Introduction: The rapid global increase in multidrug-resistant (MDR) bacteria has compromised the effectiveness of conventional antibiotics, stressing the urgent need for alternative antimicrobial strategies. CRISPR–Cas systems, originally evolved as bacterial adaptive immune mechanisms, provide programmable and highly specific tools for targeting antimicrobial resistance (AMR) determinants. Objective: This systematic review aims to evaluate the antibacterial mechanisms, delivery strategies, preclinical evidence, safety considerations, and translational potential of CRISPR–Cas systems for combating MDR bacterial infections. Methods: A systematic literature search was conducted in PubMed, Scopus, Cochrane Library, and Web of Science up to January 2026 in accordance with PRISMA 2020 guidelines. Eligible studies included original in vitro and in vivo experimental or preclinical investigations assessing CRISPR–Cas systems (Cas9, Cas12, Cas13, or related effectors) for antibacterial activity or antibiotic resensitization. Data were extracted on CRISPR effector type, bacterial target, delivery platform, and therapeutic outcome. Due to methodological heterogeneity, results were synthesized narratively. Results: Most studies reported effective killing or resensitization of MDR bacteria through chromosomal double-strand break induction, resistance plasmid curing, integron disruption, or RNA-targeted cleavage. Cas9 was the most frequently employed effector, followed by Cas12 and Cas13. Delivery strategies included bacteriophages, conjugative plasmids, and nanoparticle-based systems, with phage-mediated delivery demonstrating the most consistent efficacy in complex environments and animal models. Notably, a CRISPR-enhanced engineered bacteriophage cocktail (LBP-EC01) has advanced to clinical evaluation. Discussion: Overall, the evidence supports CRISPR–Cas antimicrobials as a promising precision-based approach for addressing AMR. However, major barriers remain, including limited host range, instability in physiological environments, emergence of escape mutations, and insufficient data on off-target effects and long-term safety. Addressing these challenges through optimized delivery platforms, multiplex targeting strategies, and standardized safety and regulatory frameworks will be essential for clinical translation. Systematic review registration: https://www.crd.york.ac.uk/PROSPERO/view/CRD420261319789 , identifier CRD4201319789.

Background: Moringa oleifera and Moringa peregrina have been extensively studied for their medicinal properties, particularly due to their rich content of bioactive phytocompounds. While their antibacterial effects on Gram-positive bacteria are well documented, there is a paucity of comparative studies on their efficacy against Gram-negative multidrug-resistant (MDR) strains found in arid regions. Methods: Leaf extracts of M. oleifera and M. peregrina, collected from Wadi Ad-Dawasir in Saudi Arabia, were prepared using ethanol, hot aqueous, and cold aqueous extraction methods. Their antibacterial properties were tested against Escherichia coli , Pseudomonas aeruginosa , Shigella sonnei , and Shigella shiga using the agar well diffusion method, minimum inhibitory concentration (MIC) assays (50–200 μg/mL), and scanning electron microscopy (SEM). Phytochemical analysis was conducted using gas chromatography–mass spectrometry (GC–MS) and standard qualitative methods. Amoxicillin–clavulanic acid and tetracycline were used as reference antibiotics. Results: All extracts showed antibacterial activity, with ethanol extracts producing the largest inhibition zones. M. peregrina was particularly effective, especially against Sh. sonnei , with an inhibition zone of 24.66 ± 0.17 mm, which surpassed that of amoxicillin–clavulanic acid (18.25 ± 1.69 mm). The MIC values were generally 50 μg/mL for both species, except for the cold extract of M. oleifera against P. aeruginosa , which had an MIC of 100 μg/mL. GC–MS analysis identified 31 compounds in M. peregrina , with stigmasterol and β -sitosterol being the most abundant. In contrast, M. oleifera was characterized by a profile rich in fatty acids. The SEM analysis revealed membrane damage in Sh. sonnei treated with the extracts, which included surface collapse, perforation, and cytoplasmic leakage. Conclusion: Both Moringa species demonstrated significant antibacterial activity against clinically important Gram-negative strains, with M. peregrina occasionally outperforming standard antibiotics. The presence of sterols in the phytochemical profile likely enhances their bactericidal action by targeting bacterial membranes. These results suggest that Moringa could serve as a viable phytotherapeutic option for addressing the growing challenge of Gram-negative bacterial resistance.

