Dynamics and short-term survival of toxic cyanobacteria species in ballast water from NOBOB vessels transiting the Great Lakes—implications for HAB invasions

We measured the presence, viability and potential toxicity of cyanobacteria in ships’ ballast tanks during three domestic voyages through the North American Great Lakes. Using molecular methods, the toxin-producing forms of Microcystis and Anabaena were monitored in ballast water after ships’ ballast tanks were filled at their first port of call, and at subsequent ports as ships transited the Great Lakes. Microcystis was detected in ballast water at intermediate and final ports of call in all three experiments, but the presence of Anabaena was more variable, suggesting low abundance or patchy distribution in ballast tanks. Both species were detected in ballast water up to 11 days old. Detection of the microcystin synthetase gene, mcyE, in ballast tanks indicated entrained cells were capable of producing microcystin, and further analyses of RNA indicated the toxin was being expressed by Microcystis, even after 11 days in dark transit. These data demonstrate within-basin transport and delivery of planktonic harmful algal bloom (HAB) species to distant ports in the world's largest freshwater reservoir, with potential implications for drinking water quality. These implications are discussed with respect to management of microbial invasions and the fate of introduced phytoplankton in their receiving environment.     

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.     

International dissemination of epidemic Vibrio cholerae by cargo ship ballast and other nonpotable waters.

In 1991 and 1992, toxigenic Vibrio cholerae O1, serotype Inaba, biotype El Tor, was recovered from nonpotable (ballast, bilge, and sewage) water from five cargo ships docked in ports of the U.S. Gulf of Mexico. Four of these ships had taken on ballast water in cholera-infected countries; the fifth took on ballast in a noninfected country. Isolates examined by pulsed-field gel electrophoresis were indistinguishable from the Latin American epidemic strain, C6707; however, they differed significantly from the endemic Gulf Coast strain (VRL 1984), the sixth-pandemic strain (569-B), and a V. cholerae non-O1 strain isolated from a ship arriving from a foreign port. On the basis of our findings, the Food and Drug Administration recommended that the U.S. Coast Guard issue an advisory to shipping agents and captains requesting that ballast waters be exchanged on the high seas before entry of ships into U.S. ports.     

Phytoplankton and bacterial assemblages in ballast water of U.S. military ships as a function of port of origin, voyage time, and ocean exchange practices

We characterized the physical/chemical conditions and the algal and bacterial assemblages in ballast water from 62 ballast tanks aboard 28 ships operated by the U.S. Military Sealift Command and the Maritime Administration, sampled at 9 ports on the U.S. West Coast and 4 ports on the U.S. East Coast. The ballast tank waters had been held for 2–176 days, and 90% of the tanks had undergone ballast exchange with open ocean waters. Phytoplankton abundance was highly variable (grand mean for all tanks, 3.21 × 10⁴ viable cells m⁻³; median, 7.9 × 10³ cells m⁻³) and was unrelated to physical/chemical parameters, except for a positive relationship between centric diatom abundance and nitrate concentration. A total of 100 phytoplankton species were identified from the ballast tanks, including 23 potentially harmful taxa (e.g. Chaetoceros concavicornis, Dinophysis acuminata, Gambierdiscus toxicus, Heterosigma akashiwo, Karlodinium veneficum, Prorocentrum minimum, Pseudo-nitzschia multiseries). Assemblages were dominated by chain-forming diatoms and dinoflagellates, and viable organisms comprised about half of the total cells. Species richness was higher in ballast tanks with coastal water, and in tanks containing Atlantic or Pacific Ocean source waters rather than Indian Ocean water. Total and viable phytoplankton numbers decreased with age of water in the tanks. Diversity also generally decreased with water age, and tanks with ballast water age >33 days did not produce culturable phytoplankton. Abundance was significantly higher in tanks with recently added coastal water than in tanks without coastal sources, but highly variable in waters held less than 30 days. Bacterial abundance was significantly lower in ballast tanks with Atlantic than Pacific Ocean source water, but otherwise was surprisingly consistent among ballast tanks (overall mean across all tanks, 3.13 ± 1.27 × 10¹¹ cells m⁻³; median, 2.79 × 10¹¹ cells m⁻³) and was unrelated to vessel type, exchange status, age of water, environmental conditions measured, or phytoplankton abundance. At least one of four pathogenic eubacteria (Listeria monocytogenes, Escherichia coli, Mycobacterium spp., Pseudomonas aeruginosa) was detected in 48% of the ballast tanks, but toxigenic strains of Vibrio cholerae were not detected. For ships with tanks of similar ballasting history, the largest source of variation in phytoplankton and bacteria abundance was among ships; for ships with tanks of differing ballasting histories, and for all ships/tanks considered collectively, the largest source of variation was within ships. Significant differences in phytoplankton abundance, but not bacterial abundance, sometimes occurred between paired tanks with similar ballasting history; hence, for regulatory purposes phytoplankton abundance cannot be estimated from single tanks only. Most tanks (94%) had adequate records to determine the source locations and age of the ballast water and, as mentioned, 90% had had ballast exchange with open-ocean waters. Although additional data are needed from sediments that can accumulate at the bottom of ballast tanks, the data from this water-column study indicate that in general, U.S. Department of Defense (DoD) ships are well managed to minimize the risk for introduction of harmful microbiota. Nevertheless, abundances of viable phytoplankton with maximum dimension >50 μm exceeded proposed International Maritime Organization standards in 47% of the ballast tanks sampled. The data suggest that further treatment technologies and/or alternative management strategies will be necessary to enable DoD vessels to comply with proposed standards.     

