Third Report of the Technical Advisory Committee
on Harmful Algal Outbreaks in Maryland

March 17, 1998

INTRODUCTION
This is the third report of the Technical Advisory Committee (TAC) established in July, 1997, by Natural Resources Secretary John R. Griffin to advise the state agencies on their efforts to monitor the occurrence and evaluate the causes of fish lesions and fish kills initially observed in the Pocomoke River. The members of the TAC are listed in Appendix 1. An Interim Report was submitted on August 14, 1997, and a Second Report on September 12, 1997. These reports were attributed to the "Technical Advisory Committee on Pocomoke River Fish Health". By September, 1997, evidence had accumulated to the point that the Committee concluded that the toxic dinoflagellate Pfiesteria piscicida or related organisms were the most probable cause of the fish kills and lesions and these maladies were also observed in two other tidal tributaries of Maryland's Eastern Shore. Additional evidence, reviewed below, provides further support to this conclusion. Reflecting these developments, the Committee herein is referred to as the "Technical Advisory Committee on Harmful Algal Outbreaks in Maryland", similar to the terminology used in Secretary Griffin's original letter of appointment.
In both the Interim and Second Reports the TAC structured its assessment around seven basic questions related to the incidence, trends, and location of fish kills and lesions; their probable causes, including whether Pfiesteria was a factor; and the environmental conditions and human activities which may underlie these problems. In general, the TAC took a very cautious position, concluding that: (1) Pfiesteria or Pfiesteria-like organisms were the most probable, but not certain, cause of observed fish kills and the lesions on the fish involved in those kills; (2) multiple other factors could be involved in causing the high incidence of lesions observed in 1997; and (3) the introduction of nutrients was the most likely suspect among the human activities which could have stimulated toxic Pfiesteria events, but this could not be concluded with great certainty. The TAC counseled that "our minds should remain open to other explanations of environmental and biological causes."
A "Technical Workshop on Pfiesteria" was convened on February 18-19, 1998, in Linthicum, Maryland, by the interagency Maryland Pfiesteria Study Team for the purpose of exchanging information among interested parties and presenting its plans for the coming warm season to the Technical Advisory Committee. During this workshop the TAC revisited the interpretation of the 1997 results for Maryland, received updates from leading Pfiesteria researchers, and evaluated the 1998 Work Plans to monitor and gain a better understanding of the causes of toxic Pfiesteria outbreaks in Maryland. After the day and a half workshop, the TAC met to develop its conclusions and offer recommendations that would strengthen the 1998 Work Plans.

ASSESSMENT UPDATE

Since the TAC's Second Report in September, 1997, public concern about the potential risks to human health of Pfiesteria outbreaks-bolstered by clear evidence that the cognitive abilities of some human subjects exposed to the affected waters last summer had been impaired -prompted action to develop policies and programs to reduce these risks. Governor Glendening appointed the Blue Ribbon Citizens Pfiesteria Action Commission (Hughes, 1997). At the Commission's request, a panel of scientific experts on nutrient effects and dinoflagellate biology prepared a statement expressing the current understanding on whether outbreaks of Pfiesteria may be reduced by controlling pollution sources. The resulting "Cambridge Consensus" (University of Maryland Center for Environmental Science, 1997) produced a number of findings ending with the following statement: "In the long term, decreases in nutrient loading will reduce eutrophication, thereby improving water quality, and in this context will likely lower the risk of toxic outbreaks of Pfiesteria-like dinoflagellates and harmful algal blooms." In essence, this group of experts, after reviewing the literature and field data collected in 1997, was less equivocal than the TAC's Second Report concerning the relationship of nutrient enrichment and Pfiesteria. A similar conclusion was reached by an expert peer-review panel convened by the State of North Carolina (Water Resources Research Institute, 1998).
Based on the presentations and discussions at the February Workshop the uncertainties concerning two other issues have also narrowed.
First, there had been limited information and some confusion on the identity of the dinoflagellates responsible for the toxic conditions. Pfiesteria piscicida had been confirmed by taxonomic analysis using scanning electron microscopy only from the Chicamacomico River and not the other fish kill sites. Subsequently, its presence has been confirmed at all kill sites (two sites in the Pocomoke River and one site in Kings Creek). Burkholder and colleagues (1998) presented evidence to suggests that the failure to confirm P. piscicida in other samples, including those from which toxin-producing forms were cultured, was due to culture methods. They also summarized results that demonstrated indications of the presence of P. piscicida in samples from each of the Pocomoke River fish kill locations. Finally, they presented methodologies for distinguishing Pfiesteria and another Pfiesteria-like dinoflagellate from other species with which they might be confused.
Secondly, there have remained questions about whether Pfiesteria is responsible for the lesions or whether they were caused by other factors, particularly infective diseases. Because the lesions on freshly killed fish were extensively invaded by bacteria and fungi, it has been suggested that these lesions were originated by other agents before the exposure of the fish to Pfiesteria-toxins. While this may be the case, it is well known that under experimental conditions Pfiesteria-toxins can produce similar lesions and it is quite possible that these lesions were initiated by earlier, sublethal exposure to Pfiesteria toxins.

