Assessment of experimental pathogenicity of avian influenza virus H9N2 isolates by intravenous pathogenicity index (IVPI) test and histopathology

A total of 1,423 number of tissue samples received from various parts of India were processed for virus isolation in chicken embryos of 9-11 d old as a part of programme to monitor Avian Influenza Virus (AIV) infection in Indian poultry population and a total number of three samples were positive by HA test and the titre of the virus isolates ranged from 1:8 to 1:64. The virus isolates had been subjected to HI test with the reference positive serum imported from NVSL, USA for various subtypes. The virus isolates gave a HI titre of 1:32 with H9 subtype specific reference positive serum indicating that the virus isolates were influenza A H9 subtype. One of the isolates (accession no. 3722/04) was subjected to neuraminidase assay and the virus was found by N2 subtype and also virus sub typing was done by RT-PCR for further confirmation. The influenza H9N2 virus isolates (accession no. 2543/04, 2544/04, 2317/04, 2424/04, and 3722/04) were used in the intravenous pathogenicity index test. In the present study no, clinical signs and/or gross lesions in experimentally inoculated chickens were observed. All the five isolates studied were found to be non-pathogenic according to the European Union definition. The organs collected from the experimental chickens did not show significant lesions, on histopathological examination, except for some minor changes in some of the organs. Immunofluorescence test was used to study the tissue tropism of avian influenza viruses’ isolates experimentally infected in chickens. The antigen was detected only in the brain and trachea of the chickens experimentally infected with the isolates having accession no. 2543/04 and intestine of chickens experimentally infected with the isolate having accession number 2544/04. Avian influenza virus re-isolation from experimentally infected chickens was attempted in 9-11 d old embryonated chicken eggs. Virus could be re-isolated from some organs, three groups infected with the isolates with accession numbers 2543/04, 2544/04 and 2317/04. Thus, the present work was envisaged to fill in the gap formed by this lack of information on avian influenza in India.