Genomic diversity of Campylobacter jejuni and Campylobacter coli isolated from the Ethiopian dairy supply chain.

Campylobacteriosis outbreaks have previously been linked to dairy foods. While the genetic diversity of Campylobacter is well understood in high-income countries, it is largely unknown in low-income countries, such as Ethiopia. This study therefore aimed to conduct the first genomic characterization of Campylobacter isolates from the Ethiopian dairy supply chain to aid in future epidemiological studies. Fourteen C. jejuni and four C. coli isolates were whole genome sequenced using an Illumina platform. Sequences were analyzed using the bioinformatics tools in the GalaxyTrakr platform to identify MLST types, and single nucleotide polymorphisms, and infer phylogenetic relationships among the studied isolates. Assembled genomes were further screened to detect antimicrobial resistance and virulence gene sequences. Among 14 C. jejuni, ST 2084 and ST 51, which belong to the clonal complexes ST-353 and ST-443, respectively, were identified. Among the 4 sequenced C. coli isolates, two isolates belonged to ST 1628 and two to ST 830 from the clonal complex ST-828. The isolates of C. jejuni ST 2084 and ST 51 carried ß-lactam resistance gene blaOXA-605, a fluoroquinolone resistance-associated mutation T86I in the gryA gene, and a macrolide resistance-associated mutation A103V in 50S L22. Only ST 2084 isolates carried the tetracycline resistance gene tetO. Conversely, all four C. coli ST 830 and ST 1628 isolates carried tetO, but only ST 1628 isolates also carried blaOXA-605. Lastly, C. jejuni ST 2084 isolates carried a total of 89 virulence genes, and ST 51 isolates carried up to 88 virulence genes. Among C. coli, ST 830 isolates carried 71 genes involved in virulence, whereas two ST 1628 isolates carried up to 82 genes involved in virulence. Isolates from all identified STs have previously been isolated from human clinical cases, demonstrating a potential food safety concern. This finding warrants further monitoring of Campylobacter in dairy foods in Ethiopia to better understand and manage the risks associated with Campylobacter contamination and transmission.

Molecular Detection of Virulence-Associated Markers in Campylobacter coli and Campylobacter jejuni Isolates From Water, Cattle, and Chicken Faecal Samples From Kajiado County, Kenya.

Campylobacter is a zoonotic foodborne pathogen that is often linked with gastroenteritis and other extraintestinal infections in humans. This study is aimed at determining the genetic determinants of virulence-encoding genes responsible for flagellin motility protein A (flaA), Campylobacter adhesion to fibronectin F (cadF), Campylobacter invasion antigen B (ciaB) and cytolethal distending toxin (cdt) A (cdtA) in Campylobacter species. A total of 29 Campylobacter coli isolates (16 from cattle, 9 from chicken, and 4 from water samples) and 74 Campylobacter jejuni isolates (38 from cattle, 30 from chicken, and 6 from water samples) described in an earlier study in Kajiado County, Kenya, were examined for the occurrence of virulence-associated genes using polymerase chain reaction (PCR) and amplicon sequencing. The correlations among virulence genes were analyzed using Pearson's correlation coefficient (R) method. Among the 103 Campylobacter strains screened, 89 were found to harbour a single or multiple virulence gene(s), giving an overall prevalence of 86.4%. C. jejuni strains had the highest prevalence of multivirulence at 64.9% (48/74), compared to C. coli (58.6%, 17/29). The ciaB and flaA genes were the most common virulence genes detected in C. jejuni (81.1% [60/74] and 62.2% [46/74], respectively) and in C. coli (each at 62.1%; 18/29). Campylobacter isolates from chicken harboured the most virulence-encoding genes. C. jejuni strains from chicken and cattle harboured the highest proportions of the cdtA and ciaB genes, respectively. All the C. coli strains from water samples harboured the cadF and flaA genes. The results obtained further revealed a significant positive correlation between cadF and flaA (R = 0.733). C. jejuni and C. coli strains from cattle, chicken, and water harbour virulence markers responsible for motility/colonization, invasion, adherence, and toxin production, evoking their important role in campylobacteriosis development among humans and livestock. The identification of cattle, chicken, and water samples as reservoirs of virulent Campylobacter spp. highlights the possible risk to human health. These data on some virulence genes of Campylobacter will assist food safety and public health officials in formulating policy statements.

