Objective: To understand the epidemiological characteristics of human infection with avian influenza A (H7N9) virus in China, and provide evidence for the prevention and control of human infection with H7N9 virus. Methods: The published incidence data of human infection with H7N9 virus in China from March 2013 to April 2017 were collected. Excel 2007 software was used to perform the analysis. The characteristics of distribution of the disease, exposure history, cluster of the disease were described. Results: By the end of April 2017, a total of 1 416 cases of human infection with H7N9 virus were confirmed in China, including 559 deaths, the case fatality rate was 39.5%. In 2016, the case number was lowest (127 cases), with the highest fatality rate (57.5%). The first three provinces with high case numbers were Zhejiang, Guangdong and Jiangsu. The median age of the cases was 55 years and the male to female ratio was 2.3∶1. Up to 66% of cases had clear live poultry exposure history before illness onset, 31% of cases had unknown exposure history and only 3% of the cases had no live poultry exposure history. There were 35 household clusters (5 in 2013, 9 in 2014, 6 in 2015, 5 in 2016, 10 in 2017), which involved 72 cases, accounting for 5% of the total cases. Conclusions: The epidemic of human infection with H7N9 virus in China during 2013-2017 had obvious seasonality and spatial distribution. There was limited family clustering. Infection cases were mostly related to poultry contact.
Adult ; Age Distribution ; Aged ; Aged, 80 and over ; Animals ; China/epidemiology* ; Cluster Analysis ; Disease Outbreaks ; Epidemics ; Female ; Humans ; Incidence ; Influenza A Virus, H7N9 Subtype/isolation & purification* ; Influenza, Human/virology* ; Male ; Middle Aged ; Poultry ; Sex Distribution

Objective: To analyze the reasons for the fluctuations in the percentage of outpatient or emergency visits for influenza-like illness (ILI) during the Spring Festival and National Day in 2014-2018 surveillance season. Methods: ILI surveillance data was collected during the period of Spring Festival and National Day in mainland China, and downloaded from Chinese Influenza Surveillance Information System, during the 2014-2018 surveillance season. Results: There was no significant difference noticed in the number of ILI reports in the festival week with weeks before or after in both the southern and northern provinces. The number of outpatient visits was much less than that of the week before and after, but the number of emergency visits was statistically significantly increased. Conclusion: In the holiday peak of ILI%, the major causes was the impact of holiday-off at sentinel hospitals, resulting in a large reduction in the number of outpatient visits in the consulting room during the festivals.
Adolescent ; Adult ; Biometry ; Child ; Child, Preschool ; China/epidemiology* ; Disease Outbreaks/prevention & control* ; Emergency Service, Hospital/statistics & numerical data* ; Holidays ; Hospitals ; Humans ; Influenza, Human/virology* ; Outpatients/statistics & numerical data* ; Population Surveillance ; Respiratory Syncytial Virus Infections/virology* ; Respiratory Syncytial Virus, Human/isolation & purification* ; Respiratory Tract Infections/virology* ; Seasons ; Young Adult

