Whole-Genome Sequencing to Detect Numerous Campylobacter jejuni Outbreaks and Match Patient Isolates to Sources, Denmark, 2015-2017.
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ABSTRACT:
SUBMITTER: Joensen KG
PROVIDER: S-EPMC7045838 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Bacterial genome sequencing has led to the development of new approaches for the analysis of food-borne epidemics and the exploration of the relatedness of outbreak-associated isolates and their separation from nonassociated isolates. Using Illumina technology, we sequenced a total of six isolates (two from patients, two from raw bulk milk, and two from dairy cattle) associated with a milk-borne Campylobacter jejuni outbreak in a farming family and compared their genomes. These isolates had identical pulsed-field gel electrophoresis (PFGE) types, and their multilocus sequence typing (MLST) type was ST-50. We used the Ma_1 isolate (milk) as the reference, and its genome was assembled and tentatively ordered using the C. jejuni NCTC 11168 genome as the scaffold. Using whole-genome MLST (wgMLST), we identified a total of three single-nucleotide polymorphisms (SNPs) and differences in poly(G or C) or poly(A or T) tracts in 12 loci among the isolates. Several new alleles not present in the database were detected. In contrast, the sequences of the unassociated C. jejuni strains P14 and 1-12S (both ST-50) differed by 420 to 454 alleles from the epidemic-associated isolates. We found that the fecal contamination of bulk tank milk occurred by highly related sequence variants of C. jejuni, which are reflected as SNPs and differences in the length of the poly(A or T) tracts. Poly(G or C) tracts are reversibly variable and are thus unstable markers for comparison. Further, unrelated strains of ST-50 were clearly separated from the outbreak-associated isolates, indicating that wgMLST is an excellent tool for analysis. In addition, other useful data related to the genes and genetic systems of the isolates were obtained.
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Project description:Campylobacter jejuni strains exhibit significant variation in the genetic content of the lipooligosaccharide (LOS) biosynthesis loci with concomitant differences in LOS structure. The C. jejuni LOS loci have been grouped into six classes based on gene content and organization. Utilizing PCR amplifications of genes from these loci, we were able to classify a majority (80%) of the LOS biosynthesis loci from 123 strains of C. jejuni that included 39 of the Penner serotype reference strains. We found that a particular LOS class was not always associated with a specific Penner serotype, and 14 of 16 Guillain-Barré syndrome-associated isolates tested in this study shared the same LOS class. The remaining isolates that could not be classified were often distinguishable from each other based on the results of gene-specific PCR and the lengths of their LOS biosynthesis loci as determined by long (XL) PCR. Sequence analysis of two of these unique XL PCR products demonstrated two new LOS classes. These results support the hypothesis that the LOS locus is a hot spot for genetic exchange and rearrangements. Analysis of the LOS biosynthesis genes by PCR assays can be used for typing C. jejuni and offers the advantage of inferring potential LOS structures.
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Project description:Campylobacter jejuni (C. jejuni) are able to colonise and infect domestic poultry and also pose a risk for humans. The aim of this study was to determine the extent of genotypic diversity among C. jejuni isolates recovered from avian and human sources in Egypt. Furthermore, the short variable region (SVR) of flagellin A (flaA) gene was analysed for the presence of allelic variants. Our results showed that C. jejuni isolates differ in their capacity to harbour each of the virulence genes alone or when present in various combinations. The flaA gene was detected in all C. jejuni strains and none of the strains had all the studied virulence genes together. When considering C. jejuni strains from the investigated sources, the cdtC gene was the most similar, while the cdtB and iam genes were the most dissimilar. We could identify 13 novel alleles in the analysed strains. The analyses of virulence gene patterns, flaA gene sequences and allelic variants showed that C. jejuni strains from different sources overlapped largely suggesting potential involvement of poultry in transmitting C. jejuni to humans. We also found that the strains isolated from the same host were highly heterogeneous, with chicken strains exhibiting the highest diversity. Moreover, the human strains were clustered closer to chicken ones than to those from pigeon. The results of this study should be taken into consideration when assessing the epidemiology and risk potential of Egyptian C. jejuni not only in poultry, but also in humans.
