Genetic diversity of Arcobacter and Campylobacter on broiler carcasses during processing

Broiler carcasses (n=325) were sampled at three sites along the processing line (prescalding, prechilling, and postchilling) in a commercial poultry processing plant during five plant visits from August to October 2004. Pulsed-field gel electrophoresis (PFGE) was used to determine the genomic fingerprints of Camospylobacter coli (n=27), Campylobacter jejuni (n=188), Arcobacter butzleri (n=138), Arcobacter cryaerophilus 1A (n=4), and A. cryaerophilus 1B (n=31) with the restriction enzymes SmaI and KpnI for Campylobacter and Arcobacter, respectively. Campylobacter species were subtyped by the Centers for Disease Control and Prevention PulseNet 24-h standardized protocol for C. jejuni. A modification of this protocol with a different restriction endonuclease (KpnI) and different electrophoresis running conditions produced the best separation of restriction fragment patterns for Arcobacter species. Both unique and common PFGE types of Arcobacter and Campylobacter strains were identified. A total of 32.8% (57 of 174) of the Arcobacter isolates had unique PFGE profiles, whereas only 2.3% (5 of 215) of the Campylobacter isolates belonged to this category. The remaining Arcobacter strains were distributed among 25 common PFGE types; only eight common Campylobacter PFGE types were observed. Cluster analysis showed no associations among the common PFGE types for either genus. Each of the eight common Campylobacter types consisted entirely of isolates from one sampling day, whereas more than half of the common Arcobacter types contained isolates from different sampling days. Our results demonstrate far greater genetic diversity for Arcobacter than for Campylobacter and suggest that the Campylobacter types are specific to individual flocks of birds processed on each sampling day.     

潜在食源性致病菌弓形菌在食品中的分布及检测研究进展

弓形菌（Arcobacter spp.）是一类重要的食源性致病菌,因其与弯曲菌（Campylobacter spp.）形态相似且亲缘关系很近,最初被称为“氧气耐受的弯曲菌”。弓形菌属内能够引起人致病的主要是布氏弓形菌、嗜低温弓形菌和斯氏弓形菌,感染致病性的弓形菌可以引起人类的肠炎、严重腹泻、败血症和菌血症等。弓形菌广泛存在于畜禽、畜禽产品和环境中,畜禽产品生产设备和水源的污染是造成其高感染率的主要途径。由于弓形菌的生理特征与弯曲菌相近,分离培养时通常较难与弯曲菌区分,分子生物学方法是弓形菌实验室诊断的主要手段。近年来,作为新型潜在的食源性致病菌,弓形菌逐渐引起研究者的关注。然而,相较于其他食源性致病菌,对弓形菌的研究尚处于起步阶段,国内的报道也相对较少。本文对弓形菌在食品中的污染现状、流行病学特点及弓形菌的实验室检测等相关研究进展进行综述,以期为广大研究者提供有价值的信息。     

Detection of seven virulence and toxin genes of Campylobacter jejuni isolates from Danish turkeys by PCR and cytolethal distending toxin production of the isolates

A total of 117 Campylobacter jejuni isolates from Danish turkeys were tested for the presence of seven virulence and toxin genes by PCR. One hundred seventeen (100%) isolates were positive for flaA, cadF, and ceuE gene primers. One hundred three (88%) isolates were positive for cdt gene cluster PCR detection (cdt gene cluster-PCR), whereas 101 (86.3%), 102 (87.2%), and 110 (94%) isolates were positive for cdtA-, cdtB-, and cdtC-PCR, respectively. Only 39 (33.3%) isolates were positive for virB11. Of 117 isolates, 114 (97.4%) produced cytolethal distending toxin (CDT) in Vero cell assays, 105 (89.7%) in Colon 205 assays, and 109 (93.2%) in chicken embryo cell assays. The CDT titers were determined in Vero cell assays. Of 117 isolates, 50 (42.7%) produced a CDT titer of 1:100, 29 (24.8%) of 1:50, and 27 (23%) of 1:5 to 1:10; 8 (6.8%) produced a CDT titer at undiluted supernatants and 3 (2.6%) produced no toxin. Twenty-nine C. jejuni isolates that were PCR negative for one or more individual cdt toxin genes also produced low or no CDT toxin. The high prevalence of the seven virulence and toxin genes demonstrates that these putative pathogenic determinants are widespread among Campylobacter isolates from turkeys and calls for further investigation for the elimination of Campylobacter infection in industrial turkey production and in industrial food chains.     

