Ichthyotoxic Alga Research Articles

Early rapid detection of ichthyotoxic algae based on fluorescent quaternion features via neural network

We developed a fast fluorescent ichthyotoxic algae identification (FFIAI) model based on a specially designed convolutional neural network to predict if ichthyotoxic algae exist in water bodies by simply inputting the initial 3D fluorescence spectrum data of an algal testing sample. Discrete excitation-emission fluorescence spectra were used to characterize the fingerprint of each ichthyotoxic algae species and applied to the fluorescent quaternion representation model to form the fluorescent quaternion feature. We trained the FFIAI model with the fluorescent quaternion features of the training database. In our experiment, the well-trained FFIAI model can achieve 100 % accuracy in identifying six species of ichthyotoxic algae, including Amphidinium carterae, Karenia mikimotoi, Phaeocystis globose, Chattonella antique, Heterosigma akashiwo, and Prymnesium parvum. The corresponding detection limits are as low as 1800, 120, 3600, 60, 2000, and 2300 cells/mL, which meets the basic detection requirements of harmful algae blooms (HABs). Moreover, this model also performed well in mixed testing samples, with the identification accuracy of ichthyotoxic algae reaching 90 % when the ichthyotoxic algae are dominant in the mixture. The model could also recognize the ichthyotoxic algae correctly even with a relatively low concentration, with an overall identification accuracy of 75 %, demonstrating its potential for precise monitoring of ichthyotoxic algae in field studies.

Identification of ichthyotoxic red tide algae based on three-dimensional fluorescence spectra and particle swarm optimization support vector machine

Acccurate identification whether red tide has ithyotoxicity is very significant for microalgae monitoring. In order to realize the rapid and non-destructive detection of ichthyotoxic red tide algae, a detection method combining three-dimensional (3D) fluorescence spectrum and particle swarm optimization support vector machine (PSO-SVM) was developed to monitor the ichthyotoxic red tide algae with cell concentrations from 104 cells/mL to 106 cells/mL. The contour maps contracted form three-dimensional fluorescence spectra of six common species of ichthyotoxic algae and eight common species of non-ichthyotoxic algae,which are analyzed to select the optimal emission and excitation wavelength span. The new feature data are acquired by using the emission spectrum data at 480 nm and 510 nm excitation wavelengths. The new feature data are used as the input of particle swarm optimization support vector machine to establish the optimal classification model of ichthyotoxic algae, which achieves an classification accuracy of 100% for the test set. The optimal classification model is successfully applied to identify the ichthyotoxicity of different algae including Heterosigma akashiwo, Chattonella marina, Phaeocystis globosa, Prorocentrum donghaiense, Karenia dunnii, Isoscelina galbana, Isosceles globosa and Skeletonema costatum.

Sublethal and antioxidant effects of six ichthyotoxic algae on early-life stages of the Japanese pearl oyster

