Ecometabolomics reveal physiological adaptations of Asiatic toads (Bufo gargarizans Cantor, 1842) to different environments along an altitudinal gradient.

Animals inhabiting altitudinal gradients exhibit a variety of adaptations to environmental variations. However, to date, changes in metabolomic profiles with altitude have been poorly characterized. Here, we used target and non-target metabolomic analyses of liver to investigate the environmental adaptations of Asiatic toads (Bufo gargarizans) along an altitudinal gradient (50 m, 1200 m, 2300 m, and 3400 m above sea level). Non-targeted metabolomics analysis identified 775 metabolites, and k-means clustering analysis showed that up-regulated metabolites along the altitudinal gradient were significantly enriched in the thiamine and sphingolipid metabolism pathways. Down-regulated metabolites were mainly enriched in alanine, aspartate and glutamate metabolism and glycolysis/glycogenesis. Weighted gene co-expression network analysis showed that metabolites positively correlated with altitude were mainly related to sphingolipid metabolism and glycerophospholipid metabolism, whereas those negatively correlated were involved in glycolysis/gluconeogenesis and glycerolipid metabolism. Moreover, a total of 52 metabolites were identified by the targeted metabolomics analysis. K-means clustering analysis showed that down-regulated metabolites along the altitudinal gradient were mainly enriched in pentose phosphate pathway and glycolysis/gluconeogenesis. In addition, toads from different altitudes exhibited significant variation in the activities of key metabolic enzymes, including phosphofructokinase, lactate dehydrogenase, and α-ketoglutarate dehydrogenase. In conclusion, the metabolic profiles of Asiatic toads along an altitudinal gradient differed significantly. These findings enhance our understanding of the physiological adaptations of toads to different environments along an altitudinal gradient.