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Project description:BackgroundCampylobacter jejuni and Campylobacter coli are human intestinal pathogens of global importance. Zoonotic transmission from livestock animals or animal-derived food is the likely cause for most of these infections. However, little is known about their general and host-specific mechanisms of colonization, or virulence and pathogenicity factors. In certain hosts, Campylobacter species colonize persistently and do not cause disease, while they cause acute intestinal disease in humans.ResultsHere, we investigate putative host-specificity using phenotypic characterization and genome-wide analysis of genetically closely related C. jejuni strains from different sources. A collection of 473 fresh Campylobacter isolates from Germany was assembled between 2006 and 2010 and characterized using MLST. A subset of closely related C. jejuni strains of the highly prevalent sequence type ST-21 was selected from different hosts and isolation sources. PCR typing of strain-variable genes provided evidence that some genes differed between these strains. Furthermore, phenotypic variation of these strains was tested using the following criteria: metabolic variation, protein expression patterns, and eukaryotic cell interaction. The results demonstrated remarkable phenotypic diversity within the ST-21 group, which however did not correlate with isolation source. Whole genome sequencing was performed for five ST-21 strains from chicken, human, bovine, and food sources, in order to gain insight into ST-21 genome diversity. The comparisons showed extensive genomic diversity, primarily due to recombination and gain of phage-related genes. By contrast, no genomic features associated with isolation source or host were identified.ConclusionsThe genome information and phenotypic data obtained in vitro and in a chicken infection model provided little evidence of fixed adaptation to a specific host. Instead, the dominant C. jejuni ST-21 appeared to be characterized by phenotypic flexibility and high genetic microdiversity, revealing properties of a generalist. High genetic flexibility might allow generalist variants of C. jejuni to reversibly express diverse fitness factors in changing environments.
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Project description:Campylobacter spp. is a significant and prevalent public health hazard globally. Campylobacter jejuni is the most frequently recovered species from human cases and poultry are considered the most important reservoir for its transmission to humans. In this study, 30 Campylobacter jejuni isolates were selected from clinical (n = 15) and broiler (n = 15) sources from a larger cohort, based on source, virulence, and antimicrobial resistance profiles. The objective of this study was to further characterise the genomes of these isolates including MLST types, population structure, pan-genome, as well as virulence and antimicrobial resistance determinants. A total of 18 sequence types and 12 clonal complexes were identified. The most common clonal complex was ST-45, which was found in both clinical and broiler samples. We characterised the biological functions that were associated with the core and accessory genomes of the isolates in this study. No significant difference in the prevalence of virulence or antimicrobial resistance determinants was observed between clinical and broiler isolates, although genes associated with severe illness such as neuABC, wlaN and cstIII were only detected in clinical isolates. The ubiquity of virulence factors associated with motility, invasion and cytolethal distending toxin (CDT) synthesis in both clinical and broiler C. jejuni genomes and genetic similarities between groups of broiler and clinical C. jejuni reaffirm that C. jejuni from poultry remains a significant threat to public health.
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Project description:Whole-genome sequencing (WGS) via next-generation sequencing (NGS) technologies is a powerful tool for determining the relatedness of bacterial isolates in foodborne illness detection and outbreak investigations. WGS has been applied to national outbreaks (for example, Listeria monocytogenes); however, WGS has rarely been used in smaller local outbreaks. The current study demonstrates the superior resolution of genetic and evolutionary relatedness generated by WGS data analysis, compared to pulsed-field gel electrophoresis (PFGE). The current study retrospectively applies WGS and a reference-free bioinformatic analysis to a Utah-specific outbreak of Campylobacter jejuni associated with raw milk and to a national multistate outbreak of Salmonella enterica subsp. enterica serovar Typhimurium associated with rotisserie chicken, both of which were characterized previously by PFGE. Together, these analyses demonstrate how a reference-free WGS workflow is not reliant on determination of a reference sequence, like WGS workflows that are based on single-nucleotide polymorphisms, or the need for curated allele databases, like multilocus sequence typing workflows.
