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

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

Genomic Diversity of Campylobacter lari Group Isolates from Europe and Australia in a One Health Context.

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.

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

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

A catalytically essential motif in external loop 5 of the bacterial oligosaccharyltransferase PglB.

Asparagine-linked glycosylation is a post-translational protein modification that is conserved in all domains of life. The initial transfer of a lipid-linked oligosaccharide (LLO) onto acceptor asparagines is catalyzed by the integral membrane protein oligosaccharyltransferase (OST). The previously reported structure of a single-subunit OST enzyme, the Campylobacter lari protein PglB, revealed a partially disordered external loop (EL5), whose role in catalysis was unclear. We identified a new and functionally important sequence motif in EL5 containing a conserved tyrosine residue (Tyr293) whose aromatic side chain is essential for catalysis. A synthetic peptide containing the conserved motif can partially but specifically rescue in vitro activity of mutated PglB lacking Tyr293. Using site-directed disulfide cross-linking, we show that disengagement of the structurally ordered part of EL5 is an essential step of the glycosylation reaction, probably by allowing sequon binding or glyco-product release. Our findings define two distinct mechanistic roles of EL5 in OST-catalyzed glycosylation. These functions, exerted by the two halves of EL5, are independent, because the loop can be cleaved by specific proteolysis with only slight reduction in activity.

Molecular structural analysis of major outer membrane protein (MOMP) gene clusters in Campylobacter lari.

Southern hybridisation shows that urease-negative (UN) Campylobacter lari JCM2530(T) carries two putative major outer membrane protein (MOMP) genes. Sequences of approximately 2.1 kbp, encoding non-coding (NC) regions, with possible open reading frames (ORFs) for MOMP (porA1 or porA2) of approximately 1.2 kbp, NC regions and partial and putative Cla_0435 or Cla_1109 ORFs were identified in all five UN C. lari isolates examined, following polymerase chain reaction (PCR) cloning and sequencing. Each putative MOMP structural gene carried start and stop codons and ribosome binding sites of 1236-1278 bp in length. The putative sigma70 transcriptional promoter and the hypothetical rho-independent transcription terminator structures were also seen. Using Northern hybridisation, there was in vivo monocistronic MOMP gene transcription. In addition, in a Japanese urease-positive thermophilic Campylobacter (UPTC) CF89-12 strain, the porA1 gene locus, including an extra gene (approximately 2000 bp in length) was identified. The extra gene may occur within the porA1 gene locus in the eight UPTC isolates of the 23 C. lari isolates examined. Thus, a genetic heterogeneity occurred within the porA1 gene locus from some of the C. lari organisms including the UPTC CF89-12.

Bloodstream infection caused by Campylobacter lari.

We describe a case of bloodstream infection (BSI) caused by Campylobacter lari in a 58-year-old man diagnosed with lumbar pyogenic spondylitis. Anaerobic blood cultures, taken on the day of admission and on hospital day 4, were positive after 30 h of incubation, although no bacteria were detected by Gram staining. After subculture on 5 % sheep blood agar for 2 days at 35 °C in a 5 % CO2 environment, capnophilic, curved, gram-negative bacteria were recovered. The bacteria were identified as C. lari using a combination of phenotypic identification methods and partial 16S rRNA gene sequencing. The BSI was eradicated following combination therapy with intravenous tazobactam/piperacillin, oral erythromycin, and sulfamethoxazole/trimethoprim. These results suggest that accurate identification, to the species level, is important to determine effective treatment of BSI caused by Campylobacter spp. and can help us to understand the epidemiology.

Molecular cloning and characterisation of the methionine sulphoxide reductase A (msrA) gene locus in Campylobacter lari organisms.

