Morphological characteristics of annual Zostera marina shoots at various germination temperatures

The timing of the transition from seed, seedlings and development into flowering is paramount importance in annual-type Zostera marina, because flowering is the first step of sexual reproduction. A majority of plants use environmental cues to regulate the transition to their developmental stages because plants must flower synchronously for successful outcrossing and must complete their sexual reproduction under favorable external conditions. The morphological characteristics (seeds and lateral shoot production, branch number, and inflorescence length) of reproductive shoots of Z. marina L. were examined in outdoor mesocosms to better understand the reproductive strategies of annual populations. Seeds in the germination experiment were divided into two groups: those exposed to cold (7 °C; vernalized group) and those left untreated (25-21 °C; non-vernalized group). All 600 seeds (300 from each group) were cultured for 2 months at 7, 10, 15, 20, and 25 °C in an indoor incubator. In the vernalized group, the germination rates were almost significantly higher than in the non-vernalized group. However, germination rates were not significantly affected by germination temperature. In outdoor mesocosms, production of vegetative shoots was observed in plants germinated at 15 and 20 °C in the vernalized group and at 10, 15 and 20 °C in the non-vernalized group. The highest number of vegetative shoots produced (35) was observed in plants germinated at 20 °C in the vernalized group, whereas seeds of either group failed to produce vegetative shoots when germinated at a low temperature (7 °C).In the flowering phase, the number of branches per shoot in the vernalized group was significantly higher than in the non-vernalized group. The total number of spadices on the 1st branches of plants in the vernalized group (germination at 20 °C) was significantly lower than that in the non-vernalized group at the same germination temperature. The total number of spadices per reproductive shoot in the vernalized group (germination at 10 °C) was also higher than in the non-vernalized group. Thus, both low temperature (vernalization) and seed germination temperature have implications for the sexual and asexual propagation of annual Z. marina populations.     

Evaluation of a mechanical seed planter for transplanting Zostera marina (eelgrass) seeds

Few seagrass transplant projects worldwide have relied on seeds, and those projects using Zostera marina (eelgrass) seeds have generally found low rates of seedling establishment (<10%). We compared seedling establishment achieved by a mechanical seed planter with seeds broadcast on the sediment surface by hand. The planter injected seeds into the sediment by pumping the seeds, suspended in a gelatin-based matrix, to a benthic sled with eight planting nozzles. As a control for the gel, seeds were also injected into the sediment without gel using a hand-held pipette. Seeds were planted at a density of 300 m⁻² into six replicate plots at each of three sites in the Chesapeake Bay region in September 2005, with seedling establishment measured in April 2006. Burying seeds, either with or without gel, had an overall positive effect on seedling establishment, but the effectiveness and the best method varied among sites. Mean seedling establishment for machine-planted seeds was significantly greater than for broadcast seeds at the Piankatank River site (4% vs. 1%), but not at the York (1.2% vs. 1.4%) or Spider Crab Bay (10.1% vs. 7.4%) sites. The effect of the gel was inconsistent among sites, with the highest seedling establishment (18.8%), resulting from seeds injected by pipette without gel at the Spider Crab site. Seed burial shows promise for increasing seedling establishment relative to seed broadcasting in the Chesapeake region, but further investigation of seed–sediment interactions at specific restoration sites is necessary. Low seedling establishment rates remain a bottleneck for seed-based eelgrass restoration.     

Digital image analysis of Zostera marina leaf injury

Current methods for assessing leaf injury in Zostera marina (eelgrass) utilize subjective indexes for desiccation injury and wasting disease. Because of the subjective nature of these measures, they are inherently imprecise making them difficult to use in quantifying complex leaf injuries from multiple sources. We have developed a method using color digital photography of eelgrass leaves which are then manipulated using image processing programs and analyzed using geographic digital image analysis. The resulting false color images are then assigned by the user into uninjured and injured groupings which may then be reported as a percentage of leaf area affected. If images are rectified, leaf area (cm²) of injured and uninjured leaf segments may be determined. Although this method is time consuming and still requires some subjective judgments, it does allow for precise analysis of highly complex leaf injuries and has the potential to be a substantial improvement over existing leaf injury indexes.     

