Comparative Assessment of the Antibacterial and Antibiofilm Actions of Benzalkonium Chloride, Erythromycin, and L(+)-Lactic Acid against Raw Chicken Meat Campylobacter spp. Isolates.

Campylobacter spp. are significant zoonotic agents, which cause annually millions of human cases of foodborne gastroenteritis worldwide. Their inclusion in biofilms on abiotic surfaces seems to play a pivotal role in their survival outside of the host, growth, and spread. To successfully mitigate the risks that arise with these bacteria, it is crucial to decrease their prevalence within the food production chain (from farm to the table), alongside the successful treatment of the resulting illness, known as campylobacteriosis. For this, the use of various antimicrobial agents remains actively in the foreground. A general-purpose biocide and cationic surfactant (benzalkonium chloride; BAC), a widely used macrolide antibiotic (erythromycin; ERY), and a naturally occurring organic acid (L(+)-lactic acid; LA) were comparatively evaluated in this work for their potential to inhibit both the planktonic and biofilm growth of 12 selected Campylobacter spp. (of which, seven were C. jejuni and five were C. coli) raw chicken meat isolates, all grown in vitro as monocultures. The inhibitory action of LA was also studied against four mixed-culture Campylobacter biofilms (each composed of three different isolates). The results showed that the individual effectiveness of the agents varied significantly depending on the isolate, growth mode (planktonic, biofilm), intercellular interactions (monocultures, mixed cultures), and the growth medium used (with special focus on blood presence). Thus, BAC exhibited minimum inhibitory concentrations (MICs), minimum bactericidal concentrations (MBCs), and minimum biofilm inhibitory concentrations (MBICs) that ranged from 0.5 to 16 µg/mL. Interestingly enough, these values varied widely from 0.25 to 1024 µg/mL for ERY. Concerning LA, the MICs, MBCs, and MBICs varied from 1024 to 4096 µg/mL, with mixed-culture biofilm formation always being more difficult to suppress when compared to biofilm monocultures. In addition, it was evident that intercellular interactions encountered within mixed-culture Campylobacter biofilms significantly influenced both the population dynamics and the tolerance of each consortium member to acid exposure. Overall, the findings of this study provide useful information on the comparative effectiveness of three well-known antimicrobial agents for the control of Campylobacter spp. under various growth modes (i.e., planktonic, biofilm, monocultures, mixed cultures) that could potentially be encountered in food production and clinical settings.

Human Campylobacter spp. infections in Italy.

PURPOSE: Campylobacter is a frequent cause of enteric infections with common antimicrobial resistance issues. The most recent reports of campylobacteriosis in Italy include data from 2013 to 2016. We aimed to provide national epidemiological and microbiological data on human Campylobacter infections in Italy during the period 2017-2021. METHODS: Data was collected from 19 Hospitals in 13 Italian Regions. Bacterial identification was performed by mass spectrometry. Antibiograms were determined with Etest or Kirby-Bauer (EUCAST criteria). RESULTS: In total, 5419 isolations of Campylobacter spp. were performed. The most common species were C. jejuni (n = 4535, 83.7%), followed by C. coli (n = 732, 13.5%) and C. fetus (n = 34, 0.6%). The mean age of patients was 34.61 years and 57.1% were males. Outpatients accounted for 54% of the cases detected. Campylobacter were isolated from faeces in 97.3% of cases and in 2.7% from blood. C. fetus was mostly isolated from blood (88.2% of cases). We tested for antimicrobial susceptibility 4627 isolates (85.4%). Resistance to ciprofloxacin and tetracyclines was 75.5% and 54.8%, respectively; resistance to erythromycin was 4.8%; clarithromycin 2% and azithromycin 2%. 50% of C. jejuni and C. coli were resistant to ≥ 2 antibiotics. Over the study period, resistance to ciprofloxacin and tetracyclines significantly decreased (p < 0.005), while resistance to macrolides remained stable. CONCLUSION: Campylobacter resistance to fluoroquinolones and tetracyclines in Italy is decreasing but is still high, while macrolides retain good activity.

