- Email: [email protected]
Isolation of influenza viruses from avian species in Great Britain
Comp.lmmun.Microbiol.infect.Dis.,Vol.3, pp. 165-170. PergamonPressLtd., 1980.Printedin GreatBritain
ISOLATION OF INFLUENZA VIRUSES FROM AVIAN SPECIES IN GREAT BRITAIN D. J. ALEXANDER Poultry Department, Central Veterinary Laboratory, New Haw, Weybridge, Surrey KT15 3NB, U.K.
Abstract--Avian influenza viruses have been isolated in Great Britain in recent years from four sources: (1) Dead birds arriving at Heathrow (London) Airport Between 1976 and 1978 62 influenza viruses were isolated, 48 of Hav7 Neq2 subtype and 14 of Hav4 (all those tested are Navl) subtype. (2) Birds hem in quarantine Between March 1976 and February 1979 30 influenza A viruses were isolated from this source. Eight had Hav7Neq2 antigens, 21 Hav4 (all those tested Navl) and one Hav2Neq I antigens. (3) Domestic poultry During 1963--1978 seven influenza isolations were made from turkeys: A/ty/England/63(Havl Nav3), A/ty/England/66(Hav6 N2), A/ty/England/69(Hav7/H3 N2), A/ty/Scofland/70(Hav? Neql), A/ty/England/N28/73 (Hay5 N2), A/ty/England/l10/77 (Hav6 N2), A/ty/England/ 647/77(Hay 1 Neql). In the early Spring of 1979 several outbreaks of influenza infections occurred on turkey farms in Norfolk. Virus isolates were predominantly of Havl subtype in combination with Nav2, N2 and Neq 1 but a virus of Hswl N1 subtype was also isolated from turkeys. The HavlNeql isolates were virulent for chickens and turkeys. (4) Wild birds An influenza virus of Hav6 Neq2 subtype was isolated from a Canada goose. Key words: Isolation, influenza A, avian, Great Britain, airport, quarantine, poultry, wild
ISOLEMENT DE VIRUS DE GRIPPE AVIAIRE A PARTIR D'ESPECES AVIAIRES EN G R A N D E - B R E T A G N E Resume--Les virus de grippe aviaire ont 6t6 isol6s en Grande-Bretagne ces ann6es derni6res partir de quatre sources: (1) Des oiseaux morts d leur arrivde d l'adroport de Heathrow (Londres). Entre 1976 et 1978, 62 virus de grippe aviaire ont &6 isol6s, dont 48 du sous-type Hav7Neq2 et 14 du sous-type Hav4 (tous ceux 6prouv6s &aient Nav 1). (2) Des oiseaux ddtenus en quarantaine Entre mars 1976 et f6vder 1979, 30 virus de grippe aviaire A ont &~ isol~s/t partir de cette source. Huit avaient des antig6nes Hav7Neq2, 21, Hay4 (tous ceux 6prouv~s, Navl), et un avait des anti#nes Hav2Neq 1. (3) Volailles domestiques Pendant la p6riode 1963-1978, la grippe aviaire a 6t6 isol6e sept fois chez des dindes: A/ ty/England/63 (HavlNav3), A/ty/England/66 (Hav6N2), A/ty/England/69 (Hav7/H3N2), A / t y / Scotland/70 (Hav?Neql), A/ty/England/N28/73 (Hav5N2), A/ty/England/110/77 (Hav6N2), A/ty/England/647/77 (Hay 1Neq 1). Au commencement du printemps 1979, plusieurs foyers de grippe aviaire se sont d6clar6s darts des 61evages de dindes dans le Norfolk. Les isolats viraux &aient principalement du sous-type Hav 1 en combinaison avec Nay2, N2 ~ Neql, mais un virus de sous-type H s w l N l a 6t6 6galement isol6 chez les dindes. Les isolats Hay 1Neq I se montraient virulents pour les poulets et les dindes. (4) Oiseaux sauvages Un virus de grippe aviaire du sous-type Hav6Neq2 a 6t6 isol6 ehez une bernache du Canada. Mots-clefs: Isolement, grippe aviaire A, aviaire, Grande-Bretagne, a~roport, quarantaine, volaiUes, sauvage 165
166
D.J. AI.EXANDER INTRODUCTION
In recent years numerous surveillance studies have been made throughout the World to examine the presence of influenza viruses in avian species due to the possible importance of animal influenza viruses in the emergence of viruses causing pandemics amongst humans [ 11. In Great Britain the presence of influenza virus has been monitored in birds from four sources: (1) Birds in transit at Heathrow (London) Airport, (2) imported captive birds held in quarantine, (3) domestic poultry, (4) wild birds. The object of this paper is to describe the influenza viruses isolated. MATERIALS AND METHODS
Virus isolations Viruses were isolated from tracheal and cloacal swabs of dead birds from Heathrow Airport, pools being made from all the dead birds in each consignment as described [2]. Viruses isolated from birds in quarantine or domestic turkeys were from suspensions of organs or cloacal swabs. All isolations were made by inoculation of material into the allantoic cavity of 9- or 10-dayold embryonated fowls' eggs.
