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1.
In this study, 100 gall bladder samples of sheep slaughtered at an abattoir in Elazi? province were examined for Campylobacter jejuni and Campylobacter coli by culture and polymerase chain reaction (PCR). Preston Campylobacter Agar supplemented with 7% horse blood and Preston Selective Supplement (Oxoid, Hampshire, UK) were used for isolation of the agents. Campylobacter spp. were isolated in 66 samples, and they were identified as 34% C. jejuni and 32% C. coli. A multiplex PCR based upon the use of ceuE gene-specific primers was applied on DNA samples extracted from C. jejuni and C. coli isolates. All C. jejuni and C. coli strains that were positive by culture were also detected to be positive by PCR. This study shows that PCR can be used an alternative, rapid and sensitive test for the identification of C. jejuni and C. coli which threaten human and animal health.  相似文献   

2.
In this study, we evaluated the applicability of cytolethal distending toxin (cdt) gene-based species-specific multiplex PCR for the direct detection and identification of Campylobacter jejuni, C. coli and C. fetus from stool specimens of patients with gastroenteritis in comparison to culture methods. A total of 711 stool specimens were examined for the isolation or detection of campylobacters by using Skirrow's selective agar culture plates, a filtration method and the multiplex PCR assay. Forty-one and 36 C. jejuni strains were isolated by culture and filtration methods, respectively. In addition, 2 and 3 C. coli strains were isolated by Skirrow and the filtration methods, respectively. However, when the multiplex PCR was employed, the cdtB genes of C. jejuni and C. coli were detected in 45 and 4 stool samples, respectively, and 9 C. jejuni PCR-positive samples by multiplex PCR were negative by culture method. Sequence analysis of the PCR products obtained from 8 stool specimens from which campylobacters were not isolated by culture method but the sequences exactly matched with that of the cdtB gene of C. jejuni strain 81-176. None of the remaining stool samples which were culture negative for campylobacters produced any amplicon. Stool samples were defined as Campylobacter-positive if detected by any method. The sensitivity of the multiplex PCR was 83%, which was higher than Skirrow (74%) and filtration method (66%). These data indicate that cdtB gene-based multiplex PCR is a rapid and more sensitive method to identify the most important species of Campylobacter for human diseases. (248).  相似文献   

3.
Altogether 16 Campylobacter (C.) isolates could be recovered from 65 Herring gulls: 5 x C. laridis, 2 x C. jejuni biovar 1, 4 x C. jejuni biovar 2 and 5 x C. coli. Campylobacter spp. were isolated from 15 out of 51 samples from Kittiwakes: 2 x C. jejuni biovar 1 and 13 x C. laridis. All C. coli isolates grew on agar containing 1.5% NaCl. Two Campylobacter isolates from 50 House sparrows differed from all other isolates by a distinct beta-hemolysis and other phenotypic characteristics and could not be associated with a certain Campylobacter species. Epidemiological aspects and the possible role of the examined birds as a source of infection for man and domestic animals are discussed.  相似文献   

4.
Thermotolerant Campylobacter spp. are frequent causes of diarrhoea in humans worldwide mostly originating from poultry. It has been suggested that extensive veterinary use of antibiotics is largely responsible for resistance in human isolates. During a 4-month period from January to April 2004, 192 Campylobacter spp. were isolated from fecal samples of 485 healthy food animals. The in vitro susceptibility to 12 antibiotics was determined by the agar disk diffusion method. Among the 192 Campylobacter spp. isolated, 135 (70.3%) were identified to be C. jejuni, 51 (26.6%) were C. coli and 6 (3.1%) were C. lari. C. jejuni was the most prevalent species in chickens (80.8%) versus 16.2% C. coli and 3.0% C. lari. All isolates found in pigs were C. coli. All strains were sensitive to chloramphenicol and ciprofloxacin and all were resistant to cephalothin. More than 90% of the strains were sensitive to clindamycin, erythromycin, gentamicin, nalidixic acid, norfloxacin, streptomycin and tetracycline. Resistance was found against ampicillin in 20% and trimethoprim-sulphamethoxazole in 37.5%. Resistance was not statistically different among C. jejuni, C. coli and C. lari (p>0.05). Multidrug resistance to two or more drugs was detected in 14.5% of strains. In conclusion, the study showed that antimicrobial resistance is found only at relatively low frequencies for most antimicrobial agents tested except for ampicillin and trimethoprim-sulphamethoxazole. The low percentages of resistance to most antimicrobial agents tested in this study may be the result of low/no usage of these agents as a growth promoters or treatment in the Ethiopian animal farm setting. The detection of multidrug resistant isolates may pose a threat to humans and further limits therapeutic options.  相似文献   

