Staphylococcus aureus response and adaptation to vancomycin.

Antibiotic resistance is an increasing challenge for the human pathogen Staphylococcus aureus. Methicillin-resistant S. aureus (MRSA) clones have spread globally, and a growing number display decreased susceptibility to vancomycin, the favoured antibiotic for treatment of MRSA infections. These vancomycin-intermediate S. aureus (VISA) or heterogeneous vancomycin-intermediate S. aureus (hVISA) strains arise from accumulation of a variety of point mutations, leading to cell wall thickening and reduced vancomycin binding to the cell wall building block, Lipid II, at the septum. They display only minor changes in vancomycin susceptibility, with varying tolerance between cells in a population, and therefore, they can be difficult to detect. In this review, we summarize current knowledge of VISA and hVISA. We discuss the role of genetic strain background or epistasis for VISA development and the possibility of strains being 'transient' VISA with gene expression changes mediated by, for example, VraTSR, GraXSR, or WalRK signal transduction systems, leading to temporary vancomycin tolerance. Additionally, we address collateral susceptibility to other antibiotics than vancomycin. Specifically, we estimate how mutations in rpoB, encoding the ß-subunit of the RNA polymerase, affect overall protein structure and compare changes with rifampicin resistance. Ultimately, such in-depth analysis of VISA and hVISA strains in terms of genetic and transcriptional changes, as well as changes in protein structures, may pave the way for improved detection and guide antibiotic therapy by revealing strains at risk of VISA development. Such tools will be valuable for keeping vancomycin an asset also in the future.

Risk factors for vancomycin treatment failure in heterogeneous vancomycin-intermediate Staphylococcus aureus bacteremia.

The incidence of heterogeneous vancomycin-intermediate Staphylococcus aureus (hVISA) infection is increasing and is associated with vancomycin treatment failures. However, studies investigating the risk factors for treatment failure in hVISA infection are limited. Patients with hVISA bacteremia treated with vancomycin over 7 days between August 2008 and June 2020 were enrolled in this study. Clinical and microbiological characteristics were compared between vancomycin treatment failure and success groups to identify the risk factors for vancomycin treatment failure. Among the 180 patients with hVISA bacteremia, 102 patients treated with vancomycin over 7 days were included. Vancomycin treatment failed in 80 (78%) patients. Patients in the vancomycin treatment failure group were older (P < 0.001) and more frequently had solid cancer (P = 0.04) than those in the vancomycin treatment success group. Solid organ transplantation (SOT) was more frequent (P < 0.001) in the vancomycin treatment success group. The Charlson comorbidity index (P = 0.01) and Acute Physiology and Chronic Health Evaluation II scores (P < 0.001) were higher in the vancomycin treatment failure group. In multivariate analysis, independent risk factors for vancomycin treatment failure were old age and severity of bacteremia. SOT and vancomycin minimal inhibitory concentration (MIC) ≤ 1.0 mg/L using the broth microdilution (BMD) method were associated with successful vancomycin treatment. Old age and infection severity were independent risk factors for vancomycin treatment failure. Vancomycin MIC using the BMD method is an important risk factor for vancomycin treatment failure, and its use should be considered in hVISA bacteremia.IMPORTANCEIn this study, we assessed the clinical and microbiological characteristics of heterogeneous vancomycin-intermediated Staphylococcus aureus (hVISA) bacteremia and identified risk factors for vancomycin treatment failure. We found that advanced age and severity of infection were independent risk factors for vancomycin treatment failure. On the other hand, solid organ transplantation and a low vancomycin minimal inhibitory concentration were associated with successful vancomycin treatment. This study highlights the importance of vancomycin minimal inhibitory concentration in hVISA bacteremia.

Diagnostic value of BHI-V4 for heterogeneous and vancomycin-intermediate Staphylococcus aureus isolates: a systematic review and meta-analysis.

