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Background Campylobacter is one of the leading causes of human bacterial gastroenteritis worldwide. Campylobacter infections are most often associated with the consumption of raw milk, undercooked poultry, and contaminated water. Objective The RapidChek®Campylobacter test system (PTM number 052201) was validated for the detection of Campylobacter jejuni, C. coli, and C. lari in raw ground chicken, chicken carcass rinse, and turkey carcass sponges. Methods The method uses a proprietary enrichment medium. Following aerobic enrichment, an immunochromatographic test strip is inserted into the tube containing the enrichment, developed for 20 min, and interpreted. Campylobacter-inoculated food samples were tested by the method, as well as the USDA/FSIS cultural reference method; Isolation and Identification of Campylobacter jejuni/coli/lari from Poultry Rinse, Sponge and Raw Product Samples MLG 41.04. The candidate method was also confirmed by an alternative cultural method. The RapidChek method was tested with 50 Campylobacter strains comprised of C. jejuni, C. coli, and C. lari, and 30 non-target strains. Results A total of 80 low-level spiked samples were tested by both methods in the study. The candidate method yielded 49 presumptive positives: all presumptive results were confirmed culturally. The reference method produced a total of 41 confirmed positive results. No difference between the alternate confirmation method and reference confirmation method was observed. Probability of detection analysis demonstrated no significant differences in the number of positive samples detected by the candidate method and cultural reference method. The RapidChek method detected all 50 Campylobacter strains and none of the 30 non-target strains, including Campylobacter spp. other than C. jejuni, C. coli, and C. lari. Conclusion The candidate method performed as well as the reference method in the detection of C. jejuni, C. coli, and C. lari in raw ground chicken, chicken carcass rinse, and turkey carcass sponges. Highlights Aerobic enrichment of selected matrixes for 48 h yielded reliable presumptive results for Campylobacter.

Background The Thermo Scientific SureTect™ Campylobacter jejuni, C. coli, and C. lari PCR Kit is a real-time PCR assay for the detection and differentiation of C. jejuni, C. coli, and C. lari from raw poultry, ready-to-cook poultry products, and environmental samples. Objective The Thermo Scientific SureTect Campylobacter jejuni, C. coli, and C. lari PCR Kit was evaluated for AOAC®Performance Tested MethodsSM certification. Methods Inclusivity/exclusivity, matrix studies, product consistency and stability, and robustness testing were conducted to assess the method’s performance. In the matrix studies, the method was validated against United States and international reference methods for Campylobacter detection. Results There were no statistically significant differences found in the matrix studies between the candidate and reference methods when analyzed by probability of detection. All 52 inclusivity strains and none of the 51 exclusivity strains tested were detected by the assay. Robustness testing demonstrated that the assay gave reliable performance with specific method deviations outside of the recommended parameters, and the real-time stability testing demonstrated that there were no statistically significant differences between kit lots, validating the stated shelf life of the kit. Conclusion The data presented support the product claims that the Thermo Scientific SureTect Campylobacter jejuni, C. coli, and C. lari PCR assay is suitable for the detection and differentiation of C. jejuni, C. coli, and C. lari from raw poultry, ready-to-cook poultry products, and environmental samples. Highlights Presumptive results can be obtained in as little as 23 h. Microaerophilic incubators are not required for enrichment.

Members of the Campylobacter lari group are causative agents of human gastroenteritis and are frequently found in shellfish, marine waters, shorebirds, and marine mammals. Within a One Health context, we used comparative genomics to characterize isolates from a diverse range of sources and geographical locations within Europe and Australia and assess possible transmission of food, animal, and environmental isolates to the human host. A total of 158 C. lari isolates from Australia, Denmark, France, and Germany, which included 82 isolates from human stool and blood, 12 from food, 14 from domestic animal, 19 from waterbirds, and 31 from the environment were analyzed. Genome-wide analysis of the genetic diversity, virulence, and antimicrobial resistance (AMR) traits was carried-out. Most of the isolates belonged to C. lari subsp. lari (Cll; 98, 62.0%), while C. lari subsp. concheus and C. lari urease-positive thermotolerant Campylobacter (UPTC) were represented by 12 (7.6%) and 15 (9.5%) isolates, respectively. Furthermore, 33 (20.9%) isolates were not assigned a subspecies and were thus attributed to distant Campylobacter spp. clades. Whole-genome sequence-derived multilocus sequence typing (MLST) and core-genome MLST (cgMLST) analyses revealed a high genetic diversity with 97 sequence types (STs), including 60 novel STs and 14 cgMLST clusters (≤10 allele differences), respectively. The most prevalent STs were ST-21, ST-70, ST-24, and ST-58 (accounting for 13.3%, 4.4%, 3.8%, and 3.2% of isolates, respectively). A high prevalence of the 125 examined virulence-related loci (from 76.8 to 98.4% per isolate) was observed, especially in Cll isolates, suggesting a probable human pathogenicity of these strains. IMPORTANCE Currently, relatedness between bacterial isolates impacting human health is easily monitored by molecular typing methods. These approaches rely on discrete loci or whole-genome sequence (WGS) analyses. Campylobacter lari is an emergent human pathogen isolated from diverse ecological niches, including fecal material from humans and animals, aquatic environments, and seafood. The presence of C. lari in such diverse sources underlines the importance of adopting an integrated One Health approach in studying C. lari population structure for conducting epidemiological risk assessment. This retrospective study presents a comparative genomics analysis of C. lari isolates retrieved from two different continents (Europe and Australia) and from different sources (human, domestic animals, waterbirds, food, and environment). It was designed to improve knowledge regarding C. lari ecology and pathogenicity, important for developing effective surveillance and disease prevention strategies.

We report a case of Campylobacter lari vertebral osteomyelitis with iliopsoas abscess. This is the first case report of vertebral osteomyelitis due to C. lari, which was identified from a vertebral biopsy sample collected by CT-guided percutaneous needle biopsy in a patient without obvious episodes of immunodeficiency. HK semisolid media played an important role in identifying the pathogen. It is important to make every possible effort to identify the causative pathogen in vertebral osteomyelitis.

