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PubMed2004

Prevalence of Arcobacter species in retail meats and antimicrobial susceptibility of the isolates in Japan.

A survey was conducted to examine the prevalence of Arcobacter species among meat samples and to investigate the antimicrobial susceptibility of the isolates in Japan. In 1998 and 1999, samples of beef (n=90), pork (n=100) and chicken meat (n=100) were purchased from seven retail shops. Arcobacter species were isolated from 2.2%, 7.0% and 23.0% of beef, pork and chicken meat samples, respectively. The rate of isolations in chicken meats was shown to be significantly higher than those of beef and pork. Species-specific polymerase chain reaction (PCR) demonstrated that the most dominant Arcobacter species was Arcobacter butzleri among the isolates examined. Multiple contaminations with different Arcobacter species were observed in 5% of the chicken samples. Almost all the strains tested showed resistance to vancomycin (100%) and methicillin (97.5%). Strains resistant to cephalothin, sulfamethoxazole-trimethoprim, nalidixic acid and chloramphenicol were detected at the rate of 81.1%, 67.2%, 53.5% and 24.6%, respectively. All Arcobacter strains examined were susceptible to ampicillin, tetracycline, streptomycin and kanamycin.

Animals↗

PubMed2001

Genotypic variation among arcobacter isolates from a farrow-to-finish swine facility.

Arcobacter spp. were isolated from nursing sows and developing pigs on three farms of a farrow-to-finish swine operation and market-age pigs at slaughter. Isolates were identified by polymerase chain reaction and genotypic fragment patterns were examined by pulsed-field gel electrophoresis (PFGE). Incidences of Arcobacter-positive samples increased progressively as the pigs aged, resulting in all of the pens at the end of the growth cycle in the finishing barn containing Arcobacter-positive feces. However, only 10 of 350 cecal samples from slaughtered pigs were positive. There was little similarity between genotypic patterns for Arcobacter collected from the three farms. The level of genotypic variation revealed by PFGE suggested that pigs in this farrow-to-finish operation were colonized by multiple Arcobacter parent genotypes that may have undergone genomic rearrangement, common to members of Campylobacteraceae, during successive passages through the animals. Additionally, the level of genotypic diversity seen among Arcobacter isolates from farms of a single farrow-to-finish swine operation suggests an important role for genotypic phenotyping as a source identification and monitoring tool during outbreaks.

Age Factors↗

PubMed1997

Detection of Arcobacter species in gastric samples from swine.

Swine stomachs were surveyed for evidence of Arcobacter spp. and Helicobacter spp. infections associated with gastric ulceration. A nested PCR test targeted to the 16S rRNA was developed to detect many Arcobacter spp. and Helicobacter spp. An internal oligonucleotide probe was used for differentiation and confirmation of the PCR product. Tissue samples were obtained from the nonglandular and glandular regions of 86 swine stomachs. Evidence of infection with these microbes was detected in 51%, with 77% of the positive samples being identified as A. butzleri using a highly specific probe. Nonglandular stomach samples (44%) were more likely to be positive by PCR than samples from the glandular (23%) region. Gross lesions of any stage of gastric ulceration, ranging from parakeratosis, erosions and ulceration, were observed in 24% of stomachs examined. Of 21 samples with lesions, 52% were positive by the broadly reactive PCR assay for Arcobacter spp. and Helicobacter spp. The majority of PCR-positive samples (75%) had no gross lesions. When a single step PCR assay that was more specific for Arcobacter spp. was used on the nonglandular stomach samples, 10.4% of the 86 samples were positive. Arcobacter spp. were cultured from four of the sample stomachs. Partial sequencing of the 16S rRNA gene identified the isolates as A. butzleri (n = 2), A. cryaerophilus, (n = 1), and a mixed culture of A. butzleri and another Arcobacter spp. (n = 1). A single step PCR assay targeted to the urease gene and culturing methods were used to screen for H. pylori or other closely related urease positive bacteria, but none were found.

Animals↗

PubMed1996

Identification of Arcobacter isolates by PCR.