BACKGROUND: Brain-heart infusion agar supplemented with 4 µg/mL of vancomycin (BHI-V4) was commonly used for the detection of heterogeneous (hVISA) and vancomycin-intermediate Staphylococcus aureus (VISA). However, its diagnostic value remains unclear. This study aims to compare the diagnostic accuracy of BHI-V4 with population analysis profiling with area under the curve (PAP-AUC) in hVISA/VISA. METHODS: The protocol of this study was registered in INPLASY (INPLASY2023120069). The PubMed and Cochrane Library databases were searched from inception to October 2023. Review Manager 5.4 was used for data visualization in the quality assessment, and STATA17.0 (MP) was used for statistical analysis. RESULTS: In total, eight publications including 2153 strains were incorporated into the meta-analysis. Significant heterogeneity was evident although a threshold effect was not detected across the eight studies. The summary receiver operating characteristic (SROC) was 0.77 (95% confidence interval [CI], 0.74-0.81). The pooled sensitivity, specificity, positive likelihood ratio, negative likelihood ratio, diagnostic score and diagnostic odds ratio were 0.59 (95% CI: 0.46-0.71), 0.96 (95%CI: 0.83-0.99), 14.0 (95% CI, 3.4-57.1), 0.43 (95%CI, 0.32-0.57), 3.48(95%CI, 2.12-4.85) and 32.62 (95%CI, 8.31-128.36), respectively. CONCLUSION: Our study showed that BHI-V4 had moderate diagnostic accuracy for diagnosing hVISA/VISA. However, more high-quality studies are needed to assess the clinical utility of BHI-V4.

Detection of Vancomycin Resistance among Methicillin-Resistant Staphylococcus aureus Strains Recovered from Children with Invasive Diseases in a Reference Pediatric Hospital.

Vancomycin is the cornerstone in treating methicillin-resistant Staphylococcus aureus (MRSA) infections. However, therapeutic failures can occur when MRSA strains with decreased susceptibility to glycopeptides (DSG) are involved. The aim of this study was to detect and characterize DSG in MRSA recovered from children with invasive diseases at a reference pediatric hospital between 2009 and 2019. Fifty-two MRSA strains were screened using agar plates with vancomycin 3 and 4 mg/L (BHI-3 and BHI-4); the VITEK2 system; and standard and macro E-tests. Suspicious hVISA were studied by population analysis profiling-area under the curve (PAP-AUC), and wall thickness was analyzed by transmission electron microscopy. Neither VRSA nor VISA were detected in this set. As only three strains met the hVISA criteria, the PAP-AUC study included 12 additional MRSA strains that grew one colony on BHI-4 plates or showed minimum inhibitory concentrations of vancomycin and/or teicoplanin ≥ 1.5 mg/L. One strain was confirmed as hVISA by PAP-AUC. The wall thickness was greater than the vancomycin-susceptible control strain; it belonged to ST30 and carried SCCmec IV. As expected, a low frequency of hVISA was found (1.9%). The only hVISA confirmed by PAP-AUC was not detected by the screening methods, highlighting the challenge that its detection represents for microbiology laboratories.

Global distribution of heterogeneous vancomycin-intermediate Staphylococcus aureus strains (1997-2021): a systematic review and meta-analysis.

BACKGROUND: Heterogeneous vancomycin-intermediate Staphylococcus aureus is considered one of the main causes in treatment failure of vancomycin, which leads to poor clinical outcomes. Herein, we comprehensively evaluated characteristics such as global prevalence, trend, and genetic backgrounds of these strains. METHODS: In this study, we conducted a meta-analysis based on PRISMA checklist 2020. In the beginning, global databases were searched to achieve the studies related to the prevalence of hVISA in clinical isolates of methicillin-resistant Staphylococcus aureus. After retrieving the eligible English studies, the prevalence of hVISA isolates and their trend changes were assessed using event rate with 95% confidence intervals. RESULTS: In the present study, the prevalence of 114 801 MRSA isolates (of 124 studies) was 64%. According to our results, although the frequency of infection with hVISA is increasing in recent years, there is not a significant difference between Asian countries and Europe/America (6.1% vs. 6.8%). In addition, infection with hVISA bacteria was higher in bacteraemic patients than other infections (9.4% vs. 5.5%), which increases hospitalization, treatment costs, and mortality in these patients. Isolates harbouring SCCmec types II and III are most common genotypes in hVISA strains. CONCLUSIONS: The prevalence of hVISA is increasing, which will reduce the effectiveness of vancomycin treatment in the coming years. The presence of hVISA stains in blood samples was higher than the other samples, which is threatening for bacteraemic patients. The results of the current study indicate a universal program to identify and control the spread of such strains in nosocomial infections.

