Comparison between the biofilm initiation of Campylobacter jejuni and Campylobacter coli strains to an inert surface using BioFilm Ring Test®

Aims: The adhesion to an inert surface (the first step of biofilm formation) of the two main pathogenic Campylobacter species, Campylobacter jejuni and Campylobacter coli, isolated from diverse origins, was compared. Methods and Results: Adhesion assays were conducted in 96‐well, polystyrene microtiter plates using the BioFilm Ring Test^® method. This new technique, based on magnetic bead entrapment, was shown to be suitable for analysing the adhesion of Campylobacter sp. strains by comparing the adhesion of four C. jejuni strains as revealed by the BioFilm Ring Test^® and immunodetection. Among the 46 strains tested, C. jejuni and C. coli displayed different adhesion capabilities ranging from no adhesion to strong adhesion. However, no strain of C. coli was strongly adherent, and statistically, C. coli adhered less to an inert surface than C. jejuni. In addition, strains isolated from animals or carcasses were less adherent than those isolated from food‐processing and clinical cases. Conclusions: These observations suggest that the food environment and the human body could have selected strains with greater adhesion. Significance and Impact of the Study: The adhesion capability of strains could partly explain the cross‐contamination or re‐contamination of food products by Campylobacter. This property could provide a mode of survival for Campylobacter in the food chain.     

Identification of a fibronectin-binding domain within the Campylobacter jejuni CadF protein

The binding of Campylobacter jejuni to fibronectin (Fn), a component of the extracellular matrix, is mediated by a 37 kDa outer membrane protein termed CadF for Campylobacter adhesion to Fn. Previous studies have indicated that C. jejuni binds to Fn on the basolateral surface of T84 human colonic cells. To further characterize the interaction of the CadF protein with Fn, enzyme-linked immunosorbent assays were performed to identify the Fn-binding domain (Fn-BD). Using overlapping 30-mer and 16-mer peptides derived from translated cadF nucleotide sequence, maximal Fn-binding activity was localized to four amino acids (AA 134-137) consisting of the residues phenylalanine-arginine-leucine-serine (FRLS). A mouse alpha-CadF peptide polyclonal antibody (M alpha-CadF peptide pAb) was generated using FRLS containing peptides and found to react with viable C. jejuni as judged by indirect fluorescent microscopy, suggesting that the FRLS residues are surface-exposed. Binding of CadF to purified Fn and INT 407 human epithelial cells was significantly inhibited with peptides containing the Fn-BD. Moreover, a CadF recombinant variant protein, in which the Phe-Arg-Leu residues (CadF AA 134-136) were altered to Ala-Ala-Gly, exhibited a 91% decrease in Fn-binding activity as compared with the wild-type CadF protein. Collectively, these data indicate that the FRLS residues (CadF AA 134-137) of the C. jejuni CadF protein possess Fn-binding activity.     

Detection and quantification of Campylobacter jejuni and Campylobacter coli using real-time multiplex PCR

Aims: We describe a real‐time quantitative multiplex polymerase chain reaction (qmPCR) assay to identify and discriminate between isolates of Campylobacter jejuni and Campylobacter coli. Methods and Results: Two novel sets of primers and hydrolysis probes were designed to amplify the unique DNA sequences within the hipO, ccoN and cadF genes that are specific to Camp. jejuni and Camp. coli. Using the designed optimized qmPCR assay conditions, the amplification efficiency is in range from 108 to 116%. These qmPCR assays are highly specific for Camp. jejuni and Camp. coli, as seen through testing of 40 Campylobacter strains and 17 non‐Campylobacter strains. In chicken juice and tap water models spiked with known quantities of Camp. jejuni, qmPCR detected 10²–10³ CFU ml^?1 within 4 h. Conclusions: The qmPCR assays developed in this study provide reliable and simultaneous detection and quantification of Camp. jejuni and Camp. coli, with good amplification reaction parameters. Significance and Impact of the Study: Following further validation, the qmPCR assay reported here has the potential to be applied to various sample types as an alternative and rapid methodology.     

