Comparative Whole-Genome Mapping To Determine Staphylococcus aureus Genome Size, Virulence Motifs, and Clonality

Abstract

cDespite being a clonal pathogen, Staphylococcus aureus continues to acquire virulence and antibiotic-resistant genes located on mobile genetic elements such as genomic islands, prophages, pathogenicity islands, and the staphylococcal chromosomal cassette mec (SCCmec) by horizontal gene transfer from other staphylococci. The potential virulence of a S. aureus strain is often determined by comparing its pulsed-field gel electrophoresis (PFGE) or multilocus sequence typing profiles to that of known epidemic or virulent clones and by PCR of the toxin genes. Whole-genome mapping (formerly optical mapping), which is a highresolution ordered restriction mapping of a bacterial genome, is a relatively new genomic tool that allows comparative analysis across entire bacterial genomes to identify regions of genomic similarities and dissimilarities, including small and large insertions and deletions. We explored whether whole-genome maps (WGMs) of methicillin-resistant S. aureus (MRSA) could be used to predict the presence of methicillin resistance, SCCmec type, and Panton-Valentine leukocidin (PVL)-producing genes on an S. aureus genome. We determined the WGMs of 47 diverse clinical isolates of S. aureus, including well-characterized reference MRSA strains, and annotated the signature restriction pattern in SCCmec types, arginine catabolic mobile element (ACME), and PVL-carrying prophage, PhiSa2 or PhiSa2-like regions on the genome. WGMs of these isolates accurately characterized them as MRSA or methicillin-sensitive S. aureus based on the presence or absence of the SCCmec motif, ACME and the unique signature pattern for the prophage insertion that harbored the PVL genes. Susceptibility to methicillin resistance and the presence of mecA, SCCmec types, and PVL genes were confirmed by PCR. A WGM clustering approach was further able to discriminate isolates within the same PFGE clonal group. These results showed that WGMs could be used not only to genotype S. aureus but also to identify genetic motifs in MRSA that may predict virulence.

Whole-Genome Mapping as a Novel High-Resolution Typing Tool for Legionella pneumophila

“…In addition, the possible effect of mobile genetic elements made us determine that maps with similarities between 95% and 98% may represent the same strain, while maps with a similarities of Ͻ95% are considered different strains. The same cutoff values were previously established for LA-MRSA and were also used in two other studies using WGM for MRSA with the USA300 genotype and Pseudomonas (16,18,23). It therefore seems that these cutoff values can be utilized for WGM in general regardless of the microorganism used.…”

Whole Genome Mapping and Re-Organization of the Nuclear and Mitochondrial Genomes of Babesia microti Isolates

“…Each of these problems can be addressed with additional approaches, including Whole Genome mapping technology (previously known as Optical Mapping) and detailed PCR and hybridization analyses. Whole Genome mapping reveals the architecture of complete genomes and has been used successfully in various areas of biology including comparative genomics, subtyping of human pathogens, epidemiology and forensics [8]–[12]. The technology has also been critical in closing the genome sequence of various human pathogens [8]–[12].…”

“…Whole Genome mapping reveals the architecture of complete genomes and has been used successfully in various areas of biology including comparative genomics, subtyping of human pathogens, epidemiology and forensics [8]–[12]. The technology has also been critical in closing the genome sequence of various human pathogens [8]–[12]. Unlike pulsed-field gel electrophoresis (PFGE) analyses, which can produce only a limited number of restriction fragments ordered by size on a gel, Whole Genome mapping involves digestion of the genomic DNA affixed to a glass substrate and allows precise mapping of contiguous fragments on each chromosome [13].…”

High Resolution Typing by Whole Genome Mapping Enables Discrimination of LA-MRSA (CC398) Strains and Identification of Transmission Events

“…In contrast, in PFGE of S. aureus only 10–15 non-ordered restriction fragments are used for the analysis [14], [15]. Although Shukla et al have previously successfully used WGM to characterize MRSA belonging to the USA300 clade [16], the number of reports in which WGM was used for molecular typing of bacterial pathogens is very limited [16]–[18].…”