Use of <em>Galleria mellonella</em> as a Model Organism to Study <em>Legionella pneumophila</em> Infection

Abstract

Citation: Harding, C.R., Schroeder, G.N., Collins, J.W., Frankel, G. Use of Galleria mellonella as a Model Organism to Study Legionella pneumophila Infection. J. Vis. Exp. (81), e50964, doi:10.3791/50964 (2013). AbstractLegionella pneumophila, the causative agent of a severe pneumonia named Legionnaires' disease, is an important human pathogen that infects and replicates within alveolar macrophages. Its virulence depends on the Dot/Icm type IV secretion system (T4SS), which is essential to establish a replication permissive vacuole known as the Legionella containing vacuole (LCV). L. pneumophila infection can be modeled in mice however most mouse strains are not permissive, leading to the search for novel infection models. We have recently shown that the larvae of the wax moth Galleria mellonella are suitable for investigation of L. pneumophila infection. G. mellonella is increasingly used as an infection model for human pathogens and a good correlation exists between virulence of several bacterial species in the insect and in mammalian models. A key component of the larvae's immune defenses are hemocytes, professional phagocytes, which take up and destroy invaders. L. pneumophila is able to infect, form a LCV and replicate within these cells. Here we demonstrate protocols for analyzing L. pneumophila virulence in the G. mellonella model, including how to grow infectious L. pneumophila, pretreat the larvae with inhibitors, infect the larvae and how to extract infected cells for quantification and immunofluorescence microscopy. We also describe how to quantify bacterial replication and fitness in competition assays. These approaches allow for the rapid screening of mutants to determine factors important in L. pneumophila virulence, describing a new tool to aid our understanding of this complex pathogen.

Legionella-protozoa-nematode interactions in aquatic biofilms and influence of Mip on Caenorhabditis elegans colonization

“…Some indications come from genomic data which suggest that certain genes might have been transferred horizontally from metazoans (Gomez-Valero et al, 2014). Further support comes from co-culture experiments with multicellular model organisms and human tissues which revealed several virulence traits and factors of L. pneumophila with a high specificity and relevance for the infection of metazoans (Bergmann and Steinert, 2015;Harding et al, 2013;Jäger et al, 2014). Especially, infection studies with the soil-dwelling nematode C. elegans and reports of potting soil and compost as sources of Legionnairesí disease strengthen the hypothesis that bacterivorous nematodes could harbor and disseminate L. pneumophila (Brassinga et al, 2010;Casati et al, 2009;Whiley et al, 2011).…”

Tyrosine Phosphorylation and Dephosphorylation in Burkholderia cenocepacia Affect Biofilm Formation, Growth under Nutritional Deprivation, and Pathogenicity

“…Ten larvae were used for each condition, and the experiment was repeated on three independent occasions. Bacterial intralarval survival was achieved as previously described (47). Five larvae were infected as described above, and then after 4 or 17 h of infection, the larvae were anesthetized by cooling at 4°C.…”

Ecosystem Services Provided by the Little Things That Run the World

“…Reared at 37°C, it serves as an excellent model system especially for bacterial infections, which could not be followed well at room temperature. Moreover, as a good correlation exists between virulence of several bacterial species in insect and mammalian models, G. mellonella is increasingly used as an infection model for human pathogens [156]. The red flour beetle Tribolium castaneum is also a popular model, since its development is more representative of other insects than D. melanogaster, a fact reflected in gene content and function.…”