mCCDA Agar Research Articles

Campylobacter magnus sp. nov., isolated from caecal contents of domestic pigs (Sus scrofa domesticus).

During the 2021 European Food Safety Authority coordinated harmonized monitoring of antimicrobial resistance in Campylobacter species in Slovenia, five Campylobacter-like strains were cultured from caeca of a total of 104 domestic pigs that could not be identified using the standard-prescribed biochemical tests or MALDI-TOF MS. The isolates were obtained using the standard ISO 10272 procedure for the isolation of thermotolerant Campylobacter with prolonged cultivation time. Small Campylobacter-like colonies were observed on mCCDA and CASA agar plates after 2-4 days of incubation; dark-field microscopy revealed relatively big spirilli-shaped bacteria exhibiting characteristic Campylobacter-like motility. The cells were 1.5-3 µm long and 0.5-0.7 µm wide, Gram-negative, oxidase-positive and catalase-positive. MALDI-TOF mass spectra were distinctive and consistent, but with low MALDI-TOF MS log scores and the closest matches being those of Campylobacter hyointestinalis and Campylobacter fetus. All five strains underwent whole-genome sequencing. Analysis of 16S rRNA gene sequences revealed that the isolates were most similar (98.3-98.4 % identity) to Campylobacter lanienae. Pairwise average nucleotide identity (ANI) values revealed that the five studied strains shared pairwise ANI of 96.2-96.5 % but were clearly distinct from the previously described Campylobacter species (ANI ≤72.8 %). The core genome-based phylogeny confirmed that the new strains form a distinct and well-supported clade within the genus Campylobacter. The conducted polyphasic taxonomic analysis confirmed that the five strains represent a novel Campylobacter species for which the name Campylobacter magnus sp. nov. is suggested, with strain 46386T (=DSM 115534T=CCUG 76865T) as the type strain.

P18 Antimicrobial resistance patterns of Campylobacter species isolated from poultry in Mathira, Nyeri County, Kenya

Background Campylobacter species are zoonotic pathogens recognized as a cause of gastroenteritis in the human population and pose a major public health threat worldwide. The most important in the genus is Campylobacter jejuni, which causes diarrhoea in patients, mainly children under the age of 1 year, young adults and immunosuppressed persons. Campylobacter occurs in the gut of most animals as normal flora and is transferred to foods from the faeces during slaughter. It is acquired through consumption of undercooked poultry and other meat as well as water and unpasteurized milk. Fruits and vegetables can be contaminated from water containing faeces from birds and other animals. Campylobacter can also be transmitted through contact with dogs and cats’ faeces as well as drinking contaminated water. Campylobacter infections are common in low- and middle-income countries as well as developed countries.ObjectivesTo assess the prevalence and determine the antimicrobial resistance patterns of Campylobacter spp. from poultry in Mathira Sub-County, Nyeri County.MethodsA total of 380 cloacal swabs were collected randomly form 57 farmers in Mathira, Kenya. They were transported in sterile Amies charcoal swabs under cold chain and stored at 4°C. They were enriched in Preston broth and incubated for 24 h at 42°C, after which they were streaked onto mCCDA agar and incubated for 24 h at 42°C under microaerobic conditions generated by Campy Gen™ packs. Typical grey moist, swarming and discrete colonies were identified using MALDI-TOF. Antimicrobial susceptibility testing was done on ampicillin, tetracycline, ciprofloxacin, gentamicin, erythromycin and nalidixic acid by the disc diffusion method and disc diameters were measured and interpreted using the CLSI guidelines.ResultsIn total, 272 out of 380 (71.32%) isolates were Campylobacter; 190 (50%) were C. jejuni while 81 (22) were Campylobacter coli. Resistance to ampicillin was 40% for C. coli and 30.53% for C. jejuni. Overall, 53.75% of C. coli isolates and 51.85% of C. jejuni isolates were resistant to tetracycline. We found 67.50% of C. coli were resistant to ciprofloxacin as compared with 38.42% of C. jejuni. Resistance to erythromycin for C. coli and C. jejuni was 10% and 16.935%, respectively. Finally, 65% of C. coli was resistant to nalidixic acid as compared with 38.42% of C. jejuni.ConclusionsThe study found that Campylobacter spp. show high levels of resistance to tetracyclines, ciprofloxacin and nalidixic acid and are MDR. Resistance to gentamicin and erythromycin is markedly low and these antibiotics can be reserved for treatment. Surveillance activities for AMR should be enhanced and laboratory capacity increased to support surveillance and diagnosis.

