Polymerase chain reaction-gene probe detection of microorganisms by using filter-concentrated samples.

To detect low levels of microorganisms in environmental samples by using polymerase chain reaction (PCR)-gene probe detection, samples were concentrated by filtration. Fluoropore (Millipore Corp.) filters were compatible with PCR DNA amplification, whereas various other filters including nitrocellulose and cellulose acetate filters inhibited PCR amplification. By concentrating cells on Fluoropore filters and releasing the DNA by freeze-thaw cycling, PCR DNA amplification could be performed without removing the filter. Concentration with Fluoropore FHLP and FGLP filters permitted the detection of single cells of microorganisms in 100-ml samples by PCR-gene probes.     

Polymerase chain reaction-gene probe detection of microorganisms by using filter-concentrated samples.

To detect low levels of microorganisms in environmental samples by using polymerase chain reaction (PCR)-gene probe detection, samples were concentrated by filtration. Fluoropore (Millipore Corp.) filters were compatible with PCR DNA amplification, whereas various other filters including nitrocellulose and cellulose acetate filters inhibited PCR amplification. By concentrating cells on Fluoropore filters and releasing the DNA by freeze-thaw cycling, PCR DNA amplification could be performed without removing the filter. Concentration with Fluoropore FHLP and FGLP filters permitted the detection of single cells of microorganisms in 100-ml samples by PCR-gene probes.     

Comparison of gauze swabs and membrane filters for isolation of Campylobacter spp. from surface water

The epidemiology of Campylobacter jejuni indicates that waterborne transmission is important; the organism has been isolated from seawater, fresh water, and estuarine sites. Membrane filtration, with and without use of an enrichment broth, has been the most common method for isolating C. jejuni from water. We evaluated two methods for isolating C. jejuni from water: membrane filtration and gauze filtration. The membrane filters evaluated included 0.22- and 0.45-micron-pore Millipore filters (Millipore Corp., Bedford, Mass.), 0.2- and 0.4-micron-pore Nuclepore filters (Nucleopore Corp., Pleasanton, Calif.), and a 0.45-micron-pore Zetapor filters (AMF Cuno, Meridian, Conn.). The gauze filters included both Moore and Spira swabs. Of the membrane filters evaluated, the 0.45-micron-pore Millipore and Zetapor filters were the most sensitive for recovery of C. jejuni from seeded waters. The 0.45-micron-pore Millipore filter placed in Oosterom broth was better for recovery of C. jejuni from seeded stationary surface waters than either the Spira or Moore swab. However, the 0.45-micron-pore Millipore filter placed on a plate or in enrichment broth was equivalent to the Spira gauze swab when used to examine water from Atlanta area streams. C. jejuni organisms were isolated from 9 of 24 surface water samples representing 5 of 12 streams.     

Comparison of gauze swabs and membrane filters for isolation of Campylobacter spp. from surface water.

The epidemiology of Campylobacter jejuni indicates that waterborne transmission is important; the organism has been isolated from seawater, fresh water, and estuarine sites. Membrane filtration, with and without use of an enrichment broth, has been the most common method for isolating C. jejuni from water. We evaluated two methods for isolating C. jejuni from water: membrane filtration and gauze filtration. The membrane filters evaluated included 0.22- and 0.45-micron-pore Millipore filters (Millipore Corp., Bedford, Mass.), 0.2- and 0.4-micron-pore Nuclepore filters (Nucleopore Corp., Pleasanton, Calif.), and a 0.45-micron-pore Zetapor filters (AMF Cuno, Meridian, Conn.). The gauze filters included both Moore and Spira swabs. Of the membrane filters evaluated, the 0.45-micron-pore Millipore and Zetapor filters were the most sensitive for recovery of C. jejuni from seeded waters. The 0.45-micron-pore Millipore filter placed in Oosterom broth was better for recovery of C. jejuni from seeded stationary surface waters than either the Spira or Moore swab. However, the 0.45-micron-pore Millipore filter placed on a plate or in enrichment broth was equivalent to the Spira gauze swab when used to examine water from Atlanta area streams. C. jejuni organisms were isolated from 9 of 24 surface water samples representing 5 of 12 streams.     

