Unmasking a looming crisis: Escalating MIC of last resort drugs against MRSA isolates from a tertiary care hospital in Central India.

PURPOSE: The Centers for Disease Control and Prevention has classified methicillin-resistant S aureus (MRSA) as a serious public health threat. The escalating minimum inhibitory concentration (MIC) of standard anti-methicillin-resistant S aureus (MRSA) drugs within the susceptible range, known as "MIC creep," jeopardizes their effectiveness against MRSA infections, posing additional challenges in managing MRSA infections. This cross-sectional study was conducted in a tertiary care hospital in Central India to assess the susceptibility trends of clinical MRSA isolates against commonly used anti-MRSA drugs and to observe MIC creep, if any, over three years (2020-2022). METHODS: The study included 158 non-repetitive clinical MRSA isolates. The MICs of vancomycin, teicoplanin, and linezolid were determined in MRSA strains using agar dilution, while the MIC of daptomycin was performed by broth microdilution. MIC creep was assessed by calculating MIC50, MIC90, Modal MIC, G-mean MIC, and susceptible and resistant percentages for the fiscal years 2020, 2021, and 2022. RESULTS: Of the 158 MRSA isolates, none were resistant to vancomycin, teicoplanin, and daptomycin, but two showed resistance to linezolid (LRSA). However, fifteen isolates showed intermediate resistance to vancomycin (VISA), and five showed intermediate resistance to teicoplanin (TISA). MIC of these anti-MRSA drugs increased in 2021 and 2022 compared to 2020. G-mean MIC for vancomycin, teicoplanin, and linezolid in MRSA strains increased significantly over the study period, while daptomycin MIC remained relatively stable, with a slight increase in 2021 and 2022. There was a high resistance rate for clindamycin, doxycycline, and chloramphenicol among VISA, TISA, and LRSA isolates compared to MRSA. CONCLUSIONS: During the three years of the study, "MIC creep" was observed in vancomycin, teicoplanin, and linezolid and, to some extent, for daptomycin in MRSA strains. The recovery of VISA, TISA, and linezolid-resistant MRSAs is worrisome, suggesting possible MRSA treatment failure and being a forerunner of resistant strains.

Heterogeneous Vancomycin Intermediate Staphylococcus aureus Infections in Diabetic and Non-Diabetic Patients - A Hospital-Based Comparative Study.

Purpose: To study the infections caused by methicillin resistant Staphylococcus aureus (MRSA) with emphasis on heterogeneous vancomycin intermediate S. aureus (hVISA) in diabetic and non-diabetic patients and their comparison. Patients and Methods: S. aureus strains isolated from diabetic and non-diabetic patients admitted in four tertiary care hospitals in Coastal Karnataka, South India, were tested for methicillin resistance and included in the present study. Demographic and clinical data of the patients were collected using structured proforma. Antimicrobial susceptibility testing was done using the Kirby-Bauer disc diffusion method, and MLSB phenotypes were identified using the D-test. The minimum inhibitory concentration (MIC) of vancomycin was determined using agar dilution. MRSA isolates were tested for hVISA using vancomycin screen agar and population analysis profile - area under the curve (PAP-AUC) test. Statistical analysis of the results was done using the chi-square test. SPSS version 29.0 was used for this purpose. Results: Out of 665 strains of S. aureus isolated, 220 (33.1%) were MRSA. Of these 220 MRSA strains, 122 (55.5%) and 98 (44.5%) were isolated from diabetic and non-diabetic patients, respectively. There was no significant difference in the antimicrobial resistance patterns of MRSA strains isolated from diabetic and non-diabetic patients. Foot infections and osteomyelitis caused by MRSA were significantly more among diabetic patients. Out of 220 strains of MRSA, 14 (6.4%) were hVISA. The rates of hVISA among MRSA isolated from diabetic and non-diabetic were 9.0% and 3.1%, respectively. This difference was statistically not significant. Conclusion: The rate of hVISA among all MRSA isolates was 6.4%. The risk of hVISA infection was three times more in diabetic patients. The results emphasize the importance of the detection of hVISA among MRSA isolates especially from diabetic patients.

Effect of Vancomycin on Cellular Fatty Acid Profiles of Vancomycin-Susceptible and Nonsusceptible Staphylococcus aureus.

