Invertebrate Grazing on Live Turtlegrass (Thalassia testudinum): A Common Interaction That May Facilitate Fungal Growth

In coastal wetlands and tropical reefs, snails can regulate foundation species by feeding on marsh grasses and hard corals. In many cases, their impacts are amplified because they facilitate microbial infection in grazer-induced wounds. Whether snails commonly graze live plants and facilitate microbial growth on plants in tropical seagrass systems is less explored. On a Belizean Caye, we examined patterns in snail-generated grazer scars on the abundant turtlegrass (Thalassia testudinum). Our initial survey showed the occurrence of snail-induced scarring on live turtlegrass blades was common, with 57% of live leaves scarred. Feeding trials demonstrated that two of five common snails (Tegula fasciata–smooth tegula and Smaragdia viridis–emerald nerite) grazed unepiphytized turtlegrass blades and that smooth tegula abundance had a positive relationship with scarring intensity. Subsequent surveys at three Caribbean sites (separated by >150 km) also showed a high occurrence of snail-induced scars on turtlegrass blades. Finally, simulated herbivory experiments and field observations of a turtlegrass bed in Florida, United States suggests that herbivore damage could facilitate fungal growth in live seagrass tissue through mechanical opening of tissue. Combined, these findings reveal that snail grazing on live turtlegrass blades in the Caribbean can be common. Based on these results, we hypothesize that small grazers could be exerting top-down control over turtlegrass growth directly via grazing and/or indirectly by facilitating microbial infection in live seagrass tissue. Further studies are needed to determine the generality and relative importance of direct and indirect effects of gastropod grazing on turtlegrass health.

Spatial Patterns of Thalassia testudinum Immune Status and Labyrinthula spp. Load Implicate Environmental Quality and History as Modulators of Defense Strategies and Wasting Disease in Florida Bay, United States

Seagrass wasting disease, caused by protists of the genus Labyrinthula, is an important stressor of the dominant macrophyte in Florida Bay (FB), United States, Thalassia testudinum. FB exhibits countervailing gradients in plant morphology and resource availability. A synoptic picture of the Thalassia-Labyrinthula relationship was obtained by assessing the activity of four immune biomarkers in conjunction with pathogen prevalence and load [via quantitative PCR (qPCR)] at 15 sites across FB. We found downregulated immune status paired with moderate pathogen load among larger-bodied host phenotypes in western FB and upregulated immunity for smaller-bodied phenotypes in eastern FB. Among the highest immune response sites, a distinct inshore-offshore loading pattern was observed, where coastal basins exposed to freshwater runoff and riverine inputs had the highest pathogen loads, while adjacent offshore locations had the lowest. To explain this, we propose a simple, conceptual model that defines a framework for testable hypotheses based on recent advances in resistance-tolerance theory. We suggest that resource availability has the potential to drive not only plant size, but also tolerance to pathogen load by reducing investment in immunity. Where resources are more scarce, plants may adopt a resistance strategy, upregulating immunity; however, when physiologically challenged, this strategy appears to fail, resulting in high pathogen load. While evidence remains correlative, we argue that hyposalinity stress, at one or more temporal scales, may represent one of many potential drivers of disease dynamics in FB. Together, these data highlight the complexity of the wasting disease pathosystem and raise questions about how climate change and ongoing Everglades restoration might impact this foundational seagrass species.

Marine Seagrass Extract of Thalassia testudinum Suppresses Colorectal Tumor Growth, Motility and Angiogenesis by Autophagic Stress and Immunogenic Cell Death Pathways