A Rapid Culture Method for the Detection of Campylobacter from Water Environments.

The natural environment and water are among the sources of Campylobacter jejuni and Campylobacter coli. A limited number of protocols exist for the isolation of campylobacters in poorly filterable water. Therefore, the goal of our work was to find a more efficient method of Campylobacter isolation and detection from wastewater and surface water than the ISO standard. In the novel rapid culture method presented here, samples are centrifuged at high speed, and the resuspended pellet is inoculated on a filter, which is placed on Campylobacter selective mCCDA agar. The motile bacteria pass through the filter pores, and mCCDA agar suppresses the growth of background microbiota on behalf of campylobacters. This culture-based method is more efficient for the detection and isolation of Campylobacter jejuni and Campylobacter coli from poorly filterable water than the ISO 17995 standard. It also is less time-consuming, taking only 72 h and comprising three steps, while the ISO standard method requires five or six steps and 144–192 h. This novel culture method, based on high-speed centrifugation, bacterial motility, and selective cultivation conditions, can be used for the detection and isolation of various bacteria from water samples.

Detection of Campylobacter sp. from Poultry Feces in Ouagadougou, Burkina Faso

Background: Campylobacter contamination in poultry and poultry product has been reported worldwide. The present study aims to determine the prevalence of Campylobacter in poultry feces using selective enrichment Bolton broth and multiplex PCR. Method: Two methods were used in this study: the first was direct plating of poultry feces into mCCDA agar plates. The second, three antibiotics were used at different concentrations to add in Bolton broth supplemented. These antibiotics were Rifampicin (Oxoid, Nepean, Ontario) with 10 mg/L, colistin (Oxoid, Nepean, Ontario) with 1 mg/mL and 2 mg/mL; trimethoprim (Oxoid, Nepean, Ontario) with 10 mg/L. The colonies with typical campylobacter morphology on blood agar (little, red and ring colonies) were further identified to the species level by multiplex polymerase chain reaction (PCR). Results: The addition of colistin (2 mg/mL) to the Bolton broth with selective supplements enhanced the selective isolation of Campylobacter strains. Out of the 52 feces samples, 18 (34.61%) were positive for campylobacter and direct plating on mCCDA 11 (21.15%) campylobacter strains (p campylobacter strains detected belonged to Campylobacter coli and 1 (5.55%) strain to Campylobacter jejuni. Conclusion: Although it is known to be difficult to isolate Campylobacter from animal feces samples, this study shows that antibiotic selective pressure improves the isolation efficiency of Campylobacter from poultry feces.

Acinetobacter baumannii detected on modified charcoal-cefoperazone-deoxycholate agar in a waste stabilization pond.

Campylobacter is a recommended reference pathogen for the verification and validation of water recycling schemes in Australia and globally. In a larger study investigating the efficacy of pathogen removal in waste stabilization ponds (WSP), we cultivated bacteria from wastewater samples on modified charcoal-cefoperazone-deoxycholate agar (mCCDA) targeting the growth of Campylobacter. A high number of colonies characteristic of Campylobacter grew on this selective medium, but this did not correlate with qPCR data. Using primers targeting the 16S rRNA gene, and additional confirmatory tests to detect VS1, ompA, blaOXA-51-like, blaOXA-23-like genes, we tested 80 random colonies from 10 WSP samples. All 80 were identified as Acinetobacter baumannii. Wastewater grab samples taken three times over 6 months throughout the WSP system showed removal of A. baumannii in the WSP at rates similar to that of Escherichia coli. Our study suggests that mCCDA agar is not a suitable medium for isolating Campylobacter from environmental samples and that A. baumannii can be used as an indicator for removal of pathogens in WSPs.