High Campylobacter diversity in retail chicken: epidemiologically important strains may be missed with current sampling methods.

Campylobacter spp. are leading bacterial gastroenteritis pathogens. Infections are largely underreported, and the burden of outbreaks may be underestimated. Current strategies of testing as few as one isolate per sample can affect attribution of cases to epidemiologically important sources with high Campylobacter diversity, such as chicken meat. Multiple culture method combinations were utilized to recover and sequence Campylobacter from 45 retail chicken samples purchased across Norwich, UK, selecting up to 48 isolates per sample. Simulations based on resampling were used to assess the impact of Campylobacter sequence type (ST) diversity on outbreak detection. Campylobacter was recovered from 39 samples (87%), although only one sample was positive through all broth, temperature, and plate combinations. Three species were identified (Campylobacter jejuni, Campylobacter coli, and Campylobacter lari), and 33% of samples contained two species. Positive samples contained 1-8 STs. Simulation revealed that up to 87 isolates per sample would be required to detect 95% of the observed ST diversity, and 26 isolates would be required for the average probability of detecting a random theoretical outbreak ST to reach 95%. An optimized culture approach and selecting multiple isolates per sample are essential for more complete Campylobacter recovery to support outbreak investigation and source attribution.

Isolation, characterization, and antimicrobial resistance profiles of Campylobacter jejuni and Campylobacter coli from raw meat of large livestock in Shahrekord, Iran.

Campylobacter spp. genera is one of the most common causes of microbial enteritis worldwide. This study aimed to find out how common Campylobacter organisms were in raw meat from large livestock in Iran, as well as to determine their antibiotic susceptibility profiles. Several 550 fresh, ready-to-eat meat samples were collected from slaughterhouses, butcher shops, and restaurants in the study region. The samples were collected from cattle (n=138), goats (n=102), camels (n=56), and sheep (n=254). Campylobacter spp. were isolated and identified using normal bacteriological methods and polymerase chain reaction (PCR). Genotyping was performed using PCR to identify virulence genes. The disc diffusion technique was used to determine antibiotic susceptibility. The two Campylobacter spp. were found in 84 (15.27%) of the 550 meat samples tested. Cattle and camel samples accounted for the highest (52.38%) and lowest (3.57%) frequencies of Campylobacter spp., respectively. There were significant differences in the prevalence of Campylobacter spp. in cattle (2=43.04 or OR=7.68, CI=3.40-17.30, P<0.01). Campylobacter jejuni and Campylobacter coli accounted for 82.14% (n=69) of Campylobacter spp. isolated from raw meat. While C. jejuni was found in 39.28% of the samples (n=33), C. coli was observed in 42.85% (n=36). Other Campylobacter spp. formed 17.85 % (n=15) of the samples. The most common genotypes observed in C. jejuni bacteria collected from different types of large animal samples were ciaB (100%) and flaA (100%). On the other hand, virbll (7.69%) was the C. jejuni strain found with the lowest incidence in different large animal samples. The most frequent genotypes found in C. coli bacteria were ciaB (100%) and flaA (100%). C. coli isolates dnaJ (0%), wlaN (0%), virbll (0%), and ceuE (0%) were detected with the lowest frequency in several samples from large livestock. Campylobacter spp. isolated from different sample types and sources were 100% sensitive to aphA-3-1 and GM10. The isolates were reported to be resistant to E15 (76.93%), cmeB (69.24%), aadE1 (69.24%), CIP5 (69.24%), and AM10 (69.24%). According to this study, Campylobacter was found in food from factory farming. Consequently, the disease can be transmitted by eating raw or undercooked meat. Therefore, proper handling and preparation of meat meals, as well as hygiene measures from the slaughterhouse to the retailer, are critical in preventing Campylobacter infections.

Campylobacter jejuni and Campylobacter coli infection, determinants and antimicrobial resistance patterns among under-five children with diarrhea in Amhara National Regional State, Northwest Ethiopia.

