Resistance to bacteriophage incurs a cost to virulence in drug resistant Acinetobacter baumannii

Introduction: Acinetobacter baumannii is a critical priority pathogen (World Health Organisation) because of the rise in nosocomial infections and its ability to evolve resistance to last resort antibiotics, which makes A. baumannii a priority target for phage therapy. Two strains of a novel, lytic bacteriophage (LemonAid and Tonic) able to infect carbapenem-resistant A. baumannii (strain NCTC 13420), were isolated from environmental water samples collected through a citizen science program.

Methods: In vitro and in vivo assays, genomics and microscopy techniques were used to characterise the phages, determine mechanisms of phage resistance and the efficacy of the phages against A. baumannii.

Results: A. baumannii developed resistance to both viruses, LemonAid and Tonic. Resistance came at a cost to virulence, with the resistant variants causing significantly reduced mortality in a Galleria mellonella larval in vivo model. A replicated 8bp insertion increased in frequency (~40% higher frequency than in the wildtype) within phage-resistant A. baumannii mutants, putatively resulting in early truncation of a protein of unknown function. Evidence from comparative genomics and an adsorption assay suggests this protein acts as a novel phage receptor site in A. baumannii. We find no evidence linking resistance to changes in capsule structure, a known virulence factor. LemonAid efficiently suppressed growth of A. baumanni in vitro across a wide range of titres. However, in vivo, while survival of A. baumannii infected larvae significantly increased with both remedial and prophylactic treatment with LemonAid (10⁷ PFU/mL), the effect was weak and not sufficient to save larvae from morbidity and mortality.

Conclusion: While LemonAid and Tonic did not prove effective as a treatment in a Galleria larvae model, there is potential to harness their ability to attenuate virulence in drug-resistant A. baumannii.