The Addition of ViriditecTM Aqueous Ozone to Peracetic Acid as an Antimicrobial Spray Increases Air Quality While Maintaining Salmonella Typhimurium, Non-pathogenic Escherichia coli, and Campylobacter jejuni Reduction on Whole Carcasses.

Currently, the most utilized antimicrobial in processing facilities is peracetic acid, PAA; however, this chemical is increasingly recognized as a hazard to human health. Preliminary evidence suggests that ozone, when introduced in a specific manner, can reduce the noxious nature of PAA. Therefore, the objective of the current study was to evaluate the efficacy of TetraClean Systems aqueous ozone, O3, in combination with PAA as an antimicrobial spray on whole chicken carcasses. This trial used 70 whole hen carcasses (7 treatments; 10 replications) that were inoculated in a 400 mL cocktail containing Salmonella, Escherichia coli, and Campylobacter (107 CFU/mL) and allowed to adhere for 60 min at 4°C for a final concentration of 105 to 106 CFU/g. The experimental 5 s (4×) spray treatments included: a no treatment negative control, TW; TW + O3 (10 ppm), TW + PAA (50 ppm), TW + PAA (500 ppm), TW + O3 + PAA (50 ppm), and TW + O3 + PAA (500 ppm). During treatment application, ambient PAA vapor was measured with a ChemDAQ Safecide PAA vapor sensor. After treatment, carcasses were immediately rinsed in 400 mL of nBPW for 2 min. Following rinsing, the dot method was utilizing for enumeration with 10 μL of rinsate being serially diluted, plated on XLD and mCCDA agar, and incubated aerobically at 37°C for 24 h or microaerophilically at 42°C for 48 h. Log-transformed counts were analyzed using ANOVA in JMP 14.0. Means were separated using Tukey’s HSD when P ≤ 0.05. There was a significant treatment effect among Salmonella, E. coli, and Campylobacter counts, and a significant treatment effect among ambient PAA (P < 0.05). TW + O3 + PAA (500 ppm), reduced Salmonella significantly compared to TW (5.71 and 6.30 log CFU/g). Furthermore, TW + PAA (500 ppm), reduced the presence of E. coli significantly compared to TW or no treated control (5.57 and 6.18 log CFU/g). Also, TW + PAA (50 ppm), TW + PAA (500 ppm), and TW + O3 + PAA (500 ppm) significantly reduced Campylobacter compared to carcasses not treated (4.80, 4.81, and 4.86 log CFU/g). Lastly, the addition of ozone significantly reduced the ambient PAA when O3 was added to 500 ppm of PAA, as TW + O3 + PAA (500 ppm) produced less ambient PAA than TW + PAA (500 ppm) (0.052 and 0.565 ppm). In conclusion, the addition of ozone to PAA may demonstrated the ability to effectively reduce ambient PAA, thus increasing employee safety.

EFICIÊNCIA DE DIFERENTES PROTOCOLOS PARA ISOLAMENTO DE Campylobacter jejuni DE CARNE DE FRANGO

Resumo Campylobacter jejuni é o principal causador de gastroenterite bacteriana aguda e a carne de frango é um importante veículo do agente. Entretanto, as metodologias convencionais de isolamento de Campylobacter muitas vezes não são eficientes, podendo levar a resultados errôneos. Sendo assim, este trabalho teve como objetivo avaliar diferentes métodos utilizados na detecção de C. jejuni em produtos de frango. Carne moída experimentalmente contaminada com três diferentes diluições do microrganismo foi analisada com diferentes protocolos para isolamento de C. jejuni. Foram feitas semeaduras diretamente nos ágares mCCDA, Columbia e ágar sangue, e após pré-enriquecimento nos caldos Bolton ou Brucella. As colônias características de Campylobacter foram identificadas e os resultados comparados a fim de avaliar qual o método foi mais eficaz. Os únicos protocolos em que foi possível recuperar o microrganismo de todos os testes foram aqueles em que foi utilizado o ágar mCCDA associado com o Caldo Bolton ou com o Caldo Brucella. Estes foram também os únicos protocolos que permitiram a recuperação de C. jejuni 24 horas após a contaminação experimental com inóculo igual a 100 UFC/25 g. Entretanto, o ágar mCCDA sem o uso de pré-enriquecimento apresentou desempenho insatisfatório, inferior ao dos demais protocolos. Conclui-se que ágar mCCDA com pré-enriquecimento em caldo Brucella ou em caldo Bolton foram mais eficientes para o isolamento de C. jejuni que os demais protocolos.

