It is not well understood why strains of community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA), a major cause of skin and soft tissue infections, became successful so quickly, overtaking the place of methicillin-sensitive S. aureus (MSSA) in many communities. To evaluate the genetic basis of differences in their virulence traits, 293 S. aureus isolates consisting of three cohorts, genotypically defined clinical CA-MRSA (n ‫؍‬ 77), clinical MSSA (n ‫؍‬ 103), and nasal carriage MSSA (n ‫؍‬ 113), collected over a 19-year period in two Midwestern states in the United States, were (i) extensively genotyped and (ii) screened for 40 known virulence genes which included those for enterotoxins, leukocidins, hemolysins, and surface proteins and several newly identified putative toxin genes from the USA400 lineage of CA-MRSA. Genotypically, nasal carriage and clinical MSSA isolates were much more diverse than was the CA-MRSA group, which was found to be of USA400 lineage only. Virulence gene profiles of the three groups showed that CA-MRSA strains harbored significantly higher percentages (>95%; P value, <0.05) of the sea, sec, sec4, seg2, seh, sek, sel, sel2, ear, ssl1, lpl10, lukSF-PV, lukD, lukE, and clfA genes than did the carriage and the clinical MSSA group (range, 0% to 58%). Genes of the enterotoxin gene cluster, seg, sei, sem, sen, and seo, were present in the clinical and carriage isolates but not in the CA-MRSA group. These results suggest that the presence of additional virulence factors in USA400 CA-MRSA strains compared to the nasal carriage and clinical MSSA strains probably contributed to their enhanced virulence.

Objective: Not much is known about the zoonotic transmission of methicillin-resistant Staphylococcus aureus (MRSA) in companion animals in the United States. We report the rate of prevalence of S. aureus and MRSA recovered from clinical samples of animals requiring treatment at veterinary clinics throughout the upper midwestern and northeastern United States. Design:We compared phenotypes, genotypes, and virulence profiles of the MRSA isolates identified in companion animals, such as cats, dogs, horses, and pigs, with typical human nosocomial and community-associated MRSA (CA-MRSA) genotypes to assess implied zoonotic transmission or zooanthroponosis. Five hundred thirty-three coagulase-positive staphylococci (CPS) isolates recovered between 2006 and 2008 from a variety of animal-source samples were screened for S. aureus by S. aureus-specific 16S rDNA primers and were screened for methicillin-resistance. All MRSA isolates were genotyped by pulsed-field gel electrophoresis (PFGE), multilocus sequence typing (MLST), and spa typing. They were also screened for common staphylococcal enterotoxin and adhesion genes by multiplex and singleplex PCR.Results: Among the 533 CPS isolates recovered, 66 (12.4%) were determined to be S. aureus and 24 (4.5%) were MRSA. The percent of animals that were positive for S. aureus were as follows: 6.6% (32 of 487) dogs, 39.6% (19 of 48) cats, 83.3% (10 of 12) horses, and 100% of pigs, rabbits, hamsters and rats. Notably, 36.4% of all S. aureus identified were MRSA. Methicillin-resistant S. aureus was present in clinical samples from 12 of 487 dogs (2.5%), 6 of 48 cats (12.5%), 5 of 12 horses (42%), and 1 of 2 pigs (50%). The 24 MRSA isolates resolved into 4 PFGE clones: USA100 (50%), USA300 (16.7%), USA500 (20.8%) and USA800 (12.5%) and 6 sequence types (ST5, ST8, ST105, ST830, and ST986) or 2 clonal complexes, CC5 and CC8. Five major virulence profiles (clusters A to E) were observed in these MRSA isolates. Genotypic and virulence profiles of cats and dogs were more similar to each other than to those of horses. A Panton-Valentine leukocidin positive isolate with ST8:USA300 background was identified in a pig causing skin and soft infection. Conclusion:The presence of human MRSA clones in these animals suggests possible reverse zoonotic transmission. This study reports the first case of a USA300 genotype in a pig. Presence of multiple virulence profiles within a MRSA genotype in these animals suggests the potential of emergence of new MRSA clones by gaining or losing additional virulence genes.

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.

Background: Prior investigations have not identified a major locus for vertebral malformations, providing evidence that there is genetic heterogeneity for this condition. WNT3A has recently been identified as a negative regulator of Notch signaling and somitogenesis. Mice with mutations in Wnt3a develop caudal vertebral malformations. Because congenital vertebral malformations represent a sporadic occurrence, linkage approaches to identify genes associated with human vertebral development are not feasible. We hypothesized that WNT3A mutations might account for a subset of congenital vertebral malformations.

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