Methicillin-resistant Staphylococcus Aureus Cells Research Articles

Micrococcal nuclease regulates biofilm formation and dispersal in methicillin-resistant Staphylococcus aureus USA300.

Biofilm formation is an important virulence factor for methicillin-resistant Staphylococcus aureus (MRSA). The extracellular matrix of MRSA biofilms contains significant amounts of double-stranded DNA that hold the biofilm together. MRSA cells secrete micrococcal nuclease (Nuc1), which degrades double-stranded DNA. In this study, we used standard methodologies to investigate the role of Nuc1 in MRSA biofilm formation and dispersal. We quantified biofilm formation and extracellular DNA (eDNA) levels in broth and agar cultures. In some experiments, cultures were supplemented with sub-MIC amoxicillin to induce biofilm formation. Biofilm erosion was quantitated by culturing biofilms on rods and enumerating detached colony-forming units (CFUs), and biofilm sloughing was investigated by perfusing biofilms cultured in glass tubes with fresh broth and measuring the sizes of the detached cell aggregates. We found that an MRSA nuc1- mutant strain produced significantly more biofilm and more eDNA than a wild-type strain, both in the absence and presence of sub-MIC amoxicillin. nuc1- mutant biofilms grown on rods detached significantly less than wild-type biofilms. Detachment was restored by exogenous DNase or complementing the nuc1- mutant. In the sloughing assay, nuc1- mutant biofilms released cell aggregates that were significantly larger than those released by wild-type biofilms. Our results suggest that Nuc1 modulates biofilm formation, biofilm detachment, and the sizes of detached cell aggregates. These processes may play a role in the spread and subsequent survival of MRSA biofilms during biofilm-related infections.IMPORTANCEInfections caused by antibiotic-resistant bacteria known as methicillin-resistant Staphylococcus aureus (MRSA) are a significant problem in hospitals. MRSA forms adherent biofilms on implanted medical devices such as catheters and breathing tubes. Bacteria can detach from biofilms on these devices and spread to other parts of the body such as the blood or lungs, where they can cause life-threatening infections. In this article, researchers show that MRSA secretes an enzyme known as thermonuclease that causes bacteria to detach from the biofilm. This is important because understanding the mechanism by which MRSA detaches from biofilms could lead to the development of procedures to mitigate the problem.

The Spx stress regulator confers high-level β-lactam resistance and decreases susceptibility to last-line antibiotics in methicillin-resistant Staphylococcus aureus.

Infections caused by methicillin-resistant Staphylococcus aureus (MRSA) are a leading cause of mortality worldwide. MRSA has acquired resistance to next-generation β-lactam antibiotics through the horizontal acquisition of the mecA resistance gene. Development of high resistance is, however, often associated with additional mutations in a set of chromosomal core genes, known as potentiators, which, through poorly described mechanisms, enhance resistance. The yjbH gene was recently identified as a hot spot for adaptive mutations during severe infections. Here, we show that inactivation of yjbH increased β-lactam MICs up to 16-fold and transformed MRSA cells with low levels of resistance to being homogenously highly resistant to β-lactams. The yjbH gene encodes an adaptor protein that targets the transcriptional stress regulator Spx for degradation by the ClpXP protease. Using CRISPR interference (CRISPRi) to knock down spx transcription, we unambiguously linked hyper-resistance to the accumulation of Spx. Spx was previously proposed to be essential; however, our data suggest that Spx is dispensable for growth at 37°C but becomes essential in the presence of antibiotics with various targets. On the other hand, high Spx levels bypassed the role of PBP4 in β-lactam resistance and broadly decreased MRSA susceptibility to compounds targeting the cell wall or the cell membrane, including vancomycin, daptomycin, and nisin. Strikingly, Spx potentiated resistance independently of its redox-sensing switch. Collectively, our study identifies a general stress pathway that, in addition to promoting the development of high-level, broad-spectrum β-lactam resistance, also decreases MRSA susceptibility to critical antibiotics of last resort.

