Thermophilic Campylobacter Research Articles

Occurrence, antimicrobial resistance and virulence properties of thermophilic Campylobacter coli originating from two different poultry settings

Thermophilic Campylobacter's (Campylobacter coli and Campylobacter jejuni) are usually zoonotic and spread by raw or undercooked meat especially poultry meat, unpasteurized milk, direct animal contact, and contaminated drinking water. In this study, 385 samples from commercial and backyard poultry settings were screened for the presence of Campylobacter spp. which revealed an isolation rate of 3.9% with higher isolation from the chicken caecum (15/113, 13.27%). On antimicrobial susceptibility testing, the highest resistance was observed for tetracycline (11/13, 84.61%) followed by macrolides (8/13, 61.53%), β-lactams (7/13, 53.84%), fluoroquinolones (5/13, 38.45%) and sulphonamide (4/13, 30.76%). Isolates from backyard poultry showed resistance to tetracycline (100%), β-lactam (66.66%), fluoroquinolone (25%), and macrolide (33.33%). Isolates from commercial poultry showed resistance to tetracycline (80%), macrolides (70%), β-lactams (51.66%), fluoroquinolones (42.5%) and sulphonamides (40%). Among the isolates, 92.30% were multi-drug resistant (MDR). Overall, tet(O) gene was observed in 6/11(54.55%) tetracycline resistant isolates. Notably, tet O gene was found in 6/8(75%) isolates from commercial poultry farms and it was absent in backyard poultry isolates. Similarly 23SrRNA and blaOXA-61 genes were detected in 4/8(50%) and 2/8(25%) macrolide and ampicillin resistant isolates, respectively. Among virulence genes, cdtC gene was noticed in all the Campylobacter isolates (100%), cadF in 93.33%, iamA in 80%, flaA in 73.33%, cdtA in 13.33% and, cdtB in 6.66% isolates while dnaJ, ciaB and pldA genes were not detected in any of the isolate. The Campylobacter isolates possessing antimicrobial resistance and virulence genes pose a serious public health threat through food chain.

Species diversity of dominant etiologically significant bacteria circulating in industrial poultry

The problem of the spread of bacterial infections of birds associated with opportunistic microorganisms occupies a significant place in the modern infectious pathology of birds.The aim of this work is to determine the species diversity of the dominant etiologically significant bacteria circulating in industrial poultry farming. Based on the results for the period 2019 to 2020, we carried out bacteriological studies of pathological material from birds in poultry farms of various technological directions.The dominant species are Escherichia coli, which accounts for 50.7%, Enterococcus faecalis - 25.4%, and Proteus mirabilis - 8.4%. Also, thermophilic Campylobacter spp. - 4.3% were isolated from the organs, including such epidemiologically dangerous species as Campylobacter jejuni. The microorganisms were virulent and differed in their cultural and biochemical properties.Cultures such as Escherichia coli, Proteus mirabilis, Pseudomonas aeruginosa, Enterococcus faecalis are isolated from many organs, which indicates the generalization of the infectious process, the culture of thermophilic Campylobacter spp. were isolated from the liver, which indicates the organotropy of these types of microorganisms.The species composition of the allocated microflora is determined by the epizootic situation in each individual farm, which results in a different percentage of the dominant species.Pure cultures of thermophilic campylobacter were obtained on selective Preston agar with cultivation for 7 days at a temperature of 42 ° C under microaerophilic conditions. However, culturing Campylobacter hepaticus is difficult because it is sensitive to a number of antimicrobial agents used in Campylobacter enrichment media and selective agar.The emergence of new Campylobacter species requires a revision of phenotypic identification algorithms. MALDI TOF mass spectrometry databases need to be updated and specific primers developed to identify new Campylobacter species.Significant variability of indicators of indication of one or another microflora from the liver and gallbladder indicates significant differences in the etiology of bacterial infections of birds.

