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Core Genome MLST for Source Attribution of Campylobacter coli.

ABSTRACT: Campylobacter species are among the leading foodborne bacterial agents of human diarrheal illness. The majority of campylobacteriosis has been attributed to Campylobacter jejuni (85% or more), followed by Campylobacter coli (5-10%). The distribution of C. jejuni and C. coli varies by host organism, indicating that the contribution to human infection may differ between isolation sources. To address the relative contribution of each source to C. coli infections in humans, core genome multilocus sequence type with a 200-allele difference scheme (cgMLST₂₀₀) was used to determine cgMLST type for 3,432 C. coli isolated from food animals (n = 2,613), retail poultry meats (n = 389), human clinical settings (n = 285), and environmental sources (n = 145). Source attribution was determined by analyzing the core genome with a minimal multilocus distance methodology (MMD). Using MMD, a higher proportion of the clinical C. coli population was attributed to poultry (49.6%) and environmental (20.9%) sources than from cattle (9.8%) and swine (3.2%). Within the population of C. coli clinical isolates, 70% of the isolates that were attributed to non-cecal retail poultry, dairy cattle, beef cattle and environmental waters came from two cgMLST₂₀₀ groups from each source. The most common antibiotic resistance genes among all C. coli were tetO (65.6%), bla _OXA _- ₁₉₃ (54.2%), aph(3')-IIIa (23.5%), and aadE-Cc (20.1%). Of the antibiotic resistance determinants, only one gene was isolated from a single source: bla _OXA _- ₆₁ was only isolated from retail poultry. Within cgMLST₂₀₀ groups, 17/17 cgMLST₂₀₀-435 and 89/92 cgMLST₂₀₀-707 isolates encoded for aph(3')-VIIa and 16/16 cgMLST₂₀₀-319 harbored aph(2')-If genes. Distribution of bla _OXA alleles showed 49/50 cgMLST₂₀₀-5 isolates contained bla _OXA _- ₄₉₈ while bla _OXA _- ₄₆₀ was present in 37/38 cgMLST₂₀₀-650 isolates. The cgMLST₂₀₀-514 group revealed both ant(6)-Ia and sat4 resistance genes in 23/23 and 22/23 isolates, respectively. Also, cgMLST₂₀₀-266 and cgMLST₂₀₀-84 had GyrAT86I mutation with 16/16 (100%) and 14/15 (93.3%), respectively. These findings illustrate how cgMLST and MMD methods can be used to evaluate the relative contribution of known sources of C. coli to the human burden of campylobacteriosis and how cgMLST typing can be used as an indicator of antimicrobial resistance in C. coli.

SUBMITTER: Harrison L

PROVIDER: S-EPMC8313984 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Campylobacter is among the most common causes of gastroenteritis worldwide. Campylobacter jejuni and Campylobacter coli are the most common species causing human disease. DNA sequence-based methods for strain characterization have focused largely on C. jejuni, responsible for 80 to 90% of infections, meaning that C. coli epidemiology has lagged behind. Here, we have analyzed the genome of 450 C. coli isolates to determine genetic markers that can discriminate isolates sampled from 3 major reservoir hosts (chickens, cattle, and pigs). These markers then were applied to identify the source of infection of 147 C. coli strains from French clinical cases. Using STRUCTURE software, 259 potential host-segregating markers were revealed by probabilistic characterization of single-nucleotide polymorphism (SNP) frequency variation in strain collections from three different hosts. These SNPs were found in 41 genes or intergenic regions, mostly coding for proteins involved in motility and membrane functions. Source attribution of clinical isolates based on the differential presence of these markers confirmed chickens as the most common source of C. coli infection in France.IMPORTANCE Genome-wide and source attribution studies based on Campylobacter species have shown their importance for the understanding of foodborne infections. Although the use of multilocus sequence typing based on 7 genes from C. jejuni is a powerful method to structure populations, when applied to C. coli, results have not clearly demonstrated its robustness. Therefore, we aim to provide more accurate data based on the identification of single-nucleotide polymorphisms. Results from this study reveal an important number of host-segregating SNPs, found in proteins involved in motility, membrane functions, or DNA repair systems. These findings offer new, interesting opportunities for further study of C. coli adaptation to its environment. Additionally, the results demonstrate that poultry is potentially the main reservoir of C. coli in France.

