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This study is the first report of the successful cultivation of Dinophysis norvegica isolated from Japanese coastal waters (7 of 48 isolated cells were established as the clonal cultures, 14.5%) and their toxin content and production. The strains were maintained at relatively high abundance (>2,000 cells mL-1) for more than one year when fed on Mesodinium rubrum with the addition of Teleaulax amphioxeia. At the end of the one-month incubation, the total amounts of pectenotoxin-2 (PTX2), dinophysistoxin-1 (DTX1) and okadaic acid (OA) were 132.0-375.0 ng mL-1 (n = 7), 0.7-3.6 ng mL-1 (n = 3) and trace level OA (n = 1), respectively. Similarly, the cell quota of PTX2 and DTX1 were 60.6-152.4 pg cell-1 (n = 7) and 0.5-1.2 pg cell-1 (n = 3), respectively. These data indicated that toxin production varies depending on the strains in this species. In the growth experiment, D. norvegica grew, but it was very slow first 12 days. However, after that they grew exponentially with the maximum growth rate of 0.56 divisions day-1 (during Day 24-27), reaching a maximum concentration of 3,000 cells mL-1 at the end of incubation (Day 36), suggesting that they have a long lag phase. In the toxin production, OA was not at a detectable level (≤ 0.010 ng mL-1) during the 36 days of incubation except for Day 6. The concentration of DTX1 and PTX2 showed similar patterns as described for vegetative growth, but the toxin production still increased on Day 36 (1.3 ng mL-1 and 154.7 ng mL-1 in DTX1 and PTX2, respectively). The findings of this study provide novel information on the toxin content and production in D. norvegica as well as details on the culturing and maintenance of the species.

This study investigated the export of dissolved iron (DFe) from river catchments with emphasis on land use and land cover (LULC). The DFe concentrations were measured at 61 and 63 sites during summer and winter, respectively, in the Kitakami Mountain in northeast Japan. LULC was classified as broadleaf forest, conifer, grassland, paddy field, cropland, and urban area using GRASS GIS software. In the downstream areas, the riverine DFe concentrations ranged from 0.84 to 115 μg/L and from 1.5 to 103 μg/L in winter and summer, respectively, whereas the DFe in the upstream areas ranged from 0.16 to 7.6 μg/L and from 1.1 to 20.0 μg/L. Regression analysis was applied to the DFe and LULC data, and the unit load for DFe was estimated for each LULC type, the unit load values being 1.6, 139, and 439 g/km2/day for broadleaf forest, paddy fields, and urban area, respectively, in winter and 61.9, 564, and 727 g/km2/day for grassland, paddy fields, and urban area, respectively, in summer, reflecting the impact of distinct seasonality. A significant positive correlation was found between urban area and riverine DFe concentration in each season, emphasizing the influence of human activity on DFe export to river catchments. Overall, our investigation provides a series of quantitative evidence showing the importance of LULC to understand DFe export from river catchments.

The Yesso scallop, Patinopecten yessoensis (Jay), is one of the most important bivalve species in the Japanese and Chinese mariculture industry. In recent years, however, high incidences of scallop shell deformity and mortality have occurred with increasing frequency, but timing of onset and underlying causes are often unclear. Here, we proposed a promising δ18Oshell-based method for constraining the onset of shell deformity and mortality of P. yessoensis. Following six months of intermediate suspension culture in Funka Bay, Northern Japan, shells from healthy, deformed and dead scallops were randomly sampled. High-resolution seawater temperature time-series computed from healthy scallop shell δ18O profiles were precisely and temporally aligned to the instrumental temperature curve, thus allowing δ18Oshell-derived temperature time-series from deformed and dead scallops to be contextualized and allowing timing of scallop deformity and death to be retrieved. Irrespective of scallop shell length, onsets of deformity were anchored in February, and since then deformed scallops grew slowly in comparison to healthy individuals. Without exception, however, dead scallops had already ceased their shell building and died before February, indicating different underlying causes of scallop deformity and mortality. Perhaps most promisingly, considering that shells do not have any isotopic turn-over and once formed, temperature information is locked in. Thus, this approach holds great promise for identifying time anchor points (onsets of deformity and death) in archived scallops collected over different time scales, especially during massive mortality events.