Assessing the potential benthic-pelagic coupling in episodic blue mussel (Mytilus edulis) settlement events within eelgrass (Zostera marina) communities

Coastal marine seagrass ecosystems are important nursery grounds for commercially and recreationally important species, and they serve as key settlement and recruitment sites for other species. We investigated several years (2001-2003) where episodic settlement events of blue mussels (Mytilus edulis) occurred in Barnegat Bay, NJ, USA. Population assessment indicated that blue mussels settled in eelgrass beds (Zostera marina) in late spring with peak densities exceeding 170,000 m⁻². Based on calculated filtration rates of M. edulis, we determined that for at least 53 days in 2001, the density and size distribution of M. edulis were sufficient to filter the water column volume in excess of twice a day, with maximum calculated filtration rates exceeding 8 m³ water m⁻² day⁻¹. While the settlement event in 2001 was very localized, in 2003, the settlement event was considerably more widespread throughout the bay, with maximum settling densities exceeding 175,000 individuals m⁻². Associated with these high densities, maximum calculated filtration rates exceeded 15 m³ water m⁻² day⁻¹. This filtration potential may have impeded the localized development of a brown-tide (Aureococcus anophagefferens) bloom in 2001, which occurred in other regions of the bay, but the widespread settlement event seen in 2003 may have impeded the development of any brown-tide blooms in Barnegat Bay during that summer. The decline in mussel densities throughout the summer may be a result of elevated water temperatures in this back bay, but at one site, the high settlement of M. edulis was followed by a substantial migration (>40 individuals m⁻²) of small sea stars (Asterias forbesii). In 2001, A. forbesii was a significant factor in reducing M. edulis density by the end of the summer at the Barnegat Inlet site and a community level assessment showed significant positive correlations between mussel aggregations and sea star densities (r=0.68-0.73, P<0.001). At this same site in 2003, the sea stars were again present in high densities (26 m⁻²) and were a potential mechanism for mussel decline. In other regions of the bay, sea star densities are very low, but numerous other predatory species exist, including blue crabs (Callinectes sapidus), green crabs (Carcinus maenus), spider crabs (Libinia spp.), and several Xanthid crabs. Given the high mussel densities seen in this study and the considerable predation by sea stars and other benthic predators, the benthic-pelagic coupling which these mussels provide in this system contributes to the high secondary production in these grass beds.     

Effects of aerobic versus anoxic conditions on glutamine synthetase activity in eelgrass (Zostera marina L.) roots: regulation of ammonium assimilation potential

Root glutamine synthetase (GS; EC 6.3.1.2) activity was measured daily (0 to 4 days) for eelgrass (Zostera marina L.) plants held under continuous darkness rooted in sediments, continuous darkness without sediments, continuous light without sediments, and control light/dark cycle (Control L/D). Roots experiencing prolonged aerobiosis exhibited lower activity in vitro than controls, whereas roots experiencing prolonged anoxia exhibited increased activity. Plants held in darkness without sediments had activity intermediate between controls and anoxic roots. One-hour pretreatment of root extracts with ATP slightly reduced in vitro glutamine synthetase activity, whereas pretreatment with ADP and AMP increased activity ≈50%. While glutamine synthetase activity increased with higher adenylate energy charge (AEC) in the reaction mixture, pretreatment of enzyme extracts at high adenylate energy charges decreased subsequent activity relative to pretreatment at lower energy charges. One-hour pretreatment with l-alanine (Ala) had little effect on enzyme activity. Pretreatment with l-glutamine (Gln), l-glutamate (Glu), and γ-amino butyric acid (GABA) increased activity ≈75%. Incubation of excised roots under anoxic conditions for 24 h nearly doubled enzyme activity. However, addition of cycloheximide to anoxic root incubations lessened or prevented the increase in activity. It appears that enhanced glutamine synthetase activity following periods of root anoxia results from interactions with metabolites that fluctuate between aerobic and anoxic conditions, particularly adenylates, and from de novo synthesis of glutamine synthetase or some other protein synthesis-dependent process.     

