Dipteran larvae and microbes facilitate nutrient sequestration in the Nepenthes gracilis pitcher plant host

The fluid-containing traps of Nepenthes carnivorous pitcher plants (Nepenthaceae) are often inhabited by organisms known as inquilines. Dipteran larvae are key components of such communities and are thought to facilitate pitcher nitrogen sequestration by converting prey protein into inorganic nitrogen, although this has never been demonstrated in Nepenthes. Pitcher fluids are also inhabited by microbes, although the relationship(s) between these and the plant is still unclear. In this study, we examined the hypothesis of digestive mutualism between N. gracilis pitchers and both dipteran larvae and fluid microbes. Using dipteran larvae, prey and fluid volumes mimicking in situ pitcher conditions, we conducted in vitro experiments and measured changes in available fluid nitrogen in response to dipteran larvae and microbe presence. We showed that the presence of dipteran larvae resulted in significantly higher and faster releases of ammonium and soluble protein into fluids in artificial pitchers, and that the presence of fluid microbes did likewise for ammonium. We showed also that niche segregation occurs between phorid and culicid larvae, with the former fragmenting prey carcasses and the latter suppressing fluid microbe levels. These results clarify the relationships between several key pitcher-dwelling organisms, and show that pitcher communities facilitate nutrient sequestration in their host.