Background/Objectives: Antimicrobial resistance (AMR) is a major public health threat, which is exacerbated by the lack of new antibiotics and the emergence of multidrug-resistant (MDR) superbugs. Comprehensive efforts and alternative strategies to combat AMR are urgently needed to prevent social, medical, and economic consequences. Pseudomonas aeruginosa is a pathogen responsible for a wide range of infections, from soft tissue infections to life-threatening conditions such as bacteremia and pneumonia. Bacteriophages have been considered as a potential therapeutic option to treat bacterial infections. Our aim was to isolate phages able to infect MDR P. aeruginosa strains. Methods: We isolated two lytic phages, using the conventional double layer agar technique (DLA), from samples obtained from the influent of a wastewater treatment plant in Concepción, Chile. The phages, designated as PaCCP1 and PaCCP2, were observed by electron microscopy and their host range was determined against multiple P. aeruginosa strains using DLA. Moreover, their genomes were sequenced and analyzed. Results: Phage PaCCP1 is a member of the Septimatrevirus genus and phage PaCCP2 is a member of the Pbunavirus genus. Both phages are tailed and contain dsDNA. The genome of PaCCP1 is 43,176 bp in length with a GC content of 54.4%, encoding 59 ORFs, one of them being a tRNA gene. The genome of PaCCP2 is 66,333 bp in length with a GC content of 55.6%, encoding 102 non-tRNA ORFs. PaCCP1 is capable of infecting five strains of P. aeruginosa, whereas phage PaCCP2 is capable of infecting three strains of P. aeruginosa. Both phages do not contain bacterial virulence or AMR genes and contain three and six putative Anti-CRISPR proteins. Conclusions: Phages PaCCP1 and PaCCP2 show promise as effective treatments for MDR P. aeruginosa strains, offering a potential strategy for controlling this clinically important pathogen through phage therapy.

There is an urgent need for alternative solutions to combat multidrug-resistant (MDR) E. coli infections. In recent years, there has been an increase in MDR strains causing urinary tract infections (UTIs), which has resulted in more challenging treatment options, increased healthcare costs and prolonged hospital stays. The utilization of bacteriophages as a prospective modality for the management of bacterial infections has garnered significant attention. The objective of this study was to isolate and describe a phage capable of infecting MDR E. coli strains isolated from the urine of patients affected with UTI. The phage EcoPhCCP1 was isolated using the plaque assay from the influent of a wastewater treatment plant. The phage was characterized by phenotypic and genomic features. Morphological characteristics such as shape and size were determined using electron microscopy, and its host range was determined against multiple MDR strains. The complete genome of the phage was subjected to whole-genome sequencing and then assembled and annotated to search for virulence or antimicrobial resistance gene (ARG). VIRIDIC was employed to compare the closest phage genomes, while VICTOR and taxMyPhage were used to construct its phylogeny. EcoPhCCP1 is a tailed phage capable of infecting and propagating in multiple MDR E. coli strains recovered from UTI. The phage genome is 44,482 bp in length, with a GC content of 50.7 mol%, and encodes 87 ORFs, 33 of which have been previously functionally annotated. Phage EcoPhCCP1 is a Kagunavirus , in the recently created Sarkviridae family. Notably, phage EcoPhCCP1 does not harbour ARGs or virulence genes, thus rendering it a promising candidate for phage therapy against clinically significant MDR E. coli strains. Moreover, phage EcoPhCCP1 possesses putative anti-CRISPR proteins.

Escherichia coli is a major pathogen responsible for urinary tract infections, septicemia, and other clinically relevant conditions, with increasing multidrug resistance (MDR) limiting available treatment options. In this context, bacteriophages represent a valuable resource for exploring novel antimicrobial and biotechnological tools. Here, we report the isolation and genomic characterization of BorMax, a novel lytic phage infecting multiple MDR E. coli. Transmission electron microscopy revealed a tailed morphology consistent with Dhillonvirus. Whole genome sequencing and de novo assembly showed a linear double-stranded DNA genome of 45,502 bp, encoding 70 predicted coding sequences (CDSs) and lacking tRNAs. Bioinformatic analyses confirmed the absence of lysogeny-associated genes, as well as virulence and antimicrobial resistance determinants. Comparative genomics using classified BorMax within the genus Dhillonvirus as a new species, sharing <77% intergenomic similarity with known members. Notably, predictions using PaCRISPR and AcRanker identified four CDSs with strong anti-CRISPR (Acr) potential, representing previously undescribed Acr candidates in this group. These genomic features highlight the novelty, safety, and potential biotechnological relevance of BorMax and contribute to the expanding genomic and functional diversity of Dhillonvirus and E. coli-infecting phages.