Methicillin-resistant Staphylococcus Aureus Survival Research Articles

Sub-inhibitory concentrations of tetrabromobisphenol A induce the biofilm formation of methicillin-resistant Staphylococcus aureus.

Biofilm formation by methicillin-resistant Staphylococcus aureus (MRSA) on indwelling medical devices complicates the treatment of infection. Tetrabromobisphenol A (TBBPA), a synthetic, lipophilic, halogenated aromatic compound widely used as an additive in plastics and electronic products, has raised environmental concerns due to its potential for bioaccumulation. This study investigated the impact of sub-inhibitory concentrations of TBBPA on MRSA biofilm formation. Crystal violet staining and confocal laser scanning microscopy analysis demonstrated that 1/8 MIC (0.5µg/mL) of TBBPA significantly stimulated MRSA biofilm formation (P < 0.0001). MTT assays indicated that the metabolic activity within the biofilms increased by 15.60-40.85% compared to untreated controls. Dot blot immunoassay, autolysis assay, and extracellular DNA (eDNA) quantification further revealed TBBPA enhanced the production of polysaccharide intercellular adhesin (PIA) and eDNA, which are key biofilm components. Additionally, TBBPA was found to enhance the production of staphyloxanthin, facilitating MRSA survival under oxidative conditions and in human whole blood. RT-qPCR analysis showed that TBBPA significantly upregulated genes associated with biofilm formation (icaA, atlA, sarA), staphyloxanthin biosynthesis (crtM and sigB), and oxidative stress responses (sodA and katA). These findings suggest that TBBPA promotes MRSA biofilm development and enhances bacterial resistance to adverse conditions, thereby potentially exacerbating risks to human health.

Inhibition of Shikimate Kinase from Methicillin-Resistant Staphylococcus aureus by Benzimidazole Derivatives. Kinetic, Computational, Toxicological, and Biological Activity Studies.

Antimicrobial resistance (AMR) is one of the biggest threats in modern times. It was estimated that in 2019, 1.27 million deaths occurred around the globe due to AMR. Methicillin-resistant Staphylococcus aureus (MRSA) strains, a pathogen considered of high priority by the World Health Organization, have proven to be resistant to most of the actual antimicrobial treatments. Therefore, new treatments are required to be able to manage this increasing threat. Under this perspective, an important metabolic pathway for MRSA survival, and absent in mammals, is the shikimate pathway, which is involved in the biosynthesis of chorismate, an intermediate for the synthesis of aromatic amino acids, folates, and ubiquinone. Therefore, the enzymes of this route have been considered good targets to design novel antibiotics. The fifth step of the route is performed by shikimate kinase (SK). In this study, an in-house chemical library of 170 benzimidazole derivatives was screened against MRSA shikimate kinase (SaSK). This effort led to the identification of the first SaSK inhibitors, and the two inhibitors with the greatest inhibition activity (C1 and C2) were characterized. Kinetic studies showed that both compounds were competitive inhibitors with respect to ATP and non-competitive for shikimate. Structural analysis through molecular docking and molecular dynamics simulations indicated that both inhibitors interacted with ARG113, an important residue involved in ATP binding, and formed stable complexes during the simulation period. Biological activity evaluation showed that both compounds were able to inhibit the growth of a MRSA strain. Mitochondrial assays showed that both compounds modify the activity of electron transport chain complexes. Finally, ADMETox predictions suggested that, in general, C1 and C2 can be considered as potential drug candidates. Therefore, the benzimidazole derivatives reported here are the first SaSK inhibitors, representing a promising scaffold and a guide to design new drugs against MRSA.

Antimicrobial activity and mechanism of anti-MRSA of phloroglucinol derivatives.