BACKGROUND: Children with under-five year age disproportionally affected with foodborne illness. Campylobacteriosis is the most common foodborne disease next to Norovirus infection. Macrolides are commonly prescribed as the first line of treatment for Campylobacter gastroenteritis, with fluoroquinolone and tetracycline as secondary options. However, resistance to these alternatives has been reported in various regions worldwide. OBJECTIVE: To determine the prevalence, associated risk-factors and antimicrobial resistance of Campylobacter jejuni and C. coli among under-five children with diarrhea. METHODS: Institution-based cross-sectional study was conducted from November, 2022 to April 2023. The study sites were selected using a random sampling technique, while the study subjects were included using a convenient sampling technique. The data were collected using a structured questionnaire. Stool samples were inoculated onto modified charcoal cefoperazone deoxycholate agar and incubated for 48 hours. The suspected colonies were analyzed using matrix-assisted laser desorption ionization-time of flight mass spectrometry to confirm the species. Antimicrobial susceptibility testing was performed using a disc diffusion technique. All potential covariates (independent variables) were analyzed one by one using bivariate logistic regression model to identify candidate variables with P value < 0.25. Multivariable logistic analysis was used to identify potential associated factors using the candidate variables. A p value ≤ 0.05 at a 95% confidence interval was statistically significant. RESULT: Among the 428 samples, 7.0% (CI: 4.5-9.3) were confirmed Campylobacter species. The prevalence of C. jejuni and C. coli among under-five children was 5.1% (CI: 3.0-7.0) and 1.9% (CI: 0.7-3.3), respectively. C. jejuni (73.3%) was dominant over C. coli (26.7%). The resident, contact with domestic animals, and parents/guardians education level were significantly associated with campylobacteriosis among under-five children. One-third of the Campylobacter isolates (33.3%, 10/30) were resistant to ciprofloxacin and tetracycline whereas 10.0% (3/30) were resistant to erythromycin. Furthermore, 3.3% (1/30) of the Campylobacter were found to be multidrug-resistant. CONCLUSION: The prevalence of Campylobacter species was 7.0%. The resistance rate of Campylobacter species of ciprofloxacin and tetracycline-resistance strains was 33.3%. Peri-urban residence, contact with domestic animals, and low parental educational statuses were significantly associated factors with increased risk of Campylobacter infection. Continuous surveillance on antimicrobial resistance and health education of personal and environmental hygiene should be implemented in the community.

High-throughput sequencing reveals genetic determinants associated with antibiotic resistance in Campylobacter spp. from farm-to-fork.

Campylobacter species are one of the most common causative agents of gastroenteritis worldwide. Resistance against quinolone and macrolide antimicrobials, the most commonly used therapeutic options, poses a serious risk for campylobacteriosis treatment. Owing to whole genome sequencing advancements for rapid detection of antimicrobial resistance mechanisms, phenotypic and genotypic resistance trends along the "farm-to-fork" continuum can be determined. Here, we examined the resistance trends in 111 Campylobacter isolates (90 C. jejuni and 21 C. coli) recovered from clinical samples, commercial broiler carcasses and dairy products in Cairo, Egypt. Multidrug resistance (MDR) was observed in 10% of the isolates, mostly from C. coli. The prevalence of MDR was the highest in isolates collected from broiler carcasses (13.3%), followed by clinical isolates (10.5%), and finally isolates from dairy products (4%). The highest proportion of antimicrobial resistance in both species was against quinolones (ciprofloxacin and/or nalidixic acid) (68.4%), followed by tetracycline (51.3%), then erythromycin (12.6%) and aminoglycosides (streptomycin and/or gentamicin) (5.4%). Similar resistance rates were observed for quinolones, tetracycline, and erythromycin among isolates recovered from broiler carcasses and clinical samples highlighting the contribution of food of animal sources to human illness. Significant associations between phenotypic resistance and putative gene mutations was observed, with a high prevalence of the gyrA T86I substitution among quinolone resistant isolates, tet(O), tet(W), and tet(32) among tetracycline resistant isolates, and 23S rRNA A2075G and A2074T mutations among erythromycin resistant isolates. Emergence of resistance was attributed to the dissemination of resistance genes among various lineages, with the dominance of distinctive clones. For example, sub-lineages of CC828 in C. coli and CC21 in C. jejuni and the genetically related clonal complexes 'CC206 and CC48' and 'CC464, CC353, CC354, CC574', respectively, propagated across different niches sharing semi-homogenous resistance patterns.

Whole genome-based characterisation of antimicrobial resistance and genetic diversity in Campylobacter jejuni and Campylobacter coli from ruminants.