We investigate the role of black-headed gulls (Chroicocephalus ridibundus), an omnivorous species that is among the most likely wild bird candidates for transmission of zoonotic agents, as a potential reservoir ofCampylobacterspp. Colonies with different anthropogenic pressures were studied to examine differences in exposure to sources ofCampylobacterbetween rural and urban birds. We recordedCampylobacterspp. in 4.87% of adult black-headed gulls and 2.22% of their chicks after analysing 1036 cloacal swabs collected over two breeding seasons in three colonies in northern Poland.Campylobacter jejuniwas found most frequently (85.72%), andCampylobacter lariandCampylobacter coliwere much scarcer. Prevalence ofCampylobacterdid not differ significantly between black-headed gulls breeding in urban (4.27%) and rural (3.80%) habitats. Almost all isolates from chicks and adults were susceptible to azithromycin (97.62%) and erythromycin (95.24%), but fewer to tetracycline (50.00%) and ciprofloxacin (47.62%).Campylobacterprevalence was unrelated to the date of sampling. Our study indicates that black-headed gulls are carriers of resistant to antibioticsCampylobacterand they can contaminate natural waterbodies with their faeces, which poses a threat to human and farm animal health.

The aim of this work was the establishment of a national laboratory sentinel surveillance service for human clinical Campylobacter in Ireland. This included detailed genomic molecular epidemiology of Campylobacter for 2019. For February-December 2019, 24 clinical microbiology laboratories in Ireland submitted all PCR/culture-positive clinical Campylobacter spp. specimens to Public Health Laboratory (PHL) Dublin one week out of every four. Antimicrobial susceptibility testing (AST) according to European Committee on Antimicrobial Susceptibility Testing (EUCAST) criteria was carried out for Campylobacter spp. isolates for ciprofloxacin, tetracycline and erythromycin. Batch whole genome sequencing (WGS) was carried out on cultures and analysis was performed to determine species, genotype, identify antimicrobial resistance (AMR) and virulence determinants and identify clusters. A total of 75 isolates and 366 PCR-positive stools were received, and 277 isolates recovered (55.7% recovery from stools). Of 257 isolates characterized by WGS, 86.4% (n = 222) were Campylobacter jejuni, 11.7% (n = 30) Campylobacter coli and 1.9% (n = 5) Campylobacter lari. There were 20 clonal complexes with ST-21 clonal complex most prevalent at 26.8% (n = 69). 50.5% (n = 140) of isolates were susceptible to all three antimicrobials tested. 39.3% (n = 109) isolates were ciprofloxacin resistant, 26.3% (n = 73) tetracycline resistant and two isolates erythromycin resistant. Congruence between phenotypic and genotypic AST was observed. There was 95.9% and 95.6% sensitivity and specificity for WGS to predict ciprofloxacin sensitivity and 98.6% and 99.5% sensitivity and specificity for WGS to predict tetracycline sensitivity. Virulence factors flaA, racR, ciaB and cdtB were detected in all isolates. WGS identified 31 potential clusters for public health alert. This sentinel surveillance of human campylobacteriosis in Ireland establishes the basis for a national reference service. Linking with other partners in a 'One Health' framework will help us better understand sources of infection to reduce disease burden and the threat of AMR.

Campylobacter is a major agent of gastroenteritis worldwide. The incidence and prevalence of campylobacteriosis have been increasing in both developed and developing countries over the last decade. In this study, 197 strains of successive Campylobacter-like were identified in French laboratories in September 2018. Bacterial isolates from clinical samples were identified with a mass spectrometer (Ultraflex III TOF/TOF and the BIOTYPER database from Bruker Daltonics). Of the 197 isolates tested, 143 were identified as Campylobacter jejuni (72.59%), 28 as Campylobacter coli (14.21%), 2 as Arcobacter butzleri (1.02%), 1 as Campylobacter fetus and 1 as Campylobacter lari with (0.51% each) by the MALDI-TOF mass spectrometry. Isolation rate of Campylobacter was highest in the 0 - 9 age group (22%). The proportion of male and female patients was 59.4% (CI 95% = 52.2-66.3) and 40.6% (CI 95% = 33.7-42.8) respectively. Sixty strains (30.5%) were resistant to tetracycline and 52 (26.4%) resistant to ampicillin. This study showed that the MALDI-TOF mass spectrometry is a rapid and accurate identification method of Campylobater spp in patients treated in private French laboratories. Key words: Campylobacter, identification, MALDI-TOF, patients, France.

The aim of this study was to assess the presence of thermotolerant Campylobacter species in shellfish production and harvesting areas of the Istrian aquatory (North Adriatic Sea), regarding the growing conditions (in terms of location and sea temperature) and shellfish species, as factors influencing increased public health risk when consuming shellfish. The study included samples of the most economically important species of live bivalve molluscs: mussels (Mytilus galloprovincialis), oysters (Ostrea edulis) and queen scallops (Aequipecten opercularis) that were collected in a period from August to October 2021, on a weekly basis at nine locations of the Istrian County, Croatia. Isolation of Campylobacter was done according to standard ISO 10272-1:2017 method, and species were identified using multiplex PCR. Isolates identified as C. jejuni and C. lari were genotyped using multilocus sequence typing (MLST). Among 108 examined samples of bivalve molluscs, mussels dominated and were the only ones found positive for the presence of Campylobacter (25.6%). The majority of Campylobacter positive samples, 85% of them (n = 17), were detected when the sea temperature was between 20 °C and 27 °C. In total, 19 C. lari and one C. jejuni strains were isolated. C. lari isolates found in this study belong to 13 sequence types (STs), and 9 of them are newly described in this research. Two out of the four previously described C. lari STs that were found in this study were previously detected in human stool. The only C. jejuni isolate was found to be sequence type 1268, which belongs to ST-1275 clonal complex that is almost exclusively found in seabirds but can sporadically cause infection in humans. Based on the obtained results, introducing surveillance of thermotolerant Campylobacter in shellfish in the Republic of Croatia is advised as an improvement for public health safety.

