Assessment of producers' response to Salmonella biosecurity issues and uptake of advice on laying hen farms in England and Wales.

High standards of biosecurity are known to reduce the risk of disease outbreaks; however, uptake of advice and implementation of biosecurity measures are dependent on many factors. This study assessed the uptake of targeted biosecurity advice by 60 laying hen farms provided during biosecurity audit visits. Advice was provided as bullet point cards focusing on specific areas identified as benefitting from improvement. These covered site entrance, site tidiness, vaccination, boot hygiene, hand hygiene, house tidiness, rodent control, fly control, red mite control and cleaning and disinfection between flocks. Background knowledge of Salmonella and biosecurity and farmers' willingness and intent to implement additional measures were assessed. About 50% of the principal decision-makers had basic background knowledge of Salmonella, with 22% considered well informed; almost all agreed that biosecurity could impact on Salmonella control and many appeared willing to implement additional biosecurity measures. Sixty-three per cent of study farms were categorised using the Defra Farmer Segmentation Model as Modern Family Businesses (MFBs), with 7-11% of farms being categorised as Custodian, Lifestyle Choice, Pragmatist or Challenged Enterprise; however, categorisation, did not determine uptake of advice. The most frequently used advice cards were boot hygiene, red mite control, hand hygiene, site entrance and cleaning and disinfection; uptake of advice ranged from 54 to 80% depending on the advice card. Uptake of advice by the farmers was encouraging, especially considering it was being provided by people other than their usual source of biosecurity information. Those who did not implement the recommended measures cited cost, difficulty of enforcement and practicality as the main reasons. However, the positive uptake of advice and implementation of recommended measures by many farmers demonstrates that targeted advice, discussed face to face with farmers, on a small number of key areas, is a potentially effective method of providing biosecurity information to complement more lengthy formal advisory reports.

A quantitative release assessment for the noncommercial movement of companion animals: risk of rabies reintroduction to the United kingdom.

In 2004, the European Union (EU) implemented a pet movement policy (referred to here as the EUPMP) under EU regulation 998/2003. The United Kingdom (UK) was granted a temporary derogation from the policy until December 2011 and instead has in place its own Pet Movement Policy (Pet Travel Scheme (PETS)). A quantitative risk assessment (QRA) was developed to estimate the risk of rabies introduction to the UK under both schemes to quantify any change in the risk of rabies introduction should the UK harmonize with the EU policy. Assuming 100 % compliance with the regulations, moving to the EUPMP was predicted to increase the annual risk of rabies introduction to the UK by approximately 60-fold, from 7.79 × 10(-5) (5.90 × 10(-5), 1.06 × 10(-4)) under the current scheme to 4.79 × 10(-3) (4.05 × 10(-3), 5.65 × 10(-3)) under the EUPMP. This corresponds to a decrease from 13,272 (9,408, 16,940) to 211 (177, 247) years between rabies introductions. The risks associated with both the schemes were predicted to increase when less than 100 % compliance was assumed, with the current scheme of PETS and quarantine being shown to be particularly sensitive to noncompliance. The results of this risk assessment, along with other evidence, formed a scientific evidence base to inform policy decision with respect to companion animal movement.

A comparison of the efficacy of different disinfection methods in eliminating Salmonella contamination from turkey houses.

AIMS: This study aimed to compare the efficacy of different disinfection methods in eliminating Salmonella contamination from turkey houses. METHODS AND RESULTS: Fifty depopulated turkey houses which had all housed Salmonella-positive flocks were visited after cleaning and disinfection. A minimum of 45 swab samples from different surfaces were taken per house and analysed for the presence of Salmonella. The sampled surfaces included intact floor surfaces, floor cracks, walls, feeders, drinkers, anteroom, nestboxes and miscellaneous items. Houses were grouped according to the disinfectant which had been used and the efficacy of the different groups of disinfectants was compared. Sixty-eight % of houses tested positive for Salmonella after C&D. Out of 4440 samples, 207 tested positive for Salmonella, giving an overall sample prevalence of 4.7%. There was no significant difference in the level of residual contamination between breeding, rearing and finishing houses. Products containing a mixture of formaldehyde, glutaraldehyde and quaternary ammonium compounds (QAC) performed significantly better than products containing hydrogen peroxide and peracetic acid. Cleaning and disinfection was least effective in nestboxes and anterooms. CONCLUSIONS: Thorough cleaning and the choice of a suitable disinfectant are crucial if Salmonella contamination of turkey houses is to be eliminated. SIGNIFICANCE AND IMPACT OF THE STUDY: This study shows that disinfectants containing a mixture of formaldehyde, glutaraldehyde and QAC perform significantly better under field conditions than oxidising products and should therefore be the first choice for disinfection of turkey premises where Salmonella is present.

