Consumer acceptability of interventions to reduce Campylobacter in the poultry food chain.

Reducing human Campylobacter cases has become a priority for the UK Government. However the public's views on acceptability of interventions to reduce Campylobacter in poultry production are poorly understood in the UK and in other countries around the world. The objective of the study was to investigate how increasing awareness and knowledge changes consumer acceptability of interventions that reduce human campylobacteriosis in the poultry food chain. This approach is readily applicable to other risks and associated interventions. It involved a survey of the views of consumers in the Grampian region in North East Scotland. This found that better hygiene practices on farm, freezing chicken meat and vaccination of chickens were acceptable to the majority of participants (95%, 53% & 52% respectively) whilst irradiation and chemical wash of chicken meat were acceptable to <50%. Increasing consumer awareness by providing information on the Campylobacter disease burden in humans increased the number of participants finding them acceptable. However, chemical wash and irradiation remained the least acceptable interventions, although highly effective at reducing Campylobacter, and were found to be never acceptable to >50% of respondents. It was found on average that food poisoning concern, previous awareness of Campylobacter and living in rural or urban areas had either no or little effect effect on the acceptability of interventions. Further, previous awareness of Campylobacter did not influence consumer concern of harmful bacteria on chicken meat. Overall, findings indicate that increasing consumer acceptability of the most effective interventions is likely to be a difficult process.

Identifying the seasonal origins of human campylobacteriosis.

Human campylobacteriosis exhibits a distinctive seasonality in temperate regions. This paper aims to identify the origins of this seasonality. Clinical isolates [typed by multi-locus sequence typing (MLST)] and epidemiological data were collected from Scotland. Young rural children were found to have an increased burden of disease in the late spring due to strains of non-chicken origin (e.g. ruminant and wild bird strains from environmental sources). In contrast the adult population had an extended summer peak associated with chicken strains. Travel abroad and UK mainland travel were associated with up to 17% and 18% of cases, respectively. International strains were associated with chicken, had a higher diversity than indigenous strains and a different spectrum of MLST types representative of these countries. Integrating empirical epidemiology and molecular subtyping can successfully elucidate the seasonal components of human campylobacteriosis. The findings will enable public health officials to focus strategies to reduce the disease burden.

A population-based exposure assessment of risk factors associated with gastrointestinal pathogens: a Campylobacter study.

A questionnaire survey was undertaken to determine the exposure of a study population to campylobacteriosis source risk factors (environmental, water, food) and results were stratified by age, population density and deprivation. Data were gathered using an exposure assessment carried out by telephone in the Grampian region of Scotland. Univariate analysis showed that children aged 5-14 years, living in low population density (0-44.4 persons/km2) and affluent areas had elevated exposure to environmental and water risk factors. Multivariate logistic regression analysis revealed that younger age groups and lower population density were significant indicators for most environmental risk factors. The results compared to reported disease incidence in Grampian showed that greater exposure to risk factors does not necessarily coincide with greater disease incidence for age groups, particularly for the 0-4 years age group. Further research is required to explain the relationship between exposure and disease incidence.

Combining risk assessment and epidemiological risk factors to elucidate the sources of human E. coli O157 infection.

E. coli O157 can be transmitted to humans by three primary (foodborne, environmental, waterborne) and one secondary (person-to-person transmission) pathways. A regression model and quantitative microbiological risk assessments (QMRAs) were applied to determine the relative importance of the primary transmission pathways in NE Scotland. Both approaches indicated that waterborne infection was the least important but it was unclear whether food or the environment was the main source of infection. The QMRAs over-predicted the number of cases by a factor of 30 and this could be because all E. coli O157 strains may not be equally infective and/or the level of infectivity in the dose-response model was too high. The efficacy of potential risk mitigation strategies to reduce human exposure to E. coli O157 using QMRAs was simulated. Risk mitigation strategies focusing on food and environment are likely to have the biggest impact on infection figures.

Source attribution, prevalence and enumeration of Campylobacter spp. from retail liver.