Background: The Bruker MALDI Biotyper® method utilizes matrix-assisted laser desorption/ionization time-of-flight MS for the rapid and accurate identification and confirmation of Gram-negative bacteria from select media types. The alternative method was evaluated in a method extension study of AOAC INTERNATIONAL First Action Official MethodSM 2017.09 using nonselective and selective agars to identify Cronobacter spp., Salmonella spp., Campylobacter spp., and select Gram-negative bacteria. Results obtained by the Bruker MALDI Biotyper were compared to the traditional biochemical methods as prescribed in the appropriate reference methods. Methods: Two collaborative studies were organized, one in the United States focusing on Cronobacter spp. and other Gram-negative bacteria and one in Europe focusing on Salmonella spp. and other Gram-negative bacteria. Fourteen collaborators from seven laboratories located within the United States participated in the first collaborative study for Cronobacter spp. Fifteen collaborators from 15 service laboratories located within Europe participated in the second collaborative study for Salmonella spp. For each target organism (either Salmonella spp. or Cronobacter spp.), a total of 24 blind-coded isolates were evaluated. In each set of 24 organisms, there were 16 inclusivity organisms (Cronobacter spp. or Salmonella spp.) and 8 exclusivity organisms (non-Cronobacter spp. and non-Salmonella spp. closely related Gram-negative organisms). For the Campylobacter spp. method extension, 17 collaborators from eight laboratories located within the United States (seven laboratories) and Canada (one laboratory) participated in the collaborative study. A total of 24 blind-coded isolates were evaluated. In each set of 24 organisms, there were 16 inclusivity organisms (Campylobacter spp.) and 8 exclusivity organisms (non-Campylobacter spp. closely related Gram-negative organisms). Results: After testing was completed, the total percentage of correct identifications from each agar type for each strain was determined at a percentage of 100.0% to the genus level for the Cronobacter study and a percentage of 100.0% to the genus level for the Salmonella study. For the Campylobacter method extension, a correct identification and confirmation rate of 100.0% was obtained for the Campylobacter organisms at the species level. For non-Cronobacter, non-Salmonella, and non-Campylobacter organisms, 100.0% were correctly identified. Conclusions: The results indicated that the alternative method produced equivalent results when compared to the confirmatory procedures specified by each reference method. Highlights: The method extension can be modified to include the identification and confirmation of Campylobacter jejuni, Campylobacter coli, and Campylobacter lari.

Highlights • This is the first case of cellulitis with bacteremia due to Campylobacter lari. • Subculture may be necessary to confirm negative conversion of the persistent C. lari bacteremia. • C. lari bacteremia was treated with oral minocycline. • The treatment regimen of C. lari bacteremia remain unclear., Campylobacter lari is an organism occasionally isolated in humans but rarely causes bacteremia. We report the first case of cellulitis with bacteremia due to C. lari in a patient undergoing mantle-cell lymphoma. A 51-year-old man presented with a two-month history of fever and bilateral leg pain and redness. Despite oral ciprofloxacin administration, his symptoms had not improved. The blood culture sample in the anaerobic bottle yielded positive results and C. lari was identified by mass spectrometry. The bacteremia did not initially respond to oral azithromycin but responded to intravenous meropenem and amikacin for five days followed by oral minocycline. This report indicates that C. lari bacteremia may be treated with oral minocycline following short-term intravenous antimicrobial therapy even among patients undergoing hematological malignancies.

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Zebra finches (Taeniopygia guttata) are laboratory animal species commonly used for modeling neurobiology and learning. Historically, using bacterial culture, biochemical analysis, and 16S ribosomal RNA gene sequencing, bacterial isolates from feces of finches housed at Massachusetts Institute of Technology had been presumptively diagnosed as Campylobacter jejuni, which is commonly isolated from both domestic and wild birds. Although the zebra finches were not clinically affected, C. jejuni is a known zoonotic pathogen that causes gastroenteritis in humans worldwide. Human transmission is predominantly foodborne and associated with the consumption of contaminated poultry; however, humans can also become infected from contact with C. jejuni-infected reservoir hosts. Because C. jejuni-infected finches pose a risk to research personnel, a study was undertaken to investigate the prevalence and taxonomic identification of Campylobacter spp. present in the finch colony. Campylobacter spp. were isolated from a total of 26 finch fecal samples collected in 2003, 2010, and 2017. 16S ribosomal RNA sequencing of all isolates determined that they shared 99% identity with either C. jejuni or Campylobacter lari. Sixteen of the isolates were subjected to further biochemical characterization and atpA and rpoB gene sequence analysis. Based on these analyses, three clusters of Campylobacter species were identified. The draft whole-genome sequences were determined for one representative isolate from each cluster. A pan-genomic phylogenetic tree, average nucleotide identity, digital DNA-DNA hybridization, and orthologous gene analyses indicated that each isolate was its own novel species, distinct from C. jejuni and other avian Campylobacter species. We have named these novel species Campylobacter taeniopygiae, Campylobacter aviculae, and Campylobacter estrildidarum, and in each novel species, we identified virulence genes suggesting their pathogenic and zoonotic potential.