Sinapic acid attenuates biofilm formation and virulence factor production in methicillin resistant Staphylococcus aureus (MRSA) isolated from raw camel milk: An in vitro and in silico approach

Camel milk is highly nutritious but contamination with biofilm forming methicillin resistant Staphylococcus aureus (MRSA) may pose a serious threat to human health and food safety. Therefore, sinapic acid (SA) was assessed for its in vitro antibiofilm and antivirulence potential against MRSA isolated from raw camel milk samples. SA reduced biofilm formation and production of key virulence functions (exopolysaccharide production, cell surface hydrophobicity and staphyloxanthin production) significantly. Pre-formed biofilms were also reduced by 38–55%. Enhanced reactive oxygen species (ROS) production in MRSA cells treated with SA could plausibly be responsible for cell death and reduced biofilm formation. In silico studies demonstrated that the overall binding energy for SA interaction with AgrA and SarA was −9.882 and −15.076 kcal mol−1, respectively. It is envisaged that SA could be exploited to mitigate biofilm formation in ensuring the safety and quality of food/dairy products and minimizing the risk of MRSA transmission through the food supply chain.

Peptide recovery from chicken feather keratin and their anti-biofilm properties against methicillin-resistant Staphylococcus aureus (MRSA).

Bacteria have the potential to adhere to abiotic surfaces, which has an undesirable effect in the food industry because they can survive for sustained periods through biofilm formation. In this study, an antibacterial peptide (ABP), with a molecular mass of 3861 Da, was purified from hydrolyzed chicken feathers using a locally isolated keratinolytic bacterium, namely Rhodococcus erythropolis, and its antibacterial and antibiofilm potential were investigated against planktonic and biofilm cells of Methicillin-Resistant Staphylococcus Aureus (MRSA). The results demonstrated that purified ABP showed the growth inhibition of MRSA cells with the minimum inhibitory concentration (MIC) of 45µg/ml and disrupted MRSA biofilm formation at a concentration of 200 ug/ml, which results were confirmed by scanning electron micrograph (SEM). Moreover, the secondary structures of the peptide were assessed as part of the FTIR analysis to evaluate its mode of action. ExPASy tools were used to predict the ABP sequence, EPCVQUQDSRVVIQPSPVVVVTLPGPILSSFPQNTA, from a chicken feather keratin sequence database following in silico digestion by trypsin. Also, ABP had 54.29% hydrophobic amino acids, potentially contributing to its antimicrobial activity. The findings of toxicity prediction of the peptide by the ToxinPred tool revealed that ABP had non-toxic effects. Thus, these results support the potential of this peptide to be used as an antimicrobial agent for the treatment or prevention of MRSA biofilm formation in feed, food, or pharmaceutical applications.

Antimicrobial activity of a natural compound and analogs against multi-drug-resistant Gram-positive pathogens.

The rapid emergence of methicillin-resistant Staphylococcus aureus (MRSA) isolates has precipitated a critical need for novel antibiotics. We have developed a one-pot synthesis method for naturally occurring compounds such as MC21-A (C58) and its chloro-analog, C59. Our findings demonstrate that these compounds kill MRSA isolates at lower or comparable concentrations to standard-of-care (SoC) antimicrobials. C59 eradicates MRSA cells in biofilms, which are notoriously difficult to treat with SoC antibiotics. Additionally, the lack of resistance development observed with C59 treatment is a significant advantage when compared to currently available antibiotics. Furthermore, these compounds are non-toxic to mammalian cell lines at effective concentrations. Our findings indicate the potential of these compounds to treat MRSA infections and underscore the importance of exploring natural products for novel antibiotics. Further investigation will be essential to fully realize the therapeutic potential of these next-generation antimicrobials to address the critical issue of antimicrobial resistance.

Enhanced Photocatalytic Properties of All-Organic IDT-COOH/O-CN S-Scheme Heterojunctions Through π-π Interaction and Internal Electric Field.