Comparative proteomic analysis of vancomycin-sensitive and vancomycin-intermediate resistant Staphylococcus aureus.

PURPOSE: The extensive use of vancomycin has led to the development of Staphylococcus aureus strains with varying degrees of resistance to vancomycin. The present study aimed to explore the molecular causes of vancomycin resistance by conducting a proteomics analysis of subcellular fractions isolated from vancomycin-intermediate resistant S. aureus (VISA) and vancomycin-sensitive S. aureus (VSSA) strains. METHODS: We conducted proteomics analysis of subcellular fractions isolated from 2 isogenic S. aureus strains: strain 11 (VSSA) and strain 11Y (VISA). We used an integrated quantitative proteomics approach assisted by bioinformatics analysis, and comprehensively investigated the proteome profile. Intensive bioinformatics analysis, including protein annotation, functional classification, functional enrichment, and functional enrichment-based cluster analysis, was used to annotate quantifiable targets. RESULTS: We identified 128 upregulated proteins and 21 downregulated proteins in strain 11Y as compared to strain 11. The largest group of differentially expressed proteins was composed of enzymatic proteins associated with metabolic and catalytic activity, which accounted for 32.1% and 50% of the total proteins, respectively. Some proteins were indispensable parts of the regulatory networks of S. aureus that were altered with vancomycin treatment, and these proteins were related to cell wall metabolism, cell adhesion, proteolysis, and pressure response. CONCLUSION: Our proteomics study revealed regulatory proteins associated with vancomycin resistance in S. aureus. Some of these proteins were involved in the regulation of cell metabolism and function, which provides potential targets for the development of strategies to manage vancomycin resistance in S. aureus.

Adaptive Laboratory Evolution of Staphylococcus aureus Resistance to Vancomycin and Daptomycin: Mutation Patterns and Cross-Resistance.

Vancomycin and daptomycin are first-line drugs for the treatment of complicated methicillin-resistant Staphylococcus aureus (MRSA) infections, including bacteremia. However, their effectiveness is limited not only by their resistance to each antibiotic but also by their associated resistance to both drugs. It is unknown whether novel lipoglycopeptides can overcome this associated resistance. Resistant derivatives from five S. aureus strains were obtained during adaptive laboratory evolution with vancomycin and daptomycin. Both parental and derivative strains were subjected to susceptibility testing, population analysis profiles, measurements of growth rate and autolytic activity, and whole-genome sequencing. Regardless of whether vancomycin or daptomycin was selected, most of the derivatives were characterized by a reduced susceptibility to daptomycin, vancomycin, telavancin, dalbavancin, and oritavancin. Resistance to induced autolysis was observed in all derivatives. Daptomycin resistance was associated with a significant reduction in growth rate. Resistance to vancomycin was mainly associated with mutations in the genes responsible for cell wall biosynthesis, and resistance to daptomycin was associated with mutations in the genes responsible for phospholipid biosynthesis and glycerol metabolism. However, mutations in walK and mprF were detected in derivatives selected for both antibiotics.

Comparative Genomics Identifies Novel Genetic Changes Associated with Oxacillin, Vancomycin and Daptomycin Susceptibility in ST100 Methicillin-Resistant Staphylococcus aureus.