Virus characterization Serotyping of isolates by haemagglutination inhibition and neuraminidase inhibition tests using chicken sera against reference strains and pathogenicity index characterization in sixweek-old chickens and turkeys was as described in 13]. One isolate, A/turkey/England/250/79 (Hswl N1) was also kindly typed by Dr. J. J. Skehel, WHO Influenza Centre, Mill Hill, London. RESULTS
1. Isolation of influenza viruses from dead birds arriving at Heathrow Airport Between May 1976 and October 1978 the dead birds from 307 consignments arriving in transit at Heathrow Airport were examined by tracheal and cloacal swabbing and 67 haemagglutinating agents isolated. Five isolates were shown to be Newcastle disease virus (NDV) and 62 were influenza A viruses. Forty-eight of the influenza viruses were identified as Hav7 Neq2 and 14 Hav4 (all seven typed so far have Navl neuraminidase). Only influenza viruses with Hav7 Neq2 antigens were isolated between May 1976 and March 1977 and only Hav4 viruses have been isolated since March 1977. Where known, the airports of origin of consignments positive for influenza virus have been exclusively in India. However the consignments have been bound for 14 different European airports representing nine different countries.
2. Isolations from captive birds in quarantine following importation into Great Britain Birds that die during the compulsory quarantine/isolation period of 35 days after importation into Great Britain are examined for haemagglutinating viruses. Between March 1976 and February 1979 62 viruses were isolated. Thirty-two were paramyxoviruses
167
Avian influenza in Great Britain
consisting of 21 NDV, six related to PMV/chicken/California/Yucalpa/56 and five to PMV/parakeet/Netherlands/449/75. Thirty were influenza A viruses, eight of Hav7 Neq2 subtype, 21 Hay4 N a v l and one Hay2 Neql. As with viruses isolated from Heathrow Airport, no Hav7 Neq2 viruses have been isolated since Spring 1977. The majority of isolations of influenza viruses were made from psittacine species or finches exported from India but the practice of re-exporting birds from intermediate countries made it difficult to ascertain the country of origin of many of the birds.
3. Isolation of influenza viruses from domestic poultry a. 1963-1978. Only seven influenza viruses were isolated from domestic poultry between 1963 and 1978, all were from turkeys and are listed in Table 1. As indicated by the pathogenicity indices, only one virus, A/ty/Eng./63, showed high virulence for turkeys producing the clinical signs and deaths typical of classical fowl plague. Another isolate A/turkey/Eng./69 produced some deaths and signs in experimentally infected birds but all other isolates were apparently avirulent in these hosts. A/ty/Eng./69 haemagglutinin and neuraminidase showed close relationships with human H3 and N2 viruses (A/Hong Kong/68) and may represent the isolation of human virus from turkeys. A/ty/Scot./70 appeared to possess a distinct haemagglutinin subtype. b. 1979. In 1979 a completely different pattern was seen compared to that of other years. Between March and May as many infections of turkeys with influenza virus were confirmed as had been reported over the previous 16 years. The characterization of the isolates obtained are shown in Table 2. On two closely situated sites (5 and 8), belonging to the same owner, the turkeys had severe clinical disease which was diagnosed as 'fowl plague' and, under Government legislation, these birds were slaughtered. In both cases the virus isolated was typed as H a v l Neq 1. At about the same time influenza viruses of low virulence were isolated from turkeys with much milder disease, not diagnosed as 'fowl plague', at other sites (3 and 4). The two sites (3 and 7) from which Hav 1 Nav2 was isolated were less then 5 km apart although different owners were involved. Another site (4) 15 km away was infected with H a v l N 2 Table 1. Influenzaviruses isolated from turkeys in Great Britain during 1963-1978 Pathogenicity: IVPIt in:
Serotypes* Isolate
County
HA
N
turkeys
chickens
A/ty/Eng/63 A/tyEng/66 A/ty/Eng/69 A/ty/Scotland/70 A/ty/Eng/N28/73 A/ty/Eng/110/77 A/ty/Eng/647/77
Norfolk Norfolk Norfolk
Havl Hay6 Hav7(H3) Hay? Hay5 Hav6 Hav I
Nay3 N2 N2 Neq 1 N2 N2 Neq 1
2.78 0.04 0.15 0.00 0.00 0.00 0.00
2.97 0.00 0.81 0.00 0.00 0.00 0.00
Norfolk Norfolk Hereford
*Viruses weretyped usinghaemagglutinin(HA) and neuraminidase(N) inhibition tests. ~'IVPI: The intravenous pathogenicity index is estimated from the time taken for six-week-oldbirds to die or show disease signs after intravenous inoculation of virus. The results are based on a scoring system in which the maximum index possible is 3.00 (10096 dead in one day) and the minimum 0.00 (no recorded signs during the I0-day observationperiod).