Comparison of sampling methods to detect Salmonella infection of turkey flocks.

AIMS: To compare the efficiency of various sampling methods for detection of Salmonella in turkey flocks. METHODS AND RESULTS: In a field study that compared various sampling methods one pair of boot swabs taken from the whole turkey house provided suitably sensitive results for fattening and rearing flocks and was no less sensitive than two pairs, each from half the house, tested as a pooled sample. The sensitivity was further enhanced by adding a dust sample. The dust sample appeared to be particularly useful in flocks with a low prevalence, especially in breeding flocks, and was more sensitive than a method which used five pairs of boot swabs per flock. Combined incubation of a boot swab and a dust sample showed no interference between the two sample types and a maximum sensitivity of detection. Litter samples and commercial sponge drag swabs provided a lower level of detection. CONCLUSIONS: A single pair of boot swabs taken from the whole house is recommended for routine sampling of commercial rearing or fattening flocks. An additional dust sample could be added to increase detection in flocks with a low prevalence or in breeding flocks, but adding an additional pair of boot swabs would not increase detection compared with a single pair. SIGNIFICANCE AND IMPACT OF THE STUDY: This study demonstrates that significant efficiencies can be made in sampling programmes for detection of Salmonella in turkey flocks without detracting from the sensitivity. Similar studies are recommended for other poultry sectors, particularly in chicken breeding flocks.

Comparison of environmental sampling methods for detecting Salmonella in commercial laying flocks in the UK.

AIMS: To investigate the performance of the Salmonella National Control Programme (NCP) sampling/testing methods in laying flocks of domestic fowl. METHODS AND RESULTS: Eighty-five visits were made to 69 flocks representative of the main production systems (cage, barn and free-range) infected with Salmonella. In each visit, three methodologies were compared: (i) the European Union (EU) baseline survey method (five faeces and two dust samples); (ii) an in-house (Veterinary Laboratories Agency, VLA) 'wet' method that involved collecting 10 dust and 10 faeces samples into jars with buffered peptone water; and (iii) a method involving two samples of pooled faeces and one of dust (cultured as one sample of each type), which has been adopted for the NCP for laying flocks across the EU. CONCLUSIONS: The 'wet' method was the most sensitive, and the NCP the least, although individual NCP samples were the most sensitive ones. SIGNIFICANCE AND IMPACT OF THE STUDY: The apparent lower sensitivity of the NCP method may be compensated by repeated sampling of flocks (twice during rear and several times during lay). Sampling using VLA methodology should be advocated for farms aiming to disclose low-level Salmonella before restrictions on the sale of eggs from Salmonella Enteritidis or Salmonella Typhimurium-infected flocks are in place.

Risk factors associated with the salmonella status of dairy farms in England and Wales.

Between October 1999 and February 2001 the salmonella status of 449 dairy farms in England and Wales was determined by environmental sampling on up to four occasions. Data were collected through interview-based questionnaires, and multivariable analyses were used to identify risk factors associated with the farms either being Salmonella positive (prevalence data) or becoming Salmonella positive (incidence data). Region, herd size, month of visit and the lack of a clean visitor parking area were significantly associated with the prevalence of Salmonella species, and there was a significant trend towards an increased risk in late summer and autumn. The introduction of six- to 24-month-old cattle into a herd was associated with a reduced prevalence, but the introduction of adult cattle only, or calves with other cattle, was associated with an increased (but not significant) risk of farms being Salmonella positive. Month of visit, the lack of a clean visitor parking area, the use of part-time workers and not feeding calves whole milk, but not region or herd size, were associated with an increased incidence of salmonella.

