BACKGROUND AND OBJECTIVE

Methicillin-resistant Staphylococcus aureus (MRSA) is one of the leading causes of hospital and community-acquired infections, showing antimicrobial resistance (AMR), which is an increasing public health concern. One of the commonly-used methods to evaluate resistance include the Kirby-Bauer disk diffusion method. However, this test is found to be time-consuming, lacking in terms of mechanization and automation, alongside its non-applicability to certain antibiotics such as vancomycin. Thus, the Clinical Laboratory Standards Institute (CLSI) recommends using the broth microdilution method in the evaluation of antibacterial activities against S. aureus. A rapid laboratory identification of MRSA is important in the treatment of patients. Therefore, this study aims to optimize and evaluate the effectiveness of a rapid microplate assay using resazurin dye as a colorimetric indicator in determining antibacterial activity against clinical isolates of MRSA and methicillin-susceptible S. aureus (MSSA).

Clinical isolates of MRSA and MSSA were obtained from the Philippine General Hospital (PGH) Microbiology Section, and American Type Culture Collection (ATCC) controls of both strains (ATCC 25923 and ATCC 43300) were acquired. These were then subjected to identification and confirmation procedures. A standardization of bacterial inoculum was performed by comparing its 24-hr growth in Mueller Hinton Broth to 0.5 McFarland Standard. The resazurin microplate assay (REMA) was set-up using two-fold serial dilution of control antibiotics such as oxacillin, vancomycin, and cefoxitin. Each plate was inoculated with standardized bacterial growth of controls and clinical isolates. To determine the time needed for the reduction of the resazurin dye, a qualitative assessment was conducted by comparing the reaction time between a 6.75 mg/mL dye with a 0.01 mg/mL dye. The plates were also subjected to different incubation times and dye concentrations, and the optical densities of the plates were compared using a microplate reader.

Results showed that there were no significant differences between the optical densities of the wells of those incubated for 5 hours and for 24 hours (p >0.05). Furthermore, there was a significant reduction in the reaction time of the dye (from 18 hours to 1 hour) when the dye concentration was reduced from 6.75 mg/mL to 0.01 mg/mL. The optimized REMA showed a significant difference between the minimum inhibitory concentrations (MICs) of the different antibiotics against the control and isolate strains of MRSA and MSSA, showing a W of -2.98 (pCONCLUSION

Based on the results presented, the researchers determined the optimal condition for the resazurin microtiter assay, which was 0.01 g/mL concentration of resazurin dye, at a 5-hour incubation period. This study has shown that an optimized REMA is an efficient and fast method to determine the antimicrobial activities of oxacillin, cefoxitin, and vancomycin against MRSA and MSSA.

Objective:Multi-drug resistant bacterial infection（MRSA） complications occurring in cochlear implant recipients is rare and of serious consequence. This paper aimed to summarize the treatment experience of a patient with MRSA infection after cochlear implantation. A patient with nasopharyngeal malignant tumor after radiotherapy developed to severe sensorineural deafness. She suffered MRSA infection nine days after cochlear implantation. Since the wound failed to heal after weeks of topical and systemic sensitive antibiotic therapy, the patient underwent surgery for wound debridement. The stimulator-receiver and the electrode of the implant was removed, negative pressure wound therapy was applied, and systemic anti-infection treatment with sensitive antibiotics for weeks, the patients recovered and was discharged from hospital 69 days after infection.
Humans ; Cochlear Implantation/adverse effects* ; Female ; Drug Resistance, Multiple, Bacterial ; Staphylococcal Infections/therapy* ; Methicillin-Resistant Staphylococcus aureus ; Cochlear Implants ; Anti-Bacterial Agents/therapeutic use* ; Postoperative Complications ; Middle Aged