168
D. J. ALEXANDER
Table 2. Influenza viruses isolated from turkeys in Great Britain during March-May 1979 Serotypes* Isolate
County site
HA
N
A/ty/Eng/328/79 A/ty/Eng/329/79 A/ty/Eng/199/79 A/ty/Eng/214A/79 A/ty/Eng/214F/79 A/ty/Eng/214K/79 A/ty/Eng/237/79 A/ty/Eng/250/79 A/ty/Eng/262/79 A/ty/Eng/272/79 A/ty/Eng/384/79
Norfolk-3 Norfolk-4 Norfolk-5a II Norfolk-5b II Norfolk-5b Norfolk-5b Norfolk-3 ¶ Norfolk-6 Norfolk-7 Norfolk-8 Hertford
Hav I Hav I Hay I Hay 1 Hay I Hav I Hay 1 Hsw I Hav I Hay 1 Hav2
Nav2§ N2 Neq I Neq I Neq I Neq I Nay2§ N1 Nav2 § Neq I Nav4
Pathogenicity: IVPI~:in: chickens 0.00 0.16 3.00 2.72 3,00 2.76 0.00 0.00 0.12 2.49 1.34
All isolations are listed in chronological order and were made within a three-week period over the end of March and beginningof April with the exception of 384/79 which was isolated in early May. *~: See footnotes to Table 1. § Virusestyped as Nav2 also showed inhibitionwith Nay3 antiserum. It 5a and 5b represent different farms on the same site. 214A, 214F and 214K are different isolates from the same flock. ¶ 328/79and 237/79 were isolated from the same site two weeks apart.
virus. A further site in Norfolk was infected with H s w l N1 virus and there was serological evidence on two sites of infection of turkeys with virus of H a v l subtype and one site with Hav2. Two other influenza outbreaks in turkeys were seen in other counties. One was in Hertfordshire from which a virus typed as Hav2 Nav4 was isolated and serological evidence of an Hav 1 infection was obtained from a farm in Suffolk near the Norfolk border. In the latter case there was no clinical evidence of disease and no virus was isolated.
4. Wild birds Studies on wild birds in Great Britain have been limited but one influenza isolation has been made in April 1979 from a C a n a d a goose: A / C a n a d a goose/England/263/79 (Hav6 Neq2) which was avirulent for chickens (IVPI 0.00). DISCUSSION The possible role played by avian species in the origins of h u m a n pandemic influenza virus has been well documented [for examples see 1 and 4]. It is therefore of considerable interest that such a high incidence of virus isolation should be made from birds in transit at Heathrow Airport and those held in quarantine. Although only dead birds were examined in these studies there was no evidence to suggest that the influenza viruses were responsible for the deaths and it seems reasonable to assume that the surviving birds in the consignments may have been infected and carried the viruses to their destinations. The likelihood of such spread and the fact that the majority of the isolates were of Hav7 subtype which has been shown to be related to the H3 subtype [1] emphasises the significance of these isolations. A further point concerning the isolations from Heathrow Airport and birds in quarantine was the change in serotype of the isolates from Hav7 to Hav4 which occurred in the early
170
D.J. ALEXANDER
3. Alexander, D. J., Allan, W. H. and Parsons, G., Characterisation of influenza viruses isolated from turkeys in Great Britain during 1963-1977, Res. vet. Sci. 26, 17-20 (1979). 4. Laver, W. G. and Webster, R. G., Ecology of influenza viruses in lower mammals and birds, Br. med. Bull. 35, 29-33 (1979). 5. Alexander, D. J., Allan, W. H., Harkness, J. W. and Hall, S. A. Isolation of influenza virus from psittaeines, Res. vet. Sci. 17, 125-127 (1974). 6. Fukumi, H., Nerome, K., Nakayama, M. and Ishida, M., Serological and virological investigations of orthomyxovirus in birds in south-east Asian area, Dev. biol. Standard. 39, 457-460 (1977). 7. Nerome, K., Nakayama, M., Ishida, M., Fukumi, H., Butterfield, W. K., Webster, R. G. and Campbell, C. H., Isolation and serological characterization of influenza A viruses from birds that were dead on arrival at Tokyo Airport, Archs Virol. 57, 261-270 (1978).