Herein, we present a distinct methodology for the in situ electrostatic assembly method for synthesizing a conjugated (IDT-COOH)/oxygen-doped g-C3N4 (O-CN) S-scheme heterojunction. The electron delocalization effect due to π-π interactions between O-CN and self-assembled IDT-COOH favors interfacial charge separation. The self-assembled IDT-COOH/O-CN exhibits a broadened visible absorption to generate more charge carriers. The internal electric field between the IDT-COOH and the O-CN interface provides a directional charge-transfer channel to increase the utilization of photoinduced charge carriers. Moreover, the active species (•O2-, h+, and 1O2) produced by IDT-COOH/O-CN under visible light play important roles in photocatalytic disinfection. The optimum 40% IDT-COOH/O-CN can kill 7-log of methicillin-resistant Staphylococcus aureus (MRSA) cells in 2 h and remove 88% tetracycline (TC) in 5 h, while O-CN only inactivates 1-log of MRSA cells and degrades 40% TC. This work contributes to a promising method to fabricate all-organic g-C3N4-based S-scheme heterojunction photocatalysts with a wide range of optical responses and enhanced exciton dissociation.

Transcriptomic analysis of cell envelope inhibition by prodigiosin in methicillin-resistant Staphylococcus aureus.

Methicillin-resistant Staphylococcus aureus (MRSA) is a leading threat to public health as it is resistant to most currently available antibiotics. Prodigiosin is a secondary metabolite of microorganisms with broad-spectrum antibacterial activity. This study identified a significant antibacterial effect of prodigiosin against MRSA with a minimum inhibitory concentration as low as 2.5 mg/L. The results of scanning electron microscopy, crystal violet staining, and confocal laser scanning microscopy indicated that prodigiosin inhibited biofilm formation in S. aureus USA300, while also destroying the structure of the cell wall and cell membrane, which was confirmed by transmission electron microscopy. At a prodigiosin concentration of 1.25 mg/L, biofilm formation was inhibited by 76.24%, while 2.5 mg/L prodigiosin significantly reduced the vitality of MRSA cells in the biofilm. Furthermore, the transcriptomic results obtained at 1/8 MIC of prodigiosin indicated that 235and 387 genes of S. aureus USA300 were significantly up- and downregulated, respectively. The downregulated genes were related to two-component systems, including the transcriptional regulator LytS, quorum sensing histidine kinases SrrB, NreA and NreB, peptidoglycan biosynthesis enzymes (MurQ and GlmU), iron-sulfur cluster repair protein ScdA, microbial surface components recognizing adaptive matrix molecules, as well as the key arginine synthesis enzymes ArcC and ArgF. The upregulated genes were mainly related to cell wall biosynthesis, as well as two-component systems including vancomycin resistance-associated regulator, lipoteichoic acid biosynthesis related proteins DltD and DltB, as well as the 9 capsular polysaccharide biosynthesis proteins. This study elucidated the molecular mechanisms through which prodigiosin affects the cell envelope of MRSA from the perspectives of cell wall synthesis, cell membrane and biofilm formation, providing new potential targets for the development of antimicrobials for the treatment of MRSA.

Ultrasound-assisted activation of PAW residual radicals in the concurrent elimination of ARB and ARGs: Process efficiency, mechanism and implication

Methicillin-resistant Staphylococcus aureus (MRSA) and antibiotic resistance gene (mecA) are considered emerging contaminants that have been extensively identified in aqueous environments due to their high resistance to disinfectants. Herein, we report for the first time the inactivation of MRSA through a sequential application of plasma-activated water and ultrasound (PAW-SU). While the impact of PAW alone on bacterial culturability and mecA integrity was negligible, the combined PAW-SU treatment outperformed the inactivation method involving PAW with direct US (PAW-DU). It exhibited a shorter lag phase and a higher maximum inactivation rate constant. We numerically demonstrated that sonolysis processes induced by external ultrasound sources considerably improved the intracellular and extracellular reactive oxygen and nitrogen species (RONS) of MRSA in PAW system, thereby increasing the PAW reactivity and applicability. The highest inactivation efficiency of PAW-SU can be attributed to the synergistic effect of enhanced intracellular RONS (i.e., peroxynitrite and singlet oxygen) and membrane disruption induced by US, resulting in irreparable oxidative damage to the Save our Soul response and energy metabolic system. In particular, the elevated arginine metabolites in MRSA cells were primarily responsible for the formation of the viable but nonculturable state (VBNC) bacteria associated with high resistance to PAW system. Surprisingly, a decrease in arginine synthesis capacity was observed to expedite the flow of the tricarboxylic acid cycle, accelerating the detrimental effects of PAW-SU on intracellular components in VBNC bacteria. Finally, the limited regrowth potential of injured MRSA caused by cyclic inactivation of PAW-SU process was further confirmed in wastewater from meat and dairy models and therefore, advocates the possible real-world applications.