Experimental manipulation of sediment organic content and water column aeration reduces Zostera marina (eelgrass) growth and survival

The effect of synergy between sediment organic enrichment and lack of night oxygen renewal in the water column on growth and survival of Zostera marina, and how it is reflected in the sulfur parameters in the plants (δ³⁴S, TS and S⁰) was studied experimentally. An experiment consisting of Z. marina mesocosms with different levels of organic enrichment and water column aeration was established, and the effects on sediment conditions, sulfide invasion and growth and survival of Z. marina were examined over a 4 week period. Shoots growing in Ambient Organic matter-sediments showed signs of sulfide invasion, as TS increased in all plant compartments and δ³⁴S of the plant tissues decreased during the experiment, but neither growth rate nor survival were significantly affected. The lack of night oxygen renewal had no evident effects in non-enriched sediments as porewater sulfide concentrations, AVS- and CRS-pools were not different from the corresponding 24 h aeration treatment. Plant growth rate and survival were neither different from the corresponding 24 h aeration treatment. On the contrary, shoots growing on High Organic matter-sediments suffered a massive sulfide invasion and it was directly correlated to the observed decrease in growth rates. Even though the lack of night oxygen renewal had no evident effects on sediment variables there were, however, strong indications that the different aeration levels affected plant performance, suggesting a lower sulfide oxidation capacity and confirming that low water column oxygen concentrations reduces the defense capacity of the shoots against sulfide invasion.Although δ³⁴S, TS and S⁰ concentrations together provided a powerful set of indicators to detect the invasion of sulfide in Z. marina shoots, this study enlightens the need for a deeper investigation of sulfide intrusion in seagrasses and the relation between plant sulfur parameters and sediment conditions.     

Ammonium uptake kinetics in root and leaf cells of Zostera marina L.

Ammonium uptake rates and the mechanism for ammonium transport into the cells have been analysed in Zostera marina L. In the cells of this species, a proton pump is present in the plasmalemma, which maintains the membrane potential. However, this seagrass shows a high-affinity transport mechanism both for nitrate and phosphate which is dependent on sodium and is unique among angiosperms. We have then analysed if the transport of another N form, ammonium, is also dependent of sodium. First, we have studied ammonium transport at the cellular level by measurements of membrane potentials, both in epidermal root cells and mesophyll cells. And second, we have monitored uptake rates in whole leaves and roots by depletion experiments. The results showed that ammonium is taken up by a high-affinity transport system both in root and leaf cells, although two different of kinetics could be discerned in mesophyll cells (with affinity constants of 2.2 ± 1.1 μM NH₄⁺, in the range 0.01-10 μM NH₄⁺, and 23.2 ± 7.1 μM NH₄⁺, at concentrations between 10 and 500 μM NH₄⁺). However, only one kinetic could be observed in epidermal root cells, which showed a K_m = 11.2 ± 1.0 μM NH₄⁺, considering the whole ammonium concentration range assayed (0.01-500 μM NH₄⁺). The higher affinity of leaf cells for ammonium was consistent with the higher uptake rates observed in leaves, with respect to roots, in depletion experiments at 10 μM NH₄⁺ initial concentration. However, when an initial concentration of 100 μM was assayed, the difference between uptake rates was reduced, but still being higher in leaves. Variations in proton or sodium-electrochemical gradient did not affect ammonium uptake, suggesting that the transport of this nutrient is not driven by these ions and that the ammonium transport mechanism could be different to the transport of nitrate and phosphate in this species.     

