Flocculation and removal of the brown tide organism, Aureococcus anophagefferens (Chrysophyceae), using clays

Previous attempts to remove the brown tideorganism, Aureococcusanophagefferens, through flocculation withclays have been unsuccessful, in spite ofadopting concentrations and dispersalprotocols that yielded excellent cellremoval efficiency (RE >90%) with otherspecies, so a study was planned to improvecell removal. Four modifications in claypreparation and dispersal were explored: 1)varying the salinity of the claysuspension; 2) mixing of the clay-cellsuspension after clay addition; 3) varyingof concentration of the initial clay stock;4) pulsed loading of the clay slurry. Theeffect of salinity was dependent on theclay mineral type: phosphatic clay (IMC-P2)had a higher RE than kaolinite (H-DP) whenseawater was used to disperse the clay, butH-DP removed cells more efficiently whensuspended in distilled water prior toapplication. Mixing after dispersalapproximately doubled RE for both clayscompared to when the slurry was layeredover the culture surface. Lowering theconcentration of clay stock and pulsing theclay loading increased RE, regardless ofmineral type. However, this increase wasmore apparent for clays dispersed inseawater than in distilled water. Ingeneral, application procedures thatdecrease the rate of self-aggregation amongthe clay particles and increase thecollision frequency between clay particlesand A. anophagefferens achieve highercell removal efficiency. These empiricalstudies demonstrated that clays might be animportant control option for the brown tideorganism, given the proper attention topreparation, dispersal methods,environmental impacts, and the hydrodynamicproperties of the system being treated. Implications for the treatment of browntides in the field are discussed.     

Elevated organic nutrient ratios associated with brown tide algal blooms of Aureococcus anophagefferens (Pelagophyceae)

Natural assemblages of Aureococcus anophagefferens collectedfrom the coastal bays of Maryland and Long Island, New Yorkoccurred under water quality conditions higher in dissolvedorganic carbon and phosphorus (relative to nitrogen) than thosesame stations when Aureococcus was not present in significantnumbers. Additional work, both in culture and in the field,confirmed the importance of a heterotrophic nutritional modein this brown tide organism.     

Development and application of a monoclonal-antibody technique for counting Aureococcus anophagefferens,an alga causing recurrent brown tides in the Mid-Atlantic United States

A method was developed for the rapid detection and enumeration of Aureococcus anophagefferens, the cause of harmful algal blooms called "brown tides" in estuaries of the Mid-Atlantic United States. The method employs a monoclonal antibody (MAb) and a colorimetric, enzyme-linked immunosorbent assay format. The MAb obtained exhibits high reactivity with A. anophagefferens and very low cross-reactivities with a phylogenetically diverse array of other protists and bacteria. Standard curves are constructed for each 96-well microtiter plate by using known amounts of a preserved culture of A. anophagefferens. This approach allows estimation of the abundance of the alga in natural samples. The MAb method was compared to an existing method that employs polyclonal antibodies and epifluorescence microscopy and to direct microscopic counts of A. anophagefferens in samples with high abundances of the alga. The MAb method provided increased quantitative accuracy and greatly reduced sample processing time. A spatial survey of several Long Island estuaries in May 2000 using this new approach documented a range of abundances of A. anophagefferens in these bays spanning nearly 3 orders of magnitude.     

Development and Application of a Monoclonal-Antibody Technique for Counting Aureococcus anophagefferens, an Alga Causing Recurrent Brown Tides in the Mid-Atlantic United States

A method was developed for the rapid detection and enumeration of Aureococcus anophagefferens, the cause of harmful algal blooms called “brown tides” in estuaries of the Mid-Atlantic United States. The method employs a monoclonal antibody (MAb) and a colorimetric, enzyme-linked immunosorbent assay format. The MAb obtained exhibits high reactivity with A. anophagefferens and very low cross-reactivities with a phylogenetically diverse array of other protists and bacteria. Standard curves are constructed for each 96-well microtiter plate by using known amounts of a preserved culture of A. anophagefferens. This approach allows estimation of the abundance of the alga in natural samples. The MAb method was compared to an existing method that employs polyclonal antibodies and epifluorescence microscopy and to direct microscopic counts of A. anophagefferens in samples with high abundances of the alga. The MAb method provided increased quantitative accuracy and greatly reduced sample processing time. A spatial survey of several Long Island estuaries in May 2000 using this new approach documented a range of abundances of A. anophagefferens in these bays spanning nearly 3 orders of magnitude.     