Avian influenza in Great Britain
169
Spring of 1977. It could be that the Indian exporters merely altered their source of birds, but this seems unlikely. Influenza viruses isolated from captive birds imported into Great Britain up to 1974 were of Hav4 subtype [5, Chu cited in 6] whereas influenza viruses isolated from caged birds imported into Japan in 1976 were exclusively Hav7 from India but Hav7 and Hav4 from South East Asian countries [6, 7]. It has been suggested that from time to time epizootics may occur amongst birds in India and S.E. Asia causing changes in the predominant subtype isolated [6]. The results obtained in our studies suggest that since early 1977 viruses of Hav4 subtype have re-emerged as the dominant influenza subtype in the birds of the Indian sub-continent. The number of isolations of influenza viruses from turkeys between 1963 and 1978 was indicative of the state of influenza in domestic poultry in Great Britain during that time. There was no evidence of influenza viruses being endemic amongst domestic or wild birds and it seemed most likely that the occasional outbreaks of viruses of low virulence were the result of fresh introductions, presumably by migratory birds [3]. The outbreaks and isolations in 1979 were therefore quite unusual. The exact origins of these outbreaks are unknown but it would seem most likely that the viruses were introduced into the turkey flocks by wild birds. The very severe winter, both in Great Britain and on the continent of Europe had led to unusual circumstances in the bird populations, particularly in Norfolk. Most noticeable were the huge flocks of starlings seen in this area, some of which were estimated as containing up to 2,000,000 birds. The high winds and heavy snowfalls of the winter had in many cases caused damage to turkey houses allowing access to wild birds. The affected area is on the migratory routes of several species of waterfowl but starlings were obviously the most popular candidates for the responsibility of immediate transmission to turkeys. It is therefore of interest that in 1978 an influenza virus with H a v l Neql antigens was isolated from starlings in Israel (Lipkind, personal communication) this is the only known isolation of influenza virus from starlings. In Great Britain there is legislation for the slaughter of birds infected with highly virulent influenza viruses. The two sites on which virulent virus was present were therefore slaughtered out and no spread of the virus to other sites has been detected. Neither has any evidence been obtained of spread from any of the premises affected by virus of low virulence despite the close proximity of many farms with apparently susceptible birds. The severity of the 1978-1979 winter apparently had much to do with number of outbreaks of influenza virus infections of turkeys both in its affect on the behaviour of wild birds and damage to the turkey houses. It is therefore notable that the last previous isolation of virulent influenza virus in Great Britain: A/ty/Eng./63 (Hav l Nay3), followed the only winter of comparable severity to that of 1978-1979 and was made from turkeys in the same locality in Norfolk as those affected with virulent virus this year. Acknowledgments-The isolations and identification of influenza viruses from birds in Great Britain which are
summarized in this paper were the result of work done by officers of the State Veterinary Service, private veterinarians, staff of the RSPCA Hostel at Heathrow Airport and, in particular, members of the Poultry Department, Central Veterinary Laboratory, Weybridge. REFERENCES 1. Webster, R. G. and Laver, W. G., Antigenic variation of influenza viruses. In The influenza Viruses and Influenza, Kilbourne, E.D. (ed.), pp. 209-314. Academic Press, New York and London (1975). 2. Alexander, D. 3., Allan, W. H. and Sillars, T., Isolation of myxovirusesfrom dead birds arriving at Heathrow Airport, London, J. Hyg., Camb. 79, 243-247 (1977).