COMBATTING METHICILLIN-RESISTANT STAPHYLOCOCCUS AUREUS (MRSA) IN THE FOOD INDUSTRY BY HARNESSING THE POWER OF NATURE: A SYSTEMATIC REVIEW

Antibiotic resistance is a critical global health concern, with Methicillin-resistant Staphylococcus aureus (MRSA) posing a significant challenge due to its resistance to commonly used antibiotics. Recent research has revealed the potential of natural compounds and microorganisms in combatting MRSA and other antibiotic-resistant bacteria. In this systematic review, we studied the effect of essential oils, bacteriophages, bacteriocins, and probiotics on S. aureus, including MRSA in particular, in the food industry. Essential oils (EOs) have gained significant attention because of their antimicrobial properties, inhibiting MRSA growth by damaging bacterial cells and inhibiting essential enzymes and compounds. Cinnamon oil liposomes caused the most significant decrease in MRSA populations among our reviewed essential oils. Bacteriophages can lyse the bacterial host. They encode peptidoglycan hydrolases called endolysins that target the bacterial cell wall. In our study, S. aureus phage (containing CHAPLysGH15 and LysGH15), and phage SA11 endolysin (LysSA11) were the most effective against S. aureus. Bacteriocins, antimicrobial peptides produced by bacteria, also show potential in combatting MRSA, mainly by generating organic acids that interfere with bacterial metabolism. According to our review, the most effective bacteriocins against S. aureus were Enterocin AS-48 with phenolic compounds or with 2NPOH, Bacteriocin isolated from Lactobacillus pentosus - Pentocin JL-1, and bacteriocin produced by S. pasteuri RSP-1, respectively. Probiotics can compete with pathogens by producing antimicrobial compounds that disrupt MRSA cell production and ultimately lead to bacterial death. In our review, the most effective probiotics were Streptomyces griseus, Pediococcus acidilactici strains A11 and C12, Lactococcus lactis, and Lactobionic acid respectively. A multi-hurdle approach combining these natural agents has shown promising results in targeting and eliminating MRSA cells. By harnessing the power of nature, we can potentially overcome the challenges posed by MRSA and other antibiotic-resistant bacteria.

Synergistic Potential of α-Melanocyte Stimulating Hormone Based Analogues with Conventional Antibiotic against Planktonic, Biofilm-Embedded, and Systemic Infection Model of MRSA.

The repotentiation of the existing antibiotics by exploiting the combinatorial potential of antimicrobial peptides (AMPs) with them is a promising approach to address the challenges of slow antibiotic development and rising antimicrobial resistance. In the current study, we explored the ability of lead second generation Ana-peptides viz. Ana-9 and Ana-10, derived from Alpha-Melanocyte Stimulating Hormone (α-MSH), to act synergistically with different classes of conventional antibiotics against methicillin-resistant Staphylococcus aureus (MRSA). The peptides exhibited prominent synergy with β-lactam antibiotics, namely, oxacillin, ampicillin, and cephalothin, against planktonic MRSA. Furthermore, the lead combination of Ana-9/Ana-10 with oxacillin provided synergistic activity against clinical MRSA isolates. Though the treatment of MRSA is complicated by biofilms, the lead combinations successfully inhibited biofilm formation and also demonstrated biofilm disruption potential. Encouragingly, the peptides alone and in combination were able to elicit in vivo anti-MRSA activity and reduce the bacterial load in the liver and kidney of immune-compromised mice. Importantly, the presence of Ana-peptides at sub-MIC doses slowed the resistance development against oxacillin in MRSA cells. Thus, this study highlights the synergistic activity of Ana-peptides with oxacillin advocating for the potential of Ana-peptides as an alternative therapeutic and could pave the way for the reintroduction of less potent conventional antibiotics into clinical use against MRSA infections.