Vancomycin is widely used for treatment of infection caused by methicillin-resistant Staphylococcus aureus (MRSA) leading to an increasing appearance of low-level vancomycin-resistant isolates called heterogeneous vancomycin-intermediate S. aureus (hVISA). The mechanism of vancomycin tolerance in hVISA is still unclear. This study aimed to investigate the fatty acid compositions of S. aureus isolates under the stress environment with vancomycin. The different responses of hVISA and vancomycin-susceptible S. aureus (VSSA) may lead to more understanding the mechanism. The bacterial lipid profiles were tested three times from three extractions of each isolate cultured on tryptic soy agar (TSA) and TSA with vancomycin. Of the 30 MRSA isolates studied, 13, 12, and 5 isolates were VSSA, hVISA, and VISA, respectively. The analysis of bacterial lipid profiles showed that under vancomycin stress, there was a reduction of straight chain fatty acids (SCFAs) in VSSA isolates but an increase in branched chain fatty acids (BCFAs). In contrast, the hVISA group exhibited an increase only in the BCFAs but not in SCFAs. Of interest, vancomycin had no effect on either BCFAs or SCFAs of the VISA cells. This study provided information of bacterial adaptation during stress with vancomycin that may be helpful to overcome the resistant bacteria.

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.

Emerging peptide antibiotics with therapeutic potential.

This review covers some of the recent progress in the field of peptide antibiotics with a focus on compounds with novel or established mode of action and with demonstrated efficacy in animal infection models. Novel drug discovery approaches, linear and macrocyclic peptide antibiotics, lipopeptides like the polymyxins as well as peptides addressing targets located in the plasma membrane or in the outer membrane of bacterial cells are discussed.

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.

The Potential Role of Sulbactam and Cephalosporins Plus Daptomycin Against Daptomycin-Nonsusceptible VISA and H-VISA Isolates: An in Vitro Study.

This study assesses the synergistic effect of the combination of cephalosporins and sulbactam with daptomycin against daptomycin-nonsusceptible, vancomycin-intermediate resistant Staphylococcus aureus (VISA) or heterogeneous vancomycin-intermediate S. aureus (h-VISA) isolates. The in vitro activity of daptomycin against daptomycin-nonsusceptible VISA/h-VISA isolates after adding cephalosporins with or without sulbactam was evaluated. The MIC of daptomycin against the VISA/h-VISA isolates was reduced after adding cephalosporins to daptomycin. Except for one VISA and two h-VISA isolates, the other VISA/h-VISA isolates became daptomycin-susceptible (MICs 1 mg/L). After adding sulbactam to each daptomycin/cephalosporin combination, the MIC of daptomycin against the VISA/h-VISA isolates decreased for 5 (33.3%), 6 (40.0%), 6 (40.0%), and 6 (40.0%) isolates with the cefazolin, cefmetazole, cefotaxime, and cefepime combinations, respectively. Synergism using the checkerboard method was noted in 100% of cefazolin and cefotaxime combinations and 87% and 80% of cefmetazole and cefepime combinations for all the VISA and h-VISA isolates. With the addition of sulbactam, synergism was noted in 100% of cefazolin, cefmetazole, and cefotaxime combinations and 93% of the cefepime combinations for all the VISA and h-VISA isolates. Almost all the FICs for the three-drug combinations were lower than those for the two-drug combinations. Using time-killing methods, a synergistic effect against five h-VISA isolates was observed. A synergistic effect of daptomycin, sulbactam, and each cephalosporin was observed for all VISA isolates. In conclusion, the activity of daptomycin against daptomycin-nonsusceptible VISA/h-VISA isolates can be enhanced by adding cephalosporins, and partially further promoted by sulbactam.

Methicillin-resistant Staphylococcus aureus (MRSA) and anti-MRSA activities of extracts of some medicinal plants: A brief review.

