Campylobacter jejuni/coli: Methodology of Isolation and Possible Interfering Factors in Primary Culture

1996 ◽  
Vol 59 (4) ◽  
pp. 429-432 ◽  
Author(s):  
MARIA HELENA C. AQUINO ◽  
JOSÉ CARLOS P. CARVALHO ◽  
ANITA TIBANA ◽  
ROBSON M. FRANCO
Keyword(s):  
Primary Culture ◽  
Campylobacter Jejuni ◽  
Campylobacter Coli ◽  
Enrichment Broth ◽  
Direct Plating ◽  
Indicator Microorganisms ◽  
Enrichment Procedure ◽  
Campylobacter Spp

The prevalence of Campylobacter jejuni and Campylobacter coli was investigated in 64 samples of fresh retail chicken purchased from commercial slaughterhouses located in Brazil. Campylobacter spp. were isolated from 40 (62.5%) of 64 analyzed samples. The strains biotyped according to Lior were classified as C. jejuni biotypes I and II, and C. coli biotypes I and II. The efficiency of different procedures for recovering Campylobacter spp. from chicken carcasses was tested. The enrichment procedure was significantly less effective than direct plating (P < 0.05), detecting 19 of 40 (47.5%) as opposed to 38 of 40 (95%) positive samples. Using direct plating the efficiency of Blaser's selective supplement was significantly more effective (P < 0.05) than Skirrow's selective supplement. To verify which factors could be affecting Campylobacter spp. growth in enrichment broth, the pH was measured after incubation for 48 h at 42°C and lactobacilli, coliforms, and enterococci were enumerated. Most of the Campylobacter-negative samples presented high levels of indicator microorganisms, which may have hindered the recovery of Campylobacter spp. during the enrichment procedure.

Enrichment Broth for the Detection of Campylobacter jejuni and Campylobacter coli in Fresh Produce and Poultry

2017 ◽  
Vol 80 (11) ◽  
pp. 1842-1850 ◽  
Author(s):  
Youmi Jo ◽  
Hye-Min Oh ◽  
Yohan Yoon ◽  
Sun-Young Lee ◽  
Ji-Hyoung Ha ◽  
...  
Keyword(s):  
Campylobacter Jejuni ◽  
Fresh Produce ◽  
International Organization ◽  
Natural Food ◽  
Campylobacter Coli ◽  
Standardization Method ◽  
Potassium Tellurite ◽  
Enrichment Broth ◽  
International Organization For Standardization ◽  
Campylobacter Spp

ABSTRACT Although campylobacteriosis caused by Campylobacter jejuni and Campylobacter coli has been increasingly reported worldwide owing to the consumption of contaminated poultry and fresh produce, the current detection protocols are not selective enough to inhibit unspecific microbes other than these pathogens. Five antibiotics were separately added to Bolton broth, and the survival rates of 18 Campylobacter spp. and 79 non-Campylobacter spp. were evaluated. The survival rate of the non-Campylobacter spp. was the lowest in Bolton broth with rifampin (6.3%), followed by cefsulodin (12.7%), novobiocin (16.5%), and potassium tellurite and sulfamethozaxole (both 17.7%). Also the most effective concentration of rifampin was found to be 12.5 mg/L, which markedly inhibited non-Campylobacter strains while not affecting the survival of Campylobacter strains. After the Campylobacter spp. were enriched in Bolton broth supplemented with 12.5 mg/L rifampin (R-Bolton broth), CampyFood Agar (CFA) was found to be better in selectively isolating the pathogens in the enrichment broth than the International Organization for Standardization method of using modified charcoal cefoperazone deoxycholate agar (mCCDA) for this step. When applied to natural food samples—here, romaine lettuce, pepper, cherry tomato, Korean leek, and chicken—the R-Bolton broth–CFA combination decreased the number of false-positive results by 50.0, 4.2, 20.8, 50.0, and 94.4%, respectively, compared with the International Organization for Standardization method (Bolton broth–mCCDA combination). These results demonstrate that the combination of R-Bolton broth and CFA is more efficient in detecting C. jejuni and C. coli in poultry and fresh produce and thus should replace the Bolton broth–mCCDA combination.