Immunofluorescence Flow Cytometry Technique for Enumeration of the Brown-Tide Alga, Aureococcus anophagefferens

A new immunologically based flow cytometry (IFCM) technique was developed to enumerate Aureococcus anophagefferens, a small pelagophyte alga that is the cause of “brown tides” in bays and estuaries of the mid-Atlantic states along the U.S. coast. The method utilizes a monoclonal antibody conjugated to fluorescein isothiocyanate (FITC-MAb) to label the surface of A. anophagefferens cells which are then detected and enumerated by using a flow cytometer. Optimal conditions for FITC-MAb staining, including solution composition, incubation times, and FITC-MAb concentrations, were determined. The FITC-MAb method was tested for cross-reactivity with nontarget, similarly sized, photoautotrophic protists, and the method was compared to an enzyme-linked immunosorbent assay (ELISA) using the same MAb. Comparisons of the IFCM technique to traditional microscopy enumeration of cultures and spiked environmental samples showed consistent agreement over several orders of magnitude (r² > 0.99). Comparisons of the IFCM and ELISA techniques for enumerating cells from a predation experiment showed a substantial overestimation (up to 10 times higher) of the ELISA in the presence of consumers of A. anophagefferens, presumably due to egested cell fragments that retained antigenicity, using the ELISA method, but were not characterized as whole algal cells by the IFCM method. Application of the IFCM method to environmental “brown-tide” samples taken from the coastal bays of Maryland demonstrated its efficacy in resolving A. anophagefferens abundance levels throughout the course of a bloom and over a large range of abundance values. IFCM counts of the brown-tide alga from natural samples were consistently lower than those obtained using the ELISA method and were equivalent to those of the polyclonal immunofluorescence microscopy technique, since both methods discriminate intact cells. Overall, the IFCM approach was an accurate and relatively simple technique for the rapid enumeration of A. anophagefferens in natural samples over a wide range of abundance values (10³ to 10⁶ cells ml⁻¹).     

Transport of the Harmful Bloom Alga Aureococcus anophagefferens by Oceangoing Ships and Coastal Boats

It is well established that cyst-forming phytoplankton species are transported in ships' ballast tanks. However, there is increasing evidence that other phytoplankton species which do not encyst are also capable of surviving ballast transit. These species have alternative modes of nutrition (hetero- or mixotrophy) and/or are able to survive long-term darkness. In our studies of no-ballast-on-board vessels arriving in the Great Lakes, we tested for the presence of the harmful algal bloom species Aureococcus anophagefferens (brown tide) in residual (i.e., unpumpable) ballast water using methods based on the PCR. During 2001, the brown tide organism was detected in 7 of 18 ballast water tanks in commercial ships following transit from foreign ports. Furthermore, it was detected after 10 days of ballast tank confinement during a vessel transit in the Great Lakes, a significant result given the large disparity between the salinity tolerance for active growth of Aureococcus (>22 ppt) and the low salinity of the residual ballast water (~2 ppt). We also investigated the potential for smaller, recreational vessels to transport and distribute Aureococcus. During the summer of 2002, 11 trailered boats from the inland bays of Delaware and coastal bays of Maryland were sampled. Brown tide was detected in the bilge water in the bottoms of eight boats, as well as in one live-well sample. Commercial ships and small recreational boats are therefore implicated as potential vectors for long-distance transport and local-scale dispersal of Aureococcus.     

Transport of the harmful bloom alga Aureococcus anophagefferens by oceangoing ships and coastal boats

It is well established that cyst-forming phytoplankton species are transported in ships' ballast tanks. However, there is increasing evidence that other phytoplankton species which do not encyst are also capable of surviving ballast transit. These species have alternative modes of nutrition (hetero- or mixotrophy) and/or are able to survive long-term darkness. In our studies of no-ballast-on-board vessels arriving in the Great Lakes, we tested for the presence of the harmful algal bloom species Aureococcus anophagefferens (brown tide) in residual (i.e., unpumpable) ballast water using methods based on the PCR. During 2001, the brown tide organism was detected in 7 of 18 ballast water tanks in commercial ships following transit from foreign ports. Furthermore, it was detected after 10 days of ballast tank confinement during a vessel transit in the Great Lakes, a significant result given the large disparity between the salinity tolerance for active growth of Aureococcus (>22 ppt) and the low salinity of the residual ballast water (approximately 2 ppt). We also investigated the potential for smaller, recreational vessels to transport and distribute Aureococcus. During the summer of 2002, 11 trailered boats from the inland bays of Delaware and coastal bays of Maryland were sampled. Brown tide was detected in the bilge water in the bottoms of eight boats, as well as in one live-well sample. Commercial ships and small recreational boats are therefore implicated as potential vectors for long-distance transport and local-scale dispersal of Aureococcus.     

