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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.

Vancomycin-resistant Staphylococcus aureus (VRSA), Vancomycin-intermediate S. aureus (VISA) and heterogeneous VISA (hVISA) are subject to vancomycin treatment failure. The aim of the present study was to determine their precise prevalence and investigate prevalence variability depending on different years and locations. Several international databases including Medline (PubMed), Embase and Web of Sciences were searched (data from 1997 to 2019) to identify studies that addressed the prevalence of VRSA, VISA and hVISA among human clinical isolates around the world. Subgroup analyses and meta-regression were conducted to indicate potential source of variation. Publication bias was assessed using Egger’s test. Statistical analyses were conducted using STATA software (version 14.0). Data analysis showed that VRSA, VISA and hVISA isolates were reported in 23, 50 and 82 studies, with an overall prevalence of 1.5% among 5855 S. aureus isolates, 1.7% among 22,277 strains and 4.6% among 47,721 strains, respectively. The overall prevalence of VRSA, VISA, and hVISA before 2010 was 1.2%, 1.2%, and 4%, respectively, while their prevalence after this year has reached 2.4%, 4.3%, and 5.3%. The results of this study showed that the frequency of VRSA, VISA and hVISA after 2010 represent a 2.0, 3.6 and 1.3-fold increase over prior years. In a subgroup analysis of different strain origins, the highest frequency of VRSA (3.6%) and hVISA (5.2%) was encountered in the USA while VISA (2.1%) was more prevalent in Asia. Meta-regression analysis showed significant increasing of VISA prevalence in recent years (p value ≤ 0.05). Based on the results of case reports (which were not included in the calculations mentioned above), the numbers of VRSA, VISA and hVISA isolates were 12, 24 and 14, respectively, among different continents. Since the prevalence of VRSA, VISA and hVISA has been increasing in recent years (especially in the Asian and American continents), rigorous monitoring of vancomycin treatment, it’s the therapeutic response and the definition of appropriate control guidelines depending on geographical regions is highly recommended and essential to prevent the further spread of vancomycin-resistant S. aureus.

A versatile method for the synthesis of new oxindoles was developed by the reaction between substituted isatins and 5-aminopyrazoles. The reaction was carried out at room temperature in ethanol using p-toluenesulfonic acid as the catalyst. The products were obtained with acceptable to excellent yields (44–96%). Structures of the new compounds were unambiguously established by spectroscopic and analytical techniques. The antibacterial activity was determined by microdilution assays. Compounds 3b, 3e, and 3g showed antistaphylococcal activity, particularly compound 3e displayed a potent activity against the vancomycin intermediate Staphylococcus aureus (VISA). Compounds 3i, 3j, and 3o inhibited Neisseria gonorrhoeae growth.

Objectives: MRSA isolates with vancomycin MIC of 1-2 μg/ml have been linked with treatment failure and heteroresistant VISA phenotype. This study was aimed at comparing two screening methods i.e. GRD Etest and Vancomycin Screen agar in detection of heteroresistance. Material & Methods: The present study was carried out on 41 Methicillin Resistant Staphylococcus aureus strains isolated from different clinical specimens collected from Lahore General Hospital, Lahore. After screening for methicillin resistance, vancomycin MIC was determined by standard E test. Isolates with a vancomycin MIC of 1-2 μg/ml were screened for heteroresistance by Glycopeptide Resistance Detection (GRD) E-test and Vancomycin screen agar. Data was entered and analyzed by using SPSS version 20.0. Study Design: Comparative Study. Setting: Pathology Department of Post Graduate Medical Institute, Lahore. Period: May 2014 to May 2015. Results: When compared with E test GRD, Vancomycin screen agar (V3) showed 100% sensitivity with a 95% CI 39.76% to 100% and the specificity was 65 % with a 95 % CI 47.46% to 79.79%. Its PPV was 23% and NPV was 100% with an overall diagnostic accuracy of 68%. When compared with E test GRD, Vancomycin screen agar (V4) showed a sensitivity of 75% with a 95% CI 19.41% to 99.37% and a specificity of 86.47% with a 95% CI 71.91 to 95.59%. Its PPV was 37.5% and NPV predictive value was 96.96% with an overall diagnostic accuracy of 85.36%. Conclusion: In developing countries like Pakistan, where E tests are costly and difficult to use in routine laboratories, a screening test, which does not miss heteroresistant VISA may be of clinical use.

The emergence of drug-resistant pathogens and the abuse of antibiotics have posed dominant threats to society. In this study, we propose an antibacterial use of bovine serum albumin-capped gold nanoclusters (BSA-AuNCs) to address the issue. BSA-AuNCs have a great antibacterial activity against MRSA and VISA which is proved by time-killing curves (TKC). The possible antibacterial mechanisms of BSA-AuNCs against MRSA and VISA are confirmed by fluorescence image observation.

