Objective: To analyze the spatial and temporal distribution characteristics of seasonal A(H3N2) influenza [influenza A(H3N2)] in China and to provide a reference for scientific prevention and control. Methods: The influenza A(H3N2) surveillance data in 2014-2019 was derived from China Influenza Surveillance Information System. A line chart described the epidemic trend analyzed and plotted. Spatial autocorrelation analysis was conducted using ArcGIS 10.7, and spatiotemporal scanning analysis was conducted using SaTScan 10.1. Results: A total of 2 603 209 influenza-like case sample specimens were detected from March 31, 2014, to March 31, 2019, and the influenza A(H3N2) positive rate was 5.96%(155 259/2 603 209). The positive rate of influenza A(H3N2) was statistically significant in the north and southern provinces in each surveillance year (all P<0.05). The high incidence seasons of influenza A (H3N2) were in winter in northern provinces and summer or winter in southern provinces. Influenza A (H3N2) clustered in 31 provinces in 2014-2015 and 2016-2017. High-high clusters were distributed in eight provinces, including Beijing, Tianjin, Hebei, Shandong, Shanxi, Henan, Shaanxi, and Ningxia Hui Autonomous Region in 2014-2015, and high-high clusters were distributed in five provinces including Shanxi, Shandong, Henan, Anhui, and Shanghai in 2016-2017. Spatiotemporal scanning analysis from 2014 to 2019 showed that Shandong and its surrounding twelve provinces clustered from November 2016 to February 2017 (RR=3.59, LLR=9 875.74, P<0.001). Conclusion: Influenza A (H3N2) has high incidence seasons with northern provinces in winter and southern provinces in summer or winter and obvious spatial and temporal clustering characteristics in China from 2014-2019.
Humans ; Influenza, Human/epidemiology* ; China/epidemiology* ; Influenza A Virus, H3N2 Subtype ; Seasons ; Cluster Analysis

BACKGROUND: The new emerging avian influenza A H7N9 virus, causing severe human infection with a mortality rate of around 41%. This study aims to provide a novel treatment option for the prevention and control of H7N9. METHODS: H7 hemagglutinin (HA)-specific B cells were isolated from peripheral blood plasma cells of the patients previously infected by H7N9 in Jiangsu Province, China. The human monoclonal antibodies (mAbs) were generated by amplification and cloning of these HA-specific B cells. First, all human mAbs were screened for binding activity by enzyme-linked immunosorbent assay. Then, those mAbs, exhibiting potent affinity to recognize H7 HAs were further evaluated by hemagglutination-inhibiting (HAI) and microneutralization in vitro assays. Finally, the lead mAb candidate was selected and tested against the lethal challenge of the H7N9 virus using murine models. RESULTS: The mAb 6-137 was able to recognize a panel of H7 HAs with high affinity but not HA of other subtypes, including H1N1 and H3N2. The mAb 6-137 can efficiently inhibit the HA activity in the inactivated H7N9 virus and neutralize 100 tissue culture infectious dose 50 (TCID50) of H7N9 virus (influenza A/Nanjing/1/2013) in vitro, with neutralizing activity as low as 78 ng/mL. In addition, the mAb 6-137 protected the mice against the lethal challenge of H7N9 prophylactically and therapeutically. CONCLUSION The mAb 6-137 could be an effective antibody as a prophylactic or therapeutic biological treatment for the H7N9 exposure or infection.
Animals ; Antibodies, Monoclonal/therapeutic use* ; Antibodies, Neutralizing/therapeutic use* ; Antibodies, Viral ; Hemagglutinins ; Humans ; Influenza A Virus, H1N1 Subtype ; Influenza A Virus, H3N2 Subtype ; Influenza A Virus, H7N9 Subtype ; Influenza Vaccines ; Influenza in Birds ; Influenza, Human/prevention & control* ; Mice