Ferrous iron (Fe[II]) oxidation by dissolved oxygen was investigated in the Shizugawa Bay watershed with particular attention given to the effect of dissolved organic matter (DOM) properties on Fe(II) oxidation. To cover a wide spectrum of DOM composition, water samples were collected from various water sources including freshwater (e.g., river water and wastewater effluent) and coastal seawater. Measurement of nanomolar Fe(II) oxidation by using luminol chemiluminescence under dark, air-saturated conditions at 25 °C indicated that spatio-temporal variation of the second-order rate constant (6.7–74.5 M−1 s−1) was partially explained by the variation of the sample pH (7.5–8.6). However, at comparable pH values, the oxidation rates for freshwater were generally greater than those for coastal seawater. The substantial decline in oxidation rate constant after the removal of humic-type (allochthonous) DOM suggested that this hydrophobic DOM is a key factor that accelerates the Fe(II) oxidation in the freshwater samples. Observed lower oxidation rates for coastal seawater compared with freshwater and organic ligand-free seawater were likely associated with microbially derived autochthonous DOM, and the variation of Fe(II) oxidation at a fixed pH was best described by fluorescence index that represents the proportion of autochthonous and allochthonous DOM in natural waters. Consistently, Fe(II) oxidation was found to be slower in the presence of cellular exudates from phytoplankton. The present study highlighted the significant effect of DOM composition on the Fe(II) oxidation in inland and coastal waters.

While cyst germination may be an important factor for the initiation of harmful/toxic blooms, assessments of the fluctuation in phytoplankton cyst germination, from bottom sediments to water columns, are rare in situ due to lack of technology that can detect germinated cells in natural bottom sediments. This study introduces a simple mesocosm method, modeled after previous in situ methods, to measure the germination of plankton resting stage cells. Using this method, seasonal changes in germination fluxes of toxic dinoflagellates resting cysts, specifically Alexandrium fundyense ( A. tamarense species complex Group I) and A. pacificum ( A. tamarense species complex Group IV), were investigated at a fixed station in Kesennuma Bay, northeast Japan, from April 2014 to April 2015. This investigation was conducted in addition to the typical samplings of seawater and bottom sediments to detect the dinoflagellates vegetative cells and resting cysts. Bloom occurrences of A. fundyense were observed June 2014 and February 2015 with maximum cell densities reaching 3.6 × 10 6 cells m −2 and 1.4 × 10 7 cells m −2 , respectively. The maximum germination fluxes of A. fundyense cysts occurred in April 2014 and December 2014 and were 9.3 × 10 3 cells m −2 day −1 and 1.4 × 10 4 cells m −2 day −1 , respectively. For A. pacificum , the highest cell density was 7.3 × 10 7 cells m −2 during the month of August, and the maximum germination fluxes occurred in July and August, reaching 5.8 × 10 2 cells m −2 day −1 . Thus, this study revealed the seasonal dynamics of A. fundyense and A. pacificum cyst germination and their bloom occurrences in the water column. Blooms occurred one to two months after peak germination, which strongly suggests that both the formation of the initial population by cyst germination and its continuous growth in the water column most likely contributed to toxic bloom occurrences of A. fundyense and A. pacificum in the bay.

A high abundance of resting cysts of the toxic dinoflagellate Alexandrium tamarense was recently reported in the vast continental shelf of the Chukchi Sea in the Arctic Ocean, suggesting that the species is widespread in the shelf. Nevertheless, little is known about the occurrence of A. tamarense vegetative cells in the water column of the arctic. Sea ice reduction and the inflow of Pacific summer water (PSW) through the Bering Strait have recently increased owing to warming in the shelf. To determine the spatial and temporal distributions of A. tamarense in the Chukchi Sea shelf and their relationship to the inflow of PSW, field samplings were conducted in the Chukchi Sea and north Bering Sea shelves three times during the summer of 2013 from July to October. Vegetative cells of A. tamarense was detected in both shelves at all sampling periods with a maximum density of 3.55 × 103 cells L−1. This species was also observed at the station at 73°N, indicating the northernmost record of this species to date. The center of the A. tamarense distribution was between the north Bering and south Chukchi Sea shelf during the first collection period, and spread to the north Chukchi Sea shelf during the second and third collection periods. The species occurrences were mainly observed at stations affected by the PSW, especially Bering shelf water. Water structure of PSW was characterized by warmer surface and bottom water temperatures, and increased temperatures may have promoted the cell growth and cyst germination of A. tamarense. Therefore, it is suggested that an increase in the PSW inflow owing to warming promotes toxic A. tamarense occurrences on the Chukchi Sea shelf.