Introduction Heteroresistant vancomycin-intermediate Staphylococcus aureus (hVISA) bacteremia may result in clinical failure of vancomycin therapy, together with prolonged infection and hospitalization. This clinical problem has resulted in a search for more effective treatment options. The current study was designed to further investigate the synergistic effect of oxacillin plus vancomycin against methicillin-resistant S. aureus (MRSA) and hVISA using checkerboard and time-kill assays. Methods Non-duplicate S. aureus isolates including hVISA (n = 29), MRSA (n = 10) and methicillin susceptible S. aureus (MSSA, n = 11) were used for combinational testing using checkerboard and time-kill assays. Results Twenty-one isolates, 15 hVISA and 6 MRSA, showed synergy between oxacillin and vancomycin by checkerboard assay with fractional inhibitory concentration indices of ≤ 0.5. The addition of oxacillin to vancomycin resulted in a reduction in baseline vancomycin MIC from 1–2 to 0.06–0.5 µg/ml against MRSA and hVISA isolates. In the time-kill assay, the combination of oxacillin and vancomycin resulted in synergistic activity against hVISA (n = 23) and MRSA (n = 7) isolates. Regrowth was observed in six hVISA isolates exposed to combination in the time-kill assay, but none of them reached the original inoculum density at 24 h. All re-growth isolates showed a onefold increase in vancomycin MIC (from 1 to 2 µg/ml) and were re-confirmed as hVISA using the population-analysis profile experiment. Overall, for hVISA and MRSA, the combination of oxacillin plus vancomycin had greater antibacterial effect than each individual drug alone. Conclusion The present study showed the potential activity of vancomycin plus oxacillin combination against hVISA and MRSA isolates. Further, continued evaluation of this combination is warranted and may have therapeutic benefits in treating complicated MRSA infections.

Prolonged vancomycin usage may cause methicillin-resistant Staphylococcus aureus to become vancomycin-intermediate S. aureus (VISA) and heterogeneous VISA (hVISA). Mechanisms of vancomycin resistance of VISA and hVISA are still unclear. In this study, analyses of nucleotide sequence variations in 30 vancomycin-sensitive S. aureus (VSSA), 41 hVISA and 16 VISA isolates revealed 29 single-nucleotide variations in 12 genes (fmtC, graR, graS, htrA, mecA, pbp2, pbp4, srtA, tcaA, upps, vicK and vraR) that are related to cell wall synthesis or the two-component system. Six of these 29 single-nucleotide variations were novel and resulted in the following amino acid changes: Q692E in FmtC; T278I, P306L and I311T in HtrA; and I63V and K101E in Upps. Since P306L and I311T in HtrA and I63V in Upps were present in the majority (76.7%-86.7%) of VSSA isolates, these three amino acid variations may not be associated with vancomycin resistance. The other three amino acid variations (T278I in HtrA, K101E in Upps and Q692E in FmtC) were present in the majority (87.5%-93.8%) of hVISA and VISA isolates, but only in a small number (22.9%-25.7%) of VSSA isolates, suggesting that they are associated with vancomycin resistance.

The importance of heterogeneous vancomycin-intermediate Staphylococcus aureus (hVISA) as a preceding stage for the development of vancomycin-resistant S. aureus is growing. We analyzed the prevalence of hVISA among bacteremia with methicillin-resistant S. aureus (MRSA) using two Etests and evaluated the clinical characteristics and outcomes. Ninety-eight MRSA isolates from blood were collected at two University hospitals in Korea. Macrodilution Etest and glycopeptide resistance detection Etests were used for detection of hVISA. Staphylococcal cassette chromosome mec (SCCmec) typing was performed by multiplex PCR. Clinical data were collected retrospectively from patient medical records. About 30% of MRSA strains were identified as hVISA. Diabetes mellitus was associated (P = 0.047) with hVISA infections. The hVISA isolates were associated with high teicoplanin MIC and multidrug resistance (P = 0.001). SCCmec type II accounted for the majority (79.3%) of hVISA strains. The prevalence of hVISA strains was increased and can lead to the development of multidrug-resistant strains. Patients with diabetes were found to have a greater risk for infection with hVISA strains. As the impact of hVISA on clinical outcome is not yet clear, large-scale studies about clinical outcomes and optimal detection methods of hVISA are needed. In conclusion, hVISA strains have a high prevalence in bloodstream MRSA infections. Awareness of the increase in hVISA strains should motivate laboratories to establish a system to detect and monitor hVISA.

