Occurrence of the polyamines caldopentamine and homocaldopentamine in axenic cultures of the red tide flagellates Chattonella antiqua and Heterosigma akashiwo (Raphidophyceae)

The polyamines caldopentamine and homocaldopentamine were detected in axenic strains of Chattonella antiqua and Heterosigma akashiwo ( Raphidophyceae ), respectively, as well as spermidine, the most abundant polyamine in both phytoplankton species. Trace amounts of putrescine, diaminopropane and norspermine were also detected in both species. Spermine was detected only from C. antiqua . These long linear polyamines are characteristic components of thermophilic bacteria. The detection from two species of Raphidophyceae indicates that the occurrence of long linear polyamines is not restricted to thermophilic microorganisms.     

Photosynthesis does influence superoxide production in the ichthyotoxic alga Chattonella marina (Raphidophyceae)

The ichthyotoxic flagellate Chattonella marina produces extraordinarylevels of the reactive oxygen species (ROS) superoxide, whichis partially controlled by electrons donated through photosyntheticelectron transfer. We demonstrate that ROS can be significantlyreduced using the photosynthesis blocking herbicide DCMU. Assuperoxide is considered a contributor to the toxicity of C.marina, fish mortality may be more prominent during daylighthours.     

Development of microsatellite markers for the red tide‐forming harmful species Chattonella antiqua,C. marina,and C. ovata (Raphidophyceae)

We have developed 11 microsatellite markers that are specific to Chattonella antiqua, C. marina, and C. ovata, the red tide‐forming harmful phytoplanktons. The 11 loci were amplified in the three species. The number of alleles per locus ranged from 5 to 16. The three species shared most microsatellite regions, although the genetic differences in specific loci were detected among them. These markers of the Chattonella species will be beneficial for biogeographical, detailed taxonomic, studies.     

Diazo-reaction positive substance observed in the cortex of Chattonella antiqua

In order to investigate the causative factors responsible for removal of mucous coat from the gill lamellae of young yellowtail, Seriola quinqueradiata by red tide, diazo-reactions were employed for planktons and their media. The concentration of NO2- in the medium containing the raphidophyceae, Chatonella antiqua (ca 2000 cells/ml), was 0.70 +/- 0.05 (mu g/ml +/- SEM). In addition, diazo-reaction positive substances (NOx) which may degenerate the mucous, was highly concentrated in the cortex (perikaryon) of Chattonella antiqua. Morphologically, mucocysts, and chloroplats were likewise present in the cortex. Mucocysts were packed with fine fibrous content. Histochemically, the mucocysts were stained with PAS and had an abundance of nitrogen oxides (NOx). We observed discharge of the fibrous material from the mucocysts. These results suggest that when Chattonella antiqua is passing between the gill lamellae, NOx discharged from the mucocysts may act on the mucous, leading to the degeneration and concomitant removal of the mucous coat from gill lamellae.     

Comparative ecophysiology of the harmful alga Chattonella marina (Raphidophyceae) form South Australian and Japanese waters

The raphidophyte flagellate Chattonella marina was successfully cultured from Boston Bay (South Australia), coincident with mass mortality of farmed bluefin tuna (Thunnus maccoyii) in April 1996. Grown under laboratory conditions at 150 mol m^-2 s^-1 irradiance, optimal growth (>0.5 day^-1) occurred at a temperature of 25C and a salinity of 30 p.s.u., but good growth (>0.3 day^-1) also occurred between a temperature of 10 and 30°C and at a salinity of 15-45 p.s.u. However, cultures grow much faster at an irradiance of 450 mol m^-2 s^-1 (1.08 day^-1). While Australian C.marina had similar temperature and salinity requirements as well-studied Japanese cultures from the Seto Inland Sea, the Australian strains exhibited a light saturation level for growth four times higher than that reported from Japan (150 mol m^-2 s^-1). An adaptation to higher light intensities was reflected in higher concentrations of microsporine-like amino acids in the Australian strains. The different light adaptation phenotypes were still apparent after long-term culturing under similar physiological conditions. Potential growth habitats for this ichthyotoxic flagellate in the Australian region and implications for finfish aquaculture industries are discussed.      

Chattonella marina (Raphidophyceae), a potentially toxic alga in the Salton Sea, California

Chattonella marina was found in high abundance in the phytoplankton of the Salton Sea from April to November, 1997. Maximum mean density was over 600 cells ml^–1 mid-lake. It was not detected from January 1997 to March 1997 or in January and February 1998. Transmission electron microscope studies show the typical Chattonella features: a teardrop-shaped nucleus, numerous radially arranged chloroplasts and lack of a cell wall. Gills of fish collected at the height of the Chattonella bloom showed damage to the primary and secondary lamellae and increased numbers of mucus cells. To our knowledge this is the first report of a species of Chattonella in a salt lake.     

