Abstract Book of EAVLD2012 - eavld congress 2012

S3 - P - 15
FIRST REPORTED LOW PATHOGENCITY AVIAN INFLEUNZA VIRUS SUBTYPE H9 INFECTION OF
DOMESTIC FOWL IN ENGLAND
C. Daniel Parker 1 , Scott M. Reid 2 , Allan Ball 1 , William J. Cox 2 , Steve C. Essen 2 , Amanda Hanna 2 ,
Marek J. Slomka 2 , Richard M. Irvine 2 , Ian H. Brown 2
1 Slate Hall Veterinary Practice Ltd, Unit 7 Highgate Farm, Over Road, Willingham, Cambridge, CB24 5EU
2 Animal Health and Veterinary Laboratories Agency-Weybridge, Woodham Lane, New Haw, Addlestone, Surrey, KT15 3NB
Avian influenza virus, H9 real-time RT-PCR, broiler breeders, UK poultry, Egg drop in poultry
Introduction
Poultry outbreaks, caused by avian influenza (AI) viruses of
H9N2 subtype, have been reported in many parts of Asia and the
Middle East since the 1990s where they may be considered as
being endemic (1, 2). Although H9 virus infections are typically
characterised as being of low pathogenicity (LP) and are not
classified as notifiable disease (3), LPAI H9N2 poultry outbreaks
in these regions continue to be characterised clinically by
respiratory disease and a reduction in egg production (4).
However, since the more recent emergence of a significant H9N2
poultry problem in Asia and the Middle East, the present paper
presents the first detailed epidemiological investigation of a H9
poultry outbreak in the UK and indeed in Europe. This occurred in
the East Anglia region in December 2010 in a broiler breeder
flock, and an important aspect of the outbreak investigation was
the successful use of molecular methods to identify and
characterise the AIV subtype.
Materials & methods
Disease suspicion was based on acute drops in egg production in
two of four sheds, poor egg shell quality and evidence of
diarrhoea. Cloacal and oropharyngeal swabs from 60 chickens
from each of two affected houses were pooled (n=48) into groups
of five swabs obtained from the same anatomical site. All swab
pools were inoculated into 10-day-old embryonated SPF fowls’
eggs, and allantoic fluids were harvested and tested for the
presence of any haemagglutinating agent (5). Four AI real-time
reverse transcription polymerase chain reaction (RRT-PCR)
assays were used to test the RNA extracts: (i) the Matrix (M)-
gene assay for generic AI detection; (ii) H5 and H7 AI virus RRT-
PCR assays to test for notifiable avian influenza (NAI); (iii) an H9
RRT-PCR; (iv) selected RNA extracts were tested by an N1 RRT-
PCR. The RNA extracts were simultaneously screened for
Newcastle disease virus (NDV) using primers and probes
targeting the L-gene of NDV. RNA extracted from the positive
cloacal swab specimen A/chicken/England/1415-51184/10 was
used for nucleotide sequencing. The HA and neuraminidase (NA)
genes were amplified by conventional reverse transcription (RT)-
PCR, sequenced and analysed phylogenetically.
poultry for 40 years vindicates the need for continued vigilance
and surveillance of AI viruses in poultry populations.
Acknowledgements
The authors thank Andrew Gibson, Mark Spelman and Simon
Rednall from the farm management team, Sue Graham of Slate
Hall Veterinary Practice Ltd for her laboratory assistance and Dr
Dennis Alexander of AHVLA for technical assistance. This study
was supported by Defra contract ED1300: Scanning Surveillance
for Disease in Poultry and Game Birds in England, Scotland and
Wales.
References
1.Alexander, DJ (2007). An overview of the epidemiology of avian
influenza. Vaccine 25:5637–5644.
2.Capua, I, Alexander, DJ (2007) Avian influenza infections in birds – a
moving target. Influenza and Other Respiratory Viruses 1:11–18.
3.OIE (World Organisation for Animal Health). (2008). World Health
Organisation for Animal Health, Terrestrial Animal Health Code, chapter
10.4 “Avian influenza”. OIE: Paris. Available at:
http://www.oie.int/eng/normes/mcode/en_chapitre_1.10.4.pdf
4.Swayne, DE, Halvorson, DA (2003). Influenza. In ”Diseases of Poultry”,
pp135-160. Eds: YM Saif, HJ Barnes, JR Glisom, AM Fadly, LR
McDougald and DE Swayne. Ames, Iowa: Iowa State University Press.
5.OIE (World Organisation for Animal Health). (2010). Manual of
Diagnostic Tests and Vaccines for Terrestrial Animals, chapter 2.3.4.
“Avian influenza”. Paris: OIE. Available at:
http://www.oie.int/fileadmin/Home/eng/Health_standards/tahm/2.03.04_AI.
p
Results
Attempted virus isolation in embryonated SPF fowls’ eggs was
unsuccessful. H9N1 LPAI virus infection was confirmed by RRT-
PCR. Sequencing of the haemagglutinin and neuraminidase
genes revealed high nucleotide identity of 93.6% and 97.9% with
contemporary North American H9 and Eurasian N1 genes,
respectively. Epidemiological investigations were conducted to
identify the source of infection and any onward spread.
Discussion & conclusions
This is the first known report of an H9 subtype avian influenza
infection in poultry in England. The infecting subtype was H9N1
and phylogenetic analysis of the HA and NA genes indicated
North American and Eurasian origins for these two genes, which
suggested that this AIV may be a reassortment that consists of
genetic segments of diverse geographic origin. The source of the
influenza virus infection could not clearly be established.
Following the infection, epidemiological investigations failed to
identify any human, inter- or intra-company contacts that could
have introduced the virus onto the farm. No other breeder or
poultry farms in the area were known to be infected. Infection
followed a period of extremely cold weather and snow which
could have compromised disease security. Wild bird activity
around the farm buildings was significantly higher during this cold
weather period as birds foraged for feed. In the present episode,
clinical signs were relatively mild in the poultry with no mortality.
However, this first reported detection of H9 LPAI virus in UK