Staphylococcus aureus strains resistant to the last line antibiotic, vancomycin, have been of clinical concern. These include heterogeneous vancomycin-intermediate S. aureus (hVISA) and VISA. The hVISA phenotype cannot be detected by routine laboratory methods. Characterization of hVISA/VISA by new technologies is necessary to differentiate them rapidly from the vancomycin-susceptible isolates (VSSA). In this study, we developed a model for discrimination of hVISA from VSSA by using attenuated total reflection-Fourier transform infrared (ATR-FTIR) spectroscopy combined with multivariate data analysis, displaying a phenotypic signature of the bacteria. ATR-FTIR spectra were acquired from a total of 59 clinical methicillin-resistant S. aureus (MRSA) isolates comprising 28 hVISA and 31 VSSA strains. Principal component analysis (PCA) and partial least square discriminant analysis (PLS-DA) were used to analyze 351 spectra of 39 isolates and develop a discrimination model for identifying hVISA and VSSA. The classification model, which was used for blind testing of 90 spectra from each of 10 hVISA, and 10 VSSA isolates, provided 100% sensitivity and specificity. The modeling revealed that the major discrimination between hVISA and VSSA phenotypes involved bands related to cell wall content (1087 and 1057 cm−1). This study showed that ATR-FTIR technique may be an alternative method for rapid detection of low-level vancomycin-resistant S. aureus.

The aim of the present study was to describe the characteristics of infections with Staphylococcus aureus with reduced vancomycin susceptibility (SARVS) including vancomycin-intermediate S. aureus (VISA) in South Korea, using data from the national sentinel surveillance system during 2014–2016. A total of 66 patients infected or colonized with SA-RVS were reported using the sentinel surveillance system. Among them, VISA was confirmed in 14 isolates (21.2%) and no vancomycin-resistant S. aureus (VRSA) was detected. Most of patients had any kind of indwelling devices (81.8%, 54/66) and underwent surgical procedures in the previous 6 months (84.8%, 56/66). Patients who admitted to an intensive care unit (ICU) in the previous 3 months were 68.2% (45/66). Furthermore, patients who used vancomycin or had MRSA in the previous 1 month were 54.5% (36/66) and 59.1% (39/66), respectively. Upon review of the medical records, 54.5% (36/66) of patients were classified as having SA-RVSassociated infection and 30-day mortality was 19.4% (7/36). Our findings revealed that there was no VRSA in South Korea. SA-RVS including VISA existed particularly in patients who had indwelling devices, history of surgical procedure, and history of ICU admission.

Objective To determine the prevalence and some genetic characteristics of clinical isolates of Staphylococcus aureus (S. aureus) with reduced susceptibility to vancomycin. Methods A total of 414 isolates of S. aureus were collected from clinical specimens from hospitals in Tehran. Vancomycin-intermediate S. aureus (VISA) was determined by brain heart infusion agar containing 4 μg/mL vancomycin screening plate and confirmed via E-test. VISA isolates were analysed for vanA, vanB, mecA, staphylococcal cassette chromosome mec types, surface protein A (Spa) types and agr specific groups. Results Brain heart infusion agar containing 4 μg/mL vancomycin screening tests revealed that 17.14% (n = 71) of S. aureus isolates were VISA phenotype. Ten of the 71 isolates were confirmed by E-test method (minimal inhibitory concentration was 4 to 8 μg/mL). All VISA isolates were susceptible to linezolid and 6 isolates (60%) were methicillin-resistant S. aureus. Five isolates belonged to agr Group II, 4 belonged to agr Group I and 1 belonged to agr Group III. Spa type t030, and staphylococcal cassette chromosome mec Type III were dominant among VISA isolates. Conclusions This study provides further evidence of the global dissemination of VISA isolates and emphasizes to vancomycin susceptibility testing prior to antibiotic therapy.

Vancomycin is the standard of care for the treatment of invasive methicillin-resistant Staphylococcus aureus (MRSA) infections. Infections with vancomycin-nonsusceptible MRSA, including vancomycin-intermediate S. aureus (VISA) and heterogeneous VISA (hVISA), are clinically challenging and are associated with poor patient outcomes. The identification of VISA in the clinical laboratory depends on standard susceptibility testing, which takes at least 24 h to complete after isolate subculture, whereas hVISA is not routinely detected in clinical labs. We therefore sought to determine whether VISA and hVISA can be differentiated from vancomycin-susceptible S. aureus (VSSA) using the spectra produced by matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS). Strains of MRSA were characterized for vancomycin susceptibility phenotype by broth microdilution and modified population analysis. We tested 21 VISA, 21 hVISA, and 38 VSSA isolates by MALDI-TOF MS. Susceptibility phenotypes were separated by using a support vector machine (SVM) machine learning algorithm. The resulting model was validated by leave-one-out cross validation. Models were developed and validated by using spectral profiles generated under various subculture conditions, as well as with and without hVISA strains. Using SVM, we correctly identified 100% of the VISA and 97% of the VSSA isolates with an overall classification accuracy of 98%. Addition of hVISA to the model resulted in 76% hVISA identification, 100% VISA identification, and 89% VSSA identification, for an overall classification accuracy of 89%. We conclude that VISA/hVISA and VSSA isolates are separable by MALDI-TOF MS with SVM analysis.