Several species of harmful algae form blooms that are detrimental to aquatic organisms worldwide with severe economic loss to several industries. The cosmopolitan ichthyotoxic dinoflagellates and raphidophytes Karenia spp., Chattonella spp., Heterosigma spp., and Margalefidinium (Cochlodinium) polykrikoides are known to cause mass mortalities of fish and invertebrates, and the dinoflagellates Heterocapsa spp. are known to cause mass mortalities of shellfish, notably bivalve molluscs. The species K. mikimotoi, K. papilionacea, H. circularisquama, H. akashiwo, M. polykrikoides, and C. marina form recurrent harmful algal blooms (HAB) in coastal aquaculture areas of shellfish, coinciding with the reproduction seasons of natural and farmed bivalve molluscs. In the present study, their effects on eggs, fertilization, embryos, and three larval stages (D-shaped, umbo and pre-settling larvae) of a model bivalve species, the Japanese pearl oyster, Pinctada fucata martensii, are reported. The harmful algae had differential negative effects on each developmental stage, and had differential effects on larvae depending on their growth stage. Eggs were more affected by M. polykrikoides, K. mikimotoi and H. circularisquama than H. akashiwo and K. papilionacea. Fertilized eggs and developing embryos were more affected by M. polykrikoides and H. circularisquama than K. mikimotoi, K. papilionacea and H. akashiwo. Mortalities as well as abnormalities were not observed in any larval stage; however, motility of d-larvae and umbo larvae was more reduced by H. circularisquama and C. marina, than M. polykrikoides. In elder, 16 day-old larvae, all harmful algae induced a significant decrease in motility with the most severe effect observed during exposures to H. circularisquama, C. marina, H. akashiwo and M. polykrikoides. The superoxidase dismutase activity in larvae was not affected by exposure to any harmful alga; however, 6- and 16-day old larvae experienced a significant increase in GST activity following 48 h of exposures, with higher sensitivity of the elder larvae to C. marina, K. mikimotoi and M. polykrikoides. These results indicate that all tested harmful algae are differentially detrimental to the early-life development of the Japanese pearl oyster, with involvement of oxidative stress. Both M. polykrikoides and H. circularisquama were the most toxic followed by C. marina, K. mikimotoi, H. akashiwo and K. papilionacea. In addition, more developed larvae were most sensitive to these harmful algae in terms of motility-avoidance behavior and oxidative stress response, suggesting that ingestion of the harmful algae might enhance the toxicity of contact-dependent effects and dissolved extracellular compounds. The results also showed that superoxide anions were not associated with effects observed in larvae. Instead cellular detoxification was induced. The differential, stage-specific and growth-specific sublethal effects on bivalve development and recruitment also warrant further investigations of the oxidative stress and antioxidant enzyme activities in larvae of bivalves, to better address the toxicity mechanisms of ichthyotoxic HAB and their impacts on the reproduction, recruitment, and fitness of bivalve molluscs.Summary: The harmful algae Heterocapsa circularisquama, Chattonella marina, Hetrosigma akashiwo, Karenia mikimotoi, K. papilionacea, Margalefidinium (Cochlodinium) polykrikoides differentially affect early life stages of Japanese pearl oyster and activate detoxification enzymes in feeding larvae.

Characterizing seston in the Penobscot River Estuary

The Penobscot River Estuary is an important system for diadromous fish in the Northeast United States of American (USA), in part because it is home to the largest remnant population of Atlantic salmon, Salmo salar, in the country. Little is known about the chemical and biological characteristics of seston in the Penobscot River Estuary. This study used estuarine transects to characterize the seston during the spring when river discharge is high and diadromous fish migration peaks in the Penobscot River Estuary. To characterize the seston, samples were taken in spring 2015 for phytoplankton identification, total suspended matter (TSM), percent organic TSM, chlorophyll a, particle size (2 μm-180 μm), particulate carbon and nitrogen concentrations, and stable carbon and nitrogen isotopes. The estuarine profiles indicate that TSM behaved non-conservatively with a net gain in the estuary. As phytoplankton constituted only 1/1000 of the particles, the non-conservative behavior of TSM observed in the estuary was most likely not attributable to phytoplankton. Particulate carbon and nitrogen ratios and stable isotope signals indicate a strong terrestrial, allochthonous signal. The seston in the Penobscot River Estuary was dominated by non-detrital particles. During a short, two-week time period, Heterosigma akashiwo, a phytoplankton species toxic to finfish, also was detected in the estuary. A limited number of fish samples, taken after the 2015 Penobscot River Estuary bloom of H. akashiwo, indicated frequent pathological gill damage. The composition of seston, along with ichthyotoxic algae, suggest the need for further research into possible effects upon resident and migratory fish in the Penobscot River Estuary.

Sublethal concentrations of ichthyotoxic alga Prymnesium parvum affect rainbow trout susceptibility to viral haemorrhagic septicaemia virus.

Ichthyotoxic algal blooms are normally considered a threat to maricultured fish only when blooms reach lethal cell concentrations. The degree to which sublethal algal concentrations challenge the health of the fish during blooms is practically unknown. In this study, we analysed whether sublethal concentrations of the ichthyotoxic alga Prymnesium parvum affect the susceptibility of rainbow trout Oncorhynchus mykiss to viral haemorrhagic septicaemia virus (VHSV). During exposure to sublethal algal concentrations, the fish increased production of mucus on their gills. When fish were exposed to the algae for 12 h prior to the addition of virus, a marginal decrease in the susceptibility to VHSV was observed compared to fish exposed to VHSV without algae. If virus and algae were added simultaneously, inclusion of the algae increased mortality by 50% compared to fish exposed to virus only, depending on the experimental setup. We concluded that depending on the local exposure conditions, sublethal concentrations of P. parvum could affect susceptibility of fish to infectious agents such as VHSV.