Campylobacter prevalence from retail liver (chicken, cattle, pig and sheep) was found to be 81%, 69%, 79% and 78% respectively. Molecular source attribution demonstrated that strains from chicken liver were most similar to those found commonly in humans. This provides further evidence of liver being a probable source of human infection.

The relationship between lay and technical views of Escherichia coli O157 risk.

Here, we bring together and contrast lay (accessible primarily through social science methodologies) and technical (via risk assessment and epidemiological techniques) views of the risk associated with the Escherichia coli O157 pathogen using two case study areas in the Grampian region of Scotland, and North Wales. Epidemiological risk factors of contact with farm animals, visiting farms or farm fields and having a private water supply were associated with postcode districts of higher than average disease incidence in the human population. However, this was not the case for the epidemiological risk factor of consumption of beef burgers, which was independent of disease incidence in the postcode district of residence. The proportion of the population expressing a high knowledge of E. coli O157 was greatest in high-incidence disease districts compared with low-incidence areas (17% cf. 7%). This supports the hypothesis that in high-disease-incidence areas, residents are regularly exposed to information about the disease through local cases, the media, local social networks, etc. or perhaps that individuals are more likely to be motivated to find out about it. However, no statistically significant difference was found between high- and low-incidence postcode districts in terms of the proportion of the population expressing a high likelihood of personal risk of infection (10% cf. 14%), giving a counterintuitive difference between the technical (epidemiological and quantitative microbiological risk assessment (QMRA)) and the lay assessment of E. coli O157 risk. This suggests that lay evaluations of E. coli O157 risk reflect intuitive and experience-based estimates of the risk rather than probabilistic estimates. A generally strong correspondence was found in terms of the rank order given to potential infection pathways, with environment and foodborne infection routes dominating when comparing public understanding with technical modelling results. Two general conclusions follow from the work. First, that integrative research incorporating both lay and technical views of risk is required in order that informed decisions can be made to handle or treat the risk by the groups concerned (e.g. the public, policy makers/risk managers, etc.). Second, when communicating risk, for example, through education programmes, it is important that this process is two-way with risk managers (e.g. including Food Standards Agency officials and communications team, public health infection control and environmental health officers) both sharing information with the public and stakeholder groups, as well as incorporating public knowledge, values and context (e.g. geographical location) into risk-management decisions.

Escherichia coli O157: comparing awareness of rural residents and visitors in livestock farming areas.

This research compared public opinions about Escherichia coli O157 (an increasing environmental hazard associated with livestock) in two farming areas with contrasting incidence of E. coli O157 disease. A questionnaire was administered in rural Grampian (10·8 cases/100,000 population per year) and North Wales (2·5 cases/100,000 population per year). Awareness was highest among farmers in Grampian (91%) and lowest among visitors to both areas (28%). Respondents were more likely to indicate vomiting (76%) than bloody diarrhoea (48%) as a common symptom. Undercooked meat and contact with farm animal faeces were identified by 60% of all respondents as risk factors who described 'basic hygiene' for risk reduction indoors. Visitors view E. coli O157 as a food hazard, not an environmental hazard that produces vomiting not dysentery. Efforts to reduce human infections in livestock farming areas could be improved with proximate reminders for visitors of the environmental pathway of E. coli O157 infection.

Putative household outbreaks of campylobacteriosis typically comprise single MLST genotypes.

During a 15-month period in Scotland a small but important number of human Campylobacter cases (3·2%) arose from 91 putative household outbreaks. Of the 26 outbreaks with known strain composition, 89% were composed of the same MLST which supports the potential use of MLST in public health epidemiology. The number of cases associated with household outbreaks is much larger than general outbreaks and there is some evidence to indicate that there may be secondary transmission, although this is relatively rare.

Multi-locus sequence types of Campylobacter carried by flies and slugs acquired from local ruminant faeces.

