Activities in 2009
Highly pathogenic avian influenza
Dr S.C. Dubey
High Security Animal Disease Laboratory, IVRI, Anand Nagar, Bhopal – 462 021, India
Tel.: (0091) 755 2759204), Fax: (0091) 755 2758842
E-mail address: scd_11@yahoo.in, website: www.hsadl.org
Summary of general activities related to the disease
1.
2.
Test(s) in use/or available for the specified disease at your laboratory
Test
For
Specificity
Total
Hemagglutination Assay
Antigen detection
-
22032
Hemagglutination
Inhibition Assay
Antigen detection
Subtype
1550
Hemagglutination
Inhibition Assay
Antibody detection
Subtype
26127
Neuramindase inhibition
Assay
Antigen detection
Subtype
26
RT-PCR
Genome detection
Group/Subtype
2850
Real time RT-PCR
Genome detection
Group/Subtype
2940
ELISA
Antibody detection
Group
395
Genome sequencing
Molecular epidemiology
–
20+2 isolates
(H5N1+H9N2)
Production and distribution of diagnostic reagents
Whole viral inactivated antigen is being used in AGID test for diagnosis of avian influenza. Approximately 100ml
of reagent was prepared for in-house use.
Chicken antiserum was raised against HPAIV H5N1for use in HA and HI assays for diagnosis of avian influenza.
Approximately 100ml of antiserum was prepared for in-house use.
Activities specifically related to the mandate
of OIE Reference Laboratories
3.
International harmonisation and standardisation of methods for diagnostic testing or the
production and testing of vaccines
As per the OIE guide lines, Department of Animal Husbandry, Government of India, maintains a strategic reserve
of killed H5N2 vaccine (Intervet). The vaccine has been tested by HSADL against challenge of Indian isolate of
Annual reports of OIE Reference Laboratories and Collaborating Centres, 2009
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Highly pathogenic avian influenza
HPAI H5N1 in SPF chickens and was found to be effective in preventing clinical disease and reduction of viral
shedding. Testing of field isolates of H5N1 as vaccine strain has also been taken up by HSADL. Though
vaccination against avian influenza has not been a part of control program at present, the testing of avian influenza
vaccine against newer isolates (H5N1) provides useful data for prediction of performance of avian influenza
vaccine in case of emergency use.
The Department of Animal Husbandry, Government of India, provides OIE validated commercial ELSA kits for
sero-surveillance of avian influenza in chickens at regional disease diagnostic laboratories after validation of the
kits by HSADL. In this activity, 2 batches of ELISA kits (Anigen, Korea) were validated with reference tests
(AGID and HI) across 200 serum samples for each batch. The kits were found to be satisfactory pertaining to
repeatability, diagnostic sensitivity and specificity.
4.
Preparation and supply of international reference standards for diagnostic tests or vaccines
1. Preparation of nucleoprotein specific recombinant antibody.
2. Preparation of NS1 protein specific mouse recombinant monoclonal antibody
5.
Research and development of new procedures for diagnosis and control
Killed H5N2 vaccine (Intervet) has been tested by HSADL against challenge of Indian isolate of HPAI H5N1 in
SPF chickens for its validation through immune response and experimental challenge using limited number of
birds.
6.
Collection, analysis and dissemination of epizootiological data relevant to international disease
control
Shared epidemiological data of avian influenza with Dr. David Castellan, Regional Veterinary Epidemiologist,
Emergency Centre for Transboundary Animal Diseases (ECTAD), FAO Regional Office for Asia and the Pacific
(FAO-RAP), Bangkok, Thailand.
Samples from migratory birds collected under guidance of the FAO officials from bird sacturies on migratory
flyways in India were tested by us and resultant data was shared with Prof. M. V. Subba Rao, National Project
consultant (AI), FAO India Office, New Delhi.
7.