The methionine sulphoxide reductase A (msrA) gene and its adjacent genetic loci from urease-negative (UN) Campylobacter lari RM2100 and urease-positive thermophilic Campylobacter (UPTC)CF89-12 strains appear to be composed of a msrA structure gene (507 base pairs [bp]) and another five-gene cluster (approximately 6300 bp) in the same strand and direction. A primer pair (F1/R4-msrA) for polymerase chain reaction (PCR) amplification was designed to generate a product of approximately 900 bp of the msrA gene, including its adjacent genetic loci for the thermophilic Campylobacter organisms and generate an amplicon with 16 C. lari isolates (n = 4 for UN C. lari; n = 12 for UPTC). Following direct nucleotide sequencing, sequence analysis and nucleotide sequence alignment analysis, the putative full-length msrA gene from the 16 C. lari isolates showed high nucleotide sequence similarities (91.8-100%) to each other and relatively low similarity (69.3-71.8%) to three reference C. jejuni and C. coli strains. In addition, the msrA gene was transcribed in both the UPTC CF89-12 and NCTC12893 cells using reverse transcription PCR. An immunoreactively positive signal was identified in the UPTC CF89-12 and NCTC12893 cells with anti-UPTC MsrA synthetic peptide antibodies.

Effects of DNA extraction methods on the real time PCR quantification of Campylobacter jejuni, Campylobacter coli, and Campylobacter lari in chicken feces and ceca contents.

Polymerase chain reaction (PCR) method was coupled with a DNA extraction to enumerate Campylobacter spp. from poultry gastrointestinal tract samples. Three experiments were conducted that included: 1) Development of a DNA standard curve related to bacterial DNA primers; 2) Design of a cell/genomic DNA extraction protocol to isolate Campylobacter spp. DNA from complex samples such as poultry feces; and 3) Comparison of PCR quantification to standard plate count methodology. The standard curve using primers for Campylobacter spp. was created for DNA extracted from environmental isolates with a linear range (R2 > 0.95) and with a high specificity for C. coli and C. jejuni recovered from poultry, swine and laboratory isolates. A 2-step extraction process of bacterial DNA from poultry feces was developed in which the cells were first concentrated using a gradient-centrifugation step followed by comparison of 4 DNA extraction methods. Two commercial DNA extraction methods (Zymo Research Quick DNA, and Invitrogen magnetic separation), a traditional phenol-chloroform DNA extraction method using proteinase K to inactivate DNAses, and an in-house isolation method for DNA extraction based on chaotropic salts were used. The middle gradient layer recovered 89% to 98% of the bacteria cells from the sample, with recovery dependent upon the Campylobacter genus. The 4 DNA extractions methods recovered 112 to 302 ug/nL of DNA. Finally, the qPCR and standard plate methods were highly correlated for enumerating Campylobacter spp. in the 2.0 to 8.0-log CFU range. Analyses of the results from this study demonstrate that the combination of the standard curve for Campylobacter spp. DNA primers, the gradient cell concentration method and DNA extraction techniques with qPCR can be used to enumerate Campylobacter spp. from poultry samples with findings similar those of traditional plate count methodology.

Expression and analysis of a cytolethal distending toxin (cdt) gene operon in Campylobacter lari.

The present study examines the expression of cytolethal distending toxin (cdt) gene encoding a cytotoxin in Campylobacter lari (n=6 urease-negative [UN] C. lari; n=4 urease-positive thermophilic Campylobacter [UPTC]). When reverse transcription polymerase chain reaction (RT-PCR) was carried out with 10 C. lari isolates using a primer pair to amplify the cdtB gene transcript segment, an approximate 260 bp RT-PCR amplicon was generated with all the isolates. In addition, cdtA, cdtB and cdtC gene operon was identified to be polycistronicly transcribed in the C. lari cells. The cdtB gene translation in the C. lari cells was also confirmed by Western blot analysis. Thus, the cdt gene operon in C. lari organisms, including UN C. lari and UPTC, was expressed at the transcriptional and translational levels in the cells. The present results suggest that all three cdt genes may be functional in the cells.