Infections due to vancomycin-intermediate S. aureus (VISA) and heterogeneous VISA (hVISA) represent a serious concern due to their association with vancomycin treatment failure. However, the underlying molecular mechanism responsible for the hVISA/VISA phenotype is complex and not yet fully understood. We have previously characterized two ST100-MRSA-hVISA clinical isolates recovered before and after 40 days of vancomycin treatment (D1 and D2, respectively) and two in vitro VISA derivatives (D23C9 and D2P11), selected independently from D2 in the presence of vancomycin. This follow-up study was aimed at further characterizing these isogenic strains and obtaining their whole genome sequences to unravel changes associated with antibiotic resistance. It is interesting to note that none of these isogenic strains carry SNPs in the regulatory operons vraUTSR, walKR and/or graXRS. Nonetheless, genetic changes including SNPs, INDELs and IS256 genomic insertions/rearrangements were found both in in vivo and in vitro vancomycin-selected strains. Some were found in the downstream target genes of the aforementioned regulatory operons, which are involved in cell wall and phosphate metabolism, staphylococcal growth and biofilm formation. Some of the genetic changes reported herein have not been previously associated with vancomycin, daptomycin and/or oxacillin resistance in S. aureus.

Reduced Susceptibility and Resistance to Vancomycin of Staphylococcus aureus: A Review of Global Incidence Patterns and Related Genetic Mechanisms.

Staphylococcus aureus is a Gram-positive bacterium causing a wide range of infections ranging from cutaneous infections to endocarditis and bacteremia. Beta-lactamases such as penicillin and, subsequently, methicillin have been used in the treatment of S. aureus infections. With the emergence of methicillin-resistant Staphylococcus aureus (MRSA), vancomycin, a bacterial cell wall synthesis inhibitor, has been used as the treatment of choice for MRSA infections. However, over the past few decades, there have been reports of reduced susceptibility and resistance of S. aureus to vancomycin globally, most recently from Michigan, United States, in July 2021. Based on the minimum inhibitory concentration (MIC) of the antibiotic against S. aureus, there are three strains of resistance, vancomycin-intermediate Staphylococcus aureus (VISA), vancomycin-resistant Staphylococcus aureus (VRSA), and heterogeneous vancomycin-intermediate Staphylococcus aureus (hVISA). The increasing prevalence of VISA and VRSA infections is a cause of global concern. This qualitative review of peer-reviewed research publications aims to describe the cases of VISA and VRSA reported in the literature globally and summarizes the genetic mechanisms implicated in their resistance. The most common mechanism implicated in VRSA infections is the vanA operon, while cell wall thickening is responsible for VISA infections. This review aims to perform a global comparison between the MIC corresponding to the strength of resistance to vancomycin and the presence of the vanA operon. In this review, VISA and VRSA are noted to be most susceptible to quinupristin-dalfopristin and linezolid, respectively. Maintaining active systemic surveillance for such infections, employing strict infection control measures, and continuing to mitigate indiscriminate and irrational use of antibiotics are some of the actions that can be undertaken to reduce the incidence and transmission of VISA, VRSA, and hVISA infections worldwide.

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.

Cell wall thickness and the molecular mechanism of heterogeneous vancomycin-intermediate Staphylococcus aureus.

Methicillin-resistant Staphylococcus aureus (MRSA) with reduced sensitivity to vancomycin (VAN) has caused many clinical cases of VAN treatment failure, but the molecular mechanism underlying the reduced sensitivity to VAN is still unclear. We isolated a heterogeneous VAN-intermediate Staphylococcus aureus (hVISA), which was also a MRSA strain with reduced sensitivity to VAN. To investigate the molecular mechanism underlying the reduced sensitivity to VAN exhibited by the hVISA strain, we compared the hVISA strain with a VAN-sensitive MRSA strain, known as the N315 strain. The images captured by transmission electron microscopy showed that the cell wall of the hVISA strain was significantly thicker than that of the N315 strain (36·72 ± 1·04 nm vs 28·15 ± 1·25 nm, P < 0·05), and the results of real-time quantitative PCR analysis suggested that the expression levels of the cell wall thickness related genes (glmS, vraR/S, sgtB, murZ and PBP4) of the hVISA strain were significantly higher than those of the N315 strain (P < 0·05). In conclusion, this study indicated that the upregulation of the expression of the genes related to cell wall synthesis might be the molecular mechanism underlying the cell wall thickening of the hVISA strain and might be related to its resistance to VAN.

