them were under negative selection.Moreover, the VP2 gene was under the strongest negative selection, while the NP1 gene was under the weakest negative selection.Conclusion All of the HBoV1 isolates circulating in Hangzhou province during 2009 to 2014 were belonged to ST2 genotype with a relatively high mutation in the area of PLA2.Despite the complete genome was conservative, its evo-lutionary rate was high.Among the four genes, the NP1 gene showed the highest substitution rate.All of the four genes were under negative selection, of which the VP2 gene was under the strongest negative selection.
ABSTRACT
<p><b>OBJECTIVE</b>To understand the molecular epidemiologic features of human metapnenmovirus (hMPV) in children with respiratory tract infection in Hangzhou.</p><p><b>METHODS</b>2 593 throat swabs were collected from patients with respiratory tract infections who visited the hospitals with sentinel surveillance programs from January 2011 to December 2013, including 1 676 outpatients and 917 inpatients. Total nucleic acid was extracted from the specimens and the fusion (F) protein gene of hMPV was amplified by quantitative real-time reverse transcription polymerase chain reaction (RT-PCR), with positive samples picked to compare with the sequence of hMPV in GenBank, after the sequence of amplification products were determined. Other two types of common respiratory virus were tested using RT-PCR.</p><p><b>RESULTS</b>The overall positive rate in this study was 6.51% (169/2 593), with 6.62% (111/1 676) in outpatients and 6.32% (58/917) in inpatients, but no statistically significant difference was found (χ(2) = 0.086, P = 0.769). The rates was 7.01% in males and 5.72% in females, with no statistically significant difference in different sex (χ(2) = 1.676, P = 0.195). The positive rate was 14.14% (28/198)in the 2-year-olds, 14.01% (22/158)in 3-year olds. The rate in 2-year olds was higher than in other groups, with statistically significant differences between the groups (χ(2) = 38.654, P = 0.000). Of the 169 positive cases, 153 (90.53%) in the younger than 5 years olds. The rates of infection with hMPV in winter and spring were statistically higher than in summer and autumn (χ(2) = 67.032, P = 0.000). The rate of co-infection was 19.52% (33/169). 88 amplified productions were selected for gene sequence analysis, and the F gene homology were 81.6%-100.0% with reference strains in GenBank. Data showed that all the 4 viral subtypes: A2 (52.27% , 46/88), B1 (37.51%, 33/88), B2 (9.09%, 8/88) and A1 (1.13%, 1/88) co-circulated during the study. However, different subtypes appeared predominant in different years:hMPV subtype B1 was in 2011 and 2012, subtype A2 in the end of 2012 and in 2013. Of the 88 specimens, gene sequences were determinate, with A genotype accounted for 67.56% (25/37), B genotype for 32.43% (12/37)in children younger than 1-year olds, and A genotype accounted for 43.13% (22/51), B genotype for 56.86% (29/51)in children above 1-year olds. Significant differences between the two groups (χ(2) = 5.143, P = 0.023) were noticed.</p><p><b>CONCLUSION</b>It was confirmed that hMPV was one of the substantial pathogens causing the respiratory tract infections. Data from our study suggested that the peak time of hMPV infection predominated during winter and spring in Hangzhou. Both hMPV subtype B1 and subtype A2 were found popular in this study, with hMPV genotype A dominating in children younger than 1-year olds.</p>
Subject(s)
Child , Child, Preschool , Female , Humans , Infant , Male , China , Epidemiology , Coinfection , Genotype , Metapneumovirus , Genetics , Virulence , Molecular Epidemiology , Paramyxoviridae Infections , Epidemiology , Real-Time Polymerase Chain Reaction , Respiratory Tract Infections , Epidemiology , Seasons , Sentinel SurveillanceABSTRACT
Objective To investigate the epidemiological characters of human metapneumovirus (hMPV) infection in children with influenza-like illness (ILI).Methods A total of 1 164 throat swabs were collected from children with ILI symptoms in Children's Hospital,Zhejiang University School of Medicine from January 2011 to December 2012.hMPV was detected by using nucleic acid assay,the fusion (F) protein gene of hMPV was amplified by RT-PCR,gene sequencing was performed and the sequences were compared with those in GenBank.Positive rates of hMPV in different age groups were compared with Chi-square test.Results Among 1 164 samples,hMPV was positive in 73 (6.27％) samples.hMPV infection was the most popular in ＞2-4 y age group (33/220,15.0％),and the positive rates of hMPV in different age groups were of statistical significance (x2 =40.69,P ＜ 0.05).hMPV infection occurred throughout the year,but it was most common in winter and spring.The highest incidence of hMPV infection was observed in December 2012 (12/51,25.53％).Among 24 samples of hMPV,14 were with genotype B1,2 were with genotype B2,and 8 were with genotype A2.The most common genotype was B1 in 2011 (10/12),and A2 in 2012 (8/12).Homology between nucleotide sequences of the 24 samples of hMPV were 81.6％ to 100.0％.Conclusions hMPV infection exists in children with ILI in Hangzhou,and the epidemic seasons are winter and spring.hMPV infection is more likely to be found in children aged 2 to 4 years old,and different genotypes may predominated alternately.