Potentiation of Antibiotics by a Novel Antimicrobial Peptide against Shiga Toxin Producing E.\xa0coli O157:H7

Juan Puño-Sarmiento,Amber J Park,Debora E Barnett Foster,Cezar M Khursigara,Erin M Anderson

doi:10.1038/s41598-020-66571-z

Abstract

Infection with Shiga toxin-producing Escherichia coli (STEC) results in hemorrhagic colitis and can lead to life-threatening sequelae including hemolytic uremic syndrome (HUS). Conventional treatment is intravenous fluid volume expansion. Antibiotic treatment is contraindicated, due in part to the elevated risk of HUS related to increased Shiga toxin (Stx) release associated with some antibiotics. Given the lack of effective strategies and the increasing number of STEC outbreaks, new treatment approaches are critically needed. In this study, we used an antimicrobial peptide wrwycr, previously shown to enhance STEC killing without increasing Stx production, in combination with antibiotic treatments. Checkerboard and time-kill assays were used to assess peptide wrwycr-antibiotic combinations for synergistic STEC killing. Cytotoxicity and real-time PCR were used to evaluate Stx production and stx expression, respectively, associated with these combinations. The synergistic combinations that showed rapid killing, no growth recovery and minimal Stx production were peptide wrwycr-kanamycin/gentamicin. Transmission electron microscopy revealed striking differences in bacterial cell morphology associated with various treatments. This study provides proof of principle for the design of an antibiotic-peptide wrwycr combination effective in killing STEC without enhancing release of Shiga toxins. It also offers a strategy for the repurposing of antibiotics for treatment of STEC infection.

Highlights

Shiga toxin-producing Escherichia coli (STEC) is a major food- and water-borne pathogen that causes bloody diarrhea, hemorrhagic colitis and can lead to life-threatening sequelae including hemolytic uremic syndrome (HUS)[1,2,3,4]
Antibiotics selected for this study include those reported to have either no change or a decrease in Stx production as well as two reported to cause an increase in Shiga toxin production[13,16,17]
We provide a proof of principle evaluation of the effectiveness of treatments of the STEC O157:H7 strain 86–24 using different combinations of antibiotics with the novel antimicrobial peptide wrwycr

Summary

Introduction

Shiga toxin-producing Escherichia coli (STEC) is a major food- and water-borne pathogen that causes bloody diarrhea, hemorrhagic colitis and can lead to life-threatening sequelae including hemolytic uremic syndrome (HUS)[1,2,3,4]. Treatment consists primarily of intravenous fluid volume expansion and supportive therapy[9,10,11], while antimicrobials generally used to treat bacterial infections are often contraindicated The reason for this counter-intuitive practice is that in vitro studies have shown that, at least for STEC O157, sublethal doses of different antibiotics and classes of antibiotics, including ampicillin, ciprofloxacin, ceftazidime, trimethoprim-sulfamethoxazole, furazolidone and the quinolones can promote the production and release of Shiga toxins that may enhance the risk of progression to HUS5,12–15. We evaluate two antibiotics that have been reported to increase Stx production/release to establish whether it is possible to find a combination of peptide wrwycr with these antibiotics that could produce a bactericidal effect without increasing Stx production In this way, we may be able to explore the repurposing of antibiotics for treatment of STEC infection. This paper provides a proof of principle for the design of an antibiotic-peptide wrwycr combination that is effective in completely killing STEC without enhancing release of Shiga toxins

Methods

Results

Conclusion