Molecular analysis of the 16S-23S rDNA internal spacer region (ISR) and truncated tRNA(Ala) gene segments in Campylobacter lari.

Following PCR amplification and sequencing, nucleotide sequence alignment analyses demonstrated the presence of two kinds of 16S-23S rDNA internal spacer regions (ISRs), namely, long length ISRs of 837-844 base pair (bp) [n = six for urease-negative (UN) Campylobacter lari isolates, UN C. lari JCM2530(T), RM2100, 176, 293, 299 and 448] and short length ISRs of 679-725 bp [n = six for UN C. lari: n = 14 for urease-positive thermophilic Campylobacter (UPTC) isolates]. The analyses also indicated that the short length ISRs mainly lacked the 156 bp sequence from the nucleotide positions 122-277 bp in long length ISRs for UN C. lari JCM2530(T). The 156 bp sequences shared 94.9-96.8 % sequence similarity among six isolates. Surprisingly, atypical tRNA(Ala) gene segment (5' end 35 bp), which was extremely truncated, occurred within the 156 bp sequences in the long length ISRs, as an unexpected tRNA(Ala) pseudogene. An order of the intercistronic tRNA genes within the short nucleotide spacer of 5'-16S rDNA-tRNA(Ala)-tRNA(Ile)-23S rDNA-3' occurred in all the C. lari isolates examined.

Molecular analysis and characterisation of the full-length flagellin C gene (flaC) from Campylobacter lari.

A degenerate polymerase chain reaction (PCR) primer pair (f-ClflaC/r-ClflaC) was constructed in silico to amplify flaC and its adjacent genetic loci from Campylobacter lari isolates. Approximately 1.45 kbp amplicons, including the sequences encoding the flaC structural gene of 750 bp, putative promoter, rho-independent intrinsic terminator regions and partial sequences of two putative open reading frames (ORFs), immediately upstream and downstream of the gene, were identified in 16 C. lari isolates (four urease-negative [UN] C. lari; 12 urease-positive thermophilic campylobacters [UPTC)]). All 16 flaC structural genes commenced with an ATG start codon and terminated with a TAA stop codon and probable ribosome-binding sites were identified in all 16 isolates. These probably indicate a monocistronic operon structure for the flaC gene in C. lari isolates. In addition, the putative flaC gene ORFs were deduced to be similar in 747 bp among all 26 thermophilic Campylobacter isolates examined, resulting in a similar calculated molecular weight of approximately 26.6-26.9 kDa. The flaC from C. lari was different from the flaA-like sequence and the shorter flaA of UPTC isolates found previously. Reverse transcription PCR and Northern blot hybridisation analyses identified flaC transcription in C. lari cells. The transcription initiation site for the flaC gene was also determined by primer extension analysis. A dendrogram constructed, based on the nucleotide sequence information of flaC from 17 C. lari isolates, demonstrated that the C. lari isolates were genetically variable and formed two minor clusters for UN C. lari and UPTC.

Rapid detection and differentiation of Campylobacter jejuni, Campylobacter coli, and Campylobacter lari in food, using multiplex real-time PCR.

A multiplex real-time PCR assay based on four differently labeled TaqMan probes for detection and differentiation of the thermophilic Campylobacter species C. jejuni, C. coli, and C. lari was established and validated in food products. This assay combines two previously published PCR assays for C. jejuni and C. coli with a newly developed detection assay for C. lari and an internal amplification control system. The selectivity of the method was determined by analyzing 70 Campylobacter strains and 43 strains of other bacteria. The sensitivity was 50 fg of C. jejuni and C. lari DNA and 500 fg of C. coli DNA per PCR. It was possible to detect 1 to 10 CFU/25 g of food before preenrichment of all three species. More than 400 samples of various foods (poultry, seafood, and meat) were analyzed after 48 h of preenrichment parallel to the conventional diagnostic method of culture and biochemical identification. Using the established real-time PCR assay, 55.4% of the samples were recognized as positive for thermophilic Campylobacter species, whereas with the conventional method only 40.3% of the samples were positive. The real-time PCR assay also detected contaminations with two different Campylobacter species in 32.6% of the analyzed poultry samples, a finding of epidemiological interest. Compared with the original PCR method, which was established for the differentiation of bacterial isolates of C. jejuni and C. coli, this new method also detects and distinguishes C. lari, was validated as an analytical tool for food analysis, and provides reliable and extensive results within 2 days.