Resensitisation of Methicillin-Resistant Staphylococcus aureus to Conventional Antibiotics in the Presence of an Engineered Enzybiotic.

Methicillin-resistant Staphylococcus aureus (MRSA) is one of the most dreadful pathogens relevant in community and nosocomial-related infections around the world. Resensitising MRSA to antibiotics, once it became resistant, was a tough choice due to the high adaptability of this bacteria to savage conditions. This study aimed to create a chimeric enzybiotic against MRSA and test its efficiency, either individually or in combination with antibiotics. The novel enzybiotic BAC100 was constructed by fusing the catalytic domain from the bacteriocin BacL1 from Enterococcus faecalis with the cell-wall-binding domain from protein P17 of Staphylococcus aureus bacteriophage ϕ44AHJD. Apart from its partial lone activity, BAC100 was found to resensitise the MRSA strain to traditional antibiotics, including ampicillin and tetracycline. Both drugs were able to reduce live MRSA cells by 85 and 90%, respectively, within 60 min of treatment together with BAC100. However, no significant activity was observed against MRSA when these drugs were tested independently, pointing to the inherent resistance of MRSA against these conventional antibiotics. To our knowledge, this is one of the first instances where an engineered enzybiotic was found to resensitise MRSA to conventional antibiotics. This study will pave the way for the development of similar peptides that can be used together with antibiotics against gruesome pathogens of clinical importance.

Ultrasensitive detection of methicillin-resistant Staphylococcus aureus using a T7 exonuclease-assisted PAM-free dual CRISPR-Cas12a biosensor

Methicillin-resistant Staphylococcus aureus (MRSA) strains are prevalent foodborne superbugs and pose significant pathogens for both humans and animals. The sensitive and specific diagnosis of MRSA holds paramount importance for its prevention and treatment. Herein, we develop a T7 exonuclease-assisted PAM-free Dual CRISPR-Cas12a Biosensor (TDCB) targeting the antibiotic resistance gene MecA for ultrasensitive and highly specific MRSA detection. After generation of single-stranded DNA (ssDNA) obtained with T7 exonuclease digestion of amplified MRSA genome, the trans-cleavage activity of dual-Cas12a-crRNA complexes was initiated without the requirement of protospacer adjacent motifs (PAM). The fluorescent signal obtained with TDCB biosensor was stronger, compared with the single-Cas12a biosensor. TDCB detects the MRSA genome and cells with a high sensitivity of 100 ag/mL and 1 cfu/mL, respectively. Moreover, TDCB can distinguish MRSA from tested 6 non-MRSA bacterial species from each other. We further validated TDCB using MRSA-infected tilapia fishes and obtained results consistent with the qPCR assay. This TDCB technology, a novel Cas12a-based biosensor that detects genomic regions without reliance on PAM motifs, can have very broad utilization in hospitals, food and feed industry, and fish and dairy farming.

Inactivation action of ultrasound-assisted cinnamaldehyde on planktonic and biofilm methicillin-resistant Staphylococcus aureus and its application in beef system

There has been a growing concern for methicillin-resistant Staphylococcus aureus (MRSA) contamination in the food industry. In the present study, the highest inactivation effect (5.80 log CFU/mL) was observed in simultaneous treatment compared with sequential treatment of ultrasound (US)-assisted cinnamaldehyde (CA). The absence of sublethal bacteria indicated US-assisted CA did not induce the tolerance or adaptation to MRSA cells. The results of SEM analysis indicated that the application of US-assisted CA resulted in severe damage to the cell membrane, as demonstrated by the highest fluorescence intensity of propidium iodide dye, intracellular substance leakage, and molecular docking analysis, as well as ATPase activity. Furthermore, US-assisted CA significantly disrupted energy metabolism, as evidenced by a reduction in ATP concentration, succinate dehydrogenase, and malate dehydrogenase activities. The exacerbation of ROS-induced damage to MRSA cells was observed due to the reduction in catalase and peroxidase activities. Additionally, a significant decrease (p < 0.05) in extracellular carbohydrates and proteins within the MRSA biofilm was noted on both glass and stainless-steel surfaces exposed to US-assisted CA. Ultimately, US-assisted CA treatment resulted in a maximum reduction (p < 0.05) of MRSA cells in beef meat, while maintaining color, pH, and sensory properties during an 8-day storage period at 4 °C. These findings suggested that simultaneous US-assisted CA treatment may serve as an alternative method for ensuring microbiological safety.

