Molecular epidemiology and phenotypes of invasive methicillin-resistant vancomycin-intermediate Staphylococcus aureus in Taiwan.

BACKGROUND: Patients with invasive infections caused by methicillin-resistant Staphylococcus aureus (MRSA), especially those with an elevated minimal inhibitory concentration (MIC) of vancomycin (VA), are likely to have treatment failure and poor outcomes. The aim of this study was to delineate and correlate the genotypes and phenotypes of clinical VA-intermediate S. aureus (VISA) from invasive infections in Taiwan. METHODS: Between 2006 and 2010, a total of 670 non-duplicate MRSA isolates were collected from patients with invasive infections, mostly from blood, as part of a nationwide antimicrobial surveillance program named Tigecycline in vitro Surveillance in Taiwan. Among them, 10 (1.5%) VISA (VA MIC = 4 mg/L) isolates were identified. Molecular typing with staphylococcal cassette chromosome mec (SCCmec), multilocus sequence typing, staphylococcal protein A (spa), mec-associated hypervariable region (dru), accessory gene regulator (agr), and pulse-field gel electrophoresis, and phenotypic analysis including antibiotic susceptibility testing, gene encoding Panton-Valentine leukocidin (pvl), and superantigenic toxin profiles, were analyzed. RESULTS: All but one isolate was defined as molecular health-care-associated MRSA: 6 as SCCmecIII-ST239-spa t037-agrI-dru7 (1 isolate) and dru14 (5 isolates), 2 as SCCmecII-ST5-spa t586-agrII-dru4, and one as SCCmecII-ST89-spa t3520-agrIII-dru7. One isolate was defined as SCCmecIV-ST59-spa t437-agrI-dru8, which was categorized as molecular community-associated MRSA. Five pulsotypes were identified; only one had a positive D-test and 3 were insusceptible to daptomycin (MIC ≧1 mg/L). Five isolates possessed sea-selk-selq, among them 4 belonged to SCCmecIII-ST239-spa t037-agrI. CONCLUSION: In this study, VISA was rarely isolated from invasive MRSA infections, and most cases harbored limited genotypes and corresponding phenotypes.

The Role of ß-Glycosylated Wall Teichoic Acids in the Reduction of Vancomycin Susceptibility in Vancomycin-Intermediate Staphylococcus aureus.

Staphylococcus aureus is an opportunistic pathogen that causes a wide range of infections. Due to the rapid evolution of antibiotic resistance that leads to treatment failure, it is important to understand the underlying mechanisms. Here, the cell wall structures of several laboratory vancomycin-intermediate S. aureus (VISA) strains were analyzed. Among the VISA strains were S. aureus VC40, which accumulated 79 mutations, including most importantly 2 exchanges in the histidine-kinase VraS, and developed full resistance against vancomycin (MIC, 64 µg/ml); a revertant S. aureus VC40R, which has an additional mutation in vraR (MIC, 4 µg/ml); and S. aureus VraS(VC40), in which the 2 vraS mutations were reconstituted into a susceptible background (MIC, 4 µg/ml). A ultraperformance liquid chromatography (UPLC) analysis showed that S. aureus VC40 had a significantly decreased cross-linking of the peptidoglycan. Both S. aureus VC40 and S. aureus VraS(VC40) displayed reduced autolysis and an altered autolysin profile in a zymogram. Most striking was the significant increase in d-alanine and N-acetyl-d-glucosamine (GlcNAc) substitution of the wall teichoic acids (WTAs) in S. aureus VC40. Nuclear magnetic resonance (NMR) analysis revealed that this strain had mostly ß-glycosylated WTAs in contrast to the other strains, which showed only the α-glycosylation peak. Salt stress induced the incorporation of ß-GlcNAc anomers and drastically increased the vancomycin MIC for S. aureus VC40R. In addition, ß-glycosylated WTAs decreased the binding affinity of AtlA, the major autolysin of S. aureus, to the cell wall, compared with α-glycosylated WTAs. In conclusion, there is a novel connection between wall teichoic acids, autolysis, and vancomycin susceptibility in S. aureus. IMPORTANCE Infections with methicillin-resistant Staphylococcus aureus are commonly treated with vancomycin. This antibiotic inhibits cell wall biosynthesis by binding to the cell wall building block lipid II. We set out to characterize the mechanisms leading to decreased vancomycin susceptibility in a laboratory-generated strain, S. aureus VC40. This strain has an altered cell wall architecture with a thick cell wall with low cross-linking, which provides decoy binding sites for vancomycin. The low cross-linking, necessary for this resistance mechanism, decreases the stability of the cell wall against lytic enzymes, which separate the daughter cells. Protection against these enzymes is provided by another cell wall polymer, the teichoic acids, which contain an unusually high substitution with sugars in the ß-conformation. By experimentally increasing the proportion of ß-N-acetyl-d-glucosamine in a closely related isolate through the induction of salt stress, we could show that the ß-conformation of the sugars plays a vital role in the resistance of S. aureus VC40.