Population genetics of Haminoea (Haloa) japonica Pilsbry, 1895, a widespread non-indigenous sea slug (Mollusca: Opisthobranchia) in North America and Europe

Haminoea japonica is an opisthobranch mollusk with a large non-indigenous range. This species is a vector for a parasite that causes the human skin disease cercarial dermatitis, and may have negative effects on populations of native species. Molecular evidence from the mitochondrial cytochrome c oxidase I gene and the histone 3 nuclear gene indicates that previously published morphology-based hypotheses on the spread of H. japonica out of Japan are correct. The most likely explanation for the current range of the species, which includes Japan, Korea, France, Spain, Italy, Canada and the USA is a recent, human-mediated dispersal from Japanese populations. The highest levels of nucleotide and haplotype diversity are found in Japan. Non-indigenous populations have low levels of genetic diversity (indicating bottlenecking). Haplotypes that were detected in the non-indigenous range of H. japonica have only been found in two localities in the native range; these two localities are in north-eastern Japan. In addition, the haplotype network structure and Spatial Analysis of Molecular Variance results confirm the origins of non-indigenous populations most likely trace to north-eastern Japan, which is where most Pacific oyster exports to North America also originated. Because there are no major shipping ports in north-eastern Japan, ballast water is less likely to be the mechanism of dispersal. The results of this study provide important data for the development of policies and regulations aimed to prevent further spread of this species in non-indigenous ranges.     

江阴港口外来船舶压载舱沉积物中甲藻包囊种类及组成

为了探究外来船舶压载舱沉积物中甲藻休眠包囊的种类组成及外来甲藻入侵风险, 本文根据包囊及其萌发细胞的形态特征辅以分子生物学信息对江阴港5艘外来船舶压载舱沉积物中的活体甲藻休眠包囊的种类进行鉴定和分析。共鉴定出甲藻休眠包囊29种(不含3种未鉴定种), 包括膝沟藻类、钙甲藻类、裸甲藻类、翼甲藻类以及原多甲藻类, 活体休眠包囊密度介于0.73-44.3 cysts/g DW。其中塔玛亚历山大藻复合种(Alexandrium tamarense species complex)、网状原角藻(Protoceratium reticulatum)、具刺膝沟藻(Gonyaulax spinifera)等有毒有害甲藻的包囊在取样的5艘船舶中均有发现。此外还发现了多种中国近海未报道的甲藻包囊种类, 其中一种经过分子手段确定为异常亚历山大藻(Alexandrium insuetum)包囊, 萌发和培养实验表明该种包囊能够在中国近海萌发并增殖, 有潜在的入侵风险和暴发赤潮的可能性。     

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.     

