Efficacy of media and methods for detecting and enumerating Campylobacter jejuni in refrigerated chicken meat.

A study was undertaken to compare several enrichment and direct isolation media for their suitability to detect and enumerate five strains of Campylobacter jejuni in refrigerated (5 degrees C) chicken meat. The influence of CO2 on survival at 5 degrees C was also investigated. Selective enrichment media evaluated included Preston broth (PB), selective semisolid brucella medium (SSBM), Campylobacter enrichment broth (CEB), VTP brucella-FBP broth (VTP), Rosef and Kapperud Campylobacter enrichment broth (RKCEB), and Doyle and Roman enrichment broth (DREB). Direct isolation agars included Campy brucella agar (CBAP), blood-free Campylobacter medium (BFCM) and modified Butzler agar (MBA). Comminuted chicken meat was inoculated with C. jejuni, sealed under atmospheric gas or CO2, and stored at 5 degrees C for up to 21 days. Viable population was determined by the most-probable-number technique (PB, SSBM, CEB, VTP, and RKCEB, followed by plating on CBAP, BFCM, and MBA), enrichment on DREB, followed by plating on CBAP, BFCM, and MBA, and direct isolation on CBAP, BFCM, and MBA. Without exception, direct plating of samples was superior to the most-probable-number technique for enumerating C. jejuni; MBA was inferior to CBAP and BFCM, and DREB performed at least as well as other enrichment media evaluated. Carbon dioxide afforded protection against death of three of the five strains of C. jejuni tested.     

Efficacy of some methods and media for detecting and enumerating Campylobacter jejuni in frozen chicken meat

Four of five strains of Campylobacter jejuni survived in chicken meat stored at -18 degrees C for 12 months. Direct plating of samples was superior to the most probable number technique for enumerating C. jejuni. Enrichment culture using the Doyle & Roman enrichment method resulted in the highest rates of detection. Packaging under an atmosphere of CO2 did not substantially influence survival.     

Efficacy of some methods and media for detecting and enumerating Campylobacter jejuni in frozen chicken meat

Four of five strains of Campylobacter jejuni survived in chicken meat stored at — 18C for 12 months. Direct plating of samples was superior to the most probable number technique for enumerating C. jejuni. Enrichment culture using the Doyle & Roman enrichment method resulted in the highest rates of detection. Packaging under an atmosphere of CO₂ did not substantially influence survival.     

Comparison of media and methods for detecting and enumerating Listeria monocytogenes in refrigerated cabbage

Direct plating, selective enrichment, and cold enrichment followed by secondary selective enrichment procedures were compared for detecting and enumerating Listeria monocytogenes in chopped cabbage stored at 5 degrees C for up to 64 days. Addition of Fe3+ to solid media enhanced detection of the organism. Cold enrichment (5 degrees C) in nutrient broth and brain heart infusion broth followed by secondary enrichment (48 h, 30 degrees C) in Trypticase soy-yeast extract-antibiotic broth and thiocyanate-nalidixic acid broth and plating on selective agar media (Doyle and Schoeni selective enrichment agar [minus acriflavin hydrochloride, supplemented with 5 micrograms of Fe3+/ml] and McBride Listeria agar) resulted in the detection of highest populations.     

Comparison of media and methods for detecting and enumerating Listeria monocytogenes in refrigerated cabbage.

Direct plating, selective enrichment, and cold enrichment followed by secondary selective enrichment procedures were compared for detecting and enumerating Listeria monocytogenes in chopped cabbage stored at 5 degrees C for up to 64 days. Addition of Fe3+ to solid media enhanced detection of the organism. Cold enrichment (5 degrees C) in nutrient broth and brain heart infusion broth followed by secondary enrichment (48 h, 30 degrees C) in Trypticase soy-yeast extract-antibiotic broth and thiocyanate-nalidixic acid broth and plating on selective agar media (Doyle and Schoeni selective enrichment agar [minus acriflavin hydrochloride, supplemented with 5 micrograms of Fe3+/ml] and McBride Listeria agar) resulted in the detection of highest populations.     

Survival of Campylobacter jejuni in chicken meat at frozen storage temperatures

The aim of this study was to determine the survival of Campylobacter jejuni in chicken meat samples at frozen temperatures and given length of incubation and to determine the impact of aerobic bacteria on the survival of C. jejuni. The chicken meat samples were inoculated with C. jejuni NCTC 11351 suspensions and stored in bags at temperatures of -20°C and -70°C. The mean value of C. jejuni from meat samples decreased from 7.52 log10 CFU/g after 30 minutes of incubation at ambient temperature, to 3.87 log10 CFU/g on the eighth week of incubation at -20°C, and to 3.78 log10 CFU/g at incubation at -70°C after the same incubation period. Both freezing temperatures, -20°C and -70°C, decreased the number of campylobacters. The presence of aerobic mesophilic bacteria did not influence the survival of C. jejuni in chicken meet samples. Keeping poultry meat at freezing temperatures is important for the reduction of C. jejuni, which has a strong influence on the prevention of occurrence of campylobacteriosis in humans.     

