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Genetic diversity of Manayunkia speciosa in the Klamath River basin By: Dan Horner Mentors: Sascha Hallett Jerri Bartholomew Background • Ceratomyxa shasta is a microscopic, spore-forming parasite • Causes intestinal necrosis and mortality of Klamath River juvenile salmonids • 60% of out migrating juvenile salmonids infected • Severe economical and ecological impacts Economic Effects of C. shasta • Low salmon numbers forced regulatory closures of 2006 salmon fishing • Loss of $150 million to the region's economy • Over $2 million spent on C. shasta research annually Salmonid Populations Salmonid species at risk: - Steelhead - Coastal cutthroat trout - Coho salmon* - Spring and fall-run chinook *Classified as an endangered species in 1997(ESA) and 2002 (CESA) C. shasta Lifecycle Salmonid Myxospore Actinospore M. speciosa Intraspecies differences could affect the suitability of this polychaete as the definitive host Hypothesis • Manayunkia speciosa in the Pacific Northwest are the same species Objective: • Determine if Klamath River polychaetes belong same species, and if so, determine if distinct strains of M. speciosa exist within the Klamath River Basin Primary Locations of Research Klamath Lake Iron Gate Dam Willamette River 1. Acted as a necessary outgroup for intraspecific phylogenetic comparisons Willamette 2. A known infectious zone for C. shasta 3. 300 miles NW of primary sample site, the Klamath. 4. Genetically isolated river basin for comparison to the Klamath Klamath Manayunkia genetic background • No genetic data exists for M. speciosa • However, Manayunkia spp. have been sequenced from Lake Baikal - M. sp.TP2002 - M. zenkewitschii - M. athalasia • All Manayunkia were sequenced using universal primers targeting the Cytochrome Oxidase c subunit 1 gene Lake Baikal Methods 1. Extract DNA from collected samples 2. Specific CO1 gene amplified through PCR 3. PCR product sequenced and analyzed for novel genetic characteristics Cytochrome Oxidase Subunit 1(CO1) • CO1 gene: - Subunit 1 is a 710 bp mitochondrial gene - Codes for 13 proteins - Could possibly be used as a universal barcode for all species • For Nematodes: - 0-6% variability: same species - 10-20% variability: different species Subunit 1 Species/Location varianced-val values Amplified # Interspecies d-val d-val bp sp2002 zenk with known athalasia consistent d-val Willamette species variability Known species M. athalasia variability 675 M. sp2002 16% 22% - 19% 19% 17% - 576 M. zenkewitschii 565 20% - - - 520 - - 19% - 505 - - 19% 2.1% - - 19% 3.2% Willamette Williamson I5 Site 530 Intraspecies variance also within acceptable level Current Genotypic Results Klamath Genotypes Willamette Genotypes Klamath and Willamette Conclusion • All the Manayunkia used in previous C. shasta research have been the species, Manayunkia speciosa. • Distinct genotypes appear to exist, but further research is necessary Ongoing Research Hudson River, NY Deschutes River, OR Fraser River, B.C. Cowlitz River, WA Lake Superior, WI Acknowledgements • • • • • • • • • • Stephen Atkinson Gerri Buckles Charlene Hurst Sarah Bjork Jill Pridgeon Sue-Jie Koo Kevin Ahern Jerri Bartholomew Sascha Hallett HHMI