Multiplex Real-Time PCR for the Detection of Tetracycline, Ciprofloxacin, and Erythromycin Resistance Determinants from Human and Foodborne Campylobacter jejuni and Campylobacter coli.

Campylobacter jejuni and Campylobacter coli are the predominant thermophilic species responsible for foodborne gastroenteritis worldwide. Elevated resistance to certain antibiotics was observed due to antimicrobial therapy in farm animals and humans, while reduced antimicrobial usage partially reduced antibiotic resistance. Monitoring the antimicrobial resistance demonstrated a substantial fraction of multi-resistant isolates, indicating the necessity of reliable tools for their detection. In this study, resistance determinants in 129 German and 21 Vietnamese isolates were selected to establish a novel multiplex real-time PCR (qPCR), facilitating the simultaneous detection of four resistance determinants. These comprised tet(O) gene variants associated with tetracycline resistance, point mutations GyrA_T86I and GyrA_T86V associated with ciprofloxacin resistance, and the erm(B) gene together with the point mutation A2075G in the 23S rRNA gene, associated with erythromycin resistance. Moreover, the performance of the qPCR assay was evaluated by comparing the results of qPCR to phenotypic antimicrobial resistance profiles, obtained with standardized EUCAMP3 microdilution panel, which showed 100% similarity (inclusivity and exclusivity). Variation in measurement methods, including qPCR machines and master mixes showed robustness, essential for laboratories. The assay can be used for the rapid detection of resistance determinants, and is beneficial for monitoring the spread of antibiotic resistance in C. jejuni and C. coli.

Prevalence and genetic characterization of Campylobacter from clinical poultry cases in China.

IMPORTANCE: Campylobacter is a major cause of campylobacteriosis worldwide, and poultry is the main reservoir for its transmission. Campylobacter was generally considered to be a harmless commensal organism in poultry without pathogenic properties. However, it was proposed that a Campylobacter-like organism may be the cause of vibrionic hepatitis, which poses a significant public health risk. The occurrence and epidemiology of Campylobacter in healthy poultry have been studied systematically, but little is known about the epidemiology of Campylobacter isolates from diseased poultry in China. Therefore, this study determined the prevalence and molecular characterization of Campylobacter from diseased chickens, ducks, and geese in Yangzhou Veterinary Hospital between December 2016 and September 2017, which was critical for improving the diagnosis and prevention of Campylobacter infections.

Bio-Mapping of Microbial Indicators and Pathogen Quantitative Loads in Commercial Broiler Processing Facilities in South America.

A bio-mapping study was conducted with the aim of creating a microbiological baseline on indicator organisms and pathogens in commercial broiler processing facilities located in a country in South America. Whole chicken carcass and wing rinses were collected from five stages of the poultry processing line: live receiving (LR), rehanger (R), post-evisceration (PE), post-chilling (PC), and wings (W). Rinses (n = 150) were enumerated using the MicroSnap™ system for total viable counts (TVC) and Enterobacteriaceae (EB), while the BAX®-System-SalQuant® and BAX®-System-CampyQuant™ were used for Salmonella and Campylobacter, respectively. TVC and EB were significantly different between stages at the processing line (p < 0.01). There was a significant reduction from LR to PC for both microbial indicators. TVC and EB counts increased significantly from PC to W. Salmonella counts at PC were significantly different from the other stages at the processing line (p = 0.03). Campylobacter counts were significantly higher than the other stages at PC (p < 0.01). The development of bio-mapping baselines with microbial indicators showed consistent reduction up to the post-chilling stage, followed by an increase at the wings sampling location. The quantification of pathogens demonstrates that prevalence analysis as a sole measurement of food safety is not sufficient to evaluate the performance of processing operations and sanitary dressing procedures in commercial processing facilities.

Campylobacter jejuni and Campylobacter coli in human stool samples: antibiotic resistance profiles, putative virulence determinants and molecular characterization of the isolates.