Campylobacter spp. have been recognized as major foodborne pathogens worldwide. An increasing frequency of antibiotic-resistant pathogens, including Campylobacter spp., have been identified to transmit from food products to humans and cause severe threats to public health. To better mitigate the antibiotic resistance crisis, rapid detection methods are required to provide timely antimicrobial resistance surveillance data for agri-food systems. Herein, we developed a polymer-based microfluidic device for the identification and antimicrobial susceptibility testing (AST) of Campylobacter spp. An array of bacterial incubation chambers were created in the microfluidic device, where chromogenic medium and antibiotics were loaded. The growth of Campylobacter spp. was visualized by color change due to chromogenic reactions. This platform achieved 100% specificity for Campylobacter identification. Sensitive detection of multiple Campylobacter species (C. jejuni, C. coli, and C. lari) was obtained in artificially contaminated milk and poultry meat, with detection limits down to 1 × 102 CFU/ml and 1 × 104 CFU/25 g, respectively. On-chip AST determined Campylobacter antibiotic susceptibilities by the lowest concentration of antibiotics that can inhibit bacterial growth (i.e., no color change observed). High coincidences (91% to 100%) of on-chip AST and the conventional agar dilution method were achieved against several clinically important antibiotics. For a presumptive colony, on-chip identification and AST were completed in parallel within 24 h, whereas standard methods, including biochemical assays and traditional culture-based AST, take several days for multiple sequential steps. In conclusion, this lab-on-a-chip device can achieve rapid and reliable detection of antibiotic-resistant Campylobacter spp. IMPORTANCE Increasing concerns of antibiotic-resistant Campylobacter spp. with regard to public health emphasize the importance of efficient and fast detection. This study described the timely identification and antimicrobial susceptibility testing of Campylobacter spp. by using a microfluidic device. Our developed method not only reduced the total analysis time, but it also simplified food sample preparation and chip operation for end users. Due to the miniaturized size of the lab-on-a-chip platform, the detection was achieved by using up to 1,000 times less of the reagents than with standard reference methods, making it a competitive approach for rapid screening and surveillance study in food industries. In addition, multiple clinically important Campylobacter species (C. jejuni, C. coli, and C. lari) could be tested by our device. This device has potential for wide application in food safety management and clinical diagnostics, especially in resource-limited regions.

Campylobacter spp. are one of the most important food-borne pathogens, which are quite susceptible to environmental or technological stressors compared to other zoonotic bacteria. This might be due to the lack of many stress response mechanisms described in other bacteria. Nevertheless, Campylobacter is able to survive in the environment and food products. Although some aspects of the heat stress response in Campylobacter jejuni are already known, information about the stress response in other Campylobacter species are still scarce. In this study, the stress response of Campylobacter coli and Campylobacter lari to elevated temperatures (46°C) was investigated by survival assays and whole transcriptome analysis. None of the strains survived at 46°C for more than 8 h and approximately 20% of the genes of C. coli RM2228 and C. lari RM2100 were differentially expressed. The transcriptomic profiles showed enhanced gene expression of several chaperones like dnaK, groES, groEL, and clpB in both strains, indicating a general involvement in the heat stress response within the Campylobacter species. However, the pronounced differences in the expression pattern between C. coli and C. lari suggest that stress response mechanisms described for one Campylobacter species might be not necessarily transferable to other Campylobacter species.

Campylobacter lariis a rare cause of extraintestinal infection in humans. There has only been one prior case report of prosthetic joint infection in an immunocompetent adult, which was rapidly fatal. We can identify no previous case reports of successful management of this condition. We discuss our management of a 63 year old stable manager that presented with 1 week history of right prosthetic knee pain and swelling. Following two washouts the patient underwent a first-stage revision of his knee. Post-operative antibiotic choice was empirical, and 28-day outcome has been promising, with complete resolution of inflammatory markers and resolution of symptoms. Outcome following planned second-stage revision will be discussed at FIS 2019.

SUMMARYThe Foodborne Diseases Active Surveillance Network (FoodNet) conducts population-based surveillance forCampylobacterinfection. For 2010 through 2015, we compared patients withCampylobacter jejuniwith patients with infections caused by otherCampylobacterspecies.Campylobacter colipatients were more often >40 years of age (OR = 1·4), Asian (OR = 2·3), or Black (OR = 1·7), and more likely to live in an urban area (OR = 1·2), report international travel (OR = 1·5), and have infection in autumn or winter (OR = 1·2).Campylobacter upsaliensispatients were more likely female (OR = 1·6), Hispanic (OR = 1·6), have a blood isolate (OR = 2·8), and have an infection in autumn or winter (OR = 1·7).Campylobacter laripatients were more likely to be >40 years of age (OR = 2·9) and have an infection in autumn or winter (OR = 1·7).Campylobacter fetuspatients were more likely male (OR = 3·1), hospitalized (OR = 3·5), and have a blood isolate (OR = 44·1). International travel was associated with antimicrobial-resistantC. jejuni(OR = 12·5) andC. coli(OR = 12) infections. Species-level data are useful in understanding epidemiology, sources, and resistance of infections.

Consumption of Campylobacter contaminated food or water is a leading cause of human acute gastroenteritis. Campylobacter jejuni, Campylobacter coli, and Campylobacter lari account for over 95% of total Campylobacter infections. A multiplex quantitative polymerase chain reaction (qPCR) for simultaneous identification of C. jejuni, C. coli, and C. lari was developed for use with the SmartCycler II system.We evaluated and combined previously described primers and probes for Campylobacter detection, designed a new internal amplification control, and optimized the multiplex qPCR for the detection of C. jejuni, C. coli, and C. lari.This method was 100% specific when tested against a panel of 32 target Campylobacter strains and 31 non-Campylobacter reference strains. Furthermore, there was no cross-reactivity with seven strains from four nontarget Campylobacter species. The amplification efficiency of each target in this multiplex qPCR was over 90%, and each coefficient of linearity was greater than 0.99. With artificially mixed genomic DNA, this method detected as few as two, three, and two genome copies of C. jejuni, C. coli, and C. lari, respectively. This method was also able to detect these three Campylobacter species in artificially contaminated milk with a sensitivity of five spiked cells of each target per reaction.The three Campylobacter targets were simultaneously identified using artificially mixed genomic DNA and spiked raw milk. This SmartCycler-based multiplex qPCR is a rapid, specific, and sensitive method to identify C. jejuni, C. coli, and C. lari.