5.
We report the isolation of Campylobacter species from the same population of feral swine that was investigated in San Benito County, California, during the 2006 spinach-related Escherichia coli O157:H7 outbreak. This is the first survey of Campylobacter in a free-ranging feral swine population in the United States. Campylobacter species were cultured from buccal and rectal-anal swabs, colonic faeces and tonsils using a combination of selective enrichment and antibiotic-free membrane filtration methods. Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF-MS, Bruker Daltonics, Inc., Billerica, MA, USA) was used to identify species followed by confirmatory multiplex PCR or 16S rRNA sequencing. Genetic relatedness of Campylobacter jejuni and Campylobacter coli strains was determined by multilocus sequence typing (MLST) and porA allele sequencing. Altogether, 12 (40%) of 30 feral swine gastrointestinal and oral cavity specimens were positive, and six species were isolated: Campylobacter coli, Campylobacter fetus, Campylobacter hyointestinalsis, Campylobacter jejuni, Campylobacter lanienae and Campylobacter sputorum. Campylobacter jejuni subtypes were closely related to MLST sequence type 21 (ST-21) and had identical porA sequences. Campylobacter coli subtypes were unrelated to isolates in the pubMLST/porA database. This feral swine population lived in close association with a 'grassfed' beef cattle herd adjacent to spinach and other leafy green row crop fields. The findings underscore the importance of protecting raw vegetable crops from faecal contamination by wild or feral animals. The study also illustrates a potential risk of Campylobacter exposure for hunters during handling and processing of wild swine meat.  相似文献   

6.
In a commercial broiler flock during rearing multiple genotypes of Campylobacter jejuni may be present as well as in gastrointestinal tracts of individual birds. The aim of this study was to optimize and apply a denaturing gradient gel electrophoresis assay of the flagellin gene (fla-DGGE) for analysis of C. jejuni and Campylobacter coli in cecal samples of broilers without prior cultivation. One C. coli and 21 C. jejuni strains isolated from broiler flocks, of which 14 typed as unique by restriction fragment length polymorphism of flaA and two undefined strains, were clustered into 9 groups when applying fla-DGGE. Spiking of cecal samples revealed that fla-DGGE is able to detect at least 4.55-5.96logCFUCampylobacter/mlcecal material. The presence of 3 strains spiked in cecal material was demonstrated by fla-DGGE as the corresponding bands were visible on the DGGE gel. Naturally contaminated cecal samples were shown to contain different types of C. jejuni and C. coli. Fla-DGGE has some potential as a cultivation-independent fast primary subtyping method for C. jejuni and C. coli in cecal samples of broilers.  相似文献   

7.
Using a newly formulated selective medium containing cefoperazone, we isolated 72 Campylobacter strains in fecal samples from 397 diarrheic dogs and cats. Of these, 39 were thermophilic catalase-negative Campylobacter species. We identified these Campylobacter strains by DNA:DNA hybridization, using digoxigenin-labeled total genomic DNA of 4 Campylobacter reference strains (C jejuni, C coli, C lari, and C upsaliensis) as a probe. The labeling was done with a commercially available kit. We could identify 66 of the 72 Campylobacter isolates to the species level with this method; identification with probes always agreed with conventional test results. Of the 66 identified strains, 33 were C upsaliensis and 33 were C jejuni. Six isolates could not be assigned to a known species with probes or conventional tests. On the basis of our findings, C upsaliensis is more resistant to cefoperazone than to cephalothin, thereby explaining the unexpected recovery of these campylobacters on cephalosporin-containing media.  相似文献   

