VETERINARY REVIEW

EQUINE INFLUENZA 1987, POST-EPIDEMIC SEROLOGICAL STUDY IN NORTH INDIA

Gurkirpal Singh, M.V.Sc., PhD

SUMMARY

A post-epidemic (equine influenza) serological study of different regions of the Punjab State and Chandigarh (Union Territory) was undertaken. One hundred and fifty serum samples were collected from horses, ponies, mules and donkeys and examined in haemagglutination inhibi- tion tests. The results indicated that 87.5% to 100% had antibodies to A/equi-1 virus and 61.9 to 88.6% to A/equi- 2 virus. Evidently, both the serotypes were widely preva- lent during the epidemic throughout the Punjab State and Chandigarh. The involvement of both serotypes of equine influenza virus in the same population was unusual.

INTRODUCTION

The epidemic of equine influenza 1987, which oc- curred in Northern and Central India in horses, ponies, etc., was due to virus A/equi-2 serotype 1,e and A-equi-1 serotype isolated in the virus laboratory of this depart- ment. In this part of India, the disease was first noted in a stable in Chandigarh (Union Territory) in the latter part of February, 1987. Within a few weeks, it was detected in all districts in the Punjab State and affected all breeds and age groups. This communication records, in brief, serological studies on equine sera collected post-epidemic from the Punjab State and Chandigarh in order to study the circula- tion of equine influenza type 1 and type 2 viruses during 1987 outbreaks.

MATERIALS AND METHODS

Sera Serum samples were obtained from 150 horses, ponies,

mules and donkeys of mixed ages, from various regions in the Punjab State and Union Territory of Chandigarh. All the animals had a history of having contacted the clinical infec-

Author's address: Senior Virologist-cum-Head, Department of Veterinary Bacteriology and virology, Punjab Agricultural U niversity, Ludhiana - 141004, India,

tion, usually with coughing. Signs of illness occurred only once in all animals except few where respiratory signs were reported to have recurred. There was no record of influenza vaccination for any of these animals. Serawere stored at-18°C.

All sera were heat-inactivated and were treated with potassium periodate to remove non-specific inhibitors be- fore being examined by HI tests.

Viruses The standard virus strains used in the HI tests were A/

equine/Prague/I/56 and A/equine/Miami/i/63. The isolates recovered from the 1987 outbreak, and

designated as A/equi-1/Ludhiana/5/87 and A/equi-2/Lu- dhiana/8/87, were used.

All of these strains were propagated in the allantoic cavity of chicken embryonated eggs. The pooled allantoic and amniotic fluids harvested were clarified by low speed centrifugation and were treated with ether prior to use?

Haemagglutination inhibition (HI) tests The HI tests were carried out in round-bottomed micro-

titre plates using 0.05 ml volumes of reactants as per the procedures described. 4 Four haemagglutinating units of vi- rus antigens were employed. Adult fowl red blood cells (0.5%) were used, with phosphate buffered saline as diluent.

RESULTS AND DISCUSSION

It will be seen from the table that HI antibodies to A/ equi-1 and A/equi-2 viruses were detected in sera from equine population in different regions in the Punjab State and in Chandigarh. The positive rate varied from 87.5% to 100% for A/equi-1 antibodies and 61.9% to 88.6% for A/equi-2 antibodies. Although not tabulated, the HI results with the A/ Miami/I/63 strain on 70 serum specimens tested were almost similar to those with local A/Ludhiana/8/87 strain.

The results indicated that both types of equine influenza virus had spread throughout the Punjab State and Chandi- garh. The spread of infection was expected since equine influenza has not been identified by clinical, serological or virological data since last report of clinically apparent cases of illness in 1964 in India, 5,e and horse population had not been vaccinated against equine influenza.

The demonstration of antibodies to A/equi-1 and A/ equi-2 viruses in post-epidemic sera as presented above, the pattern of antibody response in serial sera from affected animals to be reported elsewhere, or isolation ofA/equi- 1 and A/equi-2 viruses from animals in the acute phase of the disease from the outbreak, and evident clinical history, indicate that both types of equine influenza virus were active in different regions in the Punjab State and Chandigarh

342 JOURNAL OF EQUINE VETERINARY SCIENCE

Table. HI antibodies for both serotypes of equine influenza virus in serum specimens from equines in Punjab State and Chandigarh.

Sera positive Place Sers A/equine/ A/equi-1/ A/equi-2/

tested Prague/56 Ludhiana] Ludhiana] (HTN7) 5/87 (H7N7) 8/87 (H3N8)

*Northern region bGurdaspur & Hoshiarpur 27 27"(100) 27(100) 21 (77.7) districts, etc.