A REVIEW OF CURRENT KNOWLEDGE AND GAPS ABOUT CAMPYLOBACTER METHODS: FROM CULTURE TO CHARACTERIZATION

Campylobacteriosis is a zoonotic disease caused by Campylobacter, mostly associated with consumption of contaminated foodstuffs and water. Campylobacter jejuni and Campylobacter coli recognized as the leader of foodborne diarrheal illness in humans. The frequency of these microorganisms in poultry is fairly high than Salmonella and more challenging to measure, which represent an expensive burden charge on public health due to their difficulties to master them, especially with the fast increase rates of multidrug-resistant of thermophilic Campylobacter strains. It is well recognized that Campylobacter spp. is a fastidious cell, difficult to isolate in laboratories owing to their requirements and sensibility. That’s why; these factors must be taken into consideration during recovery protocols. A variety of phenotyping tests have been reported and widely used for confirmation and identification of Campylobacter species. Nonetheless, Whole Genome Sequencing (WGS) and Culture-Independent Diagnostic Tests (CIDTs) are new eras of hopeful technologies, mainly involved in the detection and characterization of threaten public health pathogens. This review aimed to describe the culture methods, phenotypic and genotypic schemes used to isolate, identify, and characterize Campylobacter isolates, through discussing the current knowledge and gaps related to the application of these techniques over others performed for typing this microaerophilic genus.

Genomic diversity and antimicrobial resistance of Campylobacter spp. from humans and livestock in Nigeria

BackgroundCampylobacter spp. are zoonotic pathogens, ubiquitous and are found naturally as commensals in livestock from where they can be transmitted to humans directly or through animal products. The genomic diversity and antimicrobial resistance profile of Campylobacter was investigated with a focus on C. jejuni and C. coli in humans and livestock (poultry and cattle) from Nigeria.Methods586 human stool samples and 472 faecal samples from livestock were cultured for thermophilic Campylobacter species on modified charcoal cefoperazone deoxycholate agar (mCCDA). Culture in combination with whole genome sequencing identified and confirmed the presence of Campylobacter in humans and animals from the study area. Further analysis of the sequences was performed to determine multilocus sequence types and genetic determinants of antimicrobial resistance to fluoroquinolone, betalactam, tetracycline and macrolide classes of antimicrobials.ResultsFrom the human stool samples tested, 50 (9%) were positive of which 33 (66%) were C. jejuni, 14 (28%) were C. coli while 3 (6%) were C. hyointestinalis. In livestock, 132 (28%) were positive. Thirty one (7%) were C. jejuni while 101 (21%) were C. coli. Whole genome sequencing and MLST of the isolates revealed a total of 32 sequence types (STs) identified from 47 human isolates while 48 STs were identified in 124 isolates from livestock indicating a population which was overall, genetically diverse with a few more dominant strains. The antimicrobial resistance profiles of the isolates indicated a higher prevalence of resistance in Campylobacter isolated from livestock than in humans. Generally, resistance was greatest for betalactams (42%) closely followed by fluoroquinolones (41%), tetracyclines (15%) and lastly macrolides (2%). Multidrug resistance to three or more antimicrobials was observed in 24 (13%) isolates from humans (n = 1, 4%) and chicken (n = 23, 96%).ConclusionsThis study has further contributed information about the epidemiology, genetic diversity and antimicrobial resistance profile of thermophilic Campylobacter in Nigeria.

Isolation and characterization of thermophilic Campylobacter species from geese raised in Kars region (Turkey) using cultural, molecular and mass spectrometry methods

Background:Thermophilic Campylobacters are found in the digestive tract of wild and domestic poultry and can be transmitted to humans following their fecal discharges. Aims:This study aimed to isolate thermophilic Campylobacter by culture from cloacal swabs of geese, commonly breeding in Kars region, and to identify the isolates by PCR and mass spectrometry. Antibiotics susceptibility and resistance genes of the isolates were also analysed. Methods:The study included 400 cloacal swab samples of clinically healthy geese. The samples were cultured on mCCDA medium following the pre-enrichment in Preston broth. Identification of the isolates was performed by phenotypic methods, PCR, and MALDI-TOF MS. Antibiotic susceptibility and resistance genes of the isolates were analysed with the disc diffusion method and PCR, respectively. Results:Thermophilic Campylobacter spp. were isolated from 157 (39.3%) samples. 151 (96.2%) isolates were identified Campylobacter jejuni and 6 (3.8%) Campylobacter coli by the phenotypic tests and PCR. Among 125 isolates analysed by MALDI-TOF MS, 119 (95.2%) were identified C. jejuni and 6 (4.8%) C. coli. The isolates’ resistance to ampicillin, tetracycline, ciprofloxacin, gentamicin, and azithromycin were found 33.8%, 41.4%, 75.2%, 12.1%, and 7.6%, respectively. The distributions of blaOXA61, tetO, gyrA, and aphA-3 genes were 3.2%, 90.8%, 50.8%, and 52.7%, respectively. Conclusion:Since geese are raised in pastures in the Kars region, protecting and not polluting the existing natural environment and preventing their contact with wild birds will prevent the spread of these microorganisms.