In previous studies, authors have completed the total synthesis of several phloroglucinol natural products and synthesized a series of their derivatives, which were tested with good biological activities. To discover anti-MRSA lead compound and study their mechanism of action. Phloroglucinol derivatives were tested to investigate their activities against several gram-positive strains including Methicillin-resistant Staphylococcus aureus (MRSA). The mechanism study was conducted by determining extracellular potassium ion concentration, intracellular NADPH oxidase content, SOD activity, ROS amount in MRSA and MRSA survival rate under A5 treatment. The in vitro cytotoxicity test of A5 was conducted. The activity of monocyclic compounds was stronger than that of bicyclic compounds, and compound A5 showed the best MIC value of 0.98μg/mL and MBC value of 1.95μg/mL, which were 4-8 times lower than that of vancomycin. The mechanism study of A5 showed that it achieved anti-MRSA effect through membrane damage, which is proved by increased concentration of extracellular potassium ion after A5 treatment. Another possible mechanism is the over ROS production induced cell death, which is suggested by observed alternation of several reactive oxygen species (ROS) related indicators including NADPH concentration, superoxide dismutase (SOD) activity, ROS content and bacterial survival rate after A5 treatment. The cytotoxicity results in vitro showed that A5 was basically non-toxic to cells. Acylphloroglucinol derivative A5 showed good anti-MRSA activity, possibly via membrane damage and ROS-mediated oxidative stress mechanism. It deserves further exploration to be a potential lead for the development of new anti-MRSA agent.

Loratadine Combats Methicillin-Resistant Staphylococcus aureus by Modulating Virulence, Antibiotic Resistance, and Biofilm Genes.

Methicillin-resistant Staphylococcus aureus (MRSA) has evolved to become resistant to multiple classes of antibiotics. New antibiotics are costly to develop and deploy, and they have a limited effective lifespan. Antibiotic adjuvants are molecules that potentiate existing antibiotics through nontoxic mechanisms. We previously reported that loratadine, the active ingredient in Claritin, potentiates multiple cell-wall active antibiotics in vitro and disrupts biofilm formation through a hypothesized inhibition of the master regulatory kinase Stk1. Loratadine and oxacillin combined repressed the expression of key antibiotic resistance genes in the bla and mec operons. We hypothesized that additional genes involved in antibiotic resistance, biofilm formation, and other cellular pathways would be modulated when looking transcriptome-wide. To test this, we used RNA-seq to quantify transcript levels and found significant effects in gene expression, including genes controlling virulence, antibiotic resistance, metabolism, transcription (core RNA polymerase subunits and sigma factors), and translation (a plethora of genes encoding ribosomal proteins and elongation factor Tu). We further demonstrated the impacts of these transcriptional effects by investigating loratadine treatment on intracellular ATP levels, persister formation, and biofilm formation and morphology. Loratadine minimized biofilm formation in vitro and enhanced the survival of infected Caenorhabditis elegans. These pleiotropic effects and their demonstrated outcomes on MRSA virulence and survival phenotypes position loratadine as an attractive anti-infective against MRSA.

Gold nanoparticle conjugation enhances berberine's antibacterial activity against methicillin-resistant Staphylococcus aureus (MRSA)

Nanoparticle (NP) conjugation with various biomolecules is one of the most promising approaches for targeting Methicillin-resistant Staphylococcus aureus (MRSA). In this study, berberine (BER) was conjugated with gold nanoparticles (AuNPs) to enhance its antibacterial activity against MRSA. Chemically synthesized AuNPs were characterized by UV–vis spectroscopy, size distribution and Field Emission-Scanning Electron Microscope (FE-SEM) analysis. Berberine was conjugated with AuNPs and the conjugants were characterized using UV–vis spectroscopy and Fourier Transform Infrared (FTIR). The cytotoxicity of free and conjugated BER was also investigated. Comparative studies were conducted based on the Minimum Inhibitory Concentration (MIC) and anti-biofilm activities of conjugants and free BER against MRSA isolates. To verify cell membrane disruption and intracellular imbalance following treatment exposure, reactive oxygen species (ROS) and live-dead staining experiments were performed. In vivo antibacterial efficacy of treated groups was also assessed in a BALB/c mouse-infected skin model. DLS measurement, FE-SEM, and UV–vis spectroscopy confirmed the synthesis of AuNPs with a narrow size distribution of 49.38 nm and a zeta potential of −31.9 mV. The results from UV–vis spectroscopy and FTIR provided support for the functionalization of AuNPs by BER functional groups. The In vitro antibacterial results demonstrated that the conjugated BER exhibited a lower MIC value against MRSA (109.5 μg/ml) compared to free BER (165 μg/ml). Free and conjugated BER, at their MIC concentrations, demonstrated anti-biofilm activity, resulting in biofilm eradication of 13.9 and 22.33 %, respectively. The highest level of ROS production (93 %) was associated with the conjugated BER at a concentration of 27.37 μg/ml. This finding indicates a disruption in cell membrane integrity and a reduction in bacterial viability, as demonstrated by ROS and live/dead staining assays. The cytotoxicity study on the mouse L929 fibroblast cell line revealed approximately 100 % cell viability when exposed to free or conjugated BER at their MIC concentration. This result indicates the biosafety of both of the compounds. The in vivo study in the infected skin model groups treated with conjugated and free BER revealed MRSA survival rate of 2.7 % and 26 %, respectively. These findings suggest that conjugated BER could be an effective nanoformulation candidate with a potential role in managing MRSA associated infections.