Prevalence of Arcobacter and Campylobacter on broiler carcasses during processing

Broiler carcasses (n=325) were sampled in a U.S. commercial poultry processing plant for the prevalence of Arcobacter and Campylobacter at three sites along the processing line: pre-scald, pre-chill and post-chill. Samples (75-125 broilers per site) were collected during five plant visits from August to October of 2004. Arcobacter was recovered from pre-scald carcasses more frequently (96.8%) than from pre-chill (61.3%) and post-chill carcasses (9.6%). Campylobacter was isolated from 92% of pre-scald carcasses, 100% of pre-chill carcasses, and 52% of post-chill carcasses. In total, Arcobacter was isolated from 55.1% (179 of 325), while Campylobacter was isolated from 78.5% (255 of 325) of the carcasses from the three collection sites. For Arcobacter identification, a species-specific multiplex PCR showed that A. butzleri was the most prevalent species (79.1%) followed by A. cryaerophilus 1B (18.6%). A. cryaerophilus 1A was found at low levels (2.3%). PCR identified the most common Campylobacter species as C. jejuni (87.6%) followed by C. coli (12.4%). Overall, significant contamination of broiler carcasses by Arcobacter was observed, although less than that found for Campylobacter. From pre-scald to post-chill, a far greater reduction in Arcobacter numbers was observed than for Campylobacter. Our results for Arcobacter, obtained from the same environment as the closely related pathogen Campylobacter, will aid in the development of control measures for this emerging pathogen.     

EVALUATION OF ATTACHMENT AND PENETRATION ABILITIES OF CAMPYLOBACTER JEJUNI ISOLATES OBTAINED FROM HUMANS AND CHICKEN CARCASSES DURING PROCESSING AND AT RETAIL

The differences in attachment and penetration ability of Campylobacter jejuni were determined by analyzing C. jejuni isolates obtained from chicken carcasses and from humans exhibiting symptoms of campylobacteriosis. INT 407 cells, a human cell line originating from the jejunal/ileal region, were used as the in vitro model, and attachment and penetration abilities were evaluated for each isolate. There were no significant differences between the attachment and penetration abilities of chicken isolates and human isolates (HUMN). In addition, a wide range of attachment and penetration abilities was found for the isolates, with many of the HUMN possessing low attachment and penetration abilities. These data indicate that C. jejuni attachment and penetration into the human ileal and jejunal regions may not be primary virulence factors and may only be important in causing more acute symptoms.     

Presence of Campylobacter and Arcobacter species in in-line milk filters of farms authorized to produce and sell raw milk and of a water buffalo dairy farm in Italy

The objectives of this study were to investigate the presence of Campylobacter spp. and Arcobacter spp. in dairy herds authorized for the production and sale of raw milk and in a water buffalo dairy farm, and to test the antimicrobial susceptibility of the isolates. A total of 196 in-line milk filters were collected from 14 dairy farms (13 bovine and 1 water buffalo) for detection of Campylobacter spp. and Arcobacter spp. by microbiological culture. For each farm investigated, 1 isolate for each Campylobacter and Arcobacter species isolated was tested using the Etest method (AB Biodisk, Solna, Sweden) to evaluate the susceptibility to ciprofloxacin, tetracycline, chloramphenicol, ampicillin, erythromycin, and gentamicin. A total of 52 isolates were detected in 49 milk filters in 12 farms (85.7%) out of 14 and the isolates were identified as Campylobacter jejuni (6), Campylobacter hyointestinalis ssp. hyointestinalis (8), Campylobacter concisus (1), Campylobacter fetus ssp. fetus (1), Arcobacter butzleri (22), and Arcobacter cryaerophilus (14). The small number of isolates tested for antimicrobial susceptibility precludes any epidemiological consideration but highlights that all Campylobacter isolates were susceptible to macrolides, which are the first-choice drugs for the treatment of campylobacteriosis, and that resistance to fluoroquinolones and tetracycline was detected; for Arcobacter isolates, resistance to ampicillin and chloramphenicol was detected. The sale of raw milk for human consumption by self-service automatic vending machines has been allowed in Italy since 2004 and the presence of C. jejuni in in-line milk filters confirms that raw milk consumption is a significant risk factor for human infection. The high occurrence of emerging Campylobacter spp. and Arcobacter spp. discovered in dairy farms authorized for production and sale of raw milk represents an emerging hazard for human health.     