Prevalence, antimicrobial resistance and genetic diversity of Campylobacter coli and Campylobacter jejuni in Ecuadorian broilers at slaughter age

Thermotolerant Campylobacter spp. are a major cause of foodborne gastrointestinal infections worldwide. The linkage of human campylobacteriosis and poultry has been widely described. In this study we aimed to investigate the prevalence, antimicrobial resistance and genetic diversity of C. coli and C. jejuni in broilers from Ecuador. Caecal content from 379 randomly selected broiler batches originating from 115 farms were collected from 6 slaughterhouses located in the province of Pichincha during 1 year. Microbiological isolation was performed by direct plating on mCCDA agar. Identification of Campylobacter species was done by PCR. Minimum inhibitory concentration (MIC) values for gentamicin, ciprofloxacin, nalidixic acid, tetracycline, streptomycin, and erythromycin were obtained. Genetic variation was assessed by RFLP-flaA typing and Multilocus Sequence Typing (MLST) of selected isolates. Prevalence at batch level was 64.1%. Of the positive batches 68.7% were positive for C. coli, 18.9% for C. jejuni, and 12.4% for C. coli and C. jejuni. Resistance rates above 67% were shown for tetracycline, ciprofloxacin, and nalidixic acid. The resistance pattern tetracycline, ciprofloxin, and nalidixic acid was the dominant one in both Campylobacter species. RFLP-flaA typing analysis showed that C. coli and C. jejuni strains belonged to 38 and 26 profiles respectively. On the other hand MLST typing revealed that C. coli except one strain belonged to CC-828, while C. jejuni except 2 strains belonged to 12 assigned clonal complexes (CCs). Furthermore 4 new sequence types (STs) for both species were described, whereby 2 new STs for C. coli were based on new allele sequences. Further research is necessary to estimate the impact of the slaughter of Campylobacter positive broiler batches on the contamination level of carcasses in slaughterhouses and at retail in Ecuador.

Antibiotic Resistance of Campylobacter Recovered from Faeces and Carcasses of Healthy Livestock.

Campylobacter is of major significance in food safety and human and veterinary medicine. This study highlighted resistance situation in the area of veterinary public health in Ghana. Using selective mCCDA agar, isolates were confirmed phenotypically on API CAMPY and genotypically by multiplex PCR of IpxA gene. The susceptibility profile of species to common and relevant antibiotics was determined by the Kirby-Bauer disk diffusion method. Cattle, sheep, goat, and pig faecal samples analysed, respectively, yielded 13.2% (16/121), 18.6% (22/102), 18.5% (25/135), and 28.7% (29/101) Campylobacter species while 34.5% (38/110), 35.9% (42/117), 23.9% (32/134), and 36.3% (37/102) were, respectively, recovered from the carcasses. Species identified in faeces were C. jejuni 35.8% (33/92), C. jejuni subsp. doylei 4.3% (4/92), C. coli 47.8% (44/92), and C. lari 12.0% (11/92). Species discovered in carcasses were C. jejuni 83.9% (125/149), C. jejuni subsp. doylei 2.0% (3/149), C. coli 6.0% (9/149), and C. lari 8.1% (12/149). Resistance ranged from 92 to 97% to the β-lactams, 7 to 69% to the quinolones, 0 to 44% to the aminoglycosides, 97 to 100% to erythromycin, 48 to 94% to tetracycline, 45 to 88% to chloramphenicol, and 42 to 86% to trimethoprim/sulfamethoxazole as 0% resistance was observed against imipenem.