Prevalence of Campylobacter within a Swine Slaughter and Processing Facility†,‡

2003 ◽  
Vol 66 (9) ◽  
pp. 1550-1556 ◽  
Author(s):  
R. A. PEARCE ◽  
F. M. WALLACE ◽  
J. E. CALL ◽  
R. L. DUDLEY ◽  
A. OSER ◽  
...  
Keyword(s):  
Campylobacter Jejuni ◽  
Environmental Samples ◽  
Campylobacter Coli ◽  
Direct Plating ◽  
Predominant Species ◽  
Processing Facility ◽  
The Individual ◽  
Campylobacter Spp ◽  
Broth Enrichment ◽  
Recovery Methods

In this work, the occurrence of Campylobacter in a swine slaughter and processing facility was studied. Thirty composite carcass samples, representing 360 swine carcasses, were taken immediately after exsanguination, immediately after polishing, after the final wash, and after overnight chilling at 2°C. Thirty matching composite rectal samples were also taken immediately after exsanguination, and 60 nonmatching individual colon samples were collected from the same lot of swine during evisceration. Also, 72 environmental samples were collected from equipment used in the slaughter operation (42 samples) and the processing operation (30 samples). Campylobacter was isolated by direct plating on Campy-Line agar (CLA) or Campy-Cefex agar (CCA), as well as by Bolton broth enrichment and subsequent inoculation onto CLA or CCA. For all four recovery methods combined, Campylobacter was detected on 33% (10 of 30) of the composite carcasses immediately after exsanguination, 0% (0 of 30) after polishing, 7% (2 of 30) immediately before chilling, and 0% (0 of 30) after overnight chilling. The pathogen was recovered from 100% (30 of 30) of the composite rectal samples and 80% (48 of 60) of the individual colon samples. Campylobacter was detected in 4.8% (2 of 42) and 3.3% (1 of 30) of the slaughter and processing equipment samples, respectively. The recovery rate achieved with direct plating on CLA was significantly higher (P < 0.05) than those achieved with the other three recovery methods. For the 202 isolates recovered from all of the various samples tested, Campylobacter coli was the predominant species (75%) and was followed by Campylobacter spp. (24%) and Campylobacter jejuni (1%). These results indicate that although Campylobacter is highly prevalent in the intestinal tracts of swine arriving at the slaughter facility, this microorganism does not progress through the slaughtering operation and is not detectable on carcasses after overnight chilling.

Prevalence and Antimicrobial Resistance of Campylobacter spp. Isolated from Poultry Carcasses in Poland

2013 ◽  
Vol 76 (8) ◽  
pp. 1451-1455 ◽  
Author(s):  
KINGA WIECZOREK ◽  
IWONA KANIA ◽  
JACEK OSEK
Keyword(s):  
Campylobacter Jejuni ◽  
23S Rrna ◽  
Rrna Gene ◽  
Campylobacter Coli ◽  
Genetic Determinants ◽  
Gyra Gene ◽  
23S Rrna Gene ◽  
Pcr Method ◽  
Almost All ◽  
Campylobacter Spp