Transition metal-catalyzed synthesis of new 3-substituted coumarin derivatives as antibacterial and cytostatic agents.

Aim: The aim of this study was to synthesize new coumarin-based compounds and evaluate their antibacterial and antitumor potential. Results: Using transition metal-catalyzed reactions, a series of 7-hydroxycoumarin derivatives were synthesized with aliphatic and aryl moiety attached directly at C-3 of the coumarin ring and through the ethynyl or 1,2,3-triazole linker. The 3-substituted coumarin derivative bearing bistrifluoromethylphenyl at the C-4 position of 1,2,3-triazole (33) showed strong and selective antiproliferative activity against cervical carcinoma cells. The 7-hydroxy-4-methylcoumarin with a phenyl ring directly attached to coumarin at C-3 (10) showed good potency against the methicillin-resistant Staphylococcus aureus and vancomycin-resistant strains. Conclusion: The most active coumarin derivatives owe their antiproliferative potential to the 3,5-ditrifluoromethylphenyl substituent (in 33) and antibacterial activity to the aromatic moiety (in 10); their structure can be optimized further for improved effect.

Antimicrobial Peptides with Antibacterial Activity against Vancomycin-Resistant Staphylococcus aureus Strains: Classification, Structures, and Mechanisms of Action.

The emergence of bacteria resistant to conventional antibiotics is of great concern in modern medicine because it renders ineffectiveness of the current empirical antibiotic therapies. Infections caused by vancomycin-resistant Staphylococcus aureus (VRSA) and vancomycin-intermediate S. aureus (VISA) strains represent a serious threat to global health due to their considerable morbidity and mortality rates. Therefore, there is an urgent need of research and development of new antimicrobial alternatives against these bacteria. In this context, the use of antimicrobial peptides (AMPs) is considered a promising alternative therapeutic strategy to control resistant strains. Therefore, a wide number of natural, artificial, and synthetic AMPs have been evaluated against VRSA and VISA strains, with great potential for clinical application. In this regard, we aimed to present a comprehensive and systematic review of research findings on AMPs that have shown antibacterial activity against vancomycin-resistant and vancomycin-intermediate resistant strains and clinical isolates of S. aureus, discussing their classification and origin, physicochemical and structural characteristics, and possible action mechanisms. This is the first review that includes all peptides that have shown antibacterial activity against VRSA and VISA strains exclusively.

A facile method for vancomycin C-terminus functionalization and derivatization through hydrazide.

Over 60-year clinical use of vancomycin led to the emergence of vancomycin-resistant bacteria and threatened our health. To combat vancomycin-resistant strains, numerous vancomycin analogues were developed, such as Telavancin, Oritavancin and Dalbavancin. Extra structures embedded on C-terminus has been proved to be an effective strategy to promote antibacterial activity of vancomycin against vancomycin-resistant strains. Here, we reported a facile strategy, inspired by native chemical ligation, for vancomycin C-terminus functionalization and derivatization. The introduction of C-terminal hydrazide on vancomycin not only provided us an accessible method for C-terminus functionalization through carbonyl azide and thioester, also acted as an efficient site for vancomycin structure modifications. Based on hydrazide-vancomycin, we effectively conjugated cysteine and cysteine containing peptides onto vancomycin C-terminus, and two fluorescent FITC-vancomycin were prepared through Cys-Maleimide conjugation. Meanwhile, we introduced lipophilic structures onto vancomycin C-terminus via the hydrazide moiety. The obtained vancomycin derivatives were evaluated against both Gram-positive and negative bacteria strains.

Genomic Analysis of Heterogeneous Vancomycin-Intermediate Staphylococcus aureus Strains from Different Clonal Lineages in South Korea.

