Inhibition of phage-resistant bacterial pathogen re-growth with the combined use of bacteriophages and EDTA.

The combined effects of ethylenediaminetetraacetic acid (EDTA) and bacteriophage (phage) treatment of foodborne pathogens were investigated. Although viable counts for Campylobacter jejuni decreased by 1.5 log after incubation for 8 h in the presence of phage PC10, re-growth was observed thereafter. The combination of phage PC10 and 1 mM EDTA significantly inhibited the re-growth of C. jejuni. The viable counts for C. jejuni decreased by 2.6 log (P < 0.05) compared with that of the initial count after 24 h. Moreover, EDTA at 0.67 or 1.3 mM, combined with the specific lytic phages, also effectively inhibited the re-growth of phage-resistant cells of Campylobacter coli, Salmonella enterica serovar Enteritidis, and Salmonella enterica serovar Typhimurium. In addition, the combined effects of lytic phages and EDTA were investigated on the viability of Campylobacter in BHI broth at low temperatures followed by the optimum growth temperature. The re-growth of C. coli was significantly inhibited by the coexistence of 1.3 mM EDTA, and the viable counts of surviving bacteria was about the same as the initial viable count after the incubation. This is the first study demonstrating the combined use of lytic phages and EDTA is effective in inhibiting the re-growth of phage-resistant bacteria in Gram-negative bacteria.

Staphylococcus aureus enhances biofilm formation, aerotolerance, and survival of Campylobacter strains isolated from retail meats.

In retail meat products, Campylobacter jejuni, C. coli, and Staphylococcus aureus have been reported in high prevalence. The polymicrobial interaction between Campylobacter and other bacteria could enhance Campylobacter survival during the adverse conditions encountered during retail meat processing and storage. This study was designed to investigate the potential role of S. aureus from retail meats in enhancing the survival of Campylobacter exposed to low temperature, aerobic conditions, and biofilm formation. Results indicated that viable S. aureus cells and filter-sterilized cell-free media obtained from S. aureus prolonged the survival of Campylobacter at low temperature and during aerobic conditions. Biofilm formation of Campylobacter strains was significantly enhanced in the presence of viable S. aureus cells, but the results were inconclusive when extracts from cell-free media were used. In conclusion, the presence of S. aureus cells enhances survivability of Campylobacter strains in adverse conditions such as low temperature and aerobic conditions. Further investigations are warranted to understand the interaction between Campylobacter and S. aureus, and effective intervention strategies are needed to reduce the incidence of both foodborne pathogens in retail meat products.

Survival of Campylobacter jejuni and Campylobacter coli water isolates in lake and well water.

The role of water for transmission of Campylobacter jejuni and C. coli to humans might be underestimated, as factors important for bacterial viability in water are largely unknown. We have studied water survival of seven C. jejuni and eight C. coli isolates originally isolated from Swedish waters, together with selected reference strains, over eight days at 4 °C in the dark in untreated water collected from a local lake and a private well. To study seasonality, lake water samples were collected during spring and autumn. Samples for culturable bacterial counts were taken on days 2, 4, 6, and 8 and compared to the start inoculum. For C. jejuni, a significantly better survival was observed in autumn than in spring lake water. Furthermore, C. jejuni had a significantly better survival than C. coli in autumn lake and well water samples; the rate of culturability loss was almost double for C. coli in autumn lake water. These findings contribute to a better understanding on the seasonality of waterborne Campylobacter infections and the general predominance of C. jejuni.

The Prevalence of Campylobacter spp. in Polish Poultry Meat.

The prevalence, count and molecular identification of Campylobacter spp. in Polish poultry meat were analysed. 181 samples of meat from chicken (70), turkey (47), duck (54) and goose (10) were studied. Campylobacter spp. was found in 64% of meat samples. The highest prevalence of this pathogen was detected for duck meat. On average 80% of duck samples were contaminated with Campylobacter spp. The counts of Campylobacter spp. in positive samples remained under ten colony forming units per gram of product in 59% of poultry meat. C. jejuni was more frequently detected in poultry meat than C. coli.

High survival rates of Campylobacter coli under different stress conditions suggest that more rigorous food control measures might be needed in Brazil.