The increasing emergence of multidrug-resistant infection causing microorganisms has become a significant burden globally. Despite the efforts of pharmaceuticals in producing relatively new antimicrobial drugs, they have resulted in a high rate of mortality, disability and diseases across the world especially in developing countries. Supporting this claim was the report of the Centre for Disease Control and Prevention (CDC) who estimated that over 2 million illnesses and 23,000 deaths per year are attributable to antibiotic resistant pathogens in the United States. They include Methicillin-resistant Staphylococcus aureus (MRSA), Vancomycin-intermediate Staphylococcus aureus (VISA), Vancomycin-resistant Staphylococcus aureus (VRSA), Vancomycin-resistant enterococci (VRE), Extended spectrum beta-lactamases (ESBLs) producing gram-negative bacilli, Multidrug-resistant Streptococcus pneumoniae (MDRSP), Carbapenem-resistant Enterobacteriaceae (CRE) and Multidrug-resistant Acinetobacter baumannii. For MRSA, resistance is as a result of Methicillin-sensitive S. aureus (MSSA) strains that have acquired Staphylococcal Cassette Chromosome mec (SCCmec) which carries mecA gene. The gene encodes the penicillin-binding protein (PBP2a) which confers resistance to all ß-lactam antibiotics. Vancomycin was previously the widely preferred drug for the treatment of MRSA infections. It is no longer the case with the emergence of S. aureus strains with reduced vancomycin sensitivity limiting the conventional treatment options for MRSA infections to very scanty expensive drugs. Presently, many researchers have reported the antibacterial activity of many plant extracts on MRSA. Hence, these medicinal plants might be promising candidates for treatment of MRSA infections. This work is a brief review on Methicillin-resistant Staphylococcus aureus (MRSA) and the anti-MRSA activities of extracts of selected medicinal plants.

Rapid Detection of Heterogeneous Vancomycin-Intermediate Staphylococcus aureus Based on Matrix-Assisted Laser Desorption Ionization Time-of-Flight: Using a Machine Learning Approach and Unbiased Validation.

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.

Antimicrobial Activity of Dalbavancin against Staphylococcus aureus with Decreased Susceptibility to Glycopeptides, Daptomycin, and/or Linezolid from U.S. Medical Centers.

Dalbavancin activity was assessed against a large collection of Staphylococcus aureus isolates (n = 59,903), including isolates with decreased susceptibility to vancomycin (MIC, ≥2 mg/liter; n = 1,141), daptomycin (MIC, ≥2 mg/liter; n = 48), telavancin (MIC, ≥0.12 mg/liter; n = 52), teicoplanin (MIC, ≥4 mg/liter; n = 143), and/or linezolid (MIC, ≥8 mg/liter; n = 25). Dalbavancin displayed susceptibility rates ranging from 90.4% (isolates with telavancin MIC ≥0.12 mg/liter) to 100.0% (linezolid-resistant isolates) and lower MIC values than the comparators against these resistant subsets.

Correlation between resistance mechanisms in Staphylococcus aureus and cell wall and septum thickening.

PURPOSE: The aim of the present study is to examine cell wall and septum thickening of methicillin-sensitive Staphylococcus aureus (MSSA), methicillin-resistant S. aureus (MRSA), and methicillin- and linezolid-resistant S. aureus (MLRSA) isolates by transmission electron microscopy to correlate the association of resistance mechanisms with major changes in the morphology of membrane or septum. MATERIALS AND METHODS: MSSA, MRSA, and MLRSA strains obtained from clinical samples of an outbreak that occurred in 2010 at the Intensive Care Unit of our Hospital were thawed and sown at 37°C in blood agar overnight. After that, they were washed, pelleted, and treated with a fixer solution. Pellets were dehydrated and finally embedded in resin. Transmission electron microscopy was used to characterize cell wall and septum thickening in all isolates. The comparison between the measurements obtained for each group was performed by a Kruskal-Wallis test and a post hoc Dunn-Bonferroni's pairwise comparison method. RESULTS: Differences in cell wall and septum thickness were statistically significant (P<0.001 and P<0.001, respectively) between the three groups. Moreover, significant differences were detected in wall and septum thickness between the MSSA and MRSA strains (P<0.001 and P<0.001, respectively) and between the MSSA and MLRSA strains (P<0.001 and P<0.001, respectively) but not between the MRSA and MLRSA strains (P=0.386 and P=0.117). CONCLUSION: In this analysis, we correlate the resistance mediated by alterations in the cell membrane of S. aureus (methicillin-resistant, for example) with a greater thickness of the wall or septum. The resistance added to linezolid did not determine significant changes in the characteristics of the wall or septum with respect to those strains resistant only to methicillin.

Prevalence of vancomycin resistant Staphylococcus aureus (VRSA) in methicillin resistant S. aureus (MRSA) strains isolated from burn wound infections.