Campylobacter, a leading cause of gastroenteritis in humans, asymptomatically colonises the intestinal tract of a wide range of animals.Although antimicrobial treatment is restricted to severe cases, the increase of antimicrobial resistance (AMR) is a concern. Considering the significant contribution of ruminants as reservoirs of resistant Campylobacter, Illumina whole-genome sequencing was used to characterise the mechanisms of AMR in Campylobacter jejuni and Campylobacter coli recovered from beef cattle, dairy cattle, and sheep in northern Spain. Genome analysis showed extensive genetic diversity that clearly separated both species. Resistance genotypes were identified by screening assembled sequences with BLASTn and ABRicate, and additional sequence alignments were performed to search for frameshift mutations and gene modifications. A high correlation was observed between phenotypic resistance to a given antimicrobial and the presence of the corresponding known resistance genes. Detailed sequence analysis allowed us to detect the recently described mosaic tet(O/M/O) gene in one C. coli, describe possible new alleles of blaOXA-61-like genes, and decipher the genetic context of aminoglycoside resistance genes, as well as the plasmid/chromosomal location of the different AMR genes and their implication for resistance spread. Updated resistance gene databases and detailed analysis of the matched open reading frames are needed to avoid errors when using WGS-based analysis pipelines for AMR detection in the absence of phenotypic data.

Campylobacter coli strains from Brazil can invade phagocytic and epithelial cells and induce IL-8 secretion.

Campylobacter spp. have been a predominant cause of bacterial foodborne gastroenteritis worldwide, causing substantial costs to public healthcare systems. This study aimed to assess the invasion and pro-inflammatory cytokine production capacity of Campylobacter coli strains isolated in Brazil. A total of 50 C. coli isolated from different sources in Brazil were analyzed for their capacity of invasion in Caco-2 and U-937 cell lines. The production of pro-inflammatory cytokines was quantitatively measured in response to C. coli. All the strains studied showed invasion percentage ≥ 40% in polarized Caco-2 cells. In U-937 cells assay, 35 of 50 C. coli strains studied showed invasion percentage ≥ 50%. A significant increase in IL-8 production by infected U-937 cells was observed for 17.5% of the C. coli isolates. The high percentages of invasion in Caco-2 and U-937 cells observed for all studied strains, plus the increased production of IL-8 by U-937 cells against some strains, highlighted the pathogenic potential of the C. coli studied and bring extremely relevant data since it has never been reported for strains isolated in Brazil and there are a few data for C. coli in the literature.

Isolation, identification, and antimicrobial susceptibility pattern of Campylobacter jejuni and Campylobacter coli from cattle, goat, and chicken meats in Mekelle, Ethiopia.

Campylobacter jejuni and Campylobacter coli are globally recognized as a major cause of bacterial foodborne gastroenteritis. A cross-sectional study was conducted from October 2015 to May 2016 in Mekelle city to isolate, identify, and estimate the prevalence of C. jejuni and C. coli in raw meat samples and to determine their antibiotic susceptibility pattern. A total of 384 raw meat samples were randomly collected from bovine (n = 210), goat (n = 108), and chicken (n = 66), and isolation and identification of Campylobacter spp. were performed using standard bacteriological techniques and PCR. Antibiotic susceptibility test was performed using disc diffusion method. Of the total 384 raw meat samples, 64 (16.67%) were found positive for Campylobacter spp. The highest prevalence of Campylobacter spp. was found in chicken meat (43.93%) followed by bovine meat (11.90%) and goat meat (9.25%). The most prevalent Campylobacter spp. isolated from meat samples was C. jejuni (81.25%). The overall prevalence of Campylobacter in restaurants, butcher shops, and abattoir was 43.93%, 18.30%, and 9.30%, respectively. 96.8%, 81.25%, 75%, and 71% of the Campylobacter spp. isolates were sensitive to norfloxacin, erythromycin, chloramphenicol, and sulphamethoxazole-trimethoprim, respectively. However, 96.9%, 85.9%, and 50% of the isolates were resistant to ampicillin, amoxicillin, and streptomycin, respectively. Strains that developed multi-drug resistant were 68.7%. The result of this study revealed the occurrence of Campylobacter in bovine, goat, and chicken meats. Hence, there is a chance of acquiring infection via consumption of raw or undercooked meat. Thus, implementation of hygienic practices from a slaughterhouse to the retailers, proper handling and cooking of foods of meat are very important in preventing Campylobacter infection.