Molecular characterization of Arcobacter isolates collected in a poultry slaughterhouse

In a poultry slaughterhouse, Arcobacter contamination was examined over a period of 1 week to establish possible routes of contamination. Samples were collected from the slaughter equipment and from processing water before the onset of slaughter and from the first broiler flock slaughtered on each sampling day. Characterization of 1,079 isolates by enterobacterial repetitive intergenic consensus-polymerase chain reaction and a random amplified polymorphic DNA assay resulted in the delineation of 159 Arcobacter butzleri and 139 Arcobacter cryaerophilus types. From almost all 140 neck skin samples collected before and after evisceration, A. butzleri and A. cryaerophilus were isolated simultaneously at contamination levels ranging from 10(1) to 10(4) CFU/g. Only six A. butzleri types present in the slaughterhouse environment were also present on the broiler carcasses. None of the A. cryaerophilus genotypes were detected in both the neck skin and the environmental samples. All A. butzleri types isolated from the feather samples were also isolated from broiler neck skin samples. The slaughter equipment was contaminated with arcobacters before the onset of slaughter, but it appeared unlikely that contamination through the slaughter equipment alone explained the high contamination levels on poultry products. Arcobacters were also present in processing water, but types present in water and poultry products were different. Characterization of the Arcobacter isolates did not clarify the routes of transmission, probably because of the extreme heterogeneity among Arcobacter isolates. However, the results obtained in this study brought to light insufficient decontamination at the processing plant involved in the study and confirmed the survival capacity of certain A. butzleri strains.     

Use of MIDI-fatty acid methyl ester analysis to monitor the transmission of Campylobacter during commercial poultry processing

The presence of Campylobacter spp. on broiler carcasses and in scald water taken from a commercial poultry processing facility was monitored on a monthly basis from January through June. Campylobacter agar, Blaser, was used to enumerate Campylobacter in water samples from a multiple-tank scalder; on prescalded, picked, eviscerated, and chilled carcasses; and on processed carcasses stored at 4 degrees C for 7 or 14 days. The MIDI Sherlock microbial identification system was used to identify Campylobacter-like isolates based on the fatty acid methyl ester profile of the bacteria. The dendrogram program of the Sherlock microbial identification system was used to compare the fatty acid methyl ester profiles of the bacteria and determine the degree of relatedness between the isolates. Findings indicated that no Campylobacter were recovered from carcasses or scald tank water samples collected in January or February, but the pathogen was recovered from samples collected in March, April, May, and June. Processing generally produced a significant (P < 0.05) decrease in the number of Campylobacter recovered from broiler carcasses, and the number of Campylobacter recovered from refrigerated carcasses generally decreased during storage. Significantly (P < 0.05) fewer Campylobacter were recovered from the final tank of the multiple-tank scald system than from the first tank. MIDI similarity index values ranged from 0.104 to 0.928 based on MIDI-fatty acid methyl ester analysis of Campylobacterjejuni and Campylobacter coli isolates. Dendrograms of the fatty acid methyl ester profile of the isolates indicated that poultry flocks may introduce several strains of C. jejuni and C. coli into processing plants. Different populations of the pathogen may be carried into the processing plant by successive broiler flocks, and the same Campylobacter strain may be recovered from different poultry processing operations. However, Campylobacter apparently is unable to colonize equipment in the processing facility and contaminate broilers from flocks processed at later dates in the facility.     