Campylobacters, especially C. jejuni and C. coli, have become one of the leading causes of acute gastroenteritis in humans worldwide in recent years. We aimed to investigate the presence, antimicrobial resistance, putative virulence genes, and molecular characterization of C. jejuni and C. coli recovered from human acute gastroenteritis cases in the study. In the study, suspected Campylobacter spp. isolates were obtained in 43 (5%) feces samples collected from a total of 850 patients who applied to the Erciyes University Medical Faculty acute clinic between March 2019 and February 2020. As a result of the phenotypic tests, these isolates were determined to be Campylobacter spp. According to the multiplex PCR, 33 of 43 Campylobacter spp. isolates were identified as C. jejuni (76%) and ten isolates were as C. coli (24%). In the disc diffusion test, the highest resistance rate was found in the trimethoprim/sulfamethoxazole (90.1%) and ciprofloxacin (90.1%), and the lowest rate was found in the amoxicillin-clavulanic acid (9.3%). In Campylobacter spp. isolates, the virulence genes cdtA, virB11, cdtB, cadF, iam, ceu, and flaA were found to be positive at rates of 26 (60%), 28 (65.6%), 13 (30%), 4 (9%), 27 (62%), 17 (39%), and 7 (16%), respectively. However, the cdtC gene was not detected in any of the isolates. The study searched tetO gene to examine the genetic aspect of tetracycline resistance, which was found in all Campylobacter spp. isolates. In the PCR reactions to investigate A2074C and A2075G mutations of macrolide resistance, it was determined as 7 (16%) and 21 (48%) of the isolates. To detect quinolone resistance, the rates of quinolone resistance-determining regions (QRDR) were 20 (45.4%) and the gyrA gene mutations in the mismatch amplification mutation assay PCR (MAMA-PCR), were 19 (43.1%) of isolates. In addition, the quinolone resistance gene (qnr) carried by plasmid in Campylobacter isolates was not found in the study. BlaOXA-61 and CmeB (multi-drug efflux pump) genes were detected as 28 (63.6%) and 30 (68.1), respectively. The Enterobacterial Repetitive Intergenic Consensus PCR (ERIC-PCR) used for typing the isolates revealed that the band profiles obtained from the isolates were different. In conclusion, this was a very comprehensive study revealing the presence of antibiotic-resistant C. jejuni and C. coli with various virulence genes in patients admitted to a university hospital with acute gastroenteritis. This is of utmost significance for public health. Since campylobacteria are foodborne, zoonotic pathogens and transmission occurs mostly through food. People should have detailed information about the transmission routes of campylobacteria and risky foods. In addition, staff, food processors and caterers, should be trained in hygiene.

Investigating the Significance of Non-jejuni/coli Campylobacter Strains in Patients with Diarrhea.

Campylobacter is one of the most commonly reported foodborne bacteria worldwide. Although Campylobacter jejuni and Campylobacter coli have been reported to be responsible for the great majority of campylobacteriosis, the burden of infections by species other than C. jejuni and C. coli have been increasing as a result of a transition to diagnostic test methods that enable the isolation of emerging species. The aim of the present study was to recover C. jejuni, C. coli, and emerging species from the stool samples of 500 patients with gastroenteritis and 100 healthy subjects via the use of a filtration method and culture techniques using Butzler agar and mCCDA under a microaerobic or hydrogen-enriched atmosphere, identify the species by multiplex PCR methods and assess the significance of emerging species in enteric diseases. Thirty-one (6.2%) Campylobacter spp. were isolated from the stool samples of diarrheic patients but none from healthy individuals. Of 31 isolates, 21 (67.8%), nine (29%), and one (3.2%) were identified as C. jejuni, C. coli, and Campylobacter concisus by multiplex PCR, respectively. The filtration method was superior to the culture technique using mCCDA under a microaerobic atmosphere. C. concisus was evaluated as the etiology of gastroenteritis as a result of laboratory and clinical evaluations. The present study was the first to indicate that emerging Campylobacter species are rarely detected and C. concisus is linked to acute gastroenteritis in Turkey where additional studies are warranted to clarify the significance of emerging species in gastroenteritis.

Wastewater-based epidemiology of Campylobacter spp.: A systematic review and meta-analysis of influent, effluent, and removal of wastewater treatment plants.

