R. H. Davies

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 flocks 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).

National survey for Salmonella in pigs, cattle and sheep at slaughter in Great Britain (1999-2000).

Aims:  The objective of these surveys was to estimate the prevalence of faecal carriage of Salmonella in healthy pigs, cattle and sheep at slaughter, and of pig carcase contamination with Salmonella. These data can be used as a baseline against which future change in Salmonella prevalence in these species at slaughter can be monitored.

Intestinal carriage of verocytotoxigenic Escherichia coli O157, Salmonella, thermophilic Campylobacter and Yersinia enterocolitica, in cattle, sheep and pigs at slaughter in Great Britain during 2003.

An abattoir survey was undertaken to determine the prevalence of foodborne zoonotic organisms colonizing cattle, sheep and pigs at slaughter in Great Britain. The study ran for 12 months from January 2003, involved 93 abattoirs and collected 7703 intestinal samples. The design was similar to two previous abattoir surveys undertaken in 1999-2000 allowing comparisons. Samples were examined for VTEC O157, Salmonella, thermophilic Campylobacter and Yersinia enterocolitica. The prevalence of VTEC O157 faecal carriage was 4.7% in cattle, 0.7% in sheep and 0.3% in pigs. A significant decrease in sheep was detected from the previous survey (1.7%). Salmonella carriage was 1.4% in cattle, a significant increase from the previous survey of 0.2%. In sheep, faecal carriage was 1.1% a significant increase from the previous survey (0.1%). In pigs, carriage was 23.4%, consistent with the previous study. Thermophilic Campylobacter spp. were isolated from 54.6% of cattle, 43.8% of sheep and 69.3% of pigs. Y. enterocolitica was isolated from 4.5% of cattle, 8.0% of sheep and 10.2% of pigs.

Sources of salmonella on broiler carcasses during transportation and processing: modes of contamination and methods of control

Aims: The prevalence and types of salmonella in broiler chickens during transportation and during slaughter and dressing were studied. This was part of a comprehensive investigation of salmonellas in two UK poultry companies, which aimed to find the origins and mechanisms of salmonella contamination.

Observations on Salmonella contamination of commercial laying farms before and after cleaning and disinfection

Little is known about the effectiveness of the cleaning and disinfection methods in use on commercial laying farms in Great Britain. Samples were taken from poultry house structures and equipment of five cage layer flocks, five barn egg production flocks and two free-range flocks. In the free-range houses there was a decrease in Salmonella after cleaning and disinfection, although the soil in the paddocks remained contaminated. In the barn and especially the cage layer houses, significant residual contamination remained on the surfaces of buildings and equipment. Wildlife pests were also found to be carrying Salmonella in the disinfected houses and free-range paddocks.

Persistence and clearance of different Salmonella serovars in buildings housing laying hens

We investigated factors associated with persistence of different Salmonella serovars in buildings housing laying hens in Great Britain using survival analysis. A total of 264 incidents of Salmonella detection occurring between July 1998 and August 2007 in 152 houses were recorded. For incidents involving Salmonella Enteritidis (SE), both the rodent score of the house and the type of house were positively associated with persistence. For non-SE serovars, only the type of house was associated with persistence. Persistence of SE in the houses was longest (>15 months) in step-cage and cage-scraper houses when high levels of rodents were present, and lowest in non-cage and cage-belt houses. We estimated that 42% (95% CI 23.3-63.1) of SE incidents may be cleared during the lay period, and this was related to elimination of rodents from the houses. From January 2009, EU legislation will ban the sale of fresh eggs from SE-positive and S. Typhimurium-positive flocks over their remaining lifespan. If infection is eliminated from such flocks, they would cease to represent a public health risk.