Studies on Fe uptake by phytoplankton have been often conducted using artificial culture media. However, Fe chemistry in freshwater can be influenced by riverine anthropogenic impacts and other factors causing water quality changes. In this study, therefore, Fe uptake in natural (river and reservoir) and effluent waters was investigated for the notorious bloom-forming freshwater cyanobacterium Microcystis aeruginosa. To investigate the Fe uptake mechanism, a short-term incubational assay was conducted in the presence of light, Fe(II) ligand and Fe(III) reductant, with results consistently indicating that unchelated Fe(III) is the major substrate for Fe uptake by M. aeruginosa. Further assays using various freshwater samples indicated that Fe uptake is lower in natural waters compared to that of effluent waters and, interestingly, Fe uptake was found to be limited in natural waters. These results suggest that Fe limitation can be alleviated by the inflow of effluent waters. Statistical analysis with various water quality variables indicated that Fe availability is significantly influenced by concentrations of dissolved Fe and organic matter as well as specific UV absorbance (an index of aromaticity). Overall, findings of this study highlight that watershed anthropogenic activities exert important roles in Fe uptake by freshwater cyanobacteria via alteration of Fe speciation.

Coastal waters forming an ecotoone between land and the sea include important habitats that provide many ecosystem services for human beings. Pressure brought by human activities has accumulated, however, and is beyond the capacity of coastal waters, as noted by the concept of the planetary boundary. No studies have been done about the impact of the open ocean and land-based loads from rivers and aquacultures on the environmental conditions in an open-type enclosed bay. These characteristics are different from those of a typical enclosed bay, which is influenced mainly by human activities. Few studies have been about the management methods of coastal waters, realizing that human beings can live connected with nature. Yanagi defined Satoumi as coastal waters whose productivity and biodiversity are increased by human interventions that generate fat, long, and smooth material flows. According to Yanagi, material flows can be accomplished through management of the environments in the watersheds from the mountains to the sea and the management of marine bioresources, including human consumers, who are top predators in a marine food chain. Matsuda noted that sound coastal fisheries are indispensable for maintaining healthy material flows and a sound marine environment. These two objectives form Satoumi activities, but no studies can present how to accomplish these objectives in an open-type enclosed bay. When we obtain an answer, we will be able to put them into practice for realizing a sound, prosperous, and sustainable open-type enclosed bay.

The European sea squirt, Ascidiella aspersa was first found as an alien species in 2008 from Funka Bay, Hokkaido, northern Japan, causing serious damage to the scallop aquaculture industry. We investigated A. aspersa on cultured scallops and larval occurrence from July 2010 to June 2014 to clarify life history traits and population dynamics, and consider the relation between the life history of A. aspersa and the process of scallop aquaculture. Larvae of A. aspersa were found from June to December, and recruitment on cultured scallops occurred mainly between July and October. The ascidians grew well and their weights increased until February. We found that 60–80% of A. aspersa that had settled in summer had eggs or sperm in autumn, and 90–100% of A. aspersa matured early the following summer. Maturity size in September was 17–20 mm as male, 22–24 mm as female. Scallops in Funka Bay are hung in the spring and harvested from winter to the next spring. Ascidiella aspersa settle as larvae in early summer, and grow well until winter, resulting in overgrowth on scallops in the harvest season. The linking of the process of scallop aquaculture and the life history of A. aspersa explains why this invasive ascidian has caused serious damage to the aquaculture industry in the bay. In comparison to the earlier descriptions of the native population, A. aspersa in Funka Bay has longer reproductive and growth periods, earlier initiation of reproduction, and possibly smaller maturity size.