Ayanin, a natural flavonoid inhibitor of Caseinolytic protease, is a promising therapeutic agent to combat methicillin-resistant Staphylococcus aureus infections

Antimicrobial resistance (AMR) is a global health threat. The dramatic increase of Methicillin-resistant Staphylococcus aureus (MRSA) infections emphasizes the need to find new anti-infective agents with a novel mode of action. The Caseinolytic protease (ClpP) is a central virulence factor in stress survival, virulence, and antibiotic resistance of MRSA. Here, we found ayanin, a flavonoid isolated from Callicarpa nudiflora, was an inhibitor of MRSA ClpP with an IC50 of 19.63 μM. Using quantitative real-time PCR, ayanin reduced the virulence of Staphylococcus aureus (S. aureus) by down-regulating the level of some important virulence factors, including agrA, RNAⅢ, hla, pvl, psmα and spa. The results of cellular thermal shift assay and thermal shift assay revealed a binding between ayanin and ClpP. Molecular docking showed that ASP-168, ASN-173 and ARG-171 were the potential binding sites for ClpP binding to ayanin. ClpP mutagenesis study further indicated that ARG-171 and ASN-173 were the main active sites of ClpP. The affinity constant (KD) value of ayanin with ClpP was 3.15 × 10−5 M measured by surface plasmon resonance. In addition, ayanin exhibited a significant therapeutic effect on pneumonia infection induced by S. aureus in mice in vivo, especially in combination with vancomycin. This is the first report of ayanin with in vivo and in vitro efficacy against S. aureus infection. In conclusion, ayanin is a promising therapeutic agent to combat MRSA infections by targeting ClpP.

Spinach protein-capped silver nanoparticles with low hemolytic activity inhibit methicillin-resistant Staphylococcus aureus biofilm formation as a new therapeutic approach

BackgroundThe methicillin-resistant Staphylococcus aureus (MRSA) was designated as a high-priority pathogen by the World Health Organization. Cardiovascular and thoracic surgery complications frequently involve MRSA infection. Treatment of newly emerging resistant S. aureus strains is clinically challenging due to the natural tendency of MRSA survival through biofilm formation. The failure of traditional antibiotic therapy necessitates development of next-generation therapeutic molecules. ObjectivesSilver nanoparticles were investigated to treat MRSA in order to overcome the challenges of antibiotic therapy. MethodsSpinach protein functionalized silver nanoparticles (SPAgNPs) were synthesized by reducing with sodium borohydride. The resazurin microtiter assay was used to determine the minimum inhibitory concentration (MIC) of SPAgNPs against MRSA. The antibiofilm activity was investigated using a crystal violet assay at sub-MIC levels and validated using scanning electron microscopy. To demonstrate the mechanism of action of SPAgNPs, the virulence factors involved in MRSA biofilm formation were examined. SEM was used to evaluate the hemocompatibility of human red blood cells. Results and conclusionThe synthesized SPAgNPs showed characteristic yellow colour with surface Plasmon resonance maximum at 418 nm. Powder X-ray diffraction and transmission electron microscopy analysis confirmed that the particles are crystalline with a size range from 14 to 40 nm. SPAgNPs have antibacterial activity at a minimum inhibitory concentration (MIC) of 1.25 mM, killing MRSA via disrupting membrane integrity and producing excess reactive oxygen species. Strikingly, SPAgNPs inhibit MRSA biofilm formation at sub-MIC (0.625 mM). Scanning electron microscopy study reveals that SPAgNPs trigger biofilm inhibition without causing cell damage. Hence the NPs are very effective in preventing pathogens from triggering stress responses. SPAgNPs suppress exopolysaccharide production, cell surface hydrophobicity, and staphyloxanthin biosynthesis. Testing SPAgNPs against human red blood cells reveals that NPs do not harm human cells, suggesting that the SPAgNPs are biocompatible. The findings show that SPAgNPs could be explored as next-generation materials for treating MDR pathogenic infection linked with biofilms.

