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.

Prevalence of and risk factors for Campylobacter colonisation in broiler flocks at the end of the rearing period in France.

1. A study was conducted to estimate the prevalence and quantification by species of Campylobacter infection in broiler flocks at the end of the rearing period and to identify associated risk factors. 2. A questionnaire about the rearing period was completed and caecal samples were collected from 121 broiler flocks in Brittany, France, during 2008. 3. Campylobacter was isolated in 87 out of 121 flocks--a prevalence of 71.9% (95% CI, 63.7-80.1%), including 40.5% of Campylobacter jejuni and 29.8% of Campylobacter coli. 4. The average concentration, in positive flocks, was 7.96 log10 cfu/g and ranged from 3.15 to 10.32 log10 cfu/g. 5. The average concentration by species was: 7.57 log10 cfu/g for C. jejuni and 8.44 log10 cfu/g for C. coli. 6. There was a seasonal effect, with increased risk of Campylobacter colonisation in June, July and August (odds ratio (OR) = 9.59, 95% CI 1.15-79.75). 7. The other factors, associated with lower risk of Campylobacter colonisation, were the acidification of drinking water (OR = 0.33, 95% CI 0.13-0.86), antibiotic treatment at the beginning of the rearing period (OR = 0.20, 95% CI 0.07-0.55) and rodent control around the house (OR = 0.18, 95% CI 0.03-0.95). 8. The results show that hygiene practices and biosecurity measures could lead to a reduction in Campylobacter colonisation.

Description and sources of contamination by Campylobacter spp. of river water destined for human consumption in Brittany, France.

Presence or absence of Campylobacter spp. in water of five rivers upstream from an intake point for drinking water production was investigated, and isolates genetically compared with human, pig and poultry isolates in order to determine their source. River water and drinking water obtained from these rivers were sampled one time per month, over a period of one year, and tested for Campylobacter. Isolates were typed by PFGE. Campylobacter was not detected in treated drinking water, but 50% of the river samples were contaminated. Contamination was observed on the four seasons. In total, 297 Campylobacter isolates were collected and generated 46 PFGE profiles. Campylobacter jejuni was the most frequently detected species in samples (74.1% of the isolates), followed by Campylobacter coli (17.8%) and Campylobacter lari (8.1%). Forty-two of the 46 PFGE profiles were unique. Only one genotype was detected three times in a river during the year and four genotypes in two different rivers. When compared to animal and human Campylobacter PFGE profiles, 14, 11 and one Campylobacter genotypes from water were genetically closed to human, poultry, and pig Campylobacter genotypes, respectively. The Campylobacter population displayed a high level of genetic diversity, suggesting that contamination originated from various origins. Human, poultry and pig were sources of contamination of the river by Campylobacter. Finally, no Campylobacter were detected in drinking water, indicating that the risk of outbreaks due to consumption of drinking water is low.

Comparison of genetic profiles of Campylobacter strains isolated from poultry, pig and Campylobacter human infections in Brittany, France.

Five hundred eighty-two Campylobacter isolates (177 from humans, 319 from poultry and 86 from pig) collected in Brittany, France, in 2003 and 2004 were typed by pulsed-field gel electrophoresis. The number of human cases increased during the hot season, particularly for C. jejuni. Twelve genetic groups out of 27 contained human isolates collected over the two years. These groups had 21.3 and 17.0% of the isolates obtained in 2003 and 2004, respectively. In four cases, isolates from 2003 have the same Pulsed-field gel electrophoresis (PFGE) profile as isolates from 2004. Six PFGE profiles common to poultry and human isolates were identified. Poultry isolates were found in 47 clusters containing human isolates. Caeca from farms and slaughterhouses accounted for 66% of these isolates, with chicken legs obtained from supermarkets accounting for the other 34%. Pig isolates never clustered with poultry and human isolates. In conclusion, the analysis of the genetic profiles of Campylobacter resulting from human cases showed that there were few identical or genetically close isolates between the human cases declared in 2003 and those declared in 2004. This highlighted a great genetic diversity in the isolates and indicated that it should be difficult to bind the human infections with groups of Campylobacter isolates presenting particular genetic profiles. The Campylobacter isolates obtained from the two animal production systems had different genotypes, and isolates from pigs differed genetically from isolates obtained from humans. We found that 44.6% of human Campylobacter isolates were genetically related to genotypes found in poultry and a part of these campylobacteriosis are due to contact with poultry. This is not particularly surprising in Brittany, a farming area with many animal-rearing farms and slaughterhouses. This work highlights the implication of the poultry in the French human cases and that handling of poultry is also an important risk for Campylobacter infection in humans.

Influence of bird strain on competitive exclusion of Campylobacter jejuni in young chicks.