Structural analysis and expression of the full-length cytochrome P450 gene operon in Campylobacter lari.

Two sets of PCR primers are constructed to clone the cytochrome P450 structural gene, including putative promoter and terminator structures, and its adjacent genetic loci in Campylobacter lari isolates. The putative open reading frames (ORFs) of the P450 genes from 11 C. lari isolates (n=5 for urease-negative (UN) C. lari; n=6 urease-positive thermophilic campylobacters [UPTC]) examined consisted of 1365 or 1371 bases (455 or 457 amino acid residues), differing from those of the other thermophilic campylobacters (1359 [453] for C. jejuni and C. upsaliensis; 1368 [456] for C. coli). Each of the putative ORFs from the 11 isolates examined was also shown to carry start and stop codons and ribosome binding sites. Two putative promoter structures, consisting of sequences at the -35- and -10-like regions were also identified upstream of the ORFs. A single copy of the P450 gene in the genome was identified with UN C. lari JCM2530(T) and UPTC CF89-12, based on Southern blot hybridisation analysis. In addition, when reverse transcription polymerase chain reaction (RT-PCR) analyses were carried out, the transcription of the P450 structural gene in C. lari organisms in vivo was confirmed. The transcription initiation site for the gene was also determined. High nucleotide sequence similarities (95.2-98.8%) of the full-length P450 structural gene were shown with each of the 12 C. lari isolates. The UN C. lari and UPTC organisms showed similar findings with the neighbour-joining method, based on the sequence information of the P450 structural gene.

Structural analysis of the full-length gene encoding a fibronectin-binding-like protein (CadF) and its adjacent genetic loci within Campylobacter lari.

BACKGROUND: The combined sequences encoding a partial and putative rpsI open reading frame (ORF), non-coding (NC) region, a putative ORF for the Campylobacter adhesin to fibronectin-like protein (cadF), a putative Cla_0387 ORF, NC region and a partial and putative Cla_0388 ORF, were identified in 16 Campylobacter lari isolates, using two novel degenerate primer pairs. Probable consensus sequence at the -35 and -10 regions were identified in all C. lari isolates, as a promoter. RESULTS: Thus, cadF (-like) gene is highly conserved among C. lari organisms. Transcription of the cadF (-like) gene in C. lari cells in vivo was also confirmed and the transcription initiation site was determined. A peptidoglycan-associating alpha-helical motif in the C-terminal regions of some bacterial cell-surface proteins was completely conserved amongst the putative cadF (-like) ORFs from the C. lari isolates. CONCLUSION: The putative cadF (-like) ORFs from all C. lari isolates were nine amino acid larger than those from C. jejuni, and showed amino acid residues 137 -140 of FALG (50% identity), instead of the FRLS residues of the maximal fibronectin-binding activity site demonstrated within C. jejuni CadF. A neighbor joining tree constructed based on cadF (-like) gene sequence information formed a major cluster consisting of C. lari isolates, separating from the other three thermophilic campylobacters.

Campylobacter lari: molecular and comparative analyses of the virulence-associated chromosome locus J (vacJ) gene homologue, including the promoter region.

