Background/Objectives: Antimicrobial resistance (AMR) poses a significant public health threat, leading to increased mortality. The World Health Organization has established a priority list highlighting critical multidrug-resistant (MDR) pathogens that demand urgent research on antimicrobial treatments. Considering this and the fact that new antibiotics are only sporadically approved, natural antibacterial agents have seen a resurgence in interest as potential alternatives to conventional antibiotics and chemotherapeutics. Natural antibacterials, derived from microorganisms, higher fungi, plants, animals, natural minerals, and food sources, offer diverse mechanisms of action against MDR pathogens. Here, we present a comprehensive summary of antibacterial agents from natural sources, including a brief history of their application and highlighting key strategies for using microorganisms (microbiopredators, such as bacteriophages), plant extracts and essential oils, minerals (e.g., silver and copper), as well as compounds of animal origin, such as milk or even venoms. The review also addresses the role of prebiotics, probiotics, and antimicrobial peptides, as well as novel formulations such as nanoparticles. The mechanisms of action of these compounds, such as terpenoids, alkaloids, and phenolic compounds, are explored alongside the challenges for their application, e.g., extraction, formulation, and pharmacokinetics. Conclusions: Future research should focus on developing eco-friendly, sustainable antimicrobial agents and validating their safety and efficacy through clinical trials. Clear regulatory frameworks are essential for integrating these agents into clinical practice. Despite challenges, natural sources offer transformative potential for combating AMR and promoting sustainable health solutions.

Infectious diseases remain a major global health concern, with a growing burden of antimicrobial resistance and consequent higher mortality in the human population. Accurate bacterial identification is fundamental across clinical, veterinary, agricultural, and research settings, supporting effective diagnosis, antimicrobial stewardship, infection control, food safety, and environmental monitoring; however, conventional approaches are limited by time constraints, reduced sensitivity, and challenges in detecting fastidious or uncultivable organisms. This review provides a comprehensive overview of classical and advanced methods, including microscopy, culture, biochemical testing, immunological and serological assays, proteomic and spectroscopy-based techniques, and molecular approaches, such as polymerase chain reaction (PCR), digital PCR, DNA hybridization, 16S rRNA gene sequencing, whole-genome sequencing, and metagenomics. The integration of artificial intelligence has further enhanced analytical performance. Nevertheless, harmonization of bioinformatics frameworks remains essential, as variability in algorithm-defined cut-off values limits standardized implementation of whole-genome sequencing in routine laboratories. Emerging technologies, including CRISPR-based diagnostics and phage- and nanomaterial-based detection systems, offer promising alternatives. Overall, the integration of these approaches is expected to improve the accuracy, speed, and applicability of bacterial identification across diverse settings; however, these advances should be implemented cautiously, with standardization remaining a key priority alongside technological modernization.