Bacteriological and serological investigation of persistent Salmonella enteritidis infection in an integrated poultry organisation

Bacteriological monitoring of broiler breeder farms, the hatchery, rendering plant and animal feed mill during 1991 identified a number of potential cross-contamination hazards, such as the use of processed poultry proteins in the company feed mill and contamination of egg trolleys and trays, which may have led to widespread dissemination of Salmonella enteritidis within an integrated poultry organisation. Serological monitoring of the flocks suggested that, in most cases, substantial exposure to S. enteritidis infection occurred during the mid-rearing stage whereas routine bacteriological monitoring of poultry house litter and dust samples, and meconium samples taken in the hatchery identified infection only after the onset of the laying period. At least 10 phage types and six plasmid profile types of S. enteritidis were identified in historic submissions from the organisation including one apparently specific plasmid profile type that was distributed throughout the various parts of the company. During sampling for this investigation, most of these strains were not identified, and the number of plasmid profile types was reduced to a single common UK type.

Survey of the prevalence of Salmonella species on commercial laying farms in the United Kingdom.

A survey of salmonella infection on 454 commercial layer flock holdings in the uk was carried out between October 2004 and September 2005. Fifty-four (11·7 per cent, 95 per cent confidence interval 9·3 to 14·0 per cent) were salmonella positive. The most common serovar identified was Salmonella Enteritidis at a prevalence of 5·8 per cent, and 70 per cent of these isolates were phage types 4, 6, 7 and 35. Salmonella Typhimurium was the second most prevalent serovar, found in 1·8 per cent of the farms. Of the three other serovars given top priority by the eu because of their public health significance, Salmonella Virchow and Salmonella Infantis were each isolated from one holding, but Salmonella Hadar was not isolated from any of the holdings. Analysis of antimicrobial resistance patterns revealed that over 76 per cent of the isolates were sensitive to all of the 16 drugs tested, and all the isolates were sensitive to ciprofloxacin, gentamicin, ceftazidime, apramycin, amikacin, amoxicillin/clavulanic acid, neomycin and cefotaxime.

Molecular fingerprinting evidence of the contribution of wildlife vectors in the maintenance of Salmonella Enteritidis infection in layer farms

Aims: To provide molecular fingerprinting evidence of the contribution of wildlife vectors in the on‐farm epidemiology of Salmonella Enteritidis infections.

Decreased susceptibility to ciprofloxacin in outbreak-associated multiresistant Salmonella typhimurium DT104.

