Rearing the Fruit Fly &lt;em&gt;Drosophila melanogaster&lt;/em&gt; Under Axenic and Gnotobiotic Conditions

Koyle, Melinda L.; Veloz, Madeline; Judd, Alec M.; Wong, Adam C. N.; Newell, Peter D.; Douglas, Angela E.; Chaston, John M.

doi:10.3791/54219-v

Cited by 28 publications

(16 citation statements)

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“…Differences in the established gut communities of the cohorts of flies that we used could explain some of the variation in our results (41). Using germ-free flies would eliminate these effects (52). Since our flies were housed in a laboratory setting, their diets and microbiomes do not necessarily reflect wild conditions (40), which could also impact the success of Orbaceae relative to other gut-associated species of bacteria.…”

Section: Discussionmentioning

confidence: 99%

“…Fly stocks were reared on Formula 4-24 Instant Drosophila Medium (Carolina Biological Supply Company, Burlington, NC). For experiments, stocks were swapped to a yeast-glucose agar (YGA) diet containing brewer’s yeast, D-glucose, agar, and water (52). Nystatin (10 μg/mL) and kanamycin (10 μg/mL) were added to the diet where specified to prevent the growth of fungal contaminants and ensure the maintenance of engineered strains, respectively.…”

Section: Methodsmentioning

confidence: 99%

“…Fly stocks were reared on Formula 4-24 Instant Drosophila Medium (Carolina Biological Supply Company, Burlington, NC). For experiments, stocks were swapped to a yeastglucose agar (YGA) diet containing brewer's yeast, D-glucose, agar, and water (52).…”

Section: Growth and Maintenance Of Bacterial Strains Escherichia Coli...mentioning

confidence: 99%

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The Pathfinder plasmid toolkit for genetically engineering newly isolated bacteria enables the study ofDrosophila-colonizingOrbaceae

Elston

Phillips

Leonard

et al. 2023

Preprint

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Toolkits of plasmids and genetic parts streamline the process of assembling DNA constructs and engineering microbes. Many of these kits were designed with specific industrial or laboratory microbes in mind. For researchers interested in non-model microbial systems, it is often unclear which tools and techniques will function in newly isolated strains. To address this challenge, we designed the Pathfinder toolkit for quickly determining the compatibility of a bacterium with different plasmid components. Pathfinder plasmids combine three different broad-host-range origins of replication with multiple antibiotic resistance cassettes and reporters, so that sets of parts can be rapidly screened through multiplex conjugation. We first tested these plasmids inEscherichia coli, a strain ofSodalis praecaptivusthat colonizes insects, and aRosenbergiellaisolate from leafhoppers. Then, we used the Pathfinder plasmids to engineer previously unstudied bacteria from the familyOrbaceaethat were isolated from several fly species. EngineeredOrbaceaestrains were able to colonizeDrosophila melanogasterand could be visualized in fly guts.Orbaceaeare common and abundant in the guts of wild-caught flies but have not been included in laboratory studies of how theDrosophilamicrobiome affects fly health. Thus, this work provides foundational genetic tools for studying new host-associated microbes, including bacteria that are a key constituent of the gut microbiome of a model insect species.

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Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

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The Pathfinder plasmid toolkit for genetically engineering newly isolated bacteria enables the study ofDrosophila-colonizingOrbaceae

Elston

Phillips

Leonard

et al. 2023

Preprint

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“…Preliminary tests suggested that the HS microbiome had much lower bacterial abundance, and so the C microbiome was resuspended in 6 ml sterile water and 3 ml for the HS microbiome. 50 µl of the frass wash was used to inoculate axenic eggs in sterile diets following standard protocols [20], and flies were reared in a biosafety cabinet at 25ºC and 12:12 light:dark cycle through the lifespan of the recipient flies. At 7-10 days post eclosion, recipient flies were stored at -80ºC and DNA subsequently extracted from individuals.…”

Section: Transplanting Microbiomes To Test For Adaptive Significancementioning

confidence: 99%

“…After hatching, larvae acquire their microbes through a combination of microbial persistence and colonization and form associations over the lifespan of the fly [17][18][19]. Many fly-associated microbes can be individually cultured and reconstituted in various combinations to inoculate sterile flies [14,20]. With the feasibility of microbiome manipulations and rich genetic resources, Drosophila is an ideal model to study the interplay between host genetic, microbiome, and environment (i.e., GH x GM x E interactions) in shaping host phenotypes that underlie rapid host adaptation.…”

Section: Introductionmentioning

confidence: 99%

Host evolutionary history and ecological context modulate the adaptive potential of the microbiome

Henry

Fernández

Ayroles

2020

Preprint

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The microbiome may serve as a reservoir of adaptive potential if hosts can leverage microbial adaptations in stressful environments. However, the facilitation of rapid host adaptation may be limited by interactions between host genotype, microbiome, and environment (GH x GM x E). Here, to understand how host x microbiome x environment interactions shape adaptation, we leverage >150 generations of experimental evolution in Drosophila melanogaster in a stressful environment, high sugar (HS) diet. The microbiome of HS adapted flies shifted to be dominated by one bacteria, Acetobacter pasteurianus. We next performed a fully reciprocal transplant experiment using the dominant control and HS bacteria to measure life-history and metabolic traits in flies. Mismatches between fly evolution and microbiome exerted fitness costs by slowing development and reducing fecundity, especially in the stressful HS diet. The GH x GM x E interactions observed suggest that both ecological context and host evolution shape the adaptive potential of the microbiome. We conclude by proposing future directions that highlight the benefit of using experimental evolution to study the contribution of the microbiome to host evolution.

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Wolbachia impacts microbiome diversity and fitness‐associated traits for Drosophila melanogaster in a seasonally fluctuating environment

Henry,

Fernandez,

Wolf

et al. 2024

Ecology and Evolution

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The microbiome contributes to many different host traits, but its role in host adaptation remains enigmatic. The fitness benefits of the microbiome often depend on ecological conditions, but theory suggests that fluctuations in both the microbiome and environment modulate these fitness benefits. Moreover, vertically transmitted bacteria might constrain the ability of both the microbiome and host to respond to changing environments. Drosophila melanogaster provides an excellent system to investigate the impacts of interactions between the microbiome and the environment. To address this question, we created field mesocosms of D. melanogaster undergoing seasonal environmental change with and without the vertically transmitted bacteria, Wolbachia pipientis. Sampling temporal patterns in the microbiome revealed that Wolbachia constrained microbial diversity. Furthermore, Wolbachia and a dominant member of the microbiome, Commensalibacter, were associated with differences in two higher‐order fitness traits, starvation resistance and lifespan. Our work here suggests that the interplay between the abiotic context and microbe–microbe interactions may shape key host phenotypes that underlie adaptation to changing environments. We conclude by exploring the consequences of complex interactions between Wolbachia and the microbiome for our understanding of eco‐evolutionary processes that shape host‐microbiome interactions.

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Rearing the Fruit Fly <em>Drosophila melanogaster</em> Under Axenic and Gnotobiotic Conditions

Cited by 28 publications

References 0 publications

The Pathfinder plasmid toolkit for genetically engineering newly isolated bacteria enables the study ofDrosophila-colonizingOrbaceae

The Pathfinder plasmid toolkit for genetically engineering newly isolated bacteria enables the study ofDrosophila-colonizingOrbaceae

Host evolutionary history and ecological context modulate the adaptive potential of the microbiome

Wolbachia impacts microbiome diversity and fitness‐associated traits for Drosophila melanogaster in a seasonally fluctuating environment

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