EVIDENCE FOR ASEXUAL RESTING CYSTS IN THE LIFE CYCLE OF THE MARINE PERIDINOID DINOFLAGELLATE,SCRIPPSIELLA HANGOEI1

Scrippsiella hangoei (Schiller) Larsen is a peridinoid dinoflagellate that grows during winter and spring in the Baltic Sea. In culture this species formed round, smooth cysts when strains were mixed, indicating heterothallic sexuality and hypnozygote production. However, cysts of the same morphology were also formed in clonal strains exposed to slightly elevated temperature. To better understand the role of cysts in the life cycle of S. hangoei, cyst formation and dormancy were examined in culture experiments and the cellular DNA content of flagellate cells and cysts was compared in clonal and mixed strains using flow cytometry. S. hangoei exhibited a high rate of cyst formation in culture. Cysts produced in both clonal and mixed strain cultures were thick‐walled and underwent a dormancy period of 4 months before germinating. The S. hangoei flagellate cell population DNA distributions consisted of 1C, intermediate, and 2C DNA, indicative of respective eukaryotic cell cycle phases G1, S, and G2M. The majority (>95%) of cysts had a measured DNA content equivalent to the lower 1C DNA value, indicating a haploid nuclear phase and an asexual mode of cyst formation. A small percentage (<5%) of cysts produced in the mixed strain culture had 2C DNA, and thus could have been diploid zygotes. These findings represent the first measurements of dinoflagellate resting cyst DNA content, and provide the first quantitative evidence for dinoflagellate asexual resting cysts. Asexual resting cysts may be a more common feature of dinoflagellate life cycles than previously thought.     

Life cycle,behavior and morphology of a new tide pool dinoflagellate,Gymnodinium pyrenoidosum sp. nov. (Gymnodiniales,Pyrrhophyta)

A new tide pool dinoflagellate,Gymnodinium pyrenoidosum Horiguchi et Chihara sp. nov. is described from central Japan. It was found to form dense blooms with a characteristic greenish color from April to November. The species exhibits a characteristic diurnal vertical migration and an alternation of a motile with a nonmotile phase, which are dependent on light intensity and tidal movement. Cells of the motile phase are unarmored and relatively small. They have a single, reticulate chloroplast, orange stigma situated near the sulcus and conspicuous pyrenoid in epicone. The alga reproduces itself by means of zoospores which are produced by the bipartition of protoplasm within the parent cell wall during the nonmotile stage which occurs at night. The occurrence of another type of motile cell, termed a macroswarmer, which differs from normal zoospore in size and shape has also been demonstrated.     

Characterization of biofilm-forming marine bacteria and their effect on attachment and germination of algal spores

In this work, 37 bacterial strains isolated from biofouling of marine organisms and from the Museum of Heterotrophic Bacteria of the National Scientific Center of Marine Biology were studied. The strains were identified based on their phenotypic characteristics and on the fatty acid composition of their cell wall lipids. Members of the genus Pseudoalteromonas prevailed both in associated microflora of two dinoflagellate clones and in the biofilms from marine hydrobionts. Associated microflora included also members of the CFB cluster, Bacillus, Sulfitobacter, Acinetobacter, Shewanella, and Psychrobacter. A considerable portion of strains (48.6%) exhibited antimicrobial activity. Antifouling activity against algal spores was studied using single-species bacterial biofilms and the spores of Ulva lactuca и Undaria pinnatifida, the algae most common in the Sea of Japan. Strong inhibitory effect on attachment of Ulva and Undaria spores was observed for 75 and 51% of the strains, respectively. Attached spores were, however, less sensitive to the inhibitory action of biofilms. Species specificity of algal response to bacteria was shown, with a strain having different effect on the spores of different algal species. Biotechnologically promising strains were determined, which exhibited high activity against the spores of macroalgae and could probably be used as producers of antifouling substances and as components of antifouling coatings. No relation was found between antifouling activity of bacteria and the source of their isolation. Our results indicate wide occurrence of bacteria with antifouling activity among associated microflora of marine hydrobionts and demonstrate the extent of complexity and diversity of relations between bacterial biofilms and algal spores.     

