Temporal and farm-management-associated variation in faecal pat prevalence of Arcobacter spp. in ruminants.

In a 2-year longitudinal study of adult animals on 15 dairy farms and four sheep farms in Lancashire, UK, Arcobacter spp. were isolated from all farms although not at every sampling occasion. Faecal samples were collected and cultured using standard techniques for isolation of campylobacters. Assignment to species was via PCR assays. Apparent prevalence of Arcobacter spp. was higher in dairy cattle compared to sheep (40.1% vs. 8%, P < 0.001) and in housed cattle compared to cattle at pasture (50.1% vs. 20.9%, P < 0.001). This was reflected in the higher prevalence observed in herds that were housed (n = 4) all year compared to herds that grazed cattle on pasture in the summer and housed cattle in the winter (n = 11) (55.5% vs. 36%, P < 0.001). In the case of sheep, peak prevalence was observed in autumn with increased prevalence also being associated with improving pasture quality. There was an apparent inverse association between the faecal pat prevalence of Arcobacter spp. and Campylobacter jejuni although this may in part be an artefact of laboratory test method sensitivity, whereby a relative increase in the frequency of one bacterial species would reduce the sensitivity of detecting the other.

Temporal and farm-management-associated variation in faecal-pat prevalence of Campylobacter fetus in sheep and cattle.

SUMMARY: The faecal-pat prevalence (as estimated by culture) of Campylobacter fetus from cattle and sheep on 19 farms in rural Lancashire was investigated using standard Campylobacter culture techniques and PCR during a 2-year longitudinal study. C. fetus was isolated from 9·48% [95% confidence interval (CI) 8·48-10·48] of cattle faecal pats and 7·29% (95% CI 6·21-9·62) of sheep faecal pats. There was evidence of significant differences in shedding prevalence between geographical regions; cows in geographical zone 3 had an increased risk of shedding C. fetus compared to cows in geographical zones 1 and 2 (OR 6·64, 95% CI 1·67-26·5, P = 0·007), as did cows at pasture (OR 1·66, 95% CI 1·01-2·73, P = 0·046) compared to when housed. Multiple logistic regression modelling demonstrated underlying seasonal periodicity in both species.

Comparison of Arcobacter isolation methods, and diversity of Arcobacter spp. in Cheshire, United Kingdom.

The aims of this study were, firstly, to compare five published methods for the isolation of Arcobacter spp. from animal feces in order to determine the most sensitive and specific method. Second, we analyzed the resulting isolates by multilocus sequence typing (MLST) in order to investigate the diversity of the isolates recovered. Third, we investigated the ability to recover Arcobacter spp. from frozen fecal samples. Seventy-seven fecal samples from cattle, sheep, and badgers were subjected to five isolation methods, based on published methods for the isolation of Arcobacter and Campylobacter spp. Thirty-nine Arcobacter butzleri isolates were analyzed using a multilocus sequence typing scheme. The survival of Arcobacter spp. in frozen samples was investigated by freezing the fecal samples at -80°C for 7 days and then applying the same five isolation methods. The most sensitive and specific method used an Arcobacter-specific broth in conjunction with modified charcoal cefoperazone deoxycholate agar (mCCDA) with added antibiotics. Freezing of fecal samples led to a reduction in the recovery of Arcobacter spp. by approximately 50%. The 39 allelic profiles obtained by MLST could be divided into 11 sequence types (STs). We have identified the most sensitive and specific method for the isolation of Arcobacter spp. from animal feces and demonstrated that the freezing of fecal samples prior to isolation reduces arcobacter recovery. MLST analysis of the isolates revealed a high level of diversity.

Molecular epidemiology and genetic diversity of Campylobacter jejuni in ruminants.

Multi-locus sequence typing was performed on 1003 Campylobacter jejuni isolates collected in a 2-year longitudinal study of 15 dairy farms and four sheep farms in Lancashire, UK. There was considerable farm-level variation in occurrence and prevalence of clonal complexes (CC). Clonal complexes ST61, ST21, ST403 and ST45 were most prevalent in cattle while in sheep CC ST42, ST21, ST48 and ST52 were most prevalent. CC ST45, a complex previously shown to be more common in summer months in human cases, was more prevalent in summer in our ruminant samples. Gene flow analysis demonstrated a high level of genetic heterogeneity at the within-farm level. Sequence-type diversity was greater in cattle compared to sheep, in cattle at pasture vs. housed, and in isolates from farms on the Pennines compared to the Southern Fylde. Sequence-type diversity was greatest in isolates belonging to CC ST21, ST45 and ST206.

Spatio-temporal epidemiology of Campylobacter jejuni enteritis, in an area of Northwest England, 2000-2002.

