Epigenetische DNS-Modifikation von Campylobacter coli

Abstract

Campylobacter species are the most prevalent pathogen causing acute bacterial enteritis worldwide. In contrast to Campylobacter jejuni, about 15% of Campylobacter coli strains exhibit susceptibility to restriction endonuclease digestion by DpnI cutting specifically 5’-GmATC-3’ motifs. This indicates significant differences in DNA methylation between both microbial species. DNA methylation plays an important role in the regulation of bacterial protein expression hence influencing the pathogenicity and survival traits of a bacterial cell. Studies on DNA methylation in Escherichia coli and Salmonella enterica revealed so called epigenetic switch mechanisms enabling the expression of different forms of particular proteins. The goal of the study was to examine the distribution of adenine and cytosine methylation sites in the C. coli genome and further to analyze the methylome of C. coli to identify methylation motifs and restriction modification systems (RM-systems), and compare them to related organisms like C. jejuni ssp. jejuni and Helicobacter pylori. Therefore, 95 C. coli isolates from a wide variety of sources were examined for 5’-GmATC-3’ and 5’-Cmcwgg-3’ methylation motifs by specific restriction enzyme isoschizomer digestion assays. At all 14 isolates were identified showing DNA-adenine methylase (dam) activity and 9 isolates showing DNA cytosine methylase (dcm) activity. The phylogenetic relatedness of all C. coli isolates was examined by MLST-analysis and the isolates were grouped in one of the three Clades described by Sheppard et al.. 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 and assembled into a single contig of 1.7 Mbp. The genome contains a CJIE1-like element prophage and is phylogenetically closer related 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 ten 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. 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.