Paraffin embedding and hematoxylin & eosin (H&E) staining were performed as described elsewhere [15]. scalded skin syndromes, endocarditis, osteomyelitis, and sepsis [1]. Frequent resistance to antibiotics considerably complicates treatment ofS. aureusinfections [2]. In particular, many strains ofS. aureusare resistant to methicillin and other beta-lactam antibiotics [3]. These methicillin-resistantS. aureus(MRSA) have become so abundant among nosocomial isolates ofS. aureusthat methicillin is no longer a drug of first choice for the treatment ofS. aureushospital-associated (HA) infections. Although MRSA infections were traditionally limited to hospitals, community-associated cases of MRSA (CA-MRSA) were reported starting in the late 1990s [4]. The epidemiological success of CA-MRSA strains is believed to stem from the combination of antibiotic resistance at low fitness cost [5,6] with extraordinary virulence, allowing these strains to infect otherwise healthy individuals and spread sustainably in the population [7]. Within the last decade, CA-MRSA strains have caused a pandemic of mostly skin and soft tissue infections. A particularly pronounced epidemic is seen in the United States, where CA-MRSA is the most frequent cause of skin and soft tissue infections being reported to emergency departments [8]. Importantly, almost all CA-MRSA infections in the United States are caused by closely related clones belonging to pulsedfield type USA300 [9]. CA-MRSA infections are also being reported at increasing numbers in other parts of the world [4,10,11]. Interestingly, these global CA-MRSA strains do not commonly belong to pulsed-field type USA300 but cover virtually the entire diversity ofS. aureusas a species. However, research on CA-MRSA virulence has been performed largely with strains belonging to the USA300 lineage, whereas our knowledge of virulence of other, globally occurring CA-MRSA, is MK-7145 comparatively limited. Thus, to better understand the virulence of global CA-MRSA, we here analyzed isolates belonging to major CA-MRSA lineages found worldwide and compared them to the most abundant HAMRSA strains. For this purpose, we used a rabbit model of skin infection. We chose this rabbit model because skin infections are the predominant manifestation of CA-MRSA MK-7145 disease [4]. Furthermore, it has been recently demonstrated that rabbit neutrophils have high sensitivity in vitro to the cytolytic activity of Panton-Valentine leukocidin (PVL), a leukocyte toxin epidemiologically correlated with many CA-MRSA infections [12,13]. Thus, assuming that neutrophils also are a primary target of PVL in vivo, a rabbit infection model should allow appropriate analysis of MK-7145 the relative contribution of PVL to CA-MRSA virulence. In addition, to investigate whether in vitro analyses allow predictions on the relative virulence of CA-MRSA strains, we analyzed in vitro expression of virulence factors shown or proposed to be associated with CA-MRSA virulence. == Materials And Methods == Bacterial strains and growth conditions.MRSA isolates SF8300 (pulsed-field type USA300, sequence type [ST] 8), BD02-25 (USA500, ST8), SF1497 (USA1100, ST30), SF1208 (USA200, ST36), SF1681 (USA1000, ST59), and SF2561 (USA100, ST5) were isolated from patients at San Francisco hospitals. Strain MW2 (USA400, ST1) was obtained from the Network on Antimicrobial Resistance inStaphylococcus aureus(NARSA). Strain 07-02662 (ST80) was isolated from a patient in Germany and kindly provided by Dr W. Witte, Robert Koch Institute, Wernigerode, Germany. Strain CN1 (ST72) was isolated from a patient in the Seoul area, South Korea, and kindly provided by Dr J. Kim, Hallym University College of Medicine, Republic of Korea. Bacteria were grown in tryptic soy broth (TSB) (Oxoid) or casein hydrolysate and yeast-extract containing medium (CCY) [14] at 37C, with shaking in an orbital shaker at 180 revolutions per minute (rpm). Rabbit skin abscess model.New Zealand white female rabbits (12 rabbits per strain) were used for the abscess model. All rabbits weighed between 2.0 and 2.3 kg at the time of use. Before the experiments, animals were shaved around the site of injection.S. aureusstrains were grown to mid-exponential growth phase, washed, and resuspended in sterile phosphatebuffered saline (PBS) at 5108colony-forming units (CFUs)/100 L. Rabbits were anesthetized with isoflurane and inoculated with 100 L PBS containing 5108liveS. aureusor with PBS alone in the right dorsum by intradermal injection. Abscess dimensions were measured daily with a caliper for a total of 14 Mmp13 days. Length (L) and width (W) values were used to calculate the area of lesions using the formula L W. A 0.5-mL blood sample was collected daily from the marginal ear vein of each rabbit for quantitative blood culture and measurement of cytokine concentrations, which were performed with rabbit-specific enzyme-linked immunosorbent assay (ELISA) kits (Ever Systems Biology Laboratory, Inc) according to the manufacturer’s instructions. On the fourth day.