Provision of consultant expertise to OIE or to OIE Members
The OIE recognition to this lab was accorded in May 2009. Since then technical advice was provided to DADF,
MOA, GOI on matters related to AI along with initiation of competence building activities through DADF and
SAARC Secretariat, New Delhi. However, before OIE recognition technical discussions on the points of mutual
interest related to AI diagnosis and control were carried out with counterparts from Republic of Bangladesh and a
MOU was also signed between two Governments.
8.
Provision of scientific and technical training to personnel from other OIE Members
1. Four personnel from Afghanistan, Bhutan and Sri Lanka were trained on “Molecular diagnosis of avian
influenza” during December 8-19, 2009.
2. During 2009 but before OIE recognition a group of trainees from Myanmar, Bangladesh and Nepal was also
trained for AI diagnosis under a FAO sponsored program.
9.
Provision of diagnostic testing facilities to other OIE Members
In this duration no samples were submitted to HSADL for testing and or confirmation.
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Annual reports of OIE Reference Laboratories and Collaborating Centres, 2009
Highly pathogenic avian influenza
10. Organisation of international scientific meetings on behalf of OIE or other international bodies
International workshop on “Laboratory bio-safety and bio-containment” sponsored by WHO, India office, New
Delhi, November 17-19, 2009 in which senior faculties both from medical and veterinary profession participated.
Dr. Michael Johnson, Head of Laboratory, Institute for Animal Health, Pirbright, UK and Ms Chook Mee Lan,
Head, Facilities Services & Biosafety Manager, TLL, Research Link, National University of Singapore were
invited.
11. Participation in international scientific collaborative studies
Collaborative project on “Development of diagnostics and vaccines against avian influenza” with Dr. David
Suarez, SEPRL, Athens, USA.
12. Publication and dissemination of information relevant to the work of OIE (including list of
scientific publications, internet publishing activities, presentations at international conferences)

Presentations at international conferences and meetings
1. Dubey S C. (2009) Member, Indian delegation to attend 77th General Session of OIE at Paris in which OIE
recognition for HSADL, IVRI, Bhopal as OIE reference laboratory for AI was declared. 24-29, May, Paris,
France (no presentation).
2. Kulkarni, D. D. (2009). WHO/OIE/FAO Biosafety and Laboratory biosecurity meeting for animal and human
health stake holders at OIE HQs Paris. Presented a paper entitled India’s High Security Animal Disease
Laboratory (BSL4 & ABSL4 facility). June 30th-2 July 09.
3. Tosh, C. and Nagarajan, S. (2009) both were deputed to participate and to give presentation on Role of High
Security Animal Disease Laboratory as reference laboratory for South Asia. Joint FAO/USDA regional meeting
on molecular epidemiology of origin and evolution of H5N1 highly pathogenic avian influenza virus in Asia,
Bangkok, Thailand. 09.09.2009-10.09.2009.
4. Dubey, S. C. (2009) OIE invitation for annual meeting of Laboratory Heads at Paris. Presentation on
country status of Avian influenza and Swine influenza in India. First participation after OIE recognition of
HSADL, IVRI, Bhopal. H5N1 during 15-16, Sept. Paris, France.
5. Sood, R., Swarup, D., Bhatia, S., Saini, M., Kulkarni, D.D., Dey, S., and Dubey, S.C.(2009) In-vitro antiviral
efficacies of indigenous herbs on Highly pathogenic Avian Influenza virus. ICAV 8th International Symposium on
Challenging Pandemics: Science funding and Policies. Corsica, France 3-6 Oct.
6. Murugkar, H. V. (2009) was nominated by GOI for FAO Asia Pacific for workshop cum training program on
surveillance of AI and pandemic swine influenza to harmonize the surveillance methodology between various
SEA countries & South Asian countries. 9-19 Dec. 09 at AAHL, Geelong, Australia.
7. Mishra, N. (2009) was nominated by GOI for FAO Asia Pacific workshop cum training program on Molecular
diagnostic protocols AI and pandemic swine influenza to harmonize the diagnostic methodology between various
SEA countries & South Asian countries. 9-19 December 09 at AAHL, Geelong, Australia.

