Genome-wide methylome analysis of two strains belonging to the hypervirulent Neisseria meningitidis serogroup W ST-11 clonal complex

Abstract

A rising incidence of meningococcal serogroup W disease has been evident in many countries worldwide. Serogroup W isolates belonging to the sequence type (ST)-11 clonal complex have been associated with atypical symptoms and increased case fatality rates. The continued expansion of this clonal complex in the later part of the 2010s has been largely due to a shift from the so-called original UK strain to the 2013 strain. Here we used single-molecule real-time (SMRT) sequencing to determine the methylomes of the two major serogroup W strains belonging to ST-11 clonal complex. Five methylated motifs were identified in this study, and three of the motifs, namely 5'-GATC-3', 5'-GAAGG-3', 5'-GCGCGC-3', were found in all 13 isolates investigated. The results showed no strain-specific motifs or difference in active restriction modification systems between the two strains. Two phase variable methylases were identified and the enrichment or depletion of the methylation motifs generated by these methylases varied between the two strains. Results from this work give further insight into the low diversity of methylomes in highly related strains and encourage further research to decipher the role of regions with under- or overrepresented methylation motifs.

Figure 1

Restriction-modification systems in the 13 isolates belonging to the N. meningitidis 2013 strain marked in blue and original UK strain in orange. Active genes (without inactivating mutations) are coloured: blue = active methylase, red = active restriction endonuclease (REase), purple = active very-short repair endonuclease. Motifs detected by single-molecule real-time sequencing or MspJI cleavage are black, grey = MspJI data not available. The tree on the left shows the phylogeny of the isolates based on single-nucleotide polymorphisms. Isolates from the Jamboree outbreak are marked on the right hand side in a black box, carriage isolates are marked with a star-shaped symbol.

Figure 2

Distribution of methylated bases in two N. meningitidis serogroup W isolates from the 2013 strain (top) and original UK strain (bottom), respectively. The relative positions of predicted methyltransferases are indicated on the outermost track: Type II, blue; Type III, maroon. The remaining coloured tracks display the location of methylated sites for each motif. From outer to inner: 5′-GATC-3′, green; 5′-GAAGG-3′, yellow; 5′-GCGCGC-3′ purple; 5′-ACACC-3′, grey; 5′-CGAAT-3′, pink. Tick marks display the genomic positions.

Figure 3

Phylogeny of the 13 N. meningitidis serogroup W isolates based on single-nucleotide polymorphisms (SNP), isolates belonging to the 2013 strain marked in blue and original UK strain in orange. Detected methylation motifs with single-molecule real-time sequencing are shown as black boxes. Isolates from the Jamboree outbreak are marked in a black box, carrier isolates are marked with a star-shaped symbol.

Figure 4

Venn diagram of the number of genes that had an overrepresentation (red font) and underrepresentation (blue font) of (a) 5′-ACACC-3′ motifs and (b) 5′-CGAAT-3′ motifs in each N. meningitidis strain (2013 strain in blue circles and original UK strain in orange circles). Pie charts show the functions of the genes in which the under and overrepresented motifs were found.

Figure 5

The Clusters of Orthologous Groups of proteins (COG) function classification of genes that were underrepresented (diagonal lines) or overrepresented (filled) of (a) the 5′-ACACC-3′ motif and (b) the 5′-CGAAT-3′ motif in the N. meningitidis 2013 strain and UK original strain. Yellow = metabolism, brown = information and storage processing, blue = cellular process and signalling, green = metabolism/cellular process and signalling.