Campylobacter spp. is one of the four leading causes of diarrhoeal diseases worldwide, which are generally mild but can be fatal in children, the elderly, and immunosuppressed persons. The existing disease surveillance for Campylobacter infections is usually based on untimely clinical reports. Wastewater surveillance or wastewater-based epidemiology (WBE) has been developed for the early warning of disease outbreaks and the detection of the emerging new variants of human pathogens, especially after the global pandemic of COVID-19. However, the WBE monitoring of Campylobacter infections in communities is rare due to a few large data gaps. This study is a meta-analysis and systematic review of the prevalence of Campylobacter spp. in various wastewater samples, primarily the influent of wastewater treatment plants. The results showed that the overall prevalence of Campylobacter spp. was 53.26 % in influent wastewater and 52.97 % in all types of wastewater samples. The mean concentration in the influent was 3.31 ± 0.39 log10 gene copies or most probable number (MPN) per 100 mL. The detection method combining culture and PCR yielded the highest positive rate of 90.86 %, while RT-qPCR and qPCR were the two most frequently used quantification methods. In addition, the Campylobacter concentration in influent wastewater showed a seasonal fluctuation, with the highest concentration in the autumn at 3.46 ± 0.41 log10 gene copies or MPN per 100 mL. Based on the isolates of all positive samples, Campylobacter jejuni (62.34 %) was identified as the most prevalent species in wastewater, followed by Campylobacter coli (30.85 %) and Campylobacter lari (4.4 %). These findings provided significant data to further develop and optimize the wastewater surveillance of Campylobacter spp. infections. In addition, large data gaps were found in the decay of Campylobacter spp. in wastewater, indicating insufficient research on the persistence of Campylobacter spp. in wastewater.

Comparative genomic analysis of Campylobacter hepaticus genomes associated with spotty liver disease, Georgia, United States.

Campylobacter hepaticus has re-emerged as an important cause of disease in egg laying birds worldwide, resulting in morbidity, mortality, and significant losses in eggs for the breeding and table egg laying industries. Although birds may appear asymptomatic, the disease is characterized by spots on the liver of birds and histopathological analysis reveals multifocal fibrogranulocytic necrotizing hepatitis microscopically. The re-emergence of C. hepaticus may be linked with housing practices as the disease appears more prevalent in pasture raised birds with outside exposure. Here we describe, the whole genome sequences and comparative analysis of four C. hepaticus genomes associated with an outbreak on pasture raised breeders from a farm in Georgia, United States. All four genomes were relatively similar in size and virulence genes harbored. Using these genomes, comparison with current C. hepaticus genomes available in NCBI and other databases and other members of the Campylobacter species was carried out. Using current tools available, virulence gene factor content was compared, and it was found that different tools lead to different numbers of factors identified. The four genomes from this study were relatively similar to C. hepaticus HV10 the type strain from Australia but differed from the other sequenced US strains from Iowa and Florida. C. hepaticus was found to have an overall lower gene content for genes associated with virulence and iron acquisition compared to other Campylobacter genomes and appears to cluster differently than UK genomes on phylogenetic analysis, suggesting the emergence of two lineages of C. hepaticus. This analysis provides valuable insight into the emerging pathogen C. hepaticus, its virulence factors and traits associated with disease in poultry production in the US, potentially providing insight into targets for its control and treatment for laying birds. Our analysis also confirms genes associated with iron acquisition are limited and the presence of the multidrug efflux pump CmeABC in C. hepaticus which may promote survival and persistence in the host niche - the chicken liver/bile. One unique aspect of this study was the finding of a close genetic relationship between C. hepaticus and Campylobacter fetus species and evidence of genome reduction in relation to host niche specificity.

Current State of Knowledge Regarding WHO High Priority Pathogens-Resistance Mechanisms and Proposed Solutions through Candidates Such as Essential Oils: A Systematic Review.