Protein Biomarker Discovery for Methicillin-Sensitive, Heterogeneous Vancomycin-Intermediate and Vancomycin-Intermediate Staphylococcus aureus Strains Using Label-Free Data-Independent Acquisition Proteomics.

Rapid identification of methicillin-sensitive Staphylococcus aureus (MSSA), heterogeneous vancomycin-intermediate S. aureus (hVISA), and vancomycin-intermediate S. aureus (VISA) is important for accurate treatment, timely intervention, and prevention of outbreaks. Here, 90 S. aureus isolates were analyzed for protein biomarker discovery, including MSSA, vancomycin-susceptible S. aureus (VSSA), hVISA, and VISA strains. Label-free data-independent acquisition proteomics was used to identify protein biomarkers that allow for discrimination among MSSA, hVISA, and VISA strains. There were 8786 nonredundant peptides identified, corresponding to 418 different annotated nonredundant proteins. Two VISA protein biomarkers, two hVISA protein biomarkers, and one MSSA protein biomarker with high sensitivities and specificities were discovered and verified. Data are available via MassIVE with identifier MSV000085776.

Bacteriemia por Staphylococcus aureus con sensibilidad disminuida heterogénea a vancomicina: reporte de un caso / Staphylococcus aureus bacteremia with heterogeneous intermediate-level vancomycin resistance: a case report

Staphylococcus aureusmeticilina-resistente (SAMR)es una causa frecuente de bacteriemias intrahospitalarias. Para su tratamiento se utiliza vancomicina y han emergido cepas con sensibilidad disminuida heterogénea (h-VISA) que albergan subpoblaciones con sensibi-lidad reducida a vancomicina. Se comunica un caso de bacteriemia intra-tratamiento con vancomicina por SAMR h-VISA. El aislamiento muestra sensibilidad a vancomicina (CIMvan: 1 µg/mL), sin embargo E-test GRD sugiere h-VISA (CIMvan: 2 µg/mL y CIMtei: 8 µg/mL). El análisis del perfil poblacional - área bajo la curva (PAP-AUC) valida este hallazgo. Se rota a linezolid con resolución clínica.

Methicillin-resistant Staphylococcus aureus (SAMR) is a common cause of nosocomial bacteremia. Vancomycin, a glycopeptide, is widely employed for the therapy of SAMR infections. In recent years, heterogeneous vancomycin-intermediate strains (h-VISA) have emerged. We report a case of intra-treatment bacteremia caused by SAMR h-VISA. The isolate shows susceptibility to vancomycin (MICvan: 1 µg/mL). But the GRD E-test suggests h-VISA (MICvan: 2 µg/mL and MICtei: 8 µg/mL). The population analysis profile - area under the curve (PAP-AUC) validates SAMR h-VISA. Rotation of antibiotic therapy with linezolid is done, with good clinical outcome.

Detection of heterogeneous vancomycin-intermediate Staphylococcus aureus: A preliminary report from south India.