OBJECTIVES: The increase in resistance of methicillin resistant Staphylococcus aureus (MRSA) strains to vancomycin has been perceived as a formidable threat in the therapeutic fields. The present study investigated the vancomycin resistance traits of MRSA isolates [vancomycin resistant S. aureus (VRSA)] collected from burn patients. MATERIALS AND METHODS: Twenty-nine of 40 isolates of Staphylococcus spp. were identified as S. aureus which were further tested against 20 commercially available antibiotics to determine antibiotic susceptibility patterns. RESULTS: Imipenem was the most potential antibiotic resulting in 90% sensitivity, followed by netilmicin, clindamycin, and nitrofurantoin (80% sensitivity). All isolates were found to be resistant to penicillin. Approximately 75% of them were found to be resistant to methicillin, oxacillin, azithromycin, cipro-floxacin, and tetracycline. Approximately 45% isolates exhibited resistance to amikacin, chloramphenicol, gentamycin, and tobramycin. Twenty-one of the 29 strains of S. aureus were MRSA, of which 11 were resistant to vancomycin when employing the disc diffusion method. However, when the broth micro-dilution procedure was used to measure the minimum inhibitory concentration (MIC) of vancomycin, eight isolates were resistant to vancomycin, six with an MIC of 32 µg/mL and two with an MIC of 64 µg/mL. CONCLUSION: A significant fraction of VRSA was found among MRSA strains in this study, revealing the necessity for new and effective drugs against MRSA.

Molecular characterization of Staphylococcus aureus isolates carrying the Panton-Valentine leukocidin genes from Rio de Janeiro hospitals.

In a collection of 50 pvl-positive Staphylococcus aureus isolates from 10 Rio de Janeiro hospitals, 18 (36%) were from bloodstream infections, and 31 (62%) carried the SCCmec IV. Among 25 (50%) isolates of the USA1100/ST30/CC30 lineage present in 8 hospitals, 1 isolate was characterized as vancomycin-intermediate S. aureus.

New epidemiology of Staphylococcus aureus infection in Asia.

Not only is Asia the most populous region in the world, but inappropriate therapy, including self-medication with over-the-counter antimicrobial agents, is a common response to infectious diseases. The high antibiotic selective pressure among the overcrowded inhabitants creates an environment that is suitable for the rapid development and efficient spread of numerous multidrug-resistant pathogens. Indeed, Asia is among the regions with the highest prevalence rates of healthcare-associated methicillin-resistant Staphylococcus aureus (HA-MRSA) and community-associated methicillin-resistant S. aureus (CA-MRSA) in the world. Most hospitals in Asia are endemic for multidrug-resistant methicillin-resistant S. aureus (MRSA), with an estimated proportion from 28% (in Hong Kong and Indonesia) to >70% (in Korea) among all clinical S. aureus isolates in the early 2010s. Isolates with reduced susceptibility or a high level of resistance to glycopeptides have also been increasingly identified in the past few years. In contrast, the proportion of MRSA among community-associated S. aureus infections in Asian countries varies markedly, from <5% to >35%. Two pandemic HA-MRSA clones, namely multilocus sequence type (ST) 239 and ST5, are disseminated internationally in Asia, whereas the molecular epidemiology of CA-MRSA in Asia is characterized by clonal heterogeneity, similar to that in Europe. In this review, the epidemiology of S. aureus in both healthcare facilities and communities in Asia is addressed, with an emphasis on the prevalence, clonal structure and antibiotic resistant profiles of the MRSA strains. The novel MRSA strains from livestock animals have been considered to constitute a public health threat in western countries. The emerging livestock-associated MRSA strains in Asia are also included in this review.

Vancomycin Resistance of Staphylococcus aureus in Korean Primary Hospitals

According to a United States study, 13 cases of vancomycin-resistant Staphylococcus aureus (VRSA) have been reported to date. In 2001, a survey conducted in Korea revealed that 0.5% of methicillin-resistant S. aureus (MRSA) isolates have a vancomycin minimum inhibitory concentration (MIC) of 4 microg/ml, and are thus referred to as vancomycin intermediate S. aureus (VISA). However there are no reports of VISA found in primary hospitals. We evaluated the MIC of vancomycin in MRSA samples obtained from primary hospitals to determine whether VISA was present in primary hospitals. The population analysis was performed to determine whether hetero-VISA was present in primary hospitals. As a result, twenty of the 103 isolates were S. aureus which were all MRSA and the vancomycin MIC was similar to that seen in tertiary hospitals. Population analysis confirmed that three strains were hetero-VISA, by showing that one strain grew in 8 microg/ml vancomycin and that two strains grew in 4 microg/ml vancomycin. In conclusion, hetero-VISA was detected in Korean primary hospitals, which may develop into VISA, however a larger sample size will be needed to confirm these results.