The purpose of the present study was to determine the prevalence of Campylobacter in poultry carcasses at slaughter in Poland. For the isolated strains, resistance to selected antibiotics and the associated genetic determinants were identified. A total of 498 Campylobacter isolates were obtained from 802 poultry samples during the 2-year study period. Strains were identified to species with the PCR method; 53.6% of the strains were Campylobacter jejuni and 46.4% were Campylobacter coli. A high percentage of the tested Campylobacter strains were resistant to ciprofloxacin and nalidixic acid (74.1 and 73.5%, respectively) followed by tetracycline (47.4%) and streptomycin (20.5%). Only one C. jejuni and two C. coli isolates were resistant to gentamicin. Seventy-nine (15.9%) of the 498 strains were resistant to three or more classes of antibiotics examined. Higher levels of resistance, irrespective of the antimicrobial agent tested, were found within the C. coli group. Almost all strains resistant to quinolones (99.5%) and to tetracycline (99.6%) carried the Thr-86-to-Ile mutation in the gyrA gene and possessed the tet(O) marker, respectively. All isolates resistant to erythromycin had the A2075G mutation in the 23S rRNA gene. These results reveal that poultry carcasses in Poland are a reservoir of potentially pathogenic and antimicrobial-resistant Campylobacter strains for humans, which may pose a public health risk.

Postchill Campylobacter Prevalence on Broiler Carcasses in Relation to Slaughter Group Colonization Level and Chilling System

2006 ◽  
Vol 69 (3) ◽  
pp. 495-499 ◽  
Author(s):  
M. LINDBLAD ◽  
I. HANSSON ◽  
I. VÅGSHOLM ◽  
R. LINDQVIST
Keyword(s):  
Campylobacter Jejuni ◽  
Surveillance Program ◽  
Intestinal Colonization ◽  
Direct Plating ◽  
Baseline Study ◽  
Group Data ◽  
Low Degree ◽  
High Degree ◽  
Campylobacter Spp ◽  
Skin Samples

Data from an ongoing national surveillance program of Campylobacter prevalence in broiler slaughter groups were related to results from a 1-year baseline study of broiler carcasses postchill. The goals were to establish the relation between Campylobacter prevalence in slaughter groups and on carcasses and to determine the effect of various chilling systems on Campylobacter prevalence. Pooled cloacal and neck skin samples from the surveillance program were analyzed after enrichment. Carcass rinse samples from the baseline study were analyzed after enrichment and by direct plating. Data from both studies were available for 614 carcasses. Direct-plating analyses indicated that the percentages of carcasses positive for Campylobacter jejuni and other Campylobacter spp. in slaughter groups with negative cloacal samples were 2 and 10%, respectively, whereas enrichment analyses indicated prevalences of 2% in both cases. Campylobacter prevalence in slaughter groups with a high degree of intestinal colonization (more than half of the pooled cloacal samples positive) was significantly higher than in slaughter groups with a low degree of colonization (76 to 85% and 30 to 50%, respectively, depending on Campylobacter spp. and analytical method). The prevalence of Campylobacter-positive carcasses postchill was at the same level as the prevalence of carcasses that originated from slaughter groups with positive neck skin samples at four of the six slaughterhouses. Only at one slaughterhouse, with an air-chilling system, was the postchill prevalence (13%) lower than that expected from slaughter group data (23%). The postchill prevalence (43%) was higher than that expected from slaughter group data (33%) at one slaughterhouse with immersion chilling.

scholarly journals Multiplex PCR Assay for Identifi cation and Differentiation of Campylobacter jejuni and Campylobacter coli Isolates

Folia Medica ◽  
2016 ◽  
Vol 58 (2) ◽  
pp. 95-100 ◽  
Author(s):  
Maria R. Pavlova ◽  
Elina G. Dobreva ◽  
Katucha I. Ivanova ◽  
Galina D. Asseva ◽  
Ivan N. Ivanov ◽  
...  
Keyword(s):  
Multiplex Pcr ◽  
Campylobacter Jejuni ◽  
Accurate Method ◽  
Clinical Isolates ◽  
Campylobacter Coli ◽  
Pcr Assay ◽  
Biochemical Tests ◽  
Multiplex Pcr Assay ◽  
Hydrolysis Of ◽  
Campylobacter Spp