Assessing within habitat variability in plant demography, faunal density, and secondary production in an eelgrass (Zostera marina L.) bed

This investigation addressed faunal relationships with habitat structure within a Zostera marina community targeting differences between seagrass bed edge and interior. Z. marina biomass was significantly higher from the interior portions of the bed compared to the edge, but shoot density did not vary. Additionally, leaf width and length were significantly greater in the interior of the bed, suggesting greater total leaf area. Densities of larger organisms (> 0.85 mm) were significantly greater in vegetated samples (Z. marina edge and interior) compared to unvegetated, but an analysis of similarities demonstrated significant faunal community differences among each of the identified habitats. Densities of small organisms (0.25-0.85 mm), however, were significantly greater at Z. marina edge compared to unvegetated samples and Z. marina interior. Additionally, secondary production (μg AFDW day^− 1) was estimated based on the size distribution of taxa and showed significantly greater production from samples gathered in Z. marina compared to unvegetated samples. The relative size distribution of taxa was assessed using regression analysis and results showed that the size distribution was similar for samples collected at edge and interior Z. marina, but these distributions differed significantly when compared to unvegetated samples. The results of this study suggest that although similarities exist between edge and interior portions of Z. marina beds, especially compared to unvegetated habitats, noteworthy differences in faunal density, species composition, size distribution, and secondary production exist between edge and interior Z. marina.     

Application and assessment of a nutrient pollution indicator using eelgrass (Zostera marina L.) in Barnegat Bay-Little Egg Harbor estuary, New Jersey

Eutrophication degrades numerous estuaries worldwide and a myriad of assessment metrics have been developed. Here, we apply an example of a previously developed metric (Lee et al., 2004) designed to indicate incipient estuarine eutrophication to validate this technique in an already eutrophic estuary end-member, Barnegat Bay-Little Egg Harbor, New Jersey. The metric, termed ‘Nutrient Pollution Indicator’ (NPI) uses eelgrass (Zostera marina L.) as a bioindicator and is calculated as the ratio of leaf nitrogen content (%N) to area normalized leaf mass (mg dry wt cm⁻²). Eelgrass samples were collected along the entire length of the Barnegat Bay-Little Egg Harbor from June to October 2008 to determine if leaf chemistry and morphology reflect eutrophication status and a north-south gradient of nitrogen loading from the Barnegat Bay watershed. Nitrogen content, area normalized leaf mass, and NPI values all significantly (p < 0.05) varied temporally but not spatially. NPI values did not significantly correspond to the north-south gradient of nitrogen loading from the Barnegat Bay watershed. The NPI metric is therefore not deemed to reliably indicate estuarine eutrophic status. Differences between sampling effort (number of stations) and replication did not bias the overall conclusions.     

Effect of shading of Zostera marina (eelgrass) on sulfur cycling in sediments with contrasting organic matter and sulfide pools

Eelgrass Zostera marina was collected in spring and autumn from a light-saturated environment with low-organic sediments and a light-limited environment with organic-rich sediments in Denmark. The eelgrass and sediment responses to reduced light conditions were studied in 2-week shading experiments. Z. marina responded to reduced light conditions by decreasing growth rates and a loss of above-ground biomass. The spring plants were most sensitive to light reductions and the relative leaf elongation rates were reduced with up to 58% and the shoot densities with 33-36%. There was no difference in light response in relation to sediment organic matter contents. The sulfate reduction rates were reduced in the shaded low-organic sediments with up to 67%, whereas there was no effect of shading on rates in the organic-rich sediments. The lack of effect of shading in the organic-rich sediments was attributed to a limited coupling between Z. marina production and sediment bacterial carbon cycling. In contrast to the sulfate reduction rates, the pools of reduced sulfur were increased with up to 89% in the shaded, low-organic sediments, suggesting that the reoxidation of sulfides was reduced. Shading had no effect on the pools of sulfides in the organic-rich sediments due to much larger pools of sulfides. The enhanced sensitivity of spring plants to shading was probably due to a low above- to below-ground ratio compared to the autumn plants, which limited the plant-mediated oxidation of the sediments and thus the reoxidation of sulfides. The shaded plants were possibly more exposed to anoxic and sulfidic conditions affecting their growth and survival.     