Transcriptome analysis of the euryhaline alga, Prymnesium parvum (Prymnesiophyceae): effects of salinity on differential gene expression

:There is limited information on the underlying physiological responses and molecular mechanisms involved in the ability of the ichthyotoxic alga, Prymnesium parvum, to tolerate a broad range of salinities. We report an RNA sequencing analysis of the transcriptome of P. parvum, including a de novo assembly generating 47,289 transcripts. Of them, 35.4% were identifiable, and 30.4% and 16.8% were classified into gene ontology (GO) categories and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways, respectively. Cultures were grown in media at salinities of 5 and 30 practical salinity units (psu), and the differential expression of 2507 transcripts (5.3%) was detected. Specifically, 1507 (3.2%) and 1000 (2.1%) transcripts were up- and downregulated, respectively, at 30 vs 5 psu. Relevant candidate genes identified included those whose products are involved in osmolyte production, salinity stress, ion transport, and acetate metabolism. Thirty-two different polyketide synthase (PKS) transcripts were identified, none of which were differentially expressed under our experimental conditions. However, our analysis revealed evidence of posttranscriptional regulation of polyketide prymnesin production and the contributory effects of putative hemolysins and polyunsaturated fatty acids (PUFAs) toward ichthyotoxicity. Moreover, gene transcription associated with vesicular transport was increased at the lower salinity. This comprehensive transcriptome characterization sheds light on transcriptional responses that occur following alterations in salinity in P. parvum.

Expression and standardized measurement of hemolytic activity in Heterosigma akashiwo

Heterosigma akashiwo (Hada) is a marine Raphidophyte responsible for fish deaths in temperate coastal waters. Factors leading to Heterosigma species dominance or levels of toxicity remain unresolved. This study investigated the role of several factors on the hemolytic activity of the ichthyotoxic alga Heterosigma akashiwo. Hemolytic activity was measured using the erythrocyte lysis assay, which involves incubation of an algal sample with erythrocytes and subsequent photometric measurement of hemoglobin released from lysed erythrocytes. Preliminary experiments determined the optimal assay conditions (e.g. algal sample type, cell density, incubation conditions) to detect maximal hemolytic activity in H. akashiwo. Subsequent experiments determined the roles of light; isolate variation, and growth stage. The expression of hemolytic assay was light dependent, dependent on the stage of growth and but varied little among isolates. Dramatic variations in the measurement of hemolytic activity were evident depending on the handling and preparation of the sample.

Growth and grazing of microzooplankton in response to the harmful alga Heterosigma akashiwo in prey mixtures

Microzooplankton grazers may play a role in regulating blooms of the ichthyotoxic alga Heterosigma akashiwo. This study tested the effects of H. akashiwo, when part of a mixed-prey assemblage, on the growth and feeding of microzooplankton. Laboratory cultures of 3 ciliates, Favella sp., Metacylis sp., and Strombidinopsis acuminatum, were exposed to reciprocal concentra- tions of H. akashiwo and a non-toxic prey, at saturating prey concentrations. Heterosigma akashiwo was toxic to Favella sp. and Metacylis sp. when H. akashiwo was the sole prey species; however, this toxicity was eliminated in the mixed-prey treatments, likely because of avoidance of H. akashiwo and selective feeding on non-toxic prey. In contrast, the growth rate of S. acuminatum was unaffected by H. akashiwo. Both Favella sp. and S. acuminatum ingested H. akashiwo but selected against the alga when other prey was available. In addition, natural planktonic communities, collected from East Sound, Orcas Island, northern Puget Sound, in September and October 2007, were exposed to bloom- level concentrations of H. akashiwo. Abundances of the major microzooplankton types, primarily cil- iates and Gyrodinium/Gymnodinium dinoflagellates, were unaffected by H. akashiwo, although slight changes in grazer size structure did occur. Heterosigma akashiwo was harmful to the smallest grazers, mainly aloricate ciliates and small Gyrodinium/Gymnodinium dinoflagellates, and beneficial to larger Gyrodinium/Gymnodinium dinoflagellates that were able to ingest and grow on the alga. The alga was not consumed by the majority of grazers. Preferential feeding on alternate prey reduces toxic effects of H. akashiwo on microzooplankton. Avoidance of H. akashiwo by a major group of grazers would promote bloom formation by reducing H. akashiwo mortality and focusing community grazing pressure on potential competitor species.