Combating antimicrobial resistance (AMR) is among the 10 global health issues identified by the World Health Organization (WHO) in 2021. While AMR is a naturally occurring process, the inappropriate use of antibiotics in different settings and legislative gaps has led to its rapid progression. As a result, AMR has grown into a serious global menace that impacts not only humans but also animals and, ultimately, the entire environment. Thus, effective prophylactic measures, as well as more potent and non-toxic antimicrobial agents, are pressingly needed. The antimicrobial activity of essential oils (EOs) is supported by consistent research in the field. Although EOs have been used for centuries, they are newcomers when it comes to managing infections in clinical settings; it is mainly because methodological settings are largely non-overlapping and there are insufficient data regarding EOs' in vivo activity and toxicity. This review considers the concept of AMR and its main determinants, the modality by which the issue has been globally addressed and the potential of EOs as alternative or auxiliary therapy. The focus is shifted towards the pathogenesis, mechanism of resistance and activity of several EOs against the six high priority pathogens listed by WHO in 2017, for which new therapeutic solutions are pressingly required.

Light-Up Split Broccoli Aptamer as a Versatile Tool for RNA Assembly Monitoring in Cell-Free TX-TL Systems, Hybrid RNA/DNA Origami Tagging and DNA Biosensing.

Binary light-up aptamers are intriguing and emerging tools with potential in different fields. Herein, we demonstrate the versatility of a split Broccoli aptamer system able to turn on the fluorescence signal only in the presence of a complementary sequence. First, an RNA three-way junction harbouring the split system is assembled in an E. coli-based cell-free TX-TL system where the folding of the functional aptamer is demonstrated. Then, the same strategy is introduced into a 'bio-orthogonal' hybrid RNA/DNA rectangle origami characterized by atomic force microscopy: the activation of the split system through the origami self-assembly is demonstrated. Finally, our system is successfully used to detect the femtomoles of a Campylobacter spp. DNA target sequence. Potential applications of our system include the real-time monitoring of the self-assembly of nucleic-acid-based devices in vivo and of the intracellular delivery of therapeutic nanostructures, as well as the in vitro and in vivo detection of different DNA/RNA targets.

Application of TraDIS to define the core essential genome of Campylobacter jejuni and Campylobacter coli.

Campylobacter species are the major cause of bacterial gastroenteritis. As there is no effective vaccine, combined with the rapid increase in antimicrobial resistant strains, there is a need to identify new targets for intervention. Essential genes are those that are necessary for growth and/or survival, making these attractive targets. In this study, comprehensive transposon mutant libraries were created in six C. jejuni strains, four C. coli strains and one C. lari and C. hyointestinalis strain, allowing for those genes that cannot tolerate a transposon insertion being called as essential. Comparison of essential gene lists using core genome analysis can highlight those genes which are common across multiple strains and/or species. Comparison of C. jejuni and C. coli, the two species that cause the most disease, identified 316 essential genes. Genes of interest highlighted members of the purine pathway being essential for C. jejuni whilst also finding that a functional potassium uptake system is essential. Protein-protein interaction networks using these essential gene lists also highlighted proteins in the purine pathway being major 'hub' proteins which have a large number of interactors across the network. When adding in two more species (C. lari and C. hyointestinalis) the essential gene list reduces to 261. Within these 261 essential genes, there are many genes that have been found to be essential in other bacteria. These include htrB and PEB4, which have previously been found as core virulence genes across Campylobacter species in other studies. There were 21 genes which have no known function with eight of these being associated with the membrane. These surface-associated essential genes may provide attractive targets. The essential gene lists presented will help to prioritise targets for the development of novel therapeutic and preventative interventions.

The potential role of migratory birds in the transmission of pathogenic Campylobacter species to broiler chickens in broiler poultry farms and live bird markets.