Iron (Fe) uptake kinetics by the coastal micro-algae Skeletonema marinoi-dohrnii and the macro-algae Eisenia bicyclis were investigated in the presence of Fe and dissolved organic matter (DOM) originating from terrestrial (mountainous rivers) and coastal bay water in northeast Japan. In addition, a synthetic chelator, ethylenediaminetetraacetic acid, was tested as a representative Fe-binding organic ligand with a high Fe affinity. 59Fe uptake assays showed a linear relationship between dissolved Fe concentration and Fe uptake rate by both algae at dissolved Fe concentrations ranging from 1.0 nM to 120 nM, indicating that the Fe uptake rate was limited by the concentration of Fe species. Between 8 and 69% of the Fe fraction remained as dissolved forms when river water samples were mixed with seawater. These dissolved forms were ascribed to fulvic-like components. The uptake rate of dissolved Fe originating from river water was slower than that from coastal water (i.e., ∼1.94 and ∼0.43 log for S. marinoi-dohrnii and E. bicyclis, respectively) when the same dissolved Fe concentration was compared. Thus, dissolved Fe in river water may be less bioavailable, and the Fe-binding ligand exchange from exogenous to autochthonous DOM may increase bioavailability in coastal seawater. Excitation-emission matrix-parallel factor analysis, DOC measurements, and Fe uptake tests under dark/light incubation with a Fe(II) scavenging agent indicated differences in dominant Fe uptake mechanisms between DOMs originating in river and bay samples, with photochemical reactions greatly contributing to uptake from riverine Fe-DOM and non-chromophoric DOM produced in marine environments playing an important role in coastal seawater.

This study aimed to investigate the spatial distribution of dissolved iron from river to coastal waters and iron bioavailability for coastal phytoplankton. Dissolved iron concentrations and other water quality parameters (e.g., pH, concentrations of dissolved organic carbon and trace metals, etc.) were determined in the Shizugawa Bay and its adjacent rivers, northeast Japan. Coastal dominant diatom (Chaetoceros sp.) isolated from the bay was used for incubational assay to examine growth kinetics in a range of iron concentrations. As a result, total dissolved iron concentrations of inland waters (75 ± 80 nM) were substantially higher than those of coastal waters (7.2 ± 4.8 nM). Among inland waters, iron concentrations from anthropogenic waters were relatively higher than those for forested river waters. In the bay, relatively higher concentrations of iron were observed in the inner part. From the growth experiment, half-saturation constant of iron for the growth of Chaetoceros sp. was determined to be 1.8 - 3.5 nM. The observed dissolved iron concentrations combined with growth response indicate that growth of Chaetoceros sp. is in some cases limited by iron availability. However, this study generally suggests that, while dissolved iron concentration largely decreased from river to coastal waters, terrestrial iron inputs potentially including both natural and anthropogenic sources contribute sufficient growth and iron availability by Chaetoceros sp. in the Shizugawa Bay.

The eastern Bering Sea has a vast continental shelf, which contains various endangered marine mammals and large fishery resources. Recently, high numbers of toxic A. tamarense resting cysts were found in the bottom sediment surface of the eastern Bering Sea shelf, suggesting that the blooms have recently occurred. However, little is known about the presence of A. tamarense vegetative cells in the eastern Bering Sea. This study's goals were to detect the occurrence of A. tamarense vegetative cells on the eastern Bering Sea shelf and to find a relationship between environmental factors and their presence. Inter-annual field surveys were conducted to detect A. tamarense cells and environmental factors, such as nutrients, salinity, chlorophyll a, and water temperature, along a transect line on the eastern Bering Sea shelf during the summers of 2004, 2005, 2006, 2009, 2012, and 2013. A. tamarense vegetative cells were detected during every sampling year, and their quantities varied greatly from year to year. The maximum cell densities of A. tamarense observed during the summers of 2004 and 2005 were much higher than the Paralytic shellfish poisoning warning levels, which are greater than 100–1,000 cells L-1, in other subarctic areas. Lower quantities of the species occurred during the summers of 2009, 2012, and 2013. A significant positive correlation between A. tamarense quantity and water temperature and significant negative correlations between A. tamarense quantity and nutrient concentrations (of phosphate, silicate, and nitrite and nitrate) were detected in every sampling period. The surface- and bottom-water temperatures varied significantly from year to year, suggesting that water temperatures, which have been known to affect the cell growth and cyst germination of A. tamarense, might have affected the cells' quantities in the eastern Bering Sea each summer. Thus, an increase in the Bering Sea shelf's water temperature during the summer will increase the frequency and scale of toxic blooms and the toxin contamination of plankton feeders. This poses serious threats to humans and the marine ecosystem.