Inductive and Inhibitory Effects of Light on Cell Division in Chattonella antiqua

The cell division of a red tide flagellate, Chattonella antiqua,was synchronously induced under light and dark regimes of 10L14D(a light period, L, for 10 h followed by a dark period, D, for14 h), 12L12D and l4L10D. In all regimes cell number began toincrease ca. 14 h after the onset of L and almost doubled duringone LD cycle. When the light-off timing of the last L was changedor the whole L was shifted, cells that had been synchronizedunder 12L12D invariably began to divide ca. 14 h after the onsetof L. This shows that the timing of cell division was determinedby the time of the onset of L. When cells were continuously exposed to light after a cell division,the subsequent cell division was inhibited. This effect waslimited to cells that had been synchronized under short-dayconditions. Thus it can be concluded that light has both inductive and inhibitoryeffects on cell division in this alga, the latter effect dependingupon the previously given light and dark regimes. (Received December 21, 1984; Accepted February 28, 1985)     

Assessment of EvaGreen-based quantitative real-time PCR assay for enumeration of the microalgae Heterosigma and Chattonella (Raphidophyceae)

Heterosigma akashiwo and Chattonella species (Raphidophyceae) are difficult to detect and quantify in environmental samples because of their pleomorphic and fragile cell nature. In this study, we developed a quantitative real-time polymerase chain reaction (qRT-PCR) assay for the enumeration of these algal taxa using a new DNA-binding dye, EvaGreen. Species-specific qRT PCR primers to H. akashiwo, Chattonella antiqua, Chattonella marina, Chattonella ovata, and Chattonella subsalsa were designed to target the ITS2 rRNA gene intergenic region. Primer specificities were tested via BLAST searches. In addition, specificity was verified using empirical tests, including competitive PCR. The qRT PCR assay analyzing C _t value and the log of cell number showed a significant linear relationship (r ²?≥?0.997). When light microscopy was used to monitor the population dynamics of targeted Raphidophyceae from Lake Shihwa, H. akashiwo was detected in ten samples and no Chattonella spp. were detected (70 samples collected from May, 2007 to January, 2008). In contrast, when the qRT-PCR assay was used, H. akashiwo was detected in 41 samples. C. antiqua, C. marina, and C. ovata were detected in eight samples. Most of the samples analyzed using qRT-PCR assays showed higher algal numbers than did those assayed via microscopy, suggesting that the enumeration of Raphidophyceae via classic microscopic methods most likely underestimates true algal concentration.     

Mortality in the initial ontogeny of Paralabrax maculatofasciatus (Actinopterygii, Perciformes, Serranidae) caused by Chattonella spp. (Raphidophyceae)

Fish are particularly sensitive to metabolites produced by Raphidophyte species and these have caused intensive fish kills in several countries. However, the effects on embryos of marine fish are unknown but could probably provoke an important impact on new stock recruitment and hence on fisheries. We evaluated the toxic effects of Chattonella spp. strains from the Gulf of California on three development stages of spotted sand bass (Paralabrax maculatofasciatus): embryo in segmentation stage (ES), embryo (EM), and eleutheroembryo (EL). Embryos (ES) were exposed to different cell concentrations of Chattonella subsalsa, Chattonella marina, Prorocentrum micans, and f/2 medium as control. Also, one set of embryos was tested with cell-free media for C. subsalsa cultures. Incubation lasted until embryos reached apterolarva phase. The ES was the most sensitive stage reaching 98% mortality with C. subsalsa, followed by cell-free media of C. subsalsa cultures, with mortalities close to 90%, whereas EM and EL phases presented mortalities below 60%. This work demonstrates that larval stages of P. maculatofasciatus are highly sensitive to short time exposure to all Chattonella spp. strains tested, that direct physical contact with cells is not required to cause mortality, and that the toxic effect is more pronounced when embryos hatch.     

Chemotaxis towards inorganic phosphate in the red tide alga Chattonella antiqua

The red tide flagellate, Chattonella antiqua, was shown to beattracted to inorganic phosphate in both aged seawater and artificialseawater by well-test assays.     

