In agroecosystems, bacterial antagonism is an alternative to agrochemicals, which are a threat to the environment and global health. From this perspective, a collection of environmental bacterial isolates was tested for their potential as antagonists against the phytopathogenic bacterium Xanthomonas vesicatoria, which is one of the etiological agents of bacterial spot in tomato and pepper. The isolates with the greatest in vitro antagonistic activity were selected for ex vivo assays using tomato plant leaves and further characterized for their capacity to produce hydrolytic enzymes. The results revealed promising prospects for an antagonism-based X. vesicatoria biocontrol against bacterial spot disease, thus favouring more sustainable agricultural production.

Protecting crops from bacterial diseases is a major challenge for sustainable agriculture. Bacterial spot caused by Xanthomonas arboricola pv. pruni (Xap) threatens stone fruit and almond production, while restrictions on agrochemicals in the European Union have increased the need for safe alternatives. Bacteriophages (phages) are promising biotools for controlling bacterial plant pathogens. However, there is limited research on Xap-specific phages in Europe, particularly under Mediterranean conditions. The aim of this study was to isolate and characterise phages from environmental sources with lytic activity against Spanish Xap strains from different Prunus species and geographical locations. Six Xap phages were isolated from different environmental water samples and characterised using genomic, phylogenetic, morphological and biological analyses. These studies enabled three of the phages to be assigned to different species and revealed a putative novel genus of previously undescribed phages with myovirus morphology. In vitro assays confirmed their specificity and ability to infect and lyse multiple Spanish Xap strains, significantly reducing bacterial populations by phage-based biocontrol. Five of them were selected and combined into different four-phage cocktails. The selected phages demonstrated high stability in vitro and in planta. Following optimisation of foliar spray application, two selected cocktails significantly reduced bacterial spot in Prunus plants. To our knowledge, these are the first European Xap phage cocktails to be effective in Prunus. They therefore represent promising, safe and sustainable biocontrol agents for managing bacterial spot in stone fruit and almond crops under Mediterranean climatic conditions. They could also be integrated into broader disease management strategies for woody crops and could help to reduce dependence on agrochemicals.

Background/Objectives: Escherichia coli is one of the most problematic bacterial species in hospital and community settings and a leading cause of resistant infections worldwide. This reinforces the need of new antibacterial tools like phage therapy. This study aimed to isolate and characterise lytic bacteriophages (phages) targeting clinical E. coli strains with different resistance profiles and to assess their potential for future therapeutic development. Methods: Environmental and clinical samples were used to isolate lytic phages active against clinical E. coli strains, and to evaluate their potential for future therapeutic applications. A phage collection was tested against E. coli strains from human bloodstream infections to determine their bacteriolytic activity, host range and specificity. Two phages were selected for further phenotypic and genomic characterisation. Their infectivity and antibacterial activity were evaluated under growth medium- and filtered human urine-related conditions and across different temperature and pH ranges. Results: Both phages demonstrated consistent antimicrobial activity against susceptible clinical strains, exhibiting a narrow host range and specificity. They retained infectivity under growth medium- and filtered human urine-related conditions, significantly reducing bacterial populations, and maintaining lytic activity across a range of temperatures and pH values. Genomic characterisation confirmed their lytic nature and supported their safety. Taxonomic analysis revealed that one phage was classified as a new podovirus-morphotype species within the genus Xuquatrovirus. The other displayed characteristics consistent with a novel genus, showing myovirus morphology. Conclusions: Environmental reservoirs represent a source of lytic phages active against clinically relevant E. coli strains. The new coliphages provide a basis for future development of phage-based strategies against multidrug-resistant E. coli strains.