The 2011 Great East Japan Earthquake and the subsequent huge tsunami greatly affected both human activity and the coastal marine ecosystem along the Pacific coast of Japan. The tsunami also reached Funka Bay in northern Japan and caused serious damage to the scallop cultures there, and this tsunami was believed to have affected the coastal environments in the bay. Therefore, we investigated the changes in the spatial abundance and distribution of the toxic dinoflagellates Alexandrium tamarense cysts before the tsunami (August 2010) and after the tsunami (May 2011, August 2011, May 2012 and August 2012) in the bay. Further, monthly sampling was conducted after the tsunami to identify seasonal changes of Alexandrium catenella/tamarense cysts and vegetative cells. Significant increases were observed in the populations of A. catenella/tamarense cysts, comparing the abundances before the tsunami (in August 2010; 70 ± 61 cysts g −1 wet sediment) to those just after it (in May 2011; 108 ± 84 cysts g −1 wet sediment), and both A. tamarense bloom (a maximum density was 1.3 × 10 3 cells L −1 ) and PSP (Paralytic Shellfish Poisoning) toxin contamination of scallops (9.4 mouse unit g −1 was recorded) occurred in the bay. Seasonal sampling also revealed that the encystment of A. tamarense and the supply of the cysts to bottom sediments did not occur in the bay from September to April. These results strongly suggested that the mixing of the bottom sediments by the tsunami caused the accumulation of the toxic A. tamarense cysts in the surface of bottom sediment through the process of redeposition in Funka Bay. Moreover, this cyst deposition may have contributed to the toxic bloom formation as a seed population in the spring of 2011.

article i nfo The assimilation efficiency of zooplankton is an essential parameter required to estimate energy transfer to higher trophic levels in marine ecosystems. However, little information is available for large oceanic copepods, especially the Neocalanus and Eucalanus species dominant in the subarctic Pacific. In this study, the assimilation efficiencies of the C5 stages of Neocalanus cristatus, Neocalanus flemingeri and Eucalanus bungii were evaluated using eight phytoplankton species as food. The average assimilation efficiencies of N. cristatus, N. flemingeri and E. bungii ranged between 45 and 66%, 44 and 66% and 34 and 65%, respectively. The assimilation efficiency was highly variable depending on the food phytoplankton species. In all species, the assimilation efficiency showed as ignificant negative relationship with the ash content of the phytoplankton (r 2 =0 .79-0.87, p b 0.001). The assimilation efficiency of large-body sized N. cristatus for large-sized diatoms was higher than for the other copepod species. In population models of N. cristatus, changes in assimilation efficiency affect the growth and survival rates of the population. The Lagrangian ensemble model (LEM) for N. cristatus showed that, for assimilation efficienciesless than 57%, thepopulationcouldnotbemaintained.Becausevariations inassimilation efficiency may have significant effects on the copepod population, their variability should be incorporated into marine ecosystem models in the future.

A B S T R A C T Abundance and distribution of the toxic dinoflagellate Alexandrium tamarense species complex resting cyst were investigated in the eastern Bering Sea and the Chukchi Sea for the first time. Sediment samples (top 0–3 cm depth) were collected from the continental shelf of the eastern Bering Sea (17 stations) and the Chukchi Sea (13 stations) together with a long core sample (top 0–21 cm depth) from one station in the Chukchi Sea during 2009–2012. The cysts were enumerated using the primuline staining method. Species identification of the cysts was carried out with multiplex PCR assay and the plate morphology of vegetative cells germinated from cysts in the both areas. Alexandrium cysts were widely detected in the both areas, ranging from not detected (

Artificial streams were set up to test the relationship between near-bed water velocity and periphyton growth. Periphyton community samples collected from a Japanese stream were incubated for 44 days under a light intensity of 252 ± 72 μmol·photons/m2·s, a temperature of 20–25 °C, and three near-bed water velocity classes: low (32.8 cm/s). A logistic model was applied to estimate the maximum net growth rate (μmax) and carrying capacity (Bmax). A response surface method was also applied to estimate chlorophyll a (Chl-a) and ash-free dry mass (AFDM) with respect to the independent variables (i.e., time and water velocity). We detected both the highest μmax (1.99 d−1) and highest Bmax (7.01 mg/m2) for Chl-a at the moderate water velocity. For AFDM, we observed the highest μmax (0.57 d−1) and Bmax (1.47 g/m2) at the low and moderate velocity classes, respectively. The total algae density in the region of moderate velocity at the end of the experiment was 6.47 × 103 cells/cm2, corresponding to levels 1.7 and 1.3 times higher than those at lower and higher velocities, respectively. Our findings indicated that the moderate near-bed water velocity provided favorable conditions for algal growth and corresponding biomass accumulation.

