The Galleria mellonella Waxworm Infection Model for Disseminated Candidiasis

Summary

Galleria mellonella serves as an invertebrate model for disseminated candidiasis. Here, we detail the infection protocol and provide supporting data for the model's effectiveness.

Abstract

Candida species are common fungal commensals of humans colonizing the skin, mucosal surfaces, and gastrointestinal tract. Under certain conditions, Candida can overgrow their natural niches resulting in debilitating mucosal infections as well as life-threatening systemic infections, which are a major focus of investigation due to their associated high mortality rates. Animal models of disseminated infection exist for studying disease progression and dissecting the characteristics of Candida pathogenicity. Of these, the Galleria mellonella waxworm infection model provides a cost-effective experimental tool for high-throughput investigations of systemic virulence. Many other bacterial and eukaryotic infectious agents have been effectively studied in G. mellonella to understand pathogenicity, making it a widely accepted model system. Yet, variation in the method used to infect G. mellonella can alter phenotypic outcomes and complicate interpretation of the results. Here, we outline the benefits and drawbacks of the waxworm model to study systemic Candida pathogenesis and detail an approach to improve reproducibility. Our results highlight the range of mortality kinetics in G. mellonella and describe the variables which can modulate these kinetics. Ultimately, this method stands as an ethical, rapid, and cost-effective approach to study virulence in a model of disseminated candidiasis.

Introduction

Candida species are common human commensals that are capable of emerging as opportunistic pathogens in severely immunocompromised and dysbiotic patients. Although many Candida species can cause disease, C. albicans is the most prevalent cause of disseminated candidiasis^1,2. Systemic disease results from C. albicans accessing the bloodstream through either direct penetration of previously restrictive host barriers or introduction at surgical sites and other breaches of the body³. Candida species utilize a range of pathogenic processes to cause....

Protocol

All methods described rely on use of invertebrate hosts and do not require Institutional Animal Care and Use Committee (IACUC) approval.

1. Galleria mellonella Waxworm Larvae

1. Order larvae from wholesalers and suppliers that do not introduce hormones, antibiotics, or other treatments to the larvae and which are able to ship and deliver live specimens.
   1. Be sure to purchase all larvae from the same supplier during the course of experimentation. Be careful when orde.......

Discussion

The G. mellonella waxworm model stands as an effective tool for the rapid and reproducible analysis of C. albicans virulence. This detailed protocol relies upon consistent delivery of a defined infectious dose to the same site across a batch of larvae. Infectious dose has a profound impact on G. mellonella mortality whereas use of larvae between their initial arrival and ten days following receipt produced similar results. Loss of the C. albicansMTLa allele results in .......

Materials

Name	Company	Catalog Number	Comments
Galleria mellonella	Snackworms.com		Buy twice as many worms as expected to use
10 uL, Model 1701 N SYR Cemented needle, 26G, type 2 syringe	Hamilton	80000
Petri dish, 100X15 mm, 500 pack	Fisher	FB0875712
Microcentrifuge tube, 1.7 mL, 500 pack	VWR	87003-294
Phosphate Buffered Saline (Biotechnology grade), 500 mL	VWR	97062-818
Ethanol absolute, ≥99.5% pure, 500 mL	Millipore Sigma	EM-EX0276-1S
autoclaved ddH2O