Seasonal acclimation in metabolism reduces light requirements of eelgrass (Zostera marina)

We investigated the ability of eelgrass (Zostera marina) to adjust light requirements to seasonal changes in temperature, light and nutrient conditions through changes in metabolism, pigment and nutrient content. In agreement with expectations we found that rates of respiration and light saturated photosynthesis of summer acclimated plants peaked at higher temperatures (5 °C and 2 °C higher, respectively), and were lower than of winter acclimated plants, both at sub- and supra-optimal temperatures. Moreover respiration rates were generally more sensitive to increasing temperatures than photosynthetic rates, especially so for cold acclimated plants in February (36% higher Q₁₀-values). These changes were accompanied by a reduction in chlorophyll a and nitrogen concentrations in leaves by 35% and 60% respectively from February to August. The critical light requirement (E_C) of Z. marina to maintain a positive carbon balance increased exponentially with increasing temperature but less so for summer-acclimated than for winter-acclimated plants. However, combining E_C vs temperature models for whole-plants with data on daily light availability showed that seasonal acclimation in metabolism increased the annual period, when light requirements were meet at the 2-5 m depth interval, by 32-66 days. Hence, acclimation is an important mechanism allowing eelgrass to grow faster and penetrate to deeper waters. Critical depth limits estimated for different combinations of summer temperatures and water clarity in a future climate scenario, suggested that expected increases in temperature and nutrient run-off have synergistic negative effects, especially in clear waters, stressing the importance of continued efforts to improve water clarity of coastal waters.     

Experimental test of biodeposition and ammonium excretion from blue mussels (Mytilus edulis) on eelgrass (Zostera marina) performance

Blue mussels and eelgrass have been found to coexist in many locations. However, knowledge of the interactions between these species is limited. Two experiments were conducted in the laboratory, a “Deposit” and an “Epiphyte” experiment. The Deposit experiment examined possible effects of increasing load of blue mussel (Mytilus edulis) biodeposits on sediment biogeochemistry and eelgrass (Zostera marina) performance. Z. marina mesocosms received normal or high loads of mussel biodeposits (Normal and High), while no biodeposits were added to the Control. High dosage had overall negative effects on Z. marina, which was reflected as lower leaf numbers and biomass and accumulation of elemental sulphur in rhizomes. The sediment biogeochemical conditions were altered, as the mussel biodeposits enhanced sulphate reduction rates and increased sulphide concentrations in the porewater, which may result in sulphide invasion and reduced growth of Z. marina.In the Epiphyte experiment effects of mussel excretion, with particular emphasis on ammonium, on the growth of Z. marina and their epiphytes were examined. A thick cover of epiphytes developed on Z. marina growing together with M. edulis, and the relative growth rate was reduced with 20% compared to plants from control without mussels. Overall the experiments showed negative effects on Z. marina growing together with M. edulis, thereby supporting a preceding field study by Vinther et al. [Vinther, H.F., Laursen, J.S., Holmer, M. 2008. Negative effects of blue mussel (Mytilus edulis) presence in eelgrass (Zostera marina) beds in Flensborg fjord, Denmark. Est. Coast Shelf. Sci. 77, 91-103.].     

