INTRODUCTION

In March 1998, the U.S. Fish and Wildlife Service’s (Service), Region 3 and Region 5 Great Lakes Basin Ecosystem Team (Ecoteam) identified three broad, basin-wide issues on which to focus its efforts during fiscal years 1998-99. The basin-wide issues include lake sturgeon restoration and passage, double-crested cormorant, and reauthorization of the Great Lakes Fish and Wildlife Restoration Act of 1990 (Act). These issues were selected from among several relatively urgent concerns in the Great Lakes Basin that would benefit from the combined efforts of Ecoteam members. Addressing these issues meets several Resource Goals identified by the Ecoteam. Though reauthorization of the Act has been accomplished, actions on the other two basin-wide issues are continuous and dynamic. Lake sturgeon restoration and double-crested cormorant issues are in keeping with the Service's "Fisheries’ Vision for the Future" and the "Fisheries Action Plan" which focus on interjurisdictional, native, and depleted species and related ecosystem impairments.

As a result of the Ecoteam’s priority issues for the basin, a cross-regional and cross-program committee was established to identify and address priority action items. The Lake Sturgeon Committee (Committee) (Appendix A) was formally established in July 1998 with an overall role to provide a mechanism for coordination among the programs, share information, and seek consistency from a Service perspective. Collaboration of efforts and expertise from all programs make this an extremely beneficial and efficient process. In the future, the Committee intends to develop a U.S. Fish and Wildlife Service Great Lakes Lake Sturgeon Restoration Plan. This Plan will incorporate information collected from the following priority action items identified by the Committee:

• Develop a standardized scientific database of lake sturgeon observations,
• Standardize genetic data collection and analysis,
• Inventory historic and current tributaries inhabited by lake sturgeon and obstructions to the historic range (provided in GIS format),
• Evaluate the current Habitat Suitability Index Model (HSI) for its applicability to Great Lakes populations,
• Develop a Service Great Lakes Lake Sturgeon Web Site,
• Investigate contaminant issues related to lake sturgeon restoration and passage, and
• Determine the status of Great Lakes populations for consideration under Endangered Species Act listing.

STATUS ASSESSMENT

Below are descriptions of the activities and composition of genetics samples within each agency associated with Great Lakes Basin lake sturgeon work through May 1999. See Table 1 for a summary of key components. The information has been compiled from documents, as well as correspondences between geneticists and biologists.

Ashland Fishery Resources Office - Henry Quinlin

The Ashland Fishery Resources Office (FRO) is a member of the Lake Sturgeon Sub-Committee of the Lake Superior Committee. Lake sturgeon genetic samples are obtained from several cooperators/partners including the Marquette Biological Station, Wisconsin and Minnesota Departments of Natural Resources (DNR), and Michigan Technological University (MTU). In 1995 and 1997, the Ashland FRO obtained 31 muscle tissue samples (Porter et al. 1997) and 64 fin samples, respectively from the Bad River.

Dr. Paul Fuerst at Ohio State University (OSU) (Appendix B) has received all samples obtained by the Ashland FRO. Dr. Fuerst is conducting genetic analyses with Dr. Ted Cavender of the Ohio Museum of Biological Diversity (Appendix B) and several graduate students. Their objectives include determining the genetic structure of natural Great Lakes lake sturgeon populations, providing an interpretation of this information to assist state and federal agencies, and using museum specimens to compare current genetic diversity to historical levels of genetic diversity. The current methods include obtaining fin, muscle, or blood tissue to perform biochemical (allozymes) and molecular (RAPDs, microsatellite loci, mitochondrial DNA sequences, unique variable nuclear genes) analyses. Population differentiation is determined by looking at allele frequency changes (allozymes, microsatellite, and mtDNA), band sharing (VNTR and RAPD loci), and DNA sequence divergence (mtDNA and unique variable sequences). The full analysis involves examining at least 16 populations. The document The Genetic Status of Lake Sturgeon in Lake Erie and other Populations from the Laurentian Great Lakes (Porter et al. 1995) is currently available which describes findings from some of their research. Future objectives of the project are to compare differences between sex, drainages within a lake, sites within a drainage, breeders and non-breeders, and between successive years of breeders. No date of completion has been identified for future work.