The introduction of Arcobacter spp. in poultry slaughterhouses

Despite the presence high levels of Arcobacter spp. on chicken carcasses, the source of arcobacter contamination in slaughterhouses still remains unclear. It has been hypothesised in the literature that Arcobacter species that contaminate carcasses originate in in-plant slaughterhouses and/or supply water. The present study aimed to determine the source of Arcobacter contamination in two poultry slaughterhouses in The Netherlands. Carcasses and intestinal tracts from 3 hen flocks and 2 broiler flocks were collected. Water draining off carcasses during processing in 2 slaughterhouses and supply water in one slaughterhouse were also taken. For one flock, cloacal swabs and faecal samples were taken on the farm before slaughtering. ERIC-PCR was applied to study the genetic diversity and relationship among the isolates. No Arcobacter spp. were found in the supply water but on almost all of the sampled carcasses and in carcass-draining-off water arcobacters were identified. Arcobacter spp. were detected in the gut systems of chickens, ranging from 20% to 85% in hens and 3.3% and 51% in broilers. Similar ERIC-PCR genotypes were detected in gut contents as well as on carcasses from the same flock. The present study demonstrated that Arcobacter spp. can be detected in chicken intestines at slaughter and could be brought in this way into slaughterhouses where the bacteria contaminate carcasses during processing.     

Prevalence and diversity of Arcobacter spp. in the Czech Republic

The aim of this study was to examine 634 samples of chicken, lamb, pork, beef, fish, samples from the intensive animal industry and from poultry for slaughter, as well as from the domestic breeding of poultry, horses, pigs, and lambs, from surface water, and from clinical samples for the presence of Arcobacter. All the samples were examined with a cultivation method, followed by confirmation by multiplex PCR. The method of multiplex PCR applied directly to a liquid medium after enrichment was applied only to the samples with the highest probability of the presence of arcobacters. Arcobacter spp. were detected in 11.8% of the samples, of which A. butzleri, A. cryaerophilus, and A. skirrowii were found in 6.6, 5.1, and 0.2% of the samples, respectively. The sources of the arcobacters were chicken meat from the retail market, intensive animal production facilities, domestic chicken breeding facilities, lamb raising environments, surface water and wastewater, and beef swabs taken in a meat processing factory. No occurrence of arcobacters was identified in the swabs from slaughter turkeys, ducks, and wild poultry. No arcobacters were found in horse and pig breeding environments, on pork, or on the swabs of fish. Forty-two rectal swabs taken from humans were also free of Arcobacter. Seventeen isolates of Arcobacter were further identified by sequencing the 16S rRNA gene. Varied genotypes were observed among A. butzleri from chicken meat and chicken breeds, and A. cryaerophilus from wastewater and chicken breeds. They were similar to the genotypes present in wastewater, porcine feces, human stool, and human blood obtained from databases. Our results revealed that the chicken meat from the retail market is an important source of arcobacters. Cross-contamination during handling of chicken carcass practices could play a key role in the spread of Arcobacter.     

Development of a new protocol for the isolation and quantification of Arcobacter species from poultry products.

None of the presently available selective supplements for the specific isolation of Arcobacter species allows the growth of Arcobacter butzleri, A. cryaerophilus and A. skirrowii and at the same time fully suppresses the accompanying flora present in poultry and poultry products. Furthermore, little is known about the contamination levels of poultry with Arcobacter species. In this study, a new selective supplement comprising amphotericin B (10 mg/l), cefoperazone (16 mg/l), 5-fluorouracil (100 mg/l), novobiocin (32 mg/l) and trimethoprim (64 mg/l) was developed. With a new isolation procedure, including enrichment in Arcobacter broth with the selective supplement, incubated for 24 to 48 h at 28 degrees C under microaerobic conditions, arcobacters were isolated from 100% (n = 34) of neck skin of laying hens and from 90% (n = 71) of similar samples from broilers. Of the broiler breast meat samples examined (n = 52), 65% were found to be contaminated with these bacteria. In 64% of the samples, A. butzleri was the only Arcobacter species isolated. In 9% of the samples, A. cryaerophilus was the only species present, while 11% of the samples were positive for both species simultaneously. Using direct isolation on the selective agar medium developed in this study, incubated for 24 to 48 h under microaerobic conditions at 28 degrees C. 32 out of 45 broiler carcasses and 6 out of 25 broiler breast meat samples carried a bacterial load of arcobacters of 10(2) to 10(3) cfu/g. The prevalence of Arcobacter in Belgian poultry was found higher than the prevalence of thermophilic Campylobacter species in each of the poultry categories examined. The enrichment procedure and the direct plating method were validated for the isolation of A. skirrowii. For this species, growth performance was less than the other two Arcobacter species and it was not isolated nor detected by m-PCR from the naturally contaminated poultry samples examined. This new protocol provides a fast and reliable method for the isolation of Arcobacter species from poultry and can contribute to more comprehensive epidemiological investigations.     