Experimental analysis of the response and recovery of Zostera marina (L.) and Halodule wrightii (Ascher.) to repeated light-limitation stress

Seagrasses are considered important indicators of decline in water quality resulting in increased light attenuation that negatively influences their growth and survival. Chronic light-limitation interspersed with unpredictable acute attenuation events have had poorly understood effects on seagrass recovery dynamics. Zostera marina (eelgrass) and Halodule wrightii (shoalgrass) were subject to a matrix of light-deprivation events followed by recovery periods to mimic repeated acute shading events. Plant survival, morphology, biomass, chlorophyll content, and Fv/Fm were assessed over time to determine recovery. At the end of the experiment, all plants were harvested and species-specific treatment effects were determined. Significant differences due to treatments were noted in all parameters measured. In general, responses were similar for both life-stages and between species, suggesting similar physiological tolerance to repeated acute light-attenuation events. Only plants in treatments where light-deprivation was followed by a recovery interval of at least the same duration showed signs of long-term survival. Chlorophyll fluorescence (Fv/Fm) was an important metric for assessing recovery, but it failed to detect the onset of mortality in many plants. Other metrics of plant condition need to be assessed and coupled with chlorophyll fluorescence data to assess seagrass “health”. This is of particular importance in field studies, where the history of the plants is largely unknown.     

Photosynthetic and morphological responses of eelgrass (Zostera marina L.) to a gradient of light conditions

Zostera marina L. (eelgrass) from Great Bay Estuary, New Hampshire and Maine (USA), was transplanted in outdoor mesocosms and subjected to four light treatments (100, 58, 34 and 11% surface irradiance, SI) between May and September 2003 to investigate the relationship between light availability and the growth and survival of eelgrass. Evaluating eelgrass seedlings and adult mature plants demonstrated no differences in photosynthetic response after 22 days of acclimation. During at least the first 19 days of shading, maximum electron transport rate (ETR_max) rate of eelgrass did not differ significantly between light treatments. After 40 days, a significant reduction in ETR_max and minimum saturating light was observed in plants growing at 34% SI and below. Morphological responses exhibited a linear increasing trend with greater light. 34% SI exhibited drastic reductions (to less than 25% of control) in rhizome growth, shoot density, shoot production, number of nodes per plant and plant weight at the end of the study (81 days). Shoot to root ratio at 34% SI increased by > 50%. Plants shaded to 58% SI showed no significant difference from the control in plant parameters except an increased rate of rhizome elongation. Our results link the lower shoot densities with shading to the slow growth rate of horizontal rhizomes and a total lack of lateral expansion at 11% SI. ETR_max declined over time in plants at 11% SI resulting in 81% mortality, no lateral branching and no morphological development, indicating that the minimum light required for long-term eelgrass growth and survival is greater than the previously suggested 11% SI. We demonstrate that eelgrass plants at these latitudes can persist at light levels of 58% SI and above, and are light-limited at 34% SI and below.     

Genetic characterization of Zostera asiatica on the Pacific Coast of North America

We gathered sequence information from the nuclear 5.8S rDNA gene and associated internal transcribed spacers, ITS-1 and ITS-2 (5.8S rDNA/ITS), and the chloroplast maturase K (matK) gene, from Zostera samples collected from subtidal habitats in Monterey and Santa Barbara (Isla Vista) bays, California, to test the hypothesis that these plants are conspecific with Z. asiatica Miki of Asia. Sequences from approximately 520 base pairs of the nuclear 5.8S rDNA/ITS obtained from the subtidal Monterey and Isla Vista Zostera samples were identical to homologous sequences obtained from Z. marina collected from intertidal habitats in Japan, Alaska, Oregon and California. Similarly, sequences from the matK gene from the subtidal Zostera samples were identical to matK sequences obtained from Z. marina collected from intertidal habitats in Japan, Alaska, Oregon and California, but differed from Z. asiatica sequences accessioned into GenBank. This suggests the subtidal plants are conspecific with Z. marina, not Z. asiatica. However, we found that herbarium samples accessioned into the Kyoto University Herbarium, determined to be Z. asiatica, yielded 5.8S rDNA/ITS sequences consistent with either Z. japonica, in two cases, or Z. marina, in one case. Similar results were observed for the chloroplast matK gene; we found haplotypes that were inconsistent with published matK sequences from Z. asiatica collected from Japan. These results underscore the need for closer examination of the relationship between Z. marina along the Pacific Coast of North America, and Z. asiatica of Asia, for the retention and verification of specimens examined in scientific studies, and for assessment of the usefulness of morphological characters in the determination of taxonomic relationships within Zosteraceae.     