Campylobacter jejuni FlpA Binds Fibronectin and Is Required for Maximal Host Cell Adherence

Campylobacter jejuni is one of the most frequent bacterial causes of food-borne gastrointestinal disease in developed countries. Previous work indicates that the binding of C. jejuni to human intestinal cells is crucial for host colonization and disease. Fibronectin (Fn), a major constituent of the extracellular matrix, is a ∼250-kDa glycoprotein present at regions of cell-to-cell contact in the intestinal epithelium. Fn is composed of three types of repeating units: type I (∼45 amino acids), type II (∼60 amino acids), and type III (∼90 amino acids). The deduced amino acid sequence of C. jejuni flpA (Cj1279c) contains at least three Fn type III domains. Based on the presence of the Fn type III domains, we hypothesized that FlpA contributes to the binding of C. jejuni to human INT 407 epithelial cells and Fn. We assessed the contribution of FlpA in C. jejuni binding to host cells by in vitro adherence assays with a C. jejuni wild-type strain and a C. jejuni flpA mutant and binding of purified FlpA protein to Fn by enzyme-linked immunosorbent assay (ELISA). Adherence assays revealed the binding of the C. jejuni flpA mutant to INT 407 epithelial cells was significantly reduced compared with that for a wild-type strain. In addition, rabbit polyclonal serum generated against FlpA blocked C. jejuni adherence to INT 407 cells in a concentration-dependent manner. Binding of FlpA to Fn was found to be dose dependent and saturable by ELISA, demonstrating the specificity of the interaction. Based on these data, we conclude that FlpA mediates C. jejuni attachment to host epithelial cells via Fn binding.Members of the genus Campylobacter are gram-negative, asaccharolytic, motile bacteria, which grow optimally in the laboratory at temperatures between 37 and 42°C under microaerophilic conditions. Although members of Campylobacter spp. were initially recognized to cause disease in sheep and cattle, Campylobacter jejuni was not recognized as a human pathogen until much later (25). Infection of humans with C. jejuni is characterized by a rapid onset of fever, abdominal cramps, and diarrhea. C. jejuni is now recognized as one of the leading bacterial causes of gastroenteritis in the world. In spite of the incidence of campylobacteriosis, relatively few C. jejuni virulence genes have been characterized, and our understanding of the virulence properties of C. jejuni is limited compared with that of other enteric pathogens, including Salmonella, Shigella, and Yersinia spp.The ability of C. jejuni to cause disease is a complex, multifactorial process. Virulence factors that contribute to the pathogenesis of C. jejuni are associated with motility, host (target) cell adherence, host cell invasion, protein secretion, alteration of host cell signaling pathways, induction of host cell death, evasion of host immune defenses, iron acquisition, and drug/detergent resistance (14, 18). The binding of C. jejuni to specific host cell ligands is hypothesized to play a fundamental role in host colonization and disease progression, since it prevents the organism''s clearance from the intestine by peristalsis and fluid flow. Fauchere et al. (5) reported that C. jejuni isolates recovered from individuals with fever and diarrhea adhered to cultured cells in greater numbers than isolates recovered from asymptomatic individuals. While there is no evidence indicating that C. jejuni produces fimbriae that assist in host colonization (7), a number of constitutively synthesized proteins have been proposed to act as adhesins. Bacterial adhesins are surface-exposed macromolecules that facilitate an organism''s binding to the host cell receptors. Known and putative C. jejuni adhesins include CadF, CapA, FlpA, and PorA (MOMP) (6).An emerging theme among pathogenic microorganisms is their ability to utilize host cell molecules during the infectious process to facilitate their binding and entry into host cells (27). More specifically, many bacterial pathogens have been found to bind to fibronectin (Fn), which in turn modifies host cell signaling pathways to the pathogen''s advantage. Fn exists as a dimer of nearly identical 250-kDa subunits that are linked by a pair of disulfide bonds near their C termini. Each Fn monomer is composed of three types of repeating units: type I (∼45 amino acids), type II (∼60 amino acids), and type III (∼90 amino acids) (22). In total, each monomer contains 12 type I repeats, two type II repeats, and 15 to 17 type III repeats. Fn participates in many cellular interactions, including tissue repair, embryogenesis, blood clotting, and cell migration/adhesion. Plasma Fn, which is synthesized by hepatocytes, is soluble (22). In contrast, Fn involved in host cell-extracellular matrix (ECM) interaction, which is synthesized by chondrocytes, fibroblasts, endothelial cells, macrophages, and certain epithelial cells, is present in an insoluble form (22). Fn serves as an adhesion molecule that anchors cells to ECM components, including collagen and other proteoglycan substrates.The bacterial proteins that bind to ECM components have been termed microbial surface components recognizing adhesive matrix molecules (MSCRAMMs) (23). The C. jejuni CadF protein is a member of the MSCRAMM family and one of the most extensively characterized C. jejuni virulence determinants (10-12, 15, 16, 19-21, 24, 28). CadF mediates the binding of C. jejuni to Fn, promotes bacterium-host cell interactions, and facilitates the organism''s colonization of chickens (10, 11, 15, 16, 20, 21, 28). In addition to CadF, we recently reported that a mutation in Cj1279c resulted in a C. jejuni mutant that poorly colonized broiler chickens compared with a C. jejuni wild-type strain. The product encoded by the Cj1279c gene was termed Fibronectin-like protein A (FlpA) because the protein harbors Fn type III domains (6). The goal of this study was to characterize the binding properties of FlpA and to determine if this protein is a member of the MSCRAMM family. Here we provide experimental evidence that C. jejuni FlpA is surface exposed, promotes the bacterium''s attachment to host epithelial cells, and has Fn binding activity. Assays were also performed to determine if CadF and FlpA act cooperatively to promote binding of C. jejuni to host cells and Fn. We submit that the identification of a second MSCRAMM in C. jejuni highlights the importance of Fn binding in host colonization and disease.     

