In vitro alternatives to whole animal testing. Comparative cytotoxicity studies of divalent metals in established cell lines derived from tropical and temperate water fish species in a neutral red assay

A simple vital dye (neutral red) uptake method for investigation of the acute toxicity of seven divalent metal cations to cell lines derived from the turbot (TF) and bluegill (BF-2) was evaluated. The method was extremely reproducible between replicate experiments on a week to week basis as well as being reproducible between different laboratories. Whilst there were some species-dependent differences in the relative toxicities of the metals there were good correlations between toxicity and the chemical softness (Σp) of the metal ions and also between in vivo and in vitro toxicities. These studies demonstrate that this simple and relatively inexpensive technique carried out in 96-well microplates may be extremely useful for evaluation of structure-toxicity relationships between related groups of compounds.     

Source regions for recruitment of Calanus finmarchicus to Georges Bank: evidence from molecular population genetic analysis of mtDNA

Molecular population genetic analysis has provided evidence that the copepod, Calanus finmarchicus, of the Labrador Current, Gulf of St Lawrence, Scotian Shelf, Gulf of Maine, and Georges Bank constitute a single, interbreeding population. The DNA sequence of a 350 base pair portion of the mitochondrial large subunit (16S) ribosomal RNA (rRNA) gene was determined for a total of 72 individuals collected in 1992, and 110 individuals collected in 1993 from these regions. There was significant heterogeneity in haplotype frequencies among the samples collected in 1992, but this heterogeneity did not resolve into regional patterns. The only regional differences seen were between pooled samples of the western N. Atlantic and those of the Norwegian Sea. There were no significant differences in haplotype frequencies among the samples collected in 1993, and fewer haplotypes were observed in these samples. Intraspecific molecular variation was typical of other marine species: there were 29 haplotypes among the 182 individuals sequenced. The frequency distribution of the haplotypes was highly skewed: 128 individuals shared one haplotype and 19 individuals were unique. There were 24 variable sites among the 350 bases sequenced; estimated nucleotide diversity was 0.0042. The genetic character of C. finmarchicus populations in the western N. Atlantic was stable over time in that three of the haplotypes (including the most abundant) occurred in both 1992 and 1993. However, haplotype frequencies differed significantly between the two years. The lack of regional structure in the 1992 samples and the genetic homogeneity of samples collected in 1993 across the domain from the Labrador Current to the Gulf of St Lawrence to Georges Bank and the Gulf of Maine indicated that there is significant gene flow across this region. The persistent genetic pattern suggests that the Gulf of St Lawrence may be an important source region for recruitment of C. finmarchicus to Georges Bank. Determination of zooplankton dispersal patterns within high gene flow species will provide information that may not be determined by conventional oceanographic analyses.     

Copepod grazing during spring blooms: Can Pseudocalanus newmani induce trophic cascades?

During late winter and spring of 2002 and 2003, 24 two- to three-day cruises were conducted to Dabob Bay, Washington State, USA, to examine the grazing, egg production, and hatching success rates of adult female Calanus pacificus and Pseudocalanus newmani. Here, we discuss the results of our grazing experiments for P. newmani. Each week, we conducted traditional microzooplankton dilution experiments and “copepod dilution” experiments, each from two different layers. Grazing was measured by changes in chlorophyll concentration and direct cell counts. Clearance rates on individual prey species, as calculated by cell counts, showed that Pseudocalanus are highly selective in their feeding, and may have much higher grazing rates on individual taxa than calculated from bulk chlorophyll disappearance. The grazing rates of the copepods, however, are typically an order of magnitude lower than the grazing rates of the microzooplankton community, or the growth rates of the phytoplankton. P. newmani ingested diatoms, but, at certain times fed preferentially on microzooplankton, such as ciliates, tintinnids, and larger dinoflagellates. Removal of the microzooplankton may have released the other phytoplankton species from grazing pressure, allowing those species’ abundance to increase, which was measured as an apparent “negative” grazing on those phytoplankton species. The net result of grazing on some phytoplankton species, while simultaneously releasing others from grazing pressure resulted in bulk chlorophyll-derived estimates of grazing which were essentially zero or slightly negative; thus bulk chlorophyll disappearance is a poor indicator of copepod grazing. Whether copepods can significantly release phytoplankton from the grazing pressure by microzooplankton in situ, thus causing a trophic cascade, remains to be verified, but is suggested by our study.     