Isolation of Campylobacter spp. from Client-Owned Dogs and Cats, and Retail Raw Meat Pet Food in the Manawatu, New Zealand.

Campylobacter causes acute gastroenteritis in people worldwide and is frequently isolated from food, animals and the environment. The disease is predominately food-borne but many routes of transmission and sources of infection have been described, including contact with pets. The prevalence of Campylobacter spp. in dogs and cats varies widely, and data on New Zealand pets are limited. This study aimed to investigate the prevalence of Campylobacter spp. in dogs, cats and retail raw meat pet food products in New Zealand and to characterize Campylobacter jejuni isolates using multilocus sequence typing (MLST). Ninety dogs and 110 cats examined at the Massey University Veterinary Teaching Hospital for elective procedures, and fifty locally purchased retail raw meat pet diets were sampled. Two culture protocols combining Bolton broth enrichment and mCCDA and CAT agars in a microaerobic atmosphere at 42°C and 37°C with species identification using PCR were performed. The prevalence of Campylobacter spp., C.jejuni, Campylobacter upsaliensis and Campylobacter helveticus was 36%, 13%, 23% and 1% in dogs and 16%, 5%, 5% and 7% in cats, respectively. One dog had Campylobacter lari confirmed, and three dogs and one cat had multiple Campylobacter spp. detected. Significantly more animals tested positive using CAT than mCCDA agar (P<0.001). Being neutered, vaccinated for Bordetella bronchiseptica, fed dry diets and brought in for neutering were protective factors for dogs, whereas attendance for dental treatment was a risk factor for cats. Campylobacter spp. were isolated from 28%, C.jejuni 22%, C.lari 6% and Campylobacter coli 6% of food samples. Six isolates positive by Campylobacter genus PCR were identified as Arcobacter butzleri. Poultry meat was more likely to be positive than non-poultry meat (P=0.006). Of the 13 C.jejuni pet isolates with full MLST profiles, eight were of different sequence types (ST) and all nine food isolates were of different STs.

English

Environmental waters are established sources of Campylobacter infections in humans. The aim of this study was to assess the distribution and susceptibility profiles of Campylobacter species isolated from irrigation and domestic water sources in Ghana. Samples were pre-enriched with CCDA broth and isolated on mCCDA agar. Isolates were confirmed on API CAMPY kit and with the Kirby-Bauer disk diffusion method to determine the susceptibility patterns of species. Of the 188 water samples analyzed, 42 isolates were confirmed to be contaminated with Campylobacter species, giving a prevalence rate of 22.3%. Prevalence of Campylobacter in the various water sources were 35.7% in rivers, 26.2% in streams, 21.4% in wells, 9.5% in ponds and 7.1% in boreholes. Sixty four percent (64%) of Campylobacter species were Campylobacter jejuni, followed by Campylobacter coli (19%) and Campylobacter lari (14%). Resistance was 100% to the β-lactams, 98% to erythromycin, 48-69% to the quinolones, 45-55% to the aminoglycosides, 71% to trimethoprim sulphamethoxazole, 76% to tetracycline and 90% to chloramphenicol. All the isolates (100%) were multidrug resistant. The presence of multidrug resistant Campylobacter species in the different water sources sampled in this study may indicate a significant health risk to humans and animals. Key words: Campylobacter, antibiotic resistance, water sources, Ghana.

Campylobacter in broiler slaughter samples assessed by direct count on mCCDA and Campy-Cefex agar.

Campylobacter spp. cause foodborne illnesses in humans primarily through the consumption of contaminated chicken. The aim of this study was to evaluate the United States Department of Agriculture's (USDA) recommended methodology, protocol MLG 41.02, for the isolation, identification and direct plate counting of Campylobacter jejuni and C. coli samples from the broiler slaughtering process. A plating method using both mCCDA and Campy-Cefex agars is recommended to recover Campylobacter cells. It is also possible to use this method in different matrices (cloacal swabs and water samples). Cloacal swabs, samples from pre-chiller and post-chiller carcasses and samples of pre-chiller, chiller and direct supply water were collected each week for four weeks from the same flock at a slaughterhouse located in an abattoir in southern Brazil. Samples were analyzed to directly count Campylobacter spp., and the results showed a high frequency of Campylobacter spp. on Campy-Cefex agar. For the isolated species, 72% were identified as Campylobacter jejuni and 38% as Campylobacter coli. It was possible to count Campylobacter jejuni and Campylobacter coli from different samples, including the water supply samples, using the two-agar method. These results suggest that slaughterhouses can use direct counting methods with both agars and different matrices as a monitoring tool to assess the presence of Campylobacter bacteria in their products.