Pandemic Serotypes of Vibrio cholerae Isolated from Ships’ Ballast Tanks and Coastal Waters: Assessment of Antibiotic Resistance and Virulence Genes (tcpA and ctxA)

There is concern that ships’ ballasting operations may disseminate Vibrio cholerae to ports throughout the world. Given evidence that the bacterium is indeed transported by ships, we isolated pandemic serotypes O1 and O139 from ballast tanks and characterized them with respect to antibiotic resistance and virulence genes ctxA and tcpA. We carried out concurrent studies with V. cholerae isolated from coastal waters. Of 284 isolates, 30 were serotype O1 and 59 were serotype O139. These serotypes were overrepresented in ballast tanks relative to the coastal waters sampled. All locations, whether coastal waters or ballast tanks, yielded samples from which serotype O1, O139, or both were isolated. There were three groups among the 62 isolates for which antibiotic characterization was conclusive: those exhibiting β-lactamase activity and resistance to at least one of the 12 antibiotics tested; those negative for β-lactamase but having antibiotic resistance; those negative for β-lactamase and registering no antibiotic resistance. When present, antibiotic resistance in nearly all cases was to ampicillin; resistance to multiple antibiotics was uncommon. PCR assays revealed that none of the isolates contained the ctxA gene and only two isolates, one O139 and one O1, contained the tcpA gene; both isolates originated from ballast water. These results support the bacteriological regulations proposed by the International Maritime Association for discharged ballast water.     

Transport of diatom and dinoflagellate resting spores in ships' ballast water: implications for plankton biogeography and aquaculture

Diatom and dinoflagellate species that are not endemic to aregion can be inadvertently introduced when their resistantresting stages are discharged with the ballast-tank waters andsediments of bulk cargo vessels. A survey of 343 cargo vesselsentering 18 Australian ports showed that 65% of ships were carryingsignificant amounts of sediment on the bottom of their ballasttanks. All of these samples contained diatoms, including speciesthat are not endemic to Australian waters. Diatom resting spores,especially of Chaetoceros, were also detected. Dinoflagellateresting spores (cysts) were present in 50% of the sediment samples.Of the 53 cyst species identified, 20 (including Diplopelta,Diplopsalopsis, Gonyaulax, Polykrikos, Protoperidinium, Scrippsiellaand Zygabikodinium spp.) were successfully germinated to produceviable cultures. Such diversity of diatom and dinoflagellatespecies in ships' ballast water suggests that the apparent cosmopolitanismof many coastal phytoplankton species may be due partly to theglobal transport of seawater ballast. Of considerable concernwas the detection in 16 ships of cysts of the toxic dinoflagellatesAlexandrium catenella, Alexandrium tamarense and Gymnodiniumcatenatum. One single ballast tank was estimated to contain>300 million viable A.tamarense cysts, some of which weresuccessfully germinated in the laboratory to produce toxic cultures.These toxic dinoflagellate species, which can contaminate shellfishwith paralytic shellfish poisons, pose a serious threat to humanhealth and the aquaculture industry. Ballast-water quarantinemeasures recently introduced in Australia are discussed. Mid-oceanexchange of ballast water is only partially effective in removingdinoflagellate cysts which have settled to the bottom of ballasttanks. The present work indicates that the most effective measureto prevent the spreading of toxic dinoflagellate cysts via ships'ballast water would be to avoid taking on ballast water duringdinoflagellate blooms in the water column of the world's ports.     

