Identification of a large cohort of Enterobacter jumbo phages with broad host ranges across pathogenic Gammaproteobacteria

Abstract

ESKAPE pathogens cause most hospital-acquired infections globally and often carry antibiotic resistance. Many of them have been the target of bacteriophage therapies. Enterobacter is an ESKAPE pathogen but is less frequently a target for phage therapy due to a relative lack of available phages. We isolated eight jumbo phages with genomes ranging from 223 to 366 kbp targeting Enterobacter spp. and found that they belonged to separate phage clades. Six of them formed nucleus-like structures confirmed by DAPI-staining, and were phylogenetically related to Chimalliviridae. Two jumbo phages did not form nucleus-like structures and did not cluster with Chimalliviridae. Although these jumbo phages were found on Enterobacter, many were closely related to phages with non-Enterobacter hosts. To test whether these phages may have had expanded host ranges, we examined 14 pathogenic Gammaproteobacteria and found that these phages were capable of creating plaques on 8 of them. These species included Escherichia coli, Klebsiella aerogenes, Serratia marcescens, Salmonella spp., Shigella spp., Providencia spp., Citrobacter spp., and Cronobacter sakazakii. We verified that there was phage amplification in these microbes rather than lysis from without by performing qPCR to confirm DNA replication in each species. Phages typically have narrow host ranges, a benefit for microbiome-sparing compared to antibiotics. However, the broad host ranges of these Gammaproteobacteria jumbo phages suggests that not all phages have the same risk/benefit ratios. While this broad range could aid their development as antibiotic alternatives, further study is needed to assess potential microbiome disruption. SignificanceWith the growing threat of antibiotic resistant bacteria, alternative treatments like bacteriophages have emerged. Bacteriophages typically have narrow host ranges, a disadvantage compared to antibiotics. We discovered eight jumbo phages that kill antibiotic resistant Enterobacter, but were diverse phylogenetically. Six of them formed nucleus-like structures and were members of the Chimalliviridae family and the other two did not form nucleus-like structures. We confirmed that each phage had broad host ranges capable of lysing at least eight different human Gammaproteobacteria including pathogens such as E. coli, K. aerogenes, and S. marcescens. By identifying broad host range jumbo phages that attack pathogens, we may have identified phages with spectrums of activity more similar to antibiotics than have been traditionally attributed to phages.