Increasing MβL prevalence is worrisome in various European countries; however, intrinsic resistance mechanisms in a highly genetically diverse population of carbapenem-non-susceptible P. aeruginosa are probably a matter for greater concern in these countries.

The epidemiology of Staphylococcus epidermidis in U.S. hospitals remains limited. This study aimed to address the genetic backgrounds of linezolid-susceptible and -resistant S. epidermidis strains (isolated in 2010), including cfr-carrying strains. In addition, the antimicrobial susceptibility profiles and linezolid resistance mechanisms among clonal lineages were assessed. A total of 71 S. epidermidis isolates were selected, and linezolid-resistant strains were screened for cfr and mutations in 23S rRNA, L3, and L4. All isolates were subjected to multilocus sequence typing (MLST), and the results were analyzed by eBURST. Strains showing a G2576 alteration also had M156 (7/7; 100.0%) and/or H146 (6/7; 85.7%) L3 modifications. This study provides an overview of the S. epidermidis clonal distribution and reports higher resistance rates among CC2-I strains. The results show that cfr may be acquired and expressed by both CC2 main subclusters, while 23S rRNA mutations appeared more often within CC2-I strains. Interestingly, these 23S rRNA mutants also had L3 alterations, which may act synergistically or in a compensatory manner to minimize the fitness cost while providing survival advantages under selective pressure. Staphylococcus epidermidis is ubiquitous in the human skin and mucosal microflora and causes infections in immunocompetent patients when the integrity of the skin barrier is disturbed (23). However, the vast majority of S. epidermidis infections among hospitalized patients have been associated with indwelling medical devices, such as intravascular and intrathecal catheter systems, pacemaker electrodes, urinary tract catheters, and other polymer and metal implants, which are used as vehicles for entering the host (33). It has been demonstrated that the organism possesses great capability for genetic recombination and gene acquisition, including resistance determinants (20). In fact, the rates of methicillin (oxacillin) resistance among S. epidermidis strains currently exceed 70% in many institutions worldwide (1, 7). However, although antimicrobial resistance can compromise therapy, treatment failure has been primarily associated with the species' ability to form biofilms on medical devices, which is a common feature of many nosocomial pathogens (29).Strains of S. epidermidis resistant to antimicrobial agents other than oxacillin (␤-lactams) have also been reported. In large surveillance studies, the linezolid resistance rates are still low; however, there seems to be a trend toward increased rates (7). S. epidermidis appears to be prone to accumulate linezolid resistance mechanisms, such as mutations in the 23S rRNA and in the ribosomal proteins L3 and L4 (7,10,13,18,19). In addition to alterations in the target site, a more recent linezolid resistance mechanism, the cfr gene, has been increasingly reported in the literature (6,8,30). cfr encodes a methyltransferase that catalyzes the posttranscriptional methylation of nucleotide A2503 in the 23S rRNA (6). This gene was initially detected in a transferab...

A Staphylococcus aureus surveillance program was initiated in the United States to examine the in vitro activity of ceftaroline and epidemiologic trends. Susceptibility testing by Clinical and Laboratory Standards Institute broth microdilution was performed on 4,210 clinically significant isolates collected in 2009 from 43 medical centers. All isolates were screened for mecA by PCR and evaluated by pulsed-field gel electrophoresis. Methicillin-resistant S. aureus (MRSA) were analyzed for Panton-Valentine leukocidin (PVL) genes and the staphylococcal cassette chromosome mec (SCCmec) type. All isolates had ceftaroline MICs of <2 g/ml with an MIC 50 of 0.5 and an MIC 90 of 1 g/ml. The overall resistance rates, expressed as the percentages of isolates that were intermediate and resistant (or nonsusceptible), were as follows: ceftaroline, 1.0%; clindamycin, 30.2% (17.4% MIC > 4 g/ml; 12.8% inducible); daptomycin, 0.2%; erythromycin, 65.5%; levofloxacin, 39.9%; linezolid, 0.02%; oxacillin, 53.4%; tetracycline, 4.4%; tigecycline, 0%; trimethoprim-sulfamethoxazole, 1.6%; vancomycin, 0%; and high-level mupirocin, 2.2%. The mecA PCR was positive for 53.4% of the isolates. The ceftaroline MIC 90 s were 0.25 g/ml for methicillin-susceptible S. aureus and 1 g/ml for MRSA. Among the 2,247 MRSA isolates, 51% were USA300 (96.9% PVL positive, 99.7% SCCmec type IV) and 17% were USA100 (93.4% SCCmec type II). The resistance rates for the 1,137 USA300 MRSA isolates were as follows: erythromycin, 90.9%; levofloxacin, 49.1%; clindamycin, 7.6% (6.2% MIC > 4 g/ml; 1.4% inducible); tetracycline, 3.3%; trimethoprim-sulfamethoxazole, 0.8%; high-level mupirocin, 2.7%; daptomycin, 0.4%; and ceftaroline and linezolid, 0%. USA300 is the dominant clone causing MRSA infections in the United States. Ceftaroline demonstrated potent in vitro activity against recent S. aureus clinical isolates, including MRSA, daptomycinnonsusceptible, and linezolid-resistant strains.

Human lungs are constantly exposed to bacteria in the environment, yet the prevailing dogma is that healthy lungs are sterile. DNA sequencing-based studies of pulmonary bacterial diversity challenge this notion. However, DNA-based microbial analysis currently fails to distinguish between DNA from live bacteria and that from bacteria that have been killed by lung immune mechanisms, potentially causing overestimation of bacterial abundance and diversity. We investigated whether bacterial DNA recovered from lungs represents live or dead bacteria in bronchoalveolar lavage (BAL) fluid and lung samples in young healthy pigs. Live bacterial DNA was DNase I resistant and became DNase I sensitive upon human antimicrobial-mediated killing in vitro. We determined live and total bacterial DNA loads in porcine BAL fluid and lung tissue by comparing DNase I-treated versus untreated samples. In contrast to the case for BAL fluid, we were unable to culture bacteria from most lung homogenates. Surprisingly, total bacterial DNA was abundant in both BAL fluid and lung homogenates. In BAL fluid, 63% was DNase I sensitive. In 6 out of 11 lung homogenates, all bacterial DNA was DNase I sensitive, suggesting a predominance of dead bacteria; in the remaining homogenates, 94% was DNase I sensitive, and bacterial diversity determined by 16S rRNA gene sequencing was similar in DNase I-treated and untreated samples. Healthy pig lungs are mostly sterile yet contain abundant DNase I-sensitive DNA from inhaled and aspirated bacteria killed by pulmonary host defense mechanisms. This approach and conceptual framework will improve analysis of the lung microbiome in disease.

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