Marine plants have become an inexhaustible reservoir of new phytopharmaceuticals for cancer treatment. We demonstrate in vitro/in vivo antitumor efficacy of a standardized polyphenol extract from the marine angiosperm Thalassia testudinum (TTE) in colon tumor cell lines (RKO, SW480, and CT26) and a syngeneic allograft murine colorectal cancer model. MTT assays revealed a dose-dependent decrease of cell viability of RKO, CT26, and SW480 cells upon TTE treatment with IC50 values of, respectively, 175, 115, and 60 μg/mL. Furthermore, TTE significantly prevented basal and bFGF-induced angiogenesis in the chicken chorioallantoic membrane angiogenesis assay. In addition, TTE suppressed bFGF-induced migration of endothelial cells in a wound closure assay. Finally, TTE treatment abrogated CT26 colorectal cancer growth and increased overall organism survival in a syngeneic murine allograft model. Corresponding transcriptome profiling and pathway analysis allowed for the identification of the mechanism of action for the antitumor effects of TTE. In line with our in vitro/in vivo results, TTE treatment triggers ATF4-P53-NFκB specific gene expression and autophagy stress pathways. This results in suppression of colon cancer cell growth, cell motility, and angiogenesis pathways in vitro and in addition promotes antitumor immunogenic cell death in vivo.

Patterns of Dispersion, Movement and Feeding of the Sea Urchin Lytechinus variegatus, and the Potential Implications for Grazing Impact on Live Seagrass

The sea urchin Lytechinus variegatus is a known grazer of both living and dead tissue of turtlegrass, Thalassia testudinum, occasionally denuding large areas of seagrass. Field studies have attempted to assess effects of herbivory on seagrass by enclosing urchins at various densities. However, it is unclear how unrestricted urchins affect seagrass at lower densities more typically observed in the field. This study describes movement, feeding, and distribution of L. variegatus within beds of T. testudinum in St. Joseph Bay, Florida (USA) to quantify this urchin’s impact as a seagrass grazer. Urchins were absent from portions of seagrass beds closest to shore, present at low densities midway across the bed, and at highest densities (up to ~5 individuals/m2) at the offshore edge of the bed. Urchins tended not to aggregate, moved twice as rapidly where seagrass cover was reduced, and moved > 20X faster when placed in areas of open sand. Dead seagrass tissue occurred 4—30X more frequently on oral surfaces than living seagrass tissue. Fecal pellets with dead seagrass tissue were > 3X more common than pellets with live seagrass tissue. Injury to seagrass leaves was more common along dead leaf sections than live sections (> 2—10X). Overall, spatial distributions, movement, and diet indicate that L. variegatus at densities observed in this study would tend to have minimal effects on living seagrass. Episodic periods of denuding grassbeds reported in the literature suggest L. variegatus switches to live seagrass tissue as dead tissue becomes scarce during times of high urchin density.

Polyphenolic Fraction Obtained From Thalassia testudinum Marine Plant and Thalassiolin B Exert Cytotoxic Effects in Colorectal Cancer Cells and Arrest Tumor Progression in a Xenograft Mouse Model

Marine plants are important sources of pharmacologically active metabolites. The aim of the present work was to evaluate the cytotoxic and antitumor activity of a polyphenolic fraction obtained from Thalassia testudinum marine plant and thalassiolin B in human colorectal cancer cells. Human cancer cell lines, including HCT15, HCT116, SW260, and HT29 were treated with tested products for cytotoxicity evaluation by crystal violet assay. The potential proapoptotic effect of these natural products was assessed by flow cytometry in HCT15 cells at 48 h using Annexin V-FITC/propidium iodide. In addition, reactive oxygen species (ROS) generation was measured by fluorescence using DCFH-DA staining, and sulfhydryl concentration by spectrophotometry. The in vivo antitumor activity of the polyphenolic fraction (25 mg/kg) was evaluated in a xenograft model in nu/nu mice. In vivo proapoptotic effect was also evaluated by immunohistochemistry using anti-caspase 3 and anti-Bcl-2 antibodies. The results showed that tested products exert colorectal cancer cell cytotoxicity. Besides, the tested products induced a significant increase (p < 0.05) of intracellular ROS generation, and a depletion of sulfhydryl concentration in HCT15 cells. The polyphenolic fraction arrested tumor growth and induced apoptosis in the xenograft mice model. These results demonstrate the cytotoxic activity of T. testudinum metabolites associated, at least, with ROS overproduction and pro-apoptotic effects. Here we demonstrated for the first time the antitumor activity of a T. testudinum polar extract in a xenograft mice model. These results suggest the potential use of T. testudinum marine plant metabolites as adjuvant treatment in cancer therapy.