Campylobacter jejuni survival in chicken meat as a function of temperature.

Recognition of Campylobacter fetus subsp. jejuni (referred to hereafter as C. jejuni) as an important human pathogen and its isolation from meat products indicate the need for knowledge of its survival characteristics in meats. Thermal death times (D-values) for a single strain and a five-strain composite were determined in 1% peptone and autoclaved ground chicken meat at temperatures ranging from 49 to 57 degrees C. Survival was determined for these strains in chicken meat at 4, 23, 37, and 43 degrees C. Survival was also determined on raw chicken drumsticks stored at 4 degrees C in either an ambient or a CO2 atmosphere. D-values were greater in chicken meat than in peptone in all cases. D-values in peptone for strain H-840 at 49, 51, 53, 55, and 57 degrees C were 15.2, 4.90, 1.71, 0,64, and 0.25 min, respectively. The corresponding D-values in ground chicken meat were 20.5, 8.77, 4.85, 2.12, and 0.79 min, respectively. Similar results were obtained with a composite of five strains. When sterile ground chicken meat was inoculated with approximately 10(6) to 10(7) C. jejuni cells per g and stored at 37 degrees C in an ambient atmosphere, a 1-to 2-log count increase occurred during the first 4 days, followed by a gradual decline of about 1 log during the remainder of the 17-day storage period. No growth was observed among similarly inoculated samples that were stored at 4, 23, and 43 degrees C but counts declined by about 1 to 2 logs at 4 degrees C (17 day), by 2.5 to 5 logs at 23 degrees C (17 days), and to undetectable levels at 43 degrees C (between 10 and 16 days). Survival on raw chicken drumsticks stored at 4 degrees C in CO2 and in an ambient atmosphere declined by about 1.5 and 2.0 logs, respectively, during 21 days of storage. The effect of temperature on the survival of C. jejuni in chicken meat was similar to that reported in other natural and laboratory milieus. Ordinary cooking procedures that destroy salmonellae would be expected to destroy C. jejuni.     

Prevalence of Campylobacter jejuni in chicken wings

Campylobacter jejuni was found in 82.9% of 94 chicken wing packages analyzed on the day of arrival at supermarkets and in 15.5% of 45 packages obtained from the supermarket shelves a few days later. The number of bacterial cells ranged from 10(2) to 10(3.9) per wing. The prevalence of C. jejuni in the wings varied with the brand, the day of sampling, and the age of the product.     

Efficacy of filter types for detecting Campylobacter jejuni and Campylobacter coli in environmental water samples by polymerase chain reaction.

A previously developed polymerase chain reaction (PCR) amplification of a target region in the flaA Campylobacter flagellin gene was evaluated and adapted for use with environmental water samples. The ability to detect Campylobacter jejuni or Campylobacter coli in seeded water samples was tested with various filters after concentration and freeze-thaw lysis of the bacterial cells. A nonradioactive probe for the amplified flagellin gene fragment detected as little as 1 to 10 fg of genomic DNA and as few as 10 to 100 viable C. jejuni cells per 100 ml of water filtered onto Fluoropore (Millipore Corp.) filters. No amplification was obtained with cellulose acetate filters, most likely because of binding of the DNA to the filter. Concentration and lysis of target cells on Fluoropore and Durapore (Millipore Corp.) filters allowed PCR to be performed in the same reaction tube without removing the filters. This methodology was then adapted for use with environmental water samples. The water supply to a broiler chicken production farm was suspected as the source of C. jejuni known to be endemic in grow-out flocks at the farm, despite the inability to culture the organisms by standard methods. The filtration-PCR method detected Campylobacter DNA in more than half of the farm water samples examined. Amplified campylobacter DNA was not detected in small volumes of regional surface water samples collected on a single occasion in February. The filtration-PCR amplification method provided a basis for detection of C. jejuni and C. coli in environmental waters with a high degree of specificity and sensitivity.     

肉及肉制品中空肠弯曲菌的污染情况调查

我们对辽宁地区肉及肉制品进行弯曲杆菌属检查,发现12.9%(23/177)的样品为弯曲杆菌属阳性.经API Campy鉴定系统鉴定,其中26.1%(6/23)为空肠弯曲菌(C.jejuni)阳性,同时发现1株大肠弯曲菌.该调查证实,辽宁地区进出口的肉及肉制品中,存在空肠弯曲菌的污染,如果销售不当或再加工卫生不良,会对消费者的健康构成潜在威胁.     