A polymerase chain reaction (PCR) assay was developed for the identification of the three species of Arcobacter which have been recovered from clinically ill or healthy humans and/or livestock, namely Arcobacter butzleri, Arcobacter skirrowii and Arcobacter cryaerophilus. The assay utilizes primers targeted to the genes encoding 16S rRNA of Arcobacter spp. The assay reduces the amount of time required to positively identify strains of Arcobacter.

Bacteria↗

PubMed1996

Phenotypic and ribosomal RNA characterization of Arcobacter species isolated from porcine aborted fetuses.

Aerotolerant organisms resembling Campylobacter, now designated as Arcobacter, have been described from aborted farm animals and from cases of human enteritis worldwide. The goals of this study were 1) to attempt to recover Arcobacter spp. from cases of porcine abortion, 2) to characterize these isolates by phenotype and ribotype, and 3) to compare the usefulness of ribotype and phenotype patterns for identifying Arcobacter butzleri and the DNA hybridization groups 1A and 1B of A. cryaerophilus. Isolates of Arcobacter spp. from North Carolina and Iowa were recovered from porcine tissues. In Iowa, Arcobacter spp. were recovered from 43% (13/30) of porcine abortion cases evaluated. Isolations were made from placenta (44%), kidney (44%), and stomach contents (12%), which were the only tissues examined. The most reliable biochemical tests for A. butzleri included growth in 1% glycine and in 1.5% NaCl, weak catalase activity, and resistance to cadmium chloride. Arcobacter cryaerophilus strains were characterized by strong catalase activity and sensitivity to cadmium chloride. The DNA hybridization groups 1A and 1B of A. cryaerophilus could not be distinguished by biochemical tests. This represents the first description of A. cryaerophilus DNA group 1A in animals within the United States.

Abortion, Veterinary↗

PubMed1999

Comparison of three protocols for the isolation of Arcobacter from poultry.

The microaerophilic bacterium Arcobacter has received increasing attention in recent years regarding its presence in food products. There exist a limited number of methods for the detection of this microorganism, with currently available methods being cumbersome to perform, time consuming, and limited in specificity. The objective of this study was to develop a selective enrichment broth to isolate accurately three Arcobacter spp. from concentrated chicken microflora by comparing the efficacy of various selective and growth-promoting additives in order. This newly developed enrichment broth was incorporated into an isolation protocol using a previously developed plating medium, and this new protocol was compared with two existing methods for the isolation of Arcobacter from poultry. Method 1 consisted of enrichment in Ellinghausen-McCullough-Johnson-Harris Polysorbate 80 broth followed by plating on cefoperazone-vancomycin-amphotericin B medium. Method 2 consisted of enrichment in Arcobacter selective broth and plating onto Arcobacter selective medium. Method 3 (the JM method), used a newly developed enrichment broth followed by plating on a previously described JM agar. The JM method isolated Arcobacter strains in 42 out of 50 broiler chicken samples, while methods 1 and 2 detected the organism in only 24 and 15 out of 50 samples, respectively.

Animals↗

PubMed2003

Isolation of Arcobacter butzleri and A. cryaerophilus in samples of meats and from meat-processing plants by a culture technique and detection by PCR.

A pilot survey of sources of contamination with arcobacters (representing a potential risk for humans) was done in a wide range of samples involved various kinds of meat (beef, pork, meat products, chilled chickens, etc.) from a retail level and domestic farming. Sanitary practices in slaughterhouses and production lines were checked in two different plants (a beef and pork production and a chicken processing plant). The method is based on a selective enrichment to isolate suspect strains, in combination with a PCR technique specific for arcobacters. The choice of a suitable enrichment broth and a plating agar was made with the use of pure bacterial strains and by means of real meat samples seeded with Arcobacter butzleri. The PCR technique was optimized to allow differentiation of a 1223 bp product, typical of the genus Arcobacter, and a product of 686 bp, specific for A. butzleri a total number of 198 samples were tested, of that 33 (17%) were found to be positive for the genus Arcobacter but only 22 (11%) for A. butzleri.