1. Newly hatched chicks of either layer or broiler strain were treated orally at regular intervals with either homologous or heterologous gut-flora preparations from young donor birds, in an attempt to prevent subsequent colonisation with Campylobacter jejuni by 'competitive exclusion' (CE). 2. Donors of 3 to 10 d of age were chosen to correspond with the period in which intensively reared poultry are least likely to become colonised with Campylobacter. 3. In two separate trials, material from donor layer hens (ISA Brown) protected male chicks of the same strain against a low (195 to 360 cfu/bird) Campylobacter challenge, but the same kind of material was ineffective when administered to chicks of a broiler strain (JA957). 4. Two further trials involved treatment preparations from young broilers, which failed to prevent Campylobacter colonisation of broiler chicks, even when colonisation occurred relatively slowly from a challenge of 90 to 94 cfu/bird. 5. It was concluded that any CE effect observed was strongly dependent on bird strain.

Campylobacter contamination in French chicken production from farm to consumers. Use of a PCR assay for detection and identification of Campylobacter jejuni and Camp. coli.

AIMS: Campylobacter contamination in French chicken production from the farm to the consumer was determined using a PCR assay for bacteria detection and identification. METHODS AND RESULTS: Samples were bird droppings from poultry houses, neck skins, livers, hearts, gizzards, wings, legs and escalopes from slaughterhouses and gizzards, legs, drumstick, breast and escalopes from a supermarket. Bacterial DNA extraction was performed after an enrichment step in a broth and was followed by PCR. An internal control (IC) was used for both DNA extraction and PCR. Campylobacter were detected in 79.2% of poultry houses. Of the 303 samples, 201 were Campylobacter-positive (i.e. 66.3%) including 43.2% faecal samples, 5.6% slaughterhouse samples and 17.5% supermarket samples. There was no significant difference between the molecular method and the conventional culture technique for Campylobacter detection whatever the samples. The sensitivity was 5 UFC g(-1) of samples and 1.5 x 10(3) UFC ml(-1) of enrichment broth. The use of IC revealed PCR inhibition in 13 samples and problems in the DNA extraction in five samples. CONCLUSION: Significant Campylobacter contamination affects all stages of French chicken production. SIGNIFICANCE AND IMPACT OF THE STUDY: The understanding of Campylobacter contamination at different levels of chicken production and the determination of the best place(s) for intervention are important for significantly decreasing Campylobacteriosis. Our technique is rapid and can be used on different chicken samples for Campylobacter detection and identification.

Effects of AvGard treatment on the microbiological flora of poultry carcases.

1. The efficiency of the AvGard (or Assur-Rince in the USA) trisodium phosphate poultry carcase decontamination process was evaluated during both manual and industrial trials against total aerobic mesophilic count (TAMC), thermotolerant coliforms, Pseudomonas, Enterobacteriaceae, Campylobacter, Listeria monocytogenes and Salmonella. 2. The TSP treatment proved to have significant effects on the bacterial decontamination of poultry neck skin, lowering the contamination by a factor of about 10 for TAMC and of 100 for Coliform and Pseudomonas. 3. Numeration of Salmonella with an innovative miniaturised most probable number method has proved that the effect upon these micro-organisms was also close to 2 log10 reduction. 4. The effect of TSP treatment on the ecological balance of psychrotrophic bacterial flora was also investigated to study the origin of the shelf-life flora of treated carcases (Pseudomonas being reduced to the limit of detection) and to ascertain whether L. monocytogenes might gain a competitive advantage. In fact AvGard reduced the number of L. monocytogenes on poultry carcases. 5. As a consequence of the virtual elimination of the Pseudomonas usually present, Brochothrix thermosphacta became the main species responsible for putrefaction. 6. Because the growth rate of Brochothrix thermosphacta was greater than that of L. monocytogenes at refrigeration temperature, it was considered that putrefaction would occur before the emergence of large numbers of L. monocytogenes.

Culture media for the isolation of campylobacters.

The history of the development of selective media for isolation of campylobacters, including the rationale for choice of selective agents is described. Developments have included modifications to allow incubation at 37 degrees C instead of 42 or 43 degrees C and changes in the types and concentrations of antibiotics in order not to inhibit organisms such as Campylobacter upsaliensis, C. jejuni subsp. doylei and some strains of C. coli and C. lari. When examining foods, plating media originally developed for isolation from faeces are normally used, sometimes after liquid enrichment. Most of the media include ingredients intended to protect campylobacters from the toxic effect of oxygen derivatives. Most commonly used are lysed or defibrinated blood; charcoal; a combination of ferrous sulphate, sodium metabisulphite and sodium pyruvate (FBP); and haemin or haematin. To date no medium includes an indicator system--for instance a pH indicator to show whether colonies produce acid or alkali from particular substrates. The manner in which liquid enrichment media are used has been modified for food samples to avoid inhibitory effects on sublethally damaged cells by toxic components in the formula. This is done by a preliminary period of incubation at reduced temperature and sometimes by delayed addition of antibiotics. Expensive and time-consuming methods have been proposed to achieve a microaerobic atmosphere while using liquid enrichment media. To date there is no generally accepted 'standard' method of isolating campylobacters from food.