Research Article Open Access

Identification and Subtyping of Avian Influenza Viruses by Reverse Transcription Polymerase Chain Reaction (RT-PCR) and Agarose Gel Electrophoresis

Authors

CORRESPONDING AUTHOR
B.P. Shankar
High Security Animal Disease Laboratory, IVRI, Bhopal, India
|
R. N. Sreenivas Gowda
Veterinary College, Hebbal, Bangalore-24, Karnataka, India
|
B. Pattnaik
High Security Animal Disease Laboratory, IVRI, Bhopal, India
|
B. H. Manjunatha Prabhu
Molecular Biology Genetics Units, JNCASR, Jakkur, Bangalore, India
|
B.K. Sreenivas
IAH&VB, Hebbal, Bangalore, India
|
M.K. Vinuthan
MRDG, IISc, Bangalore, India
|
D. Ranjith
Veterinary College, Hebbal, Bangalore-24, Karnataka, India
|
H.K. Pradhan
High Security Animal Disease Laboratory, IVRI, Bhopal, India
Published on: April 15, 2009
DOI 10.3923/ijps.2009.465.469
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Abstract

Avian Influenza (AI) is caused by type A influenza virus belonging to the family orthomyxoviridae, which is classified into 16 HA and 9 NA subtypes based on two surface glycoprotein’s haemagglutinin (HA) and neuraminidase (NA). In the present study we did identification and HA-subtyping of avian influenza virus by reverse transcription-PCR (RT-PCR) during the first outbreaks of AI in India during 2006. The avian influenza virus is identified by RT-PCR using a set of primers specific to the nucleoprotein (NP) gene of avian influenza virus. The HA-subtypes of avian influenza virus were determined by running with HA subtype specific primers for H5, H7 and H9 RT-PCR reactions, each using a set of primers specific to one HA-subtype. A total of 10,236 tissue / cloacal swab samples, received at the HSADL from various parts of the country, were processed for isolation of AI virus in embryonated chicken eggs. Out of these, 9 samples originating from poultry in Maharashtra (Navapur and Jalgaon) and Gujarat (Surat) states of India were found positive for H5 virus by RT-PCR. All samples received from outbreaks areas were tested by using all tree subtype specific primers(H5, H7 and H9) only H5 RT-PCR reactions was give the product of expected size, and thus the HA-subtype of the virus is determined. One sample gave the positive result with H9 subtype specific primers. The RT-PCR procedure is rapid and sensitive, and could be used for the identification and HA-subtyping of avian influenza virus in organ homogenates.

Keywords:

Avian influenza, H5N1, India, RT-PCR

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Shankar, B., Gowda, R. N. S., Pattnaik, B., Prabhu, B. H. M., Sreenivas, B., Vinuthan, M., Ranjith, D., & Pradhan, H. (2009). Identification and Subtyping of Avian Influenza Viruses by Reverse Transcription Polymerase Chain Reaction (RT-PCR) and Agarose Gel Electrophoresis. International Journal of Poultry Science, 8(5), 465–469. https://doi.org/10.3923/ijps.2009.465.469
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