8.
A total of 104 fecal specimens from 30 mammals, 12 birds, and 3 reptiles at the Phoenix Zoological Gardens, Miyazaki City, Japan, were examined for the presence of Campylobacter species. All the animals examined were healthy with no fecal abnormality. Twenty-three (22.1%) thermophilic campylobacters, (9 C. jejuni, 11 C. hyointestinalis, 2 C. coli, and 1 C. lari), were isolated from 11 animals (7 mammals and 4 birds). C. jejuni and C. hyointestinalis were the predominant species isolated from these zoo animals and C. hyointestinalis was isolated frequently from simians. As selective media influence the numbers and species of campylobacters isolated, the agar medium was not supplemented with cephalothin. Campylobacters were isolated most frequently when a combination of enrichment culture and selective agar plating was performed at 42 degrees C. For the epidemiological study, a polymerase chain reaction (PCR)-based randomly amplified polymorphic DNA (RAPD) method was used as a tool to detect the heterogeneity of amplified DNAs of Campylobacter spp. isolated from zoo animals. The two arbitrary primers used in this study enabled even closely related strains of the same Campylobacter spp. to be differentiated. RAPD analysis revealed considerable diversity among the strains, suggesting that the transmission of Campylobacter spp. among animals in a defined area occurred through different mechanisms.Examination of the genotypic diversity among the multiple clones from the same host also revealed differences between clones. These results demonstrate that campylobacter populations in zoo animals are highly divergent.  相似文献   

9.
Campylobacter jejuni and Campylobacter coli inoculation of neonatal calves   总被引：2，自引：0，他引：2  
Three groups of neonatal calves were inoculated orally with pathogenic strains of Campylobacter jejuni or C coli. The calves developed a mild, self-limiting enteritis characterized by thick mucoid feces. Bacteremia and fecal shedding of Campylobacter were sporadic in all inoculated calves. Two groups of calves were killed 1 to 3 weeks after inoculation to study the pathogenesis of infection. Postmortem culture of tissues revealed C jejuni or C coli most frequently in the ileum, cecum, colon, and blood. Clinical or pathologic differences between C jejuni-inoculated and C coli-inoculated calves were minimal.  相似文献   

10.
The isolation and antimicrobial resistance of Campylobacter jejuni and Campylobacter coli strains from broilers arriving in French slaughterhouses, were analysed according to production types (i.e. standard, export or free-range) and antimicrobial (i.e. coccidiostats, growth promoters or therapeutic agents) administration in flocks. Prevalence was 56.6% in standard, 51.3% in export and 80.0% in free-range broilers. Three hundred and ninety-three strains were identified. Two-thirds of the strains belonged to the species C. jejuni. The others were C. coli. Antimicrobial susceptibility testing was carried out for ampicillin, nalidixic acid, enrofloxacin, tetracycline, erythromycin and gentamicin according to a dilution method. The percentages of resistant strains were, 23, 25, 17, 57, 0.3 and 0% for C. jejuni and 29, 43, 40, 70, 31 and 0% for C. coli. Statistical analysis revealed significant difference in distribution of C. jejuni and C. coli and antimicrobial resistance according to production type or antimicrobial administration.  相似文献   

11.
The presence of the flaA, cadF, cdtB and iam genes of Campylobacter spp. was determined with the PCR method. The materials to investigate were 56 C. jejuni and 23 C. coli strains isolated from clinical samples (children and domestic animals). It was found that all of the Campylobacter spp. isolates from children with diarrhoea and domestic animals had cadF gene, responsible for adherence. The flaA gene was present in all Campylobacter spp. isolates derived from children and cats. Occurrence of flaA gene was confirmed in 100% of C. jejuni strains obtained from dogs. The high prevalence of the cdtB gene associated with toxin production was observed in this study (100%-Campylobacter spp. isolates obtained from dogs and cats, 97.9%-Campylobacter spp. isolates from children). The isolates showed a wide variation for the presence of iam gene. The lowest prevalence (23.5%) was detected in Campylobacter spp. obtained from dogs. The highest rates of iam detection (91.6%) were revealed in C. coli isolates from children.  相似文献   