Atopic dermatitis (AD) is a prevalent inflammatory skin disorder in which patients experience recurrent eczematous lesions and intense itching. The colonization of Staphylococcus aureus (S. aureus) is correlated with the severity of the disease, but its role in AD development remains elusive. Using single-cell RNA sequencing, we uncovered that keratinocytes activate a distinct immune response characterized by induction of Il24 when exposed to methicillin-resistant S. aureus (MRSA). Further experiments using animal models showed that the administration of recombinant IL-24 protein worsened AD-like pathology. Genetic ablation of Il24 or the receptor Il20rb in keratinocytes alleviated allergic inflammation and atopic march. Mechanistically, IL-24 acted through its heterodimeric receptors on keratinocytes and augmented the production of IL-33, which in turn aggravated type 2 immunity and AD-like skin conditions. Overall, these findings establish IL-24 as a critical factor for onset and progression of AD and a compelling therapeutic target.
Dermatitis, Atopic/genetics* ; Interleukins/metabolism* ; Animals ; Methicillin-Resistant Staphylococcus aureus/immunology* ; Mice ; Keratinocytes/microbiology* ; Humans ; Interleukin-33/immunology* ; Inflammation/microbiology* ; Staphylococcal Infections/microbiology* ; Disease Models, Animal ; Hypersensitivity/microbiology* ; Mice, Inbred C57BL

Given the increasing concern regarding antibacterial resistance, the antimicrobial properties of naphthoquinones have recently attracted significant attention. While 1,4-naphthoquinone and its derivatives have been extensively studied, the antibacterial properties of 5,8-dihydroxy-1,4-naphthoquinone derivatives remain relatively unexplored. This study presents a comprehensive in vitro and in vivo analysis of the antibacterial activity of 35 naturally sourced and chemically synthesized derivatives of 5,8-dihydroxy-1,4-naphthoquinone. Kirby-Bauer antibiotic testing identified three compounds with activity against methicillin-resistant Staphylococcus aureus (MRSA), with one compound (PNP-02) demonstrating activity comparable to vancomycin in minimum inhibitory concentration, minimum bactericidal concentration (MBC), and time-kill assays. Microscopic and biochemical analyses revealed that PNP-02 adversely affects the cell wall and cell membrane of MRSA. Mechanistic investigations, including proteomic sequencing analyses, Western blotting, and RT-qPCR assays, indicated that PNP-02 compromises cell membrane integrity by inhibiting arginine biosynthesis and pyrimidine metabolism pathways, thereby increasing membrane permeability and inducing bacterial death. In an in vivo mouse model of skin wound healing, PNP-02 exhibited antibacterial efficacy similar to vancomycin. The compound demonstrated low toxicity to cultured human cells and in hemolysis assays and remained stable during serum incubation. These findings suggest that PNP-02 possesses promising bioactivity against MRSA and represents a potential novel antibacterial agent.
Methicillin-Resistant Staphylococcus aureus/genetics* ; Anti-Bacterial Agents/chemistry* ; Naphthoquinones/administration & dosage* ; Animals ; Microbial Sensitivity Tests ; Mice ; Humans ; Staphylococcal Infections/microbiology* ; Molecular Structure

Objective: To determine the point prevalence of, and risk factors associated with MRSA carriage among resident physicians of surgical departments at the Ospital ng Maynila Medical Center. Methods: Design: Cross-sectional Study. Setting: Tertiary Government Training Hospital. Participants:51 resident physicians from different surgical departments (general surgery, obstetrics and gynecology, ophthalmology, otorhinolaryngology – head and neck surgery and dermatology) underwent nasal and pharyngeal swabs with microbial culture and sensitivity testing to identify MRSA carriers. Fisher Exact Test and logistic regression were utilized to determine associations between MRSA carriage and various risk factors including frequency of hand washing and departmental affiliation. Results: Overall prevalence rate of MRSA carriage was 9.8%. Otorhinolaryngology residents had the highest combined prevalence of MRSA of 42.9%, significantly higher compared to other departments and were used as a reference in logistic regression analyses. Notably, handwashing only once daily was associated with a 20-fold increase in the risk of MRSA carriage (OR 20.5, 95% CI: 1.82 to 230, p = .014). Other departments did not demonstrate statistically significant differences in MRSA carriage rates. Conclusions Otorhinolaryngology resident physicians had the highest combined prevalence of MRSA and nasal MRSA was found only in otorhinolaryngology residents. The surgical subspecialty and frequency of handwashing of the healthcare worker were identified as important risk factors to develop MRSA carriage. Targeted interventions (including enhanced infection control protocols and regular screening) are needed especially in high-risk departments.
Methicillin-Resistant Staphylococcus aureus ; Surgical Wound Infection