Fecal water from ileostomic patients consuming oat beta-glucan enhances immune responses in enterocytes

Yeast, fungal, and dietary beta-glucans have immune-modulating effects in vitro and in vivo, as thought, mainly by affecting leukocytes; however, effects of oat beta-glucan on enterocytes have never been studied. As recognized, supplying oat beta-glucans as such to cells in culture directly is difficult because of solubility problems. Therefore, six ileostomic patients consumed, in random order, a control diet or an oat beta-glucan enriched diet (5 g) and from the collected ileostomic content, fecal water was prepared and added to two small intestinal cell lines (INT407, Caco-2) and two colon cell lines (HT29, T84) together with a cytokine cocktail (IL-1beta + INFgamma + TNFalpha). Several parameters reflecting immune-modulation were measured. As compared to placebo fecal water, beta-glucan enriched fecal water significantly increased IL-8 production in HT29 (5.0%; p = 0.046) and INT407 cells (22.0%; p = 0.028). Intercellular adhesion molecule (ICAM)-1 expression increased in T84 (11.0%; p = 0.028) and Caco-2 cells (20.4%; p = 0.075). These immune-stimulating effects were confirmed by enhancement of inflammatory expression profiles, as determined with an antibody array. Our findings show immune enhancement by fecal water from ileostomic patients consuming oat beta-glucan both in small intestinal and colon cell lines after stimulation, which is in agreement with documented effects in leukocytes. Whether these immune-stimulating effects on enterocytes contribute to the enhanced protection of the host against invading pathogens as observed both in animals and in humans, as well as the underlying mechanism, needs further evaluation.     

Evaluation of a tissue culture-based approach for differentiating between virulent and avirulent Vibrio parahaemolyticus strains based on cytotoxicity

The ability of only a subset of Vibrio parahaemolyticus strains to cause human infection underscores the need for an analytical method that can effectively differentiate between pathogenic strains and those that do not cause disease. We tested the feasibility of a tissue culture-based assay to determine whether clinical isolates could be differentiated from nonclinical isolates based on relative isolate cytopathogenicity. To screen for cytotoxic capability, we measured relative extracellular lactate dehydrogenase as an indicator of host cell damage in five different mammalian cell lines in the presence of V. parahaemolyticus. Isolates originating from clinical sources exhibited 15.5 to 59.3% relative cytotoxicity, whereas those originating from food sources exhibited 4.4 to 54.9% relative cytotoxicity. In the presence of -1.2 x 10(6) cells, cytotoxicity was 1.6- to 3.5-fold higher (P < 0.05) for clinical isolates than for nonclinical isolates in L2, Henle 407, and Caco-2 cell lines. V. parahaemolyticus serotype O3:K6 clinical isolates had 1.6- to 2.1-fold higher cytotoxicity than did the non-O3:K6 clinical isolates, with significantly higher cytotoxicity in HeLa, J774A.1, and Henle 407 cells than in L2 and Caco-2 cells. Because V. parahaemolyticus often is found in oysters, the effect of the presence of an oyster matrix on assay efficacy was also tested with L2 cells. The cytotoxicity elicited by a highly cytotoxic V. parahaemolyticus isolate was not affected by the presence of oyster tissue, suggesting that an oyster matrix will not interfere with assay sensitivity. In the present format, this assay can detect the presence of > 10(5) cells of a virulent V. parahaemolyticus strain in an oyster matrix.     

Isolation and characterisation of Arcobacter butzleri from meat

A survey was conducted to determine the prevalence of Arcobacter in ground chicken, pork, beef and lamb meats. Meat samples were enriched in Arcobacter broth (AB) containing cefoperazone, amphotericin and teicoplanin (CAT) supplement. Samples were screened for the presence of Arcobacter spp. using a multiplex polymerase chain reaction (PCR) followed by isolation on blood and selective agar. Arcobacter butzleri was the only species of Arcobacter isolated from 35% of 88 samples of ground meats. A. butzleri was more frequently isolated from poultry (73%) than pork (29%), beef (22%) or lamb (15%) samples. No significant differences were found in the isolation rates and from the different regions sampled. Isolates were characterised by pulsed-field gel electrophoresis (PFGE) using SacII, EagI and SmaI restriction endonucleases. A number of isolates with indistinguishable PFGE fingerprints were found to be epidemiologically related, which may indicate cross-contamination of common types of Arcobacter from different meat species or between meat species. The public health significance of Arcobacter in ground meat needs to be determined.     

