GC-MS analysis and identification of moult-specific metabolites in the haemolymph of male fiddler crab, Gelasimus hesperiae

Abstract

This study characterises the moult stage-specific haemolymph metabolite profile of the male brachyuran crab Gelasimus hesperiae using gas chromatography-mass spectrometry (GC-MS) to elucidate the metabolite shift across the moulting process. Haemolymph was collected from 24 healthy crabs, comprising eight individuals each at intermoult, premoult [D2], and postmoult stages, and pooled within each stage. A total of 229 metabolites were identified, of which 61, 13, and 85 were exclusively expressed at the intermoult, premoult, and postmoult stages, respectively. Bi-plot analysis revealed substantial variations in carbohydrates, amino acids, fatty acids, nucleic acids, vitamins, and their respective derivatives. Carbohydrates, proteins, and fatty acids declined from intermoult to premoult whereas, major biochemical components or their derivatives elevated during postmoult. Amino acids involved in exoskeleton formation, calcification, and chitin-binding were present in both premoult and postmoult. Non-essential amino acids dominated in intermoult, whereas essential amino acids were higher in postmoult. Arginine, glycine, and L-aspartate remained stable throughout. Intermoult-specific metabolites were primarily linked to amino acid and fatty acid biosynthesis and the methyl farnesoate pathway. Postmoult showed substantial accumulation of fatty acids, glycine/serine/threonine pathways, carbohydrates, vitamins, nucleotide-like molecules, and chitosan biosynthesis. Overall, the results demonstrate clear moult-related metabolic shifts and elucidate the biochemical management of this energy-intensive moulting process. The differential metabolites identified may serve as potential biomarkers of moulting, endocrinological studies, and ecological monitoring of fiddler crabs.