Refining the Galleria mellonella Model by Using Stress Marker Genes to Assess Clostridioides difficile Infection and Recuperation during Phage Therapy.

Janet Y Nale,Mahananda Chutia,Jeffrey K J Cheng,Martha R J Clokie

doi:10.3390/microorganisms8091306

Janet Y Nale, Mahananda Chutia + Show 2 more

Open Access

https://doi.org/10.3390/microorganisms8091306

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Abstract

The Galleria mellonella is an effective model for probing Clostridioides difficile interactions with phages. Despite valuable insights from this model, the larvae are not easily amenable to assessing detailed clinical responses to either bacteria or phages. Here, larval survival, colonisation and toxin levels were compared to expression profiles of 17 G. mellonella stress genes to monitor Clostridiodes difficile infection (CDI), and recuperation during phage therapy. The larvae were infected with a ribotype 014/020 isolate and treated with an optimised phage cocktail. Larvae treated prophylactically with phages and the phage-control larval group were protected, showing the highest survival, and low C. difficile colonisation and toxin rates, compared to co-infection, remedial and bacterial-control larval groups. Expression of growth (9) and reproduction (2) genes were enhanced within prophylaxis and phage-control larval groups compared to the co-infection, remedial and bacterial control groups. In contrast, expression of infection (2), humoral (1) and cellular (3) immunity genes declined in the prophylactic and phage-control groups but increased in the co-infection, remedial and bacterial control larvae. The molecular markers augment the survival, colonisation and toxin data and allow detailed monitoring of CDI and recovery. This data support the use of stress marker genes as tools to analyse clinical symptoms in this model.

Highlights

Clostridioides difficile is a toxin-producing bacillus and a major cause of antibiotic-induced diarrhea [1,2]
Among the three therapy regimens investigated here, as expected, insects in the prophylaxis group were best protected from the infection, a 100% survival rate was recorded throughout the experiment, which is significant when compared to the bacterial control (p < 0.001)
Among the insects treated with a co-infection of the phage cocktail and bacteria, the first fatality was recorded in the first 24th hour, when ~10% of the insects died

Summary

Introduction

Clostridioides difficile (formerly, Clostridium difficile) is a toxin-producing bacillus and a major cause of antibiotic-induced diarrhea [1,2]. C. difficile infection (CDI) is characterised by one or more episodes of loose diarrhea, a positive stool test for C. difficile toxins A/B, and endoscopic evidence of pseudomembranous colitis, with or without a positive culture assay [3,4]. Only three antibiotics (metronidazole, vancomycin and fidaxomicin) are approved for the treatment of CDI, but evidence for reduced susceptibility and resistance to metronidazole and vancomycin, respectively, have been reported [12,13,14,15]. New antibiotics and treatments such as fecal transplant are being developed, novel anti-infectives with target specificity and minimal deleterious impact on human gut microbial niche and immune system would be most appropriate to treat CDI [17,18]

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