Antibiotic resistance patterns of chicken and human origin Campylobacter spp. in Turkey

Campylobacteriosis is of great importance for both human and chicken populations. Unconcious and overuse of antibiotics in chickens has led to the transmission of antibiotic resistance patterns to humans. The purpose of this study was to determine the prevalence of thermophilic Campylobacter species from the cecal samples at slaughter houses, and also common antibiotic resistance patterns shared between chicken origin and human origin thermophilic Campylobacter species. Isolation and identification was performed according to EN ISO 10272-1: 2017 and Real-Time Multiplex qPCR, respectively. Antibiotic susceptibility test was performed by using the Kirby Bauer Disk Diffusion Method. Of the examined randomly collected 180 cecal samples at evisceration stage in slaughterhouses, 19 (10.5%), 17 (9.44%) and 2 (1.11%) were found to harbour Campylobacter spp., C. jejuni and C. coli, respectively. The highest resistance was determined against quinolones (86.04%) and fluoroquinolones (86.04%) among the tested 43 Campylobacter spp., comprising 19 chicken origin and 24 human origin. Except for erythromycin and gentamicin, all C. jejuni isolates from chickens and humans were found to be resistant to two or three of the antibiotics tested. The same multidrug resistance profiles observed in chicken origin C. jejuni isolates for TET/CIP/NA (70.58%) and CIP/NA (29.41%) were also determined in human origin C. jejuni isolates with the rate of 25% and 50%, respectively for each. To sum up, the same resistance patterns against common antibiotics shared in both human and chicken origin C. jejuni has pose a significant public health problem.

Occurrence of thermophilic Campylobacter spp. in pigs and the assessment of biosecurity measures employed at unorganized pig farms in Thrissur, Kerala

Campylobacter spp. is considered as one of the major causes of foodborne illnesses worldwide. A total of 130 samples including faecal samples (n=40), rectal swabs (n=40) and sewage samples (n=50) were collected from the two unorganized pig farms to study the occurrence of Campylobacter spp. The biosecurity measures on the farms were also assessed. An overall occurrence of 26.15 per cent with a higher rate of isolation from rectal swabs (57.5 per cent) than faecal and sewage samples (25 per cent and 2 per cent) were observed. The occurrence of C. coli was found to be 55 per cent, while that of C. jejuni and C. coli was 5 per cent in rectal swabs collected from Farm A. Campylobacter coli could be isolated only from the sewage sample from farm B. Direct multiplex PCR screening detected C. coli in 32 per cent and 44 per cent of sewage samples from farms A and B, respectively. This indicates that the Campylobacter organisms in sewage samples might have attained viable but not culturable form. In both farms, no effective biosecurity measures were followed. The lack of biosecurity measures in farms contributes to the transmission of Campylobacter spp. from the environment to the animals. Farm workers of both the farms were unaware of hygienic practices and biosecurity measures. Furthermore, little attention was paid to personal protective measures, which could pose a significant occupational risk of contracting campylobacteriosis, resulting in complex sequelae.

Isolation and Characterization of Group III Campylobacter jejuni-Specific Bacteriophages From Germany and Their Suitability for Use in Food Production.

Campylobacter spp. are a major cause of bacterial foodborne diarrhea worldwide. While thermophilic Campylobacter species asymptomatically colonize the intestines of chickens, most human infections in industrial countries have been attributed to consumption of chicken meat or cross-contaminated products. Bacteriophages (phages) are natural predators of bacteria and their use at different stages of the food production chain has been shown to reduce the public health burden of human campylobacteriosis. However, regarding regulatory issues, the use of lytic phages in food is still under discussion and evaluation. This study aims to identify lytic phages suitable for reducing Campylobacter bacteria along the food production chain. Therefore, four of 19 recently recovered phages were further characterized in detail for their lytic efficacy against different Campylobacter field strains and their suitability under food production settings at different temperatures and pH values. Based on the results of this study, the phages vB_CjM-LmqsCP1-4 and vB_CjM-LmqsCP1-5 appear to be promising candidates for the reduction of Campylobacter jejuni in food production settings.

