The Effect of Microbiota on Lifespan in Drosophila melanogaster
Faculty Mentor: John Chaston, Plant and Wildlife Sciences
Almost all animals are affected by the bacteria found in their intestines (1). Recent research has begun to show the close relationship hosts have with their microbiome. These interactions influence metabolic, respiratory, nutritional, neurological, and immunological functions along with many other biological roles (2). The short lifespans, fruit flies make an excellent model organism for studying host/microbiome interactions (3). By better understanding the correlation between lifespan and host microbiota in D. melanogaster, I hope to help further the knowledge of the relationship between lifespan in humans and our own microbiome.
The Drosophila melanogaster lines were fed a yeast-glucose diet. Experimental flies were placed on 7.5 ml of sterile diet prepared in a 50 ml conical tube. The flies were cultured at 25⁰ C on a 12-hr light, 12-hr dark cycle.
Bacteria was grown on either modified MRS medium, potato medium, luria broth, or brain heart infusion. All cultures were grown at 30⁰ C in either aerobic (oxygenated by shaking) or anaerobic (grown static) conditions.
Preparation of axenic and gnotobiotic flies
Fruit fly eggs (≤20 hr old) were collected and rinsed with a 0.6% hypochlorite solution for two 2.5 minute washes. These were followed by three rinses with sterile water. After the rinses, the eggs were transferred to sterile diet within a biosafety cabinet. Flies that were left bacteria free are referred to as ‘axenic’. Other flies were inoculated with 1 of 42 different bacterial strains. Flies with a known microbiome are referred to as ‘gnotobiotic’.
Every 2-3 days, Drosophila was transferred to new sterile diet. The number and sex of dead flies was recorded at each transfer. The spent vials were incubated at room temperature until the eggs laid during the 2-3 day interval grew to adulthood. These flies were then homogenized and plated to check for bacterial contamination and persistence.
Meta-genome Wide Association
A meta-genome wide association study was run to predict bacterial genes that influenced lifespan. The pipeline organized similar genes found across all bacterial species into clusters of orthologous groups (COGs). Each group contained bacteria that had a similar effect on lifespan and contained a gene in common.
Figure 1: The average number of days that fruit flies survived with different bacteria types.
We found that the presence of bacteria in Drosophila melanogaster does have an effect on lifespan. Overall, microbial presence decreased lifespan when compared to micro-organism free flies. This decrease in lifespan showed variation within and across bacterial species (Figure 1) . The metagenome-wide association study gave a list of possible bacterial genes that could be responsible for the influence on lifespan. Out of 13,000 COGs, 12 significantly impacted lifespan. Among the most significant hits were genes involving vitamin B2, vitamin B12, and methionine metabolism. When 10 bacterial mutants were tested, three showed a significant impact on lifespan (Figure 2). The results strongly suggest that bacterial methionine metabolism has a role in mediating Drosophila lifespan.
Figure 2: The average number of days that flies survived with bacterial
The lifespan of Drosophila melanogaster is greatly influenced by the time of life and length of exposure to bacteria. Early exposure (first week of life) can increase lifespan while bacterial presence later in life can have deleterious effects (4). This effect must be considered when comparing different bacteria’s effect on longevity. The weekly homogenizations were able to provide data on how long the flies stayed associated with the bacteria. The length of persistence was taken into account when processing the data.
The use of a meta-genome wide association study provided a way for our lab to efficiently identify bacterial genes that influence Drosophila lifespan. This method allowed us to compare individual bacterial genes that may be not be shared within a family but can be found across different families. Only 10 mutants needed to be tested in order to confirm our predictions.
Overall, we were able to show that the presence of bacteria significantly decreased the lifespan of Drosophila melanogaster. This decrease was dependent on the bacterial strain as well as the length of time that the fly remained associated with the bacteria. These results suggest that it is possible to manipulate lifespan by simply changing the gut microbiota of the fruit fly.