Polymerase Chain Reaction confirmation and AST test of Campylobacter jejuni from sheep of Debrebirhan Agricultural Research Center North Shoa Ethiopia

Thermophilic Campylobacter have been recognized as an important cause of food born bacteria resulting in a diarrhea in different places of the world. There are few reports on the susceptibilities of thermotolerant Campylobacter strains in human, animal and foods of animal origin in Ethiopia. During a 9 month period from August 2011 to April 2012, 43 Campylobacter jejuni were isolated from fecal and carcass swab samples respectively on blood free mCCDA agar plate in micro aerobic atmosphere and were confirmed by Polymerase Chain Reaction (PCR). The antimicrobial susceptibility test was carried out for forty three clinical isolate of Campylobacter jejuni by agar disc diffusion method. Most of the isolates tested were 100 % susceptible to penicillin and nalidixic acid 97.7% gentamicin and ciprofloxacin, 95.3 % streptomycin and chloramphenicol, 90.7% to erythromycin, 74.4 % to tetracycline and 65.1% to ampicillin. Resistance was found 34.9% to ampicillin, 23.3% to tetracycline and 7% erythromycin with only 2.3% intermediate to tetracycline and erythromycin, whereas none was susceptible isolate to cephalothin. In our study area penicillin, streptomycin, and nalidixic acid show good antimicrobial activities against Campylobacter.

Method-Dependent Variability in Determination of Prevalence of Campylobacter jejuni and Campylobacter coli in Canadian Retail Poultry

Campylobacter is the most frequent cause of bacterial gastroenteritis in Canada, and the illness is commonly associated with poultry consumption. Whereas Canadian retail poultry is often contaminated with campylobacters, studies on the prevalence of this organism are inconsistent due to variability in sampling and microbiological methodology. To determine the current microbiological status of Canadian poultry, and to evaluate two commonly used microbiological methods, 348 raw poultry samples were collected at retail across Canada over a period of 3 years (2007 to 2010) and were analyzed for the presence of thermophilic Campylobacter species. The overall prevalence of Campylobacter spp. was found to be 42.8% by a combination of the two testing methods, with 33.9% of the samples positive for C. jejuni, 3.7% of the samples positive for C. coli, and 5.2% of the samples positive for both. Variability in Campylobacter spp. prevalence was observed in samples obtained from different regions across Canada and from poultry with or without skin, but this was not statistically significant. In co-contaminated samples, C. jejuni was preferentially recovered from Preston agar compared with mCCDA and Campy-Cefex agar, with an increase in recovery of C. coli on all selective media after 48 h of enrichment. A subset of 214 of the poultry rinses were analyzed by both Health Canada's standard method, MFLP-46 (enrichment in Park and Sanders broth), and a second method requiring enrichment in Bolton broth. Significantly more positive samples were obtained with the MFLP-46 method (40.6%) than with the alternate method (35.0%). This improved recovery with MFLP-46 may be due to the omission of cycloheximide from this method. These results demonstrate that determination of prevalence of Campylobacter spp. on poultry products may be significantly impacted by the choice of microbiological methods used. Canadian poultry continues to be a source of exposure to Campylobacter spp.

Surveillance of Amyloid Deposition and Bacterial Contamination in Chicken Liver from Meat Market

From the standpoint of poultry meat hygiene, amyloid deposition and bacterial contamination in livers of 197 mature white layer chickens from a slaughterhouse and 100 young broiler breeder chickens from 5 different retail butcher shops were examined. Amyloid deposition was detected in 1 liver from 2-years-old layer using alkaline Congo red staining and immunohistochemical examination, but no amyloid deposition was observed in 100 livers from young broiler. Total bacterial counts in the chicken livers were examined using nutrient, desoxycholate hydrogen sulfate and mCCDA agar plates, but no close relationship between amyloid deposition and bacterial contamination (counts and species) was observed in this study. These results suggest there is low incidence of systemic amyloid deposition in chicken liver from meat market.