Detection of small numbers of Campylobacter jejuni and Campylobacter coli cells in environmental water, sewage, and food samples by a seminested PCR assay

A rapid and sensitive assay was developed for detection of small numbers of Campylobacter jejuni and Campylobacter coli cells in environmental water, sewage, and food samples. Water and sewage samples were filtered, and the filters were enriched overnight in a nonselective medium. The enrichment cultures were prepared for PCR by a rapid and simple procedure consisting of centrifugation, proteinase K treatment, and boiling. A seminested PCR based on specific amplification of the intergenic sequence between the two Campylobacter flagellin genes, flaA and flaB, was performed, and the PCR products were visualized by agarose gel electrophoresis. The assay allowed us to detect 3 to 15 CFU of C. jejuni per 100 ml in water samples containing a background flora consisting of up to 8, 700 heterotrophic organisms per ml and 10,000 CFU of coliform bacteria per 100 ml. Dilution of the enriched cultures 1:10 with sterile broth prior to the PCR was sometimes necessary to obtain positive results. The assay was also conducted with food samples analyzed with or without overnight enrichment. As few as 相似文献   

Specific detection and confirmation of Campylobacter jejuni by DNA hybridization and PCR.

Conventional detection and confirmation methods for Campylobacter jejuni are lengthy and tedious. A rapid hybridization protocol in which a 1,475-bp chromogen-labelled DNA probe (pDT1720) and Campylobacter strains filtered and grown on 0.22-micron-pore-size hydrophobic grid membrane filters (HGMFs) are used was developed. Among the environmental and clinical isolates of C. jejuni, Campylobacter coli, Campylobacter jejuni subsp. doylei, Campylobacter lari, and Arcobacter nitrofigilis and a panel of 310 unrelated bacterial strains tested, only C. jejuni and C. jejuni subsp. doylei isolates hybridized with the probe under stringent conditions. The specificity of the probe was confirmed when the protocol was applied to spiked skim milk and chicken rinse samples. Based on the nucleotide sequence of pDT1720, a pair of oligonucleotide primers was designed for PCR amplification of DNA from Campylobacter spp. and other food pathogens grown overnight in selective Mueller-Hinton broth with cefoperazone and growth supplements. All C. jejuni strains tested, including DNase-producing strains and C. jejuni subsp. doylei, produced a specific 402-bp amplicon, as confirmed by restriction and Southern blot analysis. The detection range of the assay was as low as 3 CFU per PCR to as high as 10(5) CFU per PCR for pure cultures. Overnight enrichment of chicken rinse samples spiked initially with as little as approximately 10 CFU/ml produced amplicons after the PCR. No amplicon was detected with any of the other bacterial strains tested or from the chicken background microflora. Since C. jejuni is responsible for 99% of Campylobacter contamination in poultry, PCR and HGMF hybridization were performed on naturally contaminated chicken rinse samples, and the results were compared with the results of conventional cultural isolation on Preston agar. All samples confirmed to be culture positive for C. jejuni were also identified by DNA hybridization and PCR amplification, thus confirming that these DNA-based technologies are suitable alternatives to time-consuming conventional detection methods. DNA hybridization, besides being sensitive, also has the potential to be used in direct enumeration of C. jejuni organisms in chicken samples.     

Direct polymerase chain reaction detection of Campylobacter jejuni and Campylobacter coli in raw milk and dairy products.

A polymerase chain reaction (PCR) method designed to sensitively detect and identify Campylobacter jejuni and Campylobacter coli without the need for isolating and culturing strains is described. The intergenic sequence between the flagellin genes flaA and flaB was amplified and characterized with a triple primer or seminested primer approach. A total of 50 bacterial strains, 27 of C. jejuni and C. coli and 23 of other species, were tested, giving no false-positive or false-negative results. The detection limit as determined by ethidium bromide staining of amplification products on agarose gels was 10 bacteria or less in artificially contaminated water, milk, and soft cheese samples with the seminested primer PCR assay. As an application of the PCR system, a set of 93 samples of milk and other dairy products was screened for the presence of C. jejuni and C. coli. We identified six positive samples (6.5%), while none were found with a conventional culture method.     

Improved method for the isolation of Campylobacter jejuni and Campylobacter coli bacteriophages

A centrifugation and filtration method of isolating Campylobacter phages has been developed. Forty-nine Campylobacter phages were isolated from 272 effluent samples of which 42 produced lysis with Campylobacter jejuni strains and seven with C. coli strains. Phages were recovered from pig manure, abattoir effluents, human faeces, sewage and poultry manure. Phages were not isolated from water samples, cattle and sheep faeces or farm pasture soil.     