BACKGROUND: Campylobacter species (spp.) are one of the most important zoonotic bacteria possessing potential hazards for animal and human health worldwide. Migratory birds are implicated as significant carriers for microbes and a play very important role in the dissemination of Campylobacter to broiler chickens and their environment. The purpose of this investigation was to detect the prevalence, antibiotic resistant patterns, virulence and diversity of pathogenic Campylobacter spp. in 7 migratory bird species (Northern shoveler, Common pochard, Common teal, Northern pintail, Eared Grebe, Great Crested Grebe and Garganey) and broiler chickens that were collected from broiler poultry farms and live bird markets. RESULTS: The prevalence of Campylobacter was 12.5% (25/200), of which 15% (15/100) was recovered from 5 migratory bird species only and 10% (10/100) from broiler chickens. At the level of migratory birds, eight isolates (53.3%) were Campylobacter jejuni (C. jejuni) and 7 isolates (46.7%) were Campylobacter coli (C. coli) meanwhile, in broiler chickens C. jejuni and C. coli were 50% (5/10) for each. All isolated strains had phenotypic resistance to doxycycline, while all of the isolates were susceptible to amikacin. The multidrug resistance to three, four or five antimicrobial classes was found in 72% (18/25) of the isolated strains. The multiantibiotic resistance index between the examined isolates was 0.22-0.77, with 10 antibiotic resistance patterns. The virulence of isolated Campylobacter strains (from both migratory birds and broiler chicken birds) was detected by targeting the VirB11, ciaB and iam genes which were recorded at 16%, 52% and 100%, respectively. Additionally, 100% and 84% of the antibiotic resistance genes were identified as tetA and BlaOXA-61, respectively. CONCLUSIONS: The results of this study revealed the diversity between all the isolated strains from migratory birds and their similarity to broiler chicken isolates. The findings of the present study highlight the impact of migratory birds visiting Egypt and other countries on pathogenic Campylobacter spp. carrying pathogenic virulence and resistance genes, necessitating the application of biosecurity measures to prevent migratory birds from entering farms during their migration period.

Gold Nanoparticle and Polymerase Chain Reaction (PCR)-Based Colorimetric Assay for the Identification of Campylobacter spp. in Chicken Carcass.

Campylobacteriosis is a common cause of gastrointestinal disease. In this study, we suggest a general strategy of applying gold nanoparticles (AuNPs) in colorimetric biosensors to detect Campylobacter in chicken carcass. Polymerase chain reaction (PCR) was utilized for the amplification of the target genes, and the thiolated PCR products were collected. Following the blending of colloid AuNPs with PCR products, the thiol bound to the surface of AuNPs, forming AuNP-PCR products. The PCR products had a sufficient negative charge, which enabled AuNPs to maintain a dispersed formation under electrostatic repulsion. This platform presented a color change as AuNPs aggregate. It did not need additional time and optimization of pH for PCR amplicons to adhere to the AuNPs. The specificity of AuNPs of modified primer pairs for mapA from Campylobacter jejuni and ceuE from Campylobacter coli was activated perfectly (C. jejuni, p-value: 0.0085; C. coli, p-value: 0.0239) when compared to Salmonella Enteritidis and Escherichia coli as non-Campylobacter species. Likewise, C. jejuni was successfully detected from artificially contaminated chicken carcass samples. According to the sensitivity test, at least 15 ng/µL of Campylobacter PCR products or 1×103 CFU/mL of cells in the broth was needed for the detection using the optical method.

Multicenter Retrospective Study of Vascular Infections and Endocarditis Caused by Campylobacter spp., France.

The incidence of campylobacteriosis has substantially increased over the past decade, notably in France. Secondary localizations complicating invasive infections are poorly described. We aimed to describe vascular infection or endocarditis caused by Campylobacter spp. We included 57 patients from a nationwide 5-year retrospective study on Campylobacter spp. bacteremia conducted in France; 44 patients had vascular infections, 12 had endocarditis, and 1 had both conditions. Campylobacter fetus was the most frequently involved species (83%). Antibiotic treatment involved a ß-lactam monotherapy (54%) or was combined with a fluoroquinolone or an aminoglycoside (44%). The mortality rate was 25%. Relapse occurred in 8% of cases and was associated with delayed initiation of an efficient antimicrobial therapy after the first symptoms, diabetes, and coexistence of an osteoarticular location. Cardiovascular Campylobacter spp. infections are associated with a high mortality rate. Systematically searching for those localizations in cases of C. fetus bacteremia may be warranted.

Genomic characterization of molecular markers associated with antimicrobial resistance and virulence of the prevalent Campylobacter coli isolated from retail chicken meat in the United Arab Emirates.