Sensitivity to antibacterial preparations Campylobacter jejuni isolated from poultry products

Along with salmonellosis and Escherichiosis, intestinal campylobacteriosis caused by thermophilic campylobacter, primarily Campylobacter jejuni, is widespread. When studying the biological properties of isolated pathogens, great importance is attached to determining their sensitivity to antibacterial preparations and identifying preparation resistance. Livestock products, including poultry, are one of the most common sources of pathogenic campylobacter. Since antibiotics are often used in the process of growing and obtaining poultry meat, the acquired resistance of the campylobacter bacteria that colonize poultry products to the drugs used in veterinary medicine is likely. The spread of antibiotic-resistant strains in future can lead to decrease of the therapeutic effect in the treatment of severe forms of campylobacteriosis. In this article we present the results of determining the antibiotic resistance of C. jejuni cultures isolated from group samples of poultry products purchased in St. Petersburg in comparison with the antibiotic resistance of C. jejuni cultures isolated from group samples of chickens of the control groups, hatched without use of any antimicrobial preparations. Based on the results of our studies, we found that C. jejuni cultures isolated from group samples of poultry products purchased in St. Petersburg retail network have broad antibiotic resistance and are resistant to the effects of amoxicillin, ampicillin, carbenicillin, erythromycin, azithromycin, cephalothin, gentamycin, kanamycin, tetracycline, doxycycline, lincomycin, ciprofloxacin and co-trimoxazole, but are sensitive to the action of nalidixic acid, imipenem, meropenem, chloramphenicol and furazolidone. Cultures of C. jejuni isolated from group samples of poultry products purchased in St. Petersburg retail network have a broader antibiotic resistance to antimicrobial drugs of various pharmacological groups compared to cultures of C. jejuni isolated from group samples of chickens from the control group. Broader antibiotic resistance of C. jejuni cultures isolated from group samples of poultry products purchased in St. Petersburg retail network is most likely associated with the use of various antimicrobial drugs when growing poultry in industrial poultry farming.

Resistance to Antibiotics in Thermophilic Campylobacters.

Campylobacter jejuni (C. jejuni) is one of the most frequent causes of bacterial enterocolitis globally. The disease in human is usually self-limiting, but when complications arise antibiotic therapy is required at a time when resistance to antibiotics is increasing worldwide. Mechanisms of antibiotic resistance in bacteria are diverse depending on antibiotic type and usage and include: enzymatic destruction or drug inactivation; alteration of the target enzyme; alteration of cell membrane permeability; alteration of ribosome structure and alteration of the metabolic pathway(s). Resistance of Campylobacter spp. to antibiotics, especially fluoroquinolones is now a major public health problem in developed and developing countries. In this review the mechanisms of resistance to fluoroquinolones, macrolides, tetracycline, aminoglycoside and the role of integrons in resistance of Campylobacter (especially at the molecular level) are discussed, as well as the mechanisms of resistance to β-lactam antibiotics, sulphonamides and trimethoprim. Multiple drug resistance is an increasing problem for treatment of campylobacter infections and emergence of resistant strains and resistance are important One Health issues.

Prevalence, antibiotic spectrum and genetic relatedness in thermophilic Campylobacter species from poultry production environment of Punjab

Campylobacter is responsible for human gastroenteritis worldwide. C. jejuni and C. coli are most frequently encountered in animals, birds and man, but mainly C. jejuni is particularly adapted to poultry. The disease in humans ranges from gastroenteritis to Guillain-Barre Syndrome. To know the status of Campylobacter species especially, C. jejuni in poultry environment in Punjab, 342 poultry fecal samples (228 from layers and 114 from broilers) were collected from 30 poultry farms of Punjab. Additionally, 27 fecal samples from poultry farm workers were also collected. Analysis of the samples revealed that 21 (6.14%) fecal samples (from poultry farm) and two (7.41%) from farm workers were positive for Campylobacters based on cultural and molecular detection. Out of 23 Campylobacter positive isolates, 19 (80.95%) were C. jejuni and 4 (19.04%) were C. coli. Antibiotic resistance in the isolates was low. The majority of the isolates were sensitive to macrolide and quinolone class of antibiotics which are important for the treatment of campylobacteriosis. Pulse Field Gel Electrophoresis (PFGE) showed high genetic diversity among C. jejuni, however, it was limited in C. coli isolates. Poultry production in Punjab poses a potential risk of campylobacteriosis. However, antibiotic resistance in the isolates was low.