The central role of selenium in the biochemistry and ecology of the harmful pelagophyte,Aureococcus anophagefferens

The trace element selenium (Se) is required for the biosynthesis of selenocysteine (Sec), the 21st amino acid in the genetic code, but its role in the ecology of harmful algal blooms (HABs) is unknown. Here, we examined the role of Se in the biology and ecology of the harmful pelagophyte, Aureococcus anophagefferens, through cell culture, genomic analyses, and ecosystem studies. This organism has the largest and the most diverse selenoproteome identified to date that consists of at least 59 selenoproteins, including known eukaryotic selenoproteins, selenoproteins previously only detected in bacteria, and novel selenoproteins. The A. anophagefferens selenoproteome was dominated by the thioredoxin fold proteins and oxidoreductase functions were assigned to the majority of detected selenoproteins. Insertion of Sec in these proteins was supported by a unique Sec insertion sequence. Se was required for the growth of A. anophagefferens as cultures grew maximally at nanomolar Se concentrations. In a coastal ecosystem, dissolved Se concentrations were elevated before and after A. anophagefferens blooms, but were reduced by >95% during the peak of blooms to 0.05 nℳ. Consistent with this pattern, enrichment of seawater with selenite before and after a bloom did not affect the growth of A. anophagefferens, but enrichment during the peak of the bloom significantly increased population growth rates. These findings demonstrate that Se inventories, which can be anthropogenically enriched, can support proliferation of HABs, such as A. anophagefferens through its synthesis of a large arsenal of Se-dependent oxidoreductases that fine-tune cellular redox homeostasis.     

Molecular Cloning and Antiserum Development of Cyclin Box in the Brown Tide Alga Aureococcus anophagefferens

Cyclins can be useful cell cycle markers for growth rate studies on harmful algal blooms. In this study, a gene fragment corresponding to cyclin box was cloned for the brown tide alga Aureococcus anophagefferens. This algal gene fragment, designated as Btcycl1, was most similar to cyclin B. Oligopeptides based on the deduced amino acid sequence were synthesized and used to raise an antiserum that reacted on Western blots with a protein of about 63 kDa, the same size as cyclin B in other organisms. The cyclin B–like protein recognized by this antiserum, and the messenger RNA amplified using the primers, were more abundant in exponential cultures and decreased markedly in stationary cultures. This protein also appeared to be cell cycle dependent. Immunofluorescence labeling showed that this antiserum specifically stained a protein in Aureococcus cells and had no cross-reaction with bacteria that were present in the algal culture. The Btcycl1 sequence and the antiserum will provide a useful tool for studies on regulation of in situ growth rate for this brown tide alga. Received May 22, 2000; accepted July 13, 2000.     

Larval Sulcascaris sulcata from calico scallops, Argopecten gibbus, along the southeast coast of the United States

Adductor muscles of calico scallops, Argopecten gibbus, collected off the southeastern coast of the United States from May 1982 to December 1984 were examined for the presence of larvae of the parasitic nematode, Sulcascaris sulcata. Infection intensity, prevalence, and larval length were all positively correlated (P less than 0.001) with scallop shell height (age). Prevalence was generally highest in December and lowest in August as a result of yearly trends in the age structure of the scallop population. Prevalence was not influenced by latitude or depth over the ranges examined. The rate of infection of marine molluscs by S. sulcata is generally related to the degree and length of exposure to third-stage larvae, whose distribution is essentially that of the marine turtle definitive host. Thus, over the wide geographic area of the east coast of the U.S., overall rates of infection decrease with increasing latitude.     

Seaweed biogeography of the mid-Atlantic coast of the United States

The northern boundary of the warm temperate region of the mid-Atlantic coast of the United States is set at Cape Hatteras; the southern boundary lies at Cape Canaveral. There is some spillover of cool temperate species south of Cape Hatteras into North Carolina and spillover of warm temperate species south of Cape Canaveral toward Palm Beach. Elements of the warm temperate flora also extend into the northern Gulf of Mexico, but precise limits to the flora cannot be drawn there. Thirty-one species are endemic to the warm temperate flora. The inshore waters of North Carolina include approximately equal numbers of species with northern and southern centres of distribution; the species of the offshore waters have predominantly southern affinities, but also include most of the endemic species. Seasonal changes in the shallow water flora of North Carolina reflect eurythermal cool temperate and tropical elements in winter and summer respectively and a year-round warm temperate element. These groupings have been verified by experimental studies in which light and temperature were varied. The deep water flora is a summer flora dominated by perennial species. The inshore, eurythermal cool temperate and tropical species have a variety of cryptic stages by which they persist throughout the year.Paper presented at the Seaweed Biogeography Workshop of the International Working Group on Seaweed Biogeography, held from 3–7 April 1984 at the Department of Marine Biology, Rijks-universiteit Groningen (The Netherlands). Convenor: C. van den Hoek.     

