BACKGROUND: A clonal shift from staphylococcal cassette chromosome mec (SCCmec) type II/ST5 methicillin-resistant Staphylococcus aureus (MRSA) to SCCmec type IV/clonal complex (CC)1 MRSA has occurred rapidly in Japan. Our previous research in a geriatric hospital found SCCmec type IV/CC1 MRSA prevalence in long-term care wards. Due to intensive personal care requirements, frequent contact with healthcare providers can potentially cause unintentional nosocomial MRSA transmission. We performed polymerase chain reaction-based open reading frame typing (POT) and core genome multilocus sequence typing (cgMLST) to investigate the occurrence of nosocomial transmission and to compare the results of these methods.
METHODS: POT and whole genome sequencing were performed in 83 MRSA isolates. Commercial automated software (Ridom SeqSphere+) was used to perform cgMLST. MRSA isolates with 0 to 8 allelic differences were considered related, and medical records were consulted in these cases.
RESULTS: SCCmec type IV/CC1 MRSA was the most frequently detected clone (n=56, 67.5%), which was divided into 14 POT types, followed by SCCmec type I/ST8 (n=9) and SCCmec type IV/ST8 (n=8). Identical POT types were found across 7 of 11 wards. However, cgMLST analysis identified only three cases (six strains) of high genetic similarity, indicating nosocomial transmission; only one involved SCCmec type IV/CC1 (two strains). The mean allelic difference in the core genomes between strains with identical POT types in the same ward was 55.3 ± 22.0.
CONCLUSIONS: The cgMLST method proved more effective for identifying nosocomial transmissions compared to POT, highlighting its utility in tracking MRSA spread in healthcare settings.

Escherichia coli and Staphylococcus aureus are the most important food borne pathogen transmitting from animal meat and meat products. Therefore, it is vital to design an accurate and specific diagnostic tool for identifying those food-borne pathogens in animal meat and meat products. In the current study, E. coli, methicillin-resistant and sensitive S. aureus (MRSA and MSSA) were simultaneously detected using a developed triplex PCR-based technique. To obtain an optimal reaction parameter, the multiplex assay was optimised by changing just one parameter while holding the others constant. Specificity of the assay was assessed using several porcine bacterial template DNA. The plasmid DNA was used to test the multiplex PCR assay\'s sensitivity and interference in spiked pork samples. E. coli, MRSA, and MSSA each have PCR amplified products with sizes of 335, 533, and 209 bp, respectively. The assay detects a minimum microbial load of 102 CFU/μl for all the three pathogens and can identify bacterial DNA as low as 10-2 ng/µl. The assay was validated employing 210 pork samples obtained from retail meat shops and slaughter houses, with MRSA, E. coli, and MSSA with the occurrence rate of 1.9%, 42.38%, and 18.1%, respectively. The rate of mixed bacterial contamination in pork meat samples examined with the developed method was 6.19%, 1.43%, 1.90%, and 1.43% for MSSA & E. coli, MRSA & E. coli, MSSA & MRSA, and E. coli, MSSA & MRSA, respectively. The developed multiplex PCR assay is quick and efficient, and it can distinguish between different bacterial pathogens in a single reaction tube.

BACKGROUND: The lack of effective treatments for methicillin-resistant Staphylococcus aureus (MRSA) infection, which often leads to severe acute lung injury (ALI), poses a grave threat to human life. Sophoricoside (SOP), an isoflavone glycoside abundant in the fruit of traditional Chinese herbal Sophora japonica l., showed anti-inflammatory effects against atopic dermatitis, allergic inflammation, and lipopolysaccharide-induced ALI. However, its effect and underlying mechanism on MRSA-induced ALI remain unclear.
OBJECTIVE: The aim of this study is to assess the protective effect of SOP in MRSA-induced ALI and elucidate its underlying molecular mechanisms.
METHODS: In vivo experiments were conducted using wild-type mice to establish MRSA-induced ALI mouse model, and the effects of SOP on ALI were evaluated by hematoxylin-eosin staining, flow cytometry, quantitative real-time polymerase chain reaction, and several biochemical indicators. Adoptive transfer experiments and BTB and CNC homology 1 knockout (Bach1-/-) mice were also utilized in this study. In vitro studies employed murine macrophages RAW264.7 cells, primary bone marrow-derived macrophages (BMDMs), and primary lung macrophages to explore the underlying molecular mechanisms.
RESULTS: The administration of SOP ameliorated MRSA-induced ALI by improving pulmonary histological damages, reducing neutrophil infiltration, suppressing oxidative stress levels, and decreasing the expression of inflammatory cytokines. In isolation experiments with ALI mouse lung macrophages and macrophage adoptive transfer experiments, SOP prevented macrophage activation, thereby reducing the production of proinflammatory cytokines. In vitro experiments demonstrated that SOP decreased the expression of inflammatory mediators in lipoteichoic acid (LTA)-stimulated RAW264.7 cells, BMDMs, and primary lung macrophages. Additionally, SOP inhibited protein kinase B (Akt) phosphorylation and treatment with MK2206-a specific inhibitor of Akt-eliminated SOP\'s ability to suppress LTA-stimulated macrophage inflammation. Furthermore, stimulation with LTA or MRSA up-regulated Bach1 expression; however, deletion of Bach1 abolished the inhibitory effect of SOP on p-Akt activation as well as inflammation and ALI development.
CONCLUSIONS: This study provides the first evidence that SOP effectively mitigates MRSA-induced ALI via suppressing macrophage activation through the inhibition of Bach1/Akt pathway. These findings highlight the potential of SOP as a novel therapeutic agent for treating MRSA-induced ALI.