12.
The aim of this study was to employ a novel cytotoxicity assay based on primary porcine aortic endothelial cells in combination with a lactate dehydrogenase release assay to quantitatively determine differences in cytotoxin production between Campylobacter jejuni, C. coli, C. lari and urease-positive thermophilic campylobacters (UPTC), isolated from human faeces, animals and environmental sources. Campylobacter isolates totalling 34 and comprising of C. jejuni (n = 24) C. coli (n = 5) and UPTC (n = 4) and C. lari (n = 1) were analysed. The cytotoxic response ranged from 32.15 to 64.47% and 33.08 to 59.41%, for C. jejuni from chicken and human isolates, respectively and there was no statistically significant difference (P > 0.05) in cytotoxic response between C. jejuni isolated from humans and chicken isolates (50.78% versus 50.55% cytotoxicity, respectively). However, there was a difference in response between C. jejuni and C. coli isolated from chickens (50.78% versus 33.22% cytotoxicity, respectively). The greatest cytotoxic response was obtained with the UPTC group of organisms examined (n = 4 isolates) (mean cytotoxic response = 57.11% cytotoxicity. Employment of this cytotoxin assay may help identify virulent strains in poultry that could potentially proceed to cause clinical problems for humans and thus intervention measures targeted at the reduction or elimination of such specific strains, may be sought.  相似文献   

13.
One hundred and twenty five chickens from Grenada, consisting of 77 broilers and 48 layers were examined for carriage of thermophilic campylobacters in their ceca by culture. Seventy nine percent of chickens were positive for campylobacters, with an isolation rate of 93.5% for broilers and 56.3% for layers, the difference being significant. Sixty-four pure cultures comprising 39 Campylobacter coli, 21 Campylobacter jejuni, and 4 Campyilobacter lari isolates were tested for their resistance against 7 antibiotics using the E-test. None of the isolates were resistant to chloramphenicol and gentamicin. Resistance rates to other drugs were: ampicillin, 9.4%; ciprofloxacin, 12.5%; erythromycin, 3.1%; metronidazole, 9.4%, and tetracycline, 50% with MICs of >or=256 microg/mL for tetracycline. There were no significant differences in resistance rates between C. coli and C. jejuni. Multiple resistance to >or=2 drugs was seen in 15.6% of total isolates. All C. lari isolates were resistant to ciprofloxacin, and 3 of 4 isolates had multiple drug resistance. Overall, erythromycin, which is the drug of choice for treatment of Campylobacter infections in humans, is effective in vitro against 97% of chicken isolates in Grenada.  相似文献   

14.
AIM: To investigate the role of free-living animals such as sparrows, rodents and flies as potential reservoirs of Campylobacter spp on a dairy farm, and to assess the genetic diversity among Campylobacter isolates from the farm and an urban source. METHODS: A total of 290 samples (bovine, passerine and rodent faeces, and whole flies) were collected from a large commercial dairy farm in the Manawatu district in New Zealand, and from faeces from urban sparrows in a nearby city. Other samples collected from the dairy farm included five from silage, two from aprons worn by workers during milking, two from workers' boots and two from water in troughs in a paddock. Isolates of thermophilic Campylobacter spp were identified morphologically and phenotypically and further characterised molecularly using pulsed-field gel electrophoresis (PFGE) and the restriction enzyme SmaI. RESULTS: Campylobacter jejuni was the only Campylobacter species isolated from all samples. The highest prevalence was found in faeces from dairy cows (54%), followed by faeces from sparrows from the urban area (40%) and the farm (38%), and from rodents (11%) and whole flies (9%). Other samples from the farm environment such as silage, trough water, and workers' aprons and boots were also positive for C. jejuni. Of the 22 restriction patterns obtained, seven were common to more than one source. CONCLUSIONS: Cattle, sparrows, rodents and flies are potential reservoirs of C. jejuni on dairy farms. Identical clones of C. jejuni carried by cattle, sparrows, flies and rodents probably indicate a common source of infection. The high level of asymptomatic carriage of C. jejuni by healthy dairy cows could be sufficient to maintain infections within the dairy farm surroundings via environmental contamination.  相似文献   

15.

