Identification of antimicrobial resistance genes is important for understanding the underlying mechanisms and the epidemiology of antimicrobial resistance. As the costs of whole-genome sequencing (WGS) continue to decline, it becomes increasingly available in routine diagnostic laboratories and is anticipated to substitute traditional methods for resistance gene identification. Thus, the current challenge is to extract the relevant information from the large amount of generated data. We developed a web-based method, ResFinder that uses BLAST for identification of acquired antimicrobial resistance genes in whole-genome data. As input, the method can use both pre-assembled, complete or partial genomes, and short sequence reads from four different sequencing platforms. The method was evaluated on 1862 GenBank files containing 1411 different resistance genes, as well as on 23 de-novo-sequenced isolates. When testing the 1862 GenBank files, the method identified the resistance genes with an ID = 100% (100% identity) to the genes in ResFinder. Agreement between in silico predictions and phenotypic testing was found when the method was further tested on 23 isolates of five different bacterial species, with available phenotypes. Furthermore, ResFinder was evaluated on WGS chromosomes and plasmids of 30 isolates. Seven of these isolates were annotated to have antimicrobial resistance, and in all cases, annotations were compatible with the ResFinder results. A web server providing a convenient way of identifying acquired antimicrobial resistance genes in completely sequenced isolates was created. ResFinder can be accessed at www.genomicepidemiology.org. ResFinder will continuously be updated as new resistance genes are identified.

Staphylococcus aureus is capable of becoming resistant to all classes of antibiotics clinically available and resistance can develop through de novo mutations in chromosomal genes or through acquisition of horizontally transferred resistance determinants. This review covers the most important antibiotics available for treatment of S. aureus infections and a special emphasis is dedicated to the current knowledge of the wide variety of resistance mechanisms that S. aureus employ to withstand antibiotics. Since resistance development has been inevitable for all currently available antibiotics, new therapies are continuously under development. Besides development of new small molecules affecting cell viability, alternative approaches including anti-virulence and bacteriophage therapeutics are being investigated and may become important tools to combat staphylococcal infections in the future.

Resveratrol is a naturally occurring polyphenolic antioxidant that has received massive attention for its potential health benefits, including anticarcinogenesis, anti-aging and antimicrobial properties. The compound is well tolerated by humans and in recent years has been widely used as a nutraceutical. Its common use makes it interesting to investigate with respect to antimicrobial properties both as a single agent and in combination with conventional antibiotics. Resveratrol displays antimicrobial activity against a surprisingly wide range of bacterial, viral and fungal species. At subinhibitory concentrations, resveratrol can alter bacterial expression of virulence traits leading to reduced toxin production, inhibition of biofilm formation, reduced motility and interference with quorum sensing. In combination with conventional antibiotics, resveratrol enhances the activity of aminoglycosides against Staphylococcus aureus, whereas it antagonises the lethal activity of fluoroquinolones against S. aureus and Escherichia coli. Whilst the antimicrobial properties of the compound have been extensively studied in vitro, little is known about its efficacy in vivo. Nonetheless, following topical application resveratrol has alleviated acne lesions caused by the bacterium Propionibacterium acnes. There are currently no in vivo studies addressing its effect in combination with antibiotics, but recent research suggests that there may be a potential for enhancing the antimicrobial efficacy of certain existing antibiotic classes in combination with resveratrol. Given the difficulties associated with introducing new antimicrobial agents to the market, nutraceuticals such as resveratrol may prove to be interesting candidates when searching for solutions for the growing problem of antimicrobial resistance.

The clinical development of antimicrobial peptides (AMPs) is currently under evaluation to combat the rapid increase in MDR bacterial pathogens. However, many AMPs closely resemble components of the human innate immune system and the ramifications of prolonged bacterial exposure to AMPs are not fully understood. We show that in vitro serial passage of a clinical USA300 MRSA strain in a host-mimicking environment containing host-derived AMPs results in the selection of stable AMP resistance. Serial passage experiments were conducted using steadily increasing concentrations of LL-37, PR-39 or wheat germ histones. WGS and proteomic analysis by MS were used to identify the molecular mechanism associated with increased tolerance of AMPs. AMP-resistant mutants were characterized by measuring in vitro fitness, AMP and antibiotic susceptibility, and virulence in a mouse model of sepsis. AMP-resistant Staphylococcus aureus mutants often displayed little to no fitness cost and caused invasive disease in mice. Further, this phenotype coincided with diminished susceptibility to both clinically prescribed antibiotics and human defence peptides. These findings suggest that therapeutic use of AMPs could select for virulent mutants with cross-resistance to human innate immunity as well as antibiotic therapy. Thus, therapeutic use of AMPs and the implications of cross-resistance need to be carefully monitored and evaluated.

Staphylococcus aureus is intrinsically resistant to polymyxins (polymyxin B and colistin), an important class of cationic antimicrobial peptides used in treatment of Gram-negative bacterial infections. To understand the mechanisms underlying intrinsic polymyxin resistance in S. aureus , we screened the Nebraska Transposon Mutant Library established in S. aureus strain JE2 for increased susceptibility to polymyxin B. Nineteen mutants displayed at least 2-fold reductions in MIC, while the greatest reductions (8-fold) were observed for mutants with inactivation of either graS , graR , vraF , or vraG or the subunits of the ATP synthase ( atpA , atpB , atpG , or atpH ), which during respiration is the main source of energy. Inactivation of atpA also conferred hypersusceptibility to colistin and the aminoglycoside gentamicin, whereas susceptibilities to nisin, gallidermin, bacitracin, vancomycin, ciprofloxacin, linezolid, daptomycin, and oxacillin were unchanged. ATP synthase activity is known to be inhibited by oligomycin A, and the presence of this compound increased polymyxin B-mediated killing of S. aureus . Our results demonstrate that the ATP synthase contributes to intrinsic resistance of S. aureus towards polymyxins and that inhibition of the ATP synthase sensitizes S. aureus to this group of compounds. These findings show that by modulation of bacterial metabolism, new classes of antibiotics may show efficacy against pathogens towards which they were previously considered inapplicable. In light of the need for new treatment options for infections with serious pathogens like S. aureus , this approach may pave the way for novel applications of existing antibiotics. IMPORTANCE Bacterial pathogens that cause disease in humans remain a serious threat to public health, and antibiotics are still our primary weapon in treating bacterial diseases. The ability to eradicate bacterial infections is critically challenged by development of resistance to all clinically available antibiotics. Polymyxins constitute an important class of antibiotics for treatment of infections caused by Gram-negative pathogens, whereas Gram-positive bacteria remain largely insusceptible towards class of antibiotics. Here we performed a whole-genome screen among nonessential genes for polymyxin intrinsic resistance determinants in Staphylococcus aureus . We found that the ATP synthase is important for polymyxin susceptibility and that inhibition of the ATP synthase sensitizes S. aureus towards polymyxins. Our study provides novel insights into the mechanisms that limit polymyxin activity against S. aureus and provides valuable targets for inhibitors to potentially enable the use of polymyxins against S. aureus and other Gram-positive pathogens.