Candida auris is an emerging multidrug-resistant fungal pathogen posing serious global healthcare challenges due to its persistence, frequent misidentification, and resistance to all major antifungal drugs. This study aimed to explore natural therapeutic alternatives by investigating the antifungal and molecular properties of Moringa peregrina and Moringa oleifera leaf extracts. Ethanolic and aqueous extracts of both Moringa species were prepared and evaluated against C. auris and other Candida species ( Candida spp .) using zone of inhibition (ZI), minimum inhibitory concentration (MIC), and minimum fungicidal concentration (MFC) assays. Phytochemical composition was characterized through fourier transform infrared spectroscopy (FTIR) and gas chromatography–mass spectrometry (GC–MS) analyses, while cytotoxicity on mammalian MCF-7 cells was assessed using the MTT assay. Molecular docking and 100-ns molecular dynamics simulations were conducted to examine the binding of key bioactive compounds to C. auris dihydrofolate reductase (DHFR; PDB: 8CRH). Ethanolic extracts exhibited the strongest antifungal activity (ZI: 16–16.5 mm; MIC: 0.5–0.7 mg/mL), whereas aqueous extracts were comparatively less effective. FTIR spectra revealed prominent peaks corresponding to hydroxyl (O–H), carbonyl (C = O), and C–O functional groups, confirming the presence of alcohols, esters, phenols, and carboxylic acids, while GC–MS identified β-sitosterol, stigmasterol, dihydroxanthin, and vitamin E derivatives as predominant constituents. Docking results revealed high binding affinities for vitamin E (− 86.6 kcal/mol) and stigmasterol (− 83.5 kcal/mol), exceeding that of fluconazole (− 27.3 kcal/mol). Molecular dynamics confirmed stable protein–ligand complexes, with hydrophobic and van der Waals forces dominating the interactions. Cytotoxicity assays revealed low toxicity at antifungal concentrations (IC₅₀ ≥ 120 µg/mL). These findings demonstrate that Moringa extracts—particularly the ethanolic fractions—harbor potent bioactive compounds with promising antifungal activity against C. auris , highlighting their potential as novel, plant-derived therapeutic candidates to combat resistant fungal infections.

Pseudomonas aeruginosa (P. aeruginosa) is a multidrug-resistant opportunistic pathogen that causes both acute and chronic infections and is known for its ability to form biofilms. In the current study, we applied a hypothesis-generating framework primarily based on integrating four different datasets and applying batch correction. Weighted Gene Co-Expression Network Analysis (WGCNA) was performed in parallel with differential expression analysis using limma. Therefore, we aimed to identify potential biofilm-associated gene candidates. Significant candidate genes were subjected to functional analysis and gene ontology, followed by the construction of a protein–protein interaction network using STRING. The Pseudomonas Genome Database was used to highlight the candidate genes. A total of 271, 687, 533, and 277 significantly up-regulated differentially expressed genes (DEGs), as well as 306, 985, 472, and 312 significantly down-regulated DEGs, resulted from the exploratory analysis. Through WGCNA/limma integration, 223 common significantly up-regulated/positively correlated gene candidates were identified. Functional analysis results showed significant enrichment in virulence-related pathways, such as biofilm formation (PA0083, PA0084, hcp1, hcpC, pilH, pilI, pilJ, vfr, pqsA, pqsB, pqsC, pqsE, PA1657, and PA1658). In addition, other virulence-related pathways, such as quorum sensing, phenazine biosynthesis, the bacterial secretion system, and secondary metabolite biosynthesis, were enriched. In conclusion, our hypothesis-generating integrative analysis identifies candidate genes and potential pathways associated with biofilm formation, virulence, and other processes in P. aeruginosa. In light of this, we point out that all candidate genes presented in this study remain hypothesis-generating. Further validation is recommended, including large-scale in silico analyses and in vitro experimental studies.