A REVIEW OF CURRENT KNOWLEDGE AND GAPS ABOUT CAMPYLOBACTER METHODS: FROM CULTURE TO CHARACTERIZATION

Campylobacteriosis is a zoonotic disease caused by Campylobacter, mostly associated with consumption of contaminated foodstuffs and water. Campylobacter jejuni and Campylobacter coli recognized as the leader of foodborne diarrheal illness in humans. The frequency of these microorganisms in poultry is fairly high than Salmonella and more challenging to measure, which represent an expensive burden charge on public health due to their difficulties to master them, especially with the fast increase rates of multidrug-resistant of thermophilic Campylobacter strains. It is well recognized that Campylobacter spp. is a fastidious cell, difficult to isolate in laboratories owing to their requirements and sensibility. That’s why; these factors must be taken into consideration during recovery protocols. A variety of phenotyping tests have been reported and widely used for confirmation and identification of Campylobacter species. Nonetheless, Whole Genome Sequencing (WGS) and Culture-Independent Diagnostic Tests (CIDTs) are new eras of hopeful technologies, mainly involved in the detection and characterization of threaten public health pathogens. This review aimed to describe the culture methods, phenotypic and genotypic schemes used to isolate, identify, and characterize Campylobacter isolates, through discussing the current knowledge and gaps related to the application of these techniques over others performed for typing this microaerophilic genus.

Challenging the “gold standard” of colony-forming units - Validation of a multiplex real-time PCR for quantification of viable Campylobacter spp. in meat rinses

Campylobacter jejuni is the leading bacterial food-borne pathogen in Europe. Despite the accepted limits of cultural detection of the fastidious bacterium, the “gold standard” in food microbiology is still the determination of colony-forming units (CFU). As an alternative, a live/dead differentiating qPCR has been established, using propidium monoazide (PMA) as DNA-intercalating crosslink agent for inactivating DNA from dead, membrane-compromised cells. The PMA treatment was combined with the addition of an internal sample process control (ISPC), i.e. a known number of dead C. sputorum cells to the samples. The ISPC enables i), monitoring the effective reduction of dead cell signal by the light-activated DNA-intercalating dye PMA, and ii), compensation for potential DNA losses during processing. Here, we optimized the method for routine application and performed a full validation of the method according to ISO 16140-2:2016(E) for the quantification of live thermophilic Campylobacter spp. in meat rinses against the classical enumeration method ISO 10272-2:2017. In order to render the method applicable and cost-effective for practical application, the ISPC was lyophilized to be distributable to routine laboratories. In addition, a triplex qPCR was established to simultaneously quantify thermophilic Campylobacter, the ISPC and an internal amplification control (IAC). Its performance was statistically similar to the two duplex qPCRs up to a contamination level of 4.7 log10Campylobacter per ml of meat rinse. The limit of quantification (LOQ) of the alternative method was around 20 genomic equivalents per PCR reaction, i.e. 2.3 log10 live Campylobacter per ml of sample. The alternative method passed a relative trueness study, confirming the robustness against different meat rinses, and displayed sufficient accuracy within the limits set in ISO 16140-2:2016(E). Finally, the method was validated in an interlaboratory ring trial, confirming that the alternative method was fit for purpose with a tendency of improved repeatability and reproducibility compared to the reference method for CFU determination.Campylobacter served as a model organism, challenging CFU as “gold standard” and could help in guidance to the general acceptance of live/dead differentiating qPCR methods for the detection of food-borne pathogens.

Occurrence, Risk Factors, and Antimicrobial Susceptibility Test of Thermophilic Campylobacter Species of Bovine Carcass at Municipal Abattoir and Butcher Shops of Jimma Town, Southwest Ethiopia.

BackgroundAlthough Campylobacter jejuni (C. jejuni) and Campylobacter coli (C. coli) cause acute diarrheal diseases in people all over the world, they are most commonly seen in other mammalian species and are a seemingly healthy carrier condition. Humans in Ethiopia, on the other hand, are largely unaware of the existence of Campylobacter in food animals as potential sources of infection.ObjectiveTo determine the occurrence, risk factors, and antimicrobial susceptibility pattern of thermophilic Campylobacter species in bovine raw meat from the abattoir and butcher shops.MethodsSwab samples were collected from 177 cattle carcasses (from abattoir 93 and butcher shops 84) and cultured using standard methods. An antimicrobial susceptibility test was performed using the disk diffusion method towards eleven antimicrobial agents.ResultsThe overall prevalence of thermophilic Campylobacter species was 14 (7.9%). From these, 11 (78.6%) were identified to be C. jejuni and 3 (21.4%) were C. coli. Lack of handwashing before meat processing and after visiting the toilet, meat contact with floors, walls, or soiling during preparation, and lack of training were the most important factors independently associated with (p<0.05) the prevalence of Campylobacter species contamination. The highest level of antimicrobial resistance of the Campylobacter isolates was recorded to ampicillin (10μg) (100%), followed by amoxicillin (30μg) (78.6%) and sulphamethoxazole-trimethoprim (57.1%) while the least resisted antimicrobials were streptomycin (25μg), erythromycin (15μg), oxytetracycline (30μg) (each 28.6%), kanamycin (30μg) 14.3%, chloramphenicol (30μg) and gentamycin (10μg) (each 7.1%).ConclusionDespite the low prevalence of thermophilic Campylobacter in the current investigation, it may pose a significant public health threat. As a result, it is vital to give retailers and customers extensive education, training, and knowledge about the correct handling and cooking of animal-derived goods. Furthermore, antimicrobials should be used with caution in both veterinary and human treatment regimens as well as a wider examination of antimicrobial resistance patterns for the use of well-targeted antimicrobials.

