The use of invertebrates for in vivo studies in microbiology is well established in the scientific community. Larvae of Galleria mellonella are a widely used model for studying pathogenesis, the efficacy of new antimicrobial compounds, and immune responses. The immune system of G. mellonella larvae is structurally and functionally similar to the innate immune response of mammals, which makes this model suitable for such studies. In this review, cellular responses (hemocytes activity: phagocytosis, nodulation, and encapsulation) and humoral responses (reactions or soluble molecules released in the hemolymph as antimicrobial peptides, melanization, clotting, free radical production, and primary immunization) are discussed, highlighting the use of G. mellonella as a model of immune response to different human pathogenic microorganisms.

Probiotic strains of Lactobacillus have been studied for their inhibitory effects on Candida albicans. However, few studies have investigated the effect of these strains on biofilm formation, filamentation and C. albicans infection. The objective of this study was to evaluate the influence of Lactobacillus acidophilus ATCC 4356 on C. albicans ATCC 18804 using in vitro and in vivo models. In vitro analysis evaluated the effects of L. acidophilus on the biofilm formation and on the capacity of C. albicans filamentation. For in vivo study, Galleria mellonella was used as an infection model to evaluate the effects of L. acidophilus on candidiasis by survival analysis, quantification of C. albicans CFU/mL, and histological analysis. The direct effects of L. acidophilus cells on C. albicans, as well as the indirect effects using only a Lactobacillus culture filtrate, were evaluated in both tests. The in vitro results showed that both L. acidophilus cells and filtrate were able to inhibit C. albicans biofilm formation and filamentation. In the in vivo study, injection of L. acidophilus into G. mellonella larvae infected with C. albicans increased the survival of these animals. Furthermore, the number of C. albicans CFU/mL recovered from the larval hemolymph was lower in the group inoculated with L. acidophilus compared to the control group. In conclusion, L. acidophilus ATCC 4356 inhibited in vitro biofilm formation by C. albicans and protected G. mellonella against experimental candidiasis in vivo.

Galleria mellonella is a well-accepted insect model for the study of pathogen-host interactions and antimicrobial compounds. The main advantages of this model include the low cost of maintenance, the fast life cycle, the possibility of using a large number of caterpillars and the innate immune system, which is evolutionarily conserved relative to mammals. Because of these advantages, different research groups have been working to implement the rearing of G. mellonella in laboratory conditions. This protocol describes our experience in the rearing of G. mellonella caterpillars for experimental infection models and the influence of different artificial diets on developmental and physiological parameters. Here, we suggest a diet composition that benefits the life cycle of G. mellonella by accelerating the larval phase length and increasing the caterpillar weight. This diet also stimulated the immune system of G. mellonella by increasing the hemolymph volume and hemocyte concentration. In addition, our rearing protocol generated caterpillars that are more resistant to infection by Staphylococcus aureus, Escherichia coli and Candida albicans. A standard G. mellonella rearing protocol is fundamental to minimize external influences on the results, and this simple and easy protocol can support researchers starting to rear G. mellonella.

Streptococcus mutans and Candida albicans are found together in the oral biofilms on dental surfaces, but little is known about the ecological interactions between these species. Here, we studied the effects of S. mutans UA159 on the growth and pathogencity of C. albicans. Initially, the effects of S. mutans on the biofilm formation and morphogenesis of C. albicans were tested in vitro. Next, we investigate the influence of S. mutans on pathogenicity of C. albicans using in vivo host models, in which the experimental candidiasis was induced in G. mellonella larvae and analyzed by survival curves, C. albicans count in hemolymph, and quantification of hyphae in the host tissues. In all the tests, we evaluated the direct effects of S. mutans cells, as well as the indirect effects of the subproducts secreted by this microorganism using a bacterial culture filtrate. The in vitro analysis showed that S. mutans cells favored biofilm formation by C. albicans. However, a reduction in biofilm viable cells and inhibition of hyphal growth was observed when C. albicans was in contact with the S. mutans culture filtrate. In the in vivo study, injection of S. mutans cells or S. mutans culture filtrate into G. mellonella larvae infected with C. albicans increased the survival of these animals. Furthermore, a reduction in hyphal formation was observed in larval tissues when C. albicans was associated with S. mutans culture filtrate. These findings suggest that S. mutans can secrete subproducts capable to inhibit the biofilm formation, morphogenesis and pathogenicity of C. albicans, attenuating the experimental candidiasis in G. mellonella model.