Detection and diversity of various Arcobacter species in Danish poultry

The prevalence and diversity of different Arcobacter spp. in various poultry species in Denmark were investigated using cultural and multiplex PCR methods. A pool of three fresh droppings obtained at the production site from 70 broiler chicken flocks aged 4-5 weeks was examined. In addition, pools of 10 cloacal swabs taken at the abattoir prior to stunning from each of 15, and 37 duck and turkey flocks, respectively, were analyzed. Thirty fresh broiler chicken carcasses and 29 cloacal swabs from the respective viscera were also examined at the abattoir. Finally, 10 caecal and 10 cloacal swabs from ducks at the abattoir were analyzed individually. In total, 85 Arcobacter isolates were obtained. Of these 45, 20 and 7 were identified as Arcobacter butzleri, Arcobacter cryaerophilus and Arcobacter skirrowii, respectively, using a multiplex PCR. Interestingly, some chicken isolates of A. butzleri showed urease activity, and 6 out of seven A. skirrowi isolates were unable to hydrolyse indoxyl acetate. All chicken carcasses examined were found positive for A. butzleri and/or A. cryaerophilus, whereas 21 (72%) of the 29 chicken cloacal swabs were positive for either A. butzleri (13) or A. cryaerophilus (9). Three (4.3%) out of 70 chicken flocks analyzed were positive only for A. cryaerophilus. Of the ten ducks examined individually, 7 carried A. skirrowii and/or A. cryaerophilus in their cloacae. None of the respective caecal samples were positive. Of the remaining 15 duck flocks, 11 (73%) were positive for A. cryaerophilus (7), A. butzleri (2) or A. skirrowii (2). Four (11%) of the 37 turkey flocks analyzed harboured either A. butzleri or A. cryaerophilus. The carriage rate of Arcobacter was higher in live ducks than those of live broiler chickens and turkeys in the present study. In addition, chicken carcasses slaughtered in Denmark were found to be contaminated with Arcobacter. The presence of Arcobacter spp. both on chicken carcasses and in poultry intestine may be of significance to human health.     

Isolation of Arcobacter spp. from retail meats and cytotoxic effects of isolates against vero cells

A survey of Arcobacter spp. was conducted over a 12-month period in Guadalajara, Mexico. A total of 135 samples (45 lean ground beef samples, 45 lean ground pork samples, and 45 chicken samples, including drumsticks, gizzards, and ground or chopped breast) were collected from local butcheries. The samples were enriched in Johnson-Murano enrichment medium and then streaked onto Johnson-Murano agar plates. Typical colonies were subjected to microscopic and biochemical identification followed by polymerase chain reaction confirmation of the genus Arcobacter. All isolates confirmed to be Arcobacter isolates were then inoculated into Eagle's minimum essential medium to determine their cytotoxicity against Vero cells. Arcobacter spp. were detected in 28.8, 51.1, and 40.0% of beef, pork, and chicken samples, respectively. From these samples, 101 isolates were confirmed to be Arcobacter spp. by polymerase chain reaction. Overall, the species most frequently identified was A. butzleri, followed by A. skirrowii. A. cryaerophilus was isolated only from pork meat. Ninety-five (95%) of the Arcobacter isolates produced a virulence mechanism against Vero cells, and 38 of them induced cell elongation, indicating enterotoxin production. Eighteen isolates produced the formation of vacuoles, and 39 produced both vacuolization and elongation. The vacuolization effect may be related to a vacuolizing toxin. The production of a vacuolizing toxin by Arcobacter spp. has not previously been reported. Results obtained in this study indicate that Arcobacter spp. may show cytotoxic effects other than the recognized enterotoxin production.     

Prevalence of Campylobacter spp. in poultry and poultry meat in Germany

Of 509 samples from poultry flocks, 209 isolates (41.1%) were Campylobacter positive. The number of positive cases in broiler carcasses was 45.9%. Of 52 pheasants investigated, 25.9% were Campylobacter positive. Campylobacter jejuni was isolated from 86 (42.0%) poultry flock samples, 47 (43%) broiler samples and 15 (28%) wild pheasant samples. C. coli was found at a rate of 1.2% in poultry flocks, 13% in broilers and 21% in pheasants.     