Productivity estimation of Zostera marina on the southern coast of Korea: Effects of leaf marking height and duration

The effects of marking height and duration were examined to determine optimal marking height and duration for accurate productivity estimation of eelgrass, Zostera marina, on the southern coast of Korea. Shoots were pierced through the sheath at 2, 5, and 8 cm above the meristem and through the leaf blade at 1 cm above the blade/sheath junction (+ 1 cm blade). The marked shoots were collected weekly during periods of 4-week duration; the experiments were conducted seasonally. Leaf productivities were estimated using three measurement methods, the conventional leaf marking method (CLM), Short '87 method (Short, 1987; Gaeckle and Short, 2002), and plastochrone method. The optimal marking height and duration for leaf productivity estimation varied with the measurement method. CLM and the Short '87 method, but not the plastochrone method, were affected by marking height. The productivity estimates were lowest with a 2-cm marking height treatment due to injury of immature leaf blade tissues. The highest marking height on the sheath gives a more reliable productivity estimate with CLM, whereas any location on the sheath other than the base was appropriate as a marking point for the Short '87 method. CLM and the plastochrone method, but not the Short '87 method, were significantly affected by marking duration. CLM underestimated leaf productivity with less than a 1- or 2-week marking duration because more immature leaf tissues were counted with shorter marking durations. The marking duration should be longer than the plastochrone interval (about 2 weeks in this study) to prevent under- or overestimation of leaf productivity, but the duration should not be longer than the triple-plastochrone interval (about 6 weeks in this study) to prevent underestimation due to the sloughing off of the marked leaves.     

Effects of temperature and nitrate on phosphomonoesterase activities between carbon source and sink tissues in Zostera marina L.

Inorganic phosphorus (P_i) is important in the regulation of many carbon and nitrogen metabolic processes of plants. In this study, we examined alterations of phosphomonoesterase activity (PA; both alkaline and acid) in a submersed marine angiosperm, Zostera marina, grown in P_i non-limiting conditions under elevated temperature and/or nitrate enrichment. Control plants (ambient water-column NO₃⁻ < 2.5 μM, with weekly mean water temperatures between 26.5-27.0 °C based on a 20-yr data set in a local embayment) were compared to treated plants that were exposed to increased water-column nitrate (8 μM NO₃⁻ above ambient, pulsed daily at 0900 h), and/or increased temperature (ca. 3 °C above weekly means) over eight weeks in late summer-fall. Under both nitrate regimes, increased temperature resulted in periodic increased leaf and root-rhizome tissue carbon content, and increased acid and alkaline PA activities (AcPAs and AlPAs, respectively). There was a positive correlation between AlPA and AcPA activities and sucrose synthase activities in belowground structures, and a negative correlation between AlPA activities and sucrose concentrations. There were also periodic changes in PA partitioning between carbon source and sink tissues. In high-temperature and high-nitrate treatments, AcPAs significantly increased in leaves relative to activities in root-rhizome tissues (up to 12-fold higher in aboveground than belowground tissues in as little as 3 weeks after initiation of treatments). These responses were not observed in control plants, which maintained comparable AcPA activities in above- and belowground tissues. In addition, AlPA activity was significantly higher in leaf than in root-rhizome tissues of plants in high-temperature (weeks 3 and 6) and high temperature combined with high nitrate treatments (week 8), relative to AlPA activities in control plants. The observed changes in PAs were not related to P_i growth limitation, and may allow Z. marina to alter its carbon metabolism during periods of increased carbon demand/mobilization. This response would make it possible for Z. marina to meet short-term P requirements to maximize carbon production/allocation. Such a mechanism could help to explain the variability in PA activities that has been observed for many plant species during periods when environmental P_i exceeds requirements for optimal growth.     

