Ovarian fluid is associated with distinct sperm RNA profiles and enhanced swimming performance in a teleost fish.

In brief: For externally fertilizing species, the composition of the fertilization environment, including the presence of female reproductive fluids, has a significant influence on sperm performance. This study integrated sperm performance analyses with transcriptomics to investigate the molecular basis of environment × gamete interactions and how ovarian fluid is associated with changes in sperm function and RNA profiles. Abstract: For most externally fertilizing teleost fish, sperm remain immotile until release into the external environment, where physicochemical cues trigger motility. In many species, including blue catfish (Ictalurus furcatus), ovarian fluid (OF) present at spawning further enhances sperm swimming performance, enabling more cells to swim longer and at higher velocities than in water alone. This study aimed to identify kinematic performance and RNA profiles associated with sperm-environment interactions under three conditions: (i) sperm activated with 25% OF and 75% deionized water (dH2O) (OF-enhanced); (ii) sperm combined with 100% OF (inactive); and (iii) sperm activated with 100% dH2O (control). We utilized computer-assisted sperm analysis (CASA) in combination with RNA-sequencing to examine sperm kinematics and transcript abundance between sperm environments. The CASA results showed that sperm activated in OF (OF-enhanced) had increased sperm motility and velocity compared with sperm activated in water alone (control). Variance component analysis further identified a male × female interaction associated with variation in sperm swimming performance. RNA-sequencing revealed distinct transcript abundance patterns among sperm environments and also revealed female-associated variation in transcript abundance, providing evidence for gamete level variation on sperm performance. Functional enrichment analysis further revealed that abundant transcripts in the OF treatments were associated with cytoskeletal organization, signaling, large multiprotein complexes, and membrane-associated processes. This study provides novel observations of OF-associated sperm RNA profiles and enhanced swimming performance in a teleost fish, generating hypotheses regarding underlying molecular mechanisms of sperm activation and reaction to environmental stimuli.