Animals ; Humans ; Influenza A Virus, H5N1 Subtype ; immunology ; pathogenicity ; Poultry ; virology

Humans ; Influenza A Virus, H5N1 Subtype ; classification ; genetics ; pathogenicity ; Influenza Vaccines ; immunology ; Influenza, Human ; drug therapy ; prevention & control ; transmission ; Vaccination

Two H5N1 avian influenza viruses (AIV), A/mallard/Huadong/S/2005 (S, IVPI = 2.65, in mallard) and A/mallard/Huadong/Y/2003 (Y, IVPI = 0, in mallard), were capable of distinct in pathogenicity to non-immunized mallards (Anas platyrhynchos). There were two amino acid residues difference in the HA cleavage site between two viruses, 322 (S, Leu; Y, Gln) and 329 (S, deletion; Y, Lys). Based on the variation, a series of recombinant viruses carrying HA gene either from S or Y virus with mutation at 322 and/or 329 were constructed via reverse genetics system to explore the influence of the two amino acid residues on viral pathogenicity in mallards. Recombinant viruses with S virus backbone were completely attenuated in terms of their virulence to ducks when position 322 (L322Q) and/or position 329 (-329K) of HA gene had been mutated. The critical role that L322 and -329 of HA protein from S virus play in the high virulence to ducks were influenced by the entire background of that protein because the recombinant virus with HA gene from Y and other seven genes from S were completely attenuated even if Q322L and K329- mutations of HA gene had been achieved. Recombinant viruses with Y virus backbone significantly increased their virulence to ducks when position 322 (Q322L) and/or position 329 (K329-) of HA gene had been mutated. All recombinant viruses carrying HA gene from Y with Q322L and/or K329-mutations and other seven genes from S were completely attenuated in terms of virulence to ducks whereas all recombinant viruses carrying HA gene from Y with same mutations and other seven genes from Y gained significant virulence. It seems that the compatibility among eight genes might be an important factor for HA to exert its functions. Results indicated that the mutation at amino acid position 322 and deletion at 329 in HA cleavage site significantly influence the pathogenicity of S and Y viruses in mallard, the compatibility among eight genes also contribute to the pathogenicity of both viruses in mallard.
Animals ; Birds ; Hemagglutinin Glycoproteins, Influenza Virus ; chemistry ; genetics ; physiology ; Influenza A Virus, H5N1 Subtype ; genetics ; pathogenicity ; Structure-Activity Relationship ; Virulence

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

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 evaluate the risk of human infection after the outbreak of avian influenza H5N1 in animals, and probe the possibility for virus transmission.

METHODSBy means of field epidemiological study, molecular epidemiology, serology and emergency surveillance, persons who had ever closely contacted with sick or dead poultry were observed. While, the RT-PCR and gene sequencing method were used to detect H5 nucleic acid from environmental swabs from 4 epidemic spots, and hemagglutination inhibition assay was also used to detect H5 antibody.

RESULTSOf 22 environmental swabs detected from 4 epidemic spots, one was positive for H5 nucleic acid, and the homogeneity was 95.9% as compared with H5N1 virus A/China/GD01/2006 (H5N1) found in Guangzhou in 2006 by gene sequence analysis. 62 environmental swabs from live poultry stalls of food markets near epidemic spot were detected negative. Six of 68 blood samples of contacts were positive for H9 antibody, and all were negative for H5 antibody. 68 throat swabs of contacts were detected negative for H5 nucleic acid. No close contact was found abnormal after 7 days medical observation. 337 influenza-like cases were reported in emergency surveillance, and no suspicious case was found.

CONCLUSIONThe current outbreak of H5N1 avian influenza in water fowls has not yet caused further transmission, and human avian influenza case has not been observed. It indicates that the ability of H5N1 virus to transmit to human is not strong yet, and the risk of human infection for H5N1 is still low.

Animals ; Antibodies, Viral ; blood ; China ; epidemiology ; Disease Outbreaks ; Ducks ; Humans ; Influenza A Virus, H5N1 Subtype ; genetics ; isolation & purification ; pathogenicity ; Influenza in Birds ; epidemiology ; transmission ; Influenza, Human ; epidemiology ; transmission ; Risk Assessment

Human, birds and other mammals can be infected by avian influenza A (H5N1) virus. Bird infections spread among continents in the last 2 years. Human infections become endemic in certain areas, characterized with young patients and high mortality. An unprecedented epizootic avian influenza A (H5N1) virus that is highly pathogenic has crossed the species barrier in Asia to cause many human fatalities and poses an increasing pandemic threat. This summary describes the features of human infection with influenza A (H5N1) in epidemiology and etiology and reviews recommendations for prevention.
Animals ; Birds ; China ; epidemiology ; Humans ; Indonesia ; epidemiology ; Influenza A Virus, H5N1 Subtype ; pathogenicity ; Influenza in Birds ; prevention & control ; transmission ; Influenza, Human ; epidemiology ; prevention & control ; transmission