Effects of temperature and salinity on the growth of the red tide flagellates Heterocapsa circularisquama (Dinophyceae) and Chattonella verruculosa (Raphidophyceae)

Growth responses of the red tide flagellates, Heterocapsa circularisquama(Dinophyceae) and Chattonella verruculosa (Raphidophyceae),were examined with 36 different combinations of temperature(5–30°C) and salinity (10–35 PSU). Heterocapsacircularisquama did not grow at or below a temperature of 10°C.The maximum growth rate of H.circularisquama (1.3 divisionsday^–1) was obtained with a combination of 30°C and30 PSU. In contrast, C. verruculosa did not grow at 10 PSU andat temperatures of 25°C or more. The maximum growth rateof C. verruculosa (1.74 divisions day^–1) was obtainedwith a combination of 15°C and 25 PSU. A significant temperature-salinityinteraction on growth was found by factorial analysis. Basedon the physiological characteristics obtained in the presentstudy, these novel flagellates have a potential for future outbreaksof red tides in pre viously unaffected waters.     

Phosphate metabolism during diel vertical migration in the raphidophycean alga, Chattonella antiqua

The ecological advantage of diel vertical migration on the nutrition and accumulation of Chattonella antiqua, which is one of the dominant red-tide forming phytoplankton species in the Seto Inland Sea of Japan, was examined using a large axenic culture tank, in which vertical stratification of salinity, temperature and nutrients was maintained, analogous to natural conditions observed when red tides occur. C. antiqua was capable of migrating through very sharp salinity and temperature gradients. At night the species migrated to the deep nutrient-rich water and assimilated nutrients. During the daytime it migrated to the nutrient-depleted surface water and used the accumulated nutrients for photosynthesis. Nitrogen uptake was synchronized with phosphate uptake. ³¹P-NMR spectroscopy during the migration experiment revealed that C. antiqua has the capability of nocturnal phosphate uptake in the deep nutrient-rich water, but no capability of synthesizing polyphosphate, which was considered to be the intracellular phosphate pool. These findings were compared with those reported for another raphidophycean, Heterosigma akashiwo. Although both species carry out vertical migration and nocturnal nutrient uptake, only H. akashiwo has the capability of making an intracellular polyphosphate pool. This revised version was published online in July 2006 with corrections to the Cover Date.     

Generation of superoxide anion radicals by the marine phytoplankton organism, Chattonella antiqua

The generation of superoxide anion radical (O₂^–, believedto be a causative factor in the killing of fish by the phytoplanktonChattonella antiqua, has been examined using several methods:electrochemical technique, reduction of ferricytochrome c andfluorescent laser microscope. Evidence is presented to suggestthat these organisms release superoxide continuously while theyare living, even in the resting state. Additional generationof O₂^– accompanies the discharge of mucocysts, and istriggered when they are exposed to mucus from the gill lamellaeof fish. Such instantaneous generation of O₂^– is alsoinduced when the organisms are in contact with an electrodepoised at a potential of +0.1 V versus Ag/AgCI, which is positiveenough to oxidize O₂^– to O₂.     

Comparative studies on the fish-killing activities of Chattonella marina isolated in 1985 and Chattonella antiqua isolated in 2010, and their possible toxic factors

Chattonella antiqua isolated in 2010 showed extremely more potent fish-killing activities against red sea bream, Japanese horse mackerel, and blue damselfish than those of Chattonella marina isolated in 1985. Chemiluminescence and electron spin resonance (ESR) analyses suggested greater reactive oxygen species (ROS)-producing activity of C. antiqua than that of C. marina. Sodium benzoate, a hydroxyl radical scavenger, significantly suppressed the fish-killing activity of C. antiqua on blue damselfish. The chlorophyll level in the gill tissue of blue damselfish exposed to flagellate cells increased along with the exposure time, and the cell count of gill-associated C. antiqua estimated with chlorophyll level was higher than that of C. marina. These results suggest that the ROS-producing activity and affinity of Chattonella cells to the gill surface may be important factors influencing the fish-killing activity of Chattonella species.     

Nuclear Deformation Causes DNA Damage by Increasing Replication Stress

1. Download : Download high-res image (175KB)
2. Download : Download full-size image

     

Characterization of Sequences from Rape (Brassica napus) Nuclear DNA which Facilitate Autonomous Replication of Plasmids in Yeast

From Hind III digests of Brassica napus (rape) nuclear DNA,seven DNA fragments were obtained which conferred on plasmidsthe ability to replicate autonomously in yeast. Six of thesewere analysed by restriction enzyme mapping, yielding six differentmaps and two were further characterized by defining the minimumeffective length and by sequencing. One of these had a minimumeffective length of 220 base pairs, was 86% A.T. and containeda direct match to the consensus sequence obtained from otherDNA's (i.e. DNA from other organisms) which replicate autonomouslyin yeast. The second fragment had a minimum effective lengthof 926 base pairs, was 69% A.T. and contained a 9/11 match tothe consensus sequence. Key words: Brassica napus, nuclear DNA, autonomous replication, plasmids, yeast     