Objective: To investigate the distribution characteristics of respiratory non-bacterial pathogens in children in Ningbo from 2019 to 2021. Methods: A retrospective analysis was performed on 23 733 children with respiratory tract infection who visited the department of pediatrics of Ningbo Women and Children's Hospital from July 2019 to December 2021. There were 13 509 males (56.92%) and 10 224 females (43.08%), with an age range of 1 day to 18 years old. There were 981 cases in the neonatal group (younger than 1 month old), 5 880 cases in the infant group (1 month to younger than 1 year old), 6 552 cases in the toddler group (1 to younger than 3 years old), 7 638 cases in the preschool group (3 to younger than 7 years old), and 2 682 cases in the school-age group (7 to 18 years old). Thirteen respiratory pathogens were detected by multiple polymerase chain reaction (PCR) based on capillary electrophoresis, and SPSS 23.0 software was used for statistical analysis of the results, the count data were expressed as percentages, and the χ² test was used for comparison between groups. Results: Of the 23 733 specimens, 13 330 were positive for respiratory pathogens, with a total positive rate of 56.17%. The positive rates of human rhinovirus (HRV) 24.05% (5 707/23 733), human respiratory syncytial virus (HRSV) 10.45% (2 480/2 3733) and mycoplasma pneumoniae (Mp) 7.03% (1 668/23 733) were in the first three. The positive rates of pathogens in the male and female children were 57.47% (7 763/13 509) and 54.45% (5 567/10 224), respectively, and the difference was statistically significant (χ²=21.488, P<0.001). The positive rates in the neonatal group, infant group, toddler group, preschool group, and school-age group were 31.80% (312/981), 54.71% (3 217/5 880), 63.23% (4 143/6 552), 59.83% (4 570/7 638), 40.57% (1 088/2 682), respectively, and the difference among the groups was statistically significant (χ²=681.225, P<0.001). The single infection rate was 47.43% (11 256/23 733), the mixed infection rate of two or more pathogens was 8.74% (2 074/23 733), most of which were mixed infections of two pathogens. HRV, HADV, HCOV, Ch disseminated in the whole year. HRSV, HMPV, Boca, HPIV occurred mostly in fall and winter. The positive rates of FluA, FluB, Mp were at a low level after the corona virus disease 2019 (COVID-19) epidemic (2020 and 2021). The positive rates of FluA, H1N1, H3N2, FluB, HADV, Mp in 2020 were significantly lower than in 2019 (P<0.05). The positive rates of HPIV, HRV, HCOV, Ch in 2020 were significantly higher than in 2019 (P<0.05). The positive rates of FluA, H1N1, H3N2, HPIV, HCOV, Mp, Ch in 2021 were significantly lower than in 2020 (P<0.05). The positive rates of Boca, HMPV, HRSV in 2021 were significantly higher than in 2020 (P<0.05). Conclusion: From 2019 to 2021, the main non-bacterial respiratory pathogens of children in Ningbo City were Mp and HRV, and the detection rates of respiratory pathogens varied among different ages, seasons and genders.
Infant ; Infant, Newborn ; Child ; Child, Preschool ; Humans ; Male ; Female ; Adolescent ; Influenza A Virus, H1N1 Subtype ; Influenza A Virus, H3N2 Subtype ; Retrospective Studies ; COVID-19 ; Respiratory Tract Infections/epidemiology* ; Respiratory Syncytial Virus, Human ; Mycoplasma pneumoniae ; Coinfection

