Toxigenic effects of two benthic diatoms upon grazing activity of the sea urchin: morphological, metabolomic and de novo transcriptomic analysis

Abstract

Diatoms are unicellular algae playing a key role as photosynthetic organisms in the world’s ocean food webs. The chemical ecology of planktonic diatoms is well documented, but few studies have reported on the effects of benthic diatoms on their consumers, also due to difficulties in the collection, quantification and massive culturing of benthic species. Here for the first time we investigate the effects of feeding on two abundantly occurring benthic diatoms, Nanofrustulum shiloi and Cylindrotheca closterium, isolated from the leaves of the seagrass Posidonia oceanica, on the sea urchin Paracentrotus lividus. Adult P. lividus were fed for one month on diets of either one of the two diatoms and on the green alga Ulva rigida, used as a feeding control. By combining morphological, metabolomic and de novo transcriptomic approaches, we demonstrate toxigenic effect on embryos generated by females fed with these benthic diatoms. Furthermore, chemical analysis reveal the presence of polyunsaturated aldehydes only for N. shiloi, and a high production of other oxylipins (cytotoxic compounds on their grazers and on cancer cell lines) for both diatoms, including some additional peaks not correlated to the canonic oxylipins commonly observed in planktonic diatoms. These findings open new perspectives in the study of diatom secondary metabolites influencing their grazers.

Lipoxygenase Pathways in Diatoms: Occurrence and Correlation with Grazer Toxicity in Four Benthic Species

“…In agreement with the detrimental role of diatom LOX on marine grazers, survival of sea urchins plutei inversely correlate with levels of oxylipins and FAHs in C. closterium, N. shiloi, C. scutellum. By using the three best fits for 17 feeding experiments (Supporting Material, Figure S3) [31,32], these species cluster in separate groups that are fully consistent with their negative effects on sea urchin embryos after diatom ingestion (Figure 4). In general, the higher the quantity of oxylipins and FAHs in the benthic species, the higher the number of abnormal plutei spawned Table S4 of the Supplementary Material.…”

“…The quantities of FAHs and oxylipins measured through LC-MS analyses were correlated with biological assays of feeding experiments with the sea urchin P. lividus [31,32]. Specifically, the percentage of abnormal plutei deriving from adult sea urchins fed for one month with the four benthic diatoms (Supporting Material, Figure S2) were analyzed together with the amount of oxylipins and FAHs found in the present study.…”

“…Figure S1: GC-MS profile of N. shiloi extract with indication of PUAs. Figure S2: Bioassays reported in Ruocco et al [31,32] (a) Percentage of abnormal plutei spawned from sea urchins fed with U. rigida (CTRL) and the four benthic diatoms (TREATED); (b) Examples of normal and malformed plutei observed in CTRL and TREATED conditions, respectively. Figure S3: Effects on embryos of sea urchin P. lividus by feeding experiments with the four benthic diatoms.…”

“…The selected diatom strains were produced in axenic conditions in thermostatic chambers and further collected and incorporated into a 2% agar matrix to produce a diatom enriched feed simulating leaves of P. oceanica with diatom epiphytes that represent a common food for sea urchins. Sea urchins were fed ad libitum [31,32] and feed residuals were removed every day along with sea urchin's fecal pellets. After 30 days, the treated animals were sacrificed and their gametes were collected to be used for in vitro fertilization.…”

“…Recently, we reported that feeding on four benthic species, namely Cylindrotheca closterium, Nanofrustulum shiloi, Diploneis sp. and Cocconeis scutellum, affected reproduction in the sea urchin Paracentrotus lividus [31,32].…”