Variable and strain dependent colonisation of chickens by Escherichia coli O157.

The prevalence of enterohaemorrhagic Escherichia coli (EHEC) O157 in poultry is considered minimal compared with other species, especially ruminants. However, deliberate inoculation studies have shown that poultry are readily and persistently infected by this organism but that the mechanism of colonisation is independent of intimin, a recognised factor in host-EHEC interactions in mammalian species, and may be dependent upon flagella. Few strains of EHEC O157 have been tested in poultry and here 1-day-old and 6-week-old chicks were inoculated with seven non-toxigenic E. coli O157 strains in separate experiments. Persistence was measured semi-quantitatively by bacteriological assessment of E. coli O157 cultured from cloacal swabs (shedding score). In the 1-day-old chick model that was monitored for 43 days, all seven strains established well after inoculation. In the 6-week-old chicken model, one strain established and gave consistently high shedding for the duration of the experiment (156 days). Whereas of the remaining six strains, two persisted for 113 days, two persisted for 43 days, one persisted for 22 days and one strain was never detected.

Prevalence, incidence and geographical distribution of serovars of Salmonella on dairy farms in England and Wales.

A study of randomly selected dairy farms in England and Wales was made between October 1999 and February 2001 to estimate the prevalence and incidence of Salmonella serovars. The farms were enrolled through five milk-buying companies, which represented 63 per cent of the dairy farms in England and Wales, and they were sampled on up to four occasions (449 farms at visit 1, 272 farms at visit 2, 251 farms at visit 3 and 243 farms at visit 4). In total, 19,296 samples of pooled faecal pats and slurry were collected. The farm-specific prevalence of all serovars of Salmonella ranged from 12.1 per cent (95 per cent confidence interval [CI] 8.2 to 16.0 per cent) to 24.7 per cent (95 per cent CI 19.4 to 30.1 per cent) at each visit. The most common serovars identified were Salmonella Dublin (3.7 to 6.6 per cent farm-specific prevalence at each visit), Salmonella Agama (1.8 to 7.6 per cent) and Salmonella Typhimurium (2.6 to 4.1 per cent) The prevalence varied by region and month of sampling and increased in late summer. The incidence rate of all serovars of Salmonella was 0.43 (95 per cent CI 0.34 to 0.54) cases per farm-year at risk. There was no significant difference between the incidence rates of the common serovars S Typhimurium (0.07), S Dublin (0.06) and S Agama (0.13). A total of 29 Salmonella serovars were isolated. Few of the isolates were resistant to the 16 antimicrobial agents tested, except the isolates of S Typhimurium dt104, of which 67.9 per cent were resistant to at least five of them.

Analysis of expression of flagella by Salmonella enterica serotype typhimurium by monoclonal antibodies recognising both phase specific and common epitopes.

Monoclonal antibodies specific for phase 1 ("i" antigen), phase 2 ("1,2" antigen) and common epitopes of the flagellins of Salmonella enterica serotype Typhimurium were raised. Having confirmed their specificity, the monoclonal antibodies were used to develop semi-quantitative ELISAs in order to assess the relative expression of the two phases by strains of Typhimurium. The majority of Typhimurium strains representative of a wide cross-section of definitive types from animal and environmental sources preferentially expressed phase 1 antigen in vitro. DT40 strains were unique in expressing phase 2 preferentially. The ratio of phase 1 to phase 2 expressed by strains tended to be constant for any one strain when strains were grown on a number of conventional laboratory media. However, the ratio of phases was shown to be modulated by incubation at 42 degrees C and buffering media at pH values, notably 4.5, other than neutral. Selenite broth and Rambach media repressed flagellation.

The O-antigen of Salmonella enterica serotype Enteritidis PT4: a significant factor in gastrointestinal colonisation of young but not newly hatched chicks.