Accumulation and effects of organotin compounds in oysters and mussels: Correlation with serum biochemical and cytological factors and tissue burdens

Oysters and mussels exposed to a concentration of 0·7 ppb (μg/liter) tributyltin from painted panels in flowing seawater accumulated tin in the digestive glands to comparable levels. The mussels experienced approximately 50% mortality during the 60-day test period, but the oysters suffered virtually no deaths. There was no evidence from either bivalve of elevated numbers of hemocytes during the test period and no evidence for cellular disruption as detected by increased levels of serum lysosomal hydrolases. Serum protein of exposed mussels relative to controls increased with time of exposure to the toxicant, while oyster serum protein, normally 10 x higher than in mussels, did not. No evidence was found for elevated stress proteins (heat shock proteins) or metallothioneins in the serum hemocytes of either bivalve. Responses by these animals to fatal or near fatal doses of TBT were thus very different from responses to copper that we have reported elsewhere.^1,2     

Circulation

Low-frequency current and temperature variability on the southeast US continental shelf during summer conditions of weak wind forcing and vertical stratification was found to be similar in many aspects to previous findings for winter, when stronger wind forcing and vertical homogeneity prevails. Subtidal variability in the outer shelf is dominated by the weekly occurrence of Gulf Stream frontal eddies and meanders. These baroclinic events strongly affect the balance of momentum in the outer shelf, but not at mid-shelf. A negative alongshore sea level slope of order −10⁻⁷ is required to balance mean along-shelf momentum at the shelf edge, similar to oceanic estimates, and can contribute to the observed northward mean flow over the shelf.Low-frequency flow at mid-shelf and coastal sea level fluctuations appear to occur as a forced wave response to local alongshore wind stress events that are coherent over the shelf domain. Momentum balances indicate a trapped wave response similar to the arrested topographic wave found in the mid-Atlantic Bight (CSANADY, 1978). Density driven currents from river discharge do not appear to be significant at mid-shelf. Cold, subsurface intrusions of deeper, nutrient rich Gulf Stream waters can occasionally penetrate to mid- and inner-shelf regions north of Cape Canaveral, causing strong phytoplankton and zooplankton responses. These events were observed following the simultaneous occurrence of upwellings from northward winds and Gulf Stream frontal eddies at the shelf break during periods when the Stream was in an onshore position. Subsurface Gulf Stream intrusions to mid-shelf occur only during the summer, when the shelf is vertically stratified and cross-shelf density gradients do not present a barrier as in winter.     

A community-wide intercomparison exercise for the determination of dissolved iron in seawater

The first large-scale international intercomparison of analytical methods for the determination of dissolved iron in seawater was carried out between October 2000 and December 2002. The exercise was conducted as a rigorously “blind” comparison of 7 analytical techniques by 24 international laboratories. The comparison was based on a large volume (700 L), filtered surface seawater sample collected from the South Atlantic Ocean (the “IRONAGES” sample), which was acidified, mixed and bottled at sea. Two 1-L sample bottles were sent to each participant. Integrity and blindness were achieved by having the experiment designed and carried out by a small team, and overseen by an independent data manager. Storage, homogeneity and time-series stability experiments conducted over 2.5 years showed that inter-bottle variability of the IRONAGES sample was good (< 7%), although there was a decrease in iron concentration in the bottles over time (0.8–0.5 nM) before a stable value was observed. This raises questions over the suitability of sample acidification and storage.     

Holocene sedimentation of a wave-dominated barrierisland shoreline: Cape Lookout, North Carolina

The sedimentary record of 130 km of microtidal (0.9 m tidal range) high wave energy (1.5 m average wave height) barrier island shoreline of the Cape Lookout cuspate foreland has been evaluated through examination of 3136 m of subsurface samples from closely spaced drill holes. Holocene sedimentation and coastal evolution has been a function of five major depositional processes: (1) eustatic sea-level rise and barrier-shoreline transgression; (2) lateral tidal inlet migration and reworking of barrier island deposits; (3) shoreface sedimentation and local barrier progradation; (4) storm washover deposition with infilling of shallow lagoons; and (5) flood-tidal delta sedimentation in back-barrier environments.

