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The focus of this meeting was to discuss the suitability of using bacteriophages as alternative antimicrobials in the agrifood sector. Following a One Health approach, the workshop explored the possibilities of implementing phage application strategies in the agriculture, animal husbandry, aquaculture, and food production sectors. Therefore, the meeting had gathered phage researchers, representatives of the agrifood industry, and policymakers to debate the advantages and potential shortcomings of using bacteriophages as alternatives to traditional antimicrobials and chemical pesticides. Industry delegates showed the latest objectives and demands from consumers. Representatives of regulatory agencies (European Medicines Agency (EMA) and Spanish Agency of Medicines and Health Products (AEMPS)) presented an update of new regulatory aspects that will impact and support the approval and implementation of phage application strategies across the different sectors. [ABSTRACT FROM AUTHOR]

Due to the emergence of antibiotic resistance, phage-mediated biocontrol has become an attractive alternative for pathogen management in agriculture. While the infection characteristics of many phages can be adequately described using plaque assays and optical density, the results from phages of the apple pathogen Erwinia amylovora have low reproducibility with these techniques. Using quantitative real-time PCR (qPCR), the stage of the lytic cycle was determined through a combination of chloroform-based sampling, centrifugation, and DNase treatment. Monitoring the transition of phage genomes through the lytic cycle generates a molecular profile from which phage infection characteristics such as adsorption rate and burst size can be determined. To our knowledge, this is the first report of qPCR being used to determine these infection parameters. The characteristics of four different genera of Erwinia phages were determined. The phage ΦEa46-1-A1 was able to adsorb at a rate up to 6.6 times faster than ΦEa35-70 and ΦEa9-2. The low enrichment titer of ΦEa9-2 was shown to be due to the absence of lysis. The ΦEa46-1-A1 and ΦEa21-4 phages had the highest productivity, with burst sizes of 57 virions in 38 min and 185 virions in 98 min, respectively, suggesting these genera would make stronger candidates for the phage-mediated biocontrol of E. amylovora. [ABSTRACT FROM AUTHOR]

Bacteriophages are viruses capable of recognizing with high specificity, propagating inside of, and destroying their bacterial hosts. The phage lytic life cycle makes phages attractive as tools to selectively kill pathogenic bacteria with minimal impact on the surrounding microbiome. To effectively harness the potential of phages in therapy, it is critical to understand the phage–host dynamics and how these interactions can change in complex populations. Our model examined the interactions between the plant pathogen Erwinia amylovora, the antagonistic epiphyte Pantoea agglomerans, and the bacteriophages that infect and kill both species. P. agglomerans strains are used as a phage carrier; their role is to deliver and propagate the bacteriophages on the plant surface prior to the arrival of the pathogen. Using liquid cultures, the populations of the pathogen, carrier, and phages were tracked over time with quantitative real-time PCR. The jumbo Myoviridae phage ϕEa35-70 synergized with both the Myoviridae ϕEa21-4 and Podoviridae ϕEa46-1-A1 and was most effective in combination at reducing E. amylovora growth over 24 h. Phage ϕEa35-70, however, also reduced the growth of P. agglomerans. Phage cocktails of ϕEa21-4, ϕEa46-1-A1, and ϕEa35-70 at multiplicities of infections (MOIs) of 10, 1, and 0.01, respectively, no longer inhibited growth of P. agglomerans. When this cocktail was grown with P. agglomerans for 8 h prior to pathogen introduction, pathogen growth was reduced by over four log units over 24 h. These findings present a novel approach to study complex phage–host dynamics that can be exploited to create more effective phage-based therapies. [ABSTRACT FROM AUTHOR]

Erwinia amylovora is a globally devastating pathogen of apple, pear, and other Rosaceous plants. The use of lytic bacteriophages for disease management continues to garner attention as a possible supplement or alternative to antibiotics. A quantitative productive host range was established for 10 Erwinia phages using 106 wild type global isolates of E. amylovora, and the closely related Erwinia pyrifoliae, to investigate the potential regional efficacy of these phages within a biopesticide. Each host was individually infected with each of the 10 Erwinia phages and phage production after 8 h incubation was measured using quantitative real time PCR (qPCR) in conjunction with a standardized plasmid. PCR amplicons for all phages used in the study were incorporated into a single plasmid, allowing standardized quantification of the phage genome copy number after the infection process. Nine of the tested phages exhibited a broad host range, replicating their genomes by at least one log in over 88% of tested hosts. Also, every Amygdaloideae infecting E. amylovora host was able to increase at least one phage by three logs. Bacterial hosts isolated in western North America were less susceptible to most phages, as the mean genomic titre produced dropped by nearly two logs, and this phenomenon was strongly correlated to the amount of exopolysaccharide produced by the host. This method of host range analysis is faster and requires less effort than traditional plaque assay techniques, and the resulting quantitative data highlight subtle differences in phage host preference not observable with typical plaque-based host range assays. These quantitative host range data will be useful to determine which phages should be incorporated into a phage-mediated biocontrol formulation to be tested for regional and universal control of E. amylovora. [ABSTRACT FROM AUTHOR]