The One Health concept recognises the close interconnection between human, animal, and environmental health. In recent years, this perspective has intensified scientific focus on zoonoses. Among these, arboviruses—viruses transmitted by arthropod vectors—represent an emerging challenge, particularly in the present period strongly conditioned by climate change. Usutu virus (USUV) is a Flavivirus maintained via an enzootic bird–mosquito–bird cycle that infects other vertebrates. USUV is currently a significant animal health concern due to its expanding host range and increasing avian mortality events. Although USUV appears to be less dangerous than other emerging arboviruses in humans, the neurological disorders it can cause are alarming and increase the need for a better understanding of the spread and genetic evolution of USUV, as well as for the stronger promotion of vaccine and antiviral development. As with other arboviruses, treatment for USUV is limited to avoiding contact with mosquitoes, which is not always possible. Since vaccines do not yet exist, the use of modern OMICS sciences may provide comprehensive knowledge for developing effective control and prevention measures to avoid future pandemics and contain current epidemics.

Background/Objectives: Listeria monocytogenes is a major foodborne pathogen responsible for listeriosis, a serious illness with high morbidity and mortality, particularly in vulnerable populations. Its persistence in food processing environments and resistance to conventional preservation methods pose significant food safety challenges. Lactic acid bacteria (LAB) offer a promising natural alternative due to their antimicrobial properties, especially through the production of bacteriocins. This study investigates the competitive interactions between Lactococcus lactis and L. monocytogenes under co-culture conditions, with a focus on changes in their secretomes to better understand how LAB-derived bacteriocins can help mitigate the Listeria burden. Methods: Proteomic approaches, including Tricine-SDS-PAGE, two-dimensional electrophoresis, and shotgun proteomics, were employed to analyze the molecular adaptations of both species in response to bacterial competition. Results: Our results reveal a significant increase in the secretion of enolase by L. monocytogenes when in competition with L. lactis, suggesting its role as a stress-responsive moonlighting protein involved in adhesion, immune evasion, and biofilm formation. Concurrently, L. lactis exhibited a shift in the production of its bacteriocin, nisin, favoring the expression of Nisin Z—a variant with improved solubility and diffusion properties. This differential regulation indicates that bacteriocin production is modulated by bacterial competition, likely as a defensive response to the presence of pathogens. Conclusions: These findings highlight the dynamic interplay between LAB and L. monocytogenes, underscoring the potential of LAB-derived bacteriocins as natural biopreservatives. Understanding the molecular mechanisms underlying microbial competition could enhance food safety strategies, particularly in dairy products, by reducing reliance on chemical preservatives and mitigating the risk of L. monocytogenes contamination.