Evaluation of filters for recovery of Campylobacter jejuni from water

Campylobacter jejuni has been incriminated in several large waterborne outbreaks, but it has rarely been isolated from water itself. Better methodology is needed for the isolation of C. jejuni from water. We evaluated three types of 0.45-micron microporous filters and three different pore sizes of positively charged depth filters for their ability to recover C. jejuni from seeded, sterile tap and surface water. The microporous filters tested were Millipore HA, Gelman GN6, and Zetapor. Three pore sizes of Zeta Plus depth filters (05S, 30S, and 50S) were evaluated by using an adsorption-elution technique. The overall percent recovery in both tap and surface water by microporous filters was: Zetapor, 66%; Millipore HA, 33%; and Gelman GN6, 33%. Adsorption-elution with Zeta Plus 50S allowed 89% recovery of C. jejuni. These data suggest that both the positively charged Zetapor microporous filter and the Zeta Plus 50S depth filter are effective filters for the recovery of C. jejuni from water.     

Evaluation of filters for recovery of Campylobacter jejuni from water.

Campylobacter jejuni has been incriminated in several large waterborne outbreaks, but it has rarely been isolated from water itself. Better methodology is needed for the isolation of C. jejuni from water. We evaluated three types of 0.45-micron microporous filters and three different pore sizes of positively charged depth filters for their ability to recover C. jejuni from seeded, sterile tap and surface water. The microporous filters tested were Millipore HA, Gelman GN6, and Zetapor. Three pore sizes of Zeta Plus depth filters (05S, 30S, and 50S) were evaluated by using an adsorption-elution technique. The overall percent recovery in both tap and surface water by microporous filters was: Zetapor, 66%; Millipore HA, 33%; and Gelman GN6, 33%. Adsorption-elution with Zeta Plus 50S allowed 89% recovery of C. jejuni. These data suggest that both the positively charged Zetapor microporous filter and the Zeta Plus 50S depth filter are effective filters for the recovery of C. jejuni from water.     

The heterogeneity of Campylobacter jejuni and Campylobacter coli strains isolated from healthy cattle

AIMS: To identify campylobacters isolated from clinically healthy cattle at species level by a multiplex polymerase chain reaction (m-PCR). The heterogeneity among Campylobacter jejuni and Campylobacter coli isolates was also investigated by using a restriction fragment length polymorphism (RFLP) analysis of flagellin (flaA) gene. METHODS AND RESULTS: Samples of intestinal contents, gall bladders, liver and faeces were collected from a total number of 1154 healthy cattle. The samples were inoculated onto Preston enrichment broth and agar. Of 1154 samples, 301 (26.1%) were positive for Campylobacter spp. Using an m-PCR assay for species identification, 179 (59.5%) were positive with C. jejuni specific primers while 30 (10%) were positive with C. coli specific primers. None of the liver samples examined was positive for C. jejuni or C. coli by mPCR. All the isolates identified as C. jejuni and C. coli were successfully subtyped by flaA typing. Of the 209 isolates tested, 28 different flaA types were found. Twenty-three flaA types were identified among 179 C. jejuni isolates and the remaining five from C. coli isolates. CONCLUSIONS: Although the overall results suggest that the degree of heterogeneity among the flaA genes of thermophilic Campylobacter strains isolated from healthy cattle is relatively high, they should be treated cautiously as the number of band types for C. coli was low and band type 8 in C. jejuni was represented by a high percentage (%58). SIGNIFICANCE AND IMPACT OF THE STUDY: The findings of the present study suggest that healthy cattle can play role in the contamination of environment and human food chain by Campylobacter spp.     

Detection and genotyping of Arcobacter and Campylobacter isolates from retail chicken samples by use of DNA oligonucleotide arrays

To explore the use of DNA microarrays for pathogen detection in food, we produced DNA oligonucleotide arrays to simultaneously determine the presence of Arcobacter and the presence of Campylobacter in retail chicken samples. Probes were selected that target housekeeping and virulence-associated genes in both Arcobacter butzleri and thermotolerant Campylobacter jejuni and Campylobacter coli. These microarrays showed a high level of probe specificity; the signal intensities detected for A. butzleri, C. coli, or C. jejuni probes were at least 10-fold higher than the background levels. Specific identification of A. butzleri, C. coli, and C. jejuni was achieved without the need for a PCR amplification step. By adapting an isolation method that employed membrane filtration and selective media, C. jejuni isolates were recovered from package liquid from whole chicken carcasses prior to enrichment. Increasing the time of enrichment resulted in the isolation of A. butzleri and increased the recovery of C. jejuni. C. jejuni isolates were further classified by using an additional subset of probes targeting the lipooligosaccharide (LOS) biosynthesis locus. Our results demonstrated that most of the C. jejuni isolates likely possess class B, C, or H LOS. Validation experiments demonstrated that the DNA microarray had a detection sensitivity threshold of approximately 10,000 C. jejuni cells. Interestingly, the use of C. jejuni sequence-specific primers to label genomic DNA improved the sensitivity of this DNA microarray for detection of C. jejuni in whole chicken carcass samples. C. jejuni was efficiently detected directly both in package liquid from whole chicken carcasses and in enrichment broths.     