In 1995 and 1996, MTU provided 45 muscle samples from the Sturgeon River to Dr. Fuerst (Porter et al. 1995). Minnesota DNR and Wisconsin DNR forwarded 30 muscle, blood, fin, eye, and liver samples from St. Louis Bay lake sturgeon and 30 muscle, blood, and fin samples from Chequamegon Bay to Dr. David Philipp and Dr. Bob Fields at the Illinois Natural History Survey (Appendix B) (Fields and Philipp Unpublished). Samples from the Lake Michigan Basin (see below) and Mississippi River Basin (Flambeau, Snake Rivers) accompanied the Lake Superior Basin samples to Dr. Philipp. Dr. Philipp and Dr. Fields used a PCR-RFLP analysis of mtDNA and protein electrophoresis in their methods (Fields and Philipp Unpublished). A publication of the results is in progress.

Green Bay Fishery Resources Office - Rob Elliott

The Green Bay FRO collected lake sturgeon genetic samples from the Lake Michigan Basin in 1999.
Wisconsin and Michigan DNR have obtained samples. In 1995 and 1997, Wisconsin DNR provided 57 muscle and fin samples from Wolf River lake sturgeon to Dr. Fuerst (Porter et al. 1995). In 1995, 110 samples were obtained from Lake Winnebago fish and given to Dr. Fuerst (Porter et al. 1995). Both aforementioned waterbodies enter the Fox River, tributary to Green Bay. Furthermore, Wisconsin DNR forwarded 30 muscle, blood, and fin samples each from Wolf River/Lake Winnebago lake sturgeon and Menominee River lake sturgeon to Dr. Philipp and Dr. Fields.

In 1994 and 1996, the Michigan DNR, Marquette Field Station collected 25 muscle and blood samples and 30 fin tissue samples, respectively from the Menominee River. These samples were forwarded to Dr. Fuerst (Porter et al. 1995).

LaCrosse Fishery Resources Office - Ann Runstrom

The LaCrosse FRO obtained six lake sturgeon fin tissue samples from the Mississippi River. Although outside of the Great Lakes Basin, there is fish movement between Lake Michigan and the Mississippi River via the Illinois (Chicago) Waterway. Currently, the LaCrosse FRO holds these samples for future analysis, with no geneticist identified to conduct the work. In addition, the LaCrosse FRO initiated a cooperative project with Dr. Scott Cooper of the University of Wisconsin at LaCrosse (Appendix B) to investigate species differentiation using genetic material from eggs. Ths information was critical due to the inability to identify species’ eggs in the wild based only on appearance. Primarily, shovelnose sturgeon and paddlefish from the Mississippi drainage were studied; however, some work was conducted with pallid and lake sturgeons. The lake sturgeon eggs from the Wolf River population were collected by the Wild Rose Fish Hatchery in Wisconsin. Dr. Cooper used a polymerase chain reaction (PCR) -based test to amplify a portion of the cytochrome-B gene from the mtDNA of a single egg. The presence of distinct HincII restriction sites (markers) in the PCR product allowed rapid determination of shovelnose sturgeon, pallid sturgeon, and paddlefish. Distinct restrictions sites did not show up in the lake sturgeon eggs, which separated this species from the other three. The procedure and reagents have been forwarded to the USFWS forensics lab in Ashland, Oregon to apply towards the illegal trade of caviar (see Law Enforcement below).

Alpena Fishery Resources Office - Tracy Hill

The Alpena FRO is the coordinator of the Central Great Lakes Bi-National Lake Sturgeon Group. Lake sturgeon genetic samples have been obtained from several cooperators/partners including Michigan and Ohio DNRs and Ontario Ministry of Natural Resource offices (OMNR). The Alpena FRO currently has 20 (or more) lake sturgeon fin tissue samples from the Detroit River and approximately 10-15 from Saginaw Bay, Lake Huron fish. All samples were collected in fall 1998. Samples will be forwarded to Dr. Paul Fuerst (OSU), and to Dr. Kim Scribner (MSU) in cooperation with Michigan DNR.