Characterization of Campylobacter spp. Strains Isolated From Wild Birds in Turkey.

Turkey is an important stopover site for many migrating birds between Europe, Asia and Africa. Campylobacter spp. are frequently found in wildlife, in particular waterfowl, and distinct strains are disseminated within this reservoir. In this study, 183 wild birds of hunting areas in Turkey were collected and thermophilic Campylobacter spp. from cloacal swabs were isolated at a prevalence of 5.2% from song thrushes (6/116) and 93% from Eurasian coots (41/44). After PCR species differentiation and flaA restriction profiles determination, C. jejuni and C. coli strains were further investigated by whole genome sequencing. PCR target amplification of the ceuE gene, commonly used for C. coli species-identification was inefficient and even hampered in one isolate. A close look on the ceuE sequence revealed that various mismatches in the ceuE oligo annealing sites caused less efficient diagnostic detection. All C. coli isolates belonged to the environmental clade II and clade III, for which thirty-six novel MLST types were identified. Further single nucleotide polymorphism (SNP) analysis showed a high genomic divergence between the C. coli isolates. High variability was also implicated for putative plasmid-located genes detected in 51% of the C. coli isolates. Distinct gene variants in clades II and III C. coli were identified by a k-mer analysis. After substracting k-mers in common with C. coli clade I database, 11 and 35 distinct genes were identified in clades II and III isolates, mainly involved in surface structures and modifications as well as signal transduction, suggesting niche adaptation of C. coli strains in wild birds. All strains were susceptible against (fluoro-)quinolones, erythromycin, tetracycline, gentamicin and only one isolate was resistant against streptomycin, suggesting that the sensitive phenotype was due to absence of selective pressure and niche separation in wild birds in Turkey. We conclude that Campylobacter spp. isolates from wildlife and environmental sources are still scarce in the databases and that there is a need for more studies on thermophilic Campylobacter spp. from different places all over the world in order to complement our understanding on dissemination and adaptation to distinct niches of this global food-borne pathogen.

Genomic Characterization of Fluoroquinolone-Resistant Thermophilic Campylobacter Strains Isolated from Layer Chicken Feces in Gangneung, South Korea by Whole-Genome Sequencing.

Thermophilic Campylobacter species of poultry origin have been associated with up to 80% of human campylobacteriosis cases. Layer chickens have received less attention as possible reservoirs of Campylobacter species. Initially, the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of two archived Campylobacter isolates (Campylobacter jejuni strain 200605 and Campylobacter coli strain 200606) from layer chickens to five antimicrobials (ciprofloxacin, nalidixic acid, erythromycin, tetracycline, and gentamicin) were determined using broth microdilution while the presence of selected antimicrobial resistance genes was performed by polymerase chain reaction (PCR) using specific primers. Whole-genome sequencing (WGS) was performed by the Illumina HiSeq X platform. The analysis involved antimicrobial resistance genes, virulome, multilocus sequence typing (MLST), and phylogeny. Both isolates were phenotypically resistant to ciprofloxacin (MIC: 32 vs. 32 µg/mL), nalidixic acid (MIC: 128 vs. 64 µg/mL), and tetracycline (MIC: 64 vs. 64 µg/mL), but sensitive to erythromycin (MIC: 1 vs. 2 µg/mL) and gentamicin (MIC: 0.25 vs. 1 µg/mL) for C. jejuni strain 200605 and C. coli strain 200606, respectively. WGS confirmed C257T mutation in the gyrA gene and the presence of cmeABC complex conferring resistance to FQs in both strains. Both strains also exhibited tet(O) genes associated with tetracycline resistance. Various virulence genes associated with motility, chemotaxis, and capsule formation were found in both isolates. However, the analysis of virulence genes showed that C. jejuni strain 200605 is more virulent than C. coli strain 200606. The MLST showed that C. jejuni strain 200605 belongs to sequence type ST-5229 while C. coli strain 200606 belongs to ST-5935, and both STs are less common. The phylogenetic analysis clustered C. jejuni strain 200605 along with other strains reported in Korea (CP028933 from chicken and CP014344 from human) while C. coli strain 200606 formed a separate cluster with C. coli (CP007181) from turkey. The WGS confirmed FQ-resistance in both strains and showed potential virulence of both strains. Further studies are recommended to understand the reasons behind the regional distribution (Korea, China, and Vietnam) of such rare STs.