Defining the Roles of Pyruvate Oxidation, TCA Cycle, and Mannitol Metabolism in Methicillin-Resistant Staphylococcus aureus Catheter-Associated Urinary Tract Infection.

Methicillin-resistant Staphylococcus aureus (MRSA) is an important cause of complicated urinary tract infection (UTI) associated with the use of indwelling urinary catheters. Previous reports have revealed host and pathogen effectors critical for MRSA uropathogenesis. Here, we sought to determine the significance of specific metabolic pathways during MRSA UTI. First, we identified four mutants from the Nebraska transposon mutant library in the MRSA JE2 background that grew normally in rich medium but displayed significantly reduced growth in pooled human urine (HU). This prompted us to transduce the uropathogenic MRSA 1369 strain with the transposon mutants in sucD and fumC (tricarboxylic acid [TCA] cycle), mtlD (mannitol metabolism), and lpdA (pyruvate oxidation). Notably, sucD, fumC, and mtlD were also significantly upregulated in the MRSA 1369 strain upon exposure to HU. Compared to the WT, the MRSA 1369 lpdA mutant was significantly defective for (i) growth in HU, and (ii) colonization of the urinary tract and dissemination to the kidneys and the spleen in the mouse model of catheter-associated UTI (CAUTI), which may be attributed to its increased membrane hydrophobicity and higher susceptibility to killing by human blood. In contrast to their counterparts in the JE2 background, the sucD, fumC, and mtlD mutants in the MRSA 1369 background grew normally in HU; however, they displayed significant fitness defects in the CAUTI mouse model. Overall, identification of novel metabolic pathways important for the urinary fitness and survival of MRSA can be used for the development of novel therapeutics. IMPORTANCE While Staphylococcus aureus has historically not been considered a uropathogen, S. aureus urinary tract infection (UTI) is clinically significant in certain patient populations, including those with chronic indwelling urinary catheters. Moreover, most S. aureus strains causing catheter-associated UTI (CAUTI) are methicillin-resistant S. aureus (MRSA). MRSA is difficult to treat due to limited treatment options and the potential to deteriorate into life-threatening bacteremia, urosepsis, and shock. In this study, we found that pathways involved in pyruvate oxidation, TCA cycle, and mannitol metabolism are important for MRSA fitness and survival in the urinary tract. Improved understanding of the metabolic needs of MRSA in the urinary tract may help us develop novel inhibitors of MRSA metabolism that can be used to treat MRSA-CAUTI more effectively.

Study of SarA by DNA Affinity Capture Assay (DACA) Employing Three Promoters of Key Virulence and Resistance Genes in Methicillin-Resistant Staphylococcus aureus.

Methicillin-resistant Staphylococcus aureus (MRSA), one of the most well-known human pathogens, houses many virulence factors and regulatory proteins that confer resistance to diverse antibiotics. Although they have been investigated intensively, the correlations among virulence factors, regulatory proteins and antibiotic resistance are still elusive. We aimed to identify the most significant global MRSA regulator by concurrently analyzing protein-binding and several promoters under same conditions and at the same time point. DNA affinity capture assay (DACA) was performed with the promoters of mecA, sarA, and sarR, all of which significantly impact survival of MRSA. Here, we show that SarA protein binds to all three promoters. Consistent with the previous reports, ΔsarA mutant exhibited weakened antibiotic resistance to oxacillin and reduced biofilm formation. Additionally, production and activity of many virulence factors such as phenol-soluble modulins (PSM), α-hemolysin, motility, staphyloxanthin, and other related proteins were decreased. Comparing the sequence of SarA with that of clinical strains of various lineages showed that all sequences were highly conserved, in contrast to that observed for AgrA, another major regulator of virulence and resistance in MRSA. We have demonstrated that SarA regulates antibiotic resistance and the expression of various virulence factors. Our results warrant that SarA could be a leading target for developing therapeutic agents against MRSA infections.

Sensitizing Staphylococcus aureus to antibacterial agents by decoding and blocking the lipid flippase MprF.