Background and Objective: Methicillin-resistant Staphylococcus aureus (MRSA) is one of the leading causes of hospital and community-acquired infections, showing antimicrobial resistance (AMR), which is an increasing public health concern. One of the commonly-used methods to evaluate resistance include the Kirby-Bauer disk diffusion method. However, this test is found to be time-consuming, lacking in terms of mechanization and automation, alongside its non-applicability to certain antibiotics such as vancomycin. Thus, the Clinical Laboratory Standards Institute (CLSI) recommends using the broth microdilution method in the evaluation of antibacterial activities against S. aureus. A rapid laboratory identification of MRSA is important in the treatment of patients. Therefore, this study aims to optimize and evaluate the effectiveness of a rapid microplate assay using resazurin dye as a colorimetric indicator in determining antibacterial activity against clinical isolates of MRSA and methicillin-susceptible S. aureus (MSSA). Methods: Clinical isolates of MRSA and MSSA were obtained from the Philippine General Hospital (PGH) Microbiology Section, and American Type Culture Collection (ATCC) controls of both strains (ATCC 25923 and ATCC 43300) were acquired. These were then subjected to identification and confirmation procedures. A standardization of bacterial inoculum was performed by comparing its 24-hr growth in Mueller Hinton Broth to 0.5 McFarland Standard. The resazurin microplate assay (REMA) was set-up using two-fold serial dilution of control antibiotics such as oxacillin, vancomycin, and cefoxitin. Each plate was inoculated with standardized bacterial growth of controls and clinical isolates. To determine the time needed for the reduction of the resazurin dye, a qualitative assessment was conducted by comparing the reaction time between a 6.75 mg/mL dye with a 0.01 mg/mL dye. The plates were also subjected to different incubation times and dye concentrations, and the optical densities of the plates were compared using a microplate reader. Results: Results showed that there were no significant differences between the optical densities of the wells of those incubated for 5 hours and for 24 hours (p >0.05). Furthermore, there was a significant reduction in the reaction time of the dye (from 18 hours to 1 hour) when the dye concentration was reduced from 6.75 mg/mL to 0.01 mg/mL. The optimized REMA showed a significant difference between the minimum inhibitory concentrations (MICs) of the different antibiotics against the control and isolate strains of MRSA and MSSA, showing a W of -2.98 (p <0.05) using the Wilcoxon Rank-Sum non-parametric test. Furthermore, the REMA has shown better illustration of anti-MRSA and anti-MSSA activities as compared to the Kirby Bauer disk diffusion method. Conclusion Based on the results presented, the researchers determined the optimal condition for the resazurin microtiter assay, which was 0.01 g/mL concentration of resazurin dye, at a 5-hour incubation period. This study has shown that an optimized REMA is an efficient and fast method to determine the antimicrobial activities of oxacillin, cefoxitin, and vancomycin against MRSA and MSSA.
Methicillin Resistant Staphylococcus aureus