Invasive heteroresistant vancomycin-intermediate Staphylococcus aureus (h-VISA) isolates were identified and characterized in 10 Korean hospitals from July 2009 to June 2011. The prevalence of h-VISA infections was 3.3% (42/1,289). Most (41/42) were health care-associated infections caused by strains belonging to sequence type 5. Cases of persistent bacteremia were frequent (17/42), and 30-day mortality was high (16/40).

A 76-year-old Japanese man was admitted to hospital for treatment of fever and skin lesion at the implantation site of his pacemaker. During his hospitalization, vancomycin-intermediate Staphylococcus aureus (MIC 4 μg/mL) with reduced susceptibility to daptomycin was isolated from venous blood. This isolate was identified as methicillin-resistant S. aureus with SCCmec IV and was genotyped as sequence type 81, coa VIIa and spa type t7044, harbouring blaZ, aac(6′)-aph(2″) and enterotoxin(-like) genes sea, seb, sek, sel, selx and selw. The patient was successfully treated with daptomycin, minocycline and sulfamethoxazole/trimethoprim. We describe the identification of sequence type 81/SCCmec IV vancomycin-intermediate S. aureus from pacemaker-associated septicaemia.

Introduction: Vancomycin remains the drug of choice to treat infections caused by Methicillin Resistant Staphylococcus aureus (MRSA). Frequent therapeutic failures arise due to the emergence of strains with reduced susceptibility to vancomycin and difficulty in detecting these isolates. Therefore, improvements in screening and confirmatory tests to detect these isolates are required. Aim: To detect heteroresistant Vancomycin Intermediate Staphylococcus aureus (hVISA) isolates among Methicillin Resistant Staphylococcus aureus (MRSA) and Methicillin Sensitive Staphylococcus aureus (MSSA) obtained from various clinical samples and to study the phenotypic and genotypic charecteristics of these strains. Materials and Methods: This study was carried out in the Department of Microbiology, Chettinad Hospital and Research Institute, Kelambakkam, Chennai, Tamil Nadu, India. A total of 468 non-duplicate S.aureus strains obtained from various clinical samples during a period of two years from September 2014 to November 2016 were included in the study. MRSA isolates and hVISA/VISA isolates were detected by screening and confirmatory methods (MRSA screen by cefoxitin disc, hVISA/VISA screen by brain heart infusion agar with 6 µg vancomycin and hVISA/VISA confirmation by PAP-AUC method). Agr polymorphism in hVISA/VISA was analysed by duplex PCR. Accessory gene regulator (Agr) dysfunction in these strains was studied by their colony spreading property in soft agar and autolytic property by Triton X-100 induced lysis. The results were analysed using IBM SPSS (version 21.0) software. Results: Out of 468 S.aureus strains, 114 (24%) strains were detected as MRSA and 33 (7.0%) were hVISA/VISA strains. Among MRSA detected, 11 (9.5%) strains were confirmed as hVISA/VISA. All hVISA/VISA isolates showed reduction in autolytic activity. Colony spreading property was appreciated in 13 (39%) of these isolates. Out of the 33 hVISA/VISA strains, majority belonged to agr I, III and II and the least number of isolates belonged to agr IV. Conclusion: It was concluded that hVISA/VISA strains have diverse virulence properties and are detected in both MRSA and MSSA strains. Therefore, surveillance of these strains among S.aureus isolates having vancomycin MIC’s ≤1.5 µg/mL in laboratory settings is needed.

Heterogeneous vancomycin-intermediate Staphylococcus aureus (hVISA) is an emerging superbug with implicit drug resistance to vancomycin. Detecting hVISA can guide the correct administration of antibiotics. However, hVISA cannot be detected in most clinical microbiology laboratories because the required diagnostic tools are either expensive, time consuming, or labor intensive. By contrast, matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) is a cost-effective and rapid tool that has potential for providing antibiotics resistance information. To analyze complex MALDI-TOF mass spectra, machine learning (ML) algorithms can be used to generate robust hVISA detection models. In this study, MALDI-TOF mass spectra were obtained from 35 hVISA/vancomycin-intermediate S. aureus (VISA) and 90 vancomycin-susceptible S. aureus isolates. The vancomycin susceptibility of the isolates was determined using an Etest and modified population analysis profile–area under the curve. ML algorithms, namely a decision tree, k-nearest neighbors, random forest, and a support vector machine (SVM), were trained and validated using nested cross-validation to provide unbiased validation results. The area under the curve of the models ranged from 0.67 to 0.79, and the SVM-derived model outperformed those of the other algorithms. The peaks at m/z 1132, 2895, 3176, and 6591 were noted as informative peaks for detecting hVISA/VISA. We demonstrated that hVISA/VISA could be detected by analyzing MALDI-TOF mass spectra using ML. Moreover, the results are particularly robust due to a strict validation method. The ML models in this study can provide rapid and accurate reports regarding hVISA/VISA and thus guide the correct administration of antibiotics in treatment of S. aureus infection.