The lipopolysaccharide of Salmonella and other Gram negative pathogenic species has been implicated as a major virulence determinant and in this study we report the role of LPS of S. Enteritidis in the colonisation and persistent gastrointestinal infection of young poultry. The gene encoding the unique O-antigen ligase, waaL, was mutated by insertional inactivation in a well characterised S. Enteritidis strain, S1400/94. The waaL mutant, designated PCP, produced rough colonies on agar medium, did not agglutinate O9 antiserum, did not produce an LPS ladder on silver stained gels and was serum sensitive. PCP and a nalidixic acid marked derivative of S1400/94 (S1400/94 Nalr) were used to orally challenge young chicks, separately and together in competitive index experiments. At post-mortem examination of 1-day-old chicks challenged S1400/94 Nalr and PCP separately there were no significant differences in the numbers of S1400/94 Nalr and PCP bacteria in tissues sampled on days 1, 2, and 5. By day 42 after challenge S1400/94 Nalr bacteria were recovered in significantly higher numbers than PCP from the caecal contents (P < 0.001). In competitive index studies in the 1-day-old chick PCP colonised, invaded and persisted in lower numbers than S1400/94 Nalr. In 4-week-old chicks challenged separately, PCP bacteria were recovered from all tissues examined in significantly lower numbers than S1400/94 Nalr. In competitive index experiments in 4-week-old chicks, PCP was not detected at any site and at any time point. Therefore, the O-antigen of S. Enteritidis plays an important role in poultry infections although this role is less important in the newly hatched chick.

A longitudinal study of campylobacter infection of broiler flocks in Great Britain.

One hundred flocks associated with five integrated poultry companies were monitored for one production cycle to investigate risk factors for campylobacter infection of poultry broiler flocks. Bacteriological samples were collected from one house of birds on each site at weekly intervals from 3 to 4 weeks of age until the birds were infected with campylobacter or the flock was depopulated (whichever was sooner). Environmental samples were obtained from 20 houses after cleansing and disinfection of the site before chick arrival. Conventional methods were used for the isolation of campylobacter. Questionnaires were used to collect information on potential risk factors for campylobacter infection. Discrete-time survival analysis was used to assess the influence of various exposures on the age at which the flock was infected with campylobacter. More than 40% of flocks were infected with campylobacter by the time the chicks were 4 weeks old and >90% by 7 weeks. Infection spread rapidly to most birds in a flock. Infection was not predictable by campylobacter status of the last flock reared on the site. (However, because most flocks were infected, the power to detect such an association was poor.) There was no evidence of environmental survival of campylobacters in broiler houses after adequate cleansing and disinfection. The most important predictors of protection from campylobacter were related to effective hygiene barriers (such as housing birds in buildings in a good state of repair, appropriate usage of disinfectant boot dips and a high standard of cleansing and disinfection of the drinking-water equipment). There was no evidence that rodents were a source of infection (but most sites operated effective vermin-control programmes).

A trial of biosecurity as a means to control Campylobacter infection of broiler chickens.

We ran a controlled intervention trial to assess whether the risk of a broiler flock becoming infected with Campylobacter could be reduced by biosecurity measures. These were a standard method of cleansing and disinfecting the poultry house prior to stocking, and a standard hygiene protocol followed by all personnel who entered the study house during the flock's life. Thirty-nine flocks were allocated to intervention or control groups in a ratio of 1:2. Intervention flocks were asked to follow the specified biosecurity measures; all flocks were monitored weekly for Campylobacter infection. Analysis of infection at 42 days of age and over the life of the flock showed that the risk of thermophilic Campylobacter infection of broilers was reduced by over 50% in intervention flocks. Parts of the intervention identified as significant in the univariable analysis included twice weekly replenishment of boot dip disinfectant; potential independent risk factors identified included the location of ventilation fans and daily sanitisation of the water supply. The non-random allocation of 10 flocks to the control group may have introduced some study bias (the effect of which is discussed in the paper).

Surface disinfection tests with Salmonella and a putative indicator bacterium, mimicking worst-case scenarios in poultry houses.

