Prevalence of thermophilic Campylobacter species in ...

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Prevalence of thermophilic Campylobacter species in cats and dogs in two animal shelters in Ireland E. Acke, P. Whyte, B. R. Jones, K. McGill, J. D. Collins, S. Fanning Rectal swabs or faecal samples for the isolation of Campylobacter species were taken from 120 dogs and cats in an animal shelter in which only one kitten showed signs of gastrointestinal disease, and rectal swabs were taken from 46 dogs, 22 of which showed signs of gastrointestinal disease, in another shelter. At the first shelter, the swabs from 24 of 47 dogs (51.1 per cent) and 36 of 48 cats (75 per cent) yielded a Campylobacter species. The rate of isolation was significantly higher from dogs and cats less than six months old, and significantly higher from cats than from dogs (P≤0·05). At the second shelter Campylobacter species were isolated from 40 of 46 dogs (87 per cent), but there was no significant difference between the age groups. Campylobacter species were isolated from 19 (86·4 per cent) of the 22 dogs with signs of gastrointestinal disease and from 21 (87·5 per cent) of the 24 unaffected dogs. Several culture methods were applied to the samples collected from both shelters, and the combination significantly increased the recovery of Campylobacter species.

Veterinary Record (2006) 158, 51-54 E. Acke, CertSAM, MRCVS, P. Whyte, MSc, PhD, B. R. Jones, BVSc, FACVSc, DECVIM-CA, MRCVS, K. McGill, BSc, J. D. Collins, MVB, MVM, MS, PhD, MRCVS, S. Fanning, BSc, PhD, Faculty of Veterinary Medicine, University College Dublin, Belfield, Dublin 4, Ireland

CAMPYLOBACTERIOSIS is the most common bacterial cause of human gastroenteritis in Ireland. Campylobacter jejuni is the most frequently isolated species, followed by Campylobacter coli; asymptomatic C jejuni infections are rare in human beings in developed countries. The infective dose is low and the risk factors for infection include the ingestion or handling of undercooked meat products, especially poultry, the consumption of contaminated or unpasteurised milk and dairy products, drinking water from contaminated supplies, foreign travel, and contact with pets (Altekruse and others 1994, Altekruse and Tollefson 2003, Foley and others 2003). The clinical signs range from mild, self-limiting enteritis to dysentery. Although rare, fatal bacteraemia may occur, especially in young children and immunosuppressed and elderly people. Campylobacter species have also been associated with hepatobiliary and urogenital infections, Guillain-Barré syndrome and reactive arthritis (Skirrow and Blaser 2000). The incidence of the disease can be reduced by increased control of the processing of meat products, by improving food handling practices and by improved hygiene when handling pets (Altekruse and Tollefson 2003). Campylobacter species have been isolated from many domestic and wild animals, and the main species of veterinary importance in companion animals include C jejuni and Campylobacter upsaliensis. Asymptomatic carriers are common, but the organism has also been associated with gastrointestinal disease, especially in younger animals (Ferreira and others 1979, Blaser and others 1980, McOrist and Browning 1982, Fleming 1983, Fox and others 1983, Nair and others 1985, Burnens and others 1992). The link between gastrointestinal signs in pets and the presence of Campylobacter organisms has been investigated, but its significance is uncertain and studies are needed to determine whether Campylobacter species can cause significant disease as a primary pathogen (Bruce and Fleming 1983, Fleming 1983, Gondrosen and others 1985, Olson and Sandstedt 1987, Burnens and others 1992, Baker and others 1999, Lopez and others 2002, Sandberg and others 2002). The rates of isolation of Campylobacter species from dogs and cats vary, depending on their age and species, the isolation methods used and the specific Campylobacter species, the animals’ housing and where they live, infections with other enteropathogenic organisms, the presence of enteric disease and the season of the year. The highest prevalences have been recorded in young dogs and cats living in groups (Malik and The Veterinary Record, January 14, 2006