Use of blood-free enrichment broth in the development of a rapid protocol to detect Campylobacter in twenty-five grams of chicken meat

A Food Pathogen Enrichment (FPE) broth, which supports the growth of Campylobacter without lysed blood and CO2, was developed. The FPE broth supports the growth of Campylobacter to the same degree as Bolton and Preston broths. Using the FPE broth, we developed a novel rapid protocol to detect small numbers of Campylobacter in 25g of food. The sensitivity of FPE enrichment and PCR to detect Campylobacter spp. from spiked chicken meat was determined. The detection sensitivities for non-stressed C. jejuni and C. coli from fresh meat ranged from 5.8 to 1.1×101CFU per 25g of chicken meat, and those for freeze-stressed C. jejuni and C. coli from frozen meat ranged from 9.9×101 to 2.0×102CFU. The FPE broth enrichment culture (24h) of chicken meat, followed by PCR, resulted in a significantly higher detection score (80% positive) than conventional Bolton enrichment and subsequent colony isolation using mCCDA agar plates (18% positive). Differences between our new protocol and the Bolton enrichment method were due to the overgrowth of many resistant bacteria, especially extended-spectrum beta-lactamase-producing bacteria in the Bolton enrichment broth.

Comparison of various culture methods (Skirrow medium, a blood-free medium and a filtration system enriched in Bolton and Preston broths) for isolation of Campylobacter spp. from raw meat samples

We compared six procedures and investigated the optimal method for isolation of Campylobacter spp. from raw meat samples. Ninety-nine meat samples were enriched in Bolton broth and Preston broth, followed by plating on Skirrow, mCCDA, and blood agar (a membrane filter on its surface) media, respectively. Thirty-nine of 99 samples were positive and 71 Campylobacter were isolated by one or more methods. More than one species of Campylobacter were obtained in 8 (20.5 %) of 39 positive samples and two genotypes were yielded on the same medium (11 samples, 28.2 %) by pulsed-field gel electrophoresis (PFGE) genotyping. Enrichment by Preston broth was significantly better than by Bolton broth (P < 0.05). Moreover, the latter failed to detect Campylobacter jejuni strains. Skirrow medium was significantly less efficient than mCCDA medium and membrane filtration method (P < 0.05). Overall, the combination of PC (primary enrichment in Preston broth, followed by selective enrichment on mCCDA agar), PF (primary enrichment in Preston broth, followed by membrane filtration culture onto blood agar), and BF (primary enrichment in Bolton broth, followed by membrane filtration culture onto blood agar) methods provided the optimum isolation rate of Campylobacter spp.

&lt;i&gt;Campylobacter&lt;/i&gt; Spp. Epidemiology and Antimicrobial Susceptibility in a Developing Country, Burkina Faso (West Africa)

Data on campylobacteriosis are almost nonexistent in Burkina Faso. In this study conducted from 2006 to 2008 in Ouagadougou, stool specimens and sociodemographic data were collected from 1 246 patients attending the university teaching hospital for enteritis. Stool samples were analyzed for the presence of Campylobacter by the direct culture method on selective mCCDA agar followed by antibiotic susceptibility testing on the isolated strains. The isolation rate of Campylobacter was 2.3%,comprising of the following species C. jejuni (51.8%), C. coli (13.8%), and C. upsaliensis (3.5%). However, 30.9% of the isolates were unidentified. No resistant strain was found to gentamicin. The resistance to amoxicillin+clavulanic acid (3.4%) was lower than those (10.3-34.5%) to the other antibiotics: erythromycin (10.3%), tetracycline (10.3%), ciprofloxacin (13.8%), amoxicillin (24.1%) and ceftriaxone (34.5%), nalidixic acid (34.5%). Significant associations were found between Campylobacter enteritis and contact with animals (P=0.03), and HIV infection (P&lt;0.0001), in contrast to other sociodemographic and seasonal factors. From the data obtained Amoxicillin+clavulanic acid appear to be the first choice for treatment. The implementation of a national program may be helpful in controlling the spread of the disease and the increase of resistance to antibiotics.Keywords: Campylobacter, epidemiology, HIV, drug resistance, Burkina Faso

