B亚型禽偏肺病毒的分离鉴定及致病性研究

doi:10.11843/j.issn.0366-6964.2022.10.026

Abstract

Abstract: The purpose of this study was to identify the avian metapneumovirus (aMPV) as the pathogen of head shaking, mental depression and swollen head syndrome in some poultry farms in China. In this study, nasal turbinate and lung samples were collected from chicken farms in Shandong, Fujian and Heilongjiang. Firstly, the clinical samples were detected by aMPV specific RT-PCR method, and then the positive samples were inoculated into Vero cells for virus isolation. Secondly, the subtypes of the viruses were identified by G and F gene sequence analysis and indirect immunofluorescence assay (IFA). Finally, the pathogenicity of the isolate to SPF chickens was analyzed. Among the 220 samples, the RT-PCR results indicated that three nasal turbinate samples had specific bands at about 228 bp. After the positive samples were inoculated into Vero cells for 5 consecutive generations, the cells showed aMPV characteristic cytopathy (CPE) such as rounding, aggregation and fusion. These results showed that three isolates of aMPV were successfully isolated and named SD2001, SD2002 and HLJ2101, respectively. Phylogenetic analysis showed that the G and F genes of layer-originated SD2001, SD2002 and broiler-originated HLJ2101 isolates had high nucleotide and amino acid sequence homologies with other subtype B aMPV strains, and the nucleotide similarities were 93.4%-98.6% and 95.6%-100.0%, respectively, and the amino acid similarities were 88.7%-97.8% and 97.6%-100.0%, respectively. However, the homologies with the G and F genes of subtypes A, C and D aMPV strains were low, and the nucleotide similarities were 27.1%-61.8% and 66.8%-74.8%, respectively, and the amino acids similarities were 16.1%-36.7% and 72.5%-86.5%, respectively. These results indicated that SD2001, SD2002 and HLJ2101 isolates belonged to subtype B aMPV. Further, the specific positive serum of subtype B aMPV was used for IFA test. The specific green fluorescence signal could be observed in Vero cells inoculated with SD2001, SD2002 and HLJ2101, respectively, which furtherly confirmed that the three isolates belonged to subtype B aMPV. Next, the pathogenicity of the isolate to SPF chickens was tested. SPF chickens infected with SD2001 isolate showed clinical symptoms such as depression, head shaking and sneezing 3-6th day after infection. Histopathological analysis showed that there were pathological lesions in the turbinate, trachea and lung of infected SPF chickens. The incidence rate was 90%(18/20). In this study, three strains of subtype B aMPV were successfully isolated. Our results not only identified the pathogen of swollen head syndrome in some poultry farms in China, but also confirmed that the epidemic strain of subtype B aMPV had obvious pathogenicity to chickens. These results provided an important theoretical basis for the diagnosis and effective prevention of aMPV infection in China.

Key words: subtype B avian metapneumovirus, isolation, identify, pathogenicity

CLC Number:

S852.659.5

YU Mengmeng, BAO Yuanling, WANG Suyan, XIN Ziqi, LIU Peng, FENG Xiaoyan, MENG Lingzhai, GUO Ru, ZHANG Yanping, LIU Changjun, QI Xiaole, LI Junping, WANG Xiaomei, GAO Yulong. Isolation, Identification and Pathogenicity of Subtype B Avian Metapneumovirus[J]. Acta Veterinaria et Zootechnica Sinica, 2022, 53(10): 3540-3549.

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Isolation, Identification and Pathogenicity of Subtype B Avian Metapneumovirus

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