Animals ; Birds ; Hemagglutinin Glycoproteins, Influenza Virus ; genetics ; immunology ; Humans ; Influenza A virus ; genetics ; immunology ; pathogenicity ; Influenza in Birds ; immunology ; pathology ; virology ; Influenza, Human ; immunology ; pathology ; virology ; Virulence ; genetics

Multiple epitopes from one or more viruses can be lined up and co-expressed in one vector to generate multi-epitopes DNA vaccines. In the study, four recombinant plasmids were constructed based on HA and NP gene of avian influenza virus (AIV) (H5N1): (1) pIRES/HA, carrying the complete HA gene; (2) pIRES/tHA, carrying a truncated HA gene fragment of major neutralizing antigenic epitopes; (3) pIRES/tHA-NPep, in which three CTL epitopes of NP gene of AIV were fused to the truncated HA from the C-terminal; and (4) pIRES/tHA-NPep-IFN-gamma, which was constructed by replacing neo gene in pIRES/ tHA-NPep with IFN-y of chicken. Fifty five SPF chickens were randomly divided into five groups and immunized with the above four constructs and control plasmid. Each chicken was intramuscally immunized with 200 microg plasmid DNA three times in a two-week interval. Two weeks after the third immunization, chickens were injected with H5N1 subtype avian influenza virus. Before the virus loading no detectable antibodies to HA were found in the chicken serum; but high levels of HI antibodies were detected in the serum of the survived chickens. The percentages of CD4+ and CD8+ T lymphocyte in peripheral blood of immunized chickens increased steadily after the vaccination. After virus loading all chickens in the control group died within three to eight days, and the survival rates of the four DNA vaccine groups were as follows: pIRES/HA, 54.5%; pIRES/tHA, 30%, pIRES/ tHA-NPep, 36.3%, pIRES/tHA-NPep-IFN-gamma, 50%. These results indicated that multi-epitopes DNA immunization can induce immune response and protect chickens from homologous virus loading.
Animals ; Chickens ; Epitopes ; immunology ; Influenza A Virus, H5N1 Subtype ; immunology ; pathogenicity ; Influenza in Birds ; immunology ; prevention & control ; virology ; Vaccines, DNA ; immunology

OBJECTIVETo understand the epidemic status of avian influenza A virus in chickens and men in Guangzhou area and to prevent men suffering from avian influenza A (H5N1) virus.

METHODSEtiologic and serological surveys were conducted in chickens and men who were working in the poultry farms and slaughter house. Viruses were isolated with both MDCK cells and embryonated chicken eggs. Hemagglutination inhibition tests were performed by routine method.

RESULTSAnti-H9N2 antibody was found in 12.8% of the chickens and 5.1% of the workers.

CONCLUSIONSAvian influenza virus H9N2 subtype existed in chickens and this subtype of influenza A virus might infect men.

Animals ; Antibodies, Viral ; blood ; Chickens ; virology ; China ; Hemagglutination Inhibition Tests ; Humans ; Influenza A Virus, H9N2 Subtype ; immunology ; isolation & purification ; Influenza in Birds ; virology ; Influenza, Human ; virology

Since 2002, H7 subtype avian influenza viruses (AIVs) have caused more than 100 human infection cases in the Netherlands, Italy, Canada, the United States, and the United Kingdom, with clinical illness ranging from conjunctivitis to mild upper respiratory illness to pneumonia. On March 31st, three fatal cases caused by infection of a novel reassortant H7N9 subtype were reported in Shanghai City and Anhui Province in China. With the ability of H7 subtype to cause severe human disease and the increasing isolation of subtype H7 AIVs, we highlighted the need for continuous surveillance in both humans and animals and characterization of these viruses for the development of vaccines and anti-viral drugs.
Animals ; Chickens ; Ducks ; Humans ; Influenza A virus ; genetics ; isolation & purification ; pathogenicity ; physiology ; Influenza Vaccines ; genetics ; immunology ; Influenza in Birds ; immunology ; prevention & control ; virology ; Influenza, Human ; immunology ; prevention & control ; virology ; Poultry Diseases ; immunology ; prevention & control ; virology ; Turkeys