AIMS: To assess whether flies and slugs acquire strains of Campylobacter jejuni and Campylobacter coli present in local ruminant faeces. METHODS AND RESULTS: Campylobacter was cultured from flies, slugs and ruminant faeces that were collected from a single farm in Scotland over a 19-week period. The isolates were typed using multi-locus sequence typing (MLST) and compared with isolates from cattle and sheep faeces. Campylobacter jejuni and Camp. coli were isolated from 5·8% (n=155, average of 75 flies per pool) and 13·3% (n=15, average of 8·5 slugs per pool) of pooled fly and slug samples, respectively. The most common sequence type (ST) in flies was Camp. coli ST-962 (approx. 40%) regardless of the prevalence in local cattle (2·3%) or sheep (25·0%) faeces. Two positive slug pools generated the same ST that has not been reported elsewhere. CONCLUSIONS: Despite their low carriage rate, flies are able to acquire Campylobacter STs that are locally present, although the subset carried may be biased when compared to local source. Slugs were shown to carry a previously unreported Campylobacter ST. SIGNIFICANCE AND IMPACT OF THE STUDY: This study has demonstrated that flies carry viable Campylobacter and may contribute to the transfer of STs within and between groups of animals on farms. Further, they may therefore present a risk to human health via their contact with ready-to-eat foods or surfaces.

Campylobacter immunity and coinfection following a large outbreak in a farming community.

An outbreak of campylobacteriosis affected approximately one-half of 165 people attending an annual farmers' dance in Montrose, Scotland, in November 2005. Epidemiological investigations, including a cohort study (n = 164), identified chicken liver paté as the most likely vehicle of infection. Paté preparation involved deliberate undercooking of chicken livers by flash-frying, followed by mechanical homogenization. Typing of 32 Campylobacter strains (isolated from submitted stools) by multilocus sequence typing identified four distinct clades of Campylobacter jejuni. There was good agreement when isolates were typed by Penner serotyping, pulsed-field gel electrophoresis, and flaA short variable region sequencing but poorer agreement with phage and antibiotic susceptibility testing. At least three attendees were coinfected with two Campylobacter strains each. The outbreak was probably due to several livers contributing Campylobacter strains that survived undercooking and were dispersed throughout the paté. The study highlights improper culinary procedures as a potential human health risk and provides a striking counterexample to the "dominant outbreak strain" view of point source outbreaks of food-borne infections. It also demonstrates that previous exposure to biologically plausible sources of Campylobacter may confer protection against subsequent infection.

Microbial pollution of water by livestock: approaches to risk assessment and mitigation.

In this study, we investigate the extent to which the incidence of Escherichia coli O157:H7 can be predicted in human faeces, from human intake and infection via water contaminated by livestock and carrying this zoonotic pathogen in North-East (NE) and South-West (SW) regions of Scotland. In SW Scotland, there is a risk of coastal recreational waters failing EU standards for faecal indicator organisms, and this is assumed to be the main potential waterborne route of infection. In NE Scotland, the main waterborne route is assumed to be the many private drinking water supplies; these are mainly derived from shallow groundwater and surveys show that there is potential for significant levels of microbial contamination from livestock. The risk to human health from these sources has been assessed using a combination of process models, epidemiological risk-assessment methods and survey data. A key assumption in the calculations is the amount of mixing of pathogenic and non-pathogenic E. coli between animal faecal sources and contaminated water intake by humans. Using the probability distributions of the E. coli O157 content of individual faecal pat material (which would imply no mixing between source and human intake), based on three recent surveys of animal faeces in Scotland, led to predicted annual risks of infection slightly higher than observed human infection incidence. Using the geometric mean to represent partial mixing (which theoretically may over- or underestimate incidence with a concave dose-response curve) gave infection rates similar to those observed for two of the three faecal surveys. Using the arithmetic mean led to over-prediction of risk. This is to be expected if the true dose-response curve is (such as the Beta-Poisson curve used here) concave. Other factors that may lead to over-prediction of incidence are discussed, including under-reporting, loss of infectivity as a result of environmental exposure, immunity and the appropriateness of the Beta-Poisson curve. It is concluded that better epidemiological data for calibration of the dose-response curve, better knowledge of the degree of mixing and understanding of immunity are key requirements for progress in process model-based predictions of infection rate. The paper also explores the potential of improved farm and catchment scale management to deliver cost-effective mitigation of pollution of bathing and drinking water by livestock zoonoses.