THE 1998 WORK PLANS

The 1998 Work Plans were broken into four distinct areas: (1) Pfiesteria-Related Fish Health Investigations, (2) Fish Health, Habitat Quality and Pfiesteria Surveillance, (3) Pocomoke Event Sampling for Toxic Substances, and (4) Watershed Pollution Source Assessment. TAC comments and recommendations on each follow below:

1. Pfiesteria-Related Fish Health Investigations
It was generally felt that the fish health investigations described in this section are not necessarily directed to understanding the onset of lesions caused by Pfiesteria, but are developed to get an indication of fish health throughout the Chesapeake region. While these studies may be necessary for a multitude of reasons, the TAC only focused on the Pfiesteria-related investigations.
Experimental studies. The TAC had problems evaluating the experimental studies because no hypotheses were explicitly stated to justify the experimental design and extensive laboratory analyzes. It seemed that the experiments were still being conceived as a determination of whether Pfiesteria causes lesions, when this is well known from laboratory experiments and field observations in North Carolina. Rather, these experimental studies could prove very useful in determining whether an elevated incidence of lesions provides, as assumed in the current operational protocol, an effective warning for toxic Pfiesteria outbreaks. Experiments should be designed to test specific hypotheses regarding the degree to which lesions constitute an adequate indicator of toxic Pfiesteria conditions. While the TAC recognizes the difficulty in conducting in situ menhaden experiments, the lack of attention to the species, which appears to stimulate toxic Pfiesteria outbreaks and be most affected by them, calls into question the purpose of the experiments altogether. Experiments should concentrate ideally on menhaden, but alternately on white perch as a surrogate, and should be conducted in areas that have had toxic Pfiesteria outbreaks or areas where fish with lesions are found.
Laboratory analyzes of fish health parameters. Some members of the TAC wanted more details regarding the laboratory analyzes, particularly the stress response evaluations. Caging and handling effects on stress indicators should be thoroughly tested before any broader application of these methods.

2. Fish Health, Habitat Quality and Pfiesteria Surveillance
The Work Plan for this broad topic includes a diverse array of field observations on water quality, fish, and Pfiesteria that are based on a four-tier sampling strategy accommodating rapid response (Level I) and comprehensive assessments within previously closed tributaries (Level II), other tributaries potentially at risk (Level III), and in other areas covered by on-going monitoring programs (Level IV). This framework is a reasonable approach that provides focus and sets priorities while at the same time allows flexibility. However, coordination between the fish health and habitat quality components should be strengthened.
Fish community monitoring. The TAC recommends that fish community monitoring at least at some Level IV stations include sampling gear that can specifically sample menhaden from May through October. The TAC recognizes that much of the Level IV sampling is occurring for other specific reasons but that incorporating menhaden would add value to the ongoing efforts. The TAC also encourages the DNR to coordinate menhaden and other fish sampling efforts with other states in the mid-Atlantic region.
Habitat quality monitoring. The TAC is concerned that all Level III river systems are along the lower Eastern Shore and do not include tributaries in the upper Eastern or Western Shores of the Chesapeake Bay. Pfiesteria is known to occur in some of these areas (e.g. Choptank and Patuxent rivers). The TAC understands that additional Level III river systems may be chosen depending on funding availability and recommends that any additional Level III systems be chosen considering the following conditions: areas with relatively fine sediments, shallow bathymetry, weak currents, and high nutrient levels and where fish tend to congregate.
The TAC highly recommends that some species of dissolved organic nitrogen (DON), particularly urea, be directly measured in light of preliminary research that suggests a relationship with harmful algal species. Total DON cannot be directly measured, without measuring all of its constituents, therefore the difference method of calculating DON (TN - DIN = DON) should be continued at all Level I - III stations.
The TAC recommends that algal community sampling and analysis be done for some stations throughout the sampling season. Relatively rapid techniques such as in situ flourometry (for chlorophyl a concentrations), high-powered liquid chromotography (to identify and quantify pigment groups), and epiflourescent microscopy (for general identification) should be applied on a regular basis for all Level I - III stations. Direct counts could be completed on a subset of samples. Alan Lewitus, a TAC member, is willing to provide further details regarding specific methodology.
The TAC recommends that samples to determine the presence and abundance of Pfiesteria-like dinoflagellates should be taken and analyzed in all Level I -III river systems. This would include these Level III systems where fish kills potentially related to Pfiesteria have not yet been observed. Furthermore, the TAC feels that Maryland and its neighboring states should develop identification capabilities within the Chesapeake Bay region.
In addition to the sampling described in the Work Plan, the TAC recommends that some latitudinal sampling (i.e. shore to shore) take place around some stations and sediment DOM (pore water) measurements be taken at selected stations, if funds and staff time are available.

