Bath-Immersion Delivery of nanos-MO-Vivo During Embryogenesis Perturbs Gonadal Development in the Pacific Oyster (Crassostrea gigas)

Abstract

The regulation of germline-associated processes during early embryogenesis is an important aspect of gonadal development in aquaculture species. Here, we employed bath-immersion delivery of a nanos-targeting Morpholino-Vivo (nanos-MO-Vivo) during embryogenesis to transiently disrupt early germline development in the Pacific oyster Crassostrea gigas. Post-fertilization immersion with 100 µM nanos-MO-Vivo for 6 h achieved effective repression of germline markers while maintaining acceptable embryo viability. Transcriptomic analyses at the blastula stage revealed coordinated downregulation of PGC markers (vasa, Piwil1) and germline-associated regulatory pathways. Long-term rearing revealed that transient embryonic perturbation of nanos resulted in persistent impairment of adult gonadal development, characterized by reduced gonadal coverage and diminished gametogenic tissue. In silico analysis based on single-cell transcriptomic data further supported nanos-associated regulatory effects within the PGC population. High sequence conservation of nanos among Crassostrea species was accompanied by comparable knockdown responses in C. angulata and C. nippona, suggesting the observed effects are not restricted to a single oyster species. This study links transient embryonic perturbation of germline-associated regulation to long-term gonadal phenotypes, providing insight into reproductive modulation in molluscan aquaculture.