Agar↗

PubMed2004

Occurrence and strain diversity of Arcobacter species isolated from healthy Belgian pigs.

In this study, Arcobacter species were isolated from clinically healthy porkers and sows on four unrelated pig farms, using a quantitative isolation protocol. Isolates were identified by m-PCR, and fingerprints were distinguished by modified ERIC-PCR. The prevalence of Arcobacter in pigs ranged from 16 to 85%. Arcobacter excretion ranged from 0 to 10(4) CFUg(-1) feces. Arcobacter butzleri was the most frequently occurring species, but simultaneous shedding of two or three species occurred. Large heterogeneity among the Arcobacter species was detected in pigs and on the farms.

Animals↗

PubMed2004

Prevalence, enumeration and strain variation of Arcobacter species in the faeces of healthy cattle in Belgium.

Arcobacter species were isolated from faeces of healthy cattle on three unrelated Belgian farms, using a quantitative isolation protocol. Isolates were identified by m-PCR and characterized by modified ERIC-PCR. The Arcobacter prevalence on the three farms ranged from 7.5 to 15%. The prevalence in dairy cattle ranged from 5.9 to 11% and for young cattle and calves, the prevalence was determined as 18.9 and 27.3%, respectively. Of the 276 animals examined, eight had a bacterial load of more than 10(2) cfu/g faeces and low levels were detected in 22 animals using enrichment. The Arcobacter excretion ranged from 0 to 10(4) cfu/g faeces. Arcobacter cryaerophilus was the dominant species isolated from cows, but co-colonizations occurred in 26% of the Arcobacter excreting animals. Characterization of the 164 isolates showed a large heterogeneity and animals could be colonized with more than one genotype.

Animals↗

PubMed2003

One-step polymerase chain reaction-based typing of Arcobacter species.

A species-specific PCR assay was developed for the identification of the Arcobacter species, Arcobacter butzleri, Arcobacter cryaerophilus 1A and 1B, and Arcobacter skirrowii. The primers, which amplify the most variable areas of the 23S rRNA gene, were designed to perform species-specific identification by one-step PCR. The DNA sequence of the region from A. cryaerophilus 1B was determined, and the specific pimer for the species was designed. By using one-step PCR containing the mixed primers N.butz, N.c1.A, N.c.1B, and N.ski, species-specific amplifications were detected from the reference strains of A. butzleri, A. cryaerophilus 1A, 1B, and A. skirrowii, respectively. Primers designed in this study were also evaluated on 10 of Japanese field isolates, and all species were identified. This simple one-step PCR assay was found to be a powerful tool for the survey of Arcobacter infection.

Arcobacter↗

PubMed2003

Distribution of Arcobacter species among livestock in Japan.

A survey was conducted to examine the distribution of Arcobacter species among livestock in Japan. During May 1999 and May 2000, fecal samples from cattle (n=332) and swine (n=250), chicken cloacal swabs (n=234), and vaginal swabs of cattle (n=61) and swine (n=15) were submitted for the isolation of Arcobacter species. Arcobacter species were isolated from 3.6 and 10.0% of the cattle and swine fecal samples, respectively, along with 14.5% of chicken cloacal swabs. No significant seasonal differences were observed. Species-specific polymerase chain reaction assay showed that A. butzleri was the most prevalent species (83.3, 60.0 and 47.1% of the cattle, swine and chicken isolates, respectively), followed by A. cryaerophilus 1B (16.7, 36.0 and 55.9% of the cattle, swine and chicken isolates, respectively). Of the samples from vaginal swabs, 8.1 and 13.3% were positive for Arcobacter in cattle and swine, respectively. This is the first report demonstrating the distribution of Arcobacter species among livestock in Japan.

Animals↗

PubMed2000

Development of a combined PCR-culture technique for the rapid detection of Arcobacter spp. in chicken meat.