3. Pocomoke Event Sampling for Toxic Substances
The TAC concurs with the Work Plan that the spring sampling needs to be completed. However, further sampling may only warranted after the data are analyzed from an ecorisk perspective, including a literature search of the effects of the toxicants on a variety of species (from algae to fish) with a consideration of sub-lethal effects. If this analysis does not find any reason for concern then toxic sampling should only be conducted to address specific hypothesis-driven questions. The TAC is concerned about the continued support of the many partners enlisted to complete the toxic analysis and hopes that the partnerships will stay intact to complete the spring analysis.


4. Watershed Pollution Source Assessment
While the TAC does not have the expertise to review this section adequately, it appeared to some members as fragmentary and not well coupled with the event sampling and comprehensive assessments of tidal waters. Therefore, the Chair has offered to convene a group of state, federal and academic scientific experts to consider methods to conduct watershed assessments in a much larger context, including better integration of existing efforts and refinement of models that relate the results of small watershed modeling to estuarine loadings. The Departments of Environment, Natural Resources, and Agriculture and the U.S. Geological Survey, as well as several academic experts, have agreed to participate. This forum should take place during March or April, 1998. If scheduling conflicts occur, the TAC will seek advice regarding this section from a smaller group of experts who will solely consider this section of the Work Plan.

5. Coordination
In its review of the 1997 Work Plan, the TAC, while recognizing the necessary haste in its preparation, criticized the lack of coordination among the components of the monitoring and assessment plan. Specifically, we noted the need for a more clearly stated strategy with specific questions or hypotheses, priorities and sequences. We also recommended the requirement for a larger regional framework beyond the Pocomoke River and a strong leadership and management structure. The TAC is pleased to observe considerable improvements in the 1998 Work Plans, including the development of the four-level assessment strategy that provides both a framework for coordination of sampling but also for broader regional assessment. We are also pleased to learn that the 1998 Work Plans were produced through the active efforts of the interagency Maryland Pfiesteria Study Team. However, we also note that in complex and challenging efforts such as this there is always room for significant improvement in coordinated design, execution, and interpretation.

OTHER ISSUES
As the objectives and focus of Maryland's monitoring and assessment have evolved based on information collected and questions answered, the TAC finds itself with inadequate expertise in several important technical fields. The TAC was formed principally of individuals with expertise in fish health and algal ecology. As assessments have focused increasingly on nonpoint source inputs of nutrients and the effects of those nutrients in the estuarine ecosystem, new members should be added to the TAC to bolster its expertise in those areas. In addition, TAC expertise is needed in fish population ecology to assist in assessment of the consequences of the effects of diminished fish health and Pfiesteria-related mortalities on the resource stocks.
A number of large research proposals have been submitted to federal sponsors by Maryland investigators related to Pfiesteria. These include multi-investigator proposals to the interagency Ecology and Oceanography of Harmful Algal Bloom (ECOHAB) program to identify toxins and their effects and understand the connections between nutrient enrichment and Pfiesteria-like dinoflagellates and a proposal to the National Institute for Environmental Health for a major center to study human health-related effects. Assuming at least partial success of these research proposals, Maryland should be presented with unusual opportunities afforded by the coincidence of extensive monitoring and research activities. Effective coordination and integration of these efforts will allow experimental testing of hypotheses formulated from the observations and, conversely, will provide an extensive background context seldom afforded for research. Such effective coordination is not only an opportunity but an obligation. The TAC urges the state agency managers and university researchers and administrators to work closely toward this end.