Algicidal Effects of a Novel Marine Pseudoalteromonas Isolate (Class Proteobacteria,Gamma Subdivision) on Harmful Algal Bloom Species of the Genera Chattonella,Gymnodinium, and Heterosigma

During a bacterial survey of the Huon Estuary in southern Tasmania, Australia, we isolated a yellow-pigmented Pseudoalteromonas strain (class Proteobacteria, gamma subdivision), designated strain Y, that had potent algicidal effects on harmful algal bloom species. This organism was identified by 16S rRNA sequencing as a strain with close affinities to Pseudoalteromonas peptidysin. This bacterium caused rapid cell lysis and death (within 3 h) of gymnodinoids (including Gymnodinium catenatum) and raphidophytes (Chattonella marina and Heterosigma akashiwo). It caused ecdysis of armored dinoflagellates (e.g., Alexandrium catenella, Alexandrium minutum, and Prorocentrum mexicanum), but the algal cultures then recovered over the subsequent 24 h. Strain Y had no effect on a cryptomonad (Chroomonas sp.), a diatom (Skeletonema sp.), a cyanobacterium (Oscillatoria sp.), and two aplastidic protozoans. The algicidal principle of strain Y was excreted into the seawater medium and lost its efficacy after heating. Another common bacterial species, Pseudoalteromonas carrageenovora, was isolated at the same time and did not have these algicidal effects. The minimum concentrations of strain Y required to kill G. catenatum were higher than the mean concentrations found in nature under nonbloom conditions. However, the new bacterium showed a chemotactic, swarming behavior that resulted in localized high concentrations around target organisms. These observations imply that certain bacteria could play an important role in regulating the onset and development of harmful algal blooms.Historically, the dynamics of marine bacterial and algal populations have been studied largely in isolation. Increasing evidence is now pointing toward a close spatial and temporal association between the two and recently attention has been focused on phagocytosis of bacteria by photosynthetic flagellates (21, 28, 30). In contrast, the importance of inhibitory or predatory bacteria in regulating populations of different algal species has received relatively little attention (9, 11). Some bacteria may selectively promote bloom formation by algal species (13), while other bacteria have algicidal effects and are involved in the termination and decomposition of algal blooms (12). The latter finding has raised the possibility of bacterial control of harmful algal blooms (19). There is little data on the occurrence of marine algicidal bacteria outside Japan, where toxic blooms are frequent events (20), and algicidal bacteria have been isolated during toxic blooms of naked dinoflagellates and raphidophytes (9).Gymnodinium catenatum (a causative organism of paralytic shellfish poisoning) is thought to have been introduced into southern Tasmania via ballast water after 1973, and in some years it has a severe negative impact on the shellfish industry (16). Previous efforts to understand and predict the seasonal and interannual variability of harmful algal blooms have largely focused on the environmental factors that affect dinoflagellate growth in the water column, notably water temperature, rainfall, and water column stability (16). Rainfall and estuarine flow patterns also largely determine the allochthonous input of dissolved organic matter (DOM), which is a source of organic carbon for bacteria (27) and is possibly involved in micronutrient dynamics that promote G. catenatum growth (3, 6). As part of a study investigating DOM, bacteria, and algal interactions in the Huon Estuary (24), we isolated two bacterial strains that we tested for possible alga-bacterium interactions by using cultures of G. catenatum. Both bacteria appeared to be Pseudoalteromonas species, which are extremely common, slightly halophilic, gram-negative bacteria found in many marine ecosystems. Preliminary observations indicated that one of the strains was extremely toxic towards G. catenatum, while the other was more benign. The aims of this study were (i) to determine the taxonomic identity of the bacteria, (ii) to document by light microscopy the sequence of algal cell lysis after exposure to an algicidal Pseudoalteromonas strain and compare this lysis to the effect of the more benign Pseudoalteromonas species, (iii) to define the minimum bacterial concentrations required for algicidal effects and compare these concentrations to concentrations in natural water samples, and (iv) to investigate the range of potential target organisms for the bacterium.     

