J Vet Sci.  2018 May;19(3):406-415. 10.4142/jvs.2018.19.3.406.

Identification, sequence analysis, and infectivity of H9N2 avian influenza viruses isolated from geese

• Zhu R
• Yang X
• Zhang J
• Xu D
• Fan J
• Shi H
• Wang S
• Liu X

• KMID: 2412131
• DOI: http://doi.org/10.4142/jvs.2018.19.3.406

Abstract

The subtype H9N2 avian influenza virus greatly threatens the Chinese poultry industry, even with annual vaccination. Waterfowl can be asymptomatically infected with the H9N2 virus. In this study, three H9N2 virus strains, designated A/Goose/Jiangsu/YZ527/2011 (H9N2, Gs/JS/YZ527/11), A/Goose/Jiangsu/SQ119/2012 (H9N2, Gs/JS/SQ119/12), and A/Goose/Jiangsu/JD564/2012 (H9N2, Gs/JS/JD564/12), were isolated from domestic geese. Molecular characterization of the three isolates showed that the Gs/JS/YZ527/11 virus is a double-reassortant virus, combining genes of A/Quail/Hong Kong/G1/97 (H9N2, G1/97)-like and A/Chicken/Shanghai/F/98 (H9N2, F/98)-like; the Gs/JS/SQ119/12 virus is a triple-reassortant virus combining genes of G1/97-like, F/98-like, and A/Duck/Shantou/163/2004 (H9N2, ST/163/04)-like. The sequences of Gs/JS/JD564/12 share high homology with those of the F/98 virus, except for the neuraminidase gene, whereas the internal genes of Gs/JS/YZ527/11 and Gs/JS/SQ119/12 are closely related to those of the H7N9 viruses. An infectivity analysis of the three isolates showed that Gs/JS/SQ119/12 and Gs/JS/YZ527/11 replicated well, with seroconversion, in geese and chickens, the Gs/JS/JD564/12 did not infect well in geese or chickens, and the F/98 virus only infected chickens, with seroconversion. Emergence of these new reassortant H9N2 avian influenza viruses indicates that these viruses can infect both chicken and goose and can produce different types of lesions in each species.

Keyword

H9N2; biological characteristics; chickens; geese; infection

MeSH Terms

Animals
Asian Continental Ancestry Group
Chickens
Geese*
Humans
Influenza A Virus, H7N9 Subtype
Influenza A Virus, H9N2 Subtype
Influenza in Birds*
Neuraminidase
Population Characteristics
Poultry
Sequence Analysis*
Seroconversion
Vaccination
Neuraminidase

Figure

• 

  Fig. 1 Phylogenetic trees for all genes of avian influenza viruses (AIVs) collected from Eastern China between 2011 and 2015. Representative strains are indicated by solid triangles; the solid circles indicate strains isolated in this study; the solid rhombus indicates three commercial vaccine strains. Unrooted phylogenetic trees were generated by the distance-based neighbor-joining method using the MEGA6.0 software suite [29]. Nucleotide positions: (A) HA, 128–1540; (B) NA, 87–1263; (C) PB2, 28–2262; (D) PB1, 31–2218; (E) PA, 25–2129; (F) NP, 35–1456; (G) M, 56–960; (H) NS, 75–800. The percentage of the replicate trees in which the associated taxa clustered together in a bootstrap test with 1,000 replications are shown next to the branches; values lower than 70% are hidden.

• 

  Fig. 2 Analysis of close viral relationships of each gene from the three isolates in this study and their nucleotide homology. The colors represent the different nucleotide homology with each gene of the closest virus. The nucleotide homology in the same lineage between the isolates in this study and chosen H7N9 viruses. Eight gene segments in each of the schematic virus particles are arranged from top to bottom to represent polymerase basic 2 (PB2), polymerase basic 1 (PB1), polymerase acidic (PA), hemagglutinin (HA), nucleoprotein (NP), neuraminidase (NA), matrix (M), and nonstructural (NS) genes, and are indicated in same color with representative viruses for each lineage. The red indicates SH/F/98 lineage; the green indicates HK/G1/97 lineage; the yellow indicates ST/163/04 lineage; the blue indicates an undefined lineage, and the gray indicates another lineage.

• 

  Fig. 3 (A) Histopathology of SPF chickens and Ck/SH/F/98, Gs/JS/YZ527/11, Gs/JS/SQ119/12, and Gs/JS/JD564/12 infected geese. Lungs and tracheas were collected at 4 days post inoculation fixed in 10% formalin, embedded in paraffin, and sectioned. (B) Pathological changes in trachea and lung in chickens at 4 days after inoculation with the isolated viruses. Tissue sections were observed, and histopathological changes were scored as follows. For trachea, 0, normal; 1, congestion; 2, cilia loss; 3, a few inflammatory cell infiltration; 7, a lot of inflammatory cell infiltration. For lung, 0, normal; 1, congestion; 2, hemorrhage; 3, inflammatory cell infiltration in bronchial submucosa; 7, a lot of inflammatory cell infiltration in bronchial submucosa and alveolus. Average values for three birds are shown. Data are representative of three independent experiments. G, goose; C, chicken. Scale bars = 5 µm (A9 and A10), 10 µm (A1, A3, and A15), 20 µm (A2, A14, and A16), 50 µm (A4–8, A11–13).

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