The proliferation and dissemination of antimicrobial-resistant bacteria is an increasingly global challenge and is attributed mainly to the excessive or improper use of antibiotics. Currently, the gold-standard phenotypic methodology for detecting resistant strains is agar plating, which is a time-consuming process that involves multiple subculturing steps. Genotypic analysis techniques are fast, but they require pure starting samples and cannot differentiate between viable and non-viable organisms. Thus, there is a need to develop a better method to identify and prevent the spread of antimicrobial resistance. This work presents a novel method for detecting and identifying antibiotic-resistant strains by combining a cell sorter for bacterial detection and an elastic-light-scattering method for bacterial classification. The cell sorter was equipped with safety mechanisms for handling pathogenic organisms and enabled precise placement of individual bacteria onto an agar plate. The patterning was performed on an antibiotic-gradient plate, where the growth of colonies in sections with high antibiotic concentrations confirmed the presence of a resistant strain. The antibiotic-gradient plate was also tested with an elastic-light-scattering device where each colony\'s unique colony scatter pattern was recorded and classified using machine learning for rapid identification of bacteria. Sorting and patterning bacteria on an antibiotic-gradient plate using a cell sorter reduced the number of subculturing steps and allowed direct qualitative binary detection of resistant strains. Elastic-light-scattering technology is a rapid, label-free, and non-destructive method that permits instantaneous classification of pathogenic strains based on the unique bacterial colony scatter pattern. KEY POINTS: • Individual bacteria cells are placed on gradient agar plates by a cell sorter • Laser-light scatter patterns are used to recognize antibiotic-resistant organisms • Scatter patterns formed by colonies correspond to AMR-associated phenotypes.

Methicillin-resistant Staphylococcus aureus (MRSA) is a current global public health problem due to its increasing resistance to the most recent antibiotic therapies. One critical approach is to develop ways to revitalize existing antibiotics. Here, we show that the phytogenic compound cinnamaldehyde (CIN) and β-lactam antibiotic combinations can functionally synergize and resensitize clinical MRSA isolates to β-lactam therapy and inhibit MRSA biofilm formation. Mechanistic studies indicated that the CIN potentiation effect on β-lactams was primarily the result of inhibition of the mecA expression by targeting the staphylococcal accessory regulator sarA. CIN alone or in combination with β-lactams decreased sarA gene expression and increased SarA protein phosphorylation that impaired SarA binding to the mecA promoter element and downregulated virulence genes such as those encoding biofilm, α-hemolysin, and adhesin. Perturbation of SarA-mecA binding thus interfered with PBP2a biosynthesis and this decreased MRSA resistance to β-lactams. Furthermore, CIN fully restored the anti-MRSA activities of β-lactam antibiotics in vivo in murine models of bacteremia and biofilm infections. Together, our results indicated that CIN acts as a β-lactam adjuvant and can be applied as an alternative therapy to combat multidrug-resistant MRSA infections.

Aim: Compare two vancomycin dosing strategies in critical patients with methicillin-resistant Staphylococcus aureus (MRSA) infections, considering the heterogeneity of the dosing regimens administered and their implications for toxicity and efficacy. Materials & methods: Longitudinal retrospective observational study in two patient cohorts (standard dosing vs dosing via Bayesian algorithms). Results: The group of Bayesian algorithms received substantially higher and significantly heterogeneous doses, with an absence of nephrotoxicity. The speed of decrease observed in CRP and PCT was greater for the Bayesian strategy (p = 0.045 and 0.0009, respectively). Conclusion: Applying Bayesian algorithms to vancomycin dosage individualization allows for administering much higher doses than with standard regimens, facilitating a quicker clinical response in the absence of nephrotoxicity.
[Box: see text].