AbstractIntroduction: Campylobacter spp. are important causative agents of gastrointestinal infections in humans. The most frequently isolated strains of this bacterial genus are Campylobacter jejuni and Campylobacter coli. To date, genetic methods for bacterial identification have not been used in Bulgaria. We optimized the multiplex PSR assay to identify Campylobacter spp. and differentiate C. jejuni from C. coli in clinical isolates. We also compared this method with the routinely used biochemical methods.Aim: To identify Campylobacter spp. and discriminate C. coli from C. jejuni in clinical isolates using multiplex PCR assay.Materials and methods: Between February 2014 and January 2015 we studied 93 stool samples taken from patients with diarrheal syndrome and identified 40 species of Campylobacter spp. in them. The clinical material was cultured in microaerophilic atmosphere, the isolated strains being biochemically diff erentiated (hydrolysis of sodium hippurate for C. jejuni, and hydrolysis of indoxyl acetate for C. coli). DNA was isolated from the strains using QiaAmp MiniKit (QIAGEN, Germany). Twenty strains were tested with multiplex PCR for the presence of these genes: cadF, characteristic for Campylobacter spp., hipO for C. jejuni and asp for C. coli.Results and discussion: The biochemical tests identified 16 strains of C. jejuni, 3 strains of C. coli, and 1 strain of C. upsaliensis. After the multiplex PCR assay the capillary gel electrophoresis confirmed 16 strains of C. jejuni, 2 strains of C. coli and 2 strains of Campylobacter spp. - because of the presence of the gene cadF. C. jejuni has the gene hipO, and it is possible that this gene may not be expressed in the biochemical differentiation yielding a negative reaction as a result. In comparison, we can conclude that the genetic differentiation is a more accurate method than the biochemical tests.Conclusion: The multiplex PCR assay is a fast, accurate method for identifi cation of Campylobacter spp. which makes it quite necessary in the clinical diagnostic practice.

Detection of Campylobacter jejuni and Campylobacter coli from Broiler Chicken–Related Samples Using BAX PCR and Conventional International Organization for Standardization Culture

2008 ◽  
Vol 71 (4) ◽  
pp. 835-838 ◽  
Author(s):  
LISA K. WILLIAMS ◽  
ALISDAIR MCMEECHAN ◽  
TAMSIN BAALHAM ◽  
LAURA WARD ◽  
TOM J. HUMPHREY ◽  
...  
Keyword(s):  
Campylobacter Jejuni ◽  
Broiler Chicken ◽  
Culture Method ◽  
International Organization ◽  
Campylobacter Coli ◽  
Poultry Production ◽  
Production Chain ◽  
Enrichment Cultures ◽  
International Organization For Standardization ◽  
Campylobacter Spp

In this study, the conventional International Organization for Standardization (ISO) culture method was compared with the DuPont Qualicon BAX system, a high-throughput, rapid molecular assay that can be used to detect several bacterial species, including Campylobacter jejuni and Campylobacter coli in diverse sample types. Standard enrichment culture is a time-consuming process, taking up to 6 days to obtain a confirmed result. Rapid molecular assays have been developed that provide results within 24 h. Naturally contaminated samples from the poultry production chain were examined for the presence of Campylobacter spp. Samples from broiler chicken ceca (n = 100), fresh chicken carcass rinses (n = 60), and bootsocks (gauze sock walked through a broiler chicken house; n = 50) were enriched according to the ISO 10272 method in Bolton broth specifically designed to detect Campylobacter spp. in complex sample types. Samples were enriched without blood for use with the BAX system using the Campylobacter BAX kits for the detection of C. jejuni and C. coli. Samples also were directly plated onto modified charcoal cefperazone deoxycholate agar, and results were compared with those from the enriched samples for the ability to detect Campylobacter spp. Campylobacter spp. were isolated from 49% of samples with conventional enrichment cultures, from 48% with direct culture, from 68% with the BAX system and enrichment cultures, and from 62% with the BAX system used directly with samples. Overall, the BAX system detected more positive samples than did the conventional culture method and is an effective methodology for the rapid and reliable detection of Campylobacter spp. from diverse sample types.