OBJECTIVE: To evaluate the effect of intranasal immunization with CTA1-DD as mucosal adjuvant combined with H3N2 split vaccine. METHODS: Mice were immunized intranasally with PBS (negative control), or H3N2 split vaccine (3 μg/mouse) alone, or CTA1-DD (5 μg/mouse) alone, or H3N2 split vaccine (3 μg/mouse) plus CTA1-DD (5 μg/mouse). Positive control mice were immunized intramuscularly with H3N2 split vaccine (3 μg/mouse) and alum adjuvant. All the mice were immunized twice, two weeks apart. Then sera and mucosal lavages were collected. The specific HI titers, IgM, IgG, IgA, and IgG subtypes were examined by ELISA. IFN-γ and IL-4 were test by ELISpot. In addition, two weeks after the last immunization, surivival after H3N2 virus lethal challenge was measured. RESULTS: H3N2 split vaccine formulated with CTA1-DD could elicit higher IgM, IgG and hemagglutination inhibition titers in sera. Furthermore, using CTA1-DD as adjuvant significantly improved mucosal secretory IgA titers in bronchoalveolar lavages and vaginal lavages. Meanwhile this mucosal adjuvant could enhance Th-1-type responses and induce protective hemagglutination inhibition titers. Notably, the addition of CTA1-DD to split vaccine provided 100% protection against lethal infection by the H3N2 virus. CONCLUSION CTA1-DD could promote mucosal, humoral and cell-mediated immune responses, which supports the further development of CTA1-DD as a mucosal adjuvant for mucosal vaccines.
Adjuvants, Immunologic ; Administration, Intranasal ; Animals ; Cholera Toxin ; Female ; Immunity, Humoral ; Influenza A Virus, H3N2 Subtype ; immunology ; Influenza Vaccines ; Mice, Inbred BALB C ; Nasal Mucosa ; immunology ; Random Allocation ; Recombinant Fusion Proteins

Objective: To understand the clinical characteristics and economic burden of influenza-like illness (ILI) children aged 0-59 months in the outpatient settings in Suzhou, China, 2011-2017. Methods: From March 2011 to February 2017, we conducted a prospective surveillance program on ILI for children aged less than 5 years at Soochow University Affiliated Children's Hospital. Through standard questionnaires and follow-up survey via telephone, we collected information regarding the demographic characteristics, medical history, clinical symptoms and both direct and indirect costs associated with influenza, of the patients. We then compared clinical characteristics and economic burden of influenza A/H1N1, A/H3N2, and B infections among children with ILI. Results: We enrolled 6 310 patients with ILI from March 2011 to February 2017 and collected all their throat swabs. 791 (12.9%) of the swabs showed positive for influenza virus, including 88 (11.1%) subtype influenza A/H1N1, 288 (36.4%) subtype influenza A/H3N2, and 415(52.5%) type influenza B. The proportions of cough, rhinorrhea, wheezing, vomiting and convulsion in influenza-positive children were higher than those influenza-negative children. Except for the prevalence rates of cough (χ(2)=9.227, P=0.010), wheezing (χ(2)=7.273, P=0.026) and vomiting (χ(2)=8.163, P=0.017), other clinical symptoms appeared similar between the three viral subtypes. Among all the ILI children, the average total cost per episode of influenza was 688.4 Yuan (95%CI: 630.1-746.7) for influenza-negative children; 768.0 Yuan (95%CI: 686.8-849.3) for influenza-positive children and 738.3 Yuan (95%CI: 655.5-821.1) for influenza B. Children with influenza A/H1N1 spent much more than those with influenza A/H3N2 or influenza B in the total cost (χ(2)=7.237, P=0.028). Conclusion: Children infected influenza showed higher prevalence rates of cough, rhinorrhea, wheezing, vomiting and convulsion than those without influenza. Influenza A/H1N1 subtype caused heavier economic burden than the other two influenza subtypes.
Ambulatory Care/statistics & numerical data* ; Child ; Child, Preschool ; China/epidemiology* ; Cost of Illness ; Cough/virology* ; Female ; Fever/virology* ; Humans ; Infant ; Infant, Newborn ; Influenza A Virus, H1N1 Subtype ; Influenza A Virus, H3N2 Subtype ; Influenza, Human/epidemiology* ; Outpatient Clinics, Hospital/statistics & numerical data* ; Outpatients/statistics & numerical data* ; Prospective Studies ; Socioeconomic Factors ; Surveys and Questionnaires ; Virus Diseases