The pandemic of antibiotic resistance represents a major human health threat demanding new antimicrobial strategies. Multiple peptide resistance factor (MprF) is the synthase and flippase of the phospholipid lysyl-phosphatidylglycerol that increases virulence and resistance of methicillin-resistant Staphylococcus aureus (MRSA) and other pathogens to cationic host defense peptides and antibiotics. With the aim to design MprF inhibitors that could sensitize MRSA to antimicrobial agents and support the clearance of staphylococcal infections with minimal selection pressure, we developed MprF-targeting monoclonal antibodies, which bound and blocked the MprF flippase subunit. Antibody M-C7.1 targeted a specific loop in the flippase domain that proved to be exposed at both sides of the bacterial membrane, thereby enhancing the mechanistic understanding of bacterial lipid translocation. M-C7.1 rendered MRSA susceptible to host antimicrobial peptides and antibiotics such as daptomycin, and it impaired MRSA survival in human phagocytes. Thus, MprF inhibitors are recommended for new antivirulence approaches against MRSA and other bacterial pathogens.

Bacterial Membrane-Derived Vesicles Attenuate Vancomycin Activity against Methicillin-Resistant Staphylococcus aureus.

Methicillin-resistant Staphylococcus aureus (MRSA) has evolved numerous antimicrobial resistance mechanisms and is identified as a serious public health threat by the World Health Organization and U.S. Centers for Disease Control and Prevention. The glycopeptide vancomycin (VAN) remains a cornerstone of therapy for severe MRSA infections despite increasing reports of therapeutic failure in hospitalized patients with bacteremia or pneumonia. Recently, the role of released bacterial-derived membrane vesicles (MVs) in antibiotic resistance has garnered attention. Here we examined the effect of exogenous MRSA-derived MVs on VAN activity against MRSA in vitro, using minimum inhibitory concentration and checkerboard assays, and ex vivo, incorporating components of host innate immunity such as neutrophils and serum complement present in blood. Additionally, the proteome of MVs from VAN-exposed MRSA was characterized to determine if protein expression was altered. The presence of MVs increased the VAN MIC against MRSA to values where clinical failure is commonly observed. Furthermore, the presence of MVs increased survival of MRSA pre-treated with sub-MIC concentrations of VAN in whole blood and upon exposure to human neutrophils but not human serum. Unbiased proteomic analysis also showed an elevated expression of MV proteins associated with antibiotic resistance (e.g., marR) or proteins that are functionally linked to cell membrane/wall metabolism. Together, our findings indicate MRSA-derived MVs are capable of lowering susceptibility of the pathogen to VAN, whole-blood- and neutrophil-mediated killing, a new pharmacodynamic consideration for a drug increasingly linked to clinical treatment failures.

Human Urine Alters Methicillin-Resistant Staphylococcus aureus Virulence and Transcriptome.

Gram-positive methicillin-resistant Staphylococcus aureus (MRSA) is an emerging cause of hospital-associated urinary tract infections (UTI), especially in catheterized individuals. Despite being rare, MRSA UTI are prone to potentially life-threatening exacerbations such as bacteremia that can be refractory to routine antibiotic therapy. To delineate the molecular mechanisms governing MRSA urinary pathogenesis, we exposed three S. aureus clinical isolates, including two MRSA strains, to human urine for 2 h and analyzed virulence characteristics and changes in gene expression. The in vitro virulence assays showed that human urine rapidly alters adherence to human bladder epithelial cells and fibronectin, hemolysis of sheep red blood cells (RBCs), and surface hydrophobicity in a staphylococcal strain-specific manner. In addition, transcriptome sequencing (RNA-Seq) analysis of uropathogenic strain MRSA-1369 revealed that 2-h-long exposure to human urine alters MRSA transcriptome by modifying expression of genes encoding enzymes catalyzing metabolic pathways, virulence factors, and transcriptional regulators. In summary, our results provide important insights into how human urine specifically and rapidly alters MRSA physiology and facilitates MRSA survival in the nutrient-limiting and hostile urinary microenvironment. IMPORTANCE Methicillin-resistant Staphylococcus aureus (MRSA) is an uncommon cause of urinary tract infections (UTI) in the general population. However, it is important to understand MRSA pathophysiology in the urinary tract because isolation of MRSA in urine samples often precedes potentially life-threatening MRSA bacteremia. In this report, we describe how exposure to human urine alters MRSA global gene expression and virulence. We hypothesize that these alterations may aid MRSA in acclimating to the nutrient-limiting, immunologically hostile conditions within the urinary tract leading to MRSA UTI.

