Complete Genome Sequences of Four Mycobacteriophages Involved in Directed Evolution against Undisputed Mycobacterium abscessus Clinical Strains

Abstract

Mycobacterium; Cluster; Gene

Micobacteria; Clúster; Gen

Micobactèria; Clúster; Gen

Phage therapy is still in its infancy, but it is increasingly promising as a future alternative for treating antibiotic-resistant bacteria. To investigate the effect of phages on Mycobacterium abscessus complex (MABC), we isolated 113 environmental phages, grown them to high titres, and assayed them on MABC clinical strains through the spot test. Of all the phages, only 16 showed killing activity. Their activity was so temperate to MABC that they could not generate any plaque-forming units (PFUs). The Appelmans method of directed evolution was carried out to evolve these 16 phages into more lytic ones. After only 11 of 30 rounds of evolution, every single clinical strain in our collection, including those that were unsusceptible up to this point, could be lysed by at least one phage. The evolved phages were able to form PFUs on the clinical strains tested. Still, they are temperate at best and require further training. The genomes of one random parental phage and three random evolved phages from Round 13 were sequenced, revealing a diversity of clusters and genes of a variety of evolutionary origins, mostly of unknown function. These complete annotated genomes will be key for future molecular characterisations.

This research received financial support from Mutua Madrileña for the project ‘Formulación de bacteriófagos como liposomas y nanopartículas con aplicación potencial al tratamiento de infecciones de pacientes de fibrosis quística’, and is part of Project P122/00258, funded by Instituto de Salud Carlos III (ISCIII) and co-funded by the European Union and Centro para el Desarrollo Tecnológico Industrial (CDTI) from the Spanish Ministry of Economy, Trade and Enterprise (MINECO), with grant number IDI20200297.