Circadian regulation of reproduction and neoblast cell division in the catenulid flatworm <i>Stenostomum virginianum</i> (Platyhelminthes)

The molecular circadian clock and melatonin secretion are pivotal for coordinating homeostasis pathways in many animals. Melatonin has been shown to regulate reproduction and neoblast proliferation in triclad flatworms, yet the extent to which these mechanisms operate in other platyhelminths remains unknown. We examined light sensing, endogenous melatonin production, and circadian control of asexual reproduction in the early-branching catenulid Stenostomum virginianum. Microscopy revealed refractile bodies with photoreceptor-like structures, blue-light autofluorescence, and PAX6 immunoreactivity in cells anterior to the brain that support the presence of a photoreceptive system. High-performance liquid chromatography detected endogenous melatonin production in S. virginianum. Exogenous melatonin administered under continuous darkness suppressed reproduction and neoblast mitosis. The same treatment applied at ZT 6 during a 12 h:12 h light-dark cycle failed to affect reproduction, implying a masking or circadian compensatory effect. Phylogenetic analysis identified two broadly conserved bHLH-PAS genes (ARNT) in a transcriptome for S. virginianum; although CLOCK and CRY were not detected, they are present in the closely related Macrostomum lignano, indicating that this absence may be due to an incomplete transcriptome. These findings suggest that circadian rhythms influence reproduction and cell division in S. virginianum, extending our understanding of melatonin-mediated, light-responsive regulation in distinct branches of the platyhelminths.