Advantages of peptide nucleic acid oligonucleotides for sensitive site directed 16S rRNA fluorescence in situ hybridization (FISH) detection of Campylobacter jejuni, Campylobacter coli and Campylobacter lari

Traditionally fluorescence in situ hybridization (FISH) has been performed with labeled DNA oligonucleotide probes. Here we present for the first time a high affinity peptide nucleic acid (PNA) oligonucleotide sequence for detecting thermotolerant Campylobacter spp. using FISH. Thermotolerant Campylobacter spp, including the species Campylobacter coli, Campylobacter jejuni and Campylobacter lari, are important food and water borne pathogens. The designed PNA probe (CJE195) bound with higher affinity to a previously reported low affinity site on the 16S rRNA than the corresponding DNA probe. PNA also overcame the problem of the lack of affinity due to the location of the binding site and the variation of the target sequence within species. The PNA probe specificity was tested with several bacterial species, including other Campylobacter spp. and their close relatives. All tested C. coli, C. jejuni and C. lari strains were hybridized successfully. Aging of the Campylobacter cultures caused the formation of coccoid forms, which did not hybridize as well as bacteria in the active growth phase, indicating that the probe could be used to assess the physiological status of targeted cells. The PNA FISH methodology detected C. coli by membrane filtration method from C. coli spiked drinking water samples.     

Dissemination of fluoroquinolone-resistant Campylobacter spp. within an integrated commercial poultry production system

While characterizing the intestinal bacterial community of broiler chickens, we detected epsilon-proteobacterial DNA in the ilea of 3-day-old commercial broiler chicks (J. Lu, U. Idris, B. Harmon, C. Hofacre, J. J. Maurer, and M. D. Lee, Appl. Environ. Microbiol. 69:6816-6824, 2003). The sequences exhibited high levels of similarity to Campylobacter jejuni and Campylobacter coli sequences, suggesting that chickens can carry Campylobacter at a very young age. Campylobacter sp. was detected by PCR in all samples collected from the ilea of chicks that were 3 to 49 days old; however, it was detected only in the cecal contents of chickens that were at least 21 days old. In order to determine whether the presence of Campylobacter DNA in young chicks was due to ingestion of the bacteria in food or water, we obtained commercial broiler hatching eggs, which were incubated in a research facility until the chicks hatched. DNA sequencing of the amplicons resulting from Campylobacter-specific 16S PCR performed with the ileal, cecal, and yolk contents of the day-of-hatching chicks revealed that Campylobacter DNA was present before the chicks consumed food or water. The 16S rRNA sequences exhibited 99% similarity to C. jejuni and C. coli sequences and 95 to 98% similarity to sequences of other thermophilic Campylobacter species, such as C. lari and C. upsaliensis. The presence of C. coli DNA was detected by specific PCR in the samples from chicks obtained from a commercial hatchery; however, no Campylobacter was detected by culturing. In order to determine whether the same strains of bacteria were present in multiple levels of the integrator, we cultured Campylobacter sp. from a flock of broiler breeders and their 6-week-old progeny that resided on a commercial broiler farm. The broiler breeders had been given fluoroquinolone antibiotics, and we sought to determine whether the same fluoroquinolone-resistant strain was present in their progeny. The isolates were typed by pulsed-field gel electrophoresis, which confirmed that the parental and progeny flocks contained the same strain of fluoroquinolone-resistant C. coli. These data indicate that resistant C. coli can be present in multiple levels of an integrated poultry system and demonstrated that molecular techniques or more sensitive culture methods may be necessary to detect early colonization by Campylobacter in broiler chicks.     