Campylobacter spp. have been the most commonly reported gastrointestinal bacterial pathogen in many countries. Consumption of improperly prepared poultry meat has been the main transmission route of Campylobacter spp. Although Brazil is the largest exporter of poultry meat in the world, campylobacteriosis has been a neglected disease in the country. The aim of this study was to characterize 50 Campylobacter coli strains isolated from different sources in Brazil regarding the frequency of 16 virulence genes and their survival capability under five different stress conditions. All strains studied presented the cadF, flaA, and sodB genes that are considered essential for colonization. All strains grew at 4 °C and 37 °C after 24 h. High survival rates were observed when the strains were incubated in BHI with 7.5% NaCl and exposed to acid and oxidative stress. In conclusion, the pathogenic potential of the strains studied was reinforced by the presence of several important virulence genes and by the high growth and survival rates of the majority of those strains under different stress conditions. The results enabled a better understanding of strains circulating in Brazil and suggest that more rigorous control measures may be needed, given the importance of contaminated food as vehicles for Campylobacter coli.

Survival of selected foodborne pathogens on dry cured pork loins.

The safety of ready-to-eat products such as cured pork loins must be guaranteed by the food industry. In the present study, the efficacy of the dry curing process of pork loins obtained from free-range pigs in the reduction of three of the most important foodborne pathogens is analysed. A total of 28 pork loin segments, with an average weight of 0.57±0.12kg, were divided into four groups with three being inoculated by immersion with 7logCFU/ml of either Salmonella Typhimurium, Campylobacter coli or Listeria innocua and the last one inoculated by immersion with sterile medium (control group). The loin segments were treated with a seasoning mixture of curing agents and spices, packed in a synthetic sausage casing and cured for 64days. Microbiological analysis, pH and water activity (aw) were assessed at four stages. The values of pH and aw decreased with curing time as expected. S. Typhimurium and C. coli dropped significantly (3.28 and 2.14 log units, respectively), but limited reduction of L. innocua (0.84 log unit) was observed along the curing process. In our study, three factors were considered critical: the initial concentration of the bacteria, the progressive reduction of pH and the reduction of aw values. Our results encourage performing periodic analysis at different stages of the manufacturing of dry cured pork loins to ensure the absence of the three evaluated foodborne pathogens.

Influence of enrichment and isolation media on the detection of Campylobacter spp. in naturally contaminated chicken samples.

Investigating Campylobacter epidemiology requires adequate technique and media to ensure optimal culturing and accurate detection and isolation of Campylobacter strains. In the present study, we investigated the performances of three enrichment durations in Bolton broth (0, 24 and 48h) and compared four isolation media (mCCDA, Karmali, Butzler no. 2 and CampyFood agar (CFA)) for the detection of Campylobacter positive samples and the identification of Campylobacter species, from naturally contaminated broiler chicken samples (caeca, neck skin from carcasses, and skin from thighs). We compared our local results to those we obtained with samples from a European survey (caeca and neck skin) and a national survey (neck skin, thigh skin, and breast). Direct plating favored the detection of positive samples highly contaminated by Campylobacter (caeca and neck skin from carcasses) whatever the media. A longer enrichment reduced the rates of Campylobacter recovery except when using Butzler no. 2, more particularly for neck skin which background microflora was less important than in caeca. As a matter of fact, enrichment allowed a higher detection rate of positive samples with low Campylobacter contamination levels (breast, thigh skin), this detection being enhanced when using Butzler no. 2. When comparing the 3 other selective media, CFA was the 2nd most efficient media prior to mCCDA and Karmali. Interestingly, enrichment promoted the growth of Campylobacter coli but this promotion was least with Butzler no. 2 agar. Our study has confirmed the need to adapt the method to the types of samples for improving the detection of Campylobacter and that the method may affect the prevalence of the species.

Monitoring Antimicrobial Resistance in the Food Supply Chain and Its Implications for FDA Policy Initiatives.

In response to concerning increases in antimicrobial resistance (AMR), the Food and Drug Administration (FDA) has decided to increase veterinary oversight requirements for antimicrobials and restrict their use in growth promotion. Given the high stakes of this policy for the food supply, economy, and human and veterinary health, it is important to rigorously assess the effects of this policy. We have undertaken a detailed analysis of data provided by the National Antimicrobial Resistance Monitoring System (NARMS). We examined the trends in both AMR proportion and MIC between 2004 and 2012 at slaughter and retail stages. We investigated the makeup of variation in these data and estimated the sample and effect size requirements necessary to distinguish an effect of the policy change. Finally, we applied our approach to take a detailed look at the 2005 withdrawal of approval for the fluoroquinolone enrofloxacin in poultry water. Slaughter and retail showed similar trends. Both AMR proportion and MIC were valuable in assessing AMR, capturing different information. Most variation was within years, not between years, and accounting for geographic location explained little additional variation. At current rates of data collection, a 1-fold change in MIC should be detectable in 5 years and a 6% decrease in percent resistance could be detected in 6 years following establishment of a new resistance rate. Analysis of the enrofloxacin policy change showed the complexities of the AMR policy with no statistically significant change in resistance of both Campylobacter jejuni and Campylobacter coli to ciprofloxacin, another second-generation fluoroquinolone.