Twenty-five radiocarbon dates of subsurface peat and shell material from the Cape Lookout area are the basis for a late Holocene sea-level curve. From 9000 to 4000 B.P. eustatic sea level rose rapidly, resulting in landward migration of both barrier limbs of the cuspate foreland. A decline in the rate of sea-level rise since 4000 B.P. resulted in relative shoreline stabilization and deposition of contrasting coastal sedimentary sequences. The higher energy, storm-dominated northeast barrier limb (Core and Portsmouth Banks) has migrated landward producing a transgressive sequence of coarse-grained, horizontally bedded washover sands overlying burrowed to laminated back-barrier and lagoonal silty sands. Locally, ephemeral tidal inlets have reworked the transgressive barrier sequence depositing fining-upward spit platform and channel-fill sequences of cross-bedded, pebble gravel to fine sand and shell. Shoreface sedimentation along a portion of the lower energy, northwest barrier limb (Bogue Banks) has resulted in shoreline progradation and deposition of a coarsening-up sequence of burrowed to cross-bedded and laminated, fine-grained shoreface and foreshore sands. In contrast, the adjacent barrier island (Shackleford Banks) consists almost totally of inlet-fill sediments deposited by lateral tidal inlet migration. Holocene sediments in the shallow lagoons behind the barriers are 5–8 m thick fining-up sequences of interbedded burrowed, rooted and laminated flood-tidal delta, salt marsh, and washover sands, silts and clays.

While barrier island sequences are generally 10 m in thickness, inlet-fill sequences may be as much as 25 m thick and comprise an average of 35% of the Holocene sedimentary deposits. Tidal inlet-fill, back-barrier (including flood-tidal delta) and shoreface deposits are the most highly preservable facies in the wave-dominated barrier-shoreline setting. In the Cape Lookout cuspate foreland, these three facies account for over 80% of the sedimentary deposits preserved beneath the barriers. Foreshore, spit platform and overwash facies account for the remaining 20%.     

Abundance, distribution and patch formation of zooplankton

The goal of studies described here was to determine the responses of zooplankton taxa to phytoplankton patches which develop in and near intrusions of cold, nutrient-rich Gulf Stream water. To achieve this goal we determined the horizontal and vertical distributions of abundant mesozooplankton taxa on the south-eastern continental shelf of the USA between 29°30′ and 31°N. The study period was from June 23 to August 16, 1981. Highest concentrations of zooplankton usually occurred in and near patches of phytoplankton. Increased phytoplankton appeared to trigger the formation of patches of the calanoid copepod Temora turbinata and the cyclopoid copepods Oithona spp. and Oncaea spp. The patches of zooplankton had greater alongshore than cross-shelf dimensions. T. turbinata responded rapidly to increased concentrations of phytoplankton by reproducing and aggregating in and above intruded waters. Oithonidae which were often, but not always, abundant in phytoplankton patches eventually attained high concentrations over most of the middle and part of the inner shelf. Their concentration and that of Oncaeidae increased steadily. Oncaeidae were not abundant in recently upwelled waters, as was T. turbinata but reached high concentrations in older intrusions when the abundance of T. turbinata remained level or decreased slowly. Both cyclopoid taxa are thought to reproduce slowly (egg sacs) compared to T. turbinata. Another taxon, the doliolids, became abundant far more rapidly in intruded waters (by asexual reproduction) than did the other three taxa. Doliolids were the most opportunistic intrusion zooplankton form. They do not regularly occur in low abundance on the shelf, as do the three copepod taxa, but develop in pulses in regions where T. turbinata and Oncaea are not abundant. Of the four taxa studied the abundance of doliolids increased and decreased most rapidly, whereas Oithona and Oncaea increased slowly and did not decrease during the study period. T. turbinata and Oncaea were most abundant at 60% of all stations in the intruding water. Doliolids and Oithona on the other hand, were mostly in the thermocline and intrusion. Whereas phytoplankton patches, which developed in intrusions, were physically induced (PAFFENHÖFER and LEE, 1988), patches of zooplankton were biologically induced.