Background & objectives: Although there are reports of heterogeneous vancomycin-intermediate Staphylococcus aureus (hVISA) across the globe, there is a lack of reliable data on hVISA in India. The present study was undertaken to determine the rate of hVISA among the methicillin-resistant Staphylococcus aureus (MRSA) isolates, and to compare the brain heart infusion agar with vancomycin 4 µg/ml (BHIV4) method with population analysis profile-area under the curve (PAP-AUC) method for the detection of hVISA and to study the distribution of mobile genetic element that carries methicillin-resistance gene SCCmec (Staphylococcal cassette chromosome mec) types among these isolates. Methods: BHIV4 and PAP-AUC methods were employed to detect hVISA among 500 clinical isolates of MRSA. SCCmec typing of these isolates was performed by multiplex polymerase chain reaction. The clinical presentation, treatment with vancomycin and outcome was documented for patients with hVISA. Results: The rate of hVISA was 12.4 per cent by PAP-AUC method. Sensitivity, specificity, PPV, NPV and kappa agreement of BHIV4 with PAP-AUC was 58.06, 93.15, 54.55, 94.01 per cent and 0.498, respectively. The isolation of hVISA was significantly (P<0.01) higher in patients admitted to intensive care units and wards than in patients attending the outpatient departments. Only 38 per cent of the patients received vancomycin as therapy. Majority of the hVISA isolates carried SCCmec type V or IV. Interpretation & conclusions: The rate of hVISA isolation in our study was 12.4 per cent. The sensitivity of the BHIV4 screening test was low, and was in moderate agreement with PAP-AUC test. SCCmec type V was the predominant type seen in half of the isolates. More studies need to be done in different parts of the country on a large number of isolates to confirm our findings.

Impact of mutations in hVISA isolates on decreased susceptibility to vancomycin, through population analyses profile - area under curve (PAP-AUC).

We analyzed sequences of graSR, vraSR, walKR and rpoB genes in hVISA from Brazil. Five isolates showed mutations in at least one gene. rpoB H481N and graS T224I were the most frequent mutations, followed by graR D148Q and walK A468T. Our study reinforces the heterogeneity of genetic patterns among hVISA.

Reduced vancomycin susceptibility and increased macrophage survival in Staphylococcus aureus strains sequentially isolated from a bacteraemic patient during a short course of antibiotic therapy.

PURPOSE: The purpose of the present study was to determine the relatedness of Staphylococcus aureus strains successively isolated over a 7-day period from a single bacteraemic patient undergoing antibiotic treatment with vancomycin. METHODS: The S. aureus strains had been isolated and sequenced previously. Antibiotic susceptibility testing, population analysis profiling, and lysostaphin sensitivity and phagocytic killing assays were used to characterize these clonal isolates. RESULTS: The seven isolates (MEH1-MEH7) were determined to belong to a common multilocus sequence type (MLST) and spa type. Within the third and fifth day of vancomycin treatment, mutations were observed in the vraS and rpsU genes, respectively. Population analysis profiles revealed that the initial isolate (MEH1) was vancomycin-susceptible S. aureus (VSSA), while those isolated on day 7 were mostly heteroresistant vancomycin-intermediate S. aureus (hVISA). Supporting these findings, MEH7 was also observed to be slower in growth, to have an increase in cell wall width and to have reduced sensitivity to lysostaphin, all characteristics of VISA and hVISA strains. In addition, MEH7, although phagocytosed at numbers comparable to the initial isolate, MEH1, survived in higher numbers in RAW 264.7 macrophages. Macrophages infected with MEH7 also released more TNF-α and IFN-1ß. CONCLUSION: We report an increasing resistance to vancomycin coupled with daptomycin that occurred within approximately 3 days of receiving vancomycin and steadily increased until the infection was cleared with an alternative antibiotic therapy. This study reiterates the need for rapid, efficient and accurate detection of hVISA and VISA infections, especially in high-bacterial load, metastatic infections like bacteraemia.

Vancomycin MIC Distribution among Methicillin-Resistant Staphylococcus Aureus. Is Reduced Vancomycin Susceptibility Related To MIC Creep?