Recent genomic studies of methicillin-resistant Staphylococcus aureus (MRSA) have revealed genetic diversity in the various clonal lineages. Along with clinical concerns of MRSA infection, infection with heterogeneous vancomycin-intermediate S. aureus (hVISA) is closely associated with treatment failure. In this study, we investigated the magnitude of genetic variation and features at the genomic level of hVISA strains isolated in South Korea. Four hVISA strains were analyzed by molecular epidemiology, antimicrobial susceptibility, and whole-genome sequencing methods, and they were compared with the reference VISA and vancomycin-susceptible S. aureus strains in the same clonal lineage. The epidemiologic features of hVISA strains were closely related to the ST5 and ST239 clones. Comparative analysis of the whole genome showed genetic mutations, particularly in two-component systems (TCSs) and transcriptional regulators. Genetic mutations in walK were commonly found in both ST5- (F545L, E378K, T500K) and ST239-related (E424D, T492R) hVISA strains. hVISA strains in the ST5 clonal lineage contained mutations in TCS genes, including the walK, vraR, and agr loci, whereas ST239-related strains harbored different genetic variations in walK, lytR, and saeR. This study suggests that the diverse genetic variation of TCSs and transcriptional regulators are involved in reduced vancomycin susceptibility through different mechanisms in each clonal lineage.

Multidrug-, methicillin-, and vancomycin-resistant Staphylococcus aureus isolated from ready-to-eat meat sandwiches: An ongoing food and public health concern.

Methicillin-resistant S. aureus (MRSA) and their antimicrobial resistance pose exacerbating global health threats and endangering everyone. Thus, the prevalence, molecular characterization of virulence genes, and antimicrobial resistance patterns of strains isolated from 225 beef burger and hot dog sandwiches vended in Mansoura city, Egypt were determined. 83.1% of the sandwiches tested were contaminated with coagulase-positive S. aureus, with a mean count of 4 × 103 CFU/g. Genes encoding mecA, α-hemolysin, staphylococcal enterotoxins, and toxic shock syndrome toxin-1 were detected in 22.6%, 96.3%, 61.1%, and 0% of the strains isolated, respectively. Of the 190 coagulase-positive strains, 43 (22.6%) were confirmed as MRSA. Among them, 4 strains (2.1%) were vancomycin-resistant S. aureus (VRSA) and resistant to all antimicrobials tested. Interestingly, all isolates were resistant to at least one of the antimicrobials tested, with 75.2% being multi-drug resistant (MDR) and an average multiple antimicrobial resistance (MAR) index of 0.503. Not less important, 100%, 96.3%, 90.5%, 79.5%, 73.7%, 62.6%, and 48.9% of isolates were resistant to Kanamycin, Nalidixic acid, Cefotaxime, Sulphamethoxazole-Trimethoprim, Penicillin G, Tetracycline, and Cephalothin, respectively. The potential hazard of MDR-, MRSA-, and VRSA-contaminated sandwiches may be an indication of the presence of what is more dangerous. Hence, strict hygienic measures and good standards of food handler's personal hygiene to prevent transmission of these pathogens to consumers are imperative.

Investigation of heteroresistant vancomycin intermediate Staphylococcus aureus among MRSA isolates.

INTRODUCTION: Heteroresistant vancomycin intermediate Staphylococcus aureus (hVISA) testing is recommended when therapeutic failure is suspected in the clinics. In our research, we aimed to investigate the prevalence of hVISA among methicilline-resistant S. aureus (MRSA) isolates in our university hospital and compared three methods for detection of hVISA. METHODOLOGY: One hundred MRSA clinical isolates were collected in our medical microbiology laboratory between 01.04.2018 and 01.10.2019. For screening of hVISA, we used two screening agar plates and used one commercial medium; brain heart infusion agar (BHI) plates containing 4 µg/mL vancomycin and 16 g/Lt casein (BHIA-VC; Satola's test), BHI agar plates containing 4 µg/mLvancomycin (BHIAV), and commercially obtained vancomycin resistant Enterococci (VRE) agar for detetection of hVISA. Colonies which could grow on plates were counted manually at 24th and 48th hours. RESULTS: Among 100 MRSA isolates, 43 (43%) were found as hVISA using Satola's test. BHIAV and VRE agar screening test results were found 70% and 4%, respectively. Finally, at the step, MIC values of 20 (47%) hVISA isolates reduced to 2 µg/mL after sub culturing for the gradient test. CONCLUSIONS: We found higher rates of hVISA comparing other studies in Turkey. Both VRE agar and BHIAV screening test failed to detect hVISA properly. Meropenem in combination with vancomycin inhibited the growth of 90% hVISA isolates in our study.