Antimicrobial resistance and interspecies gene transfer in Campylobacter coli and Campylobacter jejuni isolated from food animals, poultry processing, and retail meat in North Carolina, 2018-2019.

The Center for Disease Control and Prevention identifies antimicrobial resistant (AMR) Campylobacter as a serious threat to U.S. public health due to high community burden, increased transmissibility, and limited treatability. The National Antimicrobial Resistance Monitoring System (NARMS) plays an important role in surveillance of AMR bacterial pathogens in humans, food animals and retail meats. This study investigated C. coli and C. jejuni from live food animals, poultry carcasses at production, and retail meat in North Carolina between January 2018-December 2019. Whole genome sequencing and bioinformatics were used for phenotypic and genotypic characterization to compare AMR profiles, virulence factors associated with Guillain-Barré Syndrome (GBS) (neuABC and cst-II or cst-III), and phylogenic linkage between 541 Campylobacter isolates (C. coli n = 343, C. jejuni n = 198). Overall, 90.4% (489/541) Campylobacter isolates tested positive for AMR genes, while 43% (233/541) carried resistance genes for three or more antibiotic classes and were classified molecularly multidrug resistant. AMR gene frequencies were highest against tetracyclines (64.3%), beta-lactams (63.6%), aminoglycosides (38.6%), macrolides (34.8%), quinolones (24.4%), lincosamides (13.5%), and streptothricins (5%). A total of 57.6% (114/198) C. jejuni carried GBS virulence factors, while three C. coli carried the C. jejuni-like lipooligosaccharide locus, neuABC and cst-II. Further evidence of C. coli and C. jejuni interspecies genomic exchange was observed in identical multilocus sequence typing, shared sequence type (ST) 7818 clonal complex 828, and identical species-indicator genes mapA, ceuE, and hipO. There was a significant increase in novel STs from 2018 to 2019 (2 in 2018 and 21 in 2019, p<0.002), illustrating variable Campylobacter genomes within food animal production. Introgression between C. coli and C. jejuni may aid pathogen adaption, lead to higher AMR and increase Campylobacter persistence in food processing. Future studies should further characterize interspecies gene transfer and evolutionary trends in food animal production to track evolving risks to public health.

Prevalence, molecular typing and antimicrobial susceptibility of Campylobacter spp. isolates in northern Spain.

AIM: To analyse the prevalence, genetic diversity and antimicrobial susceptibility of Campylobacter spp. in northern Spain. METHODS AND RESULTS: Campylobacter was isolated from 139 samples of broiler meat and faecal dropping of broiler and swine with a prevalence of 35·4, 62 and 42·8%, respectively. Campylobacter jejuni (n = 55) and Campylobacter coli (n = 31) were identified by multiplex-PCR in meat, faeces and human clinical samples while Campylobacter fetus (n = 3) was exclusively detected in the latter. Fingerprinting by flaA-RFLP and PFGE revealed 68 different genotypes from the 89 isolates with a Biodiversity Simpson's index of 0·98. The 86·5% of the isolates were resistant to ciprofloxacin, 85·4% to tetracycline and 49·4% to erythromycin; only three genotypes were susceptible to the three antimicrobial drugs. Multidrug resistance was detected in the 40·7% of the isolates. CONCLUSIONS: Campylobacter remains prevalent in northern Spain with a high biodiversity degree. About 93·3% of the isolates were resistant to one or more drugs. SIGNIFICANCE AND IMPACT OF THE STUDY: Although different measures are taken to control Campylobacter, the detection of isolates resistant to the drugs used in the treatment of campylobacteriosis is still high, including different species and genotypes. This evidences the need of additional strategies against this pathogen.

A Multiplex Quantitative Polymerase Chain Reaction Using Applied Biosystems 7500 Fast System for Simultaneous Identification of Three Campylobacter Species with Potential Applications to Food Analysis.