Potential Application of Silver Nanoparticles Encapsulated in Branched Amphiphilic Peptide Capsules to Circumvent Antibiotic Resistance in Methicillin-Resistant Staphylococcus Aureus: A Research Protocol

Introduction: Antibiotic resistance is a major global health concern as many fatalities occur each year due to severe infections associated with it, such as sepsis. This research protocol hopes to address the issue of antibiotic resistance in methicillin-resistant Staphylococcus aureus (MRSA) by using an alternative antibiotic therapy. Silver nanoparticles (AgNPs), which have antibiotic properties and low antibiotic resistance potential, will be administered in conjunction with stable and biocompatible branched amphiphilic peptide capsules (BAPCs). Surface functionalized ligands will be attached to the peptide capsules to target iron receptors essential to MRSA survival. By targeting its iron receptors, the bacteria will be unable to mutate in such a way that would attenuate its uptake of this antibiotic. Methods: Testing will begin in vitro on MRSA cultures to assess the minimum inhibitory concentration of the various AgNP treatment groups. Constraints determining the minimum inhibitory concentration include a minimum 3-hour delay in bactericidal effect from application, and a 95% eradication efficacy. Drug efficacy will be evaluated via ex vivo and in vivo mice experiments, with off-target toxicity effects measured via BAPC targeted immunohistochemistry, and both serological and organ-based tests. Repeated in vivo treatments will be performed against a control to measure the relative antibiotic resistance advantages this treatment proposes. As this protocol requires the use of mice, ethics approval from the Canadian Council on Animal Care will be obtained prior to experimentation. Results: It is expected that the AgNPs will be successfully encapsulated within the structurally-sound BAPCs and that the AgNPs will eradicate a significant amount of the MRSA present in the human body with minimal side effects. The in vitro, ex vivo and in vivo tests are likely to yield results that demonstrate that the most effective protocol in eradicating MRSA is to use AgNPs in BAPCs in conjunction with a vancomycin treatment. The proposal will likely satisfy the criteria for appropriate AgNP doses, which includes bactericidal effect in a time frame of hours, over 95% efficacy, and statistically significant reproducibility. Discussion: Successful BAPC mediated delivery of AgNPs prove the potential for targeted antibiotic application against multidrug bacteria with lower risk of antibiotic resistance compared to conventional antibiotics. This proof of concept protocol demonstrates the potential for applying rational design of BAPCs as a delivery vector. Certain limitations, including off-target toxicity and redundant targeting are issues to be cognizant of and controlled. Conclusion: This novel therapy can be considered as a preliminary step in overcoming antibiotic resistance. With appropriate modifications to the functional ligands attached to the BAPCs, the proposed drug mechanism could be applied to treat other bacterial strains.

Indoor Finish Material Influence on Contamination, Transmission, and Eradication of Methicillin-Resistant Staphylococcus aureus (MRSA).

Objective:The purpose of this study was to evaluate environmental surface materials used in healthcare environments for material composition, methicillin-resistant Staphylococcus aureus (MRSA) viability, and a comparison of two disinfectants, a bleach germicidal cleaner and Decon7, a novel disinfectant.Background:Contaminated environmental surfaces have been associated with outbreaks of healthcare-associated illness (HAIs). One in every 20 patients in U.S. acute care hospitals acquire a healthcare-associated illness, leading to consequences such as elevated morbidity, mortality, and a decrease in quality of life. In the patient environment, MRSA can remain viable from hours to up to 14 days.Methods:Environmental surface materials were evaluated as new and worn. Material composition and properties were assessed to evaluate surface integrity and the influence on the disinfection of MRSA. Inoculated materials were used to assess MRSA viability over time and the efficacy of a manufacturer’s recommended cleaning and disinfection product compared to a novel disinfectant.Results:Environmental surface materials respond differently in appearance and roughness, when mechanically worn. When measuring MRSA survival, at 24 hr, MRSA colony forming unit (CFU) counts were reduced on the copper sheet surface and solid surface with cupric oxide. By 72 hr, all MRSA counts were zero. Bleach and the novel disinfectant were equally effective at disinfecting MRSA from all surface types.Conclusions:This study highlights a gap in knowledge about the impact of type and wear of environmental surface materials used in healthcare environments on contamination with epidemiologically important organisms. In conclusion, environmental surface material wear, properties, and cleaning and disinfection efficacy are important factors to consider when addressing HAIs.