The cooperative action of bacterial fibronectin-binding proteins and secreted proteins promote maximal Campylobacter jejuni invasion of host cells by stimulating membrane ruffling

This study was performed to elucidate the host cell scaffolding and signalling molecules that Campylobacter jejuni utilizes to invade epithelial cells. We hypothesized that the C. jejuni fibronectin‐binding proteins and secreted proteins are required for cell signalling and maximal invasion of host cells. C. jejuni binding to host cells via the CadF and FlpA fibronectin‐binding proteins activated the epidermal growth factor (EGF) pathway, as evidenced by inhibitor studies and immunoprecipitation coupled with immunoblot analysis using antibodies reactive against total and active EGF receptor. Inhibitor studies revealed maximal C. jejuni host cell invasion was dependent upon PI3‐Kinase, c‐Src and focal adhesion kinase (FAK), all of which are known to participate in cytoskeletal rearrangements. Knockdown of endogenous Dock180, which is a Rac1‐specific guanine nucleotide exchange factor, using siRNA revealed that C. jejuni invasion was significantly reduced compared with cells treated with scrambled siRNA. We further demonstrated that the C. jejuni Cia proteins are, in part, responsible for Rho GTPase Rac1 recruitment and activation, as judged by immunofluorescence microscopy and Rac1 activation. Based on these data, we present a model that illustrates that C. jejuni utilizes a coordinated mechanism involving both adhesins and secreted proteins to promote membrane ruffling and host cell invasion.     