Patients with high vancomycin minimum inhibitory concentrations (MICs) and heteroresistant vancomycin-intermediate Staphylococcus aureus (hVISA) infection are associated with treatment failure and poor outcomes. The main purpose of this study was to investigate the effect of hVISA on patient outcome, considering both the high vancomycin MIC and the existence of heteroresistant phenotypes. From January 2005 to December 2009, consecutive meticillin-resistant S. aureus (MRSA) isolates from 284 cases of MRSA bacteraemia receiving glycopeptide therapy were collected for further MIC and hVISA testing. The demographic distribution, clinical features and outcomes in bacteraemia patients with different vancomycin MICs and hVISA status in MRSA isolates were subsequently compared. Subjects were divided into three groups: low vancomycin MIC (

In this article, we describe a case of left ventricular assist device endocarditis caused by a vancomycin-intermediate Staphylococcus aureus strain that was successfully treated with ceftaroline. Left ventricular assist device infections are briefly reviewed, and the current knowledge regarding vancomycin resistance is discussed.

We demonstrated a significant inverse correlation between vancomycin and beta-lactam susceptibilities in vancomycin-intermediate Staphylococcus aureus (VISA) and heterogeneous VISA (hVISA) isolates. Using time-kill assays, vancomycin plus oxacillin or ceftaroline was synergistic against 3 of 5 VISA and 1 of 5 hVISA isolates or 5 of 5 VISA and 4 of 5 hVISA isolates, respectively. Beta-lactam exposure reduced overall vancomycin-Bodipy ( dipy rromethene bo ron difluoride [4,4-difluoro-4-bora-3a,4a-diaza- s -indacene] fluorescent dye) binding but may have improved vancomycin-cell wall interactions to improve vancomycin activity. Further research is warranted to elucidate the mechanism behind vancomycin and beta-lactam synergy.

Vancomycin has been used as the last resort in the clinical treatment of serious Staphylococcus aureus infections. Vancomycin-intermediate S. aureus (VISA) was discovered almost two decades ago. Aside from the vancomycin-intermediate phenotype, VISA strains from the clinic or laboratory exhibited common characteristics, such as thickened cell walls, reduced autolysis, and attenuated virulence. However, the genetic mechanisms responsible for the reduced vancomycin susceptibility in VISA are varied. The comparative genomics of vancomycin-susceptible S. aureus (VSSA)/VISA pairs showed diverse genetic mutations in VISA; only a small number of these mutations have been experimentally verified. To connect the diversified genotypes and common phenotypes in VISA, we reviewed the genetic alterations in the relative determinants, including mutations in the vraTSR, graSR, walKR, stk1/stp1, rpoB, clpP, and cmk genes. Especially, we analyzed the mechanism through which diverse mutations mediate vancomycin resistance. We propose a unified model that integrates diverse gene functions and complex biochemical processes in VISA upon the action of vancomycin.

We investigated the prevalence of heterogeneous vancomycin-intermediate Staphylococcus aureus (hVISA) among methicillin-resistant S. aureus (MRSA) blood isolates collected from Korean hospitals. The hVISA prevalence in Korean hospitals has decreased during the past decade and the most significant decrease of hVISA prevalence was observed among the ST5 MRSA.

We report here the draft genome sequences of four vancomycin-intermediate Staphylococcus aureus (VISA) strains from South Korean hospitals participating in a nationwide laboratory surveillance program for vancomycin-intermediate and vancomycin-resistant Staphylococcus aureus . All strains harbor mutations in the walKR , graSR , and/or rpoB genes that are known frequently mutated determinants of VISA.

We report the complete genome sequence of vancomycin-intermediate Staphylococcus aureus (VISA) strain MI (HIP5827).

Rapid identification of community-associated (CA) methicillin-resistant Staphylococcus aureus (MRSA), hospital-associated (HA) MRSA, and vancomycin-intermediate S. aureus (VISA) is essential for proper therapy and timely intervention of outbreaks. In this study, peptide biomarkers for rapid identification of methicillin-resistant and vancomycin-intermediate S. aureus strains were discovered by matrix-assisted laser desorption ionization time-of-flight mass spectrometry. The results showed that the 1774.1 and 1792.1 m/z peaks corresponding to the phenol-soluble modulin α1 and phenol-soluble modulin α2 peptides, respectively, were present in the majority (95%, 121 of 127) of SCCmec types IV and V isolates, but only in 8% (15 of 185) of SCCmec types I-III isolates. Since SCCmec types I-III isolates are recognized as HA-MRSA and most CA-MRSA isolates belong to SCCmec types IV and V, these two peptides may serve as markers for discrimination between HA-MRSA and CA-MRSA isolates. The 1835.0 and 1863.0 m/z peaks were present in 50% (4 of 8) of heterogeneous VISA and 88% (14 of 16) of VISA isolates. The peptides of these two peaks were identified as proteolytic products of the acyl carrier protein. The results of this study provide the possibility to develop methods for identification of CA-MRSA, HA-MRSA, and vancomycin-resistant S. aureus isolates based on the presence of these peptides.