Following TA cloning and sequencing with a novel in silico-designed polymerase chain reaction (PCR) primer pair (f-ClvacJ/r-ClvacJ), approximately 750 base pairs (bp) of promoter and structural gene regions for vacJ and its adjacent genetic loci (approximately 1.14 kbp) were identified in 20 isolates of Campylobacter lari (urease-negative C. lari [n=7]; urease-positive thermophilic Campylobacter [n=13]). The nucleotide sequences of an approximately 70-bp non-coding region, including the typical promoter structure, showed sequence differences at 12 loci among 21 isolates including C. lari RM2100. The putative sigma70 promoter region upstream of the putative open reading frame (ORF), a start codon TTG and a probable ribosome binding site, AGGA, for the vacJ gene were also identified in all 21 C. lari isolates examined. Each ORF for the vacJ terminated with a TAA stop codon. No hypothetical transcriptional terminators were identified within the amplicons. The putative ORFs of the vacJ gene from 21 C. lari isolates consisted of 684 bases, similarly differing from those of the other thermophilic campylobacters (696 bases for C. jejuni RM1221 and NCTC11168 and C. coli RM2228; 690 for C. upsaliensis RM3195). Reverse transcription PCR analysis confirmed the transcription of the vacJ gene in the C. lari cells. A neighbour joining tree suggested a strong molecular discrimination efficacy between UPTC and UN C. lari employing vacJ nucleotide sequence information. The vacJ gene homologue from C. lari organisms appears not to be a lipoprotein signal peptide or a signal peptide in silico.

Demonstration of the absence of intervening sequences within 23S rRNA genes from Campylobacter lari.

Cloning, sequencing and characterization of nearly full-length 23S rRNA genes in 12 urease-positive thermophilic Campylobacter (UPTC) isolates were carried out using two novel PCR primer pairs. Nucleotide sequences of the 23S rRNA genes from the 12 isolates were first shown not to carry any intervening sequences (IVSs) in both the 25 and 45 helix regions. Then, two PCR primer sets were designed in silico for amplification of the helix 25 and 45 regions within 23S rRNA gene sequences from Campylobacter lari. No IVSs were identified within the 23S rRNA genes among a total of 53 isolates of C. lari, following PCR amplification, TA cloning and sequencing procedures. Intact 23S rRNA was identified in all 65 C. lari isolates, resulting in no production of the fragmented 23S rRNA. These data suggest that C. lari may not have any opportunity to interact with any other source of IVSs until now, or has been unable to integrate IVSs into their own genomes.

Cloning, sequencing and expression of full-length Campylobacter invasion antigen B gene operon from Campylobacter lari.

A novel PCR primer pair for amplification of full-length cia B gene from thermophilic campylobacters, generated an amplicon of approximately 2.2 kilo base pairs (kbp) with all 18 isolates (n = 7 for urease-negative (UN) C. lari; n = 9 urease-positive thermophilic Campylobacter (UPTC); n = 1 C. jejuni; n = 1 C. coli). The putative open reading frame (ORF) of the cia B from C. lari isolates consisted of 1,833 bp similarly, but differing from those of C. jejuni and C. coli isolates. The putative promoter structures consisting of a semi-conserved T -rich sequence and a consensus sequence at the -10 region were identified upstream of the putative ORF in all the C. lari isolates. A start codon ATG and a probable ribosome binding site were also identified in all the isolates. In addition, two distinctly different and taxon (UN C. lari and UPTC) dependent hypothetically intrinsic rho -independent transcriptional terminators for the cia B were identified to occur within the C. lari. Reverse transcription-PCR analysis identified the transcription of cia B gene in the C. lari cells. The neighbor joining tree suggested that the nucleotide sequence information of the cia B had molecular discrimination efficacy among UN C. lari, UPTC, C. jejuni and C. coli organisms.

The omp50 gene is transcriptionally controlled by a temperature-dependent mechanism conserved among thermophilic Campylobacter species.