Unique features of the motility and structures in the flagellate polar region of Campylobacter jejuni and other species: an electron microscopic study

Similarly to Helicobacter pylori but unlike Vibrio cholerae O1/O139, Campylobacter jejuni is non‐motile at 20°C but highly motile at ≥37°C. The bacterium C. jejuni has one of the highest swimming speeds reported (>100 μm/s), especially at 42°C. Straight and spiral bacterial shapes share the same motility. C. jejuni has a unique structure in the flagellate polar region, which is characterized by a cup‐like structure (beneath the inner membrane), a funnel shape (opening onto the polar surface) and less dense space (cytoplasm). Other Campylobacter species (coli, fetus, and lari) have similar motility and flagellate polar structures, albeit with slight differences. This is especially true for Campylobacter fetus, which has a flagellum only at one pole and a cup‐like structure composed of two membranes.     

Ribosomal operon intergenic sequence region (ISR) heterogeneity in Campylobacter coli and Campylobacter jejuni

Aims: The intergenic sequence regions (ISR) between the 16S and 23S genes of Campylobacter jejuni and Campylobacter coli are markedly different for each species. However, in the genomic sequence for Camp. coli RM2228 , two rRNA operons have an ISR that is characteristic of Camp. coli, and the third operon is characteristic of Camp. jejuni. The aim of this study was to determine the prevalence of ISR heterogeneity in these organisms. Methods and Results: PCR primers were designed to yield a 327‐base pair (bp) product for Camp. coli and 166‐bp product for Camp. jejuni. A strain like Camp. coli RM2228 should yield products of both sizes. DNA from a panel of Camp. coli (n = 133) and Camp. jejuni (n = 134) isolates were tested. All of the isolates yielded products of the predicted size for the species. To verify the data for Camp. coli RM2228 , each ribosomal operon from the isolate was individually amplified by PCR and tested with the ISR primer pair. Products of both sizes were produced as predicted. Conclusions: The cross‐species heterogeneity of the ISR seen in Camp. coli RM2228 is uncommon. Significance and Impact of the Study: The heterogeneity must have been caused by horizontal gene transfer at a frequency lower than predicted from housekeeping gene data. Thus, it can be expected that species identification based on the ISR can be confused in rare isolates.     

Transfer of Campylobacter jejuni from raw to cooked chicken via wood and plastic cutting boards

Aims: We quantified Campylobacter jejuni transferred from naturally contaminated raw chicken fillets and skins to similar cooked chicken parts via standard rubberwood (RW) and polyethylene cutting boards (PE). Methods and Results: RW and PE cutting boards (2·5 × 2·5 cm²) were constructed. RW surfaces were smooth and even, whereas PE was uneven. Scoring with scalpel blades produced crevices on RW and flaked patches on the PE boards. Raw chicken breast fillets or skin pieces (10 g) naturally contaminated with Camp. jejuni were used to contaminate the cutting boards (6·25 cm²). These were then briefly covered with pieces of cooked chicken. Campylobacter jejuni on raw chicken, the boards, and cooked chicken pieces were counted using a combined most‐probable‐number (MPN)‐PCR method. The type of cutting board (RW, PE; unscored and scored) and temperature of cooked chicken fillets and skins were examined. Unscored PE and RW boards were not significantly different in regards to the mean transfer of Camp. jejuni from raw samples to the boards. The mean transfer of Camp. jejuni from scored RW was significantly higher than from scored PE. When the chicken fillets were held at room temperature, the mean transfer of Camp. jejuni from scored RW and PE was found to be 44·9 and 40·3%, respectively. Conclusions: RW and PE cutting boards are potential vehicles for Camp. jejuni to contaminate cooked chicken. Although cooked chicken maintained at high temperatures reduced cross‐contamination via contaminated boards, a risk was still present. Significance and Impact of the Study: Contamination of cooked chicken by Camp. jejuni from raw chicken via a cutting board is influenced by features of the board (material, changes caused by scoring) and chicken (types of chicken parts and temperature of the cooked chicken).     

