Assessing strain-level variation among type I restriction-modification systems and methylation patterns in Xylella fastidiosa

Authors: O'Leary, Michael; Burbank, Lindsey; Stenger, Drake

Submitted to: American Society for Microbiology Conference
Publication Type: Abstract Only
Publication Acceptance Date: 5/20/2021
Publication Date: 6/20/2021
Citation: O'Leary, M.L., Burbank, L.P., Stenger, D.C. 2021. Assessing strain-level variation among type I restriction-modification systems and methylation patterns in Xylella fastidiosa. American Society for Microbiology Conference.

Interpretive Summary:

Technical Abstract: Xylella fastidiosa is a naturally competent gram-negative bacterial plant pathogen with five genetically distinct subspecies, i.e., fastidiosa, morus, multiplex, pauca, and sandyi. Collectively, strains of X. fastidiosa cause disease on a diverse set of host plants, including blueberry, citrus, grapevine, and olive. Genomes of X. fastidiosa strains are enriched in restriction-modification (R-M) systems, which methylate native DNA and cut unmethylated DNA at specific sequence motifs. Transformation efficiency in X. fastidiosa varies by genetic linage and is improved by inhibition of Type I R-M systems, suggesting differences among R-M system complement between strains and a role for Type I R-M systems in mediating acquisition of foreign DNA. Sequence recognition of Type I R-M systems depends on a specificity subunit, HsdS, which contains two target recognition domains (TRDs) that each recognize a 3-5 bp sequence on either side of a short nonspecific spacer. Recombination between hsdS genes can generate new TRD domain combinations, resulting in hsdS alleles with novel sequence specificities. In other bacterial pathogens where Type I R-M systems provide a barrier to transformation (e.g., Staphylococcus aureus), hsdS allele complement correlates with distinct genetic lineages. To determine if genetic lineages of X. fastidiosa have differing hsdS allele complements, Type I R-M systems were identified in 117 genome assemblies, and hsdS genes were categorized by TRD domains. Each strain contains three conserved Type I R-M systems, while strains of subspecies multiplex and pauca contain a fourth system absent from other subspecies. Three to twenty-two unique hsdS alleles and two to fourteen unique TRD domains were identified per system. In total, at least 30 combined hsdS allele profiles were identified across X. fastidiosa strains. Distribution of hsdS alleles is associated with MLST classification, but does not correlate with geographic origin or host of isolation. Genomic methylation profiles of twelve X. fastidiosa strains were derived using nanopore sequencing. Two to four methylated motifs characteristic of Type I R-M recognition sites (e.g., bipartite) were identified per strain, suggesting some of these systems are active. Variation among bipartite motif sequences was identified within and between X. fastidiosa subspecies and correlates with variation among hsdS alleles, allowing for identified motifs to be tentatively assigned to specific Type I R-M systems and hsdS alleles.