Two hundred thirty specimens of wild birds were collected from some areas in Heilongjiang Province during the period of 2003~2004, including two batches of specimens collected randomly from a same flock of mallards in Zhalong Natural Reserve in August and December, 2004, respectively. Primary virus isolation and identification for avian influenza virus (AIV) and Newcastle disease virus (NDV) were performed. The results showed that only two specimens of young mallards collected from Zhalong Natural Reserve in August, 2004 were positive to AIV (isolation rate 0.9%), and one strain (D57) of these two virus isolates was identified to be H9 subtype by hemagglutination inhibition test. Meanwhile, the two batches of blood serum samples of mallards from Zhalong were also examined for antibodies against AIV and NDV. Among 38 blood serum samples collected in August, antibodies against the hemagglutinin of H1, H3, H5, H6 and H9 subtypes of AIV were found in 1, 0, 2, 0 and 8 samples, respectively; and 11 samples were found with antibody against NDV. Whereas the NDV isolation in both two batches of specimens of mallard was negative, all of the 32 blood serum samples collected in December were negative for antibodies against AIV and NDV.
Animals ; Animals, Wild/*virology ; Antibodies, Viral/isolation&purification ; Birds/virology ; China/epidemiology ; Hemagglutination Tests ; Influenza A virus/*isolation&purification ; Influenza in Birds/epidemiology/immunology/*virology ; Newcastle Disease/epidemiology/immunology/*virology ; Newcastle disease virus/*isolation&purification ; Reverse Transcriptase Polymerase Chain Reaction

OBJECTIVETo survey the distribution of influenza A subtypes in external environment and investigate the infectious status of highly pathogenic avian influenza (H5N1) in poultry-exposed population in Wuhan.

METHODSSeventy-eight external environmental samples (water, cage surface and fecal samples) were collected from 3 habitats of wild migratory birds and 5 urban live-poultry markets in 2010. In 13 avian influenza monitoring points, 249 serum samples were collected from people living around habitats of wild migratory birds or working in live poultry markets. Real-time RT-PCR method was adopted to detect influenza A virus from external environmental samples; and multiple RT-PCR method and specific H3, H5, H7 and H9 primers were then applied to analyze the subtypes of the positive samples. The levels of H5N1 antibody in poultry-exposed population were tested by horse hemagglutination inhibition test and two avian influenza inactivated antigens: A/Hubei/1/10 and A/Anhui/1/05.

RESULTSOf the 50 external environmental samples collected from live poultry markets, 17 samples were determined to be influenza A virus positive (positive rate 34.0%), including specific subtypes as follows: 4 samples of H5 single-positive subtype, 3 samples of H9 single-positive subtype, 4 samples of H3 and H5 mixed-positive subtype, 2 samples of H3 and H9 mixed-positive subtype, 2 samples of H5 and H9 mixed-positive subtype, 2 samples of H3, H5 and H9 mixed-positive subtype, but no H7 positive subtype was found. The 28 external environmental samples collected from habitats of wild migratory birds were all influenza A virus negative. Considering different types of external environmental samples, the influenza A virus positive rates in water, cage surface and fecal samples were 37.5% (6/16), 16.7% (5/30) and 18.8% (6/32), respectively. There were total 100 samples of serum whose A/Hubei/1/10 antigen inhibiting titers ≥ 40, accounting for 40.2%; while 36 samples of serum (14.5%) whose A/Anhui/1/05 antigen inhibiting titers ≥ 40 were found. The difference had statistical significance (χ(2) = 41.433, P < 0.05). Among the 249 serum samples collected from poultry-exposed population, 5 samples were H5N1 antibody positive against A/Hubei/1/10 antigen (inhibition titer ≥ 160), which came from 4 different live poultry markets, however, no positive serum sample against A/Anhui/1/05 antigen was found.