Molecular typing of Campylobacter jejuni and C. coli from chickens and patients with gastritis or Guillain‐Barré syndrome based on multilocus sequence types and pulsed‐field gel electrophoresis patterns

Campylobacter jejuni has recently been noted as the most common cause of bacterial foodborne diseases in Japan. In the present study, we determined ST types of C. jejuni and Campylobacter coli isolated from chickens and patients with enteritis or GBS in Japan and Thailand. C. jejuni from chickens, enteritis, and GBS exhibited divergent ST types and included several novel types in addition to worldwide common types. C. coli from enteritis was also divergent. Novel ST types may represent unidentified native clones in each country. Pulsed‐field gel electrophoresis confirmed the above typing and demonstrated long‐term persistence and transmission.     

Campylobacter jejuni in Musca domestica: An examination of survival and transmission potential in light of the innate immune responses of the house flies

The house fly, Musca domestica, has been implicated as a vector of Campylobacter spp., a major cause of human disease. Little is known whether house flies serve as biological amplifying hosts or mechanical vectors for Campylobacter jejuni. We investigated the period after C. jejuni had been ingested by house flies in which viable C. jejuni colonies could be isolated from whole bodies, the vomitus and the excreta of adult M. domestica and evaluated the activation of innate immune responses of house flies to ingested C. jejuni over time. C. jejuni could be cultured from infected houseflies soon after ingestion but no countable C. jejuni colonies were observed > 24 h postingestion. We detected viable C. jejuni in house fly vomitus and excreta up to 4 h after ingestion, but no viable bacteria were detected ≥ 8 h. Suppression subtractive hybridization identified pathogen‐induced gene expression in the intestinal tracts of adult house flies 4–24 h after ingesting C. jejuni. We measured the expression of immune regulatory (thor, JNK, and spheroide) and effector (cecropin, diptericin, attacin, defensing, and lysozyme) genes in C. jejuni‐infected and ‐uninfected house flies using quantitative real time PCR. Some house fly factor, or combination of factors, eliminates C. jejuni within 24 h postingestion. Because C. jejuni is not amplified within the body of the housefly, this insect likely serves as a mechanical vector rather than as a true biological, amplifying vector for C. jejuni, and adds to our understanding of insect–pathogen interactions.     

Application of conductance techniques to the detection of thermophilic Campylobacter spp. in river water

A study of basal media identified Campylobacter enrichment broth, with (CEB+) and without (CEB) antibiotic supplement, as a suitable medium for the detection and enumeration of Campylobacter jejuni, C. coli and C. lari within aqueous samples via conductance methodology. Despite apparent differences in conductivity profiles between species in the presence of antibiotics, no significant differences (P<0.05) were detected between detection times for each species tested. CEB+ was successfully employed within a combined enrichment and conductance protocol to the detection of C. jejuni from river water at a concentration of 1 CFU ml⁻¹ from 83% of samples in under 39 h and thus demonstrated an improvement over an applied conventional membrane filtration technique.     

O‐Glycosylated 24 kDa human growth hormone has a mucin‐like biantennary disialylated tetrasaccharide attached at Thr‐60

MS was used to characterize the 24 kDa human growth hormone (hGH) glycoprotein isoform and determine the locus of O‐linked oligosaccharide attachment, the oligosaccharide branching topology, and the monosaccharide sequence. MALDI‐TOF/MS and ESI‐MS/MS analyses of glycosylated 24 kDa hGH tryptic peptides showed that this hGH isoform is a product of the hGH normal gene. Analysis of the glycoprotein hydrolysate by high‐performance anion‐exchange chromatography with pulsed amperometric detection and HPLC with fluorescent detection for N‐acetyl neuraminic acid (NeuAc) yielded the oligosaccharide composition (NeuAc₂, N‐acetyl galactosamine₁, Gal₁). After β‐elimination to release the oligosaccharide from glycosylated 24 kDa hGH, collision‐induced dissociation of tryptic glycopeptide T6 indicated that there had been an O‐linked oligosaccharide attached to Thr‐60. The sequence and branching structure of the oligosaccharide were determined by ESI‐MS/MS analysis of tryptic glycopeptide T6. The mucin‐like O‐oligosaccharide sequence linked to Thr‐60 begins with N‐acetyl galactosamine and branches in a bifurcated topology with one appendage consisting of galactose followed by NeuAc and the other consisting of a single NeuAc. The oligosaccharide moiety lies in the high‐affinity binding site 1 structural epitope of hGH that interfaces with both the growth hormone and the prolactin receptors and is predicted to sterically affect receptor interactions and alter the biological actions of hGH.     