*Southern region* CBhatinda district 12 11 (91.6) 12(100) 9(75.0)

*Eastern region bRupnagar & Patiala 29 29(100) 29(10(3) 23(79.3)

districts

*Western region bFerozepur & dFaridkot 21 19(90.4) 19(90.4) 13(61.9)

districts

*Central region bLudhiana & Jalandhar 53 51 (96.2) 53(100) 47(88.6)

districts

bChandigarh (Union Territory) 8 7(87.5) 7(87.5) 5(62.5)

Total 150 144 (96.0) 147 (98.0) 118(78.6)

The age of animals was 3-15 years or older. aArbitrary division. bTime of collection of sere was August-September, 1987. i.e. 5-6 months after illness. CSera were collected in January, 1988. dSera were collected in May and August, 1989. *Denotes the number of sere with HI antibody titre of ~20. HI antibodies ranges from 20 to 320. Figures in parentheses are percentages. Nequi-2/Ludhiana/8/87 isolate closely resembles the prototype H3N8 virus Miami/63.

(Union Territory) causing, in all probability, simultaneous infection or infections one after the other during incubation/ clinical illness in the equine population. Though equine influenza type 1 and 2 viruses are known to cause similar clinical signs and presumably would compete for the same site of replication, the detailed clinical picture and tropisms in dual infection with both serotypes need to be further studied.

After the isolation of A/equi-1 in this laboratory, and its final confirmation by Dr. J. J. Skehel, World Reference Centre, National Institute for Medical Research, London, or the demonstration of HI antibodies to A/equi-1 and A/equi-2 viruses in post-infection sera, Rao reported the presence of antibodies to A/equine-1 and A/equine-2 virus strains in horses and mules brought to Pune from the Northern States of India. 7 Earlier studies have reported widespread prevalence of A/equi-2 antibodies among the equine population in North- ern India. a A small number of serum samples were also positive for A/equi-1 antibodies. Elsewhere, studies reported prevalence of both serotypes of equine influenza in the same stud. 9

Evidently, the studies carried out in this laboratory have uncovered the aetiology of the equine influenza epidemic of 1987 earlier considered to be due to A/equi-2 virus. The involvement of A/equi-1, which is unusual, needs to be kept in mind in investigations of future outbreaks of equine influ- enza which may occur anywhere in the world. It is of interest to explore as to whether A/equi-1 virus has been going on obscurely along with type 2 equine influenza infection or in vaccinated horses. After all, some horses, supposedly vacci- nated, which were imported from abroad during January- February, 1987 and which were believed to have introduced the infection in India, could have been carrying one or both serotypes at that time.

REFERENCES

1. Uppal PK, Yadav MP, Sharma SN: Occurrence of equine influenza outbreaks in India. Indian J Comp Microbio/ Immunol Infect Dis 8:91-94,1987.

Volume 12, Number 6, 1992 343

2. Singh G, Oberoi MS, Kwatra MS, Gill SS: Isolation of influenza virus from horses in the equine influenza outbreak of 1987. Curt Sci 56:1285-1286,1987.

3. Berlin BS, McQueen JL, Minuse E, Davenport FM: A method for increasing the sensitivity of the haemagglutination inhibition test with equine influenza virus. Virol 21:665-666,1965.

4. Kendal AP, Pereira MS, Skehel JJ: Concepts andproce- dures for laboratory-based influenza surveillance. United States Department of Health and Human Services, Public Health Service, Centers for Disease Control. B 17-29,1982.

5. Feldman RA: Personal communication. (Cited by Mc- Queen JL, Steele JH, and Robinson RQ, 1967.) Influenza in animals. Advances in Vet Sci 12:299,1965.

6. Singh BS: Personal communications. (Cited by Mc- Queen JL, Steele JH, and Robinson RQ, 1967.) Influenza in animals. Advances in Vet Sci 12:299,1965.

7. Rao BL, Kadam SS, Johi MV: Serological investigation on equine influenza in Pune, Maharashtra State, India. Indian Vet J 69:103-106,1992.

8. Uppal PK, Yadav MP, Suneja SK: Virus Information Exchange News/etter for South East Asia and the Western Pacific 4:45,1987.

9. Burki F: Proceedings 1 lth Intl Symp World Assoc Vet Microb, Immuno & Sp Infect Dis, Perugia. P279,1989.

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