Consumption of Campylobacter-contaminated food is one of the most common causes of bacterial diarrhea. A previously developed quantitative polymerase chain reaction (qPCR) utilizing the SmartCycler instrument platform for identification of Campylobacter jejuni, Campylobacter coli, and Campylobacter lari had to be modified to address the recent discontinuation of the SmartCycler system. In this study, a multiplex qPCR assay was optimized on the Applied Biosystems 7500 Fast (AB7500F) platform to continue using qPCR for the identification of three target Campylobacter spp. AB7500F qPCR efficiencies obtained by testing reference genomic DNA (gDNA) were 90.9%, 86.4%, and 94.6% for C. jejuni, C. coli, and C. lari, respectively, with all correlation coefficient values >0.99. The qPCR results exhibited 100% specificity by testing gDNA samples from 37 non-target reference strains and 86 target strains (50 C. jejuni, 27 C. coli, and 9 C. lari strains) in this study. The lowest detection level using gDNA was 4, 7, and 2 genome copies per reaction for C. jejuni, C. coli, and C. lari, respectively. With a 2-day enrichment procedure, the qPCR method correctly detected target species in a spiked food matrix (frog leg, an aquaculture product). The sensitivity in 25 g food matrix was 4 colony-forming units (CFUs) for C. jejuni, 3 CFUs for C. coli, and 2 CFUs for C. lari. The results suggest that this AB7500F-based qPCR has potential applications for the identification of C. jejuni, C. coli, and C. lari in contaminated food.

Campylobacter coli bacteraemia: how common is it?

Campylobacter species are known to cause enteritis. However, over the past 40-50 years, there have been reports of varying presentations, such as cellulitis, spondylodiscitis and bacteraemia. Of the Campylobacter species, Campylobacter jejuni is the most common culprit for causing bacteraemia, however, Campylobacter coli bacteraemia is becoming more prevalent. Here, we discuss an unusual case of C. coli bacteraemia in a patient with decompensated liver cirrhosis.

Genome-Wide Identification of Host-Segregating Single-Nucleotide Polymorphisms for Source Attribution of Clinical Campylobacter coli Isolates.

Campylobacter is among the most common causes of gastroenteritis worldwide. Campylobacter jejuni and Campylobacter coli are the most common species causing human disease. DNA sequence-based methods for strain characterization have focused largely on C. jejuni, responsible for 80 to 90% of infections, meaning that C. coli epidemiology has lagged behind. Here, we have analyzed the genome of 450 C. coli isolates to determine genetic markers that can discriminate isolates sampled from 3 major reservoir hosts (chickens, cattle, and pigs). These markers then were applied to identify the source of infection of 147 C. coli strains from French clinical cases. Using STRUCTURE software, 259 potential host-segregating markers were revealed by probabilistic characterization of single-nucleotide polymorphism (SNP) frequency variation in strain collections from three different hosts. These SNPs were found in 41 genes or intergenic regions, mostly coding for proteins involved in motility and membrane functions. Source attribution of clinical isolates based on the differential presence of these markers confirmed chickens as the most common source of C. coli infection in France.IMPORTANCE Genome-wide and source attribution studies based on Campylobacter species have shown their importance for the understanding of foodborne infections. Although the use of multilocus sequence typing based on 7 genes from C. jejuni is a powerful method to structure populations, when applied to C. coli, results have not clearly demonstrated its robustness. Therefore, we aim to provide more accurate data based on the identification of single-nucleotide polymorphisms. Results from this study reveal an important number of host-segregating SNPs, found in proteins involved in motility, membrane functions, or DNA repair systems. These findings offer new, interesting opportunities for further study of C. coli adaptation to its environment. Additionally, the results demonstrate that poultry is potentially the main reservoir of C. coli in France.

Investigation of antimicrobial resistance and virulence genes of Campylobacter isolates from patients in a tertiary hospital in Edirne, Turkey.

Purpose: Campylobacter is one of the most common pathogens that cause food-borne infections worldwide. The aim of this study was to determine the antimicrobial resistance rates and the presence of multiple virulence genes in Campylobacter isolates obtained from humans. Materials and Methods: In this study, 71 Campylobacter isolates obtained from human faecal samples were used. Antimicrobial susceptibility tests were performed through the gradient strip method. The presence of virulence genes was investigated by monoplex and multiplex polymerase chain reaction. Results: The rate of resistance of the 66 Campylobacter jejuni isolates was 12.1% for erythromycin, 40.9% for tetracycline and 68.2% for ciprofloxacin. Only one of five Campylobacter coli isolates was resistant to these three antimicrobial agents. The flaB, pldA, cdtA, cadF, cdtC and ceuE genes were found in all 66 of the C. jejuni isolates. In the C. jejuni isolates, positivity rates of 92.4% for flaA, 96.7% for cdtB, 98.5% for ciaB, 90.9% for dnaJ and 96.7% for racR were observed. The flaA, flaB, ciaB, cdtA and cdtC genes were present in all C. coli isolates. Conclusions: It was detected that there is an increase in antimicrobial resistance of Campylobacter strains in our region, and most of the isolates harbour virulence genes.

