In vivo isotopic fractionation of zinc and biodynamic modeling yield insights into detoxification mechanisms in the mayfly Neocloeon triangulifer.

Diversity and biomass of aquatic insects decline in metal-rich aquatic environments, but the mechanisms by which insects from such environments cope with potentially toxic metal concentrations to survive through adulthood are less well understood. In this study, we measured Zn concentrations and isotopes in laboratory-reared diatoms and mayflies (Neocloeon triangulifer) from larval through adult stages. The larvae were fed Zn-enriched diatoms, and bio-concentrated Zn by a factor of 2.5-5 relative to the diatoms but maintained the same Zn-isotopic ratio. These results reflect the importance of dietary uptake and the greater rate of uptake relative to excretion or growth. Upon metamorphosis to subimago, Zn concentrations declined by >70%, but isotopically heavy Zn remained in the subimago bodies. We surmised that the loss of isotopically light Zn during metamorphosis was due to the loss of detoxified Zn and retention of metabolically useful Zn. Through the transition from subimago to imago, Zn concentrations and isotope ratios were virtually unchanged. Because the decrease in Zn body concentration and increase in heavier Zn are seen in the subimagos relative to the larvae, the compartmentalization of Zn must be occurring within the larvae. A biodynamic model was constructed, allowing for isotopic fractionation and partitioning of Zn between metabolically essential and detoxified Zn reservoirs within larvae. The model provides a consistent set of rate and fractionation constants that successfully describe the experimental observations. Specifically, metabolically essential Zn is isotopically heavier and is tightly held once assimilated, and excess, isotopically light Zn is sequestered, detoxified, and ultimately lost during the metamorphosis of larvae to subimagos. To our knowledge, this is the first documentation of in vivo isotopic fractionation in insects, offering an improved understanding of the mechanisms and rates by which the N. triangulifer larvae regulate excess Zn in their bodies.