Diversity and Characterization of Lytic Bacteriophage Plaque Morphotypes Targeting Escherichia coli from Anthropized Aquatic Environments in Curitiba, Brazil: Implications for Phage Therapy

The global antimicrobial resistance crisis urgently demands innovative therapeutic alternatives, with lytic bacteriophages emerging as highly specific biocontrol agents against pathogenic Escherichia coli strains in both human and veterinary medicine. This study aimed to characterize the macromorphological diversity of bacteriophage plaque phenotypes isolated from highly anthropized aquatic environments (Belém River Sites I and II, Vila Formosa River, and Barigui River) in Curitiba, Paraná, Brazil, using the reference strain E. coli ATCC 25922 as the propagation host. Environmental samples underwent standardized enrichment protocols involving Luria-Bertani/TSB broth co-cultivation, chloroform treatment, and plaque assay quantification on TSB agar, enabling morphotype discrimination. Macroscopic analysis identified six distinct plaque morphotypes: (1) large clear plaques with halos; (2) large clear plaques without halos; (3) large plaques with central clearing; (4) diffuse large plaques exhibiting central clearing with turbid edges; (5) confluent microplaques resembling rainfall patterns; and (6) discrete small pinhead plaques. Site-specific distribution patterns suggested environmental phage heterogeneity, with Types 1 and 2 predominating in heavily sewaged Belém Site I, Types 3 and 4 in Belém Site II, and Types 5 and 6 in the comparatively less contaminated Vila Formosa and Barigui river systems. These findings demonstrate that plaque morphology can serve as a rapid and cost-effective preliminary indicator of environmental bacteriophage diversity. However, the exclusive use of the reference strain E. coli ATCC 25922 may not capture the full spectrum of phages capable of infecting clinically relevant or antimicrobial-resistant E. coli strains. Therefore, the observed morphotypic richness should be interpreted as an initial biodiversity screening. Future studies incorporating host-range analysis, electron microscopy, and genomic characterization are needed to evaluate the therapeutic potential of these phages. Nevertheless, the results highlight urban aquatic environments as promising reservoirs of bacteriophages and support their future application within a One Health framework linking environmental contamination, animal health, and public health.