A transferable plasticity region in Campylobacter coli allows isolates of an otherwise non-glycolytic food-borne pathogen to catabolize glucose.

Thermophilic Campylobacter species colonize the intestine of agricultural and domestic animals commensally but cause severe gastroenteritis in humans. In contrast to other enteropathogenic bacteria, Campylobacter has been considered to be non-glycolytic, a metabolic property originally used for their taxonomic classification. Contrary to this dogma, we demonstrate that several Campylobacter coli strains are able to utilize glucose as a growth substrate. Isotopologue profiling experiments with (13) C-labeled glucose suggested that these strains catabolize glucose via the pentose phosphate and Entner-Doudoroff (ED) pathways and use glucose efficiently for de novo synthesis of amino acids and cell surface carbohydrates. Whole genome sequencing of glycolytic C. coli isolates identified a genomic island located within a ribosomal RNA gene cluster that encodes for all ED pathway enzymes and a glucose permease. We could show in vitro that a non-glycolytic C. coli strain could acquire glycolytic activity through natural transformation with chromosomal DNA of C. coli and C. jejuni subsp. doylei strains possessing the ED pathway encoding plasticity region. These results reveal for the first time the ability of a Campylobacter species to catabolize glucose and provide new insights into how genetic macrodiversity through intra- and interspecies gene transfer expand the metabolic capacity of this food-borne pathogen.

Antimicrobial activity of gallic acid against thermophilic Campylobacter is strain specific and associated with a loss of calcium ions.

Gallic acid has been suggested as a potential antimicrobial for the control of Campylobacter but its effectiveness is poorly studied. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of gallic acid against Campylobacter jejuni (n = 8) and Campylobacter coli (n = 4) strains was determined. Gallic acid inhibited the growth of five C. jejuni strains and three C. coli strains (MIC: 15.63-250 µg mL(-1)). Gallic acid was only bactericidal to two C. coli strains (MBC: 125 and 62.5 µg mL(-1)). The mechanism of the bactericidal effect against these two strains (and selected non-susceptible controls) was investigated by determining decimal reduction times and by monitoring the loss of cellular content and calcium ions, and changes in cell morphology. Gallic acid did not result in a loss of cellular content or morphological changes in the susceptible strains as compared to the controls. Gallic acid resulted in a loss of calcium ions (0.58-1.53 µg mL(-1) and 0.54-1.17 µg mL(-1), respectively, over a 180 min period) from the susceptible strains but not the controls. Gallic acid is unlikely to be an effective antimicrobial against Campylobacter in a practical sense unless further interventions to ensure an effective bactericidal mode of action against all strains are developed.

Supplementation of Bolton broth with triclosan improves detection of Campylobacter jejuni and Campylobacter coli in chicken carcass rinse.

We compared Bolton enrichment broth supplemented with antimicrobial triclosan (T-Bolton broth) and normal Bolton broth for the isolation of Campylobacter jejuni (C. jejuni) and Campylobacter coli (C. coli) from chicken carcass rinse. Whole chickens were rinsed with buffered peptone water prior to enrichment in normal Bolton broth or T-Bolton broth, followed by inoculation onto modified charcoal-cefoperazone-deoxycholate agar (mCCDA). Suspect colonies were confirmed by PCR. We observed a significantly higher number of C. jejuni or C. coli-positive samples in the T-Bolton broth (71.3%) than in the normal Bolton broth (27.5%) (p<0.05). Furthermore, the number of contaminated mCCDA plates was lower after enrichment in T-Bolton broth (3.8%) than in the normal Bolton broth (75%) (p<0.05), indicating that T-Bolton broth has higher selectivity. Finally, we identified extended-spectrum ß-lactamase-producing Escherichia coli as the predominant competing flora in normal Bolton broth. In conclusion, the use of T-Bolton broth results in significant elimination of competing bacteria.

Marinade with thyme and orange oils reduces Salmonella Enteritidis and Campylobacter coli on inoculated broiler breast fillets and whole wings.