ISOLATION OF INFLUENZA VIRUSES FROM AVIAN SPECIES IN GREAT BRITAIN D. J. ALEXANDER Poultry Department, Central Veterinary Laboratory, New Haw, Weybridge, Surrey KT15 3NB, U.K.
Abstract--Avian influenza viruses have been isolated in Great Britain in recent years from four sources: (1) Dead birds arriving at Heathrow (London) Airport Between 1976 and 1978 62 influenza viruses were isolated, 48 of Hav7 Neq2 subtype and 14 of Hav4 (all those tested are Navl) subtype. (2) Birds hem in quarantine Between March 1976 and February 1979 30 influenza A viruses were isolated from this source. Eight had Hav7Neq2 antigens, 21 Hav4 (all those tested Navl) and one Hav2Neq I antigens. (3) Domestic poultry During 1963--1978 seven influenza isolations were made from turkeys: A/ty/England/63(Havl Nav3), A/ty/England/66(Hav6 N2), A/ty/England/69(Hav7/H3 N2), A/ty/Scofland/70(Hav? Neql), A/ty/England/N28/73 (Hay5 N2), A/ty/England/l10/77 (Hav6 N2), A/ty/England/ 647/77(Hay 1 Neql). In the early Spring of 1979 several outbreaks of influenza infections occurred on turkey farms in Norfolk. Virus isolates were predominantly of Havl subtype in combination with Nav2, N2 and Neq 1 but a virus of Hswl N1 subtype was also isolated from turkeys. The HavlNeql isolates were virulent for chickens and turkeys. (4) Wild birds An influenza virus of Hav6 Neq2 subtype was isolated from a Canada goose. Key words: Isolation, influenza A, avian, Great Britain, airport, quarantine, poultry, wild
ISOLEMENT DE VIRUS DE GRIPPE AVIAIRE A PARTIR D'ESPECES AVIAIRES EN G R A N D E - B R E T A G N E Resume--Les virus de grippe aviaire ont 6t6 isol6s en Grande-Bretagne ces ann6es derni6res partir de quatre sources: (1) Des oiseaux morts d leur arrivde d l'adroport de Heathrow (Londres). Entre 1976 et 1978, 62 virus de grippe aviaire ont &6 isol6s, dont 48 du sous-type Hav7Neq2 et 14 du sous-type Hav4 (tous ceux 6prouv6s &aient Nav 1). (2) Des oiseaux ddtenus en quarantaine Entre mars 1976 et f6vder 1979, 30 virus de grippe aviaire A ont &~ isol~s/t partir de cette source. Huit avaient des antig6nes Hav7Neq2, 21, Hay4 (tous ceux 6prouv~s, Navl), et un avait des anti#nes Hav2Neq 1. (3) Volailles domestiques Pendant la p6riode 1963-1978, la grippe aviaire a 6t6 isol6e sept fois chez des dindes: A/ ty/England/63 (HavlNav3), A/ty/England/66 (Hav6N2), A/ty/England/69 (Hav7/H3N2), A / t y / Scotland/70 (Hav?Neql), A/ty/England/N28/73 (Hav5N2), A/ty/England/110/77 (Hav6N2), A/ty/England/647/77 (Hay 1Neq 1). Au commencement du printemps 1979, plusieurs foyers de grippe aviaire se sont d6clar6s darts des 61evages de dindes dans le Norfolk. Les isolats viraux &aient principalement du sous-type Hav 1 en combinaison avec Nay2, N2 ~ Neql, mais un virus de sous-type H s w l N l a 6t6 6galement isol6 chez les dindes. Les isolats Hay 1Neq I se montraient virulents pour les poulets et les dindes. (4) Oiseaux sauvages Un virus de grippe aviaire du sous-type Hav6Neq2 a 6t6 isol6 ehez une bernache du Canada. Mots-clefs: Isolement, grippe aviaire A, aviaire, Grande-Bretagne, a~roport, quarantaine, volaiUes, sauvage 165
166
D.J. AI.EXANDER INTRODUCTION
In recent years numerous surveillance studies have been made throughout the World to examine the presence of influenza viruses in avian species due to the possible importance of animal influenza viruses in the emergence of viruses causing pandemics amongst humans [ 11. In Great Britain the presence of influenza virus has been monitored in birds from four sources: (1) Birds in transit at Heathrow (London) Airport, (2) imported captive birds held in quarantine, (3) domestic poultry, (4) wild birds. The object of this paper is to describe the influenza viruses isolated. MATERIALS AND METHODS
Virus isolations Viruses were isolated from tracheal and cloacal swabs of dead birds from Heathrow Airport, pools being made from all the dead birds in each consignment as described [2]. Viruses isolated from birds in quarantine or domestic turkeys were from suspensions of organs or cloacal swabs. All isolations were made by inoculation of material into the allantoic cavity of 9- or 10-dayold embryonated fowls' eggs.