AIM: To determine the distribution of vancomycin MIC and the frequency of S. aureus strains with reduced vancomycin susceptibility among Methicillin-Resistant Staphylococcus aureus (MRSA) isolates. METHODS: MRSA isolates (n = 100) were tested for reduced susceptibility to vancomycin using MIC broth microdilution method (BMD), vancomycin screening agar with different vancomycin concentrations with and without casein, and Vitek 2 system. RESULTS: BMD detected (22%) vancomycin-intermediate S. aureus (VISA) and (78%) vancomycin-susceptible S. aureus (VSSA) but couldn't detect nine (Heterogeneous VISA) (hVISA) isolates (9%) with MIC ≤ 2 µg/ml that grew on screening agar 4 µg/ml or 6 µg/ml. Adding casein to vancomycin screening agar increased detection rate of VISA by 4.5%. Screening agar with 6 µg/ml vancomycin overall detection rate for VISA was 95.45%. Probable 'pre-hVISA'isolates (17%) showed growth on vancomycin screening agar 2 µg/ml with casein. Vitek 2 system failed to detect any VISA isolates. CONCLUSION: Vancomycin screening agar; 2 µg/ml and (4 and 6 µg/ml) were able to detect; probable "pre hVISA and (hVISA and VISA) isolates respectively based on their BMD MIC values. Decreased vancomycin susceptibility in MRSA isolates might be related to MIC creep. Analysis of vancomycin MIC values over longer periods is recommended to further study this phenomenon and its impact on vancomycin treatment failure.

An invasive marker Staphylococcus epidermidis surface protein I (SesI) harboured by a ST239 methicillin-resistant Staphylococcus aureus.

OBJECTIVES: Recently, the virulence factor sasX was described on the mobile genetic element φSPß prophage in a ST239-SCCmecIII methicillin-resistant Staphylococcus aureus (MRSA) clone. The aim of this study was to identify sesI, an sasX homologue, in an MRSA ST239 strain. METHODS: MRSA strain VB1490 was isolated from a patient with MRSA bacteraemia in India. Staphylococcal cassette chromosomemec (SCCmec) typing and whole-genome shotgun sequencing were performed. Phylogenetic analysis of VB1490 and ST239 reference genomes from the NCBI database was performed. Amplification and sequencing of the sasX gene was performed to establish allele homology. RESULTS: The sasX gene identified in isolate VB1490 was more similar to the sesI gene of Staphylococcus epidermidis RP62A than the sasX gene of ST239 MRSA strain. Whole-genome analysis revealed the presence of the sasX gene on a φSPß-like prophage that exhibited high sequence identity to that of S. epidermidis strain RP62A. CONCLUSION: These finding suggests the dissemination of the invasion-determining virulence factorsesI from S. epidermidis to a ST239 MRSA strain.

Evidence from an In Vitro Study: Is Oxacillin Plus Vancomycin a Better Choice for Heteroresistant Vancomycin-Intermediate Staphylococcus aureus?

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.

Características moleculares de Staphylococcus aureus susceptível à vancomicina poderia ajudar a prever falhas no tratamento devido a reduzida susceptibilidade à vancomicina / Molecular characteristics of vancomycin-susceptible Staphylococcus aureus could help to predict treatment failure due to reduced vancomycin susceptibility / Características moleculares de Staphylococcus aureus susceptible a la vancomicina podría ayudar a predecir fallas en el tratamiento debido a la reducida susceptibilidad a la vancomicina

Justificativa e Objetivos: Staphylococcus aureus resistente à meticilina (MRSA) é uma das causas mais frequentes de infecções relacionadas à assistência à saúde e comunitárias, e com seu avanço, a vancomicina tornou-se a principal opção terapêutica. Entretanto, o seu uso indiscriminado favoreceu o surgimento de MRSA com reduzida suscetibilidade à vancomicina, comumente associados com falhas no tratamento, bacteremia persistente, hospitalização prolongada e desfechos clínicos adversos. Este estudo avaliou a ocorrência de MRSA com reduzida suscetibilidade à vancomicina e determinou algumas características moleculares em comparação com MRSA suscetível à vancomicina (VS-MRSA). Métodos: Determinação do perfil de suscetibilidade aos antimicrobianos, a concentração inibitória mínima (CIM) e concentração bactericida mínima (CBM) para vancomicina, tolerância à vancomicina, tipagem do SCCmec e agr foram realizadas em um total de 177 MRSA. Posteriormente, foram triados para hVISA por BHIA-3V e BHIA-6V e confirmados com a Análise do Perfil Populacional - Área Abaixo da Curva (PAP-AUC). Resultados: Os fenótipos VT-MRSA e hVISA foram encontrados em 13,6% e 5,1% dos isolados clínicos de MRSA, respectivamente, e a presença de hVISA foi estatisticamente significativa entre os isolados de VT-MRSA (p<0,05). Em VT-MRSA, SCCmec tipo II foi significativamente mais frequente do que em não-VT-MRSA, assim como a presença do agr grupo II. Conclusão: Características moleculares encontradas em MRSA são importantes para a epidemiologia, bem como para demonstrar um padrão em isolados com reduzida suscetibilidade à vancomicina. Testes não-convencionais para detecção destas características podem ser realizados para evitar a identificação errada de VS-MRSA que, consequentemente, resulta em falhas no tratamento com vancomicina.(AU)

