Bacterial phylogeny explains variation in virulence but not phage efficacy in vivo

Abstract

The continuing emergence of antimicrobial resistance (AMR) poses one of the most urgent threats to public health in the 21st century, causing [~]1.2 million deaths per year. A promising alternative to traditional antimicrobials is phage therapy, which has proved effective in the treatment of antimicrobial-resistant infections, including methicillin-resistant Staphylococcus aureus (MRSA). However, considerable variation in the outcome of phage therapy remains, with phage that show high efficacy in vitro often showing little to no effect when applied in vivo. To allow for the efficient clinical use of phage, it is vital to understand how both bacterial virulence and phage efficacy vary in vivo and determine how well in vitro and in vivo measures of phage efficacy correlate. Here, we infected 4,968 Galleria mellonella larvae with 64 phylogenetically diverse Staphylococcaceae isolates, both with and without co-inoculation of the bacteriophage ISP, and recorded mortality and melanisation over 24 hours. We found that bacterial virulence varied among Staphylococcaceae strains, and that a large proportion of this variation could be explained by the evolutionary relationships between bacteria. These results indicate that pathogen phylogeny may be a useful tool for explaining variation in both the severity of clinical infections and the virulence of novel emerging pathogens. The addition of phage significantly improved the survival of G. mellonella larvae, with an average 15.2% increase in endpoint survival and 10.1% reduction in endpoint melanisation across Staphylococcaceae strains. Phage efficacy in vivo showed phylogenetic repeatability, but no detectable phylogenetic heritability across bacteria species, in contrast to previous in vitro studies of phage infections. Concurrently, we found no evidence of a correlation between in vivo and in vitro measures of phage efficacy across bacterial isolates, demonstrating that environment plays a major role in determining the outcomes of bacteria-phage interactions. This highlights that caution should be used when extrapolating from in vitro measures of phage efficacy to select phages for therapeutic uses.