Our strategy is to conduct fitness assays on Galleria mellonella larvae reared with a different microbiota status (sterile – “axenic” vs conventional larvae); and to investigate the host response following infection from two different pathogens (Bacillus thuringiensis and Metarhizium anisiopliae). At the GME unit of INRAe, France, I have focussed on bioassays using Bacillus thuringiensis, or Bt. Preliminary results revealed the axenic larvae were more susceptible to orally administered Bt than conventional larvae. At a concentration yielding approximately 98% mortality for axenic larvae, this was only half-lethal for conventional larvae. Moreover, the surviving conventional larvae were able to completely clear Bt from their gut by 96 hours. It was also found that the total life was longer in axenic larvae due to the increased average time to pupation following an early half-lethal infection of Bt compared to larvae with an intact microbiota. High variability in the time of pupation was also reported for axenic larvae whereas the majority of conventional larvae pupated within the same four days. There was no significant difference in adult life span. These findings suggest the surviving axenic larvae took longer to recover from the Bt infection resulting in delayed and variable pupation times. To further investigate the partnership between the immune response and the microbiota, immune response genes from the gut and fat body will be examined using qRT-PCR techniques and the composition and dynamics of the gut microbiota will be characterised during pathogen infection using 16S rRNA and the Illumina Miseq platform. At the SOBI laboratory of Plant and Environmental Sciences of the University of Copenhagen, Denmark, I will begin experiments using Metarhizium anisopliae to explore the role of the microbiota during topical infections.
A verification technique to evaluate the sterility of the axenic reared larvae. The entire gut of two axenic and two conventional larvae are dissected, homogenized in sterile water and serially diluted before plating onto LBA. The plates are then checked at 48 hours for growth. There is an absence of growth for axenic larvae at undiluted, 10-1 or 10-2 dilutions due to the sterile rearing conditions compared to conventional larvae which have acquired the microbiota from the diet and environment.