Optical Mapping Reveals a Large Genetic Inversion between Two Methicillin-Resistant Staphylococcus aureus Strains

Abstract

Staphylococcus aureus is a highly versatile and evolving bacterium of great clinical importance. S. aureus can evolve by acquiring single nucleotide polymorphisms and mobile genetic elements and by recombination events. Identification and location of novel genomic elements in a bacterial genome are not straightforward, unless the whole genome is sequenced. Optical mapping is a new tool that creates a high-resolution, in situ ordered restriction map of a bacterial genome. These maps can be used to determine genomic organization and perform comparative genomics to identify genomic rearrangements, such as insertions, deletions, duplications, and inversions, compared to an in silico (virtual) restriction map of a known genome sequence. Using this technology, we report here the identification, approximate location, and characterization of a genetic inversion of ϳ500 kb of a DNA element between the NRS387 (USA800) and FPR3757 (USA300) strains. The presence of the inversion and location of its junction sites were confirmed by site-specific PCR and sequencing. At both the left and right junction sites in NRS387, an IS1181 element and a 73-bp sequence were identified as inverted repeats, which could explain the possible mechanism of the inversion event.

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

“…In fact, it has been shown that Whole Genome Mapping could be routinely used as an aid to bacterial genome sequence assembly (11) and closure of sequence gaps (31). Using comparative whole-genome mapping, we (24) have also shown the presence of a large genetic inversion event in a methicillin-resistant S. aureus (MRSA). Using Salmonella enterica serovar Typhimurium isolates from an outbreak, Saunders et al (21) have shown that their WGMs distinguished variation in genome maps associated with microbial resistance and its prophage content.…”

“…WGMs of the 47 S. aureus strains were obtained. The reliability, reproducibility, and accuracy of the experimentally derived WGMs were determined using a USA400 (MW2) and a USA300 (FPR3757) strain as previously described by our group (24). Essentially, this was done by comparing the XbaI in silico maps of MW2 and USA300 (FPR3757) to the corresponding experimentally derived WGMs, which were found to be nearly identical.…”

“…Whole-genome maps (WGMs) were created by following the method described previously (24). Briefly, with the use of agarose plugs, high-molecular-weight chromosomal DNA fragments were isolated with minimal shearing.…”

“…Whole-genome mapping (previously referred to as optical mapping by OpGen, Inc.) is a technique that generates high-resolution, ordered restriction maps of whole genomes that can be used to discriminate closely related bacterial strains. The technology has been used to identify macrolevel genomic changes such as inversions, deletions, insertions, and duplications in a bacterial genome (10,12,(20)(21)(22)24). It has also been found useful in closing sequencing gaps and identifying unique sequences in prokaryotes and methylation profiles in eukaryotes (1,4).…”

“…The gene for methicillin resistance, mecA, is carried on a 21-to 67-kb element, the staphylococcal chromosomal cassette mec (SCCmec), which integrates at a conserved location in the S. aureus genome. Hospital-associated methicillin-resistant S. aureus (HA-MRSA), which continues to be a major problem in U.S. hospitals since the 1970s, tends to harbor SCCmec types I, II, and III (3,18,24). The community-associated MRSA (CA-MRSA), which was first reported in the 1990s in community settings, has been recently reported to be the most common type of MRSA in people visiting emergency departments for skin and soft tissue infections (5,7,16).…”