Surface disinfection studies mimicking worst-case scenarios in badly cleaned poultry houses were made with 3 bacterial isolates (Salmonella enteritidis, Salmonella senftenberg, and Enterococcus faecalis), and 3 1% disinfectant solutions, formaldehyde (F; 24.5% vol/vol), glutaraldehyde/benzalkonium chloride (G; Bio Komplet Plus), and a peroxygen compound (P; Virkon S), with World Health Organization (WHO) standard hard water as a control. Materials (concrete paving stones, steel feed chain links, wooden dowels, and jute egg belts) and organic matter found commonly in poultry houses (feed, fats, egg yolk) were used in the tests. Organic matter inoculated with high numbers of stationary phase cultures was added to materials and dried for 24 h at different temperatures (6, 11, 20, or 30 degrees C), immersed in solutions for set time periods (5, 15, or 30 min), and dried again for 25 h (6, 11, or 30 degrees C). Then, traditional recovery procedures (using 10-fold dilutions until 10(-4), i.e., a most probable number method) were applied. For the 2 Salmonella isolates, the efficacy of the solutions was (in decreasing order): formaldehyde > glutaraldehyde/benzalkonium chloride > peroxygen compound > WHO hard water, except when feed chain links with fats were disinfected using 30 degrees C before and after disinfection, for which the peroxygen compound seemed more effective. Enterococcus faecalis was equally or less susceptible than S. enteritidis and S. senftenberg, indicating its suitability as an indicator bacterium. For the peroxygen compound, S. senftenberg was more susceptible than S. enteritidis in spite of higher minimum inhibitory concentrations to this disinfectant for the former.

Clinical and epidemiological correlates of the neurohistology of cases of histologically unconfirmed, clinically suspect bovine spongiform encephalopathy.

The associations between three major categories of the neurohistological diagnoses and the epidemiological data were examined in unconfirmed cases of clinically suspect bovine spongiform encephalopathy (BSE). The diagnostic categories were focal spongiosis of white matter (37 cases), encephalic listeriosis (13 cases) and no significant lesions (78 cases). An additional control category of 200 confirmed cases of BSE were included for comparison. Epidemiological variables were the frequencies of specific clinical signs, the season of clinical onset, the age, the duration of the clinical signs and the geographical origin of the cases. Discriminant analysis was used to assess the contribution of these variables to the distinction between the diagnostic categories. The analyses characterised the cases of listeriosis by their shortest clinical duration, the greater prevalences of certain clinical signs and their occurrence mainly in winter and spring, consistent with current understanding of the disease. Cases of focal spongiosis, a lesion of unknown significance, but potentially with a metabolic causation, were tentatively separable from cases with no significant lesions by their winter onset. The results also confirmed that among the categories, the cases of BSE had the longest clinical duration. Despite their statistical significance, the findings do not have sufficient predictive power to be of value in making clinical decisions.

Evaluation of an ELISA for Mycobacterium bovis infection in badgers (Meles meles).

The performance of an indirect ELISA for diagnosing Mycobacterium bovis infection in live badgers was evaluated by examining blood samples collected from 1982 badgers captured during statutory badger removal operations in south west England. The Validity of the test and the factors affecting the prevalence of infection are described. The sensitivity of the ELISA was 40.7 percent, its specificity was 94.3 percent, the predictive value of a positive test was 67.5% percent and the predictive value of a negative test was 84.6 percent. Its sensitivity was significantly higher in males and animals with gross lesions typical of tuberculosis. The sensitivity and positive predictive values were enhanced when the results were grouped by control operation. Variables of significance for prevalence were the county, the time of year, the age and sex of animal, and the time after the start of a control operation. The possible use of the ELISA as a screening test is discussed.

Bovine spongiform encephalopathy: aspects of the clinical picture and analyses of possible changes 1986-1990.

A standard questionnaire was used to record the presence of specific clinical signs reported for histopathologically confirmed cases of bovine spongiform encephalopathy observed before June 30, 1990, and the frequencies of these signs were analysed. The signs most frequently recorded were apprehension, hyperaesthesia and ataxia, and there were variations in the frequency with which some signs were recorded in animals observed at different times during the epidemic. These variations were considered to be the result of differences between observers and differences in the duration of the illness, rather than a change in the clinical picture, which could possibly have occurred with a change in the nature of the agent to which the cattle had been exposed, or to a change in their response to the agent.