Susceptibility of marine teleost to the ichthyotoxic alga, Chattonella marina, is related to the fish osmoregulatory ability

Steven J.L. Xu, Summer M. Shen, Chris K.C. Wong, Norman Y.S. Woo, Doris W.T. Au Biology & Chemistry Department, City University of Hong Kong, 83 Tat Chee Avenue, Kowloon, Hong Kong Department of Biology, Baptist University of Hong Kong, 224 Waterloo Road, Kowloon, Hong Kong Department of Biology, The Chinese University of Hong Kong, Shatin, N.T., Hong Kong SAR, PR China E-mail: steven.xu@student.cityu.edu.hk

Genetic variation in bloom-forming ichthyotoxic Pseudochattonella species (Dictyochophyceae, Heterokonta) using nuclear, mitochondrial and plastid DNA sequence data

This study reports on genetic variation within bloom-forming ichthyotoxic algae in the genus Pseudochattonella. One aim of this study was to search for further molecular evidence to separate the two species, Pseudochattonella farcimen and Pseudochattonella verruculosa using DNA sequences with different levels of variability and from three different cell compartments (the nucleus, mitochondria and plastids). Secondly, the genetic variability among Pseudochattonella strains within a bloom and between blooms in successive years in the Skagerrak was assessed. We also aimed to determine the identity of Pseudochattonella strains from two other geographical locations. Using molecular data (LSU and SSU rDNA, rbcL, psbA and cox1) our results demonstrate clear delineation of the two Pseudochattonella species. The barcoding marker cox1 was well suited to separating the two closely related dictyochophyte species. Pseudochattonella strains from Germany and New Zealand were almost identical in five DNA regions to the P. verruculosa reference strain, NIES 670, suggesting that blooms in these geographical areas consisted of P. verruculosa. Comparison of DNA sequences from P. farcimen-strains within a bloom and between blooms in the Skagerrak in 2001 and 2006 suggests the existence of a homogeneous and stable population over a 5-year period. Intraspecific and intraclonal variations in the ITS rDNA were revealed within both P. farcimen and P. verruculosa eliminating ITS rDNA as a marker to differentiate among individuals and/or populations of Pseudochattonella.

The ichthyotoxic alga Chattonella marina induces Na+, K+-ATPase, and CFTR proteins expression in fish gill chloride cells in vivo

Our previous studies demonstrated that the ichthyotoxic Chattonella marina stimulated proliferation of branchial chloride cell (CC) and induced osmotic distress akin to hyperactive elimination of ions in fish (Rhabdosargus sarba). To ascertain the in vivo effects of C. marina on key CC ion transporters, the localization and expression of Na(+), K(+)-ATPase (NKA) and cystic fibrosis transmembrane conductance regulator (CFTR) proteins in response to C. marina exposure were investigated, using a quantitative immunocytochemical approach. The polarized distributions of NKA (alpha subunit) and CFTR proteins in branchial CCs of R. sarba remained unchanged under C. marina exposure. However, significant inductions of these two ion-transporters were detected in CCs of fish after 6h exposure. By real-time PCR, no significant changes in gill NKA and CFTR mRNA expressions were detected, suggesting a post-transcriptional pathway is likely involved in regulating the ion transporters abundance. This study is the first to demonstrate the in vivo effects of harmful algal toxin on NKA and CFTR protein expressions in gill transepithelial cells. Taken together, an augmentation of branchial CCs together with hyper-stimulation of NKA and CFTR in CCs attribute to the rapid development of osmotic distress in C. marina susceptible fish.