Quebrachitol inhibits biofilm formation and virulence production against methicillin-resistant Staphylococcus aureus

The present study evaluated the quebrachitol (QBC) antibiofilm and antivirulence potential against methicillin-resistant Staphylococcus aureus (MRSA). QBC inhibited MRSA biofilm formation at concentration dependent manner without affecting the bacterial growth. Then, QBC biofilm efficacy was confirmed with light and confocal laser scanning microscopy analysis. QBC treatment significantly inhibited the biofilm formation on stainless steel, titanium and silicone surfaces. Besides, QBC treatment significantly reduced the MRSA virulence productions such as lipase and hemolysis. Moreover, it reduced MRSA survival rate in the presence of hydrogen peroxide. QBC treatment inhibited the MRSA adherence on hydrophobic, hydrophilic, collagen coating and fibrinogen coating surfaces. As well as it significantly reduced the autolysin and bacterial aggregation progress. The real-time PCR analysis revealed the ability of QBC downregulated the virulence genes expression including global regulator sarA, agr and polysaccharide intracellular adhesion (PIA) encode ica. The cumulative results of the present study suggest that QBC as a potential agent to combat against MRSA pathogenesis.

Survival of Methicillin-Resistant Staphylococcus aureus in Fish and Shrimp under Different Storage Conditions

Foods that are extensively handled during preparation and stored without refrigeration are often associated with staphylococcal food poisoning. This problem is more confounding when contaminating strains belong to the methicillin-resistant Staphylococcus aureus (MRSA) group. In this study, we investigated the survivability of MRSA in two seafood matrices under different storage conditions. MRSA was inoculated at 6 and 3 log CFU/g into all sample groups of peeled shrimp (Parapeneopsis stylifera) stored at -20°C, Bombay duck fish (Harpadon nehereus) stored in ice, and dried Bombay duck fish stored at 30 ± 2°C. The populations of MRSA in frozen peeled shrimp inoculated with MRSA at 6 log CFU/g were reduced by 1.52 log CFU/g, whereas in samples inoculated with 3 log CFU/g levels remained stable after 60 days of storage. In fresh Bombay duck fish inoculated with 6 log CFU/g and stored in ice for 18 days, MRSA levels decreased by 2.75 log CFU/g. In contrast, in fresh fish inoculated with 3 log CFU/g the total viable count increased by 3.02 log CFU/g over 16 days of ice storage. In dried fish stored at 30 ± 2°C, MRSA levels declined by 3.27 log CFU/g in samples inoculated with 6 log CFU/g and by 0.91 log CFU/g in samples inoculated with 3 log CFU/g. These results suggest that the survival of MRSA depends on the temperature of storage and the inoculum level. In our study, MRSA survival was higher when inoculated at 3 log CFU/g regardless of the seafood matrix and storage temperature.

Photodynamic inactivation of methicillin-resistant Staphylococcus aureus on skin using a porphyrinic formulation.

Staphylococcus aureus is responsible for skin and soft tissue infections. Having in mind increased antibiotic resistance, in this study the efficacy of antimicrobial photodynamic therapy (aPDT) with a porphyrinic formulation (FORM) as photosensitizer (PS) to photoinactivate methicillin-resistant Staphylococcus aureus (MRSA) on skin was evaluated. Potassium iodide (KI) and iodopovidone (PVP-I) were also tested in combination with FORM as potentiator agents of FORM efficacy. The aPDT protocol was first developed in Phosphate Buffered Saline (PBS, in vitro). Porcine skin was artificially contaminated with MRSA (ex vivo) and treated with FORM, FORM + KI or FORM + PVP-I under white light. The in vitro results showed that FORM was effective to inactivate MRSA. A substantial reduction in the irradiation time, when compared to FORM alone, was observed for FORM + KI and FORM + PVP-I combinations. On skin, reductions in MRSA survival of 3.1 Log10 colony forming units (CFU) mL-1 were observed with FORM at 50 μM. Although the combined action of FORM + KI and FORM + PVP-I potentiated the aPDT efficacy in vitro, this was not observed ex vivo. Overall, the results showed that aPDT using FORM, even without coadjutants, is a promising approach for MRSA inactivation on skin.

The Fate of Methicillin-Resistant Staphylococcus aureus in a Synthetic Turf System.