Dr Boltons present address is Food Safety Microbiology Laboratory, Central Public Health Laboratory, 61 Colindale Avenue, London NW9 5HT DURING the past decade, multiresistant Salmonella typhimurium definitive phage type (DT) 104 (MR DT104) which is resistant to ampicillin, chloramphenicol, streptomycin/ spectinomycin, sulphonamides and tetracyclines (ACSSpSuT) has caused numerous infections in food animals and human beings in the UK. Infections have also been reported in many European countries as well as in the USA and Canada (Threlfall 2000). In the UK, human infections with MR DT104 are primarily zoonotic in origin (Threlfall and others 1994). Although there has been a decline in isolations from food animals since 1994, MR DT104 remains the most common salmonella strain in cattle, sheep and pigs in the UK, and is second only to Salmonella enteritidis in poultry (Anon 1999a). In human salmonellosis in England and Wales, MR DT104 has declined from a peak of 4006 isolations in 1996 (Threlfall and others 1999) to 1030 isolations in 1999 (PHLS, unpublished observations). Nevertheless, the organism remains second only to S enteritidis phage type 4 in human beings in England and Wales. Antimicrobial drugs are used under veterinary prescription for therapy and prophylaxis in food-producing animals. In 1997 over 70 per cent of isolates of S typhimurium from livestock were resistant to ampicillin, chloramphenicol, streptomycin, sulphonamides and tetracyclines, and a substantial proportion of the resistant isolates were MR DT104 (Anon 1999b). Recently, particular concern has surrounded the isolation of an increasing proportion of MR DT104 with additional resistance to the first generation quinolone nalidixic acid (Nx) in isolations from food-producing animals (Davies and others 1999). Similarly, there have been reports of decreased susceptibility to the fluoroquinolone antimicrobial ciprofloxacin in isolations of MR DT104 from human beings (Threlfall and others 1998). Ciprofloxacin is the first-line drug for the treatment of invasive salmonellosis in humans. In 1994, 1 per cent of MR DT104 from human beings in England and Wales showed decreased susceptibility to ciprofloxacin, and by 1998 this figure had risen to 16 per cent (Threlfall and others 1999). Although the ciprofloxacin minimum inhibitory concentrations (MICs) are below the breakpoint concentration of 1 mg/litre as recommended by the British Society for Antimicrobial Chemotherapy (Anon 1996), S typhimurium isolates with similar MICs have been associated with treatment failure (Piddock and others 1990). In particular, in a recent outbreak of MR DT104 in Denmark, four of 11 patients did not respond to treatment with ciprofloxacin and there were two deaths (M0lbak and others 1999). The increase in the proportion of isolates of MR DT104 with decreased susceptibility to ciprofloxacin in England and Wales is, therefore, particularly disturbing. Between August 23 and September 6, 1998, a large community outbreak of MR DT104 (86 cases) occurred in northwest Lancashire (Anon 1998), and at least four patients were admitted to hospital. Epidemiological investigations revealed that 79 per cent of cases had consumed milk from a local dairy which received raw milk supplied by two farms. MR DT104 was isolated from the milk filter and failure of on-farm pasteurisation was thought to be the cause of the outbreak. On two occasions, in March and August 1998, S typhimurium, subsequently identified as MR DT104, was isolated from dairy cattle on one of the farms supplying the dairy. Strains from human beings, the dairy cattle and the milk filter were characterised. All were of R-type ACSSpSuTNXCPL (Nx nalidixic acid, CPL ciprofloxacin [L:MIC 0.5 mg/litre]) and possessed a single 90 kilobase (kb) plasmid. When a selection of isolates were typed by pulsed-field gel electrophoresis (PFGE), all gave a single PFGE pattern (XTmi) identical to that predominant in MR DT104 (Ridley and Threlfall 1998, Prager and others 1999). Long PCR (R. A. Walker, unpublished observations) demonstrated that these isolates also possessed the chromosomally-encoded 13 kb gene cluster that confers resistance to ACSSpSuT and is characteristic of MR DT104 (Briggs and Fratamico 1999). To investigate further the clonal nature of the strains, the mechanism of resistance to nalidixic acid and ciprofloxacin was determined in isolates from human beings, dairy cattle and the milk filter. The primary mechanism of resistance to nalidixic acid and ciprofloxacin commonly involves a single nucleotide mutation of the gyrA gene (Piddock and others 1998). Mutations in the gyrA gene were therefore characterised, using a rapid LightCycler assay (Gibson and others 1999). This involved amplifying a region of gyrA by PCR followed by the detection of ciprofloxacin resistance-associated mutations using hybridisation probes and thermal analysis. All the isolates were found to have a GAC-*GGC (aspartate to glycine) substitution at codon 87, subsequently confirmed by DNA sequencing. Ten different base pair substitutions in gyrA have been identified in ciprofloxacin-resistant salmonellas. However, although the GAC-*GGC substitution has been observed in MR DT104 (Ridley and Threlfall 1998), the most common mutation in strains with decreased susceptibility to ciprofloxacin involves a GAC->AAC (aspartate to asparagine) mutation at codon 87 (Ridley and Threlfall 1998, Molbak and others 1999, R. A. Walker, unpublished observations). In this outbreak, identification of the GAC->GGC gyrA mutation at codon 87 provided further confirmation of clonal identity of the strains involved. The emergence of decreased susceptibility to ciprofloxacin in MR DT104 is subsequent to the licensing in the UK, in November 1993, of the related fluoroquinolone enrofloxacin for use in food-producing animals. However, there is considerable controversy over the apparent association between enrofloxacin usage in food animals and the emergence of strains with decreased susceptibility to ciprofloxacin in human beings. Indeed, it has been stated that without evidence of scientific data, such an association cannot be substantiated (Jones 1998). However, this investigation has demonstrated a distinct mutation in the gyrA gene in isolates of MR DT104 from dairy cattle, a milk filter and human beings in a defined outbreak situation. An identical gyrA mutation was also identified in MR DT104 isolated from dairy cattle on one of the farms four months before the outbreak. The fluoroquinolone antimicrobial marbofloxacin was in use on the farm in the months preceding the outbreak (T. Leonard, personal communication). Although it is common for subclinical infection of cattle herds with MR DT104 to persist for several months or years (Davies 1997), it is possible