A total of 969 isolates of Campylobacter jejuni originating in the Preston, Lancashire postcode district over a 3-year period were characterized using multi-locus sequence typing. Recently developed statistical methods and a genetic model were used to investigate temporal, spatial, spatio-temporal and genetic variation in human C. jejuni infections. The analysis of the data showed statistically significant seasonal variation, spatial clustering, small-scale spatio-temporal clustering and spatio-temporal interaction in the overall pattern of incidence, and spatial segregation in cases classified according to their most likely species-of-origin.

Temporal and farm-management-associated variation in the faecal-pat prevalence of Campylobacter jejuni in ruminants.

In a 2-year longitudinal study of adult animals on 15 dairy farms and four sheep farms in Lancashire, UK. C. jejuni was isolated from all farms, although not on every occasion. Faecal samples were collected and cultured using standard techniques for isolation of Campylobacter. Assignment to species was via PCR assays. Peak prevalence of C. jejuni in both cattle and sheep was observed during the summer and in cattle this apparent seasonality was associated with grazing pasture [odds ratio (OR) 2.14], while in sheep it was independent of grazing. Increased prevalence was associated with increased milk yield (OR 1.05) and herd size (OR 1.01) in dairy cattle, and with increased stocking density (OR 1.29) and pasture quality (OR 2.16) in sheep. There was considerable variation in prevalence between farms but no evidence of large-scale spatial variation. The association between C. jejuni prevalence and diet in dairy cattle deserves further investigation.

Typing of Campylobacter jejuni isolates from dogs by use of multilocus sequence typing and pulsed-field gel electrophoresis.

Campylobacter is a major cause of human gastroenteritis worldwide. Risk of Campylobacter infection in humans has been associated with many sources, including dogs. This study aimed to investigate whether C. jejuni carried by dogs could potentially be a zoonotic risk for humans and if there were common sources of C. jejuni infection for both humans and dogs. Multilocus sequence typing (MLST) together with macrorestriction analysis of genomic DNA using SmaI and pulsed-field gel electrophoresis (PFGE) were both used to analyze 33 C. jejuni isolates obtained from various dog populations, including those visiting veterinary practices and from different types of kennels. MLST data suggested that there was a large amount of genetic diversity between dog isolates and that the majority of sequence types found in isolates from these dogs were the same as those found in isolates from humans. The main exception was ST-2772, which was isolated from four samples and could not be assigned to a clonal complex. The most commonly identified clonal complex was ST-45 (11 isolates), followed by ST-21 (4 isolates), ST-508 (4 isolates), and ST-403 (3 isolates). The profiles obtained by macrorestriction PFGE were largely in concordance with the MLST results, with a similar amount of genetic diversity found. The diversity of sequence types found within dogs suggests they are exposed to various sources of C. jejuni infection. The similarity of these sequence types to C. jejuni isolates from humans suggests there may be common sources of infection for both dogs and humans. Although only a small number of household dogs may carry C. jejuni, infected dogs should still be considered a potential zoonotic risk to humans, particularly if the dogs originate from kennelled or hunt kennel dog populations, where the prevalence may be higher.

Campylobacter lari: genotype and antibiotic resistance of isolates from cattle, wildlife and water in an area of mixed dairy farmland in the United Kingdom.

Campylobacter lari is a rare human pathogen most commonly associated with birds and shellfish. Little information has been published regarding its prevalence in other environments, or on its potential role as a reservoir of antibiotic resistance. In this study, we characterized 109 C. lari isolated from a range of hosts using pulsed-field gel electrophoresis of macro-restricted chromosomal DNA, and by determining their susceptibility to a panel of four antibiotics. Pulsed-field gel electrophoresis analysis showed C. lari to be genetically diverse, particularly in isolates from wild birds and environmental water. The most common composite macro-restriction profile (cMRP) was found in multiple hosts (cattle, badgers, wild birds and rabbits), and seven other cMRPs were recovered from more than one host. All isolates were resistant to nalidixic acid and ciprofloxacin. Resistance to erythromycin and ampicillin was uncommon, but was observed in isolates from wild birds, cattle, wild mammals and water samples. The presence of the same cMRP in multiple hosts provides further evidence of transmission between livestock, wildlife and the environment, or for a common source of infection.

Prevalence and genetic diversity of Campylobacter spp. in environmental water samples from a 100-square-kilometer predominantly dairy farming area.

