# Single molecule-level detection and long read-based phasing of epigenetic variations in bacterial methylomes

## TL;DR

This paper introduces SMALR, a new method for detecting and analyzing DNA methylation at the single-molecule level in bacterial genomes, revealing epigenetic diversity.

## Contribution

SMALR provides single-molecule resolution for studying bacterial methylomes, enabling the detection of epigenetic heterogeneity.

## Key findings

- SMALR reveals distinct types of epigenetic heterogeneity in bacterial populations.
- The method offers improved resolution compared to existing SMRT sequencing-based approaches.
- SMALR enables de novo detection of DNA methylation without relying on population-level consensus.

## Abstract

Beyond its role in host defense, bacterial DNA methylation also plays important roles in the regulation of gene expression, virulence and antibiotic resistance. Bacterial cells in a clonal population can generate epigenetic heterogeneity to increase population-level phenotypic plasticity. Single molecule, real-time (SMRT) sequencing enables the detection of N6-methyladenine and N4-methylcytosine, two major types of DNA modifications comprising the bacterial methylome. However, existing SMRT sequencing-based methods for studying bacterial methylomes rely on a population-level consensus that lacks the single-cell resolution required to observe epigenetic heterogeneity. Here, we present SMALR (single-molecule modification analysis of long reads), a novel framework for single molecule-level detection and phasing of DNA methylation. Using seven bacterial strains, we show that SMALR yields significantly improved resolution and reveals distinct types of epigenetic heterogeneity. SMALR is a powerful new tool that enables de novo detection of epigenetic heterogeneity and empowers investigation of its functions in bacterial populations.

Bacterial DNA methylation is involved in many processes, from host defense to antibiotic resistance, however current methods for examining methylated genomes lack single-cell resolution. Here Beaulaurier et al. present Single Molecule Modification Analysis of Long Reads, a new tool for de novo detection of epigenetic heterogeneity.