The global rise of multidrug-resistant (MDR) pathogens has created an urgent need for alternative antimicrobial agents. In this study, we evaluated the antibacterial activity of LTX-315, a synthetic cationic 9-mer peptide originally developed as an oncolytic agent, against six ATCC reference strains and six clinical isolates, including methicillin-resistant Staphylococcus aureus (MRSA), extended-spectrum β-lactamase (ESBL)-producing Klebsiella pneumoniae, MDR Pseudomonas aeruginosa, Stenotrophomonas maltophilia, and Burkholderia cepacia. LTX-315 exhibited broad-spectrum antibacterial activity, with MICs of 8-32 μM against reference strains and as low as 4 μM against several MDR clinical isolates, whereas chloramphenicol and kanamycin were ineffective. Double-disk diffusion assays indicated synergistic interactions with aminoglycosides, chloramphenicol, and vancomycin, and checkerboard assays confirmed synergy-to-additivity interactions with gentamicin and vancomycin against S. aureus (FICI 0.38-0.75), yielding fourfold to 16-fold reductions in antibiotic dose. These findings demonstrate that LTX-315 retains potent activity against clinically relevant MDR pathogens and may serve as both a standalone antimicrobial peptide and a potential antibiotic adjuvant.

This study evaluated the antibacterial and anti-biofilm activities of selected metal ions (Al[3+], Cu[2+], Au[3+], Ag[+], and Zn[2+]) against extended-spectrum β-lactamase-producing Escherichia coli (ESBL-E. coli), Methicillin-Resistant Staphylococcus aureus (MRSA), and vancomycin-resistant Enterococcus faecalis (VRE). The rapid emergence of antimicrobial resistance (AMR) has intensified the threat posed by multidrug-resistant (MDR) pathogens, particularly biofilm-forming bacteria associated with dairy equipment surfaces. Among the tested metals, silver ions showed the strongest antibacterial activity, producing inhibition zones up to 19.00 ± 1.15 mm at 0.3125 mM and a minimum inhibitory concentration (MIC) of 0.078 mM, followed by gold ions (MIC 2.5 mM). Zinc and copper exhibited moderate activity at higher concentrations, whereas aluminium showed the least effect. Checkerboard assays demonstrated synergistic or additive interactions, particularly for Cu-Au, Zn-Cu, and Zn-Ag combinations [Fractional inhibitory concentration index (FICI) 0.5-0.75], while a ternary Cu-Zn-Ag combination showed notable additive interaction (FICI 0.512). Propidium iodide uptake assays suggested membrane damage and disruption of cell integrity. Silver also exhibited the highest biofilm eradication activity (50% biofilm eradication concentration (BEC50 0.156 mM), followed by gold and copper. On stainless steel dairy equipment surfaces, silver and gold reduced biofilm biomass by > 60% and > 40%, respectively, as confirmed by confocal laser scanning microscopy and scanning electron microscopy. Silver additionally induced the highest intracellular reactive oxygen species generation (florescence intensity 93854.92 ± 2616.2 to 165407.18 ± 148.3), indicating oxidative stress-mediated antibacterial action than other metals. This study provides a comparative evaluation of metal ions and their synergistic combinations against MDR pathogens and biofilms associated with dairy equipment surfaces and dairy-processing environments.

The rising antimicrobial resistance of clinically relevant organisms, such as Staphylococcus aureus and Escherichia coli, has triggered researchers' attention to natural medicinal plants as an alternative source of antibacterial agents. The purpose of this study was to evaluate the antibacterial, phytochemical, and toxicological activities of Cassia senna leaf. The n-hexane, methanolic, and aqueous extracts of Cassia leaves were assayed for antibacterial activity using agar well diffusion, disc diffusion, and broth microdilution assays with gentamicin, vancomycin, and chloramphenicol. The major bioactive components of the extract were identified using thin-layer chromatography (TLC) coupled with bioautography, while phytochemical screening was done on the methanolic extract. Sub-chronic (30-day repeated-dose) toxicity was evaluated in mice that received 1000 mg of the methanolic extract per kg of body weight per day for 30 consecutive days, monitoring the changes in body weight, organ appearance, blood cell count, biochemical profiles, and histology. While the S. aureus isolate was sensitive to all three reference antibiotics, the E. coli isolate was resistant to them. The methanolic extract demonstrated better antibacterial activity than the aqueous extract, as it produced bigger inhibition zones and lower minimum inhibitory concentrations than the aqueous one, while n-hexane extract did not show any antibacterial activity. The activity of Cassia senna leaf extract was higher against S. aureus than Gram-negative E. coli. TLC-bioautography revealed that there were at least four compounds in the methanolic extract that contributed to its antibacterial activity, while phytochemical screening of the methanolic extract showed that it contained flavonoids and tannins, but not phenolics, saponins, alkaloids, or terpenoids. Finally, the 30-day sub-chronic toxicity study revealed that repeated oral administration of 1000 mg/kg/day of Cassia senna leaf extract did not cause any mortality or adverse effect in mice, with the treated animals showing comparable results to the control group in terms of body weight, organ appearance, blood cell count, biochemical profile, and histology. The present study demonstrated that Cassia senna leaves had a considerable antibacterial effect, especially against S. aureus isolate, and were safe and tolerable in mice at a repeated oral dose of 1000 mg/kg/day for 30 days. Therefore, it can be suggested that Cassia leaf extract can be further explored as a promising source of antibacterial compounds.