Antimicrobial Resistance, FlaA Sequencing, and Phylogenetic Analysis of Campylobacter Isolates from Broiler Chicken Flocks in Greece.

Human campylobacteriosis caused by thermophilic Campylobacter species is the most commonly reported foodborne zoonosis. Consumption of contaminated poultry meat is regarded as the main source of human infection. This study was undertaken to determine the antimicrobial susceptibility and the molecular epidemiology of 205 Campylobacter isolates derived from Greek flocks slaughtered in three different slaughterhouses over a 14-month period. A total of 98.5% of the isolates were resistant to at least one antimicrobial agent. In terms of multidrug resistance, 11.7% of isolates were resistant to three or more groups of antimicrobials. Extremely high resistance to fluoroquinolones (89%), very high resistance to tetracycline (69%), and low resistance to macrolides (7%) were detected. FlaA sequencing was performed for the subtyping of 64 C. jejuni and 58 C. coli isolates. No prevalence of a specific flaA type was observed, indicating the genetic diversity of the isolates, while some flaA types were found to share similar antimicrobial resistance patterns. Phylogenetic trees were constructed using the neighbor-joining method. Seven clusters of the C. jejuni phylogenetic tree and three clusters of the C. coli tree were considered significant with bootstrap values >75%. Some isolates clustered together were originated from the same or adjacent farms, indicating transmission via personnel or shared equipment. These results are important and help further the understanding of the molecular epidemiology and antimicrobial resistance of Campylobacter spp. derived from poultry in Greece.

Antimicrobial Resistance Profiles, Virulence Genes, and Genetic Diversity of Thermophilic Campylobacter Species Isolated From a Layer Poultry Farm in Korea.

Thermophilic Campylobacter species are among the major etiologies of bacterial enteritis globally. This study aimed at assessing the antimicrobial resistance (AMR) profiles, virulence genes, and genetic diversity of thermophilic Campylobacter species isolated from a layer poultry farm in South Korea. One hundred fifty-three chicken feces were collected from two layer poultry farms in Gangneung, South Korea. The Campylobacter species were isolated by cultural techniques, while PCR and sequencing were used for species confirmation. Antimicrobial susceptibility testing for six antimicrobials [ciprofloxacin (CIP), nalidixic acid (NAL), sitafloxacin (SIT), erythromycin (ERY), tetracycline (TET), and gentamicin (GEN)] was carried out by broth microdilution. Three AMR and nine virulence genes were screened by PCR. Genotyping was performed by flaA-restriction fragment length polymorphism (RFLP) and multilocus sequence typing (MLST). Of the 153 samples, Campylobacter spp. were detected in 55 (35.9%), with Campylobacter jejuni and Campylobacter coli being 49 (89.1%) and six (10.9%), respectively. High-level resistance was observed for CIP (100%), NAL (100%), and TET (C. jejuni, 93.9%; C. coli: 83.3%). No resistance was observed for SIT. The missense mutation (C257T) in gyrA gene was confirmed by sequencing, while the tet(O) gene was similar to known sequences in GenBank. The rate of multidrug-resistant (MDR) strains was 8.2%, and they all belonged to C. jejuni. All Campylobacter isolates possessed five virulence genes (cdtB, cstII, flaA, cadF, and dnaJ), but none possessed ggt, while the rates for other genes (csrA, ciaB, and pldA) ranged between 33.3 and 95.9%. The flaA-RFLP yielded 26 flaA types (C. jejuni: 21 and C. coli: five), while the MLST showed 10 sequence types (STs) for C. jejuni and three STs for C. coli, with CC-607 (STs 3611) and CC-460 (ST-460) being predominant. Among the 10 STs of C. jejuni, three were newly assigned. The findings of this study highlight the increased resistance to quinolones and TET, the virulence potential, and the diverse genotypes among Campylobacter strains isolated from the layer poultry farm.

