Freshwater insects across various feeding guilds face the same dietary nutrient limitations (particularly nitrogen, N) as terrestrial insects. Thus, gut microbial N-provisioning functions have been proposed to be relevant for freshwater insects. One possible mode of microbial N-provisioning in freshwater insects is nitrate reduction to ammonium; nitrate is readily available in many streams. Microbial assimilatory nitrate reduction to ammonium (ANRA) and dissimilatory nitrate reduction to ammonium (DNRA) may provide insect host with N in the form of ammonium. However, these processes compete with denitrification (conversion of nitrate into nitrous oxide or dinitrogen). In this study, we investigated the potential for nitrate reduction to ammonium by gut microbiomes of freshwater insects from various taxa and feeding guilds, from an impaired, urban stream (Tinker’s creek) and an unimpaired stream (West Branch of the Mahoning) in Northeastern Ohio via determining the presence and expression of microbial genes involved in these processes. We sought to determine if the presence, expression, and abundance of microbial genes differed among insect feeding guilds in the same stream and/or between streams.
Overall, 22 macroinvertebrate taxa distributed into 5 feeding guilds in Tinker’s creek and 4 feeding guilds in West Branch were obtained, with more bioindicator species detected in West Branch. In Tinker’s creek, the most abundant insect families were Chironomidae (filter and deposit feeder), Hydropsychidae (filter and deposit feeder), and Elmidae (grazer and scraper). In West Branch, the most abundant families were Chironomidae (filter and deposit feeder), Elmidae (grazer and scraper), and Baetidae (grazer and scraper). Analysis of 16S rRNA gene sequences from gut microbial assemblages from the 22 freshwater insect families from both streams revealed significant differences among feeding guilds and dietary materials, suggesting diet-driven differences in gut microbial community composition. These differences may underscore differences in feeding guild specific microbial nitrate reduction processes. Microbial denitrification genes (nirK, and nosZ) have been detected in almost all sampled insect taxa from both streams, and microbial genes for ANRA (nasB) and DNRA (nrfA) have been detected in some of the sampled insect taxa, suggesting potential function. Ongoing work is focused on confirming the presence of microbial genes for ANRA and DNRA in new and remaining insect samples from both streams and quantifying expression of these genes using RNA extracted from insects used in nitrate reduction assays.