The depsidones from marine sponge-derived fungus Aspergillus unguis IB151 as an anti-MRSA agent: Molecular docking, pharmacokinetics analysis, and molecular dynamic simulation studies

Methicillin-resistant Staphylococcus aureus (MRSA) is an emerging nosocomial pathogen among hospitalized patients, with high morbidity and mortality rates. The discovery of a novel antibacterial is urgently needed to address this resistance problem. The present study aims to explore the antibacterial potential of three depsidone compounds: 2-clorounguinol (1), unguinol (2), and nidulin (3), isolated from the marine sponge-derived fungus Aspergillus unguis IB1, both in vitro and in silico. The antibacterial activity of all compounds was evaluated by calculating the Minimum inhibitory concentration (MIC) and Minimum bactericidal concentration (MBC) against MRSA using agar diffusion and total plate count methods, respectively. Bacterial cell morphology changes were studied for the first time using scanning electron microscopy (SEM). Molecular docking, pharmacokinetics analysis, and molecular dynamics simulation were performed to determine possible protein–ligand interactions and the stability of the targeting penicillin-binding protein 2a (PBP2a) against 2-clorounguinol (1). The research findings indicated that compounds 1 to 3 exhibited MIC and MBC values of 2 µg/mL and 16 µg/mL against MRSA, respectively. MRSA cells displayed a distinct shape after the addition of the depsidone compound, as observed in SEM. According to the in silico study, 2-chlorounguinol exhibited the highest binding-free energy (BFE) with PBP2a (-6.7 kcal/mol). For comparison, (E)-3-(2-(4-cyanostyryl)-4-oxoquinazolin-3(4H)-yl) benzoic acid inhibits PBP2a with a BFE less than −6.6 kcal/mol. Based on the Lipinski's rule of 5, depsidone compounds constitute a class of compounds with good pharmacokinetic properties, being easily absorbed and permeable. These findings suggest that 2-chlorounguinol possesses potential antibacterial activity and could be developed as an antibiotic adjuvant to reduce antimicrobial resistance.

Curcusinol from the fruit of Carex baccans with antibacterial activity against multidrug-resistant strains

Ethnopharmacological relevanceCarex baccans, known as Shan-Bai-Zi or Ye-Gao-Liang in China, is a traditional medicinal herb used by several ethnic groups in Yunnan Province. It is utilized for the treatment of wound infections, ulcers, and dysentery. However, there is currently a dearth of research reports on its antimicrobial potential. Aim of the studyThe substance basis of the antimicrobial activity of C. baccans will be unveiled, and the in vitro and in vivo antibacterial activities against multidrug-resistant bacteria of its major active compounds, as well as their preliminary mechanisms of action, will be investigated. Materials and methodsAn antibacterial bioactivity-guided isolation method was used to isolate and identify the active compound curcusinol from C. baccans. UPLC-DAD-MS was employed for the quantitative analysis of curcusinol. The antibacterial activity, resistance profile, synergistic effects, anti-biofilm activity, and potential mechanisms of action of curcusinol against methicillin-resistant Staphylococcus aureus (MRSA), vancomycin-resistant Enterococcus (VRE), and other multidrug-resistant bacteria (Escherichia coli, Pseudomonas aeruginosa, Acinetobacter baumannii) were investigated using various methods, including the broth microdilution method, scanning electron microscopy, time kill assay, multi-generational resistance induction assay, checkerboard synergy assay, anti-biofilm assay, and metabolomics. Furthermore, the therapeutic efficacy of curcusinol was assessed in vivo by establishing an animal skin wound infection model of MRSA. ResultsCurcusinol was isolated from the fruit of C. baccans, which accounts for 3.1% of the dry weight of the fruit. Curcusinol exhibited significant bactericidal and anti-biofilm activities against antibiotic-resistant Gram-positive bacteria in vitro. Furthermore, curcusinol acted as an antibiotic adjuvant to enhance the activity of various commonly used antibiotics against both Gram-positive and Gram-negative antibiotic-resistant bacteria without cytotoxicity to mammalian cells (A549 and RAW264.7) at 64 μM. Moreover, curcusinol affected arginine biosynthesis, cysteine and methionine metabolism, and alanine, aspartate, and glutamate metabolism significantly in MRSA cells under stress. Additionally, curcusinol effectively treated MRSA-infected mouse skin wounds and accelerated wound healing in vivo. ConclusionsThe results of this study not only support the traditional uses of C. baccans but also demonstrate that its major active compound, curcusinol, is an effective plant-derived bactericidal agent and antibacterial adjuvant with potential applications in the treatment of skin infections.