Objective: To understand the molecular characteristics of hemagglutinin (HA) and neuraminidase (NA) as well as the disease risk of influenza virus A H7N9 in Guizhou province. Methods: RNAs were extracted and sequenced from HA and NA genes of H7N9 virus strains obtained from 18 cases of human infection with H7N9 virus and 6 environmental swabs in Guizhou province during 2014-2017. Then the variation and the genetic evolution of the virus were analyzed by using a series of bioinformatics software package. Results: Homology analysis of HA and NA genes revealed that 2 strains detected during 2014-2015 shared 98.8%-99.2% and 99.2% similarities with vaccine strains A/Shanghai/2/2013 and A/Anhui/1/2013 recommended by WHO, respectively. Two strains detected in 2016 and 14 strains detected in 2017 shared 98.2%-99.3% and 97.6%-98.8% similarities with vaccine strain A/Hunan/02650/2016, respectively. Other 6 stains detected in 2017 shared 99.1%-99.4% and 98.9%-99.3% similarities with strain A/Guangdong/17SF003/2016, respectively. Phylogenetic analysis showed that all the strains were directly evolved in the Yangtze River Delta evolution branch, but they were derived from different small branch. PEVPKRKRTAR↓GLF was found in 6 of 24 strains cleavage site sequences of HA protein, indicating the characteristic of highly pathogenic avian influenza virus. Mutations A134V, G186V and Q226L at the receptor binding sites were found in the HA. All the strains had a stalk deletion of 5 amino acid residue "QISNT" in NA protein, and drug resistance mutation R294K occurred in strain A/Guizhou-Danzhai/18980/2017. In addition, potential glycosylation motifs mutations NCS42NCT were found in the NA of 9 of 24 strains. Conclusions: HA and NA genes of avian influenza A (H7N9) virus showed genetic divergence in Guizhou province during 2014-2017. The mutations of key sites might enhance the virulence of the virus, human beings are more susceptible to it. Hence, the risk of infection is increasing.
Animals ; Base Sequence ; Birds ; China/epidemiology* ; Genome, Viral ; Hemagglutinin Glycoproteins, Influenza Virus/immunology* ; Hemagglutinins/genetics* ; Humans ; Influenza A Virus, H7N9 Subtype/isolation & purification* ; Influenza in Birds ; Influenza, Human/virology* ; Neuraminidase/genetics* ; Phylogeny ; RNA, Viral/genetics* ; Sequence Analysis, DNA

The highly pathogenic avian influenza (HPAI) H5N1 virus has caused several outbreaks in domestic poultry. Despite great efforts to control the spread of this virus, it continues to evolve and poses a substantial threat to public health because of a high mortality rate. In this study, we sequenced whole genomes of eight H5N1 avian influenza viruses isolated from domestic poultry in eastern China and compared them with those of typical influenza virus strains. Phylogenetic analyses showed that all eight genomes belonged to clade 2.3.2.1 and clade 7.2, the two main circulating clades in China. Viruses that clustered in clade 2.3.2.1 shared a high degree of homology with H5N1 isolates located in eastern Asian. Isolates that clustered in clade 7.2 were found to circulate throughout China, with an east-to-west density gradient. Pathogenicity studies in mice showed that these isolates replicate in the lungs, and clade 2.3.2.1 viruses exhibit a notably higher degree of virulence compared to clade 7.2 viruses. Our results contribute to the elucidation of the biological characterization and pathogenicity of HPAI H5N1 viruses.
Animals ; China ; Influenza A Virus, H5N1 Subtype ; genetics ; isolation & purification ; pathogenicity ; Influenza in Birds ; virology ; Mice, Inbred BALB C ; Phylogeny ; Poultry

OBJECTIVETo provide a feasible and cost-effective next-generation sequencing (NGS) method for accurate identification of viral pathogens in clinical specimens, because enormous limitations impede the clinical use of common NGS, such as high cost, complicated procedures, tremendous data analysis, and high background noise in clinical samples.

METHODSViruses from cell culture materials or clinical specimens were identified following an improved NGS procedure: reduction of background noise by sample preprocessing, viral enrichment by barcoded oligonucleotide (random hexamer or non-ribosomal hexanucleotide) primer-based amplification, fragmentation-free library construction and sequencing of one-tube mixtures, as well as rapid data analysis using an in-house pipeline.

RESULTSNGS data demonstrated that both barcoded primer sets were useful to simultaneously capture multiple viral pathogens in cell culture materials or clinical specimens and verified that hexanucleotide primers captured as many viral sequences as hexamers did. Moreover, direct testing of clinical specimens using this improved hexanucleotide primer-based NGS approach provided further detailed genotypes of enteroviruses causing hand, foot, and mouth disease (HFMD) and identified other potential viruses or differentiated misdiagnosis events.

CONCLUSIONThe improved barcoded oligonucleotide primer-based NGS approach is simplified, time saving, cost effective, and appropriate for direct identification of viral pathogens in clinical practice.