Genetic environments and related transposable elements of novel cfr(C) variants in Campylobacter coli isolates of swine origin.

The cfr(C) is a cfr-like gene that confers cross-resistance to antibiotics targeting the 23S rRNA through methylation of nucleotide A2503. Here, we identified 7 C. coli isolates containing 4 novel cfr(C) variants from swine farm and slaughterhouses samples. Of the 7 cfr(C)-carrying isolates, one had a frame-shift mutation, while the other 6 had intact genes. However, one of the 6 intact genes did not show a PhLOPSA phenotype in the original isolate, but was fully functional when cloned into C. jejuni NCTC 11168. Cloning of cfr(C) variants into C. jejuni NCTC 11168 and conjugative transfer of the two cfr(C)-containing plasmids further confirmed their role in conferring resistance to PhLOPSA antimicrobials, and resulted in an 8-128-fold increase in their MICs. In all cfr(C)-carrying isolates, cfr(C) genes were located in the downstream of the kanamycin resistant gene aphA3. IS607* and IS1595-like were located immediately upstream of aphA3 gene and seemed to play a role in its recombination. A novel transposable element named ISCco7, which located immediately downstream of cfr(C) in two isolates, was probably associated with the integration of cfr(C). However, neither insertion sequence nor other transposable elements were identified near cfr(C) in the remaining five cfr(C)-positive isolates, indicating the mechanism underlying the integration of cfr(C) into plasmids or chromosomal DNA requires further investigation. These results reveal novel cfr(C) variants and their associated genetic environments in C. coli isolates and indicate the flexibility of C. coli in acquiring new antibiotic resistance genes.

Molecular characterization of Campylobacter spp. recovered from beef, chicken, lamb and pork products at retail in Australia.

Australian rates of campylobacteriosis are among the highest in developed countries, yet only limited work has been done to characterize Campylobacter spp. in Australian retail products. We performed whole genome sequencing (WGS) on 331 C. coli and 285 C. jejuni from retail chicken meat, as well as beef, chicken, lamb and pork offal (organs). Campylobacter isolates were highly diverse, with 113 sequence types (STs) including 38 novel STs, identified from 616 isolates. Genomic analysis suggests very low levels (2.3-15.3%) of resistance to aminoglycoside, beta-lactam, fluoroquinolone, macrolide and tetracycline antibiotics. A majority (>90%) of isolates (52/56) possessing the fluoroquinolone resistance-associated T86I mutation in the gyrA gene belonged to ST860, ST2083 or ST7323. The 44 pork offal isolates were highly diverse, representing 33 STs (11 novel STs) and harboured genes associated with resistance to aminoglycosides, lincosamides and macrolides not generally found in isolates from other sources. Prevalence of multidrug resistant genotypes was very low (<5%), but ten-fold higher in C. coli than C. jejuni. This study highlights that Campylobacter spp. from retail products in Australia are highly genotypically diverse and important differences in antimicrobial resistance exist between Campylobacter species and animal sources.

Prevalence and susceptibility to antibiotics from Campylobacter jejuni and Campylobacter coli isolated from chicken meat in southern Benin, West Africa.

OBJECTIVE: Poultry is commonly considered to be the primary vehicle for Campylobacter infection in humans. The aim of this study is to assess the risk of Campylobacteriosis in chicken meat consumers in southern Benin by assessing the prevalence and resistance profile of Campylobacter coli and Campylobacter jejuni isolated from chicken thigh in Southern Benin. RESULTS: The contamination rate of Campylobacter in the samples was 32.8%. From this percentage, 59.5% were local chicken thighs and 40.5% of imported chicken thighs (p = 0.045). After molecular identification, on the 256 samples analyzed, the prevalence of C. jejuni was 23.4% and 7.8% for C. coli, with a concordance of 0.693 (Kappa coefficient of concordance) with the results from phenotypic identification. Seventy-two-point seven percent of Campylobacter strains were resistant to Ciprofloxacin, 71.4% were resistant to Ampicillin and Tetracycline. 55.8% of the strains were multi-drug resistant.