Cloning and expression of the erythromycin resistance determinant from Campylobacter jejuni in Escherichia coli

The erythromycin resistance gene (Em^r) from Campylobacter jejuni ABA94 plasmid DNA was cloned into the pUC18 vector and then expressed in Escherichia coli. The location of the Em^r determinant on the chimeric plasmid was determined by restriction endonuclease mapping within a 0.8-kb EcoRI fragment. This fragment then hybridized to the 78-kb plasmid DNA but not to the 3.3-or 12.6-kb plasmid DNA of Campylobacter jejuni ABA94. Em^r in Campylobacter jejuni is therefore probably plasmid-mediated.The authors are with the Department of Genetics and Cellular Biology, University of Malaya, 59100 Kuala Lumpur, Malaysia     

Development and stability of bacteriocin resistance in Campylobacter spp

Aims: Several bacteriocins (BCNs) that were identified from chicken commensal bacteria dramatically reduced Campylobacter colonization in poultry and are being directed toward on‐farm control of this important foodborne human pathogen. A recent study has shown that BCN resistance in Campylobacter jejuni is very difficult to develop in vitro. In this study, in vivo development and stability of BCN resistance in Campylobacter was examined. Methods and Results: Chickens infected with Camp. jejuni NCTC 11168 were treated with BCN E‐760 at the dose of 5 mg kg^?1 body weight day^?1 via oral gavages for three consecutive days, which selected BCN‐resistant (BCN^r) mutants in the treated birds. However, all the in vivo‐selected mutants only displayed low levels of resistance to BCN (MIC = 2–8 mg l^?1) when compared to parent strain (MIC = 0·5 mg l^?1). Inactivation of CmeABC efflux pump of the BCN^r mutants led to increased susceptibility to BCN (8–32 fold MIC reduction). Three different BCN^rCampylobacter strains (in vitro‐ or in vivo‐derived) were examined for the stability of BCN resistance using both in vitro and in vivo systems. The low level of BCN resistance in these strains was not stable in vitro or in vivo in the absence of BCN selection pressure. Conclusions: Usage of BCN E‐760 only selected low‐level BCN^rCamp. jejuni mutants in vivo, and the low‐level BCN resistance was not stable in vitro and in vivo. Significance and Impact of the Study: The study provides helpful information for risk assessment of the future practical application of the anti‐Campylobacter BCNs in animals.     

Demonstration of persistent strains of Campylobacter jejuni within broiler farms over a 1‐year period in Lithuania

Aims: The aim of the study was to investigate the flock prevalence of Campylobacter jejuni and Campylobacter coli in broiler farms in Lithuania and to identify possible persistent strains of Camp. jejuni using amplified fragment length polymorphism (AFLP) typing method. Methods and Results: During 1 year, 42 broiler flocks from 9 broiler farms were examined to determine the prevalence of Campylobacter‐positive broiler flocks in Lithuania. Among 42 broiler flocks examined, 31 flocks (73·8%) were positive for Camp. jejuni and 17 flocks (40·48%) for Camp. coli. Campylobacter jejuni isolates were genotyped by AFLP method using BspDI and BglII restriction enzymes. Typing of 190 isolates generated 50 AFLP genotypes with the highest diversity of strains found in the summer season. Each farm showed one or more predominant AFLP types, and one AFLP type (A32) was found in five broiler farms over a 1‐year period. Conclusions: Campylobacter jejuni and Camp. coli are highly prevalent in broiler farms in Lithuania. Farm‐specific genotypes were identified in all farms examined. Type A32 was present and persisted in different broiler farms, and a common source of transmission of Camp. jejuni was suspected. Significance and Impact of the Study: For the first time, Camp. jejuni in broiler flocks has been genetically characterized in Lithuania. Persistent strains of Camp. jejuni were detected over one period at the beginning of broiler meat production chain and, therefore, the identification of contamination source of such strains and the mechanism of their particular ability to persist are crucial to establish effective control measures against Camp. jejuni infection in broiler farms.     