PCR-ELISAs for the detection of Campylobacter jejuni and Campylobacter coli in poultry samples

Campylobacter species, primarily Campylobacter jejuni and Campylobacter coli, are regarded as a major cause of human gastrointestinal disease, commonly acquired by eating undercooked chicken. We describe a PCR-ELISA for the detection of Campylobacter species and the discrimination of C. jejuni and C. coli in poultry samples. The PCR assay targets the 16S/23S ribosomal RNA intergenic spacer region of Campylobacter species with DNA oligonucleotide probes designed for the specific detection of C. jejuni, C. coli, and Campylobacter species immobilized on Nucleo-Link wells and hybridized to PCR products modified with a 5' biotin moiety. The limit of detection of the PCR-ELISA was 100-300 fg (40-120 bacterial cells) for C. jejuni and C. coli with their respective species-specific oligonucleotide probes and 10 fg (4 bacterial cells) with the Campylobacter genus-specific probe. Testing of poultry samples, which were presumptive positive for Campylobacter following culture on the Malthus V analyzer, with the PCR-ELISA determined Campylobacter to be present in 100% of samples (n = 40) with mixed cultures of C. jejuni/C. coli in 55%. The PCR-ELISA when combined with culture pre-enrichment is able to detect the presence of Campylobacter and definitively identify C. jejuni and C. coli in culture-enriched poultry meat samples.     

Molecular epidemiology of Campylobacter isolates from poultry production units in southern Ireland

This study aimed to identify the sources and routes of transmission of Campylobacter in intensively reared poultry farms in the Republic of Ireland. Breeder flocks and their corresponding broilers housed in three growing facilities were screened for the presence of Campylobacter species from November 2006 through September 2007. All breeder flocks tested positive for Campylobacter species (with C. jejuni and C. coli being identified). Similarly, all broiler flocks also tested positive for Campylobacter by the end of the rearing period. Faecal and environmental samples were analyzed at regular intervals throughout the rearing period of each broiler flock. Campylobacter was not detected in the disinfected house, or in one-day old broiler chicks. Campylobacter jejuni was isolated from environmental samples including air, water puddles, adjacent broiler flocks and soil. A representative subset of isolates from each farm was selected for further characterization using flaA-SVR sub-typing and multi-locus sequence typing (MLST) to determine if same-species isolates from different sources were indistinguishable or not. Results obtained suggest that no evidence of vertical transmission existed and that adequate cleaning/disinfection of broiler houses contributed to the prevention of carryover and cross-contamination. Nonetheless, the environment appears to be a potential source of Campylobacter. The population structure of Campylobacter isolates from broiler farms in Southern Ireland was diverse and weakly clonal.     

Use of PCR for direct detection of Campylobacter species in bovine feces

This study reports on the use of PCR to directly detect and distinguish Campylobacter species in bovine feces without enrichment. Inhibitors present in feces are a major obstacle to using PCR to detect microorganisms. The QIAamp DNA stool minikit was found to be an efficacious extraction method, as determined by the positive amplification of internal control DNA added to bovine feces before extraction. With nested or seminested multiplex PCR, Campylobacter coli, C. fetus, C. hyointestinalis, and C. jejuni were detected in all fecal samples inoculated at approximately 10(4) CFU g(-1), and 50 to 83% of the samples inoculated at approximately 10(3) CFU g(-1) were positive. At approximately 10(2) CFU g(-1), C. fetus, C. hyointestinalis, and C. jejuni (17 to 50% of the samples) but not C. coli were detected by PCR. From uninoculated bovine feces, a total of 198 arbitrarily selected isolates of Campylobacter were recovered on four commonly used isolation media incubated at three temperatures. The most frequently isolated taxa were C. jejuni (152 isolates) and C. lanienae (42 isolates), but isolates of C. fetus subsp. fetus, Arcobacter butzleri, and A. skirrowii also were recovered (相似文献   

A specific DNA probe for the identification of Campylobacter jejuni

A 6.1 kb DNA probe for the human enteric pathogen Campylobacter jejuni has been isolated from a genomic library constructed in the plasmid vector pBR322 in Escherichia coli. The DNA sequence used as a probe was identified from recombinant plasmids following immunological screening of transformants using polyclonal antisera to whole cells and to membrane antigens of C. jejuni. Restriction endonuclease fragment mapping of C. jejuni DNA inserts from three of the recombinant plasmids showed an overlapping DNA fragment. One of these recombinant plasmids, when used as a DNA probe in Southern hybridization, specifically hybridized with chromosomal DNA from all of the C. jejuni strains tested. Hybridization was not detected at high stringency between the DNA probe and chromosomal DNA from any other Campylobacter species tested except weakly with the chromosomal DNA of strains of Campylobacter coli. Hybridization was also not detected with chromosomal DNA from a range of other enteric bacteria likely to be encountered in faecal material. The intensity of hybridization with C. coli could be increased by reducing the stringency of hybridization.     

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.