Reduced vancomycin susceptibility in Staphylococcus aureus can cause serious problems relating to treatment failure and persistent infection. We investigated vancomycin susceptibility, genetic relationships and transcriptional changes of the accessory gene regulator (agr) in vancomycin-intermediate S. aureus (VISA) strains isolated from South Korea compared with vancomycin-susceptible S. aureus (VSSA) strains. Molecular characterization, population analysis profiling, agr sequencing and transcriptional profiling of RNAIII by real-time RT-PCR were performed. Of 16 VISA strains tested, eight exhibited ST5, agr II and type II SCCmec. The others exhibited ST239, agr I and type III SCCmec. A point mutation in AgrA (Asp8Gly or Ile238Lys) was found in only five VISA strains; no mutations were detected in the other strains. However, RNAIII levels markedly decreased in all VISA strains (mean of 1.39-fold change) compared with the VSSA strains (31.51-fold change) in late-exponential phases (P

The aim of this study was to observe the survivability and fitness cost of heterogeneous vancomycin-intermediate Staphylococcus aureus(hVISA) isolates. Survivability study was performed on dry cotton swab, and fitness cost was evaluated by estimating growth kinetics and generation time constant in BACTEC automated system. Total mean maximum time of recovery on primary culture was 4.1 and 7.1 weeks (P = 0.0001) for hVISA and vancomycin-sensitive S. aureus (VSSA), respectively, in dry starved condition. No significant difference between the mean value of lag phase duration (P = 0.89) was noted between hVISA and VSSA isolate in growth kinetics. However, we observed lesser generation time of hVISA isolates compared to S. aureus ATCC 29213 (P = 0.0076). This study concluded that a significant difference in generation time between VSSA and hVISA and suggests that hVISA have fitness cost compared to VSSA. However, further studies with more cases are required.

The best screening method for detecting heteroresistant vancomycin-intermediate Staphylococcus aureus (hVISA) remains unclear. Using population analysis profiling utilizing the area under the concentration-time curve (PAP-AUC) as the gold standard, we screened 458 consecutive methicillin-resistant S. aureus (MRSA) bloodstream isolates to determine the most accurate and cost-effective testing strategy to detect the presence of heteroresistance. All isolates were also tested using the macromethod Etest (MET) and glycopeptide resistance detection (GRD) Etest. The MIC was determined by several methods, including standard vancomycin Etest, vancomycin broth microdilution (BMD), and Vitek2 testing. Fifty-five (12%) hVISA and 4 (1%) VISA isolates were detected by PAP-AUC. Compared to PAP-AUC, the sensitivities and specificities of MET, GRD Etest, BMD (using a MIC cutoff of ≥2 mg/liter), and standard vancomycin Etest (using a MIC cutoff of ≥2 mg/liter) were 89 and 55%, 71 and 94%, 82 and 97%, and 71 and 94%, respectively. Combination testing increased the overall testing accuracy by reducing the number of false-positive results. Cost was determined predominately by the number of PAP-AUC runs required following a screening assay. The most cost-effective strategy was BMD (using a MIC cutoff of ≥2 μg/ml) as a standalone assay or in combination with PAP-AUC, provided that BMD testing was batched. GRD Etest remained an alternative, with 71% of hVISA isolates detected. Prevalence influenced both cost and test accuracy, with results remaining unchanged for hVISA prevalences of up to 25%. Implementation of any testing strategy would therefore be dependent on balancing cost with accuracy in a given population and clinical context.

Background Vancomycin has historically been the mainstay of therapy for MRSA bacteremia, but severe infections due to vancomycin-intermediate Staphylococcus aureus have emerged. In vitro studies have shown that the combination of a β-lactam antibiotic, such as ceftaroline with daptomycin, was synergistic against MRSA. The purpose of this study was to compare outcomes in patients who received daptomycin and ceftaroline in combination vs. vancomycin for the treatment of MRSA bacteremia. Methods This was a retrospective exploratory cohort study approved by the institutional review board at Cooper University Hospital. Patients were included if they received daptomycin/ceftaroline (cases) or vancomycin (controls) for the treatment of MRSA bacteremia between November 2010 and March 2017. Cases were matched 1:1 with controls based on source of MRSA bacteremia, age within 10 years, and renal function. The primary endpoint was clinical cure, defined as the improvement of signs and symptoms of bacteremia. Secondary endpoints included microbiologic cure, time to sterilization of blood cultures, duration of hospital stay, overall mortality, and MRSA-related mortality. Results Forty-one cases were included. There was no statistical difference between the two groups in microbiologic cure, time to sterilization of blood cultures, overall mortality, or MRSA-related mortality. There were no significant differences between patients in each group including in those with ICU admissions and who required vasopressors. Cases were significantly more likely to have hardware compared with the control group (43.9% vs. 12.2%; P = 0.0014). Clinical cure was achieved in 27 patients (65.9%) in the case group and 26 patients (63.4%) in the control group (P = 0.8173). Patients in the case group had a statistically longer mean hospital duration (29 days vs. 21 days, respectively, P = 0.0206) and more secondary complications such as bone infection (P = 0.0076). Conclusion Time to sterilization of blood cultures and overall mortality were similar in both groups. Patients in the combination group had longer hospital stays compared with vancomycin monotherapy. Daptomycin/ceftaroline combination therapy is an option for complicated MRSA bacteremia. Larger studies should be conducted to further evaluate this combination. Disclosures All authors: No reported disclosures.