Virus characterization Serotyping of isolates by haemagglutination inhibition and neuraminidase inhibition tests using chicken sera against reference strains and pathogenicity index characterization in sixweek-old chickens and turkeys was as described in 13]. One isolate, A/turkey/England/250/79 (Hswl N1) was also kindly typed by Dr. J. J. Skehel, WHO Influenza Centre, Mill Hill, London. RESULTS
1. Isolation of influenza viruses from dead birds arriving at Heathrow Airport Between May 1976 and October 1978 the dead birds from 307 consignments arriving in transit at Heathrow Airport were examined by tracheal and cloacal swabbing and 67 haemagglutinating agents isolated. Five isolates were shown to be Newcastle disease virus (NDV) and 62 were influenza A viruses. Forty-eight of the influenza viruses were identified as Hav7 Neq2 and 14 Hav4 (all seven typed so far have Navl neuraminidase). Only influenza viruses with Hav7 Neq2 antigens were isolated between May 1976 and March 1977 and only Hav4 viruses have been isolated since March 1977. Where known, the airports of origin of consignments positive for influenza virus have been exclusively in India. However the consignments have been bound for 14 different European airports representing nine different countries.
2. Isolations from captive birds in quarantine following importation into Great Britain Birds that die during the compulsory quarantine/isolation period of 35 days after importation into Great Britain are examined for haemagglutinating viruses. Between March 1976 and February 1979 62 viruses were isolated. Thirty-two were paramyxoviruses
167
Avian influenza in Great Britain
consisting of 21 NDV, six related to PMV/chicken/California/Yucalpa/56 and five to PMV/parakeet/Netherlands/449/75. Thirty were influenza A viruses, eight of Hav7 Neq2 subtype, 21 Hay4 N a v l and one Hay2 Neql. As with viruses isolated from Heathrow Airport, no Hav7 Neq2 viruses have been isolated since Spring 1977. The majority of isolations of influenza viruses were made from psittacine species or finches exported from India but the practice of re-exporting birds from intermediate countries made it difficult to ascertain the country of origin of many of the birds.
3. Isolation of influenza viruses from domestic poultry a. 1963-1978. Only seven influenza viruses were isolated from domestic poultry between 1963 and 1978, all were from turkeys and are listed in Table 1. As indicated by the pathogenicity indices, only one virus, A/ty/Eng./63, showed high virulence for turkeys producing the clinical signs and deaths typical of classical fowl plague. Another isolate A/turkey/Eng./69 produced some deaths and signs in experimentally infected birds but all other isolates were apparently avirulent in these hosts. A/ty/Eng./69 haemagglutinin and neuraminidase showed close relationships with human H3 and N2 viruses (A/Hong Kong/68) and may represent the isolation of human virus from turkeys. A/ty/Scot./70 appeared to possess a distinct haemagglutinin subtype. b. 1979. In 1979 a completely different pattern was seen compared to that of other years. Between March and May as many infections of turkeys with influenza virus were confirmed as had been reported over the previous 16 years. The characterization of the isolates obtained are shown in Table 2. On two closely situated sites (5 and 8), belonging to the same owner, the turkeys had severe clinical disease which was diagnosed as 'fowl plague' and, under Government legislation, these birds were slaughtered. In both cases the virus isolated was typed as H a v l Neq 1. At about the same time influenza viruses of low virulence were isolated from turkeys with much milder disease, not diagnosed as 'fowl plague', at other sites (3 and 4). The two sites (3 and 7) from which Hav 1 Nav2 was isolated were less then 5 km apart although different owners were involved. Another site (4) 15 km away was infected with H a v l N 2 Table 1. Influenzaviruses isolated from turkeys in Great Britain during 1963-1978 Pathogenicity: IVPIt in:
Serotypes* Isolate
County
HA
N
turkeys
chickens
A/ty/Eng/63 A/tyEng/66 A/ty/Eng/69 A/ty/Scotland/70 A/ty/Eng/N28/73 A/ty/Eng/110/77 A/ty/Eng/647/77
Norfolk Norfolk Norfolk
Havl Hay6 Hav7(H3) Hay? Hay5 Hav6 Hav I
Nay3 N2 N2 Neq 1 N2 N2 Neq 1
2.78 0.04 0.15 0.00 0.00 0.00 0.00
2.97 0.00 0.81 0.00 0.00 0.00 0.00
Norfolk Norfolk Hereford
*Viruses weretyped usinghaemagglutinin(HA) and neuraminidase(N) inhibition tests. ~'IVPI: The intravenous pathogenicity index is estimated from the time taken for six-week-oldbirds to die or show disease signs after intravenous inoculation of virus. The results are based on a scoring system in which the maximum index possible is 3.00 (10096 dead in one day) and the minimum 0.00 (no recorded signs during the I0-day observationperiod).
