Scott M. Gallager

Lipid as an index of growth and viability in three species of bivalve larvae

Abstract The larvae of three species of bivalve molluscs, Crassostrea virginica (Gmelin), Ostrea edulis (L.) and Mercenaria mercenaria (L.) were cultured under a variety of temperature and food species regimes in the laboratory. Data were collected on growth, survival, total lipid content and lipid class composition of both healthy and starved larvae. Subsamples were stained specifically for lipid content with the lipid-specific stain Oil Red O. Newly formed straight hinge larvae of all three species exhibited small lipid droplets dispersed throughout the tissues. These were resorbed within 4 days of the initiation of feeding while the digestive gland filled with exogenously supplied lipid. Embryogenesis took place at the expense of 69 and 71% of parentally derived total lipid, 92.8 and 68.3% of which was composed of triacylglycerols ( M. mercenaria and C. virginica , respectively). After 8 days of feeding on Isochrysis aff. galbana (clone T-ISO), total lipid had increased 2.7-fold in larvae of C. virginica grown at 25°C, 37.6-fold in larvae of M. mercenaria grown at 22°C, and 1.7-fold in larvae of O. edulis grown at 25°C. Shell growth in larvae of C. virginica grown at 30°C was initially higher than that at 25°C, but lipid was accumulated 2.5 times slower and survival of pediveliger larvae through metamorphosis was considerably reduced at this elevated temperature. The lipid class composition of healthy larvae of C. virginica and M. mercenaria was dominated by triacylglycerols and phospholipids in approximately equal proportions; however, triacylglycerols were preferentially catabolized during starvation and were responsible for greater than 80% of the total lipid loss. Color photographs of larvae stained with Oil Red O are presented that illustrate lipid levels in developing larvae in both healthy and poor condition. From these, a visual index of lipid content was developed to allow comparisons between larval survival and lipid levels. In general, high survival was always accompanied by a high lipid index but a high lipid index did not guarantee good survival. We suggest that the lipid index may be used to document visually the lipid content in individual bivalve larvae as an indicator of physiological condition and potential for successful metamorphosis.

Growth and survival of larvae of Mercenaria mercenaria (L.) and Crassostrea virginica (Gmelin) relative to broodstock conditioning and lipid content of eggs.

Abstract Artificial conditioning of broodstocks of Mercenaria mercenaria L. and Crassostrea virginica (Gmelin) was initiated at various times throughout the year under both laboratory and commercial hatchery conditions. Attempts were made to spawn broodstocks at weekly intervals, and data were collected on the initial total egg lipid content of eggs by analytical procedures and by visual inspection after staining with Oil Red O. The physical parameters of egg diameter and density (estimated by density gradient centrifugation) were correlated with egg lipid content. Significant correlations were evident between the initial egg lipid content and survival to both straight hinge (24 h) and pediveliger stages ( P 75%) was catabolism of a minimum of about 4.5 ng total lipid during the embryonic stages (i.e., fertilization through to prodissoconch I shell formation) in both bivalve species. Good survival to the pediveliger stage (i.e., >1%) was always predicated by eggs containing a minimum of 12% lipid of the ash-free dry weight (AFDW); however, relatively high lipid levels did not guarantee good larval survival. Species-specific egg density profiles were clearly evident; eggs of C. virginica banded unimodally at 1.075 g·cm −3 while eggs of M. mercenaria banded trimodally at 1.038, 1.059 and 1.073 g·cm −3 . Egg lipid was logarithmically related to egg diameter. The data suggest that there is a minimum, size-related, threshold lipid level in eggs necessary for optimal survival through the non-feeding embryonic stages; but environmental or genetic factors other than egg lipid content are responsible for a considerable fraction of the mortality during this period. Variations in the broodstock conditioning protocol induced large fluctuations in egg lipid levels, suggesting that strict attention should be paid to conditioning if optimal culture potential is desired. Minor variations in egg lipid levels (i.e., 4–8% AFDW) were visually discernible by staining with Oil Red O in both laboratory and hatchery environments. Evaluating eggs at the time of spawning for their potential for survival using the lipid-staining technique can provide valuable real-time information for both research and commercial hatchery applications.

Microaggregations of oceanic plankton observed by towed video microscopy.