Proteomic Changes in Methicillin-Resistant Staphylococcus aureus Exposed to Cannabinoids.

Methicillin-resistant Staphylococcus aureus (MRSA) is a major human pathogen that causes a wide range of infections. Its resistance to β-lactam antibiotics complicates treatment due to the limited number of antibiotics with activity against MRSA. To investigate development of alternative therapeutics, the mechanisms that mediate antibiotic resistance in MRSA need to be fully understood. In this study, MRSA cells were subjected to antibiotic stress from methicillin in combination with three cannabinoid compounds and analyzed using proteomics to assess the changes in physiology. Subjecting MRSA to nonlethal levels of methicillin resulted in an increased production of penicillin-binding protein 2 (PBP2). Exposure to cannabinoids showed antibiotic activity against MRSA, and differential proteomics revealed reduced levels of proteins involved in the energy production as well as PBP2 when used in combination with methicillin.

Synthesis of structure-defined β-1,4-GlcNAc-modified wall teichoic acids as potential vaccine against methicillin-resistant Staphylococcus aureus

Methicillin-resistant Staphylococcus aureus (MRSA) is a high priority pathogen due to its life-threating infections to human health. Development of prophylactic or therapeutic anti-MRSA vaccine is a potential approach to treat S. aureus infections and overcome the resistance crisis. β-1,4-GlcNAc glycosylated wall teichoic acids (WTAs) derived from S. aureus are a new type of antigen that is closely associated with β-lactam resistance. In this study, structure-defined β-1,4-GlcNAc-modified WTAs varied in chain length and numbers of GlcNAc modification were synthesized by an ionic liquid-supported oligosaccharide synthesis (ILSOS) strategy in high efficiency and chromatography-free approach. Then the obtained WTAs were conjugated with tetanus toxin (TT) as vaccine candidates and were further evaluated in a mouse model to determine the structure-immunogenicity relationship. In vivo immunological studies revealed that the WTAs-TT conjugates provoked robust T cell-dependent responses and elicited high levels of specific anti-WTAs IgG antibodies production associated with the WTAs structure including chain length as well as the β-1,4-GlcNAc modification pattern. Heptamer WTAs conjugate T6, carrying three copy of β-1,4-GlcNAc modified RboP, was identified to elicit the highest titers of specific antibody production. The T6 antisera exhibited the highest recognition and binding affinity and the most potent OP-killing activities to MSSA and MRSA cells. This study demonstrated that β-1,4-GlcNAc glycosylated WTAs are promising antigens for further development against MRSA.

Promoting the healing of methicillin-resistant Staphylococcus aureus-infected wound by a multi-target antimicrobial AIEgen of 6-Aza-2-thiothymine-decorated gold nanoclusters

