The complete methylome of Helicobacter pylori UM032

Abstract

BackgroundThe genome of the human gastric pathogen Helicobacter pylori encodes a large number of DNA methyltransferases (MTases), some of which are shared among many strains, and others of which are unique to a given strain. The MTases have potential roles in the survival of the bacterium. In this study, we sequenced a Malaysian H. pylori clinical strain, designated UM032, by using a combination of PacBio Single Molecule, Real-Time (SMRT) and Illumina MiSeq next generation sequencing platforms, and used the SMRT data to characterize the set of methylated bases (the methylome).ResultsThe N4-methylcytosine and N6-methyladenine modifications detected at single-base resolution using SMRT technology revealed 17 methylated sequence motifs corresponding to one Type I and 16 Type II restriction-modification (R-M) systems. Previously unassigned methylation motifs were now assigned to their respective MTases-coding genes. Furthermore, one gene that appears to be inactive in the H. pylori UM032 genome during normal growth was characterized by cloning.ConclusionConsistent with previously-studied H. pylori strains, we show that strain UM032 contains a relatively large number of R-M systems, including some MTase activities with novel specificities. Additional studies are underway to further elucidating the biological significance of the R-M systems in the physiology and pathogenesis of H. pylori.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-015-1585-2) contains supplementary material, which is available to authorized users.

Comparison of microbial DNA enrichment tools for metagenomic whole genome sequencing

“…While MBD2 has a high affinity for methylated CpG found in vertebrate DNA, it does still bind non-methylated CpG (Fraga et al, 2003). Bacteria and fungi also methylate cytosines at variable rates, depending on the species and conditions, though typically much less so than vertebrates, particularly at CpG motifs (Fang et al, 2012; Antequera et al, 1984; Lee et al, 2015); this low level of CpG methylation could contribute to decreased specificity. Studies from the manufacturer show that the proportions of bacterial DNA from human saliva or whole black molly fish were largely unaffected by MBD2-Fc enrichment, though some differences were measured, in particular a decrease in Neisseria fl avescens reads (Feehery et al, 2013).…”

Methylome Analysis of Two Xanthomonas spp. Using Single-Molecule Real-Time Sequencing

“…SMRT sequencing can identify methylated DNA modifications including 6mA and 4mC in real-time using fluorescent labeled nucleotides and single polymerase molecules (Eid et al, 2009). Using this technique, diverse bacterial methylomes including those of Helicobacter pylori , Lactobacillus spp., Mycobacterium tuberculosis , Escherichia coli , and Campylobacter coli have been determined, and previously unreported DNA methylation motifs have been identified (Lee et al, 2015; Powers et al, 2013; Zautner et al, 2015; Zhang et al, 2015; Zhu et al, 2016). Plant diseases caused by Xanthomonas spp.…”

Toward a genetic tool development pipeline for host-associated bacteria

“…Using DNA from E. coli overexpressing one of the methyltransferases boosted the transformation efficiency by 100-fold over unmodified plasmid, and within 10-fold of that of DNA isolated from the native B. longum strain. Methylome analysis has been conducted on other human gut strains, providing information necessary to begin bypassing restriction-modification systems in those bacteria [19,23,24]. Going forward, methyltransferases identified and characterized through these analyses could be catalogued and made available, allowing others to quickly recreate identified patterns, and transform hitherto non-transformable bacteria.…”