Essential oils have been reported to possess antimicrobial properties and therefore have potential usage as natural antimicrobials in food. In a previous study, thyme orange essential oil combination (TOC) used at the 0.5% level as a dip application on chicken cut-up parts had a significant antibacterial effect against Salmonella and Campylobacter. A study was designed to evaluate the effect of salt-phosphate marinade solution containing 0.5% TOC to 1) reduce Salmonella Enteritidis and Campylobacter coli numbers on broiler breast fillets and whole wings marinated by vacuum tumbling, and 2) reduce cross-contamination of both pathogens between inoculated and uninoculated parts during marination. A total of 52 skinless breast fillets and 52 whole wings were used for the 2 replications. For each replication, each cut-up part was randomly assigned to 1 of 5 groups: treatment 1: uninoculated parts marinated without TOC; treatment 2: inoculated parts marinated without TOC; treatment 3: uninoculated parts marinated with TOC; treatment 4: inoculated parts marinated with TOC; and control: nonmarinated inoculated parts. Samples were dipped in an inoculum containing a mixture of Salmonella Enteritidis and C. coli. The treatment samples were marinated by vacuum tumbling. All samples were immediately evaluated to determine Salmonella Enteritidis and C. coli numbers. Results indicated that TOC at the 0.5% level in the marinade solution applied by vacuum tumbling significantly reduced (P < 0.05) numbers of viable Salmonella Enteritidis by 2.6 and 2.3 log cfu/mL on broiler breast fillets and C. coli by 3.6 and 3.1 log cfu/mL on whole wings. Cross-contamination was observed as the uninoculated chicken parts marinated with inoculated parts were positive. However, the number of bacterial cells recovered from the TOC treated samples were significantly lower (P < 0.05) than the numbers recovered from the untreated samples. Marination with a salt phosphate formulation containing 0.5% TOC successfully reduced Salmonella and Campylobacter numbers on poultry products.

Inhibitory effect of high-dosage zinc oxide dietary supplementation on Campylobacter coli excretion in weaned piglets.

AIMS: This study investigated the impact of zinc oxide (ZnO) on Campylobacter coli by in vivo and in vitro assays. METHODS AND RESULTS: By in vitro growth inhibition assays, a high susceptibility of Camp. coli against ZnO could be observed. At concentrations ≥ 2.6 mmol l(-1) ZnO, a decline in cell numbers occurred. Quantitative real-time PCR assays demonstrated an up-regulation of the main oxidative stress gene (katA) in response to ZnO treatment. The expression level of katA was increased by fivefold after ZnO treatment. An experiment was carried out in pigs to elucidate the impact of ZnO as feed supplement on Camp. coli faecal excretion. Feeding a high-dosage ZnO concentration (3100 mg kg(-1) ) to piglets significantly reduced the faecal excretion of Camp. coli by up to 1 log CFU g(-1) as compared to animals receiving a low (40 mg kg(-1) ) or medium (100 mg kg(-1) ) ZnO diet. CONCLUSION: In vitro assays showed a high susceptibility of Camp. coli against ZnO. Adding high levels of ZnO to the diet of weaned piglets reduced Camp. coli excretion significantly. There is evidence for the induction of an oxidative stress response by ZnO supplementation in Camp. coli. SIGNIFICANCE AND IMPACT OF THE STUDY: Supplementation of a high-dosage ZnO diet to piglets can reduce the Camp. coli load, potentially leading to a lower contamination risk of meat during slaughter.

The effect of growth temperature on the pathogenicity of Campylobacter.

Control of Campylobacter in the food chain requires a better understanding of the behaviour of the bacteria in relevant environments. Campylobacter species are largely non-pathogenic in poultry, the body temperature of which is 42 °C. However, the bacteria are highly pathogenic in humans whose body temperature is 37 °C. The aim of this study was to examine if switching from commensal to pathogenic behaviour was related to temperature. We examined the growth, motility and invasion of T84 cells by three species of Campylobacter: C. jejuni 81116, C. jejuni M1, C. coli 1669, C. coli RM2228 and C. fetus fetus NC10842 grown at 37 and 42 °C. Our results suggest that C. jejuni isolates grow similarly at both temperatures but some are more motile at 42 °C and some are more invasive at 37 °C, which may account for its rapid spread in poultry flocks and for infection in humans, respectively. C. coli, which are infrequent causes of Campylobacter infections in humans, is less able to grow and move at 37 °C compared to 42 °C but was significantly more invasive at the lower temperature. C. fetus fetus, which is infrequently found in poultry, is less able to grow and invade at 42 °C.