Scientific publications in peer-reviewed journals
1. Nagarajan. S., Murugkar, H.V., Tosh, C., Behera, P., Jain, R., Tripathi, S., Khandia, R., Gupta, V., Kulkarni,
D.D., Dubey, S.C. (2009) Avian influenza virus (H5N1) in chickens in India. Vet Rec. January 24; 164(4): 128.
2. Nagarajan S, Rajukumar K, Tosh C, Ramaswamy V, Purohit K, Saxena G, Behera P, Pattnaik B, Pradhan HK,
Dubey SC. (2009) Isolation and pathotyping of H9N2 avian influenza viruses in Indian poultry. Vet. Microbiol.
133: 154-163.
3. Dubey, S.C., Nagarajan, S., Tosh, C., Bhatia, S., Lal Krishna (2009) Avian influenza: A long-known disease
and its current threat (Review). Ind. J. Anim. Sci. 79: 113-140.
Annual reports of OIE Reference Laboratories and Collaborating Centres, 2009
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Highly pathogenic avian influenza
4. Dubey, S. C. Venkatesh G. and Kulkarni D D. (2009). Epidemiological update on swine influenza (H1N1) in
pigs (Review). Indian Journal of Microbiology. 49 (12), 324-331, Springer India.
7. Gowthaman, V., Vanamayya, P. R., Nagarajan. S., Bhatia, S., Suba, S., Jain, R., Behera, P., Tosh, C.,
Murugkar, H. V. and Dubey, S.C. 2009. Influence of dose of inocula on outcome of clinical disease in highly
pathogenic avian influenza (H5N1) infections - An experimental study. Avian Dis. 53: xxx–xxx. (accepted).
8. Rai, M., Bhatia, S., Malik, Y.P.S., Dubey, S.C., (2009) Production and characterization of monoclonal
antibodies against NS1 protein of H5N1 avian influenza virus. Hybridoma (accepted).

Other communications
1. S. Suba, V.K. Saxena, S. Nagarajan, P.R. Vanamayya, V. Gowthaman, R. Jain, D.P. Singh and S.C. Dubey
(2009) An experimental investigation on resistance/ susceptibility to H5N1 virus in indigenous and high yielding
chickens of India. 7th International symposium on avian influenza: Avian influenza in poultry and wild birds.
Athens, Gergia, USA, 5-8 April, 2009. (Posters displayed during Session B)
2. V. Gowthaman, P.R. Vanamayya, S. Nagarajan, S. Bhatia, S. Suba, J. Rajlaxmi, B. Padmanav, C. Tosh, H.V.
Murugkar and S.C. Dubey (2009) Influence of dose of inocula on outcome of clinical disease in highly pathogenic
avian influenza (H5N1) infections – An experimental study. 7th Int. Sympo. on avian influenza: Avian influenza in
poultry and wild birds. Athens, Gergia, USA, 5-8 April, 2009. (Poster)
HSADL have sequenced and submitted 170 nucleotide sequence data to GenBank (NCBI,
http://blast.ncbi.nlm.nih.gov) of 22 avian influenza (20 H5N1 and 2 H9N2) viruses isolated from India under
the following accession numbers: GU252813-GU252834, FJ839922-FJ839923, GU083606-GU083665,
GU354056-GU354085, FJ839920-FJ839921, GU271995-GU272010, GU356580-GU356587, GU064-GU064952,
GU066386-GU066393, GQ478333, FJ966205-FJ966211 AND FJ605500-FJ605505.
13. Inscription of diagnostic kits on the OIE Register
i)
Did you participate in expert panels for the validation of candidate kits for inscription on the
OIE Register? If yes, for which kits?
No.
ii)
Did you submit to the OIE candidate kits for inscription on the OIE Register? If yes, for
which kits?
No.
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Annual reports of OIE Reference Laboratories and Collaborating Centres, 2009