The thermophilic Campylobacters are enteropathogenic for humans. We recently showed that Omp50 is a Campylobacter species-specific porin produced in Campylobacter jejuni and Campylobacter lari but not in Campylobacter coli. In the present study, we investigated regulation of the omp50 gene and found that its expression in C. jejuni was temperature-dependent, but independent of growth phase or medium viscosity. The use of RT-PCR and omp50::lacZ fusions showed that growth temperature control occurred at the transcriptional level. The promoter and the coding sequence were cloned in an Escherichia coli-Campylobacter shuttle plasmid and transferred to E. coli and to a C. jejuni Omp50-deficient strain. Regulation of omp50 gene expression by growth temperature was observed in the recombinant C. jejuni strain, but not in E. coli. The same regulation was also observed in wild-type C. lari strains and in a C. coli strain supplemented by the plasmid, suggesting that omp50 expression is controlled by a mechanism conserved among Campylobacter species.

The complete genome sequence and analysis of the human pathogen Campylobacter lari.

Campylobacter lari is a member of the epsilon subdivision of the Proteobacteria and is part of the thermotolerant Campylobacter group, a clade that includes the human pathogen C. jejuni. Here we present the complete genome sequence of the human clinical isolate, C. lari RM2100. The genome of strain RM2100 is approximately 1.53 Mb and includes the 46 kb megaplasmid pCL2100. Also present within the strain RM2100 genome is a 36 kb putative prophage, termed CLIE1, which is similar to CJIE4, a putative prophage present within the C. jejuni RM1221 genome. Nearly all (90%) of the gene content in strain RM2100 is similar to genes present in the genomes of other characterized thermotolerant campylobacters. However, several genes involved in amino acid biosynthesis and energy metabolism, identified previously in other Campylobacter genomes, are absent from the C. lari RM2100 genome. Therefore, C. lari RM2100 is predicted to be multiply auxotrophic, unable to synthesize eight different amino acids, acetyl-coA, and pantothenate. Additionally, strain RM2100 does not contain a complete TCA cycle and is missing the CydAB terminal oxidase of the respiratory chain. Defects in the amino acid biosynthetic pathways in this organism could be potentially compensated by the large number of encoded peptidases. Nevertheless, the apparent absence of certain key enzymatic functions in strain RM2100 would be expected to have an impact on C. lari biology. It is also possible that the reduction in the C. lari metabolic machinery is related to its environmental range and host preference.

Similarity of Campylobacter lari among human, animal, and water isolates in Norway.

A total of 49 isolates of Campylobacter lari from human, poultry, ducks, pigs, and water were genetically characterized. The species were identified by biotyping and multiplex polymerase chain reaction (PCR). Automatic riboprints were performed with the PstI restriction enzyme and RiboPrinter. The identification of the isolates was predicted when the corresponding pattern matched one of the patterns of the DuPont identification (DUP-ID) library and was then assigned an identification number. Thirty-five (71.4%) of the isolates were given a DUP-ID number. The isolates from water and animals showed a high degree of similarity to the human strains represented by DUP-PST1-1010, DUP-PST1-1166, DUP-PST1-1178, and DUP-PST1-1081. Some profiles (i.e., DUP-PST1-2021 and DUP-PST1-1184) were found only among the human isolates. Dendrogram analysis using BioNumerics grouped isolates into three main clusters. One of those clusters contained DUP-PST1-2021, DUP-PST1-1184, and DUP-PST1-1081, which was found in both humans and ducks. A second cluster generated DUP-PST1-1010, found in both humans and poultry, and DUP-PST1-1079, found in water. The third cluster consisted of two strains, DUP-PST1-1066 and DUP-PST1-1078, originating in humans, animals, and water. Three human strains and two poultry strains were diverse and formed their own clusters and could not be assigned a DUP-ID number. Because of the similarity of C. lari isolated from humans, poultry, ducks, pigs, and water, as well as the limited knowledge of environmental survival and its virulence factors, special hygienic precautions should be taken to avoid the risk of transmitting Campylobacter.