CONCLUSIONMultiple subtypes of avian influenza virus simultaneously prevailed in Wuhan urban poultry markets. Moreover, results from the distribution of avian influenza virus in external environment were consistent with the level of H5N1 antibody in poultry-exposed population.

Animals ; Antibodies, Viral ; blood ; Birds ; virology ; China ; Environment ; Humans ; Influenza A Virus, H5N1 Subtype ; immunology ; Occupational Exposure ; Poultry ; virology

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 recent human infection with avian influenza virus revealed that H9N2 influenza virus is the gene donor for H7N9 and H10N8 viruses infecting humans. The crucial role of H9N2 viruses at the animal-human interface might be due to the wide host range, adaptation in both poultry and mammalian, and extensive gene reassortment. As the most prevalent subtype of influenza viruses in chickens in China, H9N2 also causes a great economic loss for the poultry industry, even under the long-term vaccination programs. The history, epidemiology, biological characteristics, and molecular determinants of H9N2 influenza virus are reviewed in this paper. The contribution of H9N2 genes, especially RNP genes, to the infection of humans needs to be investigated in the future.
Animals ; Chickens ; virology ; China ; epidemiology ; Humans ; Influenza A Virus, H7N9 Subtype ; genetics ; Influenza A Virus, H9N2 Subtype ; genetics ; immunology ; physiology ; Influenza in Birds ; epidemiology ; transmission ; virology ; Influenza, Human ; epidemiology ; transmission ; virology ; Vaccination ; Viral Proteins ; classification ; metabolism

OBJECTIVETo prepare the 4 candidate vaccine strains of H5N1 avian influenza virus isolated in China.

METHODSRecombinant viruses were rescued using reverse genetics. Neuraminidase (NA) and hemagglutinin (HA) segments of the A/Xinjiang/1/2006, A/Guangxi/1/2009, A/Hubei/1/2010, and A/Guangdong/1/2011 viruses were amplified by RT-PCR. Multibasic amino acid cleavage site of HA was removed and ligated into the pCIpolI vector for virus rescue. The recombinant viruses were evaluated by trypsin dependent assays. Their embryonate survival and antigenicity were compared with those of the respective wild-type viruses.

RESULTSThe 4 recombinant viruses showed similar antigenicity compared with wild-type viruses, chicken embryo survival and trypsin-dependent characteristics.

CONCLUSIONThe 4 recombinant viruses rescued using reverse genetics meet the criteria for classification of low pathogenic avian influenza strains, thus supporting the use of them for the development of seeds and production of pre-pandemic vaccines.

Animals ; Chick Embryo ; Chickens ; China ; Hemagglutinin Glycoproteins, Influenza Virus ; genetics ; metabolism ; Influenza A Virus, H5N1 Subtype ; immunology ; Influenza Vaccines ; immunology ; Influenza in Birds ; prevention & control ; virology ; Neuraminidase ; genetics ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; Vaccines, Synthetic ; immunology

Avian influenza has emerged as one of the primary public health concern of the 21st century. Influenza strain H5N1 is capable of incidentally infecting humans and other mammals. Since their reemergence in 2003, highly pathogenic avian influenza A (H5N1) viruses have been transmitted from poultry to humans (by direct or indirect contact with infected birds) in several provinces of Mainland China, which has resulted in 22 cases of human infection and has created repercussions for the Chinese economy. People have been concerned whether a new pandemic will occur in the future. The eradication of pathogenic avian influenza viruses appears to be the most effective way to prevent an influenza pandemic. This paper will examine the features of H5N1, including incidence, infection, immunity, clinical management, prevention and control, and therapy in Mainland China.
Adolescent ; Adult ; Animals ; Birds ; Child ; China/epidemiology ; Disease Outbreaks/prevention & control ; Female ; Humans ; Incidence ; *Influenza A Virus, H5N1 Subtype ; Influenza in Birds/prevention & control ; Influenza, Human/*epidemiology/immunology/therapy ; Male ; Zoonoses/epidemiology/transmission/virology