Role of capsular polysaccharides and lipooligosaccharides in Campylobacter surface properties, autoagglutination, and attachment to abiotic surfaces.

The role of capsular polysaccharides and lipooligosaccharides in cell surface hydrophobicity, surface charge, autoagglutination (AAG), and attachment to abiotic surfaces of three strains of Campylobacter jejuni and one strain of C. coli were investigated. This was achieved by removal of capsular polysaccharides and truncation of lipooligosaccharides core oligosaccharides by inactivation of the kpsE and waaF genes, respectively. The mutants and the wild-type strains were compared after growth under planktonic (broth) and sessile (agar) conditions. Cells grown as planktonic cultures showed a significantly (p<0.05) higher degree of hydrophobicity and AAG activity but differed from their sessile counterparts with respect to surface charge and attachment counts, depending on the strain. These results suggest that prior mode of growth affects the surface properties and attachment of Campylobacter in a strain-dependent manner. There were no significant (p>0.05) differences between the three C. jejuni strains and their ΔkpsE and ΔwaaF mutants with respect to all traits tested. Inactivation of the kpsE gene significantly (p<0.05) reduced the surface charge of the C. coli strain from ∼-10 to ∼-6 mV and increased its AAG activity, while disruption of the waaF gene significantly (p<0.05) increased its surface hydrophobicity by >8° and decreased the numbers of cells attaching to stainless steel and glass by ∼0.5 log/cm². These results suggest that surface polysaccharides may influence the surface properties and attachment to abiotic surfaces of C. coli but not C. jejuni. This suggestion, however, requires further investigation using a larger number of strains of both species.

Importance of the producer on retail broiler meat product contamination with Campylobacter spp.

BACKGROUND: Campylobacter spp. are a leading cause of human bacterial gastroenteritis worldwide, with poultry meat being considered the most important source of the infection. To obtain data on broiler meat contamination with Campylobacter spp. in Lithuania, the occurrence, counts and genotypes of these pathogens on raw broiler meat products from different producers were examined. RESULTS: Out of 312 broiler meat product samples examined, 46.8% were contaminated with Campylobacter spp. Campylobacter jejuni was identified in 51.4% and Campylobacter coli in 37.7% of positive samples. Campylobacter jejuni was more frequently found in the warm period (April-October) and C. coli in the cold period (November-March) of the year (P < 0.05). The overall mean count of Campylobacter spp. was 3.55 and 3.50 log10 colony-forming units (CFU) on wings and drumsticks respectively. The occurrence and counts of Campylobacter spp. varied significantly between producers examined (P < 0.05). Analysis of flaA-RFLP genotyping revealed C. jejuni genotypes common to all producers as well as producer-specific genotypes. CONCLUSION: Both the occurrence and counts of Campylobacter spp. on broiler meat products were producer-dependent, so this should be kept in mind when risk-based control measures at national level are applied.

Experimental infection of weaned piglets with Campylobacter coli--excretion and translocation in a pig colonisation trial.

Campylobacter (C.) is one of the most common food-borne pathogen causing bacterial enteric infections in humans. Consumption of meat and meat products that have been contaminated with Campylobacter are the major source of infection. Pigs are a natural reservoir of Campylobacter spp. with C. coli as the dominant species. Even though some studies focussed on transmission of C. coli in pig herds and the excretion in faeces, little is known about the colonisation and excretion dynamics of C. coli in a complex gut microbiota present in weaned piglets and the translocation to different tissues. Therefore, an experimental trial was conducted to evaluate the colonisation and translocation ability of the porcine strain C. coli 5981 in weaned pigs. Thus, ten 35 days old piglets were intragastrically inoculated with strain C. coli 5981 (7 × 10(7)CFU/animal) encoding resistances against erythromycin and neomycin. Faecal samples were taken and C. coli levels were enumerated over 28 days. All piglets were naturally colonised with C. coli before experimental inoculation, and excretion levels ranged from 10(4) to 10(7)CFU/g faeces. However, no strain showed resistances against the additional antimicrobials used. Excretion of C. coli 5981 was seen for all piglets seven days after inoculation and highest counts were detectable ten days after inoculation with 10(6)CFU/g faeces. Post-mortem, translocation and subsequent invasion of luminal C. coli was observed for gut tissues of the small intestine and for the gut associated lymphatic tissues, such as jejunal mesenteric lymph nodes and tonsils as well as for spleen and gall bladder. In conclusion, this pig colonisation trial offers the opportunity to study C. coli colonisation in weaned piglets using the porcine strain C. coli 5981 without the need for gnotobiotic or specific pathogen-free animals.