This study aims to investigate the virological impact of the stalk region and cysteine (C) in neuraminidase (NA) of influenza A/Anhui/1/05 (H5N1) and A/Ohio/07/2009 (H1N1) viruses. The NA of A/ Anhui/1/05 (H5N1), defined as AH N1, lacked 20 amino acids (including C, defined as s20) as compared with NA of A/Ohio/07/2009 (H1N1) (defined as 09N1). We deleted s20 of 09N1 to construct 09N1-s20, and inserted s20 into AH N1 to construct AH N1+s20. To investigate the impact of C on the biological function of NA, we deleted C in 09N1 to construct 09N1-C and inserted C into AH N1 to construct AH N1-C. The pseudo-type viral particle (pp) system was used to evaluate the impact of these mutants on virology. The combination of 09N1-C and 09H1 (defined as 09H1::09N1-C) showed an infectivity 8 times that of the wild type 09H1::09N1, while the infectivity of the combination of AH N1+C and AH H5 (defined as AH H5::AH N1+C) was much lower than that of the wild type AH H5::AH N1. The infectivity of the combination of 09N1-s20 and 09H1 (defined as 09H1::09N1-s20) was 4 times that of the wild type 09H1::09N1; the infectivity of the combination of AH N1+s20 and AH H5 (defined as AH H5:: AH N1+s20) was 1/7 that of the wild type AH H5::AH N1. The co-existence of 09N1-C and AH H5 displayed 6 times the infectivity of AH H5::09N1, while the infectivity of 09H1::AH N1+C was very low. Multimer analysis showed that in the wild type 09N1, the forms of NA were dimer > tetramer > monomer; the major component of NA in 09N1-C was monomer; in 09N1-s20, the forms of NA were monomer > dimer. AH N1 was mainly composed of monomer; in AH N1+s20, the forms of NA were dimer > monomer > tetramer; in AH N1+C, the forms of NA were dimer > tetramer. Deletion of C or s20 from 09N1 did not change the expression of NA. The study suggested that deletion of C from the stalk region of NA in A/Ohio/07/2009 (H1N1) increases infectivity. Insertion of C into NA's stalk region of A/ Anhui/1/05 (H5N1) significantly decreases infectivity. Cysteine deletion in the stalk region is important for the infectivity of A/Anhui/1/05 (H5N1) and A/Ohio/07/2009 (H1N1). It may interfere with the infectivity via changes in NA polymerization.
Amino Acid Motifs ; Humans ; Influenza A Virus, H1N1 Subtype ; chemistry ; enzymology ; genetics ; pathogenicity ; Influenza A Virus, H5N1 Subtype ; chemistry ; enzymology ; genetics ; pathogenicity ; Influenza, Human ; virology ; Neuraminidase ; chemistry ; genetics ; metabolism ; Viral Proteins ; chemistry ; genetics ; metabolism ; Virulence

OBJECTIVETo analyze the spatial distribution of highly pathogenic avian influenza (HPAI) and to explore environmental factors associated with HPAI using geographic information system (GIS) techniques in Mainland China.

METHODSDatabases were set up using the information of HPAI during epidemics in 2004, and linked to digital maps at provincial and county administrative layers in the country through the ArcGIS 8.3 software. Spatial cluster analyses, spatial statistics analyses and tracking analyses on epidemic situation of HPAI were implemented. Environmental factors associated with HPAI were also analyzed on data related to weather, vegetation and migratory birds etc.

RESULTSFindings from spatial cluster analyses showed that high incidence area was centralized in 113.261 degrees ordm; east longitude and 23. 119 degrees ordm; north latitude with a radius of 1090.52 kilometers (relative risk= 2.646, P value= 0.001). Spatial statistical analyses showed that HPAI took place mainly in capital cities of provinces and surrounding areas as well as in the circumference areas of arterial rivers, lakes and seacoasts. Results also showed that HPAI occurrences were associated with low air temperature, high relative humidity and high air pressure as well as with east & central migration routes of migratory birds. The average normalized difference vegetation index was 0.36 +/- 0.11 in epidemic areas of HPAI.

CONCLUSIONHPAI was unrandomly distributed and geographically clustered in China.

Animal Migration ; Animals ; Atmospheric Pressure ; Birds ; virology ; China ; epidemiology ; Cluster Analysis ; Environment ; Geographic Information Systems ; Humidity ; Influenza A Virus, H5N1 Subtype ; pathogenicity ; Influenza in Birds ; epidemiology ; Temperature

OBJECTIVETo explore histopathologic and ultrastructural characteristics of human avian influenza (AI) infection and related etiological pathogenesis.

METHODSPostmortem lung and heart samples were collected from the patient who died of avian influenza virus infection on November 29, 2003 in China. Light and electron microscopy, immunohistochemistry and histochemistry were used to investigate the pathological changes.

RESULTSThe main pathological findings included extensive pulmonary consolidation, hemorrhage, pulmonary edema and local hemorrhagic infarct. The lamina of alveoli and bronchioles were abundantly filled with protein-rich fluid, erythrocytes, fibrin and cell debris admixed with many neutrophilis, macrophages, lymphocytes and a few of monokaryon and multinuclear giant cells. Hyaline membranes were formed. Local pulmonary tissues were heavily damaged by hemorrhage and necrosis. Alveolar septum was disintegrated. Mesenchymal edema with a few of macrophages infiltration of heart was found. Electron microscopy showed the avian influenza A virus-like particles (type C and type A) of 80 - 120 nm diameter and envelopes in the cytoplasm of pneumocytes and endothelial cells.

CONCLUSIONSFatal pneumonia associated with highly pathogenic avian influenza A virus (H5N1) infection leads to extensive pulmonary consolidation, edema and marked hemorrhagic necrosis and inflammation. Electron microscopy can identify avian influenza A virus-like particles. The findings may offer an important theoretical basis for clinical diagnosis and treatment.

Animals ; Autopsy ; methods ; Birds ; China ; Humans ; Influenza A Virus, H5N1 Subtype ; pathogenicity ; ultrastructure ; Influenza A virus ; classification ; Influenza in Birds ; pathology ; Influenza, Human ; diagnostic imaging ; pathology ; surgery ; virology ; Microscopy, Electron ; Ultrasonography ; Virulence Factors