Green Autofluorescence in Dinoflagellates, Diatoms, and Other Microalgae and Its Implications for Vital Staining and Morphological Studies

Green autofluorescence (GAF) has been described in the short flagellum of golden and brown algae, the stigma of Euglenophyceae, and cytoplasm of different life stages of dinoflagellates and is considered by some researchers a valuable taxonomic feature for dinoflagellates. In addition, green fluorescence staining has been widely proposed or adopted to measure cell viability (or physiological state) in areas such as apoptosis of phytoplankton, pollutant stresses on algae, metabolic activity of algae, and testing treatment technologies for ships' ballast water. This paper reports our epifluorescence microscopic observations and quantitative spectrometric measurements of GAF in a broad phylogenetic range of microalgae. Our results demonstrate GAF is a common feature of dinoflagellates, diatoms, green algae, cyanobacteria, and raphidophytes, occurs in the cytoplasm and particularly in eyespots, accumulation bodies, spines, and aerotopes, and is caused by molecules other than chlorophyll. GAF intensity increased with time after cell death or fixation and with excitation by blue or UV light and was affected by pH. GAF of microalgae may be only of limited value in taxonomy. It can be strong enough to interfere with the results of green fluorescence staining, particularly when stained samples are observed microscopically. GAF is useful, however, for microscopic study of algal morphology, especially to visualize cellular components such as eyespots, nucleus, aerotopes, spines, and chloroplasts. Furthermore, GAF can be used to visualize and enumerate dinoflagellate cysts in marine and estuarine sediments in the context of anticipating and monitoring harmful algal blooms and in tracking potentially harmful dinoflagellates transported in ships' ballast tanks.     

Retrospective eDNA assessment of potentially harmful algae in historical ship ballast tank and marine port sediments

Microalgal bloom events can cause major ecosystem disturbances, devastate local marine economies, and endanger public health. Therefore, detecting and monitoring harmful microalgal taxa is essential to ensure effective risk management in waterways used for fisheries, aquaculture, recreational activity, and shipping. To fully understand the current status and future direction of algal bloom distributions, we need to know how populations and ecosystems have changed over time. This baseline knowledge is critical for predicting ecosystem responses to future anthropogenic change and will assist in the future management of coastal ecosystems. We explore a NGS metabarcoding approach to rapidly identify potentially harmful microalgal taxa in 63 historic and modern Australian marine port and ballast tank sediment samples. The results provide a record of past microalgal distribution and important baseline data that can be used to assess the efficacy of shipping guidelines, nutrient pollution mitigation, and predict the impact of climate change. Critically, eDNA surveys of archived sediments were able to detect harmful algal taxa that do not produce microscopic fossils, such as Chattonella, Heterosigma, Karlodinium, and Noctiluca. Our data suggest a potential increase in Australian harmful microalgal taxa over the past 30 years, and confirm ship ballast tanks as key dispersal vectors. These molecular mapping tools will assist in the creation of policies aimed at reducing the global increase and spread of harmful algal taxa and help prevent economic and public‐health problems caused by harmful algal blooms.     

GENETIC VARIATION AMONG STRAINS OF THE TOXIC DINOFLAGELLATE GYMNODINIUM CATENATUM (DINOPHYCEAE)

The toxic dinoflagellate Gymnodinium catenatum Graham has formed recurrent toxic blooms in southeastern Tasmanian waters since its discovery in the area in 1986. Current evidence suggests that this species might have been introduced to Tasmania prior to 1973, possibly in cargo vessel ballast water carried from populations in Japan or Spain, followed by recent dispersal to mainland Australia. To examine this hypothesis, cultured strains from G. catenatum populations in Australia, Spain, Portugal, and Japan were examined using allozymes and randomly amplified polymorphic DNA (RAPD). Allozyme screening detected very limited polymorphism and was not useful for population comparisons; however, Australian, Spanish, Portuguese, and Japanese strains showed considerable RAPD diversity, and all strains examined represented unique genotypes. Multidimensional scaling analysis (MDS) of RAPD genetic distances between strains showed clear separation of strains into three nonoverlapping regional clusters: Australia, Japan, and Spain/Portugal. Analysis of genetic distances between strains from the three regional populations indicated that Australian strains were almost equally related to both the Spanish/Portuguese population and the Japanese population. Analysis of molecular variance (AMOVA) found that genetic variation was partitioned mainly within populations (87%) compared to the variation between the regions (8%) and between populations within regions (5%). The potential source population for Tasmania’s introduced G. catenatum remains equivocal; however, strains from the recently discovered mainland Australian population (Port Lincoln, South Australia, 1996) clustered with Tasmanian strains, supporting the notion of a secondary relocation of Tasmanian G. catenatum populations to the mainland via a shipping vector. Geographic and temporal clustering of strains was evident among the Tasmanian strains, indicating that genetic exchange between neighboring estuaries is limited and that Tasmanian G. catenatum blooms are composed of localized, estuary-bound subpopulations.     