Prevalence, genotypic diversity and detection of virulence genes in thermotolerant Campylobacter at different stages of the poultry meat supply chain.

Thermotolerant Campylobacter is the leading bacterial cause of foodborne illness in humans worldwide. The objectives of this study were to estimate prevalence and to identify and characterize potential sources of thermotolerant Campylobacter contamination in broilers on farms and at the slaughterhouse; to evaluate the clonal relationship among thermotolerant Campylobacter isolates from different stages of the broiler meat supply chain, and to analyze the presence of virulence genes in different sources of thermotolerant Campylobacter. A total of 1210 samples were collected from three broiler meat supply chains in Santa Fe, Argentina. At the farms, the sampling collection included broilers one week prior to slaughter, wild-living birds, domestic dogs, wild rodents, farm workers' boots, litter, feed, drinking water, flies, and darkling beetles (Alphitobius diaperinus). At the slaughtering line, the samples taken were from the evisceration zone (broiler cecum, working surfaces, evisceration knives and workers' hands), from the chiller zone (surfaces and direct supply water) and from the packing zone (work surfaces, workers' hands and broiler carcasses). The samples taken along each supply chain were in the same batch. The isolates obtained were identified to the species level (C. jejuni and C. coli) by multiplex PCR and were analyzed using pulsed-field gel electrophoresis to compare different profiles according to the source. Finally, the presence of 11 virulence genes was examined (cadF, cdtA, cdtB, cdtC, ciaB, flaA, flhA, iam, wlaN, virB11, racR). From 254 isolates, 128 (50.4%) were Campylobacter jejuni and 126 (49.6%) Campylobacter coli. C. jejuni was the species most prevalent in farm and C. coli the species most prevalent at the slaughterhouse. We detected thermotolerant Campylobacter in samples of wild birds, darkling beetles, farm workers' boots, flies and litter. At the slaughterhouse, the prevalence varied along the process line. By analyzing PFGE results, C. jejuni showed 21 profiles with three predominant genotypes, while C. coli showed 14 profiles with four predominant genotypes. A high genotype diversity was found; however, relationships between isolates from different stages of the broiler meat chain, between broiler and potential sources of thermotolerant Campylobacter contamination and between strains in the farm and in the slaughterhouse were detected. Furthermore, there was evidence of cross-contamination at the slaughterhouse. FlaA, flhA genes were detected in all strains, and the third most prevalent virulence gene was cadF. Only those strains obtained from flies, wild-living birds and broiler carcass samples harbored 10 of 11 pathogenic genes. The prevalence of some pathogenic genes between C. jejuni and C. coli was different. This evidence should contribute the scientific basis to implement risk management measures in public health.

Research Note: A baseline survey of thermotolerant Campylobacter in retail chicken in southern Brazil.

Chicken is a leading source of thermotolerant Campylobacter, which triggers human foodborne enteritis. This study evaluated thermotolerant Campylobacter contamination of retail chicken in southern Brazil, using qualitative and quantitative analyses. Selective enrichment in Bolton broth for 24 and 48 h after plating onto modified charcoal-cefoperazone-deoxycholate (mCCD) agar and Preston agar was assessed. The combined results of the detection and enumeration methods revealed a frequency of 70% occurrence of thermotolerant Campylobacter in chicken samples. Campylobacter was enumerated in 60% of the samples, whereas 46% of the samples were positive in the qualitative analysis. Quantitative analysis showed average counts of 3.10 ± 0.15 log10 CFU/sample. Higher numbers of Campylobacter-positive samples were found using 24-h enrichment before plating onto Preston agar (46%) than onto mCCD agar (2%). The majority of isolated strains were identified as Campylobacter jejuni, and Campylobacter coli was also found but to a lesser extent. Subtyping revealed a clear distinction between strains isolated from different chicken sources. The enriched samples plated onto mCCD agar showed extensive spreading of nonproducing extended-spectrum ß-lactamases Proteus mirabilis that hampered the identification of Campylobacter colonies. P. mirabilis strains showed resistance to cefoperazone, trimethoprim, and polymyxin B present in broth and plate media used and were inhibited by rifampicin present in Preston agar. The results underline the effect of the spread of contaminant strains on Campylobacter cultures, which might be prevented using a recently revised International Organization for Standardization method for qualitative analysis of chicken.