Antimicrobial Susceptibility and Genomic Analysis of Aliarcobacter cibarius and Aliarcobacter thereius, Two Rarely Detected Aliarcobacter Species.

Aliarcobacter cibarius and Aliarcobacter thereius are two rarely detected Aliarcobacter species. In the study, we analyzed the antimicrobial susceptibility and provide detailed insights into the genotype and phylogeny of both species using whole-genome sequencing. Thermophilic Campylobacter species are the most common bacterial foodborne pathogens causing gastroenteritis in humans worldwide. The genus Aliarcobacter is part of the Campylobacteraceae family and includes the species Aliarcobacter butzleri, Aliarcobacter cryaerophilus, Aliarcobacter skirrowii, and the rarely described Aliarcobacter cibarius, Aliarcobacter faecis, Aliarcobacter lanthieri, Aliarcobacter thereius, and Acrobarter trophiarum. Aliarcobacter are emergent enteropathogens and potential zoonotic agents. Here, we generated, analyzed, and characterized whole-genome sequences of Aliarcobacter cibarius and Aliarcobacter thereius. They were isolated from water poultry farms in Germany, cultured and identified by MALDI-TOF MS. With PCR the identity was verified. Antibiotic susceptibility testing was carried out with erythromycin, ciprofloxacin, doxycycline, tetracycline, gentamicin, streptomycin, ampicillin, and cefotaxime using the gradient strip method (E-test). Whole-genome sequences were generated including those of reference strains. Complete genomes for six selected strains are reported. These provide detailed insights into the genotype. With these, we predicted in silico known AMR genes, virulence-associated genes, and plasmid replicons. Phenotypic analysis of resistance showed differences between the presence of resistance genes and the prediction of phenotypic resistance profiles. In Aliarcobacter butzleri, the nucleotide sequence of the gyrA gene (DQ464331) can show a signature mutation resulting in an amino acid change T85>I. Acrobarter cibarius and Acrobarter thereius showed the same gene as assessed by similarity annotation of the mutations 254C>G. Most of the isolates were found to be sensitive to ciprofloxacin. The ciprofloxacin-resistant Aliarcobacter thereius isolate was associated with the amino acid change T85>I. But this was not predicted with antibiotic resistance databases, before. Ultimately, a phylogenetic analysis was done to facilitate in future outbreak analysis.

Biotyping of Multidrug Resistant &amp;lt;i&amp;gt;Campylobacter jejuni&amp;lt;/i&amp;gt; from Poultry and Humans in Northern Region of Ghana

Thermophilic Campylobacters are zoonotic bacteria which are universally famous for causing enteritis in humans. They are normally found as commensals in the digestive tract of food animals with poultry being a major reservoir of the pathogenic species. This study highlighted the presence of Campylobacter in poultry (commercial and domestic) and humans (patients and asymptomatic individuals) and characterized strains by biotyping and susceptibility test in the Northern region of Ghana where animal husbandry is commonly practiced but zoonotic studies are limited. A total of 1087 specimens from stools of humans and cloacal swabs of poultry were screened from 25th October, 2017 to 7th May, 2019. Samples were cultured on modified charcoal-cefoperazone-deoxycholate agar and isolates identified using standard microbiological procedures and Lior Biotyping, while the antibiogram of isolates was determined by the Kirby-Bauer disk diffusion method. The frequency of Campylobacter was 43.1% and 12.9% respectively in poultry and humans. Campylobacter jejuni were recovered from 84% of commercial birds and 64% of domestic birds and in humans significantly fewer strains were observed in patients than asymptomatic individuals (p 0.05). Biotype distribution revealed C. jejuni biotype I prevalence in domestic birds, patients and asymptomatic individuals whereas Bioytype II was largely found in commercial poultry. All isolated strains of Campylobacter were resistant to tetracycline and 69.4% of Campylobacter jejuni strains were resistant to erythromycin. Imipenem and the aminoglycosides were relatively effective as resistance of 10% and below 20% were respectively obtained. None of the endorsed treatment drugs (erythromycin, ciprofloxacin, and tetracycline) can be admitted in this region due to common resistance found among strains against these agents.