Relevant aspects of Arcobacter spp. as potential foodborne pathogen

Arcobacter species are Gram-negative spiral-shaped organisms belonging to the family Campylobacteraceae that can grow microaerobically or aerobically. The Arcobacter organisms also have the ability to grow at 15 degrees C, which is a distinctive feature that differentiates Arcobacter species from Campylobacter species. Cultural detection of Arcobacter is generally performed by an enrichment step and takes 4 to 5 days. In the last few years, several studies comparing different culture-based protocols have been published. Furthermore, DNA-based assays have also been established for rapid and specific identification of Arcobacter spp. Recent evidence suggests that Arcobacter, especially A. Butzleri, may be involved in human enteric diseases. Moreover, A. butzleri has also occasionally been found in cases of human extraintestinal diseases. However, up to now, little is known about the mechanisms of pathogenicity or potential virulence factors of Arcobacter spp. There is evidence that livestock animals may be a significant reservoir of Arcobacter spp. and over the last few years, the presence of these organisms in raw meat products as well as in surface and ground water has received increasing attention. In view of control measures to be used to prevent or to eliminate the hazard of Arcobacter spp. in food, several treatments have been evaluated for their effectiveness. While the role of Arcobacter spp. in human disease awaits further evaluation, a precautionary approach is advisable. Measures aimed at reduction or eradication of Arcobacter from the human food chain should be encouraged. With this article, we review the recent literature on this organism with a special emphasis on the information relevant to food safety.     

Arcobacter spp. enumeration in poultry meat using a combined PCR-ELISA assay

A rapid assay for enumeration of Arcobacter spp. in chicken meat was developed by using the polymerase chain reaction (PCR) coupled to an enzyme-linked immunosorbent assay (ELISA). Following a short selective enrichment of poultry samples, bacterial DNA was extracted and amplified using digoxigenin-labelled primers specific for 16S rRNA sequences of Arcobacter spp. Amplified fragments were heat denatured before being quantified by an ELISA. In this technique, a biotinylated probe immobilized onto streptavidin-coated microplates was used to capture the digoxigenin-PCR products. A peroxidase antidigoxigenin conjugate was added to the plate and, in the presence of substrate, PCR products were quantitated based on an optical density reading. Distinct absorbance differences were obtained when assaying poultry samples containing Arcobacter spp. in the range 10-10(4) CFU/g.     

Distribution and characterization of Campylobacter spp. from Russian poultry

The distribution of Campylobacter spp. on 13 poultry farms (broiler chicken, quail, pheasant, peacock, and turkey) from eight regions (Vladimir, Vologda, Voronezh, Kaluga, Liptsk, Moscow, Orenburg, and Orel) in Russia was surveyed. Intestinal materials were plated onto Campylobacter-selective medium and plates were incubated microaerobically at 42 degrees C for 24 or 48 h. Identification was based on colonial morphology, microscopic examination, and biochemical tests; latex agglutination assays were used for confirmation. In total, 116 isolates were derived from 370 samples. Isolation rates were similar, regardless of whether the birds were from small or large broiler production farms. Susceptibility of 48 representative (from these production sources) strains of Campylobacter spp. to 38 antimicrobial compounds was determined by disk diffusion assays. All strains tested were sensitive to amikacin, gentamycin, sisomycin, chloramphenicol, imipenem, oleandomycin, erythromycin, azitromycin, and ampicillin. The strains were also sensitive to 100 microg/disk of carbenicillin, fluoroquinolones, and to nitrofurans. Fluoroquinolone sensitivity was most notable and may be related to its limited application in poultry production within Russia. Hippurate and ribosomal RNA gene primers were developed and used to distinguish Campylobacter jejuni and Campylobacter coli and to provide a measure of strain discrimination. The combination of PCR analysis and randomly amplified polymorphic DNA (RAPD) typing were conducted for selected isolates. The various poultry species and the different locations yielded Campylobacter isolates with discrete randomly amplified polymorphic DNA patterns. The distribution and substantial diversity of Campylobacter spp. isolates appears similar to that previously reported in other countries.