A modified assay to determine hemolytic toxin variability among Karenia clones isolated from the Gulf of Mexico

Karenia brevis (formerly Gymnodinium breve) is a toxic marine dinoflagellate generally restricted to the Gulf of Mexico and is the main causative organism in fish kills, shellfish intoxications and respiratory distress in humans following bloom events. K. mikimotoi is a morphologically similar co-occurring species which is toxic in other parts of the world oceans, but has not been recognized as a major contributor in toxicity of blooms within the Gulf of Mexico. Recently there has been increasing evidence of the simultaneous production of a variety of bioactive compounds in addition to potent neurotoxins (brevetoxin) in Karenia brevis isolates. These compounds are potentially ichthyotoxic and have been shown to cause hemolysis in several bioassays [Eshbach, E., Scharsack, J., John, U., Medlin, L., 2001. Improved erythrocyte lysis assay in microtitre plates for the sensitive detection and efficient measurement of haemolytic compounds from ichthyotoxic algae. J. Appl. Toxicol. 21, 513–519; Kirkpatrick, B., Fleming, L.E., Squicciarini, D., Backer, L.C., Clark, R., Abraham, W., Benson, J., Cheng, Y.S., Johnson, D., Pierce, R., Zaias, J., Bossart, G.D., Baden, D.G., 2004. Literature review of Florida red tide: implications for human health effects. Harmful Algae 3, 99–115]. Presence of hemolytic compounds may therefore add to the overall toxicity levels of bloom events. Current monitoring methods include assays which are highly sensitive in brevetoxin detection and yet may not target other harmful compounds. By adapting protocols developed by Eshbach et al. [Eshbach, E., Scharsack, J., John, U., Medlin, L., 2001. Improved erythrocyte lysis assay in microtitre plates for the sensitive detection and efficient measurement of haemolytic compounds from ichthyotoxic algae. J. Appl. Toxicol. 21, 513–519], Red drum ( Sciaenops ocellatus) erythrocytes were used to create a modified bioassay to detect hemolytic activity of crude algal extracts. Red drum was selected because it is endemic to coastal areas throughout the Gulf of Mexico and is sensitive to Karenia blooms, and thus makes this species a valid ecological target. Preliminary data has shown this method is sensitive for use in assessing hemolysis induced by laboratory cultures down to levels of 1 × 10 3 cells mL −1. Results showed an unexpectedly high level of hemolytic activity among K. mikimotoi clones, with one Texas strain inducing significantly higher hemolysis compared to Florida K. brevis isolates. Using this approach, future research efforts will examine the difference in production of hemolytic compounds among various Karenia clones.

Improved erythrocyte lysis assay in microtitre plates for sensitive detection and efficient measurement of haemolytic compounds from ichthyotoxic algae.

Haemolytic substances produced by ichthyotoxic algae often are unknown in molecular structure or specific mechanism of toxicity. Detection and quantification of such substances are dependent on bioassays, using markers that are sensitive for haemolytic impairment and generation of a recordable response. The erythrocyte lysis assay (ELA) represents an advantageous bioassay in this respect, because the lytic response can be measured photometrically by the amount of released haemoglobin. The aim of the present study was to establish an improved assay based on the ELA principle, for sensitive determination of haemolytic substances of microalgae and for high sample throughput. For this purpose we adapted the ELA to a 96-well microtitre plate format, which significantly reduced the sample volumes and allowed rapid processing of samples. Further improvement was achieved by measuring absorption of lysed erythrocytes at 414 nm, which significantly increased the sensitivity of the ELA compared to the measurements at 540 nm that are usually applied in this type of assay. Using carp (Cyprinus carpio) erythrocytes it was possible to detect haemolysis induced by 4 microg ml(-1) of saponin and as little as two haemolytic Alexandrium tamarense cells. It is suggested that this improved ELA in microtitre plates be used as a low-cost monitoring tool for detection and analysis of potential harmful algae. Furthermore, this ELA can be useful as a sensitive screening system for substances of pharmacological interest, e.g. selectively acting cytolytic antibiotics.

