Environmental Quality Assessment Using Fecal Metabolomics in Waterfowl from Protected Wetlands in Southwest Spain

The study of environmental stress requires an understanding of biological responses to pollutants, which can be difficult to interpret due to multiple influencing factors. This study investigates the fecal metabolome of waterfowl as bioindicators of environmental quality in two protected wetlands in southwestern Spain: the Odiel Marshes Biosphere Reserve (OMBR) and the Cádiz Bay Important Bird Area (CBIBA). Using untargeted UHPLC-QTOF-MS metabolomics, fecal samples were analyzed from five species: spoonbills, black-headed gulls, yellow-legged gulls, lesser black-backed gulls, and storks. Significant alterations were observed in key metabolites, including fatty acids, steroids, glycerophospholipids, and sphingolipids, reflecting differences in diet, habitat use, and pollutant exposure. In spoonbills, steroids and fatty acids represented the largest fractions of the detected metabolites, while comparisons between gull species revealed variations in fatty acid and glycerophospholipid levels. Multivariate analyses showed a clear separation between species and sites. Pathway analysis identified several altered metabolic routes, primarily involving fatty acid, bile acid, and sphingolipid metabolism. Overall, these results demonstrate that fecal metabolomics is a robust noninvasive tool for assessing pollution-related physiological changes, confirming waterbirds as effective bioindicators of wetland ecosystem health.