Enterococcus Phage Abitsa: Biological Characterization, Antibiofilm Activity and Evolutionary Insights into the Genus...

Enterococcus faecalis is an important opportunistic pathogen associated with persistent oral infections, biofilm formation, and antimicrobial tolerance, which makes phages targeting this species of both therapeutic and evolutionary interest. Here, we describe the isolation and characterization of Abitsa, a novel lytic Enterococcus phage representing a new species within the genus Efquatrovirus. Abitsa formed small plaques and displayed siphovirus-like morphology. It showed an optimal multiplicity of infection of 0.01, a short latent period of 10 min and a burst size of 28 ± 5 virions per infected cell. The phage efficiently suppressed planktonic growth of E. faecalis over a broad multiplicity of infection (MOI) range and significantly disrupted pre-formed biofilms, with the strongest effect observed at MOI 0.1, resulting in an 82.14% reduction in biofilm biomass. Abitsa remained stable at pH 4-8, at 5-50 °C for 1 h, and in up to 75% chloroform, but exhibited a narrow host range, lysing only one additional clinical E. faecalis isolate among 34 tested enterococcal isolates. Genome analysis showed that Abitsa has a 41,581-bp linear genome lacking lysogeny-associated genes and represents a novel Efquatrovirus species. Comparative structural and phylogenetic analyses additionally supported mosaic evolution and extensive domain shuffling in receptor-binding and lysis-related proteins of Efquatrovirus-like phages. Together, these results identify Abitsa as a biologically unusual and evolutionarily informative lytic phage with antibiofilm activity.