Swine influenza viruses (SwIVs) cause considerable morbidity and mortality in domestic pigs, resulting in a significant economic burden. Moreover, pigs have been considered to be a possible mixing vessel in which novel strains loom. Here, we developed and evaluated a novel M2e-multiple antigenic peptide (M2e-MAP) as a supplemental antigen for inactivated H3N2 vaccine to provide cross-protection against two main subtypes of SwIVs, H1N1 and H3N2. The novel tetra-branched MAP was constructed by fusing four copies of M2e to one copy of foreign T helper cell epitopes. A high-yield reassortant H3N2 virus was generated by plasmid based reverse genetics. The efficacy of the novel H3N2 inactivated vaccines with or without M2e-MAP supplementation was evaluated in a mouse model. M2e-MAP conjugated vaccine induced strong antibody responses in mice. Complete protection against the heterologous swine H1N1 virus was observed in mice vaccinated with M2e-MAP combined vaccine. Moreover, this novel peptide confers protection against lethal challenge of A/Puerto Rico/8/34 (H1N1). Taken together, our results suggest the combined immunization of reassortant inactivated H3N2 vaccine and the novel M2e-MAP provided cross-protection against swine and human viruses and may serve as a promising approach for influenza vaccine development.
Animals ; Antibodies, Viral/blood ; Antigens, Viral/genetics/*immunology ; Body Weight ; Cross Protection/*immunology ; Disease Models, Animal ; Epitopes, T-Lymphocyte/genetics/immunology ; Female ; Influenza A Virus, H3N2 Subtype/genetics/*immunology ; Influenza Vaccines/*immunology ; Mice ; Mice, Inbred BALB C ; Orthomyxoviridae Infections/*immunology/mortality/pathology/prevention & control ; Peptides/genetics/*immunology ; Random Allocation ; Survival Analysis ; Vaccines, Synthetic/immunology ; Virus Replication

Influenza A virus is an enveloped virus that belongs to the Orthomyxoviridae family. It has 8 negative RNA segments that encode 16 viral proteins. The viral polymerase consists of 3 proteins (PB 1, PB2 and PA) which plays an important role in the transcription and replication of the influenza A virus. Polymerase basic protein 1 (PB 1) is a critical member of viral polymerase complex. In order to further study the function of PB1, we need to prepare the PB1 antibody with good quality. Therefore, we amplified PB1 conserved region (nt1648-2265) by PCR and cloned it into pET-30a vector, and transformed into Escherichia coli BL2 1. The expression of His tagged PB 1 protein was induced by IPTG, and His-PB 1 proteins were purified by Ni-NTA resin. For preparation of PB 1 protein antiserum, rabbits were immunized with His-PB 1 fusion protein 3 times. Then the titer of PB 1 polyclonal antibody was measured by indirect ELISA. The antibody was purified by membrane affinity purification and subjected to immunoblotting analysis. Data showed that PB1 antibody can recognize PB 1 protein from WSN virus infected or pCMV FLAG-PB 1 transfected cells. Meanwhile, PB 1 antibody can also recognize specifically other subtype strains of influenza A virus such as H9N2 and H3N2. PB 1 polyclonal antibody we generated will be a useful tool to study the biological function of PB1.
Animals ; Antibodies, Viral ; biosynthesis ; Cloning, Molecular ; Enzyme-Linked Immunosorbent Assay ; Escherichia coli ; metabolism ; Genetic Vectors ; Influenza A Virus, H3N2 Subtype ; Influenza A Virus, H9N2 Subtype ; Plasmids ; Rabbits ; Viral Proteins ; immunology

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.
Amino Acid Substitution ; China ; epidemiology ; Evolution, Molecular ; Genetic Variation ; Humans ; Influenza A Virus, H1N1 Subtype ; classification ; genetics ; isolation & purification ; Influenza A Virus, H3N2 Subtype ; classification ; genetics ; isolation & purification ; Influenza, Human ; epidemiology ; virology ; Phylogeny ; Selection, Genetic ; Viral Matrix Proteins ; genetics ; Viral Proteins ; chemistry ; genetics