Nuclear DNA Amounts in Mosses (Musci)

A comparative investigation into nuclear DNA amounts using flowcytometry and video-based Feulgen densitometry was carried outin 289 accessions of 138 different moss taxa (Bryatae), originatingfrom Austria, Switzerland, Spain, Mexico and the USA. Samplingincluded species from all major moss clades (except Sphagnum).Flow cytometry data agreed highly with the Feulgen data, whichonce again demonstrates the high reliability of both methodsfor DNA amount determination. For the first time, extensivedata on absolute C-values of mosses are available. Haploid DNAcontents (1C) ranged from 0.174 to 2.16 pg, which representsonly about a 12-fold variation. This low C-value variation isremarkable when compared to angiosperms which vary approx. 1000-fold.C-values are usually relatively constant within genera and evenfamilies; however, genera with varying C-values also exist.From the low frequency observed, secondary polyploidy playsonly a minor role in mosses. Possible reasons for the low C-valuevariation are discussed. Copyright 2000 Annals of Botany Company Mosses, Bryatae, genome size, nuclear DNA amounts, C-value variation, Feulgen, flow cytometry, densitometry, image analysis     

Nuclear DNA Amounts in Palms (Arecaceae)

Nuclear DNA amounts are reported for 83 species and 53 genera of palms, covering all of the six subfamilies. 4C DNA contents range between 3.89 and 55.62 pg in diploids, showing an approximate 14.3-fold variation in genome size. Polyploids have DNA contents of up to 156.40 pg/4c which demonstrates a 40.2-fold variation. Diploids with high DNA contents occur in three subfamilies of palms (Coryphoideae, Calamoideae, Arecoideae), and seem to be further restricted to particular tribes or subtribes (Thrinacinae, Borasseae, Lepidocaryeae, Caryoteae, some subtribes of Areceae). Palms from the subfamilies Nypoideae and Phytelephantoideae have the lowest DNA amounts, followed by the Phoeniceae and the Corypheae: Livistoninae from the subfamily Coryphoideae. Although DNA amounts in some genera and subtribes are usually constant, e.g., in Phoenix, Phytelephas, the Livistoninae, Dypsidinae, diploid Butiinae), considerable variation occurs at the diploid level in some large and apparently actively evolving genera such as Chamaedorea, Pinanga, Cenoma and possibly Bactris. Formaldehyde fixation is recommended for palms, as conventional ethanol-acetic acid fixation has proved to be unsuitable for DNA estimation of Feulgen-stained nuclei by microdensitometry, since it can lead to errors up to 2.5-fold in extent. Chromosome counts are reported for 72 of the species studied, of which 42 are new.     

DNA Replication in the Face of (In)surmountable Odds

We describe here a model for sequential recruitment of various enzymatic systems that maintain DNA replication fidelity in cells with damaged bases, especially those formed by ultraviolet (UV) irradiation. Systems of increasing complexity but decreasing fidelity are recruited to restore replication of damaged DNA. The first and most accurate response is nucleotide excision repair (NER) that is cell cycle-independent; next come various delaying cell cycle checkpoints that provide an extended time window for NER. These delay the onset of the S phase at the G1/S boundary, and inhibit the initiation of individual replicating units (replicons and clusters of replicons) within the S phase. When checkpoints fail to operate completely, DNA replication forks must negotiate damage and the loss of coding information on the parental DNA strands. Replication can be resumed using bypass polymerases, or alternative bypass mechanisms. Finally, if all else fails, replication forks may degrade to double strand breaks and recombinational processes then allow their reconstruction. A network of signaling kinases modulates the efficiency of many damage responsive proteins to tailor their activities and subcellular localizations by phosphorylation and dephosphorylation.     

Nuclear DNA content inCelosia (Amaranthaceae)

Nuclear DNA amount of five species ofCelosia ranging from 2x to 12x varies from 3.26 (2x) to 9.70pg (12x). The diploidC. trigyna has twice as much DNA/basic genome as other taxa, which is commensurate with its taxonomic position and genetic isolation. There is insignificant variation in DNA/basic genome among 4x, 8x, and 12x taxa. Therefore, DNA/nucleus shows a strong positive correlation with ploidy level. The different accessions of 4x taxa show constancy of DNA amounts. There is no correlation of seed weight with DNA amount.