Clinical Laboratory Techniques ; DNA Barcoding, Taxonomic ; DNA Primers ; Enterovirus ; classification ; genetics ; isolation & purification ; Herpesvirus 4, Human ; genetics ; isolation & purification ; Humans ; Influenza B virus ; genetics ; isolation & purification ; Real-Time Polymerase Chain Reaction ; Sequence Analysis, DNA ; methods ; Sequence Analysis, RNA ; methods

Rapid and accurate identification of an influenza outbreak is essential for patient care and treatment. We describe a next-generation sequencing (NGS)-based, unbiased deep sequencing method in clinical specimens to investigate an influenza outbreak. Nasopharyngeal swabs from patients were collected for molecular epidemiological analysis. Total RNA was sequenced by using the NGS technology as paired-end 250 bp reads. Total of 7 to 12 million reads were obtained. After mapping to the human reference genome, we analyzed the 3-4% of reads that originated from a non-human source. A BLAST search of the contigs reconstructed de novo revealed high sequence similarity with that of the pandemic H1N1 virus. In the phylogenetic analysis, the HA gene of our samples clustered closely with that of A/Senegal/VR785/2010(H1N1), A/Wisconsin/11/2013(H1N1), and A/Korea/01/2009(H1N1), and the NA gene of our samples clustered closely with A/Wisconsin/11/2013(H1N1). This study suggests that NGS-based unbiased sequencing can be effectively applied to investigate molecular characteristics of nosocomial influenza outbreak by using clinical specimens such as nasopharyngeal swabs.
Databases, Genetic ; Genotype ; High-Throughput Nucleotide Sequencing ; Humans ; Influenza A Virus, H1N1 Subtype/classification/*genetics/isolation & purification ; Influenza, Human/diagnosis/*virology ; Nasopharynx/*virology ; Nucleic Acid Amplification Techniques ; Phylogeny ; RNA, Viral/analysis/metabolism ; Sequence Analysis, RNA ; Viral Proteins/genetics

OBJECTIVETo evaluate the effect of supply of fresh poultry products on reducing environment contamination of avian influenza virus (AIV) in markets in Guangzhou.

METHODSA total of 40 markets, including 20 selling alive poultry and 20 selling fresh poultry products, were selected randomly in Guangzhou to conduct environment surveillance in 80 poultry stalls every 4 months from July 2014 to April 2015. Four smear samples were collected from different sites of each poultry stall to detect nucleic acid of AIV. The positive samples were further detected for AIV subtype H5, H7 and H9 nucleic acids.

RESULTSAmong 40 alive poultry stalls, 95.0% (38/40) kept alive poultry overnight, 25.0% (10/40) were disinfected daily, 95.0% (38/40) were cleaned up weekly, 95.0% (38/40) were closed for one day every month. Among 40 fresh poultry product stalls, 20.0% (8/40) were disinfected daily, 90.0% (36/40) were cleaned up weekly, and 96.0% (38/40) ever sold dressed poultry from alive poultry markets. The positive rate of AIV in alive poultry markets was 40.4% (252/623), higher than that in fresh poultry product markets (32.3%, 197/610), the difference was significant (χ(2)=8.85, P=0.003), and the positive rate of subtype H9 virus in alive poultry markets was 28.6% (178/623), higher than that in fresh poultry product markets (16.2%, 99/610), the difference was significant (χ(2)=26.95, P<0.001). In fresh poultry product markets, the positive rate of AIV in stalls selling dressed poultry was 37.3% (180/482), higher than that in stalls selling no dressed poultry (13.3%, 17/128), the difference was significant (χ(2)=26.78, P<0.001), and the positive rate of subtype H9 virus in stalls selling dressed poultry was 19.1% (92/482), higher than that in stalls selling no dressed poultry (5.5%, 7/128), the difference was significant (χ(2)=13.80, P<0.001). Both the positive rate of AIV and the positive rate of subtype H9 virus were highest in the second round surveillance (October 2014). The differences in AIV and its subtype H5, H7 and H9 virus positive rates of environmental samples from four different sites were not significant, respectively. In the same sample site, the positive rate of subtype H9 virus in alive poultry markets was higher than that in fresh poultry product markets the difference was significant (P<0.05).

CONCLUSIONSThe supply of fresh poultry products could effectively reduce the level of environment contamination of AIV in markets. Dressed poultry supplement caused the risk of AIV spread in fresh poultry product markets.