A biological survey of ballast water in container ships entering Hong Kong

The role of ballast water in the introduction of exotic species has recently received extensive attention. The aim of this study is to assess the importance of ballast water discharge as a vector for the introduction of exotic species into Hong Kong waters. Twelve ballast water samples were collected from 5 container ships entering Hong Kong between June1994 and October 1995. The ballast water originated from ports on both sides of the Pacific Ocean. At least 81 species from 8 animal phyla and 5 protist phyla were found. Most of the major marine taxonomic groups were represented and many planktonic larval stages were included. Species richness in the ballast tanks decreased with the age of ballast water. Copepoda was the most diverse and abundant taxonomic group. The density of calanoid and cyclopoid copepods decreased with the age of ballast water, but that of harpacticoid copepods did not change significantly with time. Bivalve, crustacean, polychaete and ascidian larvae from ballast water samples were observed to settle in laboratory culture tanks. The mussel Mytilopsis sallei which was introduced to Hong Kong in 1980, was one of the bivalves that settled readily. Results of this study indicate that ballast water can be a major source for the introduction of exotic species to Hong Kong waters. Regulatory guidelines on the discharge of ballast water should be established. This revised version was published online in August 2006 with corrections to the Cover Date.     

Reflections on the ballast water dispersal—harmful algal bloom paradigm

The ballast water dispersal—HAB paradigm, increasingly invoked circumstantially to explain puzzling and unaccountable HAB species outbreaks when lacking the multiple tests of confirmation recommended by Bolch and de Salas (2007), is evaluated. The types and examples of natural dispersions and taxon cycles are compared to exotic species bloom behavior linked to ballast water vectoring. The regional spreading, bloom behavior and disjunct distributions of the brown tide pelagophyte Aureococcus anophagefferens and the toxic dinoflagellate Gymnodinium catenatum, attributed to ballast water vectoring, are used as representative examples to evaluate the general application of the ballast water—HAB paradigm and associated interpretative problems. Human-aided emigration has a seeding and colonization ecology that differs from bloom ecology. For self-sustaining blooms to occur, these two ecologies must be accommodated by habitat growth conditions. The three stages that a non-native species must pass through (pioneering, persistence, community entry) to achieve colonization, community maintenance, and to bloom, and the niche-related factors and role of habitat disturbance are discussed. The relevance of cryptic occurrences, cyst deposits, dormancy periods and bloom rhythms of HAB species to their blooms attributed to ballast water-assisted introductions is also sketched. The different forms of HAB species rarity, their impact on the ballast water dispersal—HAB paradigm, and the dispersion and blooms of specialist and generalist HAB species are discussed. The remarkable novel and, often, monospecific blooms of dinoflagellate HAB species are being paralleled by similar eruptive bloom behavior cutting across phylogenetic lines, and being found also in raphidophytes, haptophytes, diatoms, silicoflagellates, etc. These blooms cannot be explained only as seeding events. An ecological release of ‘old barriers’ appears to be occurring generally at coastal bloom sites, i.e. something significant is happening ecologically and embedded within the ballast water—HAB paradigm. There may be a relationship between Life Form type [Smayda, T.J., Reynolds, C.S., 2001. Community assembly in marine phytoplankton: application of recent models to harmful dinoflagellate blooms. J. Plankton Res. 23, 447–461] and mode of expatriation; HAB dinoflagellate species commonly reported to produce ballast water-assisted toxic blooms invariably are members of cyst-producing Life Forms IV, V, VI. Ballast water vectoring of Life Forms I, II, III is rarely reported, even though many produce cysts, and where their novel introductions do occur they are more likely to be ichthyotoxic and vectored in shellfish stock consignments. The relevance of, and need to distinguish between morphospecies and their geographic/ribotype clades are discussed based on the Alexandrium tamarense/catenella/fundyense complex. Morphospecies-level ballast water dispersions are probably minor compared to the dispersal of the different ribotypes (toxic/non-toxic clades) making up HAB morphospecies; the redistribution and admixture of genotypes should be the focus. Ballast water-assisted expatriations impact the global occurrence of HABs through the direct transfer of previously absent species or introduction of genetic strains from the donor habitat that are ecologically favored over resident strains. The hybridization of species may be of potentially greater impact, resulting from the (1) mating of individuals from the donor and recipient habitats, or (2) through the interbreeding of strains introduced from two different donor sites into the recipient site, and whose progeny have greater ecological fitness than indigenous strains. Exceptional ecological changes of some sort appear to be occurring globally which, in combination with the genetically altered ecophysiological behavior of HAB species linked to ballast water dispersion and admixture, underpins the global HAB phenomenon. The impact of ballast water and shellfish transplantation on HABs and phytoplankton community ecology, generally, is considerably greater than the current focus on HAB species distributions, vectoring, and blooms. The methodological, investigative and conceptual potential of the ballast water—HAB paradigm should be exploited by developing a GEOHAB type intiative to advance quantification of global HAB ecology.     

