Insect flight metabolic rate revealed by bolus injection of the stable isotope (13)C

Measuring metabolic rate (MR) poses a formidable challenge in free-flying insects who cannot breathe into masks or be trained to fly in controlled settings. Consequently, flight MR has been predominantly measured on hovering or tethered insects flying in closed systems. Stable isotopes such as labelled water allow measurement of MR in free-flying animals but integrates the measurement over long periods exceeding the average flight duration of insects. Here, we applied the ‘bolus injection of isotopic (13)C Na-bicarbonate’ method to insects to measure their flight MR and report a 90% accuracy compared to respirometry. We applied the method on two beetle species, measuring MR during free flight and tethered flight in a wind tunnel. We also demonstrate the ability to repeatedly use the technique on the same individual. Therefore, the method provides a simple, reliable and accurate tool that solves a long-lasting limitation on insect flight research by enabling the measurement of MR during free flight.