Background: Ardisia serrata (Aunasin) is an endemic Philippine plant of the family Primulaceae, with several studiesshowing the genus Ardisia as having potential antibacterial, antiangiogenic, cytotoxic, and antipyretic properties. Objective: This study aims to determine the antibacterial and antibiofilm-forming activity of Ardisia serrata ethanolic and aqueous extracts on Escherichia coli, Methicillin-Sensitive Staphylococcus aureus (MSSA), and Methicillin-Resistant Staphylococcus aureus (MRSA). Methods: This is an experimental study testing the activity against bacterial strains of E. coli, MSSA, and MRSA using ethanolic and aqueous extracts of A. serrata leaves. Microtiter susceptibility and biofilm inhibition assays were done with two-fold dilutions of the extract against the selected strains using spectrophotometry with optical density (OD) at 600 nm and 595 nm, respectively, to quantify bacterial growth and biofilm inhibition. The bacterial susceptibility and biofilm inhibition activity was reported as percent inhibition (PI). Minimum inhibitory concentration (MIC), and minimum biofilm inhibition concentration (MBIC) values were obtained using logarithmic regression of the PI values. Results: A. serrata ethanolic extracts showed weak growth inhibitory activity against MSSA and MRSA with minimum inhibitory concentration (MIC) values of 2.6192 and 3.2988 mg/mL, respectively, but no biofilm inhibition activity was noted, while the aqueous extracts exhibited negligible biofilm inhibition activity against MSSA and MRSA with minimum biofilm inhibition concentration (MBIC) values of 13.5972 and 8964.82 mg/mL, respectively, and with no growth inhibition activity. Both ethanolic and aqueous extracts showed no growth inhibition and biofilm inhibition activities against E. coli. Conclusion Staphylococcus aureus is susceptible to the bioactivity of the leaf extracts of A. serrata and has potential to be used as an antibacterial in the treatment of infectious diseases.
Methicillin-resistant Staphylococcus aureus ; Escherichia coli ; natural product ; biological products

Objective: To understand the population structure of food-borne Staphylococcus (S.) aureus in China. Methods: Whole genome sequencing was used to analyze 763 food-borne S. aureus strains from 16 provinces in China from 2006 to 2020. Multilocus sequence typing (MLST), staphylococcal protein A gene (spa) typing, and staphylococcal chromosome cassettemec (SCCmec) typing were conducted, and minimum spanning tree based on ST types (STs) was constructed by BioNumerics 7.5 software. Thirty-one S. aureus strains isolated from imported food products were also included in constructing the genome phylogenetic tree. Results: A total of 90 STs (20 novel types) and 160 spa types were detected in the 763 S. aureus isolates. The 72 STs (72/90, 80.0%) were related to 22 clone complexes. The predominant clone complexes were CC7, CC1, CC5, CC398, CC188, CC59, CC6, CC88, CC15, and CC25, accounting for 82.44% (629/763) of the total. The STs and spa types in the predominant clone complexes changed over the years. The methicillin-resistant S. aureus (MRSA) detection rate was 7.60%, and 7 SCCmec types were identified. The ST59-t437-Ⅳa (17.24%, 10/58), ST239-t030-Ⅲ (12.07%, 7/58), ST59-t437-Ⅴb (8.62%, 5/58), ST338-t437-Ⅴb (6.90%, 4/58) and ST338-t441-Ⅴb (6.90%, 4/58) were the main types in MRSA strains. The genome phylogenetic tree had two clades, and the strains with the same CC, ST, and spa types clustered together. All CC7 methicillin sensitive S. aureus strains were included in Clade1, while 21 clone complexes and all MRSA strains were in Clade2. The MRSA strains clustered according to the SCCmec and STs. The strains from imported food products in CC398, CC7, CC30, CC12, and CC188 had far distances from Chinese strains in the tree. Conclusions: In this study, the predominant clone complexes of food-borne strains were CC7, CC1, CC5, CC398, CC188, CC59, CC6, CC88, CC15, and CC25, which overlapped with the previously reported clone complexes of hospital and community-associated strains in China, suggesting that close attention needs to be paid to food, a vehicle of pathogen transmission in community and food poisoning.
Humans ; Staphylococcus aureus/genetics* ; Methicillin-Resistant Staphylococcus aureus/genetics* ; Multilocus Sequence Typing ; Phylogeny ; Staphylococcal Infections/epidemiology* ; China/epidemiology*