Antimicrobial and antibiofilm photodynamic therapy against vancomycin resistant Staphylococcus aureus (VRSA) induced infection in vitro and in vivo.

Biofilm mediated infection caused by multi-drug resistant bacteria are difficult to treat since it protects the microorganisms by host defense system, making them resistant to antibiotics and other antimicrobial agents. Combating such type of nosocomial infection, especially in immunocompromised patients, is an urgent need and foremost challenge faced by clinicians. Therefore, antimicrobial photodynamic therapy (aPDT) has been intensely pursued as an alternative therapy for bacterial infections. aPDT leads to the generation of reactive oxygen species (ROS) that destroy bacterial cells in the presence of a photosensitizer, visible light and oxygen. Here, we elucidated a possibility of its clinical application by reducing the treatment time and exposing curcumin to 20 J/cm2 of blue laser light, which corresponds to only 52 s to counteract vancomycin resistant Staphylococcus aureus (VRSA) both in vitro and in vivo. To understand the mechanism of action, the generation of total reactive oxygen species (ROS) was quantified by 2'-7'-dichlorofluorescein diacetate (DCFH-DA) and the type of phototoxicity was confirmed by fluorescence spectroscopic analysis. The data showed more production of singlet oxygen, indicating type-II phototoxicity. Different anti-biofilm assays (crystal violet and congo red assays) and microscopic studies were performed at sub-MIC concentration of curcumin followed by treatment with laser light against preformed biofilm of VRSA. The result showed significant reduction in the preformed biofilm formation. Finally, its therapeutic potential was validated in skin abrasion wistar rat model. The result showed significant inhibition of bacterial growth. Furthermore, immunomodulatory analysis with rat serum was performed. A significant reduction in expression of proinflammatory cytokines TNF-α and IL-6 were observed. Hence, we conclude that curcumin mediated aPDT with 20 J/cm2 of blue laser treatment (for 52 s) could be used against multi-drug resistant bacterial infections and preformed biofilm formation as a potential therapeutic approach.

Near-Infrared-Controlled Nanoplatform Exploiting Photothermal Promotion of Peroxidase-like and OXD-like Activities for Potent Antibacterial and Anti-biofilm Therapies.

Nanozymes that mimic peroxidase (POD) activity can convert H2O2 into bactericidal free radicals, which is referred to as chemodynamic therapy (CDT). High glutathione (GSH) levels in the infectious tissue severely limit the performance of CDT. Herein, we report a near-infrared-controlled antibacterial nanoplatform that is based on encapsulating tungsten sulfide quantum dots (WS2QDs) and the antibiotic vancomycin in a thermal-sensitive liposome. The system exploits the photothermal sensitivity of the WS2QDs to achieve selective liposome rupture for the targeted drug delivery. We determined that WS2QDs show a strong POD-like activity under physiological conditions and the oxidase-like activity, which can oxidate GSH to further improve the CDT efficacy. Moreover, we found that increased temperature promotes multiple enzyme-mimicking activities of WS2QDs. This platform exerts antibacterial effects against Gram-positive Mu50 (a vancomycin-intermediate Staphylococcus aureus reference strain) and Gram-negative Escherichia coli and disrupts biofilms for improved penetration of therapeutic agents inside biofilms. In vivo studies with mice bearing Mu50-caused skin abscess revealed that this platform confers potent antibacterial activity without obvious toxicity. Accordingly, our work illustrates that the photothermal and nanozyme properties of WS2QDs can be deployed alongside a conventional therapeutic to achieve synergistic chemodynamic/photothermal/pharmaco therapy for powerful antibacterial effects.

The combined antibacterial effects of sodium new houttuyfonate and berberine chloride against growing and persistent methicillin-resistant and vancomycin-intermediate Staphylococcus aureus.