Oceanic plankton have been hypothesized to occur in micropatches (<10 meters) that can have a large impact on marine ecosystem dynamics. Towed video microscopy was used to unobtrusively determine distributions of oceanic plankton over a continuum of scales from microns to hundreds of meters. Distinct, taxa-specific aggregations measuring less than 20 centimeters were found for copepods but not for nonmotile (cyanobacterial colonies) or asexual (doliolid phorozooids) forms, which suggests that these small patches are related to mating. Significant patchiness was also found on larger scales and was correlated among taxa, indicating physical control. These video observations provide new insights into basic plankton ecology by allowing quantitative assessment of individual plankton in their natural, undisturbed state.

Physiological and biochemical energetics of larvae of Teredo navalis L.and Bankia gouldi (Bartsch) (Bivalvia:Teredinidae).

Abstract The biochemical composition of larvae of Teredo navalis L. and Bankia gouldi (Bartsch) (Bivalvia: Teredinidae) was examined throughout larval development at 23°C and 30–32%. salinity in the presence of the phytoplankton food Isochrysis aff. galbana (clone T-ISO), during a delay of metamorphosis in the presence of food but absence of a wood substratum and during periods of enforced starvation. Newly released Teredo navalis larvae had a mean length ( L ) and height ( H ) of 89.3 and 76.1 μm respectively. The first appearance of pediveliger larvae at 212.1 μm L and 230.0 μm H occurred 27 days after release. Larval dry weight increased from 0.29 μg to 1.96 μg during this period. Newly formed straight hinge larvae of Bankia gouldi had dimensions of 62.8 μm L and 49.8 μm H . Metamorphically competent B. gouldi larvae had dimensions of 230.0 μm L and 282.9 μm H and were first observed 20 days after fertilization. Larval dry weight increased from 0.06 μg to 2.20 μg during this period. During enforced delay of metamorphosis the ash-free dry weight of Teredo navalis larvae decreased whereas the ash free dry weight of Bankia gouldi larvae increased. During the early period of shelled larval development both species showed similar decreases in lipid, protein and carbohydrate levels (μg·mg dry weight −1 ); however, this was reflected in a decrease in biochemical content (μg·larva −1 ) only in Teredo navalis . During enforced starvation the major proportion of both the weight and caloric losses were due to protein. Lipid also contributes significantly to these losses whereas the contribution of carbohydrate was small. Larval oxygen consumption rates were determined directly by manometry and indirectly by estimates of decrease in caloric content during periods of enforced starvation. Direct and indirect determinations for T. navalis are described by the relationships R = 1.16 W 1.05 and R = 0.98 W 1.24 respectively where R is the respiration rate in nl O 2 · larva −1 · h −1 and W is dry weight inclusive of shell in μg. Direct and indirect determinations for Bankia gouldi are described by the relationships R = 1.37 W 1.25 and R = 1.81 W 1.25 respectively. When data for both assay procedures are combined for each species the relationships R = 1.10 W 1.07 and R = 1.44 W 1.18 are obtained for Teredo navalis and Bankia gouldi respectively.

Rapid visualization of plankton abundance and taxonomic composition using the Video Plankton Recorder

Abstract Traditional methods for determining spatial distributions of planktonic taxa involve net, pump, and bottle collections followed by the tedious and time-consuming task of plankton sample analysis. Thus, plankton ecologists often require months or even years to process samples from a single study. In this paper, we present a method that allows rapid visualization of the distribution of planktonic taxa while at sea. Rapid characterization of plankton distributions is essential in the dynamic physical environment, where biological and physical patterns can change quickly. Such a “sample-and-observe” capability is necessary for mapping ephemeral features (such as patches, eddies, jets, plumes) and determining appropriate locations to conduct more localized sampling, including in situ observational studies. We describe the techniques used in imaging the plankton, analyzing the video, and visualizing the data. We present an example of at-sea data analysis conducted aboard R.V. Columbus Iselin on Georges Bank in May 1994 and visualizations of the 3-dimensional distribution of selected planktonic taxa in a 2 × 2 km × 90 m volume of seawater.

