Stress Response of the Juvenile Tri‐Spine Horseshoe Crab <i>Tachypleus tridentatus</i> to Hypoxia: Insights From Phenotypic, Metabolomic, and Microbial Analyses

Abstract

Over the past few decades, ocean hypoxia has been increasing due to human activities. Hypoxic stress, characterized by a reduced level of dissolved oxygen, is an escalating threat to marine ecosystems, with potentially devastating effects on the viability of endangered species such as the tri-spine horseshoe crab Tachypleus tridentatus. Even though this species is remarkably resilient to low oxygen levels, persistent hypoxia can negatively impact its population's survivability. The objective of this research was to examine the impact of short-term hypoxia on the behavior, gut microbiota, and metabolomics of juvenile T. tridentatus. First instar juvenile horseshoe crabs were subjected to hypoxic stress (2 mg O₂/L) for 14 days and then recovered for 7 days in an environment with normal dissolved oxygen. The findings demonstrated that short-term hypoxia reduced the rates of swimming and burrowing of horseshoe crabs, and induced the change of tissue metabolites and intestinal flora malfunction. Additionally, in the hypoxia groups on day 14, 86 distinct metabolites showed a trend of downregulation, while 29 metabolites showed an upregulation trend. Arginine biosynthesis; histidine metabolism; vitamin B6 metabolism; aminoacyl-tRNA biosynthesis; and alanine, aspartate, and glutamate metabolism were the top five metabolic KEGG pathways (p < 0.05) enriched with 8 metabolites. In conclusion, our results provided new insights related to the behaviors, microbiota, and metabolites involved in juvenile T. tridentatus exposed to short-term hypoxic conditions and confirmed that hypoxia impairs their behavioral and physiological status.