168
D. J. ALEXANDER
Table 2. Influenza viruses isolated from turkeys in Great Britain during March-May 1979 Serotypes* Isolate
County site
HA
N
A/ty/Eng/328/79 A/ty/Eng/329/79 A/ty/Eng/199/79 A/ty/Eng/214A/79 A/ty/Eng/214F/79 A/ty/Eng/214K/79 A/ty/Eng/237/79 A/ty/Eng/250/79 A/ty/Eng/262/79 A/ty/Eng/272/79 A/ty/Eng/384/79
Norfolk-3 Norfolk-4 Norfolk-5a II Norfolk-5b II Norfolk-5b Norfolk-5b Norfolk-3 ¶ Norfolk-6 Norfolk-7 Norfolk-8 Hertford
Hav I Hav I Hay I Hay 1 Hay I Hav I Hay 1 Hsw I Hav I Hay 1 Hav2
Nav2§ N2 Neq I Neq I Neq I Neq I Nay2§ N1 Nav2 § Neq I Nav4
Pathogenicity: IVPI~:in: chickens 0.00 0.16 3.00 2.72 3,00 2.76 0.00 0.00 0.12 2.49 1.34
All isolations are listed in chronological order and were made within a three-week period over the end of March and beginningof April with the exception of 384/79 which was isolated in early May. *~: See footnotes to Table 1. § Virusestyped as Nav2 also showed inhibitionwith Nay3 antiserum. It 5a and 5b represent different farms on the same site. 214A, 214F and 214K are different isolates from the same flock. ¶ 328/79and 237/79 were isolated from the same site two weeks apart.
virus. A further site in Norfolk was infected with H s w l N1 virus and there was serological evidence on two sites of infection of turkeys with virus of H a v l subtype and one site with Hav2. Two other influenza outbreaks in turkeys were seen in other counties. One was in Hertfordshire from which a virus typed as Hav2 Nav4 was isolated and serological evidence of an Hav 1 infection was obtained from a farm in Suffolk near the Norfolk border. In the latter case there was no clinical evidence of disease and no virus was isolated.
4. Wild birds Studies on wild birds in Great Britain have been limited but one influenza isolation has been made in April 1979 from a C a n a d a goose: A / C a n a d a goose/England/263/79 (Hav6 Neq2) which was avirulent for chickens (IVPI 0.00). DISCUSSION The possible role played by avian species in the origins of h u m a n pandemic influenza virus has been well documented [for examples see 1 and 4]. It is therefore of considerable interest that such a high incidence of virus isolation should be made from birds in transit at Heathrow Airport and those held in quarantine. Although only dead birds were examined in these studies there was no evidence to suggest that the influenza viruses were responsible for the deaths and it seems reasonable to assume that the surviving birds in the consignments may have been infected and carried the viruses to their destinations. The likelihood of such spread and the fact that the majority of the isolates were of Hav7 subtype which has been shown to be related to the H3 subtype [1] emphasises the significance of these isolations. A further point concerning the isolations from Heathrow Airport and birds in quarantine was the change in serotype of the isolates from Hav7 to Hav4 which occurred in the early
170
D.J. ALEXANDER
3. Alexander, D. J., Allan, W. H. and Parsons, G., Characterisation of influenza viruses isolated from turkeys in Great Britain during 1963-1977, Res. vet. Sci. 26, 17-20 (1979). 4. Laver, W. G. and Webster, R. G., Ecology of influenza viruses in lower mammals and birds, Br. med. Bull. 35, 29-33 (1979). 5. Alexander, D. J., Allan, W. H., Harkness, J. W. and Hall, S. A. Isolation of influenza virus from psittaeines, Res. vet. Sci. 17, 125-127 (1974). 6. Fukumi, H., Nerome, K., Nakayama, M. and Ishida, M., Serological and virological investigations of orthomyxovirus in birds in south-east Asian area, Dev. biol. Standard. 39, 457-460 (1977). 7. Nerome, K., Nakayama, M., Ishida, M., Fukumi, H., Butterfield, W. K., Webster, R. G. and Campbell, C. H., Isolation and serological characterization of influenza A viruses from birds that were dead on arrival at Tokyo Airport, Archs Virol. 57, 261-270 (1978).