Purpose: Cutaneous infection in epidermolysis bullosa (EB) can cause significant morbidity, mortality, and dangerous sequelae. This review article aims to delve into the known epidemiology of EB, highlight the disease\'s primary causative agents and their antimicrobial resistance spectrum.Materials and methods: A thorough literature search was conducted using Medline, EMBASE, JBI and PubMed to gather data on the microbial landscape of EB wounds. The focus was on identifying the most common bacteria associated with EB infections and assessing their antimicrobial resistance profiles.Results: The analysis revealed that Staphylococcus aureus is the most frequently identified bacterium in EB wounds, with a notable prevalence of methicillin-resistant strains (MRSA). Specific studies on mupirocin resistance further indicated rising rates of mupirocin-resistant Staphylococcus aureus, with one study reporting rates as high as 16.07%. Additionally, high resistance to other antibiotics, such as levofloxacin and trimethoprim/sulfamethoxazole, was observed in MRSA isolates.Conclusions: The findings highlight the critical need for regular resistance surveillance and the prudent use of mupirocin to manage infections effectively in EB. The multi-drug resistant nature of pathogens in EB presents a significant challenge in treatment, highlighting the importance of antimicrobial stewardship. Ultimately, given the sparse literature and the rarity of large-scale studies, further longitudinal research on the antimicrobial resistance profile of bacteria isolated from EB wounds is essential.

In this study, we investigated the genomic changes in a major methicillin-resistant Staphylococcus aureus (MRSA) clone following a significant outbreak at a hospital. Whole-genome sequencing of MRSA isolates was utilized to explore the genomic evolution of post-outbreak MRSA strains. The epidemicity of the clone declined over time, coinciding with the introduction of multimodal infection control measures. A genome-wide association study (GWAS) identified multiple genes significantly associated with either high or low epidemic success, indicating alterations in mobilome, virulence, and defense mechanisms. Random Forest models pinpointed a gene related to fibrinogen binding as the most influential predictor of epidemicity. The decline of the MRSA clone may be attributed to various factors, including the implementation of new infection control measures, single nucleotide polymorphisms accumulation, and the genetic drift of a given clone. This research underscores the complex dynamics of MRSA clones, emphasizing the multifactorial nature of their evolution. The decline in epidemicity seems linked to alterations in the clone\'s genetic profile, with a probable shift towards decreased virulence and adaptation to long-term carriage. Understanding the genomic basis for the decline of epidemic clones is crucial to develop effective strategies for their surveillance and management, as well as to gain insights into the evolutionary dynamics of pathogen genomes.

We determined antibiotic susceptibility and employed Oxford Nanopore whole-genome sequencing to explore strain diversity, resistance, and virulence gene carriage among methicillin-resistant Staphylococcus aureus (MRSA) strains from different infection sites and timepoints in a tertiary Kenyan hospital. Ninety-six nonduplicate clinical isolates recovered between 2010 and 2023, identified and tested for antibiotic susceptibility on the VITEK ID/AST platform, were sequenced. Molecular typing, antibiotic resistance, and virulence determinant screening were performed using the relevant bioinformatics tools. The strains, alongside those from previous studies, were stratified into two periods covering 2010-2017 and 2018-2023 and comparisons were made. Mirroring phenotypic profiles, aac(6\')-aph(2″) [aminoglycosides]; gyrA (S84L) and grlA (S80Y) [fluoroquinolones]; dfrG [anti-folates]; and tet(K) [tetracycline] resistance determinants dominated the collection. While the proportion of ST239/241-t037-SCCmec III among MRSA reduced from 37.7% to 0% over the investigated period, ST4803-t1476-SCCmec IV and ST152-t355-SCCmec IV were pre-eminent. The prevalence of Panton-Valentine leucocidin (PVL) and arginine catabolic mobile element (ACME) genes was 38% (33/87) and 6.8% (6/87), respectively. We observed the displacement of HA-MRSA ST239/241-t037-SCCmec III with the emergence of ST152-t355-SCCmec IV and a greater clonal heterogeneity. The occurrence of PVL+/ACME+ CA-MRSA in recent years warrants further investigations into their role in the CA-MRSA virulence landscape, in a setting of high PVL prevalence.

Silver has been shown to improve the antibiotic effects of other drugs against both Gram- positive and -negative bacteria. In this study, we investigated the antibiotic potential of cannabidiol (CBD), cannabichromene (CBC) and cannabigerol (CBG) and their acidic counterparts (CBDA, CBCA, CBGA) against Gram-positive bacteria and further explored the additive or synergistic effects of silver nitrate or silver nanoparticles using 96-well plate growth assays and viability (CFUs- colony-forming units). All six cannabinoids had strong antibiotic effects against MRSA with minimal inhibitory concentrations (MICs) of 2 mg/L for CBG, CBD and CBCA; 4 mg/L for CBGA; and 8 mg/L for CBC and CBDA. Using 96-well checkerboard assays, CBC, CBG and CBGA showed full or partial synergy with silver nitrate; CBC, CBDA and CBGA were fully synergistic with silver nanoparticles against MRSA. Using CFU assays, combinations of CBC, CBGA and CBG with either silver nitrate or silver nanoparticles, all at half or quarter MICs, demonstrated strong, time-dependent inhibition of bacterial growth (silver nitrate) and bactericidal effects (silver nanoparticles). These data will lead to further investigation into possible biomedical applications of specific cannabinoids in combination with silver salts or nanoparticles against drug-resistant Gram-positive bacteria.