Enterococcus helps E. coli ‘armor up’ in dog, poultry co-infections

A new study finds that two subtypes of pathogenic Escherichia coli (E. coli) produce five to 16 times more protective capsular “slime” when Enterococcus faecalis (EF) is present. The finding could lead to targeted therapies for E. coli infections specific to dogs and poultry.

The E. coli in question – uropathogenic E. coli (UPEC) and avian pathogenic E. coli (APEC) – cause urinary tract infections in dogs and bloodstream infections in poultry, respectively.

“Urinary tract infections, while not usually fatal to dogs, are extremely common and one of the leading reasons antibiotics are prescribed in small animal medicine,” says Grayson Walker, former DVM/Ph.D. student at North Carolina State University and corresponding author of the study. Walker is currently a veterinary medical officer with the U.S. Department of Agriculture.

“On the other hand, APEC is a leading cause of poultry death worldwide,” Walker says. “And both infections are made more severe when there is a co-infection with EF. Previous studies have shown that part of the reason for this is that EF helps E. coli survive in low-iron environments such as the urinary tract or the bloodstream. We wanted to see what else might be happening.”

The research team began by growing different APEC and UPEC strains progressively closer to Enterococcus in an iron-restricted culture system. They identified EF-responsive strains of APEC and UPEC and noted that these strains grew faster and produced more exopolysaccharide, a slimy, protective capsule, when they were in closer proximity to Enterococcus.

Then they looked at EF-responsive and non-responsive strains in a chicken embryo model of co-infection and found increased mortality in embryos coinfected with EF-responsive strains compared to those coinfected with non-responsive strains or with APEC or EF alone.

They compared the genomes of EF-responsive and non-responsive strains and found that, in addition to iron acquisition genes, responsive strains had genes associated with virulence and capsule production specifically.

“For these infections in dogs and poultry, Enterococcus is acting as E. coli’s armorer,” Walker says. “We already knew that coinfection overcomes low-iron environments. Now we know it also enables E. coli to better protect itself.

“Hopefully this study will lead to the identification of new targets for vaccines or therapeutics against these coinfections of Enterococcus and pathogenic E. coli.”

The paper appears in PLOS ONE and was supported by the U.S. Food and Drug Administration GenomeTrakr program (1U18FD00678801) and the U.S. Department of Agriculture Animal Plant Health Inspection Service National Bio and Agro-Defense Facility Scientist Training Program. NC State co-authors include research specialist Mitsu Suyemoto, and professors Megan Jacob and Siddhartha Thakur. Former NC State associate professor Luke Borst also contributed to the work.

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Note to editors: An abstract follows.

“Canine uropathogenic and avian pathogenic Escherichia coli harboring conjugative plasmids exhibit augmented growth and exopolysaccharide production in response to Enterococcus faecalis”

DOI:  10.1371/journal.pone.0312732

Authors: Grayson K. Walker, M. Mitsu Suyemoto, Megan E. Jacob, Siddhartha Thakur, and Luke B. Borst, North Carolina State University
Published: Nov. 27, 2024 in PLOS ONE

Abstract:
Uropathogenic Escherichia coli (UPEC) and avian pathogenic Escherichia coli (APEC) are extraintestinal pathogenic Escherichia coli (ExPEC) that infect dogs and poultry. These agents occur both as single-species infections and, commonly, in co-infection with Enterococcus faecalis (EF); however, it is unclear how EF co-infections modulate ExPEC virulence. Genetic drivers of interspecies interactions affecting virulence were identified using macrocolony co-culture, chicken embryo co-infection experiments, and whole-genome sequence analysis of ExPEC and EF clinical isolates. Ten of 11 UPEC strains originally co-isolated with EF exhibited a growth advantage when co-cultured with EF on iron-limited, semi-solid media in contrast to growing alone (P < 0.01). Phylogenetic analyses of these UPEC and 18 previously screened APEC indicated the growth-response phenotype was conserved in ExPEC despite strain diversity. When genomes of EF-responsive ExPEC were compared to non-responsive ExPEC genomes, EF-induced growth was associated with siderophore, exopolysaccharide (EPS), and plasmid conjugative transfer genes. Two matched pairs of EF-responsive and non-responsive ExPEC were selected for further characterization by macrocolony proximity and chicken embryo lethality assays. EF-responsive ExPEC produced 5 to 16 times more EPS in proximity to EF and were more lethal to embryos alone and during co-infection with EF compared to non-responsive ExPEC (P < 0.05). A responsive APEC strain cured of its conjugative plasmid lost the enhanced growth and EPS production response to EF. These data demonstrate that ExPEC growth augmentation by EF occurs in UPEC and APEC strains and is linked to conjugative virulence plasmids and EPS production, which are widely conserved ExPEC virulence determinants.