Polymicrobial communities where bacteria must compete with each other to persist can serve as a source of uncharacterized antibacterial compounds to develop drugs for the treatment of drug-resistant infections. This study investigates interbacterial competition between bacteria found in the oral cavity, where Streptococcus species comprise a large portion of the resident oral microbiota, and Enterococcus faecalis is a pathobiont that is commonly found in root canal infections. We used co-cultures to determine whether oral streptococci and E. faecalis compete with each other. Our experiments revealed that multiple strains of Streptococcus mutans, an important cariogenic bacterium, reduce the viability of vancomycin-resistant Enterococcus and other gram-positive bacteria, including Staphylococcus epidermidis and methicillin-resistant Staphylococcus aureus. Furthermore, reduction in viability of bacteria by some strains of S. mutans requires the production of the non-ribosomal cyclic lipopeptide mutanobactin, while another strain inhibits independently of mutanobactin. We determined that S. mutans mutanobactin production increases target cell membrane permeability and that reduction of target cell viability is contact dependent. We also determined that the secreted E. faecalis protease gelatinase (GelE) is required for recovery from mutanobactin-mediated reduction in viability. Additionally, our data show that S. mutans mutanobactin production prevents E. faecalis biofilm formation and kills biofilm-associated E. faecalis. Together, this work demonstrates how natural products from a common oral bacterium contribute to competition in polymicrobial environments, which will inform future strategies to treat and prevent bacterial infections.IMPORTANCEAntimicrobial resistance requires new therapeutics to treat drug-resistant infections. Novel antimicrobial compounds can be discovered in polymicrobial communities, where bacterial natural products promote competitive fitness. The oral cavity hosts a microbial consortium, and we investigated interactions between oral streptococci and Enterococcus faecalis, a common cause of root canal infections. We demonstrate antibacterial activity of streptococcal mutanobactin against gram-positive pathogens, including antibiotic-resistant isolates. We further show that the E. faecalis virulence factor gelatinase promotes recovery from mutanobactin-mediated reduction in viability, and that mutanobactin prevents and kills E. faecalis biofilms. By probing interactions between bacteria that occupy the same niche and characterizing antibacterial activity of a bacterial product, this work contributes to broader efforts to identify and develop antibiotics to treat clinically relevant drug-resistant infections.

Treatment of healthcare-associated infections is becoming increasingly challenging due to rising antimicrobial resistance (AMR). As clinical microbiology and public health continue to transition toward whole-genome sequencing (WGS) for outbreak detection, diagnostics, and surveillance, the ability to accurately predict antimicrobial resistance from genomic sequence data will become crucial. In this study, in silico antimicrobial resistance prediction with the Staramr tool was refined from whole-genome sequencing data for four top-priority healthcare-associated bacteria: carbapenemase-producing Escherichia coli, carbapenemase-producing Klebsiella pneumoniae, methicillin-resistant Staphylococcus aureus, and vancomycin-resistant Enterococcus faecium. Isolates (n = 4,876) were collected in Canada within two national surveillance programs: the Canadian Nosocomial Infection Surveillance Program and the Canadian Ward Study. Broth microdilution was used as the gold standard to evaluate the performance of Staramr. The average sensitivity, specificity, and concordance of categorical antimicrobial resistance prediction compared to broth microdilution were 90.8%, 90.1%, and 92.4% for E. coli (n = 1,095); 87.7%, 91.7%, and 90.6% for K. pneumoniae (n = 687); 95.5%, 98.9%, and 96.3% for methicillin-resistant S. aureus (n = 2,150); and 95.4%, 97.9%, and 97.2% for vancomycin-resistant Enterococcus faecium (n = 944). Additionally, this internal tool refinement identified areas of improvement for antimicrobial resistance prediction. The average sensitivity, specificity, and concordance of antimicrobial resistance prediction were high (>90%) for most combinations of antimicrobials and species that were evaluated. While AMR prediction from WGS was very promising, it did not correlate well with current Clinical and Laboratory Standards Institute breakpoints for non-susceptibility for some antimicrobial-organism combinations. As in silico prediction of antimicrobial resistance has been improved in this study through tool refinement, it shows future promise for molecular methods to complement phenotypic antimicrobial susceptibility testing. This will allow for more precise tracking of the molecular epidemiology of resistance and cost savings when isolates are being sequenced for strain typing.

