Genomic Insights of First erm B-Positive ST338-SCC mec VT/CC59 Taiwan Clone of Community-Associated Methicillin-Resistant Staphylococcus aureus in Poland

Abstract

We report the first Polish representative of community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA), lukS/F-PV-positive, encoding the ermB gene, as a genetic determinant of constitutive resistance to macrolides, lincosamides, and streptogramin B antibiotics, cMLS-B. This is the first detection of the CA-MRSA strain responsible for nosocomial infection in the Warsaw Clinical Hospital. Resistance to β-lactams associates with a composite genetic element, SCCmec cassette type V_T (5C2&5). We assigned the strain to sequence type ST338 (single-locus variant of ST59), clonal complex CC59, spa-type t437, and agr-type I. Genomic-based comparison was designated SO574/12 as an international Taiwan clone, which has been so far described mainly in the Asia-Pacific region. The ermB gene locates on the chromosome within the 14,690 bp mobile element structure, i.e., the MES_PM1-like structure, which also encodes aminoglycoside- and streptothricin-resistance genes. The MES_PM1-like structure is a composite transposon containing Tn551, flanked by direct repeats of IS1216V insertion sequences, which probably originates from Enterococcus. The ermB is preceded by the 273 bp regulatory region that contains the regulatory 84 bp ermBL ORF, encoding the 27 amino acid leader peptides. The latest research suggests that a new leader peptide, ermBL2, also exists in the ermB regulatory region. Therefore, the detailed function of ermBL2 requires further investigations.

Keywords: CA-MRSA; IS1216V insertion sequence; Panton–Valentine leukocidin (PVL); ST338/CC59; Taiwan clone; ermB gene; macrolides lincosamides and streptogramin B (MLS-B); mobile element structure (MES PM1).

Figure 1

Circular genome view (CGV) of SO574/12 MRSA, with the essential genes’ assignment.

Figure 2

Visualization of global large-scale genome alignment according to progressive Mauve algorithm for genomes of CA-MRSA ZY05 and CA-MRSA SO574/12 strains. Each type of locally collinear block (LCB), is marked with a different color. The pair of LCBs contain the high conserved homology regions are assigned with the same stain. The homology regions that were defined within the aligned genomes are connected with the line in the same color. Both the contig 33 in genome of CA-MRSA SO574/12 isolate, which contains ermB gene, and its matching pair LCB in genome of CA-MRSA ZY05 strain are marked on the diagram in light blue.

Figure 3

Small-scale local comparison of three ermB encoding structures and its genetic organization: (A)—ermB carrying LCB from CA-MRSA ZY05; (B)—ermB carrying contig 33 from CA-MRSA SO574/12; and (C)—ermB encoded region from E. faecalis plasmid pEflis48. Strips in the same blue color inform that aligned structures display each other as homologues and occur with a high level of similarity, coverage, and structure organization.

Figure 4

Genetic structure and organization of MES _PM1-like in genome of SO574/12 strain. MES—chromosomal mobile element structure; LCB—locally collinear block; IS1216V—insert on sequence IS1216V; tnp—transposase of Tn551 gene; tnpR—DNA-invertase gene; ermB-AP—ermB-associated protein (unknown function) gene; ermB—23S rRNA (adenine(2058)-N(6))-dimethyltransferase gene; ermBL—leader peptide region of ermB gene; aph(3′)-III—aminoglycoside 3′-phosphotransferase gene; Δsat-4—partly deleted streptothricin acetyltransferase gene; ant(6)-I—aminoglycoside 6-nucleotidyltransferase gene; ubiE—methyltransferase gene, UbiE/COQ5 family; DNA polymerase—DNA polymerase gene, β-like region; HTH-domain—helix-turn-helix domain protein gene; rec-SS—site-specific recombinase gene; Δrec—partly deleted recombinase gene; HP-gene—hypothetical protein gene; DNA topoisomerase III—DNA topoisomerase III; res—resolvase gene.

Figure 5

Multiple sequence alignment of ermB containing genomic regions (partial visualization of aligned sequences) of four strains: (1) MES_PM1-like of CA-MRSA SO574/12 SCCmecVb/ST338/CC59 clinical isolate; (2) MES_PM1-like of CA-MRSA ZY05 SCCmecVb/ST338/CC59 reference Taiwan clone (GenBank accession number: CP045472.1); (3) MES_PM1 of CA-MRSA PM1 SCCmecVb/ST59/CC59 reference clone (GenBank accession number: AB699882.1); and (4) Enterococcus faecalis N48 plasmid pEflis48 (GenBank accession number: MT877066.1). Part (A) shows alignment of 228 bp 5′-end of analyzed regions. In the case of strain number (1), (2), and (3), the nucleotide similarity and coverage were 100%; in the case of strain (4), the sequence was shorter (Δ127 bp), the identity 99.99%, and the coverage of the rest of DNA region was 99.15%, because of one point mutation (substitution). Part (B) shows alignment of internal regions of four analyzed strains to visualize transition mutation site 6165A:41,483G (CDS of aph(3′)-III gene).