Poultry meat and products are major transmission routes of human campylobacteriosis. The aim of this study was to determine the numbers and antibiogram profile of Campylobacter isolates from slaughtered broiler and layer birds. One hundred and sixty caecal and one hundred and thirty two carcasses were randomly sampled at the Kejetia poultry slaughter, isolated on charcoal-cefoperazone-deoxycholate agar (CCDA) and confirmed by API CAMPY and their resistance profiles assessed by Kirby-Bauer disk diffusion. Prevalence was 22.5 and 21.9% in the faecal and carcasses, respectively with no significant differences. Species identified among faecal isolates were Campylobacter jejuni (42%), Campylobacter coli (28%), Campylobacter lari (22%) and Campylobacter hyo-intestinalis (8%) while 79% C. jejuni, 14% C. coli, 4% C. jejuni sub sp. doylei, and 3% C. lari were obtained from the carcasses. Resistance to the β-lactams ranged from 75 to 100%, 41 to 86% to the quinolones, 14 to 36% to the aminoglycosides, 100% to erythromycin, 97 to 100% to tetracycline, 72 to 83% to chloramphenicol and 90 to 94% to trimethoprim sulfamethoxazole. All species were sensitive to imipenem, but 100% of isolates were multidrug resistant. Contamination of carcasses with multidrug resistant strains of Campylobacter is a threat to handlers and consumers and of major public health issue. Key words: Multidrug resistance, faeces, carcass, poultry, Kumasi, Ghana.

The aim of this study was to identify and characterize thermophilic Campylobacter species in faecal samples from goats in Khartoum State, Sudan, by application of multiplex polymerase chain reaction. Campylobacteriosis is a zoonotic disease of global concern, and the organisms can be transmitted to human via food, water and through contact with farm animals and pets. There are five clinically related Campylobacter species: Campylobacter jejuni (C. jejuni). Campylobacter coli, Campylobacter lari, Campylobacter upsaliensis and Campylobacter fetus. Conventional cultural methods to diagnose campylobacteriosis are tedious and time consuming. Wide ranges of genes have been reported to be used for PCR-based identification of Campylobacter spp. We used a multiplex PCR assay to simultaneously detect genes from the major five clinically significant Campylobacter spp. The genes selected were hipO (hippuricase) and 23S rRNA from glyA (serine hydroxymethyl transferase) from each of C. jejuni. C. coli, C. lari, and C. upsaliensis; and sapB2 (surface layer protein) from C. fetus subsp. fetus. The assay was used to identify Campylobacter isolates recovered from 336 cultured faecal samples from goats in three localities in Khartoum State. C. coli was the most predominant isolate (234; 69.6%), followed by C. jejuni (19; 5.7%), C. upsaliensis (13; 3.9%), C. fetus subsp. fetus (7; 2.1%) and C. lari (6; 1.8%). Twenty-nine goats showed mixed infection with Campylobacter spp., 21 of which harbored two Campylobacter spp., while eight animals were infected with three species. Ten out of twelve goats that displayed diarrhea harbored C. coli only. C. coli, C. jejuni and C. upsaliensis showed significant variation with localities. The prevalence of C. coli was significantly higher (87; 25.9%) in goats from Omdurman, whereas C. jejuni and C. upsaliensis were significantly higher (11; 3.3%, 9; 2.7%) in goats from Khartoum. The multiplex PCR assay was found to be rapid and easy to perform and had a high sensitivity and specificity for characterizing the isolates, even in mixed cultures. The study demonstrated the significance of goats as reservoirs in the dissemination of Campylobacter spp. which could be considered as potential agent of caprine enteritis and abortion as well as contamination of the wider environment posing serious public health concern in Khartoum State.

Campylobacter spp. are recovered from a wide variety of sources, including birds, livestock, shellfish, and human clinical samples. We present here the complete genomic data for the type strains of Campylobacter fetus subsp. venerealis, Campylobacter lari subsp. concheus, Campylobacter sputorum bv. sputorum, and Campylobacter volucris.

To better understand public health implications of waterfowl as reservoirs for zoonotic sources of Campylobacter in recreational waters, we developed a Gallus gallus (chick) model of infection to assess the pathogenicity of environmental isolates of Campylobacter. This method involved exposure of 1-day-old chicks through ingestion of water, the natural route of infection. Viable Campylobacter from laboratory-infected animals were monitored by using a modified non-invasive sampling of fresh chick excreta followed by a passive polycarbonate-filter migration culture assay. The method was used to evaluate the infectivities of three laboratory strains of Campylobacter spp. (Campylobacter coli, Campylobacter jejuni, and Campylobacter lari), three clinical isolates of C. jejuni, and four environmental Campylobacter spp. isolated from California gulls (Larus californicus). The results revealed that chicks were successfully infected with all laboratory and clinical isolates of Campylobacter spp. through ingestion of Campylobacter-spiked water, with infection rates ranging from 90% in a dose-dependent manner. More importantly, exposure of chicks with Campylobacter spp. isolated from Gallus gallus excreta also resulted in successful establishment of infection (≤90%). Each monitored Campylobacter spp. contained ≥7.5 × 104 CFU⋅g–1 of feces 7 days post-exposure. These results suggest that a G. gallus model can be used to assess infectivity of Campylobacter isolates, including gull and human clinical isolates. Use of an avian animal model can be applied to assess the importance of birds, such as the G. gallus, as potential contributors of waterborne-associated outbreaks of campylobacteriosis.

