Summary: Use of Bacteriophages as Antimicrobials Against Avian Pathogenic Escherichia coli Infections in Poultry: from Isolation to Therapeutic Evaluation

Use of Bacteriophages as Antimicrobials Against Avian Pathogenic Escherichia coli Infections in Poultry: from Isolation to Therapeutic Evaluation The poultry industry is affected by huge economic losses due to APEC infections. Colibacillosis is a cause of increased mortality and morbidity, decreased productivity and carcass rejection. The application of phage therapy to the large food animal industry would considerably reduce the extensive global use of antibiotics. Because of the increasing need for therapies offering an alternative to the use of antibiotics, the therapeutic use of a phage cocktail, composed of phages selected on their in vitro characteristics, was assessed. A broad phage collection of 76 phages, isolated from surface water samples, was constituted. During the analysis of their phenotypic characteristics, strictly lytic phages with a broad and complementary spectrum of infection were selected. Some of them displayed the valuable characteristics of generating low cross-resistance and serogroup O78-specificity. A total of nine phages was selected, among which four were able to grow on the challenge strain CH2, and thus later included in the cocktail. Site-directed mutagenesis revealed that three of these recognize the O78 antigen as receptor, while the fourth showed by transposon-mutagenesis to have OmpF and putatively the outer core of the lipopolysaccharide as receptor. The genomes of the selected phages underwent a further in silico analysis, where no predicted proteins revealed similarity with any known toxin, nor the presence of conserved toxin domains. The N4and T4-like nature of the cocktail phages confirmed their strictly lytic behavior. Moreover, a thorough genomic and proteomic analysis was performed on the phage PhAPEC6, as it possesses the third largest sequenced phage genome. The therapeutic efficiency of the in silico validated phage cocktail was further assessed by oral and intratracheal administration. Although the cocktail phages could be re-isolated from the lung and heart of chickens that were euthanized, none of the treated groups showed a significant decrease in mortality, lesion scores or weight loss, compared to the untreated group. The efficiency of the phage cocktail used in treating CH2-infected chickens in vivo is negligible, even though in vitro, the phages in the cocktail were able to efficiently lyse the APEC strain CH2.