Amplicon sequencing of ITS2 region of (1) fecal material of Drosophila melanogaster raised in fruit and vegetable-based microcosms perpetuating field collected microbes (n=26); (2) Pooled ontogenetic environment samples of 10 vials harboring a single fly until eclosion for three different substrates (n=9); (3) Samples of fecal material from flies on laboratory diet and flies transferred to fruit from laboratory diet, as well as blanks are included for comparison.

Microcosms based on different breeding substrates of Drosophila melanogaster (apple, banana, grape, plum, lemon, onion, tomato) were set up from pieces of substrate colonized by insect associated microbes in the field (Riedel & Rohlfs 2025, in preparation). Microbes were maintained by D. melanogaster through ingestion and reinoculation by fecal droplet deposition on fresh substrates provided ad libitum. DNA was extracted from fecal material of groups of female flies, and from substrates after fly development in an experimental set-up. Fungal amplicon regions were used to identify fly associated microorganism from said fecal material using the primers ITS3_KYO2 (forward) and ITS4_KY03 (reverse) covering the Internal spacer region two (ITS2, Toju et al., 2012). The raw demultiplexed sequences were created on an Illumina MiSeq v3.0 (2 × 300-bp) run conducted at Advanced Identification Methods (AIM), Leipzig, Germany. For more detailed description of the datasets see Riedel et al. 2025 (https://doi.org/10.1101/2025.10.14.682149) and Riedel & Rohlfs 2025 in preparation.