A DNA hybridization test was investigated for application to the detection of Campylobacter spp. in poultry samples. The test chemistry involves solution phase hybridization and detection by means of an enzymatically generated colorimetric endpoint. DNA probes used in the test system are targeted to unique sequences of ribosomal RNA and are specific for Campylobacter jejuni , Campylobacter coli , Campylobacter lari , and Campylobacter fetus subsp. fetus . Initial experiments with pure cultures of C. jejuni established the sensitivity limit of the DNA hybridization assay at approximately 106-7 CFU per sample. Experiments were designed to define optimal conditions for recovery and selective enrichment of Campylobacter spp. from chicken carcasses for use in conjunction with the DNA hybridization assay. Following overnight enrichment, cultures were swabbed onto Campy-Cefex agar plates and allowed to incubate for 24 h. This overnight growth was then suspended and assayed with the DNA probe. The remainder of the overnight enrichment was centrifuged and the resulting pellet was analyzed. Thirty-eight chicken carcasses were assayed for Campylobacter spp. by DNA probe and culture methodology employing culture enrichment and selective plating. Culture procedures isolated Campylobacter spp. from 23 carcasses, while the DNA probe assay detected the organism from 21 carcasses. The DNA probe registered five "false" positives and seven "false" negatives relative to the cultural bacteriologic approach.

Aim : The study was conducted to determine the prevalence and viability of Campylobacter species isolates from different species of poultry and humans in Sokoto State, Nigeria. Materials and Methods : A cross-sectional study was performed in the live birds markets, humans on admission and at outpatient clinics in the randomly selected hospitals in Sokoto State. Isolation and characterization of Campylobacter species were performed using standard culture isolation techniques and biochemical characterization. A total of 798 (506 cloacal and 292 fecal) swabs from poultry and humans, respectively, were collected and analyzed. The viability of 307 isolates stored in 15% glycerol and 85% tryptone broth at −20°C was determined after 7-13 months. Results : A total of 312 (39%) were positive for Campylobacter species which comprises 119 (30%), 20 (30%), 3 (14%), 9 (56%), 1 (50%), and 160 (55%) in chicken, guinea fowls, pigeons, ducks, turkey, and humans, respectively. The total of 38 (24%), 63 (39%), and 59 (37%) humans and 29 (19%), 79 (52%), and 44 (29%) poultry isolates were positive for Campylobacter jejuni , Campylobacter Coli , and Campylobacter Lari, respectively. A total of 261 (85%) of the stored isolates were still viable on re-isolation with the viability rates of 41 (95%), 67 (85%), and 17 (59%) at 7, 9, and 13 months of storage, respectively. There was a negative correlation between months of storage and viability rates. However, there was no significant statistical association (p>0.05) between prevalence rate and species of poultry. Conclusion : Campylobacter species have been detected with varying degree of prevalence in both poultry and humans and their ability to survive freezing at −20°C (95%) for up to 7 months has been revealed in the study. This is not only a concern to food and livestock industries but also a concern to the public health at large, especially, in view of the study area being considered one of the largest livestock producers in Nigeria. Campylobacteriosis is known to be associated with the cost of gastroenteritis management, antimicrobial resistance, food contamination, and complications such as a paralytic condition called Guillain-Barre syndrome.

Campylobacter lari strain SCHS02, a novel hyper-aerotolerant strain that survives under aerobic conditions, was isolated from retail duck meat. The genome is a single chromosome of 1,520,838 base pairs, with a mean GC content of 29.7%. It harbors 1546 protein-coding sequences and 45 tRNA and 9 rRNA genes. Genes associated with the oxidative stress response, including perR, bcp, ahpC, and sodB, were identified in the genome. Furthermore, 68 virulence-related genes were identified and sorted into 9 classes and 14 subclasses. The virulence gene profile of SCHS02 was similar to those of two human clinical C. lari isolates. Comparative genomic analysis of strain SCHS02 and 18 C. lari strains retrieved from a public database revealed the core and accessory gene profiles of C. lari strains, as well as putative core gene involved in halotolerance. Phylogenetic analysis revealed that strain SCHS02 is genetically related to isolates from bird samples and human clinical isolates, rather than to isolates from other environmental sources. These findings reveal essential genomic information about the newly identified hyper-aerotolerant C. lari strain isolated from a duck source, providing a basis for future studies of the strain considering its potential threat to public health and further research of the pathogenicity of C. lari.

Background: The objective of this study was to determine the prevalence of thermophilic Campylobacter spp. in asymptomatic school-going children and establish the antibiotic resistance patterns of the isolates towards the drugs used to treat campylobacteriosis, including macrolides, quinolones and tetracycline. Campylobacter spp. are a leading cause of enteric illness and have only recently shown resistance to antibiotics. Methods: This study isolated Campylobacter spp., including Campylobacter coli, Campylobacter jejuni and Campylobacter lari, in stool samples from asymptomatic school-going children in one of the biggest urban slums in Kenya. The disc diffusion method using EUCAST breakpoints was used to identify antibiotic-resistant isolates, which were further tested for genes encoding for tetracycline resistance using primer-specific polymerase chain reaction. Results: In total, 580 stool samples were collected from 11 primary schools considering both gender and age. Subjecting 294 biochemically characterized Campylobacter spp. isolates to genus-specific PCR, 106 (18.27% of stool samples) isolates were confirmed Campylobacter spp. Out of the 106 isolates, 28 (4.83%) were Campylobacter coli, 44 (7.58%) were Campylobacter jejuni while 11 (1.89%) were Campylobacter lari. Campylobacter jejuni had the highest number of isolates that were multi-drug resistant, with 26 out of the 28 tested isolates being resistant to ciprofloxacin (5 mg), nalidixic acid (30 mg), tetracycline (30 mg) and erythromycin (15 mg). Conclusions: In conclusion, asymptomatic school going children in the study area were found to be carriers of multidrug resistant Campylobacter coli, Campylobacter jejuni and Campylobacter lari at 84%. A one-health approach, which considers overlaps in environment, animals and human ecosystems, is recommended in addressing multidrug resistane in Campylobacter, since animals are the main reservoirs and environmental contamination is evident.