Vancomycin-intermediate Staphylococcus aureus (VISA) organisms have minimum inhibitory concentrations (MICs) of 4 to 8 microg/mL and are often associated with vancomycin treatment failure. Detection of VISA has remained problematic. A comparison of 4 methods to detect VISA was done. Of the 20 VISA isolates, the Clinical and Laboratory Standards Institute (CLSI) broth microdilution method yielded susceptible end points of 2 microg/mL for 7, MicroScan (Siemens Healthcare Diagnostics, West Sacramento, CA) for 2, Trek Sensititre (Trek Diagnostic Systems, Cleveland, OH) for 1, and Etest (AB Biodisk North America, Piscataway, NJ) for none. Comparison with the CLSI method showed essential agreement for 95% or more for the Etest, MicroScan, and Trek methods; categorical agreement was as follows: Etest, 60%; MicroScan, 65%; and Trek, 60%. Reliance on a single automated method for determining vancomycin MICs could lead to misclassification of some VISA isolates as vancomycin susceptible. At least 2 methods, including the Etest, should be used when confirming VISA because of slight differences in results from different methods around the end points of 2 and 4 microg/mL .

We compared the results obtained with six commercial MIC test systems (Etest, MicroScan, Phoenix, Sensititre, Vitek Legacy, and Vitek 2 systems) and three reference methods (agar dilution, disk diffusion, and vancomycin [VA] agar screen [VScr]) with the results obtained by the Clinical and Laboratory Standards Institute broth microdilution (BMD) reference method for the detection of VA-intermediate Staphylococcus aureus (VISA). A total of 129 S. aureus isolates (VA MICs by previous BMD tests, ≤1 μg/ml [ n = 60 strains], 2 μg/ml [ n = 24], 4 μg/ml [ n = 36], or 8 μg/ml [ n = 9]) were selected from the Centers for Disease Control and Prevention strain collection. The results of BMD with Difco Mueller-Hinton broth were used as the standard for data analysis. Essential agreement (percent ±1 dilution) ranged from 98 to 100% for all methods except the method with the Vitek Legacy system, for which it was 90.6%. Of the six commercial MIC systems tested, the Sensititre, Vitek Legacy, and Vitek 2 systems tended to categorize VISA strains as susceptible (i.e., they undercalled resistance); the MicroScan and Phoenix systems and Etest tended to categorize susceptible strains as VISA; and the Vitek Legacy system tended to categorize VISA strains as resistant (i.e., it overcalled resistance). Disk diffusion categorized all VISA strains as susceptible. No susceptible strains (MICs ≤ 2 μg/ml) grew on the VScr, but all strains for which the VA MICs were 8 μg/ml grew on the VScr. Only 12 (33.3%) strains for which the VA MICs were 4 μg/ml grew on VScr. The differentiation of isolates for which the VA MICs were 2 or 4 μg/ml was difficult for most systems and methods, including the reference methods.

Heterogeneous vancomycin intermediate Staphylococcus aureus (hVISA) strains are increasingly reported, and their association with vancomycin treatment failure is a well-known problem worldwide. A total of 500 clinical isolates of methicillin-resistant S. aureus were screened for hVISA by four different methods from May 2011 to May 2014. The number of strains screened for hVISA from May to April in 2011-2012, 2012-2013, and 2013-2014 were 198, 123, and 179, respectively. hVISA strains were studied by transmission electron microscopy (TEM) for cell wall thickness and also for their ability to form biofilm on a polystyrene microtiter plate. hVISA strains detected by four different methods-brain heart infusion agar with vancomycin with 4 mg/L/gradient plate/macro E-test/and glycopeptide resistance detection (GRD) E test-were as follows: 11.6%/10%/9%, and 9.5% in 2011-2012, 12.1%/9.7%/8.9%, and 10.5% in 2012-2013, and 13.9%/11.7%/11.1%, and 12.8% in 2013-2014, respectively. Population analysis profile-area under curve analysis confirmed hVISA in 4.5% (9/198), 6.5% (8/123), and 6.7% (12/179) in respective years; 24% (7/29) of hVISA isolates were nonsusceptible to daptomycin. TEM showed a significant increase in cell wall thickness of hVISA isolates (p0.001) with a distinct reduction in their biofilm formation ability.