Marine bacteria antagonistic to the harmful algal bloom species Alexandrium tamarense (Dinophyceae)

As part of efforts to enhance the strategies employed to manage and mitigate algal blooms and their adverse effects, algicidal bacteria have shown promise as potential suppressors of these events. Nine strains of bacteria algicidal against the toxic dinoflagellate, Alexandrium tamarense, were isolated from the East Sea area, China. Sequence analysis of 16S rDNA showed that all the algicidal bacteria belonged to the γ-proteobacteria subclass and the genera Pseudoalteromonas (strain SP31 and SP44), Alteromonas (strain DH12 and DH46), Idiomarina (strain SP96), Vibrio (strain DH47 and DH51) and Halomonas (strain DH74 and DH77). To assess the algicidal mode of these algicidal bacteria, bacterial cells and the filtrate from bacterial cultures were inoculated into A. tamarense cultures, and fluorescein diacetate vital stain was applied to monitor the growth of the algal cells. The results showed that all the algicidal bacteria exhibited algicidal activity through an indirect attack since algicidal activity was only detected in cell free supernatants but not the bacterial cells. This is the first report of bacteria from the genus Idiomarina showing algicidal activity to the toxic dinoflagellate A. tamarense and these findings would increase our knowledge of bacterial–algal interactions and the role of bacteria during the population dynamics of HABs.     

In situ hatching of invertebrate diapausing eggs from ships' ballast sediment

Ships that enter the Great Lakes laden with cargo carry only residual ballast water and sediment in ballast tanks. These ships are designated ‘no ballast on board’ (NOBOB) and constitute > 90% of inbound traffic. We conducted in situ experiments using emergence traps to assess the viability and the introduction potential of invertebrate diapausing stages present in ships’ ballast sediment. All trials commenced while vessels operated on the lower lakes (Erie, Ontario) and were completed 6–11 days later at ports on the upper lakes (Michigan, Lake Superior). Eight trials were conducted on four ships using five different ballast sediments. Hatching was observed on every ship, although not from all sediments on all ships. Overall hatch rates were very low (0.5 individuals per 500 g sediment), typically involving activation of < 0.05% of total eggs present. Five species of rotifers and copepod nauplii were hatched from ballast sediments, although only one or two species typically hatched from any one sediment. Results of this study indicate that hatching of diapausing eggs contained in ballast sediment of NOBOB ships poses a relatively low risk of invasion to the Great Lakes. However, as reproduction may occur in tanks, and non‐indigenous species may be involved in numerous introduction events, the risk posed by this vector is small but potentially important. While dormancy is a characteristic enabling enhanced survival during transportation in ballast tanks, it becomes a hindrance for introduction.