Oligosaccharyltransferase (OST) is a key enzyme of the N-glycosylation pathway, where it catalyzes the transfer of a glycan from a lipid-linked oligosaccharide (LLO) to an acceptor asparagine within the conserved sequon N-X-T/S. A previous structure of a ternary complex of bacterial single subunit OST, PglB, bound to a non-hydrolyzable LLO analog and a wild type acceptor peptide showed how both substrates bind and how an external loop (EL5) of the enzyme provided specific substrate-binding contacts. However, there was a relatively large separation of the substrates at the active site. Here we present the X-ray structure of PglB bound to a reactive LLO analog and an inhibitory peptide, revealing previously unobserved interactions in the active site. We found that the atoms forming the N-glycosidic bond (C-1 of the GlcNAc moiety of LLO and the –NH2 group of the peptide) are closer than in the previous structure, suggesting that we have captured a conformation closer to the transition state of the reaction. We find that the distance between the divalent metal ion and the glycosidic oxygen of LLO is now 4 Å, suggesting that the metal stabilizes the leaving group of the nucleophilic substitution reaction. Further, the carboxylate group of a conserved aspartate of PglB mediates an interaction network between the reducing-end sugar of the LLO, the asparagine side chain of the acceptor peptide, and a bound divalent metal ion. The interactions identified in this novel state are likely to be relevant in the catalytic mechanisms of all OSTs. ISSN:2045-2322

1. Rebecca G. Same, MD* 2. Pranita D. Tamma, MD, MHS* 1. *Division of Pediatric Infectious Diseases, Department of Pediatrics, The Johns Hopkins University School of Medicine, Baltimore, MD * Abbreviations: CDC: : Centers for Disease Control and Prevention CIDT: : culture-independent diagnostic testing GBS: : Guillain-Barre syndrome HIV: : human immunodeficiency virus IBD: : inflammatory bowel disease Ig: : immunoglobulin Campylobacter is one of the 2 most common causes of foodborne illness in the United States. It most commonly occurs in children younger than 5 years of age. Campylobacter species can cause a wide range of syndromes, from asymptomatic infections to severe systemic infections. After completing this article, readers should be able to: 1. Recognize that Campylobacter is a common cause of foodborne illness in the United States and internationally. 2. Understand the indications for testing and the treatment of Campylobacter infection. Campylobacter species are an important cause of infection throughout the world, especially in young children. Campylobacter jejuni is the most common Campylobacter species in the United States, where it is 1 of the top 2 causes of foodborne illness. (1) Other important species include Campylobacter upsaliensis , Campylobacter lari , and Campylobacter fetus , which can cause serious systemic infections in all age groups, particularly in immunocompromised patients. Campylobacter species infections cause gastroenteritis and typically present with diarrhea that may or may not be bloody, emesis, and abdominal pain. Campylobacter infection in children can mimic intussusception, appendicitis, or inflammatory bowel disease (IBD). Immune-mediated complications of Campylobacter infection include reactive arthritis and Guillain-Barre syndrome (GBS). Campylobacter infection is a nationally notifiable disease, and confirmed cases should be reported through the Centers for Disease Control and Prevention (CDC) Foodborne Diseases Active Surveillance Network. Most Campylobacter infections are mild and self-limited and require only supportive care, although some may lead to severe dehydration. Serious infection or infections in immunocompromised hosts may benefit from treatment with macrolide antibiotics. Increasing …

The objective of this study was to examine the efficacy of three antibiotic mixtures for use as supplements in a selective medium that could be used to isolate Campylobacter from mixed bacterial cultures during aerobic incubation. A non-selective, basal broth medium was prepared and supplemented with Bolton, Cefex, or Skirrow antibiotic mixtures. The ability of pure cultures of Campylobacter coli, Campylobacter fetus, Campylobacter jejuni, and Campylobacter lari to grow in basal broth and basal broth supplemented with each one of the antibiotic mixtures after aerobic incubation for 24 and 48 h at 37°C was determined. Also, the ability of pure cultures of Escherichia coli, Enterococcus faecalis, Listeria monocytogenes, Pseudomonas aeruginosa, Salmonella Kentucky, and Staphylococcus aureus to grow in basal and supplemented broth medium was determined. Additionally, the recovery of bacteria from mixed cultures containing one Campylobacter isolate with the other 6 non-Campylobacter isolates after aerobic incubation for 48 h at 37°C in basal broth and broth supplemented with the Bolton antibiotic mixture was examined. Results indicated that there was significant (p ≤ 0.05) growth of most Campylobacter isolates in basal broth and in broth supplemented with Bolton, Cefex, or Skirrow antibiotic mixtures after 24 or 48 h of aerobic incubation. Although, there was significant growth non-Campylobacter isolates in basal broth, growth of most these bacteria was inhibited in media supplemented with antibiotic mixtures. Also, Campylobacter growth was generally significantly less than other bacteria in mixed bacterial cultures grown in basal broth for 48 h at 37°C, but significantly more of most Campylobacter isolates were recovered from mixed bacterial cultures grown in media supplemented with the Bolton antibiotic mixture. Findings indicate that basal medium containing this supplement should be studied as a selective medium that may be utilized with aerobic incubation to isolate Campylobacter from environmental samples containing other bacteria.

Campylobacter spp. is one of the main cause of bacterial gastroenteritis in the world. The increase of antibiotic resistance in this species is a threat to public health. A Campylobacter spp. surveillance study was performed in Italy in the 2013–2016 period by the Enter-Net Italia network. The most prevalent Campylobacter species identified causing gastroenteritis was Campylobacter jejuni (73.4%) and 45% of all the annual cases of campylobacteriosis were reported in the summer period. High rates of ciprofloxacin and tetracycline resistance in Campylobacter spp. have been observed. An increasing percentage of Campylobacter coli strains simultaneously resistant to ciprofloxacin, tetracycline and erythromycin has been found. Molecular mechanisms of resistance have been investigated and the role of efflux pumps evaluated. Antibiotic resistance in Campylobacter spp. is an increasing serious threat that requires coordinated action to minimize the emergence and spread of antimicrobial resistant strains from animals to humans throughout the food chain.