Genotyping of clinical and chicken isolates of Campylobacter jejuni and Campylobacter coli

Genomic DNA from 58 strains of Campylobacter made up of 48 Campylobacter jejuni and ten Campylobacter coli were digested with Sma I and analysed by pulsed-field gel electrophoresis (PFGE). The cleavage of DNA by Sma I gave 22 distinct hybridization patterns. The two Campylobacter species were subtyped by PFGE. The average genomic size for C. jejuni by Sma I digestion was 1.73 Mb, while that of C. coli gave 1.7 Mb. Results from this study indicate that PFGE analysis by Sma I digested genomic DNA provides a reliable means of differentiating between and within species of Campylobacter and provides a practical approach to epidemiological studies of Campylobacter.     

Evaluation of methods for the enumeration of Campylobacter jejuni and Campylobacter coli bacteriophages

The Spiral Plater System used for enumeration of bacteria was evaluated for the titration of Carnpylobacter phages. Twelve phages of C. jejuni were titrated using the conventional surface droplet technique, soft agar overlay-pour plate technique, and the Spiral Plater System. Phage counts obtained by the three methods were similar but the Spiral Plater System showed greater precision than the other methods.     

Efficacy of agar media for enumerating two Saccharomyces species in sucrose syrups

Experiments were carried out to determine the suitability of acidified and antibiotic-supplemented agars for supporting colony formation by cells ofSaccharomyces cerevisiae andSaccharomyces rouxii. Yeasts had been suspended in sucrose syrups buffered at pH 3.0, 5.0 and 7.0, and stored at 4 and 21°C for periods of time ranging to 10 weeks. TheSaccharomyces species were recovered in equal numbers, regardless of the type of enumeration agar. Sucrose, at concentrations up to 60%, protected cells against inactivation during storage, and survival was greater at 21° than at 4°C.     

Efficacy of agar media for enumerating two Saccharomyces species in sucrose syrups

Experiments were carried out to determine the suitability of acidified and antibiotic-supplemented agars for supporting colony formation by cells ofSaccharomyces cerevisiae andSaccharomyces rouxii. Yeasts had been suspended in sucrose syrups buffered at pH 3.0, 5.0 and 7.0, and stored at 4 and 21°C for periods of time ranging to 10 weeks. TheSaccharomyces species were recovered in equal numbers, regardless of the type of enumeration agar. Sucrose, at concentrations up to 60%, protected cells against inactivation during storage, and survival was greater at 21° than at 4°C.     

Comparison of methods for isolating Campylobacter jejuni from raw milk

The method of Doyle and Roman (Appl. Environ. Microbiol. 43:1343-1353, 1982) was compared with that of Lovett et al. (Appl. Environ. Microbiol. 46:459-462, 1983) for the ability to recover Campylobacter jejuni strains inoculated into raw milk at a concentration of less than 1 cell per g. The method of Lovett et al. gave significantly greater recovery proportions.     

Comparison of 2-day-old and 14-day-old chicken colonization models for Campylobacter jejuni

In this study, we compared two types of chicken infection models for Campylobacter jejuni in terms of infectious dose required to colonize the chickens and the susceptibility of chickens of different ages to persistent colonization by C. jejuni. In one model, chickens at day 2 posthatching were used, and in the other, 14-day-old chickens were used. The minimum C. jejuni cell number required to colonize 14-day-old chickens was 5 x 10(4) cells, and that for 2-day-old chickens was 5 x 10(3). The ability of various C. jejuni strains to colonize the chicken gastrointestinal tract was the same in both models.     

Survival of cold-stressed Campylobacter jejuni on ground chicken and chicken skin during frozen storage

Campylobacter jejuni is prevalent in poultry, but the effect of combined refrigerated and frozen storage on its survival, conditions relevant to poultry processing and storage, has not been evaluated. Therefore, the effects of refrigeration at 4 degrees C, freezing at -20 degrees C, and a combination of refrigeration and freezing on the survival of C. jejuni in ground chicken and on chicken skin were examined. Samples were enumerated using tryptic soy agar containing sheep's blood and modified cefoperazone charcoal deoxycholate agar. Refrigerated storage alone for 3 to 7 days produced a reduction in cell counts of 0.34 to 0.81 log10 CFU/g in ground chicken and a reduction in cell counts of 0.31 to 0.63 log10 CFU/g on chicken skin. Declines were comparable for each sample type using either plating medium. Frozen storage, alone and with prerefrigeration, produced a reduction in cell counts of 0.56 to 1.57 log10 CFU/g in ground chicken and a reduction in cell counts of 1.38 to 3.39 log10 CFU/g on chicken skin over a 2-week period. The recovery of C. jejuni following freezing was similar on both plating media. The survival following frozen storage was greater in ground chicken than on chicken skin with or without prerefrigeration. Cell counts after freezing were lower on chicken skin samples that had been prerefrigerated for 7 days than in those that had been prerefrigerated for 0, 1, or 3 days. This was not observed for ground chicken samples, possibly due to their composition. C. jejuni survived storage at 4 and -20 degrees C with either sample type. This study indicates that, individually or in combination, refrigeration and freezing are not a substitute for safe handling and proper cooking of poultry.