Abundant cyst distributions of the toxic dinoflagellate Alexandrium fundyense (previous A. tamarense north American clade) were recently observed on the north Chukchi Sea shelf and on the eastern Bering Sea shelf, suggesting that A. fundyense is both highly adapted to the local environments in the high latitude areas and might cause toxin contamination of plankton feeders. However, little is known about the physiological characteristics and toxin profiles of A. fundyense in these areas, which are characterized by low water temperatures, weak sunlight, and more or less permanent ice cover during winter. To clarify the physiological characteristics of A. fundyense, the effects of water temperature and light intensity on the vegetative growth and toxin profiles of this species were examined using A. fundyense strains isolated from one sediment sample collected from each area. Using the same sediments samples, seasonal changes of the cyst germination in different water temperatures were investigated. Vegetative cells grew at temperatures as low as 5 °C and survived at 1 °C under relatively low light intensity. They also grew at moderate water temperatures (10–15 °C). Their cysts could germinate at low temperatures (1 °C) and have an endogenous dormancy period from late summer to early spring, and warmer water temperatures (5–15 °C) increased germination success. These physiological characteristics suggest that A. fundyense in the Chukchi Sea and eastern Bering Sea is adapted to the environments of high latitude areas. In addition, the results suggest that in the study areas A. fundyense has the potential to germinate and grow when water temperatures increase. Cellular toxin amounts of A. fundyense strains from the eastern Bering Sea and Chukchi Sea were ranged from 7.2 to 38.2 fmol cell−1. These toxin amounts are comparable with A. fundyense strains isolated from other areas where PSP toxin contamination of bivalves occurs. The dominant toxin of the strains isolated from the Chukchi Sea was saxitoxin, while most A. fundyense strains from the eastern Bering Sea are dominated by the C2 toxin. Toxin profiles similar to those detected in Chukchi Sea have not been reported by any previous research. The dominance of a highly toxic PST variant in Chukchi A. fundyense suggests that presence of the species at low cell concentrations may cause toxin contamination of predators. This study revealed that abundant A. fundyense cysts deposited on the eastern Bering Sea and Chukchi Sea shelves potentially germinate and grow with PSP toxin contents in the local environments. In conclusion, a high risk of PSP occurrences exists on the eastern Bering Sea and Chukchi Sea shelves.

This study investigated the role of snowmelt runoff on water circulation, water mass formation, and the production of cultured scallop larvae, as a part of a land-sea linkage, by analyzing hydrological data in conjunction with nutrient data and by conducting sensitivity experiments based on a coupled land-sea model of Funka Bay, Japan, a typical semi-enclosed bay. A comparison between observed data and the simulated runoff showed that, using newly compiled datasets of nutrient concentrations in rivers and groundwater, the model was sufficiently accurate to estimate the terrestrial dissolved inorganic nitrogen (DIN) flux from the river and submarine groundwater discharges (SGDs). The average volume flux from the SGDs accounted for 26% of the riverine runoff flux. The DIN flux from SGDs accounted for a maximum of 40% of the total DIN loading to the bay before the snowmelt period. Sensitivity experiments using an ocean simulation indicated that the freshwater flux supplied by snowmelt runoff not only enhances clockwise circulations along with upwelling along the coast, but also modifies the distributions of wintertime water masses in the bay. However, the snowmelt runoff has little effect on larvae transport since wind forcing, rather than riverine buoyancy, dominates the circulation patterns. The annual density of scallop spat was highly correlated with snowmelt runoffs associated with high DIN concentrations, which suggested that riverine nutrients can increase the biomass of phytoplankton in near-shore seas and improve food availability for scallop spawners, resulting in increased egg production in March to April. Therefore, the nutrient flux from agricultural source areas through the large snowmelt runoff has an important role in larvae production. Land-sea linkages need to be identified to design sustainable and synergetic systems of aquaculture and agriculture for the integrated management of coastal regions.