Diversity of Escherichia coli O157 in a longitudinal farm study using multiple-locus variable-number tandem-repeat analysis.

AIMS: To perform a longitudinal study of the diversity of Escherichia coli O157 from a ruminant pasture/stream environment using multiple-locus variable-number tandem-repeat analysis (MLVA). METHODS AND RESULTS: Samples of faecal droppings from grazing ruminants and from an adjacent stream were tested longitudinally for E. coli O157 by enrichment and immunomagnetic separation (IMS). Using MLVA, 24 different profiles were identified from a total of 231 E. coli O157 isolates, of which 80 were included in a similarity analysis. Four main clusters with several subclusters were observed. Although there was close contact between sheep and cattle during the study period, E. coli O157 was surprisingly not detected from cattle faeces. CONCLUSIONS: The cluster analysis indicated both unrelated and closely related E. coli O157 strains. The choice of loci to target in MLVA is important for the subtyping result, as loci with high diversities are essential for discriminating between closely related isolates. SIGNIFICANCE AND IMPACT OF THE STUDY: There is a lack of data available on the use of MLVA to describe E. coli O157 diversity and changes over time in the animal reservoirs and the environment. Such data are needed in order to further develop MLVA as a typing method.

Leaching of bioluminescent Escherichia coli O157:H7 from sheep and cattle faeces during simulated rainstorm events.

AIMS: Development of a novel inoculation technique to improve the current methods of determining the leaching of Escherichia coli O157:H7 from faeces. METHODS AND RESULTS: Ruminant faeces were inoculated with a high [c. 10(7) colony forming units (CFU) g(-1)] or low (c. 10(4) CFU g(-1)) load of a lux-marked strain of E. coli O157:H7 via injection, and subjected to four simulated heavy rainfall events. The population density and metabolic activity of E. coli O157:H7 recovered within the leachate was determined following each simulated rain event and compared with the indigenous E. coli population. The concentration of E. coli O157:H7 in the leachates followed a similar trend to that of nonpathogenic E. coli. Significantly greater densities of generic and pathogenic E. coli were recovered in the leachates generated from sheep faeces compared with cattle faeces. Pathogen metabolic activity was also significantly greater in sheep faeces. CONCLUSIONS: Our findings show that E. coli O157:H7 may readily leach from ruminant faeces during rain events. The bacterium leaches more freely from sheep faeces than from cattle faeces and displays greater metabolic activity within sheep leachate. SIGNIFICANCE AND IMPACT OF THE STUDY: A novel inoculation technique was developed that allowed the determination of both population density and cellular activity of E. coli O157:H7 in leachate derived from faeces.

Sexual dimorphism in campylobacteriosis.

Sexual dimorphism in infectious diseases whereby disease incidence is more prevalent in one gender has been reported repeatedly in the scientific literature. Both behavioural and physiological differences have been suggested as a cause of this gender bias but there is a paucity of data to support either of these viewpoints. Here it is hypothesized that for campylobacteriosis physiological factors play an important role in the higher incidence in males. We demonstrate in the human population (from several countries in three continents) that this bias exists in young children (<1 year) where behavioural differences between genders are likely to be minimal. Further we demonstrate this difference in an animal model where both infection rates and shedding rates of the organism are greater in male mice.

Hierarchical dose response of E. coli O157:H7 from human outbreaks incorporating heterogeneity in exposure.

