Rearing the Fruit Fly Drosophila melanogaster Under Axenic and Gnotobiotic Conditions

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

doi:10.3791/54219

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Method Article

Rearing the Fruit Fly Drosophila melanogaster Under Axenic and Gnotobiotic Conditions

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DOI:

10.3791/54219

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July 30th, 2016

Melinda L. Koyle¹ , Madeline Veloz¹ , Alec M. Judd¹ , Adam C.-N. Wong²^,⁴ , Peter D. Newell²^,⁵ , Angela E. Douglas²^,³ , John M. Chaston¹^,²

¹Department of Plant and Wildlife Sciences, Brigham Young University, ²Department of Entomology, Cornell University, ³Department of Molecular Biology and Genetics, Cornell University, ⁴Division of Infectious Diseases, Boston Children's Hospital, Harvard Medical School, ⁵Biological Sciences, SUNY Oswego

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Summary

A method for rearing Drosophila melanogaster under axenic and gnotobiotic conditions is presented. Fly embryos are dechorionated in sodium hypochlorite, transferred aseptically to sterile diet, and reared in closed containers. Inoculating diet and embryos with bacteria leads to gnotobiotic associations, and bacterial presence is confirmed by plating whole-body Drosophila homogenates.

Abstract

The influence of microbes on myriad animal traits and behaviors has been increasingly recognized in recent years. The fruit fly Drosophila melanogaster is a model for understanding microbial interactions with animal hosts, facilitated by approaches to rear large sample sizes of Drosophila under microorganism-free (axenic) conditions, or with defined microbial communities (gnotobiotic). This work outlines a method for collection of Drosophila embryos, hypochlorite dechorionation and sterilization, and transfer to sterile diet. Sterilized embryos are transferred to sterile diet in 50 ml centrifuge tubes, and developing larvae and adults remain free of any exogenous microbes until the vials are opened. Alternatively, flies with a defined microbiota can be reared by inoculating sterile diet and embryos with microbial species of interest. We describe the introduction of 4 bacterial species to establish a representative gnotobiotic microbiota in Drosophila. Finally, we describe approaches for confirming bacterial community composition, including testing if axenic Drosophila remain bacteria-free into adulthood.

Introduction

Most animals are intimately associated with bacteria ('microbiota') from birth to death¹. Comparisons of microorganism-free ('axenic') and microorganism-associated ('conventional') animals have shown microbes influence diverse aspects of animal health, including metabolic, nutritional, vascular, hepatic, respiratory, immunological, endocrine, and neurological function². The fruit fly Drosophila melanogaster is a key model for understanding many of these processes in the presence of microbes^3,4 and for studying microbiota influence on animal health^5,6. No bacterial species is present in every ....