High-resolution observations of plankton spatial distributions correlated with hydrography in the Great South Channel, Georges Bank

During a cruise to Georges Bank in May 1992, the Video Plankton Recorder (VPR) was towyoed while non-invasively obtaining images of the plankton and environmental (CTD) data. Data from an 8 h transect across the Great South Channel (GSC) were analyzed on a continuum of spatial scales from coarse-scale (100 km) to micro-scale (mm). Abundance was determined for 12 taxonomic groups including: invertebrate larvae (ophiopluteus larvae, anthozoa larvae: Cerianthus sp.), hydroids, copepods (Calanus sp., Pseudocalanus sp.), pteropods (Limacina retroversa, Clione sp.), ctenophores (Mnemiopsis sp., Pleurobrachia sp.), larvacea (Oikopleura sp.), chaetognatha (Sagitta sp.), and diatom colonies (Chaetoceros socialis). Species-specific plots of the positions of individual plankton in the water column and plots of the temperature and salinity at which the plankton were observed (temperature-salinity-plankton plots) showed that major taxonomic groups were patchy at coarse scales because of their association with specific water masses of different origin and associated temperature/density discontinuities (pycnocline and fronts). Analysis of the T-S characteristics of water types indicated that diatom colonies and ophiopluteus larvae of echinoderms were transported to GSC in a band of cold water originating on the south flank of Georges Bank. Within this band, diatom colonies formed an intense patch at a front reaching a density of 5 ml−1. Within each water mass, fine-scale (10s of meters) plankton patchiness was associated with regions of vertical stability as indicated by the association of plankton with regions of high gradient Richardson number. Aggregation of plankton at the microscale (<1 m) occurred significantly only for plankton capable of active swimming, suggesting a dynamic interaction between biological and physical variables at this spatial scale. On occasion, veliger larvae of Limacina retroversa were found in spawning patches at concentrations exceeding 600 mk−1 within a few centimeters of the air-sea interface. The ability to observe and quantify local concentrations of plankton together with micro-scale physics, but over broad spatial scales, will help provide information on the coupling between spatial scales necessary to understand how individuals interact to form populations and communities in the world oceans.

Automatic Plankton Image Recognition

Plankton form the base of the food chain in the ocean and are fundamental to marine ecosystem dynamics. The rapid mapping of plankton abundance together with taxonomic and size composition is very important for ocean environmental research, but difficult or impossible to accomplish using traditional techniques. In this paper, we present a new pattern recognition system to classify large numbers of plankton images detected in real time by the Video Plankton Recorder (VPR), a towed underwater video microscope system. The difficulty of such classification is compounded because: 1) underwater images are typically very noisy, 2) many plankton objects are in partial occlusion, 3) the objects are deformable and 4) images are projection variant, i.e., the images are video records of three-dimensional objects in arbitrary positions and orientations. Our approach combines traditional invariant moment features and Fourier boundary descriptors with gray-scale morphological granulometries to form a feature vector capturing both shape and texture information of plankton images. With an improved learning vector quantization network classifier, we achieve 95% classification accuracy on six plankton taxa taken from nearly 2,000 images. This result is comparable with what a trained biologist can achieve by using conventional manual techniques, making possible for the first time a fully automated, at sea-approach to real-time mapping of plankton populations.

Video Plankton Recorder estimates of copepod, pteropod and larvacean distributions from a stratified region of Georges Bank with comparative measurements from a MOCNESS sampler

Abstract A two-vessel exercise was conducted over the southern flank of Georges Bank during the onset of vernal stratification in May 1992. The Video Plankton Recorder (VPR), a towed video system, was used to map out the fine-scale distributions of zooplankton to a depth of 70 m along a trackline which described a regular grid (3.5 × 4.5 km) in Lagrangian space. A second vessel following a parallel course conducted Multiple Opening/Closing Net and Environmental Sensing System (MOCNESS) sampling during the last section of the grid, which provided an opportunity to compare data from the two systems. Both the VPR and the MOCNESS provided similar data on the taxonomic composition of the plankton which was numerically dominated by copepods ( Calanus, Pseudocalanus, Oithona ), pteropods ( Limacina ) and larvaceans ( Oikopleura ). The absence of rare ( −3 ) species from the VPR dataset was a consequence of the small volume sampled (0.0694 m 3 ) by the high magnification camera, while fragile gelatinous taxa were undersampled by the MOCNESS. Estimates of copepod and pteropod concentrations were comparable for the two gear types. While the species composition of the plankton did not change statistically along the grid, abundances of the dominant taxa varied along the transect and each taxon demonstrated pronounced fine-scale vertical patterns that appeared to be related to hydrographic features. The VPR represents a powerful tool for rapid surveys of the micro- to fine-scale structure of zooplankton assemblages either alone, or in conjunction with other sampling techniques.

