Effects Of Conspesific Cues And Bacterial Biofilms On Settlement Of Ulva Intestinalis Spores

The marine seaweed Ulva is the important fouling macroalgae and the main green tide-forming algae,causing serious economic and ecological problems.In the marine environment,Ulva usually forms dense aggregation.Settlement of spores plays an important role in Ulva aggregation and is the key stage in its life cycle,which changes the life phase from planktonic(spores)into sessile form.Settlement of larvae of marine benthic invertebrates is usually suggested to be guided by the habitat-specific cues,but so far there is only a few of reports on cues for settlement of macroalgae.In present study,we explored cues on settlement of Ulva intestinalis spores,which may be released by conspecific fronds(interspecies)or bacteria(interspecific)because both of them existed in high abundance in habitats of U.intestinalis.The fronds of U.intestinalis were extracted using water and organic solvents and subsequently tested for their activity on settlement of U.intestinalis.The compounds active in inducing settlement of spores were analysed,purified and identified.To understand the role of bacteria on settlement of spores,the bacterial community associated with U.intestinalis were analyzed and the effect of biofilm of bacteria isolates were also tested for their activity on settlement of spores.The main results were as the followings.1.The extracts of U.intestinalis fronds by organic solvents were found to contain chemical cues that significantly induce settlement of spores.By bioassay-guided isolation,fatty acid C16:4(hexadeca-4,7,10,13-tetraenoic acid)was isolated from the hexane extract.This fatty acid significantly induced settlement of U.intestinalis spores at the concentration as low as 5 ?g mL-1.Another chemical compound was successfully isolated from the dichloromethane extract,which was identified as dibutyl phthalate.That compound was also active in inducing settlement of spores at 0.5?10 ?g mL-1.2.The analysis on composition of fatty acid related compounds in active fractions(HU,FI,and FII)from U.intestinalis showed that they all contained saturated fatty acids,monounsaturated fatty acids and polyunsaturated fatty acids.Fatty acids from 8 carbon atoms to 22 carbon atoms in chain length were found.Furthermore,seven fatty acids(including six fatty acids founded in U.intestinalis)were tested for their activity on settlement of U.intestinalis spores.The effect of fatty acids on settlement of spores was specific to fatty acid.The effect fatty acid on settlement of spores depends on a combination of chain length and the degree of unsaturation.Fatty acids C16:1,C16:4 and C18:2 were found to significantly enhance settlement of U.intestinalis spores,indicating they may be the natural inducers from conspecific fronds for controlling settlement and aggregation of spores.This the first report about chemical cues from conspecific fronds to guide the settlement of Ulva spores.3.Analysis of bacterial community structure associated with U.intestinalis from Xiamen coastline showed the dominance of phyla Proteobacteria,Bacteriodetes and Cyanobacteria.The result of hightrouput sequencing analysis revealed that bacteria community on algae surface and surrounding rock surface were different in class levels.As much as 35 strains of bacteria were isolated from algae surface and surrounding rock surface.Three isolates(Hyunsoonleella pacifica,Erythrobacter vulgaris and Shewanella loihica)significantly induced settlement and aggregation of U.intestinalis spores.Analysis of bacteria community revealed that Hyunsoonleella and Erythrobacter were in high abundance associated with U.intestinalis.It was found here that the bacteria from the U.intestinalis habitat could induce settlement and aggregation of U.intestinalis spores.