Multispecies protocol for the induction of sporulation in Ulva spp.

Sporogenesis and gametogenesis (later defined as “sporulation”) in Ulva species are important stages of their life cycle. However, the developmental and environmental factors controlling Ulva sporulation are poorly understood. In the present study we evaluated six protocols involving abiotic stresses, i.e. fragmentation, dehydration, chemical stress, nutrient starvation and cold shock, to induce the formation of reproductive tissues in strains from three Ulva species, Ulva lacinulata , Ulva compressa , and Ulva intestinalis . The most successful protocol combined fragmentation, cold shock (4 °C for 1 h) and nutrient depletion, achieving an overall success rate of 80%, across 32 Ulva strains, where U. compressa , U. intestinalis , and U. lacinulata exhibited success rates of 100%, 80% and 70%, respectively. The mean final reproductive tissue percentage (RT%), defined as the maximum percentage of cells within a thallus undergoing sporulation, did not differ significantly among the three species. However, U. compressa , gametophytes ( n = 8) exhibited significantly higher mean RT% than sporophytes ( n = 6), with values of 91.0 ± 8.1% and 66.2 ± 11.4%, respectively. Also, the gametophytes of U. compressa exhibited earlier sporulation onset (day 3) relative to sporophytes (day 6). The difference in duration of sporulation between U. lacinulata and U. compressa was statistically insignificant. Our findings provide new insights into the species and life stage specific responses of Ulva to abiotic stresses, and offer a reliable protocol for inducing reproduction in three Ulva species.