In 1993-1996, Michigan DNR provided approximately 40 muscle, fin, and blood samples from Lake St. Clair lake sturgeon to Dr. Fuerst as well as 30 fin tissues from Black Lake in 1997 (Porter et al. 1997). In 1998, they obtained 95 fin samples from the St. Clair River and 140 from Lake St. Clair. These were forwarded to Dr. Scribner who will utilize microsatellite and mtDNA analysis techniques. The mtDNA work will begin with direct DNA sequencing, possibly followed by PCR-based tests and restriction sites.

Ohio DNR provided 14 muscle, fin, and blood samples from Lake Erie to Dr. Fuerst in 1993-1995 (Porter et al. 1997).

In 1995, the OMNR, Lake Huron Management Unit provided 15 blood and muscle samples each from the North Channel and southern Lake Huron lake sturgeon, Canada (Porter et al. 1997). In 1996, the Lake Erie Management Unit provided 25 fin samples from western Lake Erie lake sturgeon to Dr. Fuerst (via Lower Great Lakes FRO). These 25 samples, in addition to eleven 1997 fin samples were also provided to Dr. Bernie May (Appendix B). See Lower Great Lakes FRO paragraphs below for information regarding Dr. May’s work.

Lower Great Lakes Fishery Resources Office - Chris Lowie

The Lower Great Lakes FRO has obtained 29 lake sturgeon fin tissues from several agencies. Tissue samples were collected from the following water bodies; eastern Lake Erie (6), upper Niagara River (2), lower Niagara River (11), and Lake Ontario (10). Samples have been collected by the Lower Great Lakes FRO since 1994, with the majority being obtained in 1996 and 1998. Samples from 1994 to 1996 were forwarded directly to Dr. Fuerst, and to Dr. Bernie May (University of California at Davis) via Harold Kincaid at the USGS- BRD laboratory in Wellsboro, PA (Appendix B). The 1997 and 1998 samples were only forwarded to Dr. May. In addition, the 1998 samples were sent to Dr. Tim King at the USGS-BRD, Leetown Science Center in West Virginia (LSC) (Appendix B).

The objectives of Dr. May and Dr. Kincaid’s work includes: (1) characterize stocks with genetic markers, (2) develop standardized DNA analysis for sturgeon broodstocks (see Discussion below), (3) measure genetic diversity in sturgeon species, (4) provide a centralized database for genetic information, and (5) assist fisheries management agencies develop genetic management programs. Methods involve using fin tissue to develop primers for microsatellite loci to characterize within- and between-population variation. Dr. May and his colleagues have published their lake sturgeon markers, which can be used by other geneticists (May et al. 1997). A study to identify the inheritance mechanisms of microsatellite loci in lake sturgeon has been performed by the investigators, with a manuscript being prepared for publication in 1999. Also, final results of the microsatellite screening and population differentiation are expected to be completed in 1999.

The Leetown Science Center has primarily been working with Atlantic sturgeon; however, more recently has performed lake sturgeon genetic analysis. The LSC has developed markers for Atlantic sturgeon and have tested them on our lake sturgeon samples. The Atlantic sturgeon markers are available at Genbank, a clearinghouse through the National Library of Medicine and a publication identifying the markers is currently in peer review. The Atlantic sturgeon markers and Dr. May’s lake sturgeon markers can be utilized by the LSC to evaluate future samples. Methods include using fin tissue and the microsatellite loci primers to determine genetic variation.

Between 1994-1996, the New York State Department of Environmental Conservation provided Dr. Fuerst with 15 blood samples from St. Lawrence River lake sturgeon below the Robert Moses Power Project and 7 blood samples from Grasse River lake sturgeon. Barbel samples collected in 1997 and 1998 have been forwarded to Dr. Rejean Fortin (Appendix B) in cooperation with lower St. Lawrence River studies (see sections below).

