Campylobacter jejuni is the leading cause of bacterial gastro-enteritis in the developed world. It is thought to infect 2–3 million people a year in the US alone, at a cost to the economy in excess of US $4 billion. C. jejuni is a widespread zoonotic pathogen that is carried by animals farmed for meat and poultry. A connection with contaminated food is recognized, but C. jejuni is also commonly found in wild animals and water sources. Phylogenetic studies have suggested that genotypes pathogenic to humans bear greatest resemblance to non-livestock isolates. Moreover, seasonal variation in campylobacteriosis bears the hallmarks of water-borne disease, and certain outbreaks have been attributed to contamination of drinking water. As a result, the relative importance of these reservoirs to human disease is controversial. We use multilocus sequence typing to genotype 1,231 cases of C. jejuni isolated from patients in Lancashire, England. By modeling the DNA sequence evolution and zoonotic transmission of C. jejuni between host species and the environment, we assign human cases probabilistically to source populations. Our novel population genetics approach reveals that the vast majority (97%) of sporadic disease can be attributed to animals farmed for meat and poultry. Chicken and cattle are the principal sources of C. jejuni pathogenic to humans, whereas wild animal and environmental sources are responsible for just 3% of disease. Our results imply that the primary transmission route is through the food chain, and suggest that incidence could be dramatically reduced by enhanced on-farm biosecurity or preventing food-borne transmission.

Responsible for the majority of bacterial gastroenteritis in the developed world, Campylobacter jejuni is a pervasive pathogen of humans and animals, but its evolution is obscure. In this paper, we exploit contemporary genetic diversity and empirical evidence to piece together the evolutionary history of C. jejuni and quantify its evolutionary potential. Our combined population genetics–phylogenetics approach reveals a surprising picture. Campylobacter jejuni is a rapidly evolving species, subject to intense purifying selection that purges 60% of novel variation, but possessing a massive evolutionary potential. The low mutation rate is offset by a large effective population size so that a mutation at any site can occur somewhere in the population within the space of a week. Recombination has a fundamental role, generating diversity at twice the rate of de novo mutation, and facilitating gene flow between C. jejuni and its sister species Campylobacter coli. We attempt to calibrate the rate of molecular evolution in C. jejuni based solely on within-species variation. The rates we obtain are up to 1,000 times faster than conventional estimates, placing the C. jejuni–C. coli split at the time of the Neolithic revolution. We weigh the plausibility of such recent bacterial evolution against alternative explanations and discuss the evidence required to settle the issue.

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.

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.

SUMMARYIn 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.