Development of Oncomelania hupensis quadrasi embryonic cell primary culture

Oncomelania hupensis quadrasi is the obligate intermediate host of Schistosoma japonicum in the Philippines and is central to sustaining environmental transmission. Although its distribution and infection dynamics have been well characterized, parasite-host interactions at the cellular level remain largely unexplored due to the lack of suitable in vitro systems. This study describes the establishment and optimization of primary embryonic cell cultures of O. h. quadrasi embryonic cell primary culture, providing foundational tools for cellular and molecular investigations. Morphologically identified embryos at the onset of shell formation were enzymatically dissociated using collagenase, and trypsin-EDTA and maintained under a range of conditions, including varying osmolalities (∼125-275 mOsm/kg H₂O), basal media (Roswell Park Memorial Institute medium and Schneider's Insect Medium), and nutritional supplements, particularly fetal calf serum (FCS). Culture performance was assessed by trypan blue exclusion and resazurin and XTT reduction assays. Culture conditions using Schneider's Insect Medium (SIM) supplemented with lactalbumin hydrolysate (LH) and 20% fetal calf serum (FCS) at approximately 200 mOsm/kg H₂O and 27 °C supported the maintenance of viable embryonic cell populations and metabolic activity. Under minimally disturbed conditions, viable cell populations were maintained for up to approximately 30 days. Although this observation was derived from limited biological replication, it establishes experimental conditions that warrant independent validation. Increases in viable cell counts were observed during early culture but did not correspond to doubling of total cell numbers, and no direct evidence of cell division was obtained. These findings define culture conditions that supported the establishment and maintenance of O. h. quadrasi embryonic cell populations under the experimental conditions evaluated. Further studies incorporating increased biological replication and orthogonal assays for cell division are needed to confirm reproducibility and characterize long-term culture performance. This work provides a methodological foundation for the development of species-specific in vitro systems to support schistosomiasis and molluscan biology research.