Water samples were taken systematically from a 100-km2 area of mainly dairy farmland in northwestern England and examined for Campylobacter spp. Pulsed-field gel electrophoresis-restriction fragment length polymorphism (PFGE-RFLP) and flaA strain typing of Campylobacter jejuni and Campylobacter coli isolates were done. Data on the water source and the adjacent environment were recorded and examined as explanatory variables. Campylobacter spp. were isolated from 40.5% (n = 119) of the water samples tested. C. jejuni was isolated from 14.3%, C. coli was isolated from 18.5%, and Campylobacter lari was isolated from 4.2% of the samples. Campylobacter hyointestinalis was not isolated from any water source. The difference in prevalence between water types (trough, running, and standing) was significant (P = 0.001). C. jejuni was the species most commonly isolated from trough-water and running-water sources, while C. coli was the most frequently isolated from standing water (P < 0.001). No association was found between the presence of Escherichia coli and that of Campylobacter spp. The final multivariable logistic regression model for Campylobacter spp. included the following variables: water source, soil type, aspect, and amount of cattle fecal material in the environment (fecal pat count). Strain typing demonstrated a diverse population of C. jejuni and the presence of a common C. coli flaA type that was widely distributed throughout the area. Most of the isolates within the common flaA type were discriminated by PFGE-RFLP. These findings suggest a possible role for environmental water in the epidemiology of Campylobacter spp. in a farming environment.

Frequency and spatial distribution of environmental Campylobacter spp.

Humans are exposed to Campylobacter spp. in a range of sources via both food and environmental pathways. For this study, we explored the frequency and distribution of thermophilic Campylobacter spp. in a 10- by 10-km square rural area of Cheshire, United Kingdom. The area contains approximately 70, mainly dairy, farms and is used extensively for outdoor recreational activities. Campylobacter spp. were isolated from a range of environmental samples by use of a systematic sampling grid. Livestock (mainly cattle) and wildlife feces and environmental water and soil samples were cultured, and isolates were presumptively identified by standard techniques. These isolates were further characterized by PCR. Campylobacter jejuni was the most prevalent species in all animal samples, ranging from 11% in samples from nonavian wildlife to 36% in cattle feces, and was isolated from 15% of water samples. Campylobacter coli was commonly found in water (17%) and sheep (21%) samples, but rarely in other samples. Campylobacter lari was recovered from all sample types, with the exception of sheep feces, and was found in moderate numbers in birds (7%) and water (5%). Campylobacter hyointestinalis was only recovered from cattle (7%) and birds (1%). The spatial distribution and determinants of C. jejuni in cattle feces were examined by the use of model-based spatial statistics. The distribution was consistent with very localized within-farm or within-field transmission and showed little evidence of any larger-scale spatial dependence. We concluded that there is a potentially high risk of human exposure to Campylobacter spp., particularly C. jejuni, in the environment of our study area. The prevalence and likely risk posed by C. jejuni-positive cattle feces in the environment diminished as the fecal material aged. After we took into account the age of the fecal material, the absence or presence of rain, and the presence of bird feces, there was evidence of significant variation in the prevalence of C. jejuni-positive cattle feces between grazing fields but no evidence of spatial clustering beyond this resolution. The spatial pattern of C. jejuni is therefore consistent with that for an organism that is ubiquitous in areas contaminated with cattle feces, with a short-scale variation in infection intensity that cannot be explained solely by variations in the age of the fecal material. The observed pattern is not consistent with large-scale transmission attributable to watercourses, wildlife territories, or other geographical features that transcend field and farm boundaries.

Genotypic and antibiotic susceptibility characteristics of a Campylobacter coli population isolated from dairy farmland in the United Kingdom.

Campylobacter infections are the most common cause of bacterial enteritis in humans, and nearly 8% of such infections are caused by Campylobacter coli. Most studies have concentrated on Campylobacter jejuni, frequently isolated from intensively farmed poultry and livestock production units, and few studies have examined the spread and relatedness of Campylobacter across a range of geographical and host boundaries. Systematic sampling of a 100-km2 area of mixed farmland in northwest England yielded 88 isolates of C. coli from a range of sample types and locations, and water was heavily represented. Screening for antibiotic resistance revealed a very low prevalence of resistance, while genotyping performed by using three methods (flaA PCR restriction fragment length polymorphism [RFLP], pulsed-field gel electrophoresis [PFGE], and fluorescent amplified fragment length polymorphism [fAFLP]) provided insights into the genomic relatedness of isolates from different locations and hosts. Isolates were classified into 23 flaA groups, 34 PFGE groups, and five major fAFLP clusters. PFGE banding analysis revealed a high level of variability and no clustering by sample type. fAFLP and flaA analyses successfully grouped the isolates by sample type. We report preliminary findings suggesting that there is a strain of C. coli which may have become adapted to survival or persistence in water and that there is a group of mainly water-derived isolates from which unusual flaA PCR fragments were recovered.