Galleria mellonella as a host model to study Candida glabrata virulence and antifungal efficacy

Candida species are common human fungal pathogens causing a wide range of clinical diseases, ranging from superficial infections to life-threatening systemic disease. Superficial infections include vaginal candidiasis which affects over 75% of women during their lifetime with 5% of women suffering debilitating recurrent infections. Life-threatening systemic Candida disease is the fourth most common nosocomial blood stream infection, affecting those undergoing chemotherapy, recovering from surgical procedures or major burns, transplant recipients and AIDs patients. The crude mortality rate associated with these infections is high, ranging from 46–75%, and current estimates suggest at least 400,000 life-threatening infections occur annually. Candida albicans is the predominant cause of invasive candidiasis, although in the last 3 decades there has been a rise in the incidence of non-albicans Candida species with Candida glabrata, Candida parapsilosis and Candida tropicalis being the other main agents causing disease. Of these, C. glabrata is the second most common cause of invasive candidiasis in the USA and Central and Northern Europe, and it has been associated with higher hospital costs. The basis of this increasing incidence of C. glabrata is not fully understood, however, it could be partially attributed to the higher innate tolerance C. glabrata displays to azole antifungals alongside its greater potential to develop drug resistance coincident with therapy. Murine models of infection are typically viewed as the gold standard for fungal virulence studies. However, although these models allow the host-pathogen interaction to be studied in vivo they do come with caveats associated with cost, legislation, and careful ethical considerations. Furthermore, with the development of large scalemutant libraries alternative, more ethically acceptable, models are required to identify interesting virulence targets while limiting the use of mice. Given the caveats associated with murinemodels of infectionmini-host models, mainly invertebrates, have been explored as alternative models for fungal infection. These models include amoeba (Dictyostellium discoideum), nematodes (Caenorhabditis elegans), fruit fly (Drosophila melanogaster) and the greater wax moth larvae (Galleria mellonella). G. mellonella, a lepidopteran, was first described as a mini-host for Candida species by Kavanagh and coworkers, and has received particular attention as an alternative host as it displays some important advantages. The G. mellonella larvae can be incubated at 37 C, allowing virulence to be studied at human body temperature, and an exact inoculum of the pathogen can be delivered by injection. Furthermore, the assays are inexpensive and simple to perform, allowing large numbers of larvae to be infected and thus increasing the statistical power of the assay. Finally some aspects of the G. mellonella immune response show similarities with the innate immune response of mammals. Given these advantages the model has now been developed for a wide range of fungal pathogens, including severalCandida species. To study C. glabrata infection in mice immunosuppression is usually required, and fungal burdens and persistence are normally used as a parameter for virulence due to the absence of mortality. This, along with the recent development of large scale mutant libraries, makes alternative models for studying C. glabrata virulence an attractive proposition. Initial reports on establishing the G. mellonella model for testing the virulence of Candida species only reported a low level of killing of larvae by C. glabrata. However, we and others have now shown that a faster rate of killing by C. glabrata is seen when using a higher pathogen concentration. In this work we provide the first detailed report on the ability of C. glabrata to grow and cause lethal infections in G. mellonella in a dose dependent manner. Furthermore, we have shown that this model can be used to assess the