BACKGROUND: Infections caused by drug-resistant Staphylococcus aureus, especially vancomycin-intermediate Staphylococcus aureus (VISA), leave clinicians with limited therapeutic options for treatment. Persister cells is a leading cause of recalcitrant infection and antibiotic treatment failure, and there is no drug in clinical use that specifically targets persister cells currently. Here, we report a promising combination therapy of sodium new houttuyfonate (SNH) and berberine chloride (BBR) which is able to eradicate both growing and persistent drug-resistant Staphylococcus aureus. RESULTS: The susceptibility test showed SNH exhibited anti-MRSA activity with MIC90 at 64 µg/mL, while BBR showed weak anti-MRSA activity with MIC90 at 512 µg/mL. MICs of BBR in combination with 1/2 MIC SNH decreased by 4 to 64 folds compared with MICs of BBR alone. The results of time-killing assays revealed that the combined use of sub-MIC SNH and BBR offered an in vitro synergistic action against growing MRSA (including pathogenic MRSA) and VISA strains. More importantly, the combination of SNH and BBR was able to eradicate VISA Mu50 and pathogenic MRSA persister cells. The synergistic effect is likely related to the interruption of the cell membrane caused by SNH, which is confirmed by scanning electron microscope and membrane potential and permeability analysis. CONCLUSIONS: Our study provide a promising clinical curative strategy for combating drug-resistant S. aureus infections, especially for recalcitrant infections caused by persister cells.

Unveiling the Mechanism of Action of 7α-acetoxy-6ß-hydroxyroyleanone on an MRSA/VISA Strain: Membrane and Cell Wall Interactions.

The number of cases of failure in the treatment of infections associated with resistant bacteria is on the rise, due to the decreasing efficacy of current antibiotics. Notably, 7α-Acetoxy-6ß-hydroxyroyleanone (AHR), a diterpene isolated from different Plectranthus species, showed antibacterial activity, namely against Methicillin-resistant Staphylococcus aureus (MRSA) strains. The high antibacterial activity and low cytotoxicity render this natural compound an interesting alternative against resistant bacteria. The aim of this study is to understand the mechanism of action of AHR on MRSA, using the MRSA/Vancomycin-intermediate S. aureus (VISA) strain CIP 106760, and to study the AHR effect on lipid bilayers and on the cell wall. Although AHR interacted with lipid bilayers, it did not have a significant effect on membrane passive permeability. Alternatively, bacteria treated with this royleanone displayed cell wall disruption, without revealing cell lysis. In conclusion, the results gathered so far point to a yet undescribed mode of action that needs further investigation.

An Update on Staphylococcus aureus NorA Efflux Pump Inhibitors.

BACKGROUND: Methicillin-resistant and vancomycin-resistant Staphylococcus aureus are pathogens causing severe infectious diseases that pose real public health threats problems worldwide. In S. aureus, the most efficient multidrug-resistant system is the NorA efflux pump. For this reason, it is critical to identify efflux pump inhibitors. OBJECTIVE: In this paper, we present an update of the new natural and synthetic compounds that act as modulators of antibiotic resistance through the inhibition of the S. aureus NorA efflux pump. RESULTS: Several classes of compounds capable of restoring the antibiotic activity have been identified against resistant-S. aureus strains, acting as NorA efflux pump inhibitors. The most promising classes of compounds were quinolines, indoles, pyridines, phenols, and sulfur-containing heterocycles. However, the substantial degree structural diversity of these compounds makes it difficult to establish good structure- activity correlations that allow the design of compounds with more promising activities and properties. CONCLUSION: Despite substantial efforts put forth in the search for new antibiotic adjuvants that act as efflux pump inhibitors, and despite several promising results, there are currently no efflux pump inhibitors authorized for human or veterinary use, or in clinical trials. Unfortunately, it appears that infection control strategies have remained the same since the discovery of penicillin, and that most efforts remain focused on discovering new classes of antibiotics, rather than trying to prolong the life of available antibiotics, and simultaneously fighting mechanisms of bacterial resistance.

Opposite effect of vancomycin and D-Cycloserine combination in both vancomycin resistant Staphylococcus aureus and enterococci.