Life history and habitat use of the speckled worm eel, Myrophis punctatus, along the east coast of the United States

Many species of fishes along the east coast of the United States have complex life histories, especially those that move over hundreds of kilometers across ocean and estuarine habitats. To further unravel the life history of one of these, the speckled worm eel, Myrophis punctatus we examined samples from extensive time series and discrete samples collected in the ocean and estuaries between Florida and Massachusetts. We now surmise spawning occurs between fall and early winter in the ocean south of Cape Hatteras, North Carolina and in the vicinity of the Bahamas. The pelagic leptocephalus larvae (10?C80 mm Total Length [TL]) are transported north in the Gulf Stream and across the continental shelf to arrive at estuarine inlets at ages of 53 ?C 110 days. Their estuarine immigration and abundance varies along the east coast, with higher levels occurring at inlets in South Carolina (North Inlet), and North Carolina (Beaufort Inlet), during the winter and early spring. Much lower abundances occur in New Jersey (Little Egg Inlet) in winter and spring and again in the summer. These ingressing individuals were euryodontic leptocephali and metamorphic stages and were shrinking to lengths of 76?C52 mm TL as these stages progressed. Metamorphic individuals and glass eels subsequently settle and burrow in estuarine sediments, as do all subsequent stages, and thereby become relatively unavailable to many sampling gears. In estuaries they attained sizes up to 440 mm TL. Later, they presumably enter the ocean to spawn because that is where the smallest larvae are found.     

Patterns of plant species richness in pasture lands of the northeast United States

Pasture lands are an important facet of land use in the northeast United States, yet little is known about their recent diversity. To answer some fundamental questions about the diversity of these pasture lands, we designed a broad survey to document plant species richness using an intensive, multi scale sampling method. We also wanted to learn whether environmental (soils or climate) or land management variables could help explain patterns of species richness. A total of 17 farms, encompassing 37 pastures, were sampled in New York, Pennsylvania, Vermont, Maryland, Massachusetts and Connecticut during July and August 1998. We positively identified a total of 161 different plant species across the study region. Species richness averaged 31.7±1.1 on pastures. Infrequent, transient species that were mostly perennial and annual forbs accounted for 90% of the species richness. Except for a subjective rating of grazing intensity, land management methods were not good predictors of species richness. Over time, it appears that grazing neither reduces nor increases species richness in pastures. Of the environmental variables measured, only soil P explained a significant amount of the variation in species richness. Soil P was inversely related to species richness at the 1m² scale. Percent SOM was positively associated with species richness at this scale, although weakly. At larger spatial scales, we suggest that patterns of species richness are best explained by the species diversity of soil seed banks, or seed rain, and stochastic recruitment of these species into existing vegetation.     

Characterization of red tide aerosol on the Texas coast

In the fall of 2000, there was a red tide episode in the Gulf of Mexico near Corpus Christi, TX. We sampled at the Marine Science Institute in Port Aransas on 25 October 2000. Between 25–27 October 2000, we sampled at the Texas State Aquarium (TSA) near Corpus Christi Bay. Two high-volume samplers were equipped with a filter and a five-stage impactor, respectively. Because the amounts of brevetoxin (PbTx) collected in the air samples were low, we developed a LC/MS technique to analyze the PbTxs. Personal exposure was estimated with a personal filter sampler placed in the lapel of field workers. Concentrations of PbTx-2 and -3 were detected in the samples taken at the TSA; however, PbTx was not detected in the samples from the Marine Science Institute. The concentration of PbTx-2 was between 1.5 and 5.9 ng m⁻³ and much lower concentrations for PbTx-3. The ratio of PbTx-2 and -3 was 8.7±5.2. In the highest exposure period (26–27 October), PbTx-6 was also detected. No respiratory symptoms were reported at the Marine Science Institute, whereas at the TSA, symptoms including irritation in the nose and throat, and itchy skin were reported among seven field study workers. The PbTx concentrations estimated from both high-volume impactor and filter samplers were similar. The mass median aerodynamic diameters were between 7 and 9 μm (geometric standard deviation of 1.6), a relatively large size for inhaled ambient particles. Inhaled particles of this size would be predominantly deposited in the upper respiratory tract (nasal, oral, and pharyngeal area), and subsequent respiratory irritation could result from the presence of the particles themselves or from toxins associated with the particles. Information gained from these studies will aid in evaluations of the human risk associated with inhalation of red tide aerosols.     