Incidence of Campylobacter jejuni/coli on Pork Carcasses in the Northeast Georgia Area

1985 ◽  
Vol 48 (9) ◽  
pp. 808-810 ◽  
Author(s):  
A. J. BRACEWELL ◽  
J. O. REAGAN ◽  
J. A. CARPENTER ◽  
L. C. BLANKENSHIP
Keyword(s):  
Campylobacter Jejuni ◽  
Campylobacter Coli ◽  
Skin Area ◽  
Isolation Rate ◽  
Direct Plating ◽  
Isolation Methods ◽  
Before And After

One hundred and twelve freshly slaughtered pork carcasses from three packing plants were sampled before and after chilling for the presence of Campylobacter jejuni/coli by the use of two isolation methods (Preston enrichment and Skirrow direct plating). Preston enrichment media gave the highest isolation rate, 12.5%, on freshly slaughtered carcasses. No isolations were obtained from chilled carcasses. More isolates were obtained from the ham skin area compared with the jowl area. All isolates were confirmed as Campylobacter coli.

Evaluation of TECRA Broth, Bolton Broth, and Direct Plating for Recovery of Campylobacter spp. from Broiler Carcass Rinsates from Commercial Processing Plants†

2009 ◽  
Vol 72 (5) ◽  
pp. 972-977 ◽  
Author(s):  
L. J. RICHARDSON ◽  
N. A. COX ◽  
J. S. BAILEY ◽  
M. E. BERRANG ◽  
J. M. COX ◽  
...  
Keyword(s):  
The United States ◽  
Culture Broth ◽  
Enrichment Broth ◽  
Direct Plating ◽  
Conventional Culture ◽  
Chicken Carcass ◽  
Processing Plants ◽  
Commercial Processing ◽  
Laboratory Procedures ◽  
Campylobacter Spp

The purpose of this study was to compare a conventional culture broth method (Bolton enrichment), a newly developed proprietary broth method (TECRA Campylobacter enrichment), and direct plating for recovery of Campylobacter spp. from chicken carcass rinsates. Whole carcass rinses were taken from 140 carcasses at rehang (immediately after defeathering but before evisceration) and from 140 carcasses at postchill from eight different processing plants in the United States. The rinsate samples were packed in ice and shipped overnight to the laboratory. Aliquots of the rinsate were transferred into Bolton and TECRA enrichment broths and were direct plated. Standard laboratory procedures with Campy-cefex plates were followed for recovery of Campylobacter spp. For rehang carcasses, 94% were positive for Campylobacter spp. with the TECRA enrichment broth and 74% were positive with the Bolton enrichment broth. For postchill carcasses, 74% were positive for Campylobacter spp. with the TECRA enrichment broth and 71% were positive with the Bolton enrichment broth. Compared with the Bolton enrichment broth, TECRA enrichment broth significantly suppressed non-Campylobacter microflora (P < 0.05). Overall, TECRA enrichment broth yielded an 11% higher total number of Campylobacter-positive samples compared with the Bolton enrichment broth. Campylobacter spp. detection in postchill samples was significantly greater (P < 0.05) by enrichment (84%) than by direct plating (19%). The high number of Campylobacter-positive samples obtained with all procedures indicated that 99% of the carcass rinsates obtained at rehang and 84% obtained at postchill contained Campylobacter spp.