A combined PCR-culture technique was developed to detect Arcobacter spp. in fresh chicken meat. Following a short selective enrichment of chicken samples, bacterial DNA was extracted and amplified using primers targeted at the genes encoding 16S rRNA of Arcobacter spp. The selected primers amplify a 181-bp fragment from all Arcobacter spp., whereas no PCR product is generated for other bacteria, including the closely related Campylobacter and Helicobacter species. The assay was used to screen 96 retail-purchased chicken samples for the presence of Arcobacter spp. Fifty-three percent of the samples analysed were positive for this micro-organism. The assay is simple and sensitive and reduces the amount of time required to positively detect Arcobacter spp. in poultry meat.

Animals↗

PubMed2006

Detection and identification of food-borne pathogens of the genera Campylobacter, Arcobacter and Helicobacter by multiplex PCR in poultry and poultry products.

The objective of this study was to develop a multiplex polymerase chain reaction (PCR) to detect and differentiate food-borne pathogens of the three genera Campylobacter, Arcobacter and Helicobacter in a single step procedure. One common reverse primer and three genus-specific forward primers were designed by hybridizing to the 16S rRNA of selected reference strains. Besides the species with significance as food-borne pathogens isolated from poultry meat--Campylobacter jejuni, Campylobacter coli, Arcobacter butzleri and Helicobacter pullorum--several other members of these genera were tested to determine the specificity of the designed multiplex PCR. In total, 20 ATCC and NCTC reference strains of Campyobacter, Arcobacter and Helicobacter were used to evaluate the PCR. Specific amplificates were obtained from all thermophilic species of Campylobacter as well as from species of Arcobacter and Helicobacter. No amplification product was obtained from the non-thermophilic Campylobacter, C. hyointestinalis and C. fetus. Furthermore, a total of 43 field strains of the three genera isolated from poultry, pigs, cattle and humans were investigated using this PCR. To confirm the classification of 10 H. pullorum strains the 16S rRNAs were sequenced. The developed PCR is a helpful diagnostic tool to detect and differentiate Campylobacter, Arcobacter and Helicobacter isolated from poultry and poultry products.

Animals↗

PubMed2002

Assessment of the genetic diversity among arcobacters isolated from poultry products by using two PCR-based typing methods.

In this study, enterobacterial repetitive intergenic consensus PCR (ERIC-PCR) and randomly amplified polymorphic DNA PCR (RAPD-PCR) were optimized for characterization of Arcobacter butzleri, Arcobacter cryaerophilus, and Arcobacter skirrowii. In addition, a simple and rapid DNA extraction method was tested for use in both typing procedures. Both methods had satisfactory typeability and discriminatory power, but the fingerprints generated with ERIC-PCR were more reproducible and complex than those obtained with RAPD-PCR. The use of nondiluted boiled cell suspensions as DNA templates was found to be very useful in ERIC-PCR. Characterization of large numbers of Arcobacter isolates is therefore preferably performed by the ERIC-PCR procedure. Isolates for which almost identical ERIC fingerprints are generated may subsequently be characterized by RAPD-PCR, although adjustment and standardization of the amount of the DNA template are necessary. In the second part of this study, the genotypic diversity of arcobacters present on broiler carcasses was assessed by using both typing methods. A total of 228 cultures from 24 samples were examined after direct isolation and enrichment. The isolates were identified by using a multiplex PCR as A. butzleri (n = 182) and A. cryaerophilus (n = 46). A total of 131 types (91 A. butzleri types and 40 A. cryaerophilus types) were discerned without discordance between the two typing techniques. The analysis of the poultry isolates showed that poultry products may harbor not only more than one species but also multiple genotypes. All genotypes were confined to one poultry sample, and only three genotypes were found after simultaneous enrichment and direct isolation. These results demonstrate that different outcomes can be obtained in epidemiological studies depending on the isolation procedure used and the number of isolates characterized.

Arcobacter↗

PubMed2002

Sensitivity of three methods used in the isolation of Arcobacter spp. in raw ground pork.