Avian influenza in Great Britain
169
Spring of 1977. It could be that the Indian exporters merely altered their source of birds, but this seems unlikely. Influenza viruses isolated from captive birds imported into Great Britain up to 1974 were of Hav4 subtype [5, Chu cited in 6] whereas influenza viruses isolated from caged birds imported into Japan in 1976 were exclusively Hav7 from India but Hav7 and Hav4 from South East Asian countries [6, 7]. It has been suggested that from time to time epizootics may occur amongst birds in India and S.E. Asia causing changes in the predominant subtype isolated [6]. The results obtained in our studies suggest that since early 1977 viruses of Hav4 subtype have re-emerged as the dominant influenza subtype in the birds of the Indian sub-continent. The number of isolations of influenza viruses from turkeys between 1963 and 1978 was indicative of the state of influenza in domestic poultry in Great Britain during that time. There was no evidence of influenza viruses being endemic amongst domestic or wild birds and it seemed most likely that the occasional outbreaks of viruses of low virulence were the result of fresh introductions, presumably by migratory birds [3]. The outbreaks and isolations in 1979 were therefore quite unusual. The exact origins of these outbreaks are unknown but it would seem most likely that the viruses were introduced into the turkey flocks by wild birds. The very severe winter, both in Great Britain and on the continent of Europe had led to unusual circumstances in the bird populations, particularly in Norfolk. Most noticeable were the huge flocks of starlings seen in this area, some of which were estimated as containing up to 2,000,000 birds. The high winds and heavy snowfalls of the winter had in many cases caused damage to turkey houses allowing access to wild birds. The affected area is on the migratory routes of several species of waterfowl but starlings were obviously the most popular candidates for the responsibility of immediate transmission to turkeys. It is therefore of interest that in 1978 an influenza virus with H a v l Neql antigens was isolated from starlings in Israel (Lipkind, personal communication) this is the only known isolation of influenza virus from starlings. In Great Britain there is legislation for the slaughter of birds infected with highly virulent influenza viruses. The two sites on which virulent virus was present were therefore slaughtered out and no spread of the virus to other sites has been detected. Neither has any evidence been obtained of spread from any of the premises affected by virus of low virulence despite the close proximity of many farms with apparently susceptible birds. The severity of the 1978-1979 winter apparently had much to do with number of outbreaks of influenza virus infections of turkeys both in its affect on the behaviour of wild birds and damage to the turkey houses. It is therefore notable that the last previous isolation of virulent influenza virus in Great Britain: A/ty/Eng./63 (Hav l Nay3), followed the only winter of comparable severity to that of 1978-1979 and was made from turkeys in the same locality in Norfolk as those affected with virulent virus this year. Acknowledgments-The isolations and identification of influenza viruses from birds in Great Britain which are
summarized in this paper were the result of work done by officers of the State Veterinary Service, private veterinarians, staff of the RSPCA Hostel at Heathrow Airport and, in particular, members of the Poultry Department, Central Veterinary Laboratory, Weybridge. REFERENCES 1. Webster, R. G. and Laver, W. G., Antigenic variation of influenza viruses. In The influenza Viruses and Influenza, Kilbourne, E.D. (ed.), pp. 209-314. Academic Press, New York and London (1975). 2. Alexander, D. 3., Allan, W. H. and Sillars, T., Isolation of myxovirusesfrom dead birds arriving at Heathrow Airport, London, J. Hyg., Camb. 79, 243-247 (1977).