Ontogenetic changes in the vertical distribution of giant scallop larvae, Placopecten magellanicus, in 9-m deep mesocosms as a function of light, food, and temperature stratification

To understand how thermal stratification and food abundance affects the vertical distribution of giant scallop larvae Placopecten magellanicus (Gmelin), a mesocosm study was conducted in January and February 1992. The position of larvae was followed over 55 d in replicated 9-m deep tanks in relation to a sharp thermocline and the presence or absence of phytoplankton. Growth and vertical position of larvae were monitored in separate treatments which included phytoplankton added above the thermocline, below the thermocline, throughout the mesocosm, or absent from the mesocosm. Changes in the vertical position of larvae over time were quantified with a new, profiling, video-optical instrument capable of semi-automatically identifying, counting and sizing larvae. The strong diurnal migration of scallop larvae resulted in aggregations at two interfaces: the air/water interface during the night, and at the thermocline during the day. At times, the concentration of larvae within cm of the surface was > 100 times that in the remaining water column. The formation of bioconvective cells of swimming larvae at the air/water interface allowed larval aggregations to persist throughout the period of darkness. Regardless of the distribution of food, larvae remained above the thermocline during most of the experiment. Therefore, only in those treatments where food was also present above the thermocline was larval growth relatively high. Larger larvae penetrated the thermocline only after reaching a shell length of about 200 μm; thus larval size, rather than chronological age, was more important in describing their vertical distribution. The rapid increase in kinematic viscosity with decreasing water temperature at the thermocline may retard the movement of larvae and contribute to aggregation at this interface. The influence of larval size on their vertical distribution, and the resulting potential for horizontal transport to settlement sites, points to the importance of persistent hydrographic features as critical factors contributing to settlement variance in scallops.

Efficient grazing and utilization of the marine cyanobacterium Synechococcus sp. by larvae of the bivalve Mercenaria mercenaria

Efficient grazing by marine bivalve larvae has been thought to be limited to particles larger than 4 μm in diameter, thereby eliminating photosynthetic and non-photosynthetic picoplankton as contributers to larval diets. Documentation of ingestion, carbon retention and growth of laboratory-reared larvae of the bivalve Mercenaria mercenaria L. on Synechococcus sp. (WH7803), a small unicellular cyanobacterium 1 μm in diameter, was facilitated using 14C-labelled cells in pulse/chase experiments and growth of larvae on diets of cell mixtures of both Synechococcus sp. and the haptophyte Isochrysis aff. galbana (TISO). Clearance rates on Synechococcus sp. ranged between 2 and 23 μl larva-1 h-1 depending on ambient cell concentration and larval age. Retention efficiency of cell carbon after gut evacuation was about 55% for both prey species. Growth rates of larvae fed on monocultures of Synechococcus sp. at typical summer concentrations in coastal waters (1×105 cells ml-1, ∼29 μg C l-1) was two-fold lower than on monocultures of Isochrysis galbana at 1×104 cells ml-1 (∼120 μg C l-1). Larval growth was inhibited and atrophy of the digestive gland was observed when Synechococcus sp. was offered at concentrations at or exceeding 8.6×105 cells ml-1. Larval growth was enhanced, however, in the presence of Synechococcus sp. (5×104 cells ml-1) when Isochrysis galbana was limiting. During the diurnal study of Synechococcus sp. population dynamics conducted by Waterbury et al. (1986) in Vinyard Sound, Massachusetts, the abundance of bivalve larvae was sufficient to account for 12 to 24% of the calculated grazing activity on Synechococcus sp. When nanoplankton are scarce, invertebrate larvae may exert considerable grazing pressure on Synechococcus sp. and derive benefit from ingestion of these cyanobacteria.