Grantee Research Project Results
2004 Progress Report: Investigation of Toxic Raphidophyte Population Dynamics Using Molecular and Physiological Tools
EPA Grant Number: R831041Title: Investigation of Toxic Raphidophyte Population Dynamics Using Molecular and Physiological Tools
Investigators: Hutchins, David A. , Cary, S. Craig , Coyne, Kathryn J. , Doblin, Martina
Institution: University of Delaware , Old Dominion University
EPA Project Officer: Packard, Benjamin H
Project Period: September 1, 2003 through August 31, 2006
Project Period Covered by this Report: September 1, 2003 through August 31, 2004
Project Amount: $567,331
RFA: Ecology and Oceanography of Harmful Algal Blooms (2002) RFA Text | Recipients Lists
Research Category: Water Quality , Aquatic Ecosystems , Water
Objective:
The objectives of this research project are to: (1) gain a better understanding of the effects of environmental perturbations and grazing pressure on raphidophyte community dynamics; (2) identify environmental factors that stimulate the growth of raphidophytes relative to other algal taxa; and (3) investigate the potential of raphidophyte cyst distributions as an indicator of seasonal bloom “hot spots.”
Progress Summary:
Year 1 of the project has seen progress on all of the major objectives. Quantitative polymerase chain reaction (qPCR) probes have been developed and tested on culture and field samples of Delaware Inland Bays’ (DIB) Chattonella subsalsa, Heterosigma akashiwo, and Fibrocapsa japonica (by Coyne and Cary). A new probe currently is being tested for C. verruculosa. We discovered that a supposedly “universal” eukaryotic primer, EukA, does not amplify raphidophytes. We have been able to develop another primer that does amplify raphidophytes, and this fortuitous discovery now allows us to amplify the rest of the algal community and the raphidophytes separately, offering a unique set of tools to compare the harmful algal bloom (HAB) group with sympatric species. Using Genescan, we also demonstrated the existence of several distinct strains of H. akashiwo in the DIB.
Three sets of laboratory vertical migration column experiments (by Handy and Hutchins) have been conducted, as well as three field mesocosm experiments. In the laboratory, we used columns designed to mimic the water column height of the DIB (~ 1 m). We followed the migration patterns of C. subsalsa and H. akashiwo using our qPCR methods and microscopy . We discovered that these species vertically migrate from the surface to the sediments on a diel cycle. These species apparently spend the night hours on the sediment’s surface and are near the water surface during midday. The DIB are stratified at least periodically in the summer. Phytoplankton that can migrate vertically throughout the day would have a competitive advantage for available nutrients, regardless of how the nutrients are distributed throughout the water column. In addition, understanding raphidophyte movement is essential to sampling these harmful species correctly.
Two sets of grazing experiments with collected mesograzers (copepods) were carried out using natural bloom samples (by Doblin). Preliminary evidence suggests that copepods are not exerting a major grazing pressure on raphidophytes, although more studies will be needed. Copepod abundance in the DIB is highly variable, affected by time of day and salinity (influenced by tide), as well as other factors. Consequently, grazing pressure by copepods is likely to be highly variable. Ten micrograzing dilution experiments (by Demir and Hutchins) suggest that protozoans at times may consume Heterosigma at significant rates, although these also are highly variable.
To study the raphidophyte cyst populations within the DIB, molecular techniques that characterize species presence and abundance were employed (Portune and Coyne). Multiple benthic sites within the DIB were examined during the past year. The presence of particular raphidophyte species and changes in their abundances were measured to determine whether these sediments provided possible seedbeds for future blooms. The changes in abundance also were correlated with the overlying water column during environmental mesocosm experiments to determine if excystment of cells out of the sediments had any effect on the initiation of blooms. Raphidophyte abundances within the sediments were determined using extracted DNA in quantitative real-time PCR (QRT-PCR). Laboratory experiments confirmed that QRT-PCR was very effective in accurately measuring cell abundances, especially at very low concentrations. Reverse transcriptase PCR also was employed to determine if DNA was actively being transcribed into RNA to determine cell viability and eliminate the possibility of residual DNA from dead cells in the sediments. Initial results from the Bloom Catcher 3 experiment (in which raphidophyte blooms were contained in an environmental mesocosm and its progression was monitored over the time of the bloom) confirmed that initial cell abundances were significantly greater than the concentrations after the bloom was over, possibly indicating a large-scale excystment event.
We completed the laboratory experiments examining the effects of nitrate, ammonium, phosphate, urea, and glutamic acid on growth rates of both C. subsalsa and H. akashiwo isolated from the DIB (by Zhang and Hutchins). We also examined the salinity and temperature tolerances of both species. Our completed salinity and temperature experiments suggest that Chattonella has much narrower salinity and temperature tolerances than Heterosigma, especially at lower ranges. Chattonella growth virtually is halted below 16°C and 15 ppt salinity, whereas Heterosigma continues rapid growth down to at least 5°C and 5 ppt. This suggests that Heterosigma blooms may occur earlier and continue later in the growing season and can occur considerably farther up in low salinity tributaries than Chattonella blooms. This also is consistent with the existing data set on bloom occurrences that has been collected by the Volunteer Phytoplankton Monitoring Group over the past several years.
We expect to be able to make similar comparisons between the two species’ nutrient requirements when the current data analysis is finished. We have already noted some interesting differences. For example, although Chattonella appears unable to use urea as a nitrogen source, Heterosigma grows very well on this organic nitrogen source. The same set of analyses and predictive information for Heterosigma will be available as soon as the data workup is completed. This information should be valuable in predicting the times and places where HABs of each species may occur.
Future Activities:
We will continue as planned over the next 12 months. The work is considerably ahead of the original schedule. Three papers are in the advanced stages of preparation and should be submitted for publication in the near future.
Journal Articles:
No journal articles submitted with this report: View all 32 publications for this projectSupplemental Keywords:
marine science, estuary, ecology, monitoring, Mid-Atlantic, ecosystem protection/environmental exposure and risk, water, ecological risk assessment, ecology and ecosystems, environmental chemistry, environmental monitoring, monitoring/modeling, oceanography, algal blooms, Chattonella, Delaware, DE, Delaware Inland Bays, algal bloom detection, algal growth, algal pigments, aquatic ecosystem, aquatic toxins, bloom dynamics, marine biogeochemistry, marine ecosystem, nutrient kinetics,, RFA, Scientific Discipline, Water, Ecosystem Protection/Environmental Exposure & Risk, Environmental Chemistry, Oceanography, Monitoring/Modeling, algal blooms, Environmental Monitoring, Ecological Risk Assessment, Ecology and Ecosystems, marine ecosystem, aquatic ecosystem, bloom dynamics, pigment analysis, nutrient kinetics, algal growth, aquatic toxins, marine biogeochemistry, Raphidophyte blooms, algal pigments, Delaware, ChattonellaProgress and Final Reports:
Original AbstractThe perspectives, information and conclusions conveyed in research project abstracts, progress reports, final reports, journal abstracts and journal publications convey the viewpoints of the principal investigator and may not represent the views and policies of ORD and EPA. Conclusions drawn by the principal investigators have not been reviewed by the Agency.