It is widely recognized that conservation of biological diversity is essential for preserving existing biological systems. Biological diversity has been traditionally measured at the interspecific community level, but the concept can also be applied at varying levels of intraspecific organization including population and life history diversity. Identifying patterns of hierarchical population structure and diversity are critical steps toward setting conservation priorities and understanding processes of evolution and speciation. Neutral genetic molecular markers such as allozymes, mitochondrial DNA (mtDNA), and microsatellite DNA are commonly used to detect genetic diversity, each with increasing temporal resolution. At the finest temporal scale, microsatellite markers are being used to identify population structure and help interpret historical demographic processes that shape current patterns of diversity.

Conservation of salmonid biological diversity in the Pacific Northwest has focused on recognizing fundamental units of intraspecific genetic diversity that reflect population structure in relation to reproductive isolation. Intraspecific diversity has been defined at broad levels with formal terms such as subspecies and evolutionary significant units (ESU), but increasingly sophisticated molecular techniques can detect population structure at an even finer scale. Understanding fine-scale population structure and evolutionary relationships among populations is fundamental for preserving current and future ecological and evolutionary processes.

In the Pacific Northwest, small populations of coastal cutthroat trout (Onchorhynchus clarki clarki) are commonly the only salmonid species present in headwater streams above barriers to anadromy. Headwater streams are highly dynamic in space and time and natural environmental stocasticities such as floods, drought, channel desiccation, landslides and debris flows are common system processes. Coastal cutthroat trout are adapted to natural stochastic processes but remain sensitive to habitat alteration and are considered indicators of ecosystem integrity. Furthermore, populations isolated above barriers to anadromy are not influenced by marine fluctuations, and are better direct indicators of the effects of landscape alteration.

In order to persist in isolated headwater environments, coastal cutthroat trout populations must maintain genetic heterogeneity in spite of demographic and environmental stocasticities. Demographic factors such as founder effects, bottlenecks, and genetic drift, reduce heterogeneity, and affect small populations with little or infrequent gene flow disproportionately. Genetic population structure is shaped by historical biogeographic events, spatial environmental heterogeneity, life history differences and/or differential levels of human-mediated habitat manipulation. Characterizing genetic structure within populations will lend insight into the affects of historical and current processes on isolated coastal cutthroat populations. In this study, environmental variables that are related to genetic heterogeneity will be identified. Results of this study will give fisheries and forest managers insight into the genetic population structure and the relative effects of landscape variables on genetic diversity and population viability of coastal cutthroat trout. Specific objectives include:

1. Use microsatellite molecular markers to assay the degree of spatial genetic differentiation as a function of geographic location.
2. Test for concordance in spatial genetic structure among different ecological requirements, i.e. connectivity, basin size above barrier, etc.
3. Investigate identifiable population genetic subunits that can be tied to ecological and environmental variables and spatial heterogeneity within habitats with varying human-mitigated histories.

Methods

As part of an earlier statewide geographic information system (GIS) survey, 280 third order basins containing barrier-isolated populations of coastal cutthroat trout were identified and 40 populations were randomly selected for extensive field studies (see Gresswell and Bateman, CFER Annual Report 2001). From 1999 through 2002, genetic samples were collected from the 40 populations and preserved for future genetic analysis. Fish were sampled using single-pass electrofishing techniques starting in the lowest portions of the basin and proceeding upstream. Tissue samples were collected from up to 100 cutthroat trout per population. Each fish was captured with a backpack electro-fishing unit, anesthetized to reduce handling stress, weighed and measured, and a small portion of the caudal fin was removed and stored in a desiccant or a buffer solution.
For this genetic analysis, 25 coastal cutthroat trout populations are currently being characterized using microsatellite molecular marker techniques. Tissue collections from each basin were randomly subsampled to 95 individual fish to allow for adequate statistical power. For collections with less than 95 samples, the entire collection was used. DNA was amplified using polymerase chain reaction (PCR) techniques, and genotypes and allele frequencies were determined using electrophoresis methodologies. Genetic indexes such as Fst will be used to determine how genetic diversity is organized among and within the fish populations. Landscape variables will be derived from GIS databases and field data and will be compared to genetic diversity using multivariate techniques, and phylogenetic relationships will be examined by comparing genetic distance matrices.

Research Results and Management Implications

Between 1998 and 2002, 3554 coastal cutthroat trout tissue samples were collected from 40 basins. Of these, 25 basins and 2255 individual fish were randomly subsampled for analysis. Currently, DNA has been extracted from over 1300 individual fish and multiple microsatellite loci are being screened and optimized for use in this study.

Study Timeline

Tissue collection was completed in July 2002. DNA extraction, amplification and characterization will continue through spring 2003. A final report and manuscript for peer review will be completed by September 2003.