The use of conventional antibiotic therapies is in question owing to the emergence of drug-resistant pathogenic bacteria. Therefore, novel, highly efficient antibacterial agents to effectively overcome resistant bacteria are urgently needed. Accordingly, in this work, we described a novel class luminogen of 6-Aza-2-thiothymine-decorated gold nanoclusters (ATT-AuNCs) with aggregation-induced emission property that possessed potent antimicrobial activity against methicillin-resistant Staphylococcus aureus (MRSA). Scanning electron microscopy was performed to investigate the interactions between ATT-AuNCs and MRSA. In addition, ATT-AuNCs exhibited excellent ROS generation efficiency and could effectively ablate MRSA via their internalization to the cells. Finally, tandem mass tag-labeling proteome analysis was carried out to investigate the differential expression proteins in MRSA strains. The results suggested that ATT-AuNCs killed MRSA cells through altering the expression of multiple target proteins involved in DNA replication, aminoacyl-tRNA synthesis, peptidoglycan and arginine biosynthesis metabolism. Parallel reaction monitoring technique was further used for the validation of these proteome results. ATT-AuNCs could also be served as a wound-healing agent and accelerate the healing process. Overall, we proposed ATT-AuNCs could serve as a robust antimicrobial aggregation-induced emission luminogen (AIEgen) that shows the ability to alter the activities of multiple targets for the elimination of drug-resistant bacteria.

Photothermal and natural activity-based synergistic antibacterial effects of Ti3C2Tx MXene-loaded chitosan hydrogel against methicillin-resistant Staphylococcus aureus

Methicillin-resistant Staphylococcus aureus (MRSA) has strong resistance to antibiotic therapy. In this regard, developing antibiotic-free antibacterial agents is of great significance to treat MRSA infections. Herein, we loaded Ti3C2Tx MXene nanomaterial in the non-crosslinked chitosan (CS) hydrogel. The obtained MX-CS hydrogel is expected to not only adsorb MRSA cells via CS-MRSA interactions, but also gather the MXene-induced photothermal hyperthermia, achieving the efficient and intensive anti-MRSA photothermal therapy. As a result, under NIR irradiation (808 nm, 1.6 W/cm2, 5 min), MX-CS showed a greater photothermal effect than MXene alone did (30 μg/mL, 49.9 °C for MX-CS and 46.5 °C for MXene). Importantly, MRSA cells were rapidly adsorbed on MX-CS hydrogel (containing 30 μg/mL MXene) and completely inhibited (99.18 %) under NIR irradiation for 5 min. In contrast, MXene (30 μg/mL) and CS hydrogel alone only inhibited 64.52 % and 23.72 % MRSA, respectively, significantly lower than the inhibition caused by MX-CS (P < 0.001). Interestingly, when the hyperthermia was depleted by a 37 °C water bath, the bacterial inhibition rate of MX-CS significantly decreased to 24.65 %. In conclusion, MX-CS hydrogel has a remarkable synergistic anti-MRSA activity by gathering MRSA cells and MXene-induced hyperthermia, and may have great potentials in treating MRSA-infected diseases.

Disinfection of methicillin-resistant Staphylococcus Aureus on flat surface by 460 nm light and hydrogen peroxide combination

Staphyloxanthin (STX) is a carotenoid pigment produced by Staphylococcus aureus to protect the bacteria from oxidation stress. Eliminating Staphyloxanthin from S. aureus cell membrane by inducing the pigment photolysis using 460nm light, then killing weakened bacteria with an H2O2 solution could be a new approach to develop anti - Staphylococcus aureus procedure. A model to prove the feasibility of this combination to kill Methicillin-Resistant Staphylococcus Aureus (MRSA) bacteria on a flat surface was tested. Material and method: Staphyloxanthin pigment extracted from MRSA bacteria was treated with 460nm light at different light intensities to evaluate the photolysis potential of 460nm light. The change in MRSA shape after 460nm light treatment was also investigated by Scanning electron microscopy. Using glass cover-slips as an emulated model for contact surface in public, the combination of different 460nm light intensities and H2O2 0.75% was tested on the surface loaded with MRSA living cells, and the number of MRSA cells survived after treatment was enumerated. Result: Higher intensity and longer light treatment yielded a higher photolysis effect, as 100 and 200 mW/cm2 light intensity could degrade 77.50% to 83.45% of STX pigment after 20 minutes of irradiation. Also, MRSA cells had significant changes as more wrinkles and bumps appeared under high-intensity 460nm light. When tested on the flat surface of the coverslip, the strongest MRSA eradication effect can be observed in the combination of 200 mW/cm2 light treatment with 0.75% H2O2 solution, as 100% MRSA cells were completely killed after 20 minutes of treatment.