The infectivity of pathogenic microorganisms is a key factor in the transmission of an infectious disease in a susceptible population. Microbial infectivity is generally estimated from dose-response studies in human volunteers. This can only be done with mildly pathogenic organisms. Here a hierarchical Beta-Poisson dose-response model is developed utilizing data from human outbreaks. On the lowest level each outbreak is modelled separately and these are then combined at a second level to produce a group dose-response relation. The distribution of foodborne pathogens often shows strong heterogeneity and this is incorporated by introducing an additional parameter to the dose-response model, accounting for the degree of overdispersion relative to Poisson distribution. It was found that heterogeneity considerably influences the shape of the dose-response relationship and increases uncertainty in predicted risk. This uncertainty is greater than previously reported surrogate and outbreak models using a single level of analysis. Monte Carlo parameter samples (alpha, beta of the Beta-Poisson model) can be readily incorporated in risk assessment models built using tools such as S-plus and @ Risk.

Use of multilocus sequence typing to investigate the association between the presence of Campylobacter spp. in broiler drinking water and Campylobacter colonization in broilers.

The presence of campylobacters in broiler chickens and throughout the broiler water delivery systems of 12 farms in northeastern Scotland was investigated by sensitive enrichment methods and large-volume filtration. Campylobacter presence was independent of the water source and whether the water was treated. The genotypes of Campylobacter jejuni isolates recovered from chickens and various locations within the water delivery systems were compared by multilocus sequence typing. Matching strains in shed header tanks and birds were found at 1 of the 12 farms investigated. However, the sequence of contamination or whether the source was within or outside the shed was not determined. Nevertheless, these data provide evidence that drinking water could be associated with broiler infection by campylobacters.

Concentration and prevalence of Escherichia coli O157 in sheep faeces at pasture in Scotland.

AIMS: To study the presence, numbers and virulence profiles of Escherichia coli O157 in sheep faeces and validate the microbiological methods used to attain these data. METHODS AND RESULTS: Flock level prevalence was found to be 40% (six from 15) and 6.5% of faecal samples tested were found to be positive. Two farms gave samples defined as high shedding (>10(4) CFU g(-1)), one of which comprised 91% positive samples with 13/33 at the high shedding level. CONCLUSIONS: These data confirmed that sheep are an important reservoir of E. coli O157. SIGNIFICANCE AND IMPACT OF THE STUDY: Sheep play a significant role in the maintenance and dispersal of E. coli O157 in the farming environment and are an important source of human infection.

A stochastic model simulating the capture of pathogenic micro-organisms by superparamagnetic particles in an isodynamic magnetic field.

The method of immunomagnetic separation (IMS) has become an established technique to concentrate and separate animal cells, biologically active compounds and pathogenic micro-organisms from clinical, food and environmental matrices. One drawback of this technique is that the analysis is only possible for small sample volumes. We have developed a stochastic model that involves numerical simulations to optimize the process of concentration of pathogenic micro-organisms onto superparamagnetic carrier particles (SCPs) in a gradient magnetic field. Within the range of the system parameters varied in the simulations, optimal conditions favour larger particles with higher magnetite concentrations. The dependence on magnetic field intensity and gradient together with concentration of particles and micro-organisms was found to be less important for larger SCPs but these parameters can influence the values of the collision time for small particles. These results will be useful in aiding the design of apparatus for immunomagnetic separation from large volume samples.

Magnetizable needles and wires--modeling an efficient way to target magnetic microspheres in vivo.

The in vivo targeting of tumors with magnetic microspheres is currently realized through the application of external non-uniform magnetic fields generated by rare-earth permanent magnets or electromagnets. Our theoretical work suggests a feasible procedure for local delivery of magnetic nano- and microparticles to a target area. In particular, thin magnetizable wires placed throughout or close to the target area and magnetized by a perpendicular external uniform background magnetic field are used to concentrate magnetic microspheres injected into the target organ's natural blood supply. The capture of the magnetic particles and the building of deposits thereof in the blood vessels of the target area were modeled under circumstances similar to the in vivo situation. This technique could be applied to magnetically targeted cancer therapy or magnetic embolization therapy with magnetic particles that contain anticancer agents, such as chemotherapeutic drugs or therapeutic radioisotopes.