OBJECTIVE: To analyze the distribution and drug resistance of pathogens in oral mucositis associated with chemotherapy in hospitalized patients with malignant hematopathy, so as to provide scientific evidences for rational selection of antibiotics and infection prevention and control. METHODS: From July 2020 to June 2022, 167 patients with malignant hematopathy were treated with chemical drugs in the Department of Hematology, Hainan Hospital, and secretions from oral mucosal infected wounds were collected. VITEK2 COMPECT automatic microbial identification system (BioMerieux, France) and bacterial susceptibility card (BioMerieux) were used for bacterial identification and drug susceptibility tests. RESULTS: A total of 352 strains of pathogens were isolated from 167 patients, among which 220 strains of Gram-positive bacteria, 118 strains of Gram-negative bacteria and 14 strains of fungi, accounted for 62.50%, 33.52% and 3.98%, respectively. The Gram-positive bacteria was mainly Staphylococcus and Streptococcus, while Gram-negative bacteria was mainly Klebsiella and Proteus. The resistance of main Gram-positive bacteria to vancomycin, ciprofloxacin and gentamicin was low, and the resistance to penicillin, cefuroxime, ampicillin, cefotaxime, erythromycin and levofloxacin was high. The main Gram-negative bacteria had low resistance to gentamicin, imipenem and penicillin, but high resistance to levofloxacin, cefotaxime, cefuroxime, ampicillin and vancomycin. The clinical data of oral mucositis patients with oral ulcer (severe) and without oral ulcer (mild) were compared, and it was found that there were statistically significant differences in poor oral hygiene, diabetes, sleep duration less than 8 hours per night between two groups (P<0.05). CONCLUSION Gram-positive bacteria is the main pathogen of oral mucositis in patients with malignant hematopathy after chemotherapy. It is sensitive to glycopeptide antibiotics and aminoglycosides antibiotics. Poor oral hygiene, diabetes and sleep duration less than 8 hours per night are risk factors for oral mucositis with oral ulcer (severe).
Humans ; Vancomycin/therapeutic use* ; Cefuroxime ; Levofloxacin ; Oral Ulcer/drug therapy* ; Drug Resistance, Bacterial ; Anti-Bacterial Agents/adverse effects* ; Ampicillin ; Penicillins ; Cefotaxime ; Gram-Positive Bacteria ; Gram-Negative Bacteria ; Gentamicins ; Stomatitis/drug therapy*

Objective: To explore the impact of traditional Chinese medicine berberine (BBR) on membrane integrity and permeability of Methicillin-resistant Staphylococcus aureus (MRSA) and the change of bacterial cell wall structure, laying a foundation for the clinical application of berberine in antibacterial. Methods: This study used a non-randomized concurrent controlled trial. The 3 MRSA strains were isolated and cultured from lower respiratory tract samples of geriatric patients from Shanghai Eighth People's Hospital between 2019 and 2020.The Meirier VETEK MS fully automated rapid microbial mass spectrometry detection system and VETEK 2 Compact fully automated microbial identification instrument were used to identify bacterial drug sensitivity experiments to detect bacterial species and drug sensitivity. The minimal inhibitory concentration (MIC) of BBR on MRSA strains was determined by broth microdilution. This study used conductivity tests to assess the changes in membrane permeability in response to different concentration of BBR on MRSA, while also investigating the changes in MRSA morphology by transmission electron microscopy. GraphPad Prism5 was used to analyze the differences in the electrical conductivity experimental results. Results: The MIC of BBR on MRSA was 64 μg/ml. After co-culturing MRSA with BBR for 4 h at 8 μg/ml, 16 μg/ml, 32 μg/ml, 64 μg/ml and 128 μg/ml, respectively, the electrical conductivity increased, compared with the control group, by 24.49%,34.59%,208.92%,196.40% and 208.68%, respectively. By transmission electron microscopy, This study found that low concentration of BBR (8 μg/ml,1/8 MIC) caused no significant damage to MRSA, and the bacterial structure of MRSA remained intact. The cell wall of MRSA became thinner after treatment with berberine at medium concentration (64 μg/ml,1 MIC), while high concentration of BBR (512 μg/ml,8 MIC) induced the destruction and dissolution of MRSA cell wall structure and the leakage of bacterial contents, leading to bacterial lysis. Conclusion: Berberine can kill bacteria by altering the permeability of MRSA cell membrane and destroying and dissolving the structure of the cell wall.
Methicillin-Resistant Staphylococcus aureus ; Berberine/pharmacology* ; China ; Anti-Bacterial Agents/pharmacology* ; Cell Membrane ; Microbial Sensitivity Tests