Retention of Campylobacter (Campylobacterales: Campylobacteraceae) in the House Fly (Diptera: Muscidae)

The house fly (Musca domestica L.) may transmit Campylobacter to broiler flocks. We assessed the retention time of house flies for Campylobacter jejuni at five temperatures and three doses. Flies were inoculated individually at their proboscis with 1.6 x 10(7) CFU (colony forming units) of C. jejuni and incubated at 15, 20, 25, 30, and 35 degrees C. Furthermore, a dose experiment was conducted at 25 degrees C where individual flies were inoculated in three series: 6.5 x 10(6), 6.0 x 10(4), and 8.2 x 10(2) C.jejuni CFU. Whole flies were tested for C. jejuni carriage at 0, 6, 12, 18, and 24 h by initial preenrichment in Bolton broth, which afterwards was streaked on modified mCCDA agar plates and incubated under micro-aerobic conditions. The results showed that the time C. jejuni remained in flies declined over time with ascending temperatures and when reducing the inoculation dose. All flies stayed Campylobacter positive 24 h postinoculation at 15 degrees C whereas only one-third of the flies were positive at 20 degrees C and few to none at 25, 30, and 35 degrees C. When combinations of temperature and retention time were expressed as accumulated day-degrees, data could be adequately fitted using a generalized linear mixed model that included a linear effect of day-degrees and the difference between the lowest and the two highest doses. Based on model predictions of selected combinations of temperature and dose, the time for 50% and 1% of flies containing Campylobacter was calculated. It is suggested that house flies are mainly short distance carriers of C. jejuni.

A Comparison of Different Culture Methods for the Recovery ofCampylobacterSpecies from Pets

Five culture methods for the recovery of Campylobacter species (spp.) were evaluated on 361 rectal swabs collected from cats and dogs in Ireland. Speciation using PCR methods was performed on all isolates to assess the sensitivity of each culture method for isolation of Campylobacter spp., and to establish the prevalence of Campylobacter jejuni, C. coli, C. upsaliensis, C. lari and C. helveticus. Overall 163 of 361 (45.2%) samples were confirmed Campylobacter spp. positive. Direct plating onto modified charcoal cefoperazone deoxycholate agar (mCCDA) with cefoperazone, amphotericin and teicoplanin (CAT) selective supplement yielded a significantly higher prevalence of Campylobacter spp. (33.0%) than each of the other four methods (P < or = 0.05). This method was also the most sensitive method for isolation of C. upsaliensis compared with any of the other four methods used in the current study (P < or = 0.05). A direct plating method onto mCCDA agar with CCDA selective supplement and a filtration method onto blood agar after pre-enrichment in CAT supplemented broth yielded lower Campylobacter spp. prevalences of 19.7% and 17.5% respectively. A filtration method onto CAT agar and pre-enrichment in Preston broth before plating onto mCCDA agar were less sensitive for the isolation of Campylobacter spp. Speciation results of Campylobacter isolates revealed the majority of Campylobacter isolates were C. upsaliensis (50.0%) and C. jejuni (41.9%). A small number of isolates were C. coli (2.6%), C. lari (1.5%) and C. helveticus (1.1%). The overall detection of Campylobacter spp. in the 361 pets sampled was significantly increased by using a combination of isolation methods (P < or = 0.05), producing a more accurate determination of the prevalence of Campylobacter spp. in pets in Ireland and of the actual Campylobacter species. As the majority of Campylobacter spp. were recovered by direct plating onto mCCDA agar with CAT supplement, this method is the method of choice if only a single method is selected for isolation of the most common Campylobacter spp. detected in pets and humans.