The increasing spread of antibiotic resistant bacteria is a major human health concern. The challenging development of new effective antibiotics has led to focus on seeking synergistic antibiotic combinations. Vancomycin (VAN) is a glycopeptide antibiotic used to treat Staphylococcus aureus and enterococci infections. It is targeting D-Alanyl-D-Alanine dimers during peptidoglycan biosynthesis. D-cycloserine (DCS) is a D-Alanine analogue that targets peptidoglycan biosynthesis by inhibiting D-Alanine:D-Alanine ligase (Ddl). The VAN-DCS combination was found to be synergistic in VAN resistant S. aureus strains lacking van genes cluster. We hypothesize that this combination leads to opposite effects in S. aureus and enterococci strains harboring van genes cluster where VAN resistance is conferred by the synthesis of modified peptidoglycan precursors ending in D-Alanyl-D-Lactate. The calculated Fractional Inhibitory Concentration of VAN-DCS combination in a van- vancomycin-intermediate, VanA type, and VanB type strains were 0.5, 5 and 3, respectively. As a result, VAN-DCS combination leads to synergism in van-lacking strains, and to antagonism in strains harboring van genes cluster. The VAN-DCS antagonism is due to a mechanism that we named van-mediated Ddl inhibition bypass. Our results show that antibiotic combinations can lead to opposite effects depending on the genetic backgrounds.

New thiazolyl-pyrazoline derivatives bearing nitrogen mustard as potential antimicrobial and antiprotozoal agents.

A new series of N-substituted pyrazoline derivatives 6a-g, 7a-g, 8a-g, and 9a-g was synthetized by reaction of hydrazine derivatives and chalcone-thiazole hybrids bearing nitrogen mustard 5a-g. The chalcones 5a-g were obtained by Claisen-Schmidt condensation of thiazole-2-nitrogen mustard 3 and selected acetophenones 4a-g. These new compounds 6/7/8/9a-g were screened for their antifungal activity against Cryptococcus neoformans, with IC50 values of 3.9-7.8 µg/ml for the N-3,5-dichlorophenyl pyrazolines 9e-g. Interestingly, those compounds show low cytotoxic effects toward erythrocytes (RBC). In addition, N-acetyl (6a,b) and N-formyl pyrazolines (7a, 7b, 7c, and 7g) showed inhibitory activity against methicillin-susceptible Staphylococcus aureus, methicillin-resistant S. aureus, and vancomycin-intermediate S. aureus, with the most important minimum inhibitory concentration values ranging from 31.25 to 125 µg/ml. Regarding the antiprotozoal activity, thiazolyl-pyrazolines 9g, 8f, and 7c display high activity against Plasmodium falciparum, Leishmania (V) panamensis, and Trypanosoma cruzi, with EC50 values of 11.80, 6.46, and 4.98 µM, respectively, and with 7c being approximately 2.6-fold more potent than benznidazole with a selectivity index of 1.61 on U-937 human cells, showing promising potential as a novel antitrypanosomal agent.

Vancomycin intermediate resistant Staphylococcus aureus in the nasal cavity of asymptomatic individuals: a potential public health challenge.

BACKGROUND: The potential of transmitting multidrug resistant Staphylococcus aureus from asymptomatic individuals to healthy individuals could constitute a great challenge to antimicrobial therapy. METHODS: The antibiograms of the S. aureus from asymptomatic individuals were determined by disk diffusion and agar dilution assay techniques with different antibiotics and vancomycin. RESULTS: Of the 152 S. aureus isolated, (59)38.8% isolates were multi-drug resistant strains. Streptomycin was the most effective and inhibited (135)88.82% of the isolates while ceftazidime inhibited (24)15.8% of the isolates. While (82)54.0% of the isolates inhibited by cefuroxime had resistant colonies within their inhibition zones (Rc) and ofloxacin inhibited (100)65.8% of the isolates without having resistant colonies within the inhibition zones, ceftazidime inhibited (7)4.6% of the isolates with resistant colonies within the inhibition zones. Subjecting the isolates to vancomycin showed that (27)17.8% were resistant to 2 µg/ml, (43)28.3% were resistant to 4 µg/ml and (27)17.8% of the isolates were simultaneously resistant to both concentrations of vancomycin. Although (100)65.8% of the isolates had MARindex ≥0.2, (52)34.2% of the isolates had MARindex ≤ 0.2 and (65)428% of the isolates were considered multidrug resistant strains. CONCLUSION: The isolation of multi-drug and vancomycin intermediate resistant strains of S. aureus in high percentage, in this study, presents a great threat to clinicians and general populace. The vancomycin intermediate resistant S. aureus (VISA) in asymptomatic individuals could be a critical concern to the therapeutic dilemma to be added to the presence of multi-drug resistance. A more sustainable therapy must be in place to prevent its dissemination or the outbreak of its infection.