Isolation and Identification of H9N2 influenza and Newcastle disease viruses co-infections in chicken

Abstract

Co-infections with Newcastle disease (ND) and avian influenza (AI) highly pathogenic viruses causes high mortality and high economic losses. In this study, 322 samples were collected from 50 chicken flocks suffering from respiratory symptoms, severe drop in egg production and mortalities from December, 2013 to December, 2014. Real time polymerase chain reaction (RT-PCR) was performed to detect the M protein of H9 gene of AI and F gene of velogenic ND virus (vNDV) co-infection. H9 PCR was 20 and 34 in cloacal and pharyngeal swabs with total percentage of 6.2 % and 10.5%, respectively. The H9 sequences showed low pathogenic RSSR/GLF motif. The phylogenetic tree showed that H9 sequences belong to the G1 lineage which clustered with 2015 -2016 Egyptian H9N2. Partial sequences of the Fusion gene revealed that the isolated NDV isolates belong to class II genotype VIIj. The cleavage site of the F protein showed the presence of a polybasic amino acid motif (112RRQKRF117) suggesting that our two isolates were vNDV strain. The molecularly identified H9 (1 isolate) and NDV Genotype VII (2 isolates) were named AI/CHICKEN/EGYPT/48Ob/NRC-CU/2014(H9), NDV/CHICKEN/EGYPT/48Ob/NRC-CU/2014, and NDV/CHICKEN/EGYPT/ 66Oc/NRC-CU/2014, and nucleotide sequences were submitted to GenBank and given accession numbers MG966504, MG966505, and MG966506, respectively. Conclusion: Natural mixed infections with AIV H9N2 and virulent NDV genotype VIIj viruses were detected in chicken flocks suffering from respiratory symptoms and mortalities in Egypt.