Food web analysis of an eelgrass (Zostera marina L.) meadow and neighbouring sites in Mitsukuchi Bay (Seto Inland Sea,Japan) using carbon and nitrogen stable isotope ratios

The contribution of benthic microalgal production has been compared both within and outside a coastal eelgrass (Zostera marina L.) meadow. Carbon and nitrogen stable isotope ratios of suspended particulate organic matter (POM), epiphytic and epilithic organic matter (EOM), leaves of Z. marina (inside the meadow only) and two secondary consumer species (small crustaceans and fish) were measured inside and outside a meadow in Mitsukuchi Bay, Northwest Seto Inland Sea, Japan. Inside the meadow, primary producers (epiphyton) and consumers showed higher δ¹³C signatures than outside. Primary and secondary consumers inside the meadow were mainly dependent on epiphyton on the leaves of Z. marina, while consumer species outside the meadow were basically dependent on epilithon.     

Using light-permeable grating to mitigate impacts of residential floats on eelgrass Zostera marina L. in Puget Sound, Washington

This study evaluated whether light-permeable deck grating could mitigate impacts of residential mooring floats constructed over eelgrass (Zostera marina L.) in Puget Sound, Washington. Eelgrass shoot densities in undisturbed control areas and underneath and adjacent to 11 residential floats (16–50% of each float was grated) were monitored prior to float installation and annually for 3 years following installation. Using linear regression analysis, a decline in eelgrass shoot densities relative to controls was detected underneath three floats (eelgrass was eliminated under only one float) and adjacent to two floats. When control data were used to represent 100% grated, there was a weak relationship between eelgrass bed quality and percent of the deck grated (r = 0.46, p = 0.032), but no relationship when the range of grating was 16–50% (p = 0.90). The percent of a float deck grated did not contribute significantly to a multiple regression model examining change in eelgrass density that included five other dependent variables associated with the design of the floats. We conclude that either there was no effect of grating up to 50% of a float deck or we could not detect an effect. We hypothesize that the large number of site and landscape scale variables associated with a float influenced the effect (and our ability to detect it) of any one variable (such as grating). Consequently, we recommend that managers manipulate as many attributes of a float as possible (including grating) in order to reduce risks to eelgrass.     

Dormancy of Medicago marina (L.) seed

Medicago marina (L.) is a Mediterranean species whose seeds show strong dormancy that prevents germination. We used an integrated approach of physiological analyses and proteomics to investigate the mechanisms that control M. marina dormancy/germination and that underlie stress tolerance. First, we evaluated the effects on dormancy breaking of the following treatments: mechanical scarification, freezing at −20 °C, storage for 4 months and heating at 100 °C for 1 h. Mechanical scarification and freezing were the most effective treatments in overcoming dormancy. The role of abscisic acid (ABA) in M. marina dormancy was studied by ELISA immuno-enzymatic assay. The ABA content of germinated and non-germinated mature (control) and treated seeds was determined. The level of ABA was higher in treated seeds than in control seeds; the most significant increase occurred in the heated seeds. A comparison of the ABA level in the germinated, control and treated seeds suggests that different mechanisms modulate ABA content in response to different stresses, and that a specific ABA-signalling pathway regulates germination. Proteomic analysis revealed 46 proteins differentially expressed between treated and untreated seeds; 14 of these proteins were subsequently identified by mass spectrometry. Several of the proteins identified are important factors in the stress response, and are involved in such diverse functions as lipid metabolism, protein folding and chromatin protection. Lastly, an analysis of the phosphoproteome maps showed that the function of many proteins in seeds subjected to temperature treatment is modulated through post-translational modifications.