REFERENCES

Brukholder, J.M., H.B. Glasgow, and E.K. Hannon. 1998. The Toxic Pfiesteria Complex vs. Mistaken Identities in the Albemarle-Pamlico and the Chesapeake. Summary Position Paper, February 1998.

Jordan, S.J., and E.B. May. 1998. Histological and Microbiological Findings Studied on Fish Taken from the Pocomoke River and Adjacent Tributaries. Maryland Department of Natural Resources, Oxford, Maryland.

Hedrick, J.D., and F.J. Margraf. 1998. Distribution, Progression, and Species Specific Incidence of Fish Skin Abnormalities in the Pocomoke River System. Maryland Cooperative Fish and Wildlife Research Unit, University of Maryland Eastern Shore, Princess Anne, Maryland.

Hughes, H.R. (Chair). 1997. Blue Ribbon Citizens Pfiesteria Action Commission, Final Report. Office of the Governor, Annapolis, Maryland.

Maryland Department of the Environment. 1998a. Pocomoke Watershed Pollution Assessment: Background and Summary of Results to Date. Baltimore, Maryland.

Maryland Department of the Environment. 1998b. Summary of Results of Monitoring for Chemical Contamination in the Lower Pocomoke River. Baltimore, Maryland.

Maryland Department of Natural Resources. 1998a. Histological Investigations--Pfiesteria--1996/1997. Oxford, Maryland.

Maryland Department of Natural Resources. 1998b. Water Quality and Pfiesteria Studies of 1997. Annapolis, Maryland.

Maryland Departments of Natural Resources, Environment, and Agriculture. 1998. 1998 Work Plans.

Technical Advisory Committee. 1997a. Interim Report of the Technical Advisory Committee on Pocomoke River Fish Health. Maryland Department of Natural Resources, Annapolis.

Technical Advisory Committee. 1997b. Second Report of the Technical Advisory Committee on Pocomoke River Fish Health. Maryland Department of Natural Resources, Annapolis.

Water Resources Research Institute of The University of North Carolina. 1998. The Raleigh Report 1998: Pfiesteria Research Needs and Management Actions. Report No. SRS-19. Raleigh, NC.

University of Maryland Center for Environmental Science. 1997. The Cambridge Consensus: Forum on Land-Based Pollution and Toxic Dinoflagellates in Chesapeake Bay. Cambridge, Maryland.

Appendix 1

Technical Advisory Committee on Harmful Algal Outbreaks in Maryland

Dr. Donald F. Boesch*, Chair University of Maryland Center for Environmental Science

Dr. Robert Anderson* (immunology) Chesapeake Biological Laboratory University of Maryland Center for Environmental Science

Dr. JoAnn Burkholder* (algal ecology) North Carolina State University

Dr. Eugene Burreson (diseases of marine organisms)

Dr. Larry Haas, alternate* (phytoplankton ecology, water quality) Virginia Institute of Marine Science

Dr. Mary Haasch* (fish toxicology) Chesapeake Biological Laboratory University of Maryland Center for Environmental Science

Dr. Sherwood Hall* (monitoring harmful algal blooms) U.S. Food and Drug Administration

Dr. Reginal Harrell (fish culture and physiology) Horn Point Laboratory University of Maryland Center for Environmental Science

Dr. Michael Hirshfield* (environmental conservation) Chesapeake Bay Foundation

Dr. Andrew Kane* (aquatic pathology and toxicology) Aquatic Pathobiology Center University of Maryland School of Medicine

Mr. Richard Lacouture* (phytoplankton monitoring) Environmental Research Center Academy of Natural Sciences

Dr. Alan Lewitus* (algal culture and ecology) Belle Baruch Institute University of South Carolina

Dr. Harold Marshall* (phytoplankton monitoring) Old Dominion University

Dr. Kevin Sellner* (algal bloom ecology) Coastal Ocean Program National Oceanic and Atmospheric Administration

Dr. Karen Steidinger (ecology and management of harmful algal blooms) Marine Research Institute Florida Department of Environmental Protection

Dr. Diane Stoecker* (heterotrophic microplankton) Horn Point Laboratory University of Maryland Center for Environmental Science

Dr. Daniel Terlizzi (aquaculture) Maryland Cooperative Extension Service University of Maryland, College Park

Dr. Patricia Tester (dynamics of algal blooms) Beaufort Laboratory National Oceanic and Atmospheric Administration

*Attended the February 18-19, 1998 workshop.

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