Avian community structure along elevational gradients in the northeastern United States

Summary Breeding birds were censused along four elevational gradients in the Adirondack Mountains, New York, and the Green Mountains, Vermont. The bird communities of the four gradients were basically similar in species composition, richness and amplitude patterns. Three measures of species diversity decreased with increasing elevation. Low-elevation communities contained higher proportions of rare species and the relative abundances conformed to the broken-stick distribution. At higher elevations the communities showed greater dominance and the dominance-diversity curves approached geometric series. The species characteristic of high-elevation communities had the broadest altitudinal distributions.The upper and lower distributional limits of most species were independent of one another except at ecotones where marked changes in vegetation structure occurred. On each mountain, slightly more than half of the species limits coincided with ecotones. This is a significantly greater proportion than has been found in similar studies of tropical forest bird communities. In further contrast to tropical communities, we found no convincing cases of altitudinal competitive exclusion between species. Interspecific competition in the past seems to have been translated primarily into differences in habitat selection by temperate forest birds.Many of the differences between temperate forest breeding bird communities and tropical ones can be understood in terms of the migratory nature of most of the temperate species and the lower species richness in temperature forests.     

An AFLP-based survey of genetic diversity among accessions of sea oats (Uniola paniculata, Poaceae) from the southeastern Atlantic and Gulf coast states of the United States

Uniola paniculata, commonly known as sea oats, is a C₄ perennial grass capable of stabilizing sand dunes. It is most abundant along the Gulf of Mexico and southeastern Atlantic coastal regions of the United States. The species exhibits low seed set and low rates of germination and seedling emergence, and so extensive clonal reproduction is achieved through production of rhizomes, which may contribute to a decline in genetic diversity. To date, there has been no systematic assessment of genetic variability and population structure in naturally occurring stands in the USA. This study was conducted to assess the genetic relationship and diversity among nineteen U. paniculata accessions representing eight states: Texas, Louisiana, Mississippi, Alabama, Florida, South Carolina, North Carolina, and Virginia, using amplified fragment length polymorphism (AFLP). Twelve AFLP EcoRI+MseI primer combinations generated a wide range of polymorphisms (42–81%) with a mean of 59%. Overall, the sea oats plants exhibited a low range of genetic similarity. Florida accessions, FL-33 and FL-39, were most genetically diverse and the accessions from both Carolinas and Virginia (NC-1, NC-11, SC-15, and VA-53) harbored less genetic variability. Cluster analysis using the UPGMA approach separated U. paniculata plants into four major clusters which were also confirmed by principal coordinate analysis (PCO). Further examination of the different components of genetic variation by analysis of molecular variance (AMOVA) indicated the largest proportion of variability at the state level (47.8%) followed by the variation due to the differences among the genotypes within an accession (34.4%), and the differences among the accessions within a state (17.8%). The relationship between genetic diversity and geographic source of sea oats populations of the United States as revealed through this comprehensive study will be helpful to resource managers and commercial nurseries in identifying suitable plant materials for restoration of new areas without compromising the adaptation and genetic diversity.     

Distribution of seagrasses along the Indian coast

Seagrass environments, from the main coast of India, Lakshadweep and Andaman Islands, were surveyed for seagrass and marine algal composition. Extensive seagrass meadows and the maximum number of species (seven genera and 12 species) occurred along the Tamil Nadu coast. Seagrasses were observed from intertidal to subtidal regions down to 8 m depth. Thalassia hemprichii (Ehrenberg) Aschers. and Cymodocea serrulata (R. Brown) Aschers. & Magnus were the dominant seagrasses in the subtidal zones. Halophila beccarii Aschers. was restricted to the intertidal mudflats in association with mangroves. The rich growth of seagrasses along the Tamil Nadu coast and Lakshadweep can be attributed mainly to high salinity, clarity of the water and sandy substratum. One hundred species of marine algae were recorded from the seagrass environments of India.     

The capacity for change and the unpredictability of the phytoplankton of the east coast of the United States

Differences in the change of numerical abundance from time totime through the year and place to place along the U.S. eastcoast show Skeletonema costatum to have a greater capacity forchange than Thalassionema nitzschioides. These differences emergefrom the unpredictable occurrence and absence of Skeletonemafrom time to time through the year and from year to year atWoods Hole and from place to place along the U.S. east coast.Skeletonema's greater capacity for change than Thatassionema'stransmit an inherently variable capacity for change to the wholephytoplankton, and the unpredictability of these species transmitsthis characteristic to the whole population too.