Methicillin-resistant Staphylococcus aureus (MRSA) is responsible for one-third of infectious outbreaks reported among competitive athletes at high school and collegiate levels of competition, while the frequency of infections among National Football League players is nearly 400-fold higher than in the general population. The increased prevalence of MRSA infections among such athletes necessitates the study of football-specific environments as MRSA reservoirs. The recovery and viability of MRSA will differ between turf infill and turf fibers. Descriptive laboratory study. Level 4. Each of 6 infill types and 3 turf fiber types were inoculated with MRSA, sacrificed over time, and enumerated after bacteria extraction and dilution plating to determine the incubation time at which 50% of inoculated MRSA are recoverable and viable (here termed A50). The role of infill toxicity on MRSA survival was assessed using a dialysis assay. On average, MRSA was available for up to 96 hours on infill (average A50: 13 hours) and 24 hours on turf fibers (A50: 4 hours) (P < 0.001). The A50 for each infill type was also variable among infill groups (P < 0.001), averaging 2 hours (ethylene propylene diene monomer [EPDM] rubber), 7 hours (cork-based material), 9 hours (polymer-coated materials), 12 hours (crumb rubber), 13 hours (thermoplastic elastomer [TPE] rubber), and 27 hours (sands). MRSA remained available on slit-film and nylon fiber types for 12 hours postdeposition and for 24 hours on monofilament fibers. Toxicity assays showed that over 90% of MRSA cells remained viable after 6 hours of exposure to cork infill and sands, while 79%, 71%, 68%, and 17% of MRSA remained viable after exposure to polymer-coated materials, crumb rubber, TPE rubber, and EPDM rubber, respectively. Our data also indicated that organic and sand infills exhibited minimal toxicity to MRSA, while high relative toxicity drives limited MRSA availability in EPDM rubber infill. MRSA recovery and viability varied among infill types but not among differing turf blades. The results of this study can inform appropriate athlete and field management practices.

Caspase‐11 counteracts mitochondrial ROS‐mediated clearance of Staphylococcus aureus in macrophages

Methicillin‐resistant Staphylococcus aureus (MRSA) is a growing health concern due to increasing resistance to antibiotics. As a facultative intracellular pathogen, MRSA is capable of persisting within professional phagocytes including macrophages. Here, we identify a role for CASP11 in facilitating MRSA survival within murine macrophages. We show that MRSA actively prevents the recruitment of mitochondria to the vicinity of the vacuoles they reside in to avoid intracellular demise. This process requires CASP11 since its deficiency allows increased association of MRSA‐containing vacuoles with mitochondria. The induction of mitochondrial superoxide by antimycin A (Ant A) improves MRSA eradication in casp11−/− cells, where mitochondria remain in the vicinity of the bacterium. In WT macrophages, Ant A does not affect MRSA persistence. When mitochondrial dissociation is prevented by the actin depolymerizing agent cytochalasin D, Ant A effectively reduces MRSA numbers. Moreover, the absence of CASP11 leads to reduced cleavage of CASP1, IL‐1β, and CASP7, as well as to reduced production of CXCL1/KC. Our study provides a new role for CASP11 in promoting the persistence of Gram‐positive bacteria.

Longistylin A, a natural stilbene isolated from the leaves of Cajanus cajan, exhibits significant anti-MRSA activity

Longistylin A (LLA) is an abundant stilbene isolated from the leaves of Cajanus cajan (L.) Millsp. However, the antibacterial effect of LLA is not yet understood. Therefore, in this study, a detailed investigation of the antibacterial effect of LLA, particularly against methicillin-resistant Staphylococcus aureus (MRSA), was conducted. In vitro, LLA exhibited strong antibacterial activity against MRSA with a minimum inhibitory concentration (MIC) of 1.56 µg/mL and displayed much more rapid bactericidal activity (3-log decrease in MRSA survival within 8 h) than vancomycin. A membrane-targeting experiment suggested that the antibacterial activity of LLA is associated with perturbation of the bacterial membrane potential and increased membrane permeability. Notably, LLA had relatively weak cytotoxicity to murine macrophages [50% cytotoxic concentration (CC50)=8.61 ± 0.57 µg/mL]. In vivo, topical treatment of a skin injury with LLA improved wound healing and closure in an MRSA-infected wound healing mouse model. After 3 days treatment, LLA decreased MRSA bacterial counts in the wounded region, reduced the accumulation of immune cells at the injury site, and alleviated induction of the inflammatory cytokines tumour necrosis factor-alpha (180.74 ± 10.78 pg/mL vs. 606.57 ± 68.99 pg/mL) and interleukin-6 (87.25 ± 10.19 pg/mL vs. 280.58 ± 42.27 pg/mL) in serum.