| S-EPMC6204689 | biostudies-literature
Project description:Transcriptional regulation mediates adaptation of pathogens to environmental stimuli and is important for host colonisation. The Campylobacter jejuni genome sequence reveals a surprisingly small set of regulators, mostly of unknown function, suggesting an intricate regulatory network. Interestingly, C. jejuni lacks the homologues of ubiquitous regulators involved in stress response found in many other Gram-negative bacteria. Nonetheless, cj1000 is predicted to code for the sole LysR-type regulator in the C. jejuni genome, and thus may be involved in major adaptation pathways. A cj1000 mutant strain was constructed and found to be attenuated in its ability to colonise 1-day old chicks. Complementation of cj1000 mutation restored the colonisation ability to that of wild type levels. The mutant strain was also outcompeted in a competitive colonisation assay of the piglet intestine. High resolution oxygraphy was carried out for the first time on C. jejuni and revealed a role for Cj1000 in controlling O2 consumption. Furthermore, microarray analysis of the cj1000 mutant revealed both direct and indirect regulatory targets, including genes involved in energy metabolism and oxidative stress defences. These results highlight the importance of Cj1000 regulation in host colonisation and in major physiological pathways.
2013-07-03 | GSE43792 | GEO
Project description:Growth of Campylobacter jejuni in butyrate, including deletions of a TCS involved in sensing this response
2021-01-01 | GSE142852 | GEO
Project description:Campylobacter jejuni is one of the leading causes of bacterial gastroenteritis worldwide. In the United States, New Hampshire was one of the 18 states that reported cases in the 2016 to 2018 multistate outbreak of multidrug-resistant C. jejuni Here, we aimed to elucidate the baseline diversity of the wider New Hampshire C. jejuni population during the outbreak. We used genome sequences of 52 clinical isolates sampled in New Hampshire in 2017, including 1 of the 2 isolates from the outbreak. Results revealed a remarkably diverse population composed of at least 28 sequence types, which are mostly represented by 1 or a few strains. A comparison of our isolates with 249 clinical C. jejuni from other states showed frequent phylogenetic intermingling, suggesting a lack of geographical structure and minimal local diversification within the state. Multiple independent acquisitions of resistance genes from 5 classes of antibiotics characterize the population, with 47/52 (90.4%) of the genomes carrying at least 1 horizontally acquired resistance gene. Frequently recombining genes include those associated with heptose biosynthesis, colonization, and stress resistance. We conclude that the diversity of clinical C. jejuni in New Hampshire in 2017 was driven mainly by the coexistence of phylogenetically diverse antibiotic-resistant lineages, widespread geographical mixing, and frequent recombination. This study provides an important baseline census of the standing pangenomic variation and drug resistance to aid the development of a statewide database for epidemiological studies and clinical decision making. Continued genomic surveillance will be necessary to accurately assess how the population of C. jejuni changes over the long term.
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Project description:The Gram-negative pathogen Campylobacter jejuni is the most common cause of bacterial foodborne disease worldwide. The mechanisms that lead to bacterial invasion of eukaryotic cells and massive intestinal inflammation are still unknown. In this study, we report that C. jejuni infection of mouse macrophages induces upregulation of pro-IL-1? transcript and secretion of IL-1? without eliciting cell death. Immunoblotting indicated cleavage of caspase-1 and IL-1? in infected cells. In bone marrow-derived macrophages from different knockout mice, IL-1? secretion was found to require NLRP3, ASC, and caspase-1/11 but not NLRC4. In contrast to NLRP3 activation by ATP, C. jejuni activation did not require priming of these macrophages. C. jejuni also activated the NLRP3 inflammasome in human macrophages as indicated by the presence of ASC foci and caspase-1-positive cells. Analysis of a vast array of C. jejuni mutants with defects in capsule formation, LPS biosynthesis, chemotaxis, flagella synthesis and flagellin (-like) secretion, type 6 secretion system needle protein, or cytolethal distending toxin revealed a direct correlation between the number of intracellular bacteria and NLRP3 inflammasome activation. The C. jejuni invasion-related activation of the NLRP3 inflammasome without cytotoxicity and even in nonprimed cells extends the known repertoire of bacterial inflammasome activation and likely contributes to C. jejuni-induced intestinal inflammation.
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