Campylobacter is a major cause of gastroenteritis worldwide, with broiler meat accounting for most illnesses. Antimicrobial intervention is recommended in severe cases of campylobacteriosis. The emergence of antimicrobial resistance (AMR) in Campylobacter is a concerning food safety challenge, and monitoring the trends of AMR is vital for a better risk assessment. This study aimed to characterize the phenotypic profiles and molecular markers of AMR and virulence in the prevalent Campylobacter species contaminating chilled chicken carcasses sampled from supermarkets in the United Arab Emirates (UAE). Campylobacter was detected in 90 (28.6%) out of 315 tested samples, and up to five isolates from each were confirmed using multiplex PCR. The species C. coli was detected in 83% (75/90) of the positive samples. Whole-genome sequencing was used to characterize the determinants of AMR and potential virulence genes in 45 non-redundant C. coli isolates. We identified nine resistance genes, including four associated with resistance to aminoglycoside (aph(3')-III, ant(6)-Ia, aph(2″)-Ib, and aac(6')-Im), and three associated with Beta-lactam resistance (blaOXA-61, blaOXA-193, and blaOXA-489), and two linked to tetracycline resistance (tet(O/32/O), and tet(O)), as well as point mutations in gyrA (fluoroquinolones resistance), 23S rRNA (macrolides resistance), and rpsL (streptomycin resistance) genes. A mutation in gyrA 2 p.T86I, conferring resistance to fluoroquinolones, was detected in 93% (42/45) of the isolates and showed a perfect match with the phenotype results. The simultaneous presence of blaOXA-61 and blaOXA-193 genes was identified in 86.6% (39/45) of the isolates. In silico analysis identified 7 to 11 virulence factors per each C. coli isolate. Some of these factors were prevalent in all examined strains and were associated with adherence (cadF, and jlpA), colonization and immune evasion (capsule biosynthesis and transport, lipooligosaccharide), and invasion (ciaB). This study provides the first published evidence from the UAE characterizing Campylobacter virulence, antimicrobial resistance genotype, and phenotype analysis from retail chicken. The prevalent C. coli in the UAE retail chicken carries multiple virulence genes and antimicrobial resistance markers and exhibits frequent phenotype resistance to macrolides, quinolones, and tetracyclines. The present investigation adds to the current knowledge on molecular epidemiology and AMR development in non-jejuni Campylobacter species in the Middle East and globally.

Relevance and Importance of Biofilms in the Resistance and Spreading of Campylobacter spp. Within the Food Chain.

Campylobacter spp. are Gram-negative microaerophilic and non-spore-forming bacteria that primarily live as commensal organisms in the gastrointestinal tract of many domestic and wild birds and mammals. These frequently provoke foodborne illness, being responsible for the most common cause of acute bacterial gastroenteritis worldwide. Although these are typically fastidious and slow-growing microorganisms which are sensitive to desiccation and other stresses (e.g., extreme pH, freezing, UV, disinfectants), these can still survive outside their host for long periods, with their attachment to surfaces and the formation of and/or inclusion into biofilms to be considered quite important for their environmental survival and widespread dissemination. In this chapter, the most representative studies on the existence, relevance, and importance of biofilms in the resistance and spreading of Campylobacter spp. within the food chain are presented. Hopefully, such accumulated and focused knowledge is expected to highlight the problem and trigger more effective ways for its mitigation, improving the safety of our food supply and protecting public health.

Quantitative assessment of Campylobacter spp. levels with real-time PCR methods at different stages of the broiler food chain.

The importance of thermotolerant Campylobacter spp. as a food-borne pathogen continues to increase and there is a great need for rapid quantitative results in routine diagnostics. However, currently, only the culture-based ISO method is authorized for use in the context of official food control. The present study therefore aimed to assess the suitability of a qPCR method for a rapid quantitative determination of Campylobacter spp. at different stages in the poultry production chain and its equivalence with the culture-based method. Samples from two processors were collected and evaluated both separately and together. Censored regression (Tobit) models have been used to establish a relationship between Campylobacter qPCR counts on the carcasses and explanatory variables of processor and meat counts. Further, correlations of qPCR Campylobacter spp. counts at the different stages of production were calculated. In addition, the comparative data between microbiological enumeration and qPCR results were statistically analyzed. In the correlation calculation of the qPCR results, a highly significant relationship between the Campylobacter spp. counts of the neck skin samples to breast fillet and leg samples could be calculated, indicating a good prediction of Campylobacter spp. loads in these samples. The intercalating dye ethidium monoazide (EMA) was used to see whether the correlations between microbiological counts and qPCR results were improved by pretreating fecal and cecal samples before qPCR analysis. It was shown that the observed values of scatter plots between the qPCR-based and the culture-based methods were strongly correlated. However, on average, the qPCR results were two log10 CFU/mL levels higher than the microbiological counts. However, the classical culture-based method for food hygiene risk assessment cannot be replaced one-to-one by the qPCR or EMA-qPCR. The qPCR method can rather be used for the rapid identification of particularly highly contaminated flocks.