Studies of embryo transfer from cattle clinically affected by bovine spongiform encephalopathy (BSE).

Semen from 13 bulls, eight with clinical bovine spongiform encephalopathy (BSE), was used to artificially inseminate (AI) 167 cows with clinical BSE, and their resultant embryos were collected non-surgically seven days after AI. The viable and non-viable embryos with intact zonae pellucidae were washed 10 times (as recommended by the International Embryo Transfer Society) then frozen. Later, 587 of the viable embryos were transferred singly into 347 recipient heifers imported from New Zealand, and 266 live offspring were born of which 54.1 per cent had a BSE-positive sire and a BSE-positive dam. The recipients were monitored for clinical signs of BSE for seven years after the transfer, and the offspring were monitored for seven years after birth. Twenty-seven of the recipients and 20 offspring died while being monitored but none showed signs of BSE. Their brains, and the brains of the recipients and offspring killed after seven years, were examined for BSE by histopathology, PrP immunohistochemistry, and by electron microscopy for scrapie-associated fibrils. They were all negative. In addition, 1020 non-viable embryos were sonicated and injected intracerebrally into susceptible mice (20 embryos per mouse) which were monitored for up to 700 days, after which their brains were examined for spongiform lesions. They were all negative. It is concluded that embryos are unlikely to carry BSE infectivity even if they have been collected at the end-stage of the disease, when the risk of maternal transmission is believed to be highest.

Tuberculin manufacturing source and breakdown incidence rate of bovine tuberculosis in British cattle, 2005-2009.

The single intradermal comparative cervical tuberculin (SICCT) test is the primary test used for surveillance for bovine tuberculosis (bTB) in cattle in Great Britain (GB). The tuberculin used can, with other factors, influence test accuracy. In this analysis, the detection of infected cattle in GB 2005-2009 was compared between SICCT tests using tuberculins manufacturered by different manufacturers. Higher rates of reactors (adjusted rate 209 vs 186 per 100,000 tests, P = 0.003) and herd bTB incidents (adjusted total breakdown rate 5.1 vs 4.5 per 100 herd-years at risk, P < 0.001) were detected using tuberculin manufactured at Weybridge compared with Lelystad. However, confirmation of infection in reactors by postmortem evidence was higher with Lelystad tuberculin (adjusted percent 44.1 vs 47.1, P = 0.018). The findings, overall, suggest slightly higher test sensitivity and lower test specificity associated with Weybridge tuberculin compared with Lelystad. Assuming effective adjustment for confounding, the overall impact of tuberculin manufacturing source (2007-2009), was calculated to range somewhere between 315 false positive breakdowns, and 1086 bTB breakdowns missed (624 confirmed) as a result of using Weybridge and Lelystad tuberculin, respectively. However, animals that tested negative to the SICCT were not slaughtered at the time of the tests, so definitive conclusions are not possible.

Geographical association between the genotype of bovine tuberculosis in found dead badgers and in cattle herds.

In a survey, 457 badgers that had been found dead in Wales were postmortem-examined, and samples were examined by histology and by extended culture (for up to 12 weeks). Mycobacterium bovis was cultured from 55 badgers (12.0 per cent), and the histology typical of M bovis infection was seen in a further six (1.3 per cent). The prevalence in badgers in each of 10 geographical areas varied between 0 and 26 per cent (P<0.001), and was associated with the incidence of confirmed M bovis infection in cattle herds in the same areas (P<0.01). In northern Wales, bTB was rare in both hosts. An infected badger was 12.3 times more likely to be within 5 km of a confirmed cattle bTB breakdown than an uninfected badger. The M bovis isolates from badgers belonged to one of four genotypes defined by spoligotype and variable number tandem repeat type. These genotypes were also found in 290 concurrent confirmed herd breakdowns, and tended to be similar to the genotypes in badgers in the same geographical areas. When badgers and cattle no more than 30 km apart were compared, the genotype diversity was greater in cattle than in badgers (P=0.016), suggesting that the movement of cattle plays a greater part in the spatial distribution of M bovis than the movement of badgers.