Passage of Campylobacter jejuni and Campylobacter coli Subtypes through 0.45- and 0.65-Micrometer-Pore-Size Nitrocellulose Filters

2017 ◽  
Vol 80 (12) ◽  
pp. 2029-2032 ◽  
Author(s):  
Mark E. Berrang ◽  
Richard J. Meinersmann ◽  
Nelson A. Cox
Keyword(s):  
Pore Size ◽  
Campylobacter Jejuni ◽  
Soft Agar ◽  
Filter Method ◽  
Campylobacter Coli ◽  
Direct Plating ◽  
Complex Samples ◽  
Liquid Cultures ◽  
Cellular Density ◽  
The Mean

ABSTRACT Campylobacter can be difficult to recover from complex samples due to overgrowth by background bacteria. A 0.45- or 0.65-μm-pore-size filter overlaid on agar plates can be used as a means to separate Campylobacter from confounding non-Campylobacter cells, facilitating detection on solid plating media. It is unclear what percentage of cells in a Campylobacter suspension passes through a filter and results in visible colonies. The objective of this study was to compare the number of Campylobacter cells detected by the filter method with those detected by direct plating and determine if the filter method can be used to estimate cellular density of an unknown Campylobacter in suspension. Overnight liquid cultures of six subtypes of Campylobacter jejuni and six of Campylobacter coli, all originally detected in chicken samples, were used for this study. Motility of isolates was tested by using a stab into soft agar, incubating plates, and measuring colony size. Each subtype was applied to Campy-Cefex agar directly and through a 0.45- or 0.65-μm-pore-size filter. Filters were removed, plates were incubated, and colonies were counted; three replications were conducted. Mean recovery by direct plating was 8.3 log CFU/mL. Regardless of pore size, the overall mean number of Campylobacter detected by using the filter method was significantly less than that using direct plating (P < 0.05). The mean difference between direct plating and plating though a 0.65-μm-pore-size filter for motile Campylobacter was log 2.4 CFU/mL, with a 95% confidence interval of ±0.2 log CFU/mL.

Prevalence of putative virulence markers inCampylobacter jejuniandCampylobacter coliisolated from hospitalized children, raw chicken, and raw beef in Tehran, Iran

2011 ◽  
Vol 57 (2) ◽  
pp. 143-148 ◽  
Author(s):  
Mohammad Hamidian ◽  
Maryam Sanaei ◽  
Mehdi Bolfion ◽  
Hossein Dabiri ◽  
Mohammad-Reza Zali ◽  
...  
Keyword(s):  
Campylobacter Jejuni ◽  
Rrna Gene ◽  
Campylobacter Coli ◽  
Hospitalized Children ◽  
Unequal Distribution ◽  
Exact Test ◽  
Virulence Markers ◽  
Dnaj Gene ◽  
Different Origins ◽  
Campylobacter Spp

The incidence of the virulence-associated genes cdtA, cdtB, cdtC, cadF, dnaJ, racR, and pldA has been investigated in Campylobacter jejuni and Campylobacter coli collected from raw chicken and beef from retailers in Tehran, Iran, and from hospitalized children (age, ≤14 years) suffering from diarrhea. Campylobacter spp. were collectively identified by morphological and biochemical methods. Campylobacter jejuni and C. coli were discriminated from other Campylobacter spp. by amplification of a specific conserved fragment of the 16S rRNA gene. The distinction between C. jejuni and C. coli was subsequently made by molecular determination of the presence of the hipO gene in C. jejuni or the ask gene in C. coli. Fragments of the studied virulence-associated genes, cdtA, cdtB, cdtC, cadF, racR, dnaJ, and pldA, were amplified by PCR and subjected to horizontal gel electrophoresis. A total of 71 isolates of C. jejuni and 24 isolates of C. coli from meat were analyzed, while the numbers of isolates from the hospitalized children were 28 and 9, respectively. The unequal distribution of C. jejuni and C. coli in the samples has also been reported in other studies. Statistical analyses by the use of the two-tailed Fisher’s exact test of the occurrence of the virulence genes in the isolates of different origins showed that the occurrence of the dnaJ gene was consistently significantly higher in all C. jejuni isolates than in C. coli. The occurrence of the other virulence markers did not differ significantly between species in the majority of the isolates. The PCR results also showed that the occurrence of the virulence markers in the analyzed isolates was much lower than in other studies, which may be caused by a divergent genomic pool of our isolates in comparison with others.

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