This study focused on the isolation, characterization, and evaluation of the antimicrobial and antioxidant activities of a crude extract from Bacillus subtilis isolated from rhizosphere soil. Through biochemical and physiological assessments, followed by whole genome sequencing, the isolate was confirmed as Bacillus subtilis BSP1. We examined the antimicrobial activity of B. subtilis BSP1 metabolites against various pathogenic bacteria and fungi. To enhance its antibacterial efficacy, we optimized the fermentation medium to maximize the secretion of antibacterial agents. Our findings demonstrated that the crude extract exhibited notable antimicrobial properties against various pathogenic bacterial and fungal isolates. The antioxidant test revealed a dose-dependent increase in the extract’s DPPH scavenging activity and reducing power, with an impressive 98.9% DPPH scavenging activity at 30 mg/mL. Importantly, safety assessments indicated a lack of hemolytic activity on human red blood cells, with only 1.3% hemolysis at 100 mg/mL, suggesting its potential suitability for practical applications. In summary, Bacillus subtilis BSP1, isolated from soil, appears to be a promising candidate for antibiotic production. Its significant antimicrobial and antioxidant properties, combined with its safety profile, highlight its potential applications in medicine, agriculture, and biotechnology.

(1) Background: The emergence of multidrug-resistant (MDR) Salmonella enterica poses a major threat to global public health, underscoring the urgent need for alternative therapeutic strategies. Bacteriophages represent a promising alternative due to their high specificity and potent ability to lyse MDR strains. (2) Methods: In this study, we isolated a novel MDR Salmonella Enteritidis-targeting bacteriophage from Lebanese sewage and characterized its host range, thermal and pH stability, and infection dynamics. Whole-genome sequencing was performed using Illumina technology to determine its genetic features and taxonomic classification. (3) Results: the bacteriophage was classified within the genus Jerseyvirus and the class Caudoviricetes with a 43 kb dsDNA genome encoding 66 open reading frames (ORFs). It demonstrated remarkable thermal stability, retaining infectivity after prolonged incubation at 65 °C, and showed a broad host range. The phage formed large, clear plaques, displayed rapid adsorption (>97% within 3 min), a short latent period (20 min), and a burst size of ~32 PFU per cell. Genome analysis revealed no lysogeny, virulence, or resistance genes, confirming its strictly lytic nature and supporting its potential use as a biocontrol agent. (4) Conclusions: These findings identify SA01 as a novel, strictly lytic, and thermally stable bacteriophage with strong potential as a biocontrol agent against multidrug-resistant Salmonella Enteritidis. Its broad host range suggests potential activity also against other Salmonella enterica serovars, supporting its applicability in food safety and biotechnology.

Introduction: Carbapenem-resistant Gram-negative bacteria (CR-GNB) are rapidly spreading pathogens that increase morbidity and mortality in hospital settings and significantly restrict available treatment options worldwide. The lack of molecular epidemiological data and the limited use of next-generation sequencing (NGS) in South Lebanon have hindered comprehensive surveillance efforts. This study represents the first molecular characterization of CR-GNB in this region. Methods: A total of 477 clinical Gram-negative bacterial isolates were collected from intensive care unit (ICU) patients admitted to various hospitals in South Lebanon in 2023. Of these, 131 CR-GNB were subjected to whole-genome sequencing using the Illumina MiSeq platform. K-mer-based species identification, multilocus sequence typing (MLST), antimicrobial resistance (AMR) gene profiling, and plasmid analysis were performed using multiple bioinformatic tools. Phylogenetic analysis was conducted using SaffronTree. Results: K-mer-based identification revealed that the predominant species among CR-GNB isolates were Pseudomonas spp. and Escherichia coli (26.7% each), followed by Klebsiella pneumoniae (19.8%), Acinetobacter baumannii (17.6%), Proteus mirabilis (4.6%), Enterobacter cloacae (2.3%), Achromobacter spp. (1.5%), and Citrobacter freundii (0.8%). Based on antimicrobial susceptibility testing, isolates were classified as follows: 0.8% as pan drug-resistant (PDR), 40.5% as extensively drug-resistant (XDR), and 52.7% as multidrug-resistant (MDR) and 6.1% as antimicrobial-resistant (AMR). All isolates harbored AMR genes, with the following distribution: 2% bla_VIM, 5% bla_NDM-1, 27% bla_NDM-5, 65% bla_OXA-type, and 1% bla_DIM-1. Plasmid-associated AMR genes were detected in 58% of isolates; among these, 96% carried Inc-family plasmids, 57% Col plasmids, and 11% replication-associated elements (rep). Phylogenetic analysis demonstrated that certain isolates exhibited both hospital-specific and shared genetic profiles, indicating widespread dissemination across multiple healthcare facilities, as well as evidence of local emergence and ongoing transmission. Conclusions: The high prevalence of CR-GNB harboring resistance genes and plasmids underscores the urgent need for NGS-based genomic surveillance in South Lebanon. Implementing such strategies is essential for tracking resistance genes, identifying clonal outbreaks, and guiding effective infection control interventions to mitigate the spread of CR-GNB.