Diagnostic accuracy of multiplex nucleic acid amplification tests for Campylobacter infection: a systematic review and meta-analysis.

Campylobacter infection is one of the most frequently reported foodborne diseases with approximately 230,000 and 1.5 million cases each year in Europe and the USA, respectively. Culture methods are the reference for the diagnosis of Campylobacter infections; however, these methods are complex and time-consuming. Multiplex nucleic acid amplification test is favored due to its rapidity, automatization in the procedure followed and the quick simultaneous testing of numerous foodborne pathogens. The aim of this meta-analysis was to evaluate the accuracy of these tests for the diagnosis of Campylobacter infection. Scopus, Science Direct, PubMed, Web of Science, and Mendeley were searched for peer-reviewed articles. The split component synthesis method with the use of the inverse variance heterogeneity model was chosen for the quantitative meta-analysis. Sensitivity analysis was performed by age category and index test. The literature search found 34 studies involving 28,105 patients with suspected gastroenteritis. The sensitivity and specificity were 95.3% (92.3; 97.1) and 97.1% (95.1; 98.3), respectively, and AUC (area under the curve) was 0.963 (0.947; 0.974). Pediatric patients had a lower sensitivity (87.4, 48.2; 98.1) and higher specificity (99.2, 91.6; 99.9) estimate compared to all ages category (sensitivity 95.3, 91.3; 97.5, specificity 96.7, 93.7; 98.3). Among the various index tests, Seeplex/Allplex and Amplidiag/Novodiag had the lowest estimate for sensitivity (88.9, 73.8; 95.8) and specificity (95.2, 86; 98.4), respectively. BDMax had the highest (sensitivity 98.1, 96.1; 99 and specificity 98.5, 97; 99.3). Multiplex nucleic acid tests showed excellent accuracy and could play an influential role in diagnosing Campylobacter infections.

Analysis of Campylobacter spp. contamination and drug resistance in poultry sold in Jiading District， Shanghai from 2019 to 2021 / 上海预防医学

ObjectiveTo investigate the contamination status and drug resistance of Campylobacter spp. in poultry sold in Jiading District， Shanghai. MethodsFour types of poultry meats （chickens， ducks， geese and pigeons） were sampled from commercial markets， and potential Campylobacter spp. contamination was isolated and identified. Furthermore， resistance of isolated Campylobacter spp. to 15 commonly used antibiotics was tested. ResultsTotally 29 Campylobacter jejuni strains and 34 Campylobacter. coli were isolated from 236 commercial poultry samples. The most severe contamination of Campylobacter spp. was found in chicken samples， with a detection rate of 34.04%， while the lowest detection rate of Campylobacter spp. was found in duck （19.67%）. Contamination status was categorized with different storage conditions. The lowest detection rate of 6.67% was noted under frozen condition， while highest detection rate of 41.27% was noted under cold storage. Campylobacter jejuni was completely resistant to cefazolin， ciprofloxacin， nalidixic acid and tetracycline， and Campylobacter coli was completely resistant to cefazolin， cefoxitin and nalidixic acid； Campylobacter spp. showed the lowest resistance to imipenem. Multi-drug resistant strains accounted for 100.00% of the isolated strains. 96.83% of the strains were resistant to more than 5 drugs， with the highest number reaching 14 kinds of antibiotics. ConclusionThere is a significant difference in the contamination status and drug resistance of Campylobacter spp. isolated from four types of poultry meats sold in Jiading District， Shanghai， and the drug resistance is serious. It is strongly recommended that the use of antibiotics should be strictly controlled. Freezing can effectively reduce Campylobacter spp. pollution.