To analyze the antigenic and genetic characteristics of the influenza A (H3N2) virus in mainland China during the surveillance year of 2013-2014, the antigenic characteristics of H3N2 virus were analyzed using reference ferret anti-sera. The nucleotide sequences of the viruses were determined by Sanger dideoxy sequencing, phylogenetic trees were constructed with the neighbor-joining method, and the genetic characteristics of the viruses were determined in comparison to current vaccine strains. The results showed that most of the H3N2 viruses were antigenically closely related to the A/Victoria/361/2011 vaccine strain cell-propagated prototype virus (99.6%). Using the A/Texas/50/2012 egg isolate as the reference antigen, 15.1% of the viruses were found to be closely antigenically related to it, while 11.9% of strains were closely antigenically related to the egg-propagated epidemic strain, A/Shanghai-Changning/1507/2012. Phylogenetic analysis of HA genes indicated that the A(H3N2) viruses in this surveillance year were in the same clade, but no drug resistant mutation was identified in the NA genes. During the 2013-2014 influenza surveillance year, no significant genetic change was detected in either the HA or NA genes of the A(H3N2) viruses, while significant mutations were found in egg isolates resulting from their adaptation during propagation in eggs. The antigenic and genetic changes should be investigated in a timely manner to enable the selection of an appropriate vaccine strain in China.
Animals ; Antigenic Variation ; Base Sequence ; Chick Embryo ; China ; Genetic Variation ; Hemagglutinin Glycoproteins, Influenza Virus ; genetics ; immunology ; Humans ; Influenza A Virus, H3N2 Subtype ; genetics ; immunology ; isolation & purification ; Influenza, Human ; virology ; Molecular Sequence Data ; Mutation ; Phylogeny

OBJECTIVETo analyze the susceptibility of influenza A (H3N2) viruses to neuraminidase inhibitors during 2011-2012 in Mainland China.

METHODSAll the tested viruses were obtained from the Chinese National Influenza Surveillance Network, which covers 31 provinces in mainland China, including 408 network laboratories and 554 sentinel hospitals. In total 1 903 viruses were selected with isolation date from January 1, 2011 to December 31, 2012 in Mainland China, among these viruses, 721 were confirmed to be influenza A (H3N2) virus by Chinese National Influenza Center and tested for the susceptibility to oseltamivir and zanamivir using chemiluminescence-based assay. The neuraminidase inhibitor sensitive reference virus A/Washington/01/2007 (119E) and oseltamivir resistant virus A/Texas/12/2007 (E119V) were used as control in this study. The t -test was used to compare the difference of NAI susceptibility of viruses isolated from different years.

RESULTSThe half maximal inhibitory concentration (IC₅₀) of A/Washington/01/2007 for oseltamivir and zanamivir was (0.10 ± 0.02) and (0.30 ± 0.05) nmol/L, respectively. The IC₅₀ of A/Texas/12/2007 for oseltamivir and zanamivir was (4.27 ± 1.60) and (0.20 ± 0.03) nmol/L, respectively. Among the 721 influenza A (H3N2) viruses, 132 influenza A (H3N2) viruses were isolated in 2011 and 589 influenza A (H3N2) viruses were isolated in 2012. The IC50 for oseltamivir ranged from 0.04 to 0.62 nmol/L for viruses isolated in 2011 and ranged from 0.02 to 0.95 nmol/L for viruses in 2012, and the IC₅₀ of all the viruses tested was within 10-fold IC₅₀ (1.0 nmol/L) of the neuraminidase inhibitor sensitive reference virus A/Washington/01/2007. The IC50 of zanamivir ranged from 0.12 to 0.80 nmol/L for viruses in 2011 and ranged from 0.04 to 0.72 nmol/L for viruses in 2012, and was within 10-fold IC₅₀ (3.0 nmol/L) of the neuraminidase inhibitor sensitive reference virus A/Washington/01/2007.

CONCLUSIONThe influenza A(H3N2) viruses isolated during 2011-2012 in Mainland China were tested to be sensitive to oseltamivir and zanamivir.

Antiviral Agents ; China ; Drug Resistance, Viral ; Enzyme Inhibitors ; Epidemiological Monitoring ; Humans ; Influenza A Virus, H3N2 Subtype ; Influenza, Human ; Neuraminidase ; Oseltamivir ; Zanamivir