A multiplex PCR assay for simultaneous detection and differentiation of Campylobacter jejuni, Campylobacter coli, and Campylobacter lari was developed and validated to assess the occurrence of these bacteria in fresh chicken meat and by-products in Bangkok, Thailand, by using a new combination of four previously published PCR primers for C. jejuni, C. coli, C. lari, and a universal 16S rDNA gene as an internal control. The specificity was determined by using 13 strains of other bacteria. With pure culture DNA, the detection limit was 0.017 ng/PCR for C. jejuni and C. coli and was 0.016 ng/PCR for C. lari. It can detect 10 CFU of C. jejuni, C. coli, and C. lari in 2 g of chicken meat within a 16-h enrichment time. Our multiplex PCR assay was applied for identification of Campylobacter spp. in 122 supermarket samples and 108 fresh market samples. Of the 230 samples evaluated by multiplex PCR, 54.0, 3.3, and 10.7% of supermarket samples were positive for C. jejuni, C. coli, and mixed C. jejuni and C. coli, respectively, and 56.5 and 33.3% of fresh market samples were positive for C. jejuni and mixed C. jejuni and C. coli, respectively. No sample was positive for C. lari. Fresh market samples had significantly higher C. jejuni and C. coli contamination than those from supermarkets (relative risk: 1.3; P = 0.0001). Compared with the culture method (a gold standard), the sensitivity, specificity, positive predictive value, negative predictive value, and diagnostic accuracy of multiplex PCR were 97.7, 86.8, 96.1, 92.0, and 95.2%, respectively. No significant difference was observed between results from two methods (P = 0.55). Therefore, the established multiplex PCR was not only rapid and easy to perform but had a high sensitivity and specificity to distinguish between C. jejuni, C. coli, and C. lari, even in samples containing mixed contamination. Our study indicated that fresh chicken meat and by-products from fresh markets were significantly less hygienic than those from supermarkets.

Full-length tlyA gene and its adjacent genetic loci from the urease-positive thermophilic Campylobacter (UPTC) CF89-12 [approximately 15,000 base pairs (bp) in length], as well as a reference strain Campylobacter lari RM2100 (approximately 9,000 bp), were analyzed. The possible open-reading frame of tlyA from UPTC CF89-12 was shown to have 720 bp with a calculated molecular mass of approximately 26.7 kDa. Using a primer pair designed in silico, a total of approximately 1.1 kbp consisting of putative promoter region, structural gene for tlyA, and its adjacent genetic loci were identified in all 17 C. lari isolates [n = 13 for UPTC; n = 4 for urease-negative (UN) C. lari]. Although sequence differences were demonstrated at approximately 20 loci within the 90 bp non-coding (NC) region, including the putative promoter structure candidates immediately upstream of the tlyA gene among the 18 isolates including C. lari RM2100, no sequence differences were identified within the NC region among the five UN C. lari isolates examined. A start codon ATG and a probable ribosome-binding site, AGGC(T)GG(A), for the tlyA gene were identified in all 18 isolates, including C. lari RM2100. The putative intrinsic ρ-independent transcriptional terminator structure candidate was also identified for the tlyA gene in both UPTC CF89-12 and C. lari RM2100. Additionally, the hemolysis assay was performed with some of the C. lari isolates. The tlyA gene nucleotide sequence data may possibly be useful for discrimination between UN C. lari and UPTC organisms, as well as for the differentiation among the four thermophilic Campylobacter species.

Aim: The study was carried out to survey and determine the prevalence of Campylobacter species among pigs and humans within the pig rearing areas of Zuru Kebbi State, Nigeria. Materials and Methods: A cross-sectional study was conducted among household pigs and humans between September, 2013 and February, 2014. Isolation and characterization of Campylobacter species were performed using standard culture isolation techniques and biochemical characterization. A total of 450 fecal samples comprised of 300 from pigs and 150 from humans was collected and analyzed. Results: Prevalence of Campylobacter were 278 (92.67%) and 94 (62.67%) for pigs and humans respectively. The most encountered Campylobacter species in both cases was Campylobacter coli (276 [74.19]) followed by Campylobacter jejuni (62 [16.66]). The least isolated species in pigs was Campylobacter hyointestinalis 5 (1.8%) while Campylobacter lari 2 (2.13%) was least isolated in humans. C. lari was not found in pigs. No significant association (p>0.05) existed between Campylobacter isolates and the age and sex of both pigs and humans in this study. Conclusion: Both pigs and humans within the pig rearing areas of Zuru have been shown to harbor Campylobacter species and this might be due to extensive system of pig farming with indiscriminate defecation by pigs coupled with unhygienic disposal of human wastes in the environment and poor personal hygiene.

1. The aim of the present study was to evaluate the antimicrobial resistance of Campylobacter strains (C. jejuni, C. coli and C. lari) isolated from broiler carcasses processed in the State of Paraná, Brazil. 2. Rates of microbial resistance and susceptibility were assessed by both Disk Diffusion (DD) and Etest (Minimum Inhibitory Concentration) techniques. Antibiotics were tested using DD (12 antibiotics) and/or MIC (7 antibiotics) methods. 3. A total of 95.8% of the strains were resistant to at least two agents. In terms of multidrug resistance, 75% of strains were resistant to three or more groups of antibiotics. The highest rates of resistance were detected for cefalotin, ciprofloxacin, tetracycline and nalidixic acid. A high rate of susceptibility of the strains to erythromycin (95.8%) was found confirming that this is considered the agent of choice for treating campylobacteriosis. Comparison of the microbial resistance and susceptibility, as determined simultaneously by the two methods, found the techniques to be statistically equivalent for 5 out of the 6 antibiotics tested. 4. The results of this study suggest the need for adopting measures to control the use of antibiotics in broiler production to prevent multidrug resistance of Campylobacter strains and reduce the risk of serious human diseases caused by the consumption of contaminated chicken meat.