Background

Campylobacter is the most commonly reported bacterial cause of enteritis in humans in the EU Member States and other industrialized countries. One significant source of infection is broilers and consumption of undercooked broiler meat. Campylobacter jejuni is the Campylobacter sp. predominantly found in infected humans and colonized broilers. Sequence analysis of the 16S rRNA gene is very useful for identification of bacteria to genus and species level. The objectives in this study were to determine the degree of intraspecific variation in the 16S rRNA genes of C. jejuni and C. coli and to determine whether the 16S rRNA sequence types correlated with genotypes generated by PFGE analysis of SmaI restricted genomic DNA of the strains.

Methods

The 16S rRNA genes of 45 strains of C. jejuni and two C. coli strains isolated from broilers were sequenced and compared with 16S rRNA sequences retrieved from the Ribosomal Database Project or GenBank. The strains were also genotyped by PFGE after digestion with SmaI.

Results

Sequence analyses of the 16S rRNA genes revealed nine sequence types of the Campylobacter strains and the similarities between the different sequence types were in the range 99.6–99.9%. The number of nucleotide substitutions varied between one and six among the nine 16S rRNA sequence types. One of the nine 16S rRNA sequence profiles was common to 12 of the strains from our study and two of these were identified as Campylobacter coli by PCR/REA. The other 10 strains were identified as Campylobacter jejuni. Five of the nine sequence types were also found among the Campylobacter sequences deposited in GenBank. The three 16S rRNA genes in the analysed strains were identical within each individual strain for all 47 strains.

Conclusion

C. jejuni and C. coli seem to lack polymorphisms in their 16S rRNA gene, but phylogenetic analysis based on 16S rRNA sequences was not always sufficient for differentiation between C. jejuni and C. coli. The strains were grouped in two major clusters according to 16S rRNA, one cluster with only C. jejuni and the other with both C. jejuni and C. coli. Genotyping of the 47 strains by PFGE after digestion with SmaI resulted in 22 subtypes. A potential correlation was found between the SmaI profiles and the 16S rRNA sequences, as a certain SmaI type only appeared in one of the two major phylogenetic groups.  相似文献   

16.
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.  相似文献   

17.
A dramatic rise in the number of resistant Campylobacter to quinolones has been documented in human patients and domestic animals. In this study, the mechanism of acquisition of quinolone resistance was studied by detecting point mutations in the gyrA gene of Campylobacter strains obtained from broilers and strains with in vitro-induced resistance. The minimal inhibitory concentrations (MICs) of norfloxacin (NFLX) and ofloxacin (OFLX) for the strains that had no point mutation were slightly increased from the source strain (Campylobacter jejuni ATCC 33560). The MICs of nalidixic acid (NA), NFLX, and OFLX for the strains that had the point mutation at Thr-86 were 100 or 200 microg/ml, 50 microg/ml, and 25 microg/ml, respectively. The MIC of NA for the strain that had a point mutation at Asp-90 higher than those for the strains that had the point mutation at Thr-86, but the MICs of NFLX and OFLX were relatively lower than those for the strains that had point mutation at Thr-86. These findings suggest that the degree of antimicrobial resistance against NA, NFLX, and OFLX in the in vitro-induced C. jejuni strains was associated with the location of the point mutation in gyrA. On the other hand, a point mutation in all seven resistant strains isolated from broilers was located only at Thr-86, while the MICs of the three quinolones varied in each wild strain. This suggests that another mechanism might also be involved in the acquisition of quinolone resistance in C. jejuni wild strains.  相似文献   

18.
AIM: To investigate the role of free-living animals such as spar- rows, rodents and flies as potential reservoirs of Campylobacter spp on a dairy farm, and to assess the genetic diversity among Campylobacter isolates from the farm and an urban source.