Project description:Campylobacter is among the most common worldwide causes of bacterial gastroenteritis. This organism is part of the commensal microbiota of numerous host species, including livestock, and these animals constitute potential sources of human infection. Molecular typing approaches, especially multilocus sequence typing (MLST), have been used to attribute the source of human campylobacteriosis by quantifying the relative abundance of alleles at seven MLST loci among isolates from animal reservoirs and human infection, implicating chicken as a major infection source. The increasing availability of bacterial genomes provides data on allelic variation at loci across the genome, providing the potential to improve the discriminatory power of data for source attribution. Here we present a source attribution approach based on the identification of novel epidemiological markers among a reference pan-genome list of 1,810 genes identified by gene-by-gene comparison of 884 genomes of Campylobacter jejuni isolates from animal reservoirs, the environment, and clinical cases. Fifteen loci involved in metabolic activities, protein modification, signal transduction, and stress response or coding for hypothetical proteins were selected as host-segregating markers and used to attribute the source of 42 French and 281 United Kingdom clinical C. jejuni isolates. Consistent with previous studies of British campylobacteriosis, analyses performed using STRUCTURE software attributed 56.8% of British clinical cases to chicken, emphasizing the importance of this host reservoir as an infection source in the United Kingdom. However, among French clinical isolates, approximately equal proportions of isolates were attributed to chicken and ruminant reservoirs, suggesting possible differences in the relative importance of animal host reservoirs and indicating a benefit for further national-scale attribution modeling to account for differences in production, behavior, and food consumption.IMPORTANCE Accurately quantifying the relative contribution of different host reservoirs to human Campylobacter infection is an ongoing challenge. This study, based on the development of a novel source attribution approach, provides the first results of source attribution in Campylobacter jejuni in France. A systematic analysis using gene-by-gene comparison of 884 genomes of C. jejuni isolates, with a pan-genome list of genes, identified 15 novel epidemiological markers for source attribution. The different proportions of French and United Kingdom clinical isolates attributed to each host reservoir illustrate a potential role for local/national variations in C. jejuni transmission dynamics.

Project description:Although the majority of previous work on campylobacteriosis has centered on the species Campylobacter jejuni, Campylobacter coli, the sister group to C. jejuni, is also a significant problem, but remains a much less studied organism. The purpose of this study was to develop and apply an expanded 16 locus MLST genotyping scheme to a large collection of C. coli isolates sampled from a wide range of host species, and to complete microarray comparative genomic hybridizations for these same strains, in order to: (1) determine whether host specific clones, genotypes, or clonal complexes are evident and (2) evaluate whether there are particular genes comprising the dispensable portion of the C. coli genome that are more commonly associated with certain host species. Genotyping and ClonalFrame analyses of the expanded MLST data suggest that (1) host preferred groups have tended to evolve in the diversification of C. coli, (2) this has happened repeatedly, at different times, throughout the evolutionary history of the species, and (3) recombination has played varying roles in the diversification of the different groups. Concomitant with the information on evolutionary history derived from the MLST data, the microarray data suggests that a combination of common ancestry in some cases and lateral gene transfer in others are behind a tendency for sets of genes to be common to isolates derived from particular hosts. Keywords: comparative genomic hybridization Combimatrix CustomArray™ 4X2K was used in this study. This array is divided into 4 sectors, each of which contains 2,240 in situ synthesized oligonucleotide probes (spots) with the same probe design and layout. Based on the sequence of Campylobacter coli strain RM2228, oligonucleotide probes were designed to have a similar annealing temperature of 56ºC and a length 35-40 bp. Two separate designs were used in this study; both included 100 control probes (20 negative controls with sequences from plant and phage, each with 5 replicate spots) as well as loci from the RM2228 genome. Because of the strict criteria for probe design, not all ORFs could be covered in this analysis. The first design included 1942 of the 1967 protein coding genes described in the unfinished sequence of C. coli strain RM2228. The second-generation design was based on genes that were not clearly present (loci with low intensity or no hybridization for at least one strain) in the hybridization results involving the first design. The second design included additional two or five probes, separated from one another in order to span the entire gene, for these 615 ambiguous loci, synthesized in situ to occupy the 2,240 independent microarray spots. Replicate microarrays were hybridized for every 65 strains tested in this study.

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Core Genome MLST for Source Attribution of Campylobacter coli.

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