Arcobacter, an aerotolerant Campylobacter-like organism, has been designated an emerging pathogen because of its newly recognized ability to cause diarrheal illness in both humans and animals and its presence in the human food supply. Because there is no standard isolation method for its detection, the true occurrence of this pathogen is largely unknown. In addition, the lack of a standardized isolation protocol limits the ability of investigators to compare field data. Arcobacter has been detected in whole muscle and ground pork at various levels by two different isolation methods (those of deBoer and Collins). In this study, these methods were tested along with the Johnson-Murano (JM) method, developed in our laboratory. The sensitivity of each method was tested for ground pork inoculated with Arcobacter butzleri and Arcobacter cryaerophilus IA at levels of 10(4), 10(3), 10(2), and 10(1) CFU/g. Controls included tubes with uninoculated pork and broth tubes without pork. All samples that were morphologically similar to Arcobacter were analyzed by Gram staining and by catalase and oxidasereactions. Presumptive positive samples were confirmed by the polymerase chain reaction. The JM method was determined to be the most sensitive, detecting A. butzleri down to a level of 10(1) CFU/g in 100% of the samples and detecting A. cryaerophilus IA at a level of 10(1) CFU/g in 75% of samples. In a pure buffer system, the Collins method was as effective as the JM method in isolating both organisms to levels of 10(1) cells per g.

Animals↗

PubMed1998

Diversity and prevalence of Arcobacter spp. in broiler chickens.

Ninety-nine strains of Arcobacter spp., isolated from 10 chicken carcasses purchased from a supermarket and 15 chicken carcasses collected from a poultry abattoir, were speciated using a variety of phenotypic identification methods. All were tested using API Campy test strips and the 16-test (Preston) identification scheme developed for campylobacters. Fifty strains were selected for examination using a more comprehensive probabilistic identification scheme, and the identity of representative strains confirmed by protein profiling using SDS-PAGE. All 25 carcasses yielded Arcobacter butzleri. Three supermarket and 10 abattoir carcasses also carried A. cryaerophilus, and two abattoir carcasses carried A. skirrowii. The API Campy scheme proved unsatisfactory for identifying these strains: only 20 of 99 strains were accurately identified, all of which were A. cryaerophilus, the only Arcobacter sp. included in the database. Moreover, 76 of 99 strains were misidentified. The 16-test scheme identified all the arcobacter strains as A. cryaerophilus, since neither A. butzleri nor A. skirrowii had been described when the scheme was developed. The computer-assisted probabilistic scheme succeeded in identifying all but one strain, the identity of which was clarified by the use of SDS-PAGE. To our knowledge this is the first time that arcobacters other than A. butzleri have been reported in poultry meat or any other food of animal origin. Their high prevalence in poultry products may be of significance to public health.

Animals↗

PubMed1998

Isolation of Arcobacter butzleri in raw water and drinking water treatment plants in Germany.

A total of 147 "Campylobacter-like" strains were isolated from six drinking water treatment plants within a two year investigation period. The strains were characterized by bio-, and serotyping according the scheme for Arcobacter butzleri. The result was that 100 strains were typed as Arcobacter butzleri, 17 strains as Arcobacter butzleri-like and 6 strains as Campylobacter jejuni/coli. 24 strains were typed as Arcobacter spp. The strains isolated from the treatment plants showed the same serotypes as described for human isolates. Therefore the spread of Arcobacter organisms via the drinking water path must be suspected.

Bacterial Typing Techniques↗

PubMed2003

Isolation of Arcobacter species from animal feces.

A previously developed Arcobacter isolation protocol for poultry skin and meat was validated for the isolation of Arcobacter from feces of livestock animals. Good repeatability, in-lab reproducibility and sensitivity were achieved and the specificity was improved by additional incorporation of cycloheximide and increase of the novobiocin concentration in the selective supplement. The limit of detection of quantitative and qualitative analysis was 10(2) and 10(0) cfu g(-1) feces, respectively. From fecal samples collected at slaughterhouse, Arcobacter was isolated from 43.9% of porcine, 39.2% of bovine, 16.1% of ovine and 15.4% of equine samples. All three animal-associated Arcobacter species were isolated and levels up to 10(3) cfu g(-1) feces were determined.

Abattoirs↗