High levels of isotope elimination improve precision and allow individual-based measurements of metabolic rates in animals using the doubly labeled water method

Abstract

Doubly labeled water (DLW) can be used to measure energy expenditure in free-ranging animals, but questions have been raised about its accuracy in different species or contexts. We investigated whether differences in the extent of isotope elimination affects the precision and accuracy of the DLW method, which can vary according to the experimental design or metabolic rate of the species. Estimated total energy expenditure by the DLW method (TEEdlw) was compared with actual total energy expenditure simultaneously measured via respirometry (TEEresp) in streaked shearwaters Calonectris leucomelas, a pelagic seabird. Subjects were divided into three groups with different experimental conditions: at rest on the ground for 24 h (Group A) or for 48 h (Group B), and at rest on the water for 24 h (Group C). TEEdlw in Group A matched TEEresp, whereas there was an overestimation of TEEdlw in both Groups B and C compared with TEEresp. However, compared with Group A, TEEdlw in Groups B and C had reduced the isotopic analytical variability and thus higher precision. The best regression model (TEEdlw = 1.37 TEEresp − 14.12) showed a high correlation (R2 = 0.82) between TEEdlw and TEEresp and allows a correction factor for field metabolic rates in streaked shearwaters. Our results demonstrate that the commonly made assumption that the DLW method is not appropriate for individual-based estimates may be incorrect in certain circumstances. Although a correction factor may be necessary when using the DLW method to estimate metabolic rate, greater levels of isotope eliminations provides DLW estimates with high precision, which can adequately represent relative individual estimates. Nevertheless, the DLW method, should be used with caution when characterizing interspecies difference of energy expenditures.

Field and laboratory metabolism and thermoregulation in rhinoceros auklets

“…The method allowed an estimation of initial isotope enrichment from a single blood sample and was a less invasive technique with lower impact on the behavior of study subjects ( Schultner et al, 2010 ; Niizuma & Shirai, 2015 ). Recent validation studies have demonstrated that the precision of the DLW technique can be increased by using a longer sampling interval and/or by applying it to a species with a higher metabolic rate ( Shirai et al, 2015 ; Kume et al, 2019 ). The DLW injectate used in our study contained 21.0 atom percent 18 O, 10.5 atom percent 2 H, and 0.9% NaCl.…”

“…We diluted the serum and injectate samples with distilled water measured with an electronic balance (Mettler-Toledo, Columbus, OH, USA) to the nearest 0.01 mg. We analyzed the 2 H and 18 O isotope concentrations of the serum, DLW injectate, and distilled water using isotope ratio mass spectrometry (IRMS; Hydra 20-20, Sercon, Crewe, UK; Shirai et al, 2012 ; Shirai et al, 2015 ). We used the water equilibration method ( Horita et al, 1989 ) to analyze the serum, DLW injectate, and distilled water in duplicate.…”

“…Oxygen consumption rate (Vo 2 ) was measured using an open-flow respirometry system composed of an acrylic metabolic chamber and an oxygen analyzer (Xentra 4100, Servomex Ltd, UK) as previously described in Shirai et al (2015). For the measurement of RMR in air, a 20-L metabolic chamber (20 cm long ×25 cm high ×40 cm wide) was submerged in a thermostatic water bath and maintained at 4.7-20.7 • C. For the measurement of RMR on water, a 72-L metabolic chamber (30 cm long ×60 cm high ×40 cm wide) was filled with freshwater (to a depth of 30 cm) maintained at 5.5-17.5 • C.…”

Peer Review #3 of "Field and laboratory metabolism and thermoregulation in rhinoceros auklets (v0.3)"

“…The fieldwork was permitted by the Ministry of the Environment (21-26-0291 0292, 21-27-0367 0368 0369 0370 0371, 21-28-0344 035 036 037) and the Agency of Cultural Affairs (26-4-2188, 27-4-1928, 29-4-18). Measurements of resting metabolic rate using respirometry in air and on water Oxygen consumption rate (Vo 2 ) was measured using an open-flow respirometry system composed of an acrylic metabolic chamber and an oxygen analyzer (Xentra 4100, Servomex Ltd, UK) as previously described in Shirai et al (2015). For the measurement of RMR in air, a 20-L metabolic chamber (20 cm long × 25 cm high × 40 cm wide) was submerged in a thermostatic water bath and maintained at 4.7-20.7℃.…”

“…The method allowed an estimation of initial isotope enrichment from a single blood sample and was a less invasive technique with lower impact on the behavior of study subjects (Schultner et al, 2010;. Recent validation studies have demonstrated that the precision of the DLW technique can be increased by using a longer sampling interval and/or by applying it to a species with a higher metabolic rate (Shirai et al, 2015;Kume et al, 2019). The DLW injectate used in our study contained 21.0 atom percent 18 O, 10.5 atom percent 2 H, and 0.9% NaCl.…”