Fibronectin‐binding proteins of Clostridium perfringens recognize the III1‐C fragment of fibronectin

The Clostridium perfringens strain 13 genome contains two genes (fbpA, fbpB) that encode putative Fbp. Both rFbpA and rFbpB were purified and their reactivity with human serum Fn was analyzed. To determine the region of the Fn molecule recognized by rFbp, a plate binding assay using N‐terminal 70‐kDa peptide, III₁‐C peptide, and 110‐kDa peptide containing III_2–10 of Fn was performed. Both rFbp bound to the III₁‐C peptide of Fn but not to the other peptides. However, the III₁‐C fragment of Fn is known to be cryptic in serum Fn. Then, rFbp‐BP from Fn were purified by rFbp‐affinity chromatography. The yield of purified proteins was approximately 1% of the applied Fn on a protein basis. Western blotting analysis of the rFbp‐BP, using four different anti‐Fn monoclonal antibodies, revealed that the rFbp‐BP carried partial Fn antigenicity. Bindings of rFbp to rFbp‐BP were inhibited by the presence of the III₁‐C peptide, suggesting that rFbp‐BP express the III₁‐C fragment. The binding of Fn to III₁‐C was inhibited by the presence of either rFbpA or rFbpB. This result that suggests C. perfringens Fbps may inhibit the formation of Fn‐matrix in vivo.     

Occurrence of thermotolerant Campylobacter species in surface waters of a Mediterranean area and in its prevailing pollution sources

Aims: To determine the prevalence of Campylobacter in surface waters of a highly populated Mediterranean area. Methods and Results: Surface water and wastewater samples were collected from an area in the north‐east of Spain during a 2‐year study. All the samples were analysed using the MPN method and Multiplex PCR to quantify and identify Campylobacter. It was detected in 82% of the samples from the Llobregat River with a mean of 1·3 MPN 100 ml^?1. The lowest counts were obtained in summer. Campylobacter coli was the predominant species in this river. The bacteria were isolated from marsh water but not from seawater samples. The highest counts of campylobacters were found in poultry wastewater where Camp. jejuni was the predominant species, as in urban sewage. In pig slurry, Camp. coli was the only species detected. Conclusions: Campylobacter jejuni and Camp. coli are present and widely distributed in the surface water of the studied area. The two species co‐exist, with Camp. coli being predominant. In river water, campylobacter counts presented a seasonal distribution. No relationship with faecal indicators was found. Significance and Impact of the Study: This study provides the first data on the occurrence and concentrations of thermotolerant campylobacter species in surface water in a Mediterranean area.     

Crystal structure of the Campylobacter jejuni CmeC outer membrane channel

As one of the world's most prevalent enteric pathogens, Campylobacter jejuni is a major causative agent of human enterocolitis and is responsible for more than 400 million cases of diarrhea each year. The impact of this pathogen on children is of particular significance. Campylobacter has developed resistance to many antimicrobial agents via multidrug efflux machinery. The CmeABC tripartite multidrug efflux pump, belonging to the resistance‐nodulation‐cell division (RND) superfamily, plays a major role in drug resistant phenotypes of C. jejuni. This efflux complex spans the entire cell envelop of C. jejuni and mediates resistance to various antibiotics and toxic compounds. We here report the crystal structure of C. jejuni CmeC, the outer membrane component of the CmeABC tripartite multidrug efflux system. The structure reveals a possible mechanism for substrate export.