Vancomycin resistance in Staphylococcus aureus has recently emerged as an important clinical problem with implications for laboratory detection and clinical management of patients infected with resistant strains. To date, low-level vancomycin resistance in the form of vancomycin-intermediate S. aureus (VISA) and heterogenous vancomycin-intermediate S. aureus (hVISA) have been more common, with only four cases of true vancomycin resistant S. aureus (VRSA) reported. This article reviews current knowledge about the epidemiology, clinical manifestations and optimal management of hVISA and VISA infections. VISA and hVISA have now been reported from many countries, and these strains tend to occur in patients with significant comorbidities and previous antibiotic exposure. Despite the difficulties in laboratory detection, there are increasing data linking VISA and hVISA to failure of glycopeptide antimicrobial therapy. Aggressive surgical intervention and non-glycopeptide-based antimicrobial therapy appears to improve outcomes for patients infected with these low-level vancomycin-resistant strains. Clinicians and diagnostic laboratories need to be aware of VISA and hVISA as a clinical problem, and consider aggressive surgical debridement and non-glycopeptide-based therapy where infections with such strains are suspected or proven.

The development of reduced vancomycin susceptibility in Staphylococcus aureus in many cases appears to be associated with characteristic changes. These changes may have pitfall of identifying S. aureus by automated testing methods like Vitek 32. In this study, we retested 24 heterogeneous vancomycin-intermediate Staphylococcus haemolyticus (h-VISH) collected in 2008-2010 at the Department of Clinical Microbiology by conventional biochemical tests and polymerase chain reaction (PCR). The heterogeneous vancomycin-intermediate S. aureus (hVISA) reversion test and electron microscopic examination were also used. Six isolates of 24 h-VISH possessed nuc, coa, and 16S rRNA genes, and could be reversed into S. aureus. It suggested that biochemical and morphological changes in hVISA and vancomycin-intermediate S. aureus (VISA) should be considered, and the detection of S. aureus, especially reduced vancomycin susceptibility isolates, requires more attention and different techniques.

We report a case of clearance of persistent bacteremia due to daptomycin non-susceptible, vancomycin-intermediate Staphylococcus aureus native mitral valve endocarditis with a combination of ceftaroline and daptomycin, in an 81-year-old medically complex patient who was not an operative candidate.

Background Staphylococcus aureus (S. aureus), especially Methicillin-resistant S. aureus (MRSA) remains a major cause of serious infection and is associated with increased morbidity and mortality. Vancomycin (VAN) has been the mainstay of therapy for MRSA infections. However, decades of selective pressure have led to increasing concerns regarding the efficacy of VAN against MRSA. In vitro data suggest the potential for potent synergy between several Β-lactams (BLs) and VAN. The objective of this study is to further explore the synergistic effect between BLs and VAN against MRSA strains with varying susceptibility to VAN. Methods Fifty randomly selected clinical MRSA strains from the Anti-Infective Research Laboratory library with varying susceptibility to VAN were evaluated for VAN alone and VAN in combination with Cefazolin (CFZ), Cefepime (FEP), Ceftaroline (CPT), and Nafcillin (NAF) minimum inhibitory concentration (MIC) by microdilution in duplicate. The potential for synergy was assessed by 24 hours time-kills (TK). Synergy was defined as >2 log10 CFU/mL difference between combination and the most active single agent at 24 hours. Results BLs reduced VAN MIC values against all strains (4–16 fold reduction). In TK studies against MRSA, all BLs demonstrated a similar extent of killing at 24 hours and showed synergy with VAN against all strains. Each combination was superior to any single agent alone, and each was bactericidal (3.42 ± 0.26 log10 CFU/cm2 reduction, P < 0.001 for all comparisons). All single agent exposures demonstrated no activity at 24 hours. Phenotype (Number of Strains) VAN MIC Range (µg/mL) VAN MIC Range with CFZ VAN MIC Range with FEP VAN MIC Range with CPT VAN MIC Range with NAF MRSA (15) 0.5 – 2 0.0625 – 0.25 0.0625 – 1 0.0625 – 0.25 0.125 – 0.5 hVISA (20) 0.5 – 2 0.125 – 0.5 0.0625 – 0.5 0.0625 – 0.5 0.125 – 1 VISA (15) 4 (all) 0.0625 – 0.5 0.5 – 2 0.0625 – 0.25 0.125 – 1 Conclusion The combination of VAN and BLs significantly improved antibacterial activity against MRSA, hVISA, and VISA compared with any agent alone, supporting the potential use of Vancomycin/BL combination therapy in infections caused by MRSA. Further clinical research is warranted to investigate the synergistic activity of vancomycin against these Staphylococcus strains. Disclosures M. J. Rybak, Allergen: Scientific Advisor, Consulting fee