Animals ; China ; Commerce ; statistics & numerical data ; Disinfection ; statistics & numerical data ; Environmental Microbiology ; Influenza A virus ; isolation & purification ; Poultry ; Poultry Products ; supply & distribution

Animals ; China ; Influenza A virus ; genetics ; isolation & purification ; Influenza in Birds ; virology ; Nucleic Acids ; genetics ; Poultry ; virology

OBJECTIVETo compare the epidemiological and clinical features of lower respiratory tract infection (LRTI) caused by influenza virus A (IVA) and influenza virus B (IVB) in children.

METHODSThe clinical data of 366 children with LRTI caused by influenza virus (IV), who were hospitalized in Yuying Children′s Hospital of Wenzhou Medical University between 2010 and 2014, were analyzed retrospectively, and there were 272 cases caused by IVA and 94 cases caused by IVB.

RESULTSIV was mainly prevalent from December to March of the next year, with the predominance of IVA. There were small peaks of IVA prevalence in July or September every other year, and IVB was prevalent from December to March of the next year every other year. The children with LRTI caused by IVA alone had a significantly higher white blood cell (WBC) count and significantly higher percentages of children with increased WBC, abnormal serum sodium, and abnormal serum potassium than those caused by IVB alone (P<0.05). However, there were no significant differences in age, sex, underlying diseases, clinical manifestations, and co-infection rate with bacteria or atypical pathogens between the two groups (P>0.05). The rate of co-infection with respiratory syncytial virus (RSV) was significantly higher in the IVB group than in the IVA group (P<0.01).

CONCLUSIONSIVA is prevalent in winter and spring every year and has small peaks in summer every other year, while IVB is prevalent in winter and spring every other year. Compared with IVB, IVA causes more cases of increased WBC and electrolyte disturbance. The children infected with IVB are more likely to be co-infected with RSV. The children with LRTI caused by IVA and IVB have similar clinical manifestations.

Child ; Child, Preschool ; China ; epidemiology ; Female ; Humans ; Infant ; Infant, Newborn ; Influenza A virus ; genetics ; isolation & purification ; physiology ; Influenza B virus ; genetics ; isolation & purification ; physiology ; Influenza, Human ; diagnosis ; epidemiology ; virology ; Male ; Respiratory Tract Infections ; diagnosis ; epidemiology ; virology ; Retrospective Studies ; Seasons

A quail-origin subtype of the influenza virus was isolated from a human-infecting H7N9 subtype of the avian influenza virus found in a live poultry market and was given the name A/Quail/Hangzhou/1/ 2013 (H9N2). We analyzed the whole genome of this virus and its biologic characteristics. Sequence analyses suggested that the: HA and NS genes belonged to a CK/BJ/1/94-like lineage; NA, NP, PA and PB1 genes belonged to a SH/F/98-like lineage; M and PB2 genes belonged to a G1-like lineage. Analyses of key amino acids showed that the cleavage site in HA protein was PSRSSR ↓ GL, and that the HA protein had a human receptor-binding site with Leu226. Deletion of amino acids 69 - 73 was detected in the stalk of NA protein, the M2 protein had an Asn31 mutation, and the NS1 protein had two mutations at Ser42, Ala149. The intravenous pathogenicity of this virus was 0.36. A study in chickens suggested that all inoculated birds shed the virus from the trachea and cloaca on the third day post-infection (p. i. ) until 11 days. All chickens that had direct contact shed the virus on the second day p. i. until 8 days. Results of virus reisolation suggested that lung and tracheal tissues could shed the virus in 5 days, whereas the other organs could shed the virus in 3 days. These results suggest that this virus strain is H9N2 subtype LPAIV, whose lineage is prevalent in mainland China. This research provides evidence on how to monitor and prevent the H9N2 subtype of the avian influenza virus.
Animals ; Chick Embryo ; Chickens ; China ; Genotype ; Influenza A Virus, H9N2 Subtype ; classification ; genetics ; isolation & purification ; Influenza in Birds ; virology ; Phylogeny ; Quail ; virology