Bacteria's ability to resist current antibiotics has grave implications for future treatment options. Herein, we report novel pyrimidine-sulfonyl chloride derivatives as potential antimicrobial agents against multi-drug resistance, synthesised through a molecular hybridisation technique. FTIR, [1]H-NMR and [13]C-NMR analysis data corroborated the structural confirmation of all synthesised compounds. The compounds were screened against a panel of Gram-positive and Gram-negative bacterial strains, including Mycobacterium tuberculosis strains. Cell permeability assays were also performed for MRSA and S. aureus. Among the tested compounds, 6l and 6m exhibited the most potent antibacterial activity against MRSA with a MIC value of 0.2 μg mL[-1], more potent than both ciprofloxacin (MIC = 3.91 μg mL[-1]) and vancomycin (MIC = 0.49 μg mL[-1]). Further 6j and 6m also displayed good antibacterial activity against clinically isolated MRSA with MIC values of 0.97 and 7.8 μg mL[-1], respectively. In contrast, compounds 6g and 6q showed the best activity against Gram-negative bacteria E. hormaechei (MIC = 0.97 μg mL[-1]), comparable to ciprofloxacin (MIC = 0.2 μg mL[-1]). In the antimycobacterial studies, compounds 6a, 6c, 6i, 6k and 6l were more potent (MIC = 3.91 μg mL[-1]) against the extensively drug-resistant Mycobacterium tuberculosis (XDR) strain than the standard drugs used and against H37Rv, compounds 6i and 6k exhibited the best potency with a MIC value of 3.91 μg mL[-1]. The cell penetration experiment demonstrated that compounds 6h-6m achieved greater than 90% propidium iodide penetration in S. aureus (translating to cell death) at the stated MIC well, whereas less than 80% cell penetration was achieved for methicillin-resistant S. aureus. Computational simulation studies were conducted to validate the biological findings, showing potential binding affinity at the binding site of MurB and FemA enzymes. Thus, pyrimidine-based sulfonyl chloride derivatives exhibited promising anti-microbial and anti-mycobacterial activities.

A series of novel 2-aminopyrimidine-thiazolidinone hybrids were designed, synthesized, and evaluated for antimicrobial activity against selected bacterial and fungal pathogens. Structural characterization was accomplished using FT-IR, NMR, and mass spectrometry. Among the synthesized compounds, SPRB21 exhibited the most potent antibacterial activity, with MIC values of 125 μg/mL against Escherichia coli ATCC 25922 and ATCC 11775 and 250 μg/mL against Pseudomonas aeruginosa ATCC BAA-427. SPRB17 also demonstrated promising activity, with MIC values of 250, 250, and 500 μg/mL, respectively, showing activity comparable to vancomycin against E. coli and superior activity against P. aeruginosa. In antifungal studies, SPRB27 displayed the highest activity against Candida albicans ((MICf = 12.15 μg/mL; MFC = 61.42 μg/mL),), while SPRB23 exhibited the best fungicidal activity against Aspergillus niger (MICf = 53.75 μg/mL; MFC = 206.5 μg/mL). Although these compounds were less potent than the standard antifungal drug, fluconazole, they demonstrated notable antifungal efficacy within the synthesized series. Molecular docking against dihydrofolate reductase (DHFR) revealed strong binding affinities for SPRB22 and SPRB23, while SAR studies indicated that electron-withdrawing substituents enhanced antimicrobial potency. SwissADME predictions confirmed favorable drug-likeness and oral bioavailability, and 300 ns molecular dynamics simulations validated the stable binding of SPRB21 and SPRB23 within bacterial and fungal DHFR active sites, respectively. These findings identify the 2-aminopyrimidine-thiazolidinone scaffold as a promising platform for the development of novel antimicrobial agents.