METHODS: A total of 290 samples (bovine, passerine and ro- dent faeces, and whole flies) were collected from a large com- mercial dairy farm in the Manawatu district in New Zealand, and from faeces from urban sparrows in a nearby city. Other samples collected from the dairy farm included five from silage, two from aprons worn by workers during milking, two from workers' boots and two from water in troughs in a paddock. Isolates of thermophilic Campylobacter spp were identified mor- phologically and phenotypically and further characterised mo- lecularly using pulsed-field gel electrophoresis (PFGE) and the restriction enzyme SmaI.

RESULTS: Campylobacter jejuni was the only Campylobacter species isolated from all samples. The highest prevalence was found in faeces from dairy cows (54%), followed by faeces from sparrows from the urban area (40%) and the farm (38%), and from rodents (11%) and whole flies (9%). Other samples from the farm environment such as silage, trough water, and work- ers' aprons and boots were also positive for C. jejuni. Of the 22 restriction patterns obtained, seven were common to more than one source.

CONCLUSIONS: Cattle, sparrows, rodents and flies are po- tential reservoirs of C. jejuni on dairy farms. Identical clones of C. jejuni carried by cattle, sparrows, flies and rodents prob- ably indicate a common source of infection. The high level of asymptomatic carriage of C. jejuni by healthy dairy cows could be sufficient to maintain infections within the dairy farm sur- roundings via environmental contamination.  相似文献   

19.
Campylobacter jejuni and Campylobacter coli strains were isolated from feces of dairy cattle at farms with no known problem due to campylobacteria. Farms were located in the northeast, desert southwest, and Pacific west. Twenty isolates were identified by ribotyping with a RiboPrinter. The ability of these bovine isolates to colonize the ceca of chicks was determined by challenge inoculation and reisolation of the challenge strain from the ceca at 1 and 2 wk after challenge. Isolates recovered from chick ceca were examined by ribotyping to assure they matched the challenge strain. One hundred percent of the bovine-derived challenge strains were capable of colonizing chicks. These results indicate that dairy cattle may be asymptomatic Campylobacter carriers and potential sources of campylobacteria contamination of poultry facilities.  相似文献   

20.
Thermotolerant Campylobacter spp., in particular Campylobacter jejuni, are among the most frequently identified pathogens, found to be causing human gastrointestinal infections in Europe, with the Czech Republic being no exception. The presented work aimed at assessing results of the first nationwide monitoring of prevalence and antibiotic resistance of Campylobacter spp. in broiler flocks in the Czech Republic, including a comparison of antibiotic resistance of C. jejuni isolates collected from poultry and the human community. The monitoring was carried out in poultry slaughterhouses in 2006 and 2007. From broilers, cloacal swabs were collected and examined. The human isolates of C. jejuni were acquired from rectal swabs in community patients with diarrhoeal diseases. Suspected isolates of both animal and human origin were confirmed by the PCR methods. Antibiotic resistance to selected anti-microbial agents was tested by the microdilution method. In the monitored period, the prevalence of thermotolerant Campylobacter spp. in broilers in the Czech Republic reached almost 50%. In 2006, C. jejuni was detected in 46% and Campylobacter coli in 3% of the tested samples. In 2007, C. jejuni was found in 43% and C. coli in 2% of the samples. The results of anti-microbial susceptibility testing of C. jejuni showed higher resistance in animals when compared with humans. The only exception was tetracycline with higher resistance in isolates of human origin. The highest resistance detected was to quinolone antibiotics. Resistance to oxolinic acid was 77% in animal and 60% in human isolates, to ciprofloxacin 72% in isolates from poultry and 55% in those from humans. In ampicillin, 26% of poultry isolates and 16% of human isolates were resistant. Moreover, 9% of animal isolates demonstrated resistance to streptomycin, undetected in human isolates. In erythromycin, resistance was found in 6% of poultry and 1% of human isolates.  相似文献   

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