Sensitive and rapid detection of Campylobacter jejuni and Campylobacter coli using loop-mediated isothermal amplification.

Loop-mediated isothermal amplification (LAMP) is an established nucleic acid amplification method offering rapid, accurate, and cost-effective diagnosis of infectious diseases. From the beginning of DNA extraction to final detection of Campylobacter jejuni and Campylobacter coli, the assay requires less than 50 and 90 min from a colony on selective media, and human feces, respectively. For chicken meat samples, the assay requires approximately 24-48 h from the beginning of the enrichment culture to final detection. The sensitivity of the LAMP assay is tenfold higher than that of the equivalent PCR assay. LAMP amplification can be judged by both turbidimeter analysis and visual assessment with the unaided eye. The LAMP assay is a powerful tool for rapid, simple, and sensitive detection of C. jejuni and C. coli, which may facilitate the investigation of C. jejuni and C. coli contamination in chicken, as well as the early diagnosis of C. jejuni and C. coli infection in humans.

Effects of lactic acid and commercial chilling processes on survival of Salmonella, Yersinia enterocolitica, and Campylobacter coli in pork variety meats.

Current industry chilling practices with and without the application of 2% L-lactic acid were compared for their effectiveness at reducing levels of Salmonella, Yersinia enterocolitica, and Campylobacter coli on pork variety meats. Pork variety meats (livers, intestines, hearts, and stomachs) were inoculated individually with one of the three pathogens and subjected to five different treatment combinations that included one or more of the following: water wash (25°C), lactic acid spray (2%, 40 to 50°C), chilling (4°C), and freezing (-15°C). Samples were analyzed before treatment, after each treatment step, and after 2, 4, and 6 months of frozen storage. Results showed that when a lactic acid spray was used in combination with water spray, immediate reductions were approximately 0.5 log CFU per sample of Salmonella, 0.8 log CFU per sample of Y. enterocolitica, and 1.1 log CFU per sample of C. coli. Chilling, both alone and in combination with spray treatments, had little effect on pathogens, while freezing resulted in additional 0.5-log CFU per sample reductions in levels of Salmonella and Y. enterocolitica, and an additional 1.0-log CFU per sample reduction in levels of C. coli. While reductions of at least 1 log CFU per sample were observed on variety meats treated with only a water wash and subsequently frozen, samples treated with lactic acid had greater additional reductions than those treated with only a water spray throughout frozen storage. The results of this study suggest that the use of lactic acid as a decontamination intervention, when used in combination with good manufacturing practices during processing, causes significant reductions in levels of Salmonella, Y. enterocolitica, and C. coli on pork variety meats.

An enhanced technique combining pre-enrichment and passive filtration increases the isolation efficiency of Campylobacter jejuni and Campylobacter coli from water and animal fecal samples.

Improved isolation techniques from environmental water and animal samples are vital to understanding Campylobacter epidemiology. In this study, the efficiency of selective enrichment in Bolton Broth (BB) followed by plating on charcoal cefoperazone deoxycholate agar (CCDA) (conventional method) was compared with an approach combining BB enrichment and passive filtration (membrane method) adapted from a method previously developed for testing of broiler meat, in the isolation of thermophilic campylobacters from surface water and animal fecal samples. The conventional method led to recoveries of Campylobacter from 36.7% of the water samples and 78.0% of the fecal samples and similar numbers, 38.3% and 76.0%, respectively, were obtained with the membrane method. To investigate the genetic diversity of Campylobacter jejuni and Campylobacter coli obtained by these two methods, isolates were analyzed using Comparative Genomic Fingerprinting, a high-resolution subtyping technique. The conventional and membrane methods yielded similar numbers of Campylobacter subtypes from water (25 and 28, respectively) and fecal (15 and 17, respectively) samples. Although there was no significant difference in recovery rates between the conventional and membrane methods, a significant improvement in isolation efficiency was obtained by using the membrane method, with a false-positive rate of 1.6% compared with 30.7% obtained using the conventional method. In conclusion, although the two methods are comparable in sensitivity, the membrane method had higher specificity, making it a cost-effective procedure for the enhanced isolation of C. jejuni and C. coli from water and animal fecal samples.