Epizootic mortality in the pilchard Sardinops sagax neopilchardus in Australia and New Zealand in 1995. I. Pathology and epizootiology

DAO Diseases of Aquatic Organisms Contact the journal Facebook Twitter RSS Mailing List Subscribe to our mailing list via Mailchimp HomeLatest VolumeAbout the JournalEditorsSpecials DAO 28:1-15 (1997) - doi:10.3354/dao028001 Epizootic mortality in the pilchard Sardinops sagax neopilchardus in Australia and New Zealand in 1995. I. Pathology and epizootiology Whittington RJ, Jones JB, Hine PM, Hyatt AD A large-scale epizootic occurred in the Australasian pilchard Sardinops sagax neopilchardus between March and September 1995 over more than 5000 km of the Australian coastline and 500 km of the New Zealand coastline. Affected fish died within a few minutes of clinical signs of respiratory distress and death was associated with hypoxaemia and hypercapnea. Significant lesions were confined to the gills and comprised acute to subacute inflammation followed by bizarre epithelial hypertrophy and hyperplasia. The lesions were initially focal but progressed to become generalised over about 4 d. Pathological changes in affected fish from western Australia, eastern Australia and New Zealand were similar, suggesting a common aetiology. The lesions were unlike those associated with ichthyotoxic algae, siliceous algae, physicochemical factors, fungi, bacteria, dinoflagellates, amoebae, other protozoa and metazoa. A herpesvirus was consistently present in gills of affected fish and absent from unaffected pilchards and is proposed as the aetiological agent. The rate of spread of the mortality front (approximately 30 km d-1) in relation to the migration rate of pilchards and prevailing currents suggests that a vector was involved. The disease may have been newly introduced into Australian waters. Clupeoidei · Pilchard · Sardinops sagax neopilchardus · Gill diseases · Pathology · Epizootiology · Mortality · Herpesvirus Full text in pdf format NextExport citation RSS - Facebook - Tweet - linkedIn Cited by Published in DAO Vol. 28, No. 1. Publication date: January 16, 1997 Print ISSN:0177-5103; Online ISSN:1616-1580 Copyright © 1997 Inter-Research.

Blowflies (Calliphora vomitoria) as an in Vivo Bioassay for Diarrhetic and Hemolytic Algae Toxins

Using blowflies (Calliphora vomitoria) as test subjects, a bioassay method was developed for the detection of diarrhea-causing and hemolytic (i.e., ichthyotoxic) algae toxins in biological material. Flies were immobilized by refrigeration, and injected in series of 10 with the required doses of the toxins in question. Following injection, lethality of flies was monitored at 27°C over a 24-h period. Okadaic acid (OA), the most common diarrhea-causing toxin, gave a detectable lethality of flies after 2 h of monitoring. For this exposure time, a 50% lethal dose (LD50) of 666 μg OA/kg was detected, while an LD50 of 274 μg OA/kg was found after 4 h of surveillance. These figures are comparable to the lethal doses reported within a 24-h surveillance period for OA in mice, suggesting the fly bioassay as equivalent in sensitivity to the mouse bioassay for diarrhea-causing toxins, or even more sensitive by the use of a moderately longer time of surveillance than 4 h. Sensitivity of the fly bioassay appears limited by time, however, since high lethality of control flies was occasionally found after 16-24 h. Moderately toxic extracts of mussels exposed to the ichthyotoxic alga Prymnesium parvum gave detectable lethality in flies within 1 h of surveillance, suggesting an even faster toxic effect of hemolytic toxins in flies than that of diarrhea-causing toxins.

Effect of toxin ofprymnesium patelliferumon neurotransmitter transport mechanisms: Development of a sensitive test method

A crude extract of the ichthyotoxic phytoflagellate Prymnesium patelliferum strongly inhibited the uptake of neurotransmitters into isolated nerve endings (synaptosomes) and synaptic vesicles. These systems were about 100 and 10 times more sensitive toward the algal toxin, respectively, than a standard hemolysis assay often used for testing the toxicity of Prymnesium spp. and other ichthyotoxic algae. Prymnesium patelliferum grown in phsophorus-deficient (-P) medium was about five times more active than when grown in phosphorus sufficient (+P) medium. The inhibition ratio between the high-affinity synaptosomal uptake of L-glutamate and gamma-aminobyturic acid (GABA) was 1/2.7 for the -P culture and 1/1.9 for the +P culture. The inhibition ratios for the low-affinity vesicular uptake of L-glutamate, GABA, and dopamine (DA) were 1/5.8/0.3 and 1/1.7/0.2, respectively. The synaptosomal transport of L-glutamate is a rapid, simple, and sensitive test method for toxicity determination of Prymnesium spp. and will be a useful tool in the further isolation and purification of the toxic principles of this and other related algae. It is suggested that the toxin interferes with ion channels or acts as an ionopore.