Love 1989, Burnens and others 1992, Torre and Tello 1993, Baker and others 1999, Lopez and others 2002, Sandberg and others 2002). The aims of this study were to establish the prevalence of Campylobacter species in cats and dogs at two animal shelters, to investigate the possible risk factors for the infection, and to establish any correlation with gastrointestinal signs. MATERIALS AND METHODS Shelter 1 In autumn 2002, rectal swabs were taken from 47 dogs and 48 cats, and faecal samples were collected from 15 dogs and 10 cats, at a shelter in the Republic of Ireland. The dogs were of different ages, breeds and sex, and the cats were mainly domestic shorthair, of different ages and sex; they were housed in a cattery in a separate building from the dogs. The animals had been in the shelter for between one day and several months. Only one sample was collected from each of the 120 animals. None of the dogs showed clinical signs, but one kitten had diarrhoea. All the animals over eight weeks old had been vaccinated with a multivalent vaccine and treated with a broad-spectrum anthelminthic when they entered the shelter. Each dog, except the young puppies, was kennelled alone and the dogs had access to a run with concrete flooring and an area of grass. There were separate quarantine facilities for sick animals. The average population consisted of 25 dogs and 40 cats. There were also a few farm animals on the premises. Shelter 2 In spring 2003, rectal swabs were collected from 46 dogs in a shelter in Northern Ireland, which held approximately 50 dogs of various ages, breeds and sex. They had been in the shelter for periods ranging from one day to several years and they were sampled once. Aggressive dogs were not sampled. All the dogs had been fully vaccinated with a multivalent vaccine and had been treated with a broad-spectrum anthelminthic. They were kennelled alone or with one or two other dogs and had access to a run with concrete flooring and an area of grass. No quarantine measures were taken for apparently healthy dogs that entered the shelter. Twenty-two (47·8 per cent) of the dogs had diarrhoea; those with severe diarrhoea were iso-

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lated, but those with mild diarrhoea were not. Faecal samples from 12 of the animals showing clinical signs were collected and tested for gastrointestinal parasites, including Giardia and Cryptosporidium species, by the sugar and zinc sulphate flotation method described by Colville (1991), and for Salmonella species (Whyte and others 2003). Five animals with severe signs were tested for parvovirus infection by ELISA (IDEXX). Three staff members at the shelter developed acute diarrhoea, but their signs resolved and they had not sought medical advice. Culture methods The rectal swabs were transported in an Amies transport medium (Copan) and cultured within 48 hours at the Veterinary Public Health and Food Safety Laboratory, University College Dublin. Five different culture methods for Campylobacter species were used, including direct plating and enrichment techniques. In shelter 1, direct plating and filtration on to cefoperazone amphotericin teicoplanin (CAT) agar and modified charcoal cefoperazone deoxycholate agar (mCCDA) was applied to all 120 samples. In addition, 55 samples were filtered on to blood agar, and 96 samples were cultured on to mCCDA after enrichment in Preston broth. All five methods were used on the 46 samples from the dogs in shelter 2. These selective solid media were prepared by using blood-free mCCDA (Oxoid), to which either CAT (SR174) or charcoal cefoperazone deoxycholate agar (CCDA) (SR155E) selective supplements were added in accordance with the manufacturer’s instructions (Oxoid). Liquid selective enrichment media were prepared by using either the Preston broth or CAT broth formulations. Twentyfive grams of the basal medium (Nutrient broth number 2; Oxoid) was dissolved in 950 ml of deionised water and autoclaved; the solution was cooled to 55°C and Preston or CAT Campylobacter-selective supplements (SR204E or SR174, respectively) and a growth supplement (SR804E) were added, together with 50 ml of saponin-lysed defibrinated horse blood. Direct plating The rectal swabs were streaked on to mCCDA

and CAT agar plates and incubated at 37°C in a microaerophilic environment (Genbox microaer sachets; bioMérieux) for 48 hours for the mCCDA and 96 hours for the CAT plates. The plates were examined after 48 hours, and if negative the CAT plates were incubated for a further 48 hours. Any suspect colonies were subcultured on to blood agar plates and incubated for 48 hours. Selective enrichment The rectal swabs were placed in tubes

containing 5 ml of 0·1 per cent peptone water and vortexed for one minute; 1 ml samples of the vortexed diluent were then placed in 10 ml of Preston broth and 10 ml of CAT broth and incubated for 24 hours at 37°C. After the incubation, 200 µl of the CAT broth was filtered through a 47 mm diameter, 0·65 µm pore size cellulose acetate filter (Sartorius) in situ on CAT agar and blood agar plates. After one hour, the filter was removed, the filtrate was spread over the plate with a sterile glass hockey stick, and the blood agar plates were incubated at 37°C for 48 hours and the CAT agar plates were incubated for 96 hours. Suspect colonies on the CAT plates were transferred to blood agar plates and incubated for 48 hours, and then examined. The enriched samples in Preston broth were subcultured on to mCCDA agar and incubated at 37°C for 48 hours, and suspect colonies were transferred on to blood agar. Faecal samples and rectal swabs were cultured directly on to mCCDA and CAT agar plates and subcultured. For enrichment, 1 g of faecal material was diluted in 9 ml of 0·1 per cent peptone water, vortexed for one minute and processed in the same way as the rectal swabs.

A subpopulation of suspect isolates was screened and confirmed as Campylobacter species on the basis of the results of a multiplex PCR assay (Wang and others 2002). Statistical analysis The prevalences of Campylobacter species recovered from the samples in each group were compared by using chi-squared analysis, with significance defined at the 95·0 per cent level (P