LOWER ST. LAWRENCE RIVER and LAKE CHAMPLAIN

LAW ENFORCEMENT - Ashland, Oregon - Steve Fain

The forensics laboratory has currently performed species identification on 17 sturgeon species using mtDNA from the eggs. Regarding lake sturgeon, most work has been performed on meat samples rather than caviar and they have no difficulty identifying them. In the future, the lab will use microsatellite techniques on the North American sturgeon species. This will help identify populations within a species.

DISCUSSION

This report provides an inventory of lake sturgeon genetic samples in the Great Lakes Basin as well as a brief overview of analysis techniques being used. Based on the information reported by participating agencies, it is apparent that enhanced interagency coordination is necessary to guide future research on this topic. Standardization of lake sturgeon genetics work is possible; however, the issues presented below must be addressed in the near future.

The best collection, fixing, and preserving method should be identified and standardized among agencies to obtain clean and functional samples. Several tissue types have been obtained for genetic analysis (Table 1). Fortunately, tissue type does not cause a conflict in comparing analyses. However, different collection, fixing, and preserving methods are being used, which requires extensive care in the field and laboratory to avoid contamination of the sample. A contaminated sample can result in an inaccurate analysis, leading to false comparisons. In 1999, a recommendation was made to all agencies to continue collecting and preserving tissue samples as instructed by their respective geneticist; however, to keep their samples until enhanced coordination could occur.

The results of mitochondrial DNA (mtDNA) and microsatellite analyses are not directly comparable. There are both advantages and disadvantages of using each technique for genetic stock identification; however, Ferguson and Danzmann (1998) suggest both genome analyses should be surveyed extensively to obtain the best representation of a population. The two results can be combined rather than compared.

If the same analysis method is being used between geneticists, discrepancies can still occur when different markers, which are developed by the individual geneticist, are used. In brief, mtDNA analysis is usually performed by direct base pair sequencing or by using restriction enzymes (markers), which recognizes and cuts the DNA at a particular base pair sequence (i.e. allele). Microsatellite analysis looks for a specific base pair sequence and quantifies the number of repeats of that sequence. Again, comparing separate analyses becomes difficult because the markers identify specific alleles that can differ depending on the base pair sequences being utilized, and it is uncertain if all geneticists are using the same base pair sequence (e.g. allelic identification). Often an automated sequencer is used for precision; however, two sequencers can also differ in allele identification. Phospho-imaging is another technique that can be a source of variation in allelic designations.

A coordinated effort to identify lake sturgeon genetic needs must occur among state, federal, and provincial resource managers and biologists in the Great Lakes Basin. These needs must be articulated to research geneticists, who in turn can provide practical information needed to manage lake sturgeon populations. In general, to guide restoration and enhancement efforts in the Great Lakes, resource managers need answers regarding population differentiation. Regardless of using mtDNA, microsatellite, or both, information from the available markers has not been fully utilized to assess population structure. Also, additional markers may be needed. A sturgeon genetic marker library is being developed, using microsatellite markers, to compile available sturgeon markers and determine their utility to assess population structure. Researchers contributing to this effort include Dr. Bob Sheehan at Southern Illinois University (shovelnose sturgeon), Dr. May (lake sturgeon), and Dr. King (Atlantic sturgeon). Potentially, this is a positive step toward standardizing analyses due to a large number of markers being developed. In addition, it will allow multiple geneticists to use the same identified alleles, and likely provide enough information for definitive population differentiation.

The Lower Great Lakes Fishery Resources Office is in the process of coordinating a genetics workshop, including resource biologists, managers, and geneticists, to discuss standardization of Great Lakes lake sturgeon genetics activities. The expected outcomes from this workshop include: (1) sharing current genetics capabilities/technology with all participants, (2) identifying information and research needs, (3) identifying the best collection and analysis methods, (4) establishing a network of communication between agencies and geneticists, and (5) identifying funding sources to conduct the necessary research.

ACKNOWLEDGEMENTS

The Great Lakes Basin Ecosystem Team, Lake Sturgeon Committee would like to thank all of the participating agencies and contact personnel who provided information on their lake sturgeon genetics activities. Also, a special thanks is given to the geneticists for their time and patience during numerous conversations to ensure accuracy in the information presented. Also, the author thanks all reviewers for their time critiquing previous drafts.

REFERENCES