Background and Objectives: Methicillin-resistant Staphylococcus aureus (MRSA) is one of the most frequent causes of healthcare-associated and community-acquired infections and with its advancement, vancomycin became the main therapeutic option. However, its indiscriminate use favored the emergence of MRSA with reduced susceptibility to vancomycin, commonly associated with vancomycin treatment failure, persistent bacteremia, prolonged hospitalization and adverse clinical outcome. This study evaluated the occurrence of MRSA with reduced vancomycin susceptibility and determined some molecular characteristics in comparison with vancomycin-susceptible MRSA (VS-MRSA). Methods: Determination of antimicrobial susceptibility profile, the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) for vancomycin, vancomycin-tolerance, SCCmec and agr typing were performed in a total of 177 MRSA. Thereafter, they were screened for hVISA by BHIA-3V and BHIA-6V and confirmed with population analysis profile - area under the curve method (PAP-AUC). Results: VT-MRSA and hVISA phenotypes were found in 13.6% and 5.1% of clinical isolates of MRSA, respectively, and the presence of hVISA was statistically significant among VT-MRSA isolates (p<0.05). In T-MRSA, SCCmec type II was significantly more frequent than in non-VT-MRSA, as well as the presence of agr group II. Conclusion: Molecular characteristics found in MRSA are important for epidemiology, as well as demonstrate a pattern in reduced vancomycin susceptibility isolates. Non-conventional tests for detection of these characteristics might be performed to prevent misidentification of VS-MRSA that, consequently, results in vancomycin treatment failures.(AU)

Justificación y objetivos: Staphylococcus aureus resistente a la meticilina (MRSA) es una de las causas más frecuentes de infecciones relacionadas con la asistencia sanitaria y comunitarias, y con su avance, a la vancomicina se ha convertido en la principal opción terapéutica. Sin embargo, su uso indiscriminado favoreció el surgimiento de MRSA con reducida susceptibilidad a la vancomicina, comúnmente asociados con fallas en el tratamiento, bacteriemia persistente, hospitalización prolongada y resultados clínicos adversos. Este estudio evaluó la ocurrencia de MRSA con reducida susceptibilidad a la vancomicina y determinó algunas características moleculares en comparación con MRSA susceptible a la vancomicina (VS-MRSA). Métodos: Determinación del perfil de susceptibilidad a los antimicrobianos, la concentración inhibitoria mínima (CIM) y la concentración bactericida mínima (CBM) para vancomicina, tolerancia a la vancomicina, tipificación del SCCmec y agr se realizaron en un total de 177 MRSA. Resultados: Los fenotipos VT-MRSA y hVISA se encontraron en el 13,6% y el 5,1% de los aislados clínicos de MRSA, respectivamente, y la presencia de hVISA fue estadísticamente significativa entre los aislados de VT-MRSA (p<0.05). En VT-MRSA, SCCmec tipo II fue significativamente más frecuente que en no-VT-MRSA, así como la presencia del agr grupo II. Conclusión: Características moleculares encontradas en MRSA son importantes para la epidemiología, así como para demostrar un patrón en aislados con reducida susceptibilidad a la vancomicina. Pruebas no convencionales para la detección de estas características pueden realizarse para evitar la identificación errónea de VS-MRSA que, consecuentemente, resulta en fallas en el tratamiento con vancomicina.(AU)