The overuse of carbapenems has driven the rise of carbapenem-resistant Pseudomonas aeruginosa (CRPA) and Enterobacterales (CRE), against which ceftazidime–avibactam (CAZ-AVI) offers an alternative treatment. This study phenotypically determined resistance profiles of P. aeruginosa (PA), Escherichia coli (EC), and Klebsiella pneumoniae (KP) clinical isolates to CAZ-AVI and investigated molecular resistance mechanisms genotypically. A total of 394 PA, 90 EC, and 84 KP isolates were collected from the American University of Beirut Medical Center. Antimicrobial susceptibility testing (AST) and whole-genome sequencing (WGS) were performed on 30 isolates per species. Results showed that 100% of KP, 63% of EC, and 100% of PA isolates were carbapenem-resistant. Among these, 73% of KP, 79% of EC, and 60% of PA were CAZ-AVI-resistant. WGS revealed diverse sequence types, plasmids, and antimicrobial resistance genes. Additionally, 100% of KP, 93% of EC, and 89% of PA isolates produced metallo-β-lactamases (MBLs). Mutations in ampD, ampR, and mexR were identified in CAZ-AVI-resistant, non-MBL-producing PA, whereas mutations in ompC, marR, and ampC were detected in CAZ-AVI-resistant, non-MBL-producing EC. While CAZ-AVI remains effective against most CRE and CRPA lacking MBLs, resistance to CAZ-AVI is multifactorial, commonly involving overexpression of efflux pumps and AmpC β-lactamases, loss of porin channels, and the presence of oxacillinases.

1) Background: The emergence of multidrug-resistant (MDR)-Salmonella enterica poses a major threat to global public health underscoring the urgent need for alternative therapeutic strategies where bacteriophages are a promising candidate due to their high specificity and potent ability to lyse MDR strains.2) Methods: In this study, we isolated a novel MDR-Salmonella Enteritidis-targeting bacteriophage from Lebanese sewage and characterized its host range, thermal and PH stability, and infection dynamics. Whole-genome sequencing was performed by Illumina to determine genetic features and taxonomic position.3) Results: the bacteriophage belongs to genus Jerseyvirus and Class Caudoviricetes with a 43kb dsDNA genome and 66 ORFs. It demonstrated extraordinary thermal stability, retaining infectivity after prolonged incubation at 65 °C, showed a broad host range, produced large, clear plaques, displayed rapid adsorption (>97% within 3 min), a short latent period (20 min), and a burst size of ~320 PFU per cell. Genome analysis revealed no lysogeny, virulence, or resistance genes, confirming its strictly lytic nature and supporting its potential use as a biocontrol agent. 4) Conclusions: These findings identify SA01 as a novel, strictly lytic, and thermally stable bacteriophage with strong potential as a biocontrol agent against multidrug-resistant Salmonella Enteritidis, supporting its potential activity also against other Salmonella enterica serovars for future applications in food safety and biotechnology.

The overuse of carbapenems has driven the rise of carbapenem-resistant Pseudomonas aeruginosa (CRPA) and Enterobacterales (CRE), against which ceftazidime-avibactam (CAZ-AVI) offers an alternative treatment. This study phenotypically determined resistance profiles of P. aeruginosa (PA), Escherichia coli (EC), and Klebsiella pneumoniae (KP) clinical isolates to CAZ-AVI and investigated molecular resistance mechanisms genotypically. A total of 394 PA, 90 EC, and 84 KP isolates were collected from the American University of Beirut Medical Center. Antimicrobial susceptibility testing (AST) and whole-genome sequencing (WGS) were performed on 30 isolates per species. Results showed 100% of KP, 63% of EC, and 100% of PA isolates were carbapenem-resistant. Among these, 73% of KP, 79% of EC, and 60% of PA were CAZ-AVI-resistant. WGS revealed diverse sequence types, plasmids, and antimicrobial resistance genes. Additionally, 100% of KP, 93% of EC, and 89% of PA isolates produced metallo-β-lactamases (MBLs). Mutations in ampD, ampR, and mexR were identified in CAZ-AVI-resistant, non-MBL-producing PA, whereas mutations in ompC, marR, and ampC were detected in CAZ-AVI-resistant, non-MBL-producing EC. While CAZ-AVI remains effective against most CRE and CRPA lacking MBLs, resistance to CAZ-AVI is multifactorial, commonly involving overexpression of efflux pumps and AmpC β-lactamases, loss of porin channels, and the presence of oxacillinases.