The therapeutic potential of bacteriophages targeting gram-negative bacteria using Galleria mellonella infection model

Prasanth Manohar,Ramesh Nachimuthu,Bruno S. Lopes

doi:10.1186/s12866-018-1234-4

Prasanth Manohar, Ramesh Nachimuthu + Show 1 more

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https://doi.org/10.1186/s12866-018-1234-4

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Abstract

BackgroundPhage therapy is the therapeutic use of bacteriophages to treat highly drug resistant bacterial infections. The current surge in bacteriophage therapy is motivated mainly because of the emergence of antibiotic-resistant bacteria in clinics. This study evaluated the therapeutic potential of three bacteriophages isolated against Escherichia coli ec311, Klebsiella pneumoniae kp235 and Enterobacter cloacae el140 strains using Galleria mellonella. The in vitro activity of three different phages belonging to Podoviridae and Myoviridae families was studied by the double agar overlay method against multi-drug resistant strains. Larval survivability studies were performed to evaluate the potential of phages against infection using G. mellonella.ResultsAll the three phages were found to have potential to infect the host bacterial strains. For in vivo studies it was observed that E. coli and E. cloacae infected larvae, should be treated with three phage doses (20 μL, 104 PFU/mL) at 6 h interval to achieve 100% survival rate. But in the case of K. pneumoniae, a single phage dose treatment showed promising outcome. When mixed bacterial infections (all three bacterial cultures at 108 CFU/mL) were tested, minimum of four doses of phage cocktail (three phages) at 6 h interval was necessary to recover the larvae. All the results were confirmed by enumerating bacteria from the larvae.ConclusionOur data shows that although in vitro studies showed high infectivity of phages, for in vivo models multiple phage doses were required for effective treatment.

Highlights

Phage therapy is the therapeutic use of bacteriophages to treat highly drug resistant bacterial infections
We examined the in vivo therapeutic efficiency of three bacteriophages; Escherichia phage ECP311 (ECP311), Klebsiella phage KPP235 (KPP235) and Enterobacter phage ELP140 (ELP140) against E. coli, K. pneumoniae and E. cloacae strains, using G. mellonella as a model organism
ECP311 belonged to Phieco32likevirus (Podoviridae family) and has the adsorption velocity of 1.1 × 10− 9 mL/min, latency period of 26 min and a burst size of 180 phage particles/infected cell, KPP235 belonged to Podoviridae of phages with the adsorption velocity of 4.35 × 10− 9 mL/min, latent period of 40 min and a burst size of 120 phage particles/infected cell while ELP140 belonged to Myoviridae and has an adsorption velocity, latent period and burst size of 2.8 × 10− 9 mL/min, 11 min and 135 phages/infected cell (Additional file 1: Figure S1)

Summary

Introduction

Phage therapy is the therapeutic use of bacteriophages to treat highly drug resistant bacterial infections. This study evaluated the therapeutic potential of three bacteriophages isolated against Escherichia coli ec311, Klebsiella pneumoniae kp235 and Enterobacter cloacae el140 strains using Galleria mellonella. Phage therapy involves the use of bacteriophages for the treatment of bacterial infections and has gained a renewed interest because of antibiotic resistance in bacteria [7–9]. We examined the in vivo therapeutic efficiency of three bacteriophages; Escherichia phage ECP311 (ECP311), Klebsiella phage KPP235 (KPP235) and Enterobacter phage ELP140 (ELP140) against E. coli, K. pneumoniae and E. cloacae strains, using G. mellonella as a model organism. We investigated the effective dosage of target phages required to reduce the lethality of bacterial infections in G. mellonella model and evaluated the dose dependent potential of phage cocktail (three phages each at 104 PFU/mL) to target multiple bacterial infection

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