Dataset Information

Comparison of different technologies for the decipherment of the whole genome sequence of Campylobacter jejuni BfR-CA-14430.

ABSTRACT: Background:Campylobacter jejuni is a zoonotic pathogen that infects the human gut through the food chain mainly by consumption of undercooked chicken meat, raw chicken cross-contaminated ready-to-eat food or by raw milk. In the last decades, C. jejuni has increasingly become the most common bacterial cause for food-born infections in high income countries, costing public health systems billions of euros each year. Currently, different whole genome sequencing techniques such as short-read bridge amplification and long-read single molecule real-time sequencing techniques are applied for in-depth analysis of bacterial species, in particular, Illumina MiSeq, PacBio and MinION. Results:In this study, we analyzed a recently isolated C. jejuni strain from chicken meat by short- and long-read data from Illumina, PacBio and MinION sequencing technologies. For comparability, this strain is used in the German PAC-CAMPY research consortium in several studies, including phenotypic analysis of biofilm formation, natural transformation and in vivo colonization models. The complete assembled genome sequence most likely consists of a chromosome of 1,645,980 bp covering 1665 coding sequences as well as a plasmid sequence with 41,772 bp that encodes for 46 genes. Multilocus sequence typing revealed that the strain belongs to the clonal complex CC-21 (ST-44) which is known to be involved in C. jejuni human infections, including outbreaks. Furthermore, we discovered resistance determinants and a point mutation in the DNA gyrase (gyrA) that render the bacterium resistant against ampicillin, tetracycline and (fluoro-)quinolones. Conclusion:The comparison of Illumina MiSeq, PacBio and MinION sequencing and analyses with different assembly tools enabled us to reconstruct a complete chromosome as well as a circular plasmid sequence of the C. jejuni strain BfR-CA-14430. Illumina short-read sequencing in combination with either PacBio or MinION can substantially improve the quality of the complete chromosome and epichromosomal elements on the level of mismatches and insertions/deletions, depending on the assembly program used.

SUBMITTER: Epping L

PROVIDER: S-EPMC6913002 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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Comparison of different technologies for the decipherment of the whole genome sequence of <i>Campylobacter jejuni</i> BfR-CA-14430.

Epping Lennard L Golz Julia C JC Knüver Marie-Theres MT Huber Charlotte C Thürmer Andrea A Wieler Lothar H LH Stingl Kerstin K Semmler Torsten T

Gut pathogens 20191216

<h4>Background</h4><i>Campylobacter jejuni</i> is a zoonotic pathogen that infects the human gut through the food chain mainly by consumption of undercooked chicken meat, raw chicken cross-contaminated ready-to-eat food or by raw milk. In the last decades, <i>C. jejuni</i> has increasingly become the most common bacterial cause for food-born infections in high income countries, costing public health systems billions of euros each year. Currently, different whole genome sequencing techniques such ...[more]

PMID: 31890037

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Project description:Campylobacter species are the most prevalent bacterial pathogen causing acute enteritis worldwide. In contrast to Campylobacter jejuni, about 5 % of Campylobacter coli strains exhibit susceptibility to restriction endonuclease digestion by DpnI cutting specifically 5'-G(m)ATC-3' motifs. This indicates significant differences in DNA methylation between both microbial species. The goal of the study was to analyze the methylome of a C. coli strain susceptible to DpnI digestion, to identify its methylation motifs and restriction modification systems (RM-systems), and compare them to related organisms like C. jejuni and Helicobacter pylori.Using one SMRT cell and the PacBio RS sequencing technology followed by PacBio Modification and Motif Analysis the complete genome of the DpnI susceptible strain C. coli BfR-CA-9557 was sequenced to 500-fold coverage and assembled into a single contig of 1.7 Mbp. The genome contains a CJIE1-like element prophage and is phylogenetically closer to C. coli clade 1 isolates than clade 3. 45,881 6-methylated adenines (ca. 2.7 % of genome positions) that are predominantly arranged in eight different methylation motifs and 1,788 4-methylated cytosines (ca. 0.1 %) have been detected. Only two of these motifs correspond to known restriction modification motifs. Characteristic for this methylome was the very high fraction of methylation of motifs with mostly above 99 %.Only five dominant methylation motifs have been identified in C. jejuni, which have been associated with known RM-systems. C. coli BFR-CA-9557 shares one (RAATTY) of these, but four ORFs could be assigned to putative Type I RM-systems, seven ORFs to Type II RM-systems and three ORFs to Type IV RM-systems. In accordance with DpnI prescreening RM-system IIP, methylation of GATC motifs was detected in C. coli BfR-CA-9557. A homologous IIP RM-system has been described for H. pylori. The remaining methylation motifs are specific for C. coli BfR-CA-9557 and have been neither detected in C. jejuni nor in H. pylori. The results of this study give us new insights into epigenetics of Campylobacteraceae and provide the groundwork to resolve the function of RM-systems in C. coli.

Project description:Whole genome sequencing (WGS) has been used to assess the phylogenetic relationships, virulence and metabolic differences, and the relationship between gene carriage and host or niche differentiation among populations of C. jejuni isolates. We previously characterized the presence and expression of CJIE4 prophage proteins in four C. jejuni isolates using WGS and comparative proteomics analysis, but the isolates were not assessed further. In this study we compare the closed, finished genome sequences of these isolates to the total proteome. Genomes of the four isolates differ in phage content and location, plasmid content, capsular polysaccharide biosynthesis loci, a type VI secretion system, orientation of the ~92 kb invertible element, and allelic differences. Proteins with 99% sequence identity can be differentiated using isobaric tags for relative and absolute quantification (iTRAQ) comparative proteomic methods. GO enrichment analysis and the type of artefacts produced in comparative proteomic analysis depend on whether proteins are encoded in only one isolate or common to all isolates, whether different isolates have different alleles of the proteins analyzed, whether conserved and variable regions are both present in the protein group analyzed, and on how the analysis is done. Several proteins encoded by genes with very high levels of sequence identity in all four isolates exhibited preferentially higher protein expression in only one of the four isolates, suggesting differential regulation among the isolates. It is possible to analyze comparative protein expression in more distantly related isolates in the context of WGS data, though the results are more complex to interpret than when isolates are clonal or very closely related. Comparative proteomic analysis produced log2 fold expression data suggestive of regulatory differences among isolates, indicating that it may be useful as a hypothesis generation exercise to identify regulated proteins and regulatory pathways for more detailed analysis.

Dataset Information

Comparison of different technologies for the decipherment of the whole genome sequence of Campylobacter jejuni BfR-CA-14430.

Publications

Comparison of different technologies for the decipherment of the whole genome sequence of <i>Campylobacter jejuni</i> BfR-CA-14430.

Similar Datasets

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