TEMPORAL GENETIC ANALYSIS OF STEELHEAD (ONCORHYNCHUS MYKISS) REVEALS HATCHERY-INDUCED DRIFT IN CAPTIVITY by Melissa R. Reneski A Thesis Presented to The Faculty of Humboldt State University In Partial Fulfillment Of the Requirements for the Degree Masters of Science In Natural Resources: Fisheries June, 2011 ABSTRACT Temporal Genetic Analysis of Steelhead (Oncorhynchus mykiss) Reveals Hatchery-Induced Drift in Captivity Melissa R. Reneski The primary evolutionary goal of many hatchery supplementation programs is to minimize genetic change and fitness loss associated with captive breeding. The objective of this study was to assess the genetic stability of a hatchery steelhead (Oncorhynchus mykiss) population from the Mad River (California, USA) over 35 years of captive breeding. The genetic population structure of historical (1974-1975) and contemporary (2000-2010) hatchery and wild stocks were compared using 14 microsatellite loci. Contributions from four non-indigenous populations transplanted to the Mad River were also assessed. The founding steelhead population artificially propagated at Mad River Hatchery was resolved as genetically similar to wild steelhead from natural areas in the Mad River and those from the Eel River. From this starting point, the captive population diverged over 35 years while the wild population retained the historical condition but was also highly admixed (17-44%) with the newly formed hatchery population. Analyses indicated that contributions from non-local populations were not responsible for divergence of the hatchery population. Instead a lower effective population size in the hatchery (Ne =246-285) in comparison to the wild (Ne=1,935-4,356) and closure of the hatchery population to immigration from the wild suggests drift in isolation is responsible for divergence. The small number of spawners used in many years at the hatchery along with variance in reproductive success likely explains the observed level of drift. The evolutionary potential of both hatchery and wild populations may be constrained unless iii efforts are implemented to reduce divergence among them and mitigate for the deleterious effects of genetic drift in the hatchery population. iv ACKNOWLEDGEMNTS Funding for this research project was provided by California Department of Fish and Game’s Steelhead Fishing Report and Restoration Card program. Additional funding was provided through scholarships by the Marin Rod and Gun Club, Eureka Rotary Club, Golden West Women Flyfishers Foundation, and Granite Bay Flycasters. I especially thank Dr. Andrew Kinziger for his immeasurable guidance and discussions throughout this process. Sincere thanks to Dr. Carlos Garza for providing laboratory space and materials, genotype data and feedback during the writing process. I would also like to thank members of the NMFS Santa Cruz genetics lab (Libby Gilbert-Horvath, Alicia Abadia, Devon Pearse and Anthony Clemento) for all their time in assisting during the genotyping and analysis components of the project. Many thanks to folks at the California Department of Fish and Game in Arcata (Mark Zuspan, Seth Ricker and Justin Garwood). Your support was instrumental in initiating my graduate career, devising field logistics and providing archived scale samples from the Northern California Scale Archive. Sample collections were also provided by the Washington Department of Fish and Wildlife. I am grateful for the comments provided by Drs. David Hankin and Eric Loudenslager. A sincere thank you to Ken Lindke for providing constant support and encouragement throughout this process. v TABLE OF CONTENTS Page ABSTRACT ....................................................................................................................... iii AKNOWLEDGMENTS ......................................................................................................v LIST OF TABLES ............................................................................................................ vii LIST OF FIGURES ......................................................................................................... viii LIST OF APPENDICES .................................................................................................... ix INTRODUCTION ...............................................................................................................1 MATERIALS AND METHODS .........................................................................................6 Sample Collection ....................................................................................................6 DNA Extraction and Microsatellite Analysis ..........................................................8 Statistical Analysis ...................................................................................................9 Data Quality .................................................................................................9 Genetic Diversity .........................................................................................9 Population Structure...................................................................................10 Effective Population Size and Migration ...................................................12 RESULTS ..........................................................................................................................14 Data Quality ...........................................................................................................14 Genetic Diversity ...................................................................................................16 Population Structure...............................................................................................16 Effective Population Size and Migration ...............................................................23 DISCUSSION ....................................................................................................................26 Temporal Genetic Change .....................................................................................26 Introgression by Non-local strains .........................................................................27 Asymmetric Gene Flow .........................................................................................28 Drift in Isolation .....................................................................................................29 CONCLUSIONS AND RECOMMENDATIONS ............................................................31 LITERATURE CITED ......................................................................................................33 vi LIST OF TABLES Table Page 1 Summary of O. mykiss sample populations collected from the Mad River and comparative basins including population identification (ID), sampling location, sample size, sample status, sample type (T=tissue, S=scale), and collection year(s) ....................................................................................................................7 2 Sample populations, sample size (n), percent missing data, mean number of alleles per locus (A), rarified allelic richness (Ar), rarified private allelic richness (Ap), expected heterozygosity (HE) and observed heterozygosity (HO) for Mad River and comparative populations of steelhead. ................................................15 3 Pairwise genetic distance values (FST) for all Mad River and comparative O. mykiss populations. All FST estimates were significant except those in bold. ....17 4 Demographic estimates of for Mad River Hatchery steelhead from 1975 to 2009 based on methods of Waples (1990). is the number of female spawners within a generation and is the harmonic mean of the number of total spawners in generation ......................................................................................24 vii LIST OF FIGURES Figure Page 1 Sample populations including the (A) locations of the comparative out-of-basin populations Washougal (WA), Eel (EEL), Russian (RUSS), and San Lorenzo (SAN_LOR) rivers, and (B) location of the Mad River (MAD) basin (Northern California) including the Mad River Hatchery (triangle) and Matthews dam (circle) forming the upstream limit of anadromy ..................................................3 2 Discriminant analysis of principle componenets (DAPC) of multi-locus genotype data for all sample populations, including Mad River and comparative stocks. Axis one and two accounted for 39.7% and 32.2% of the variability in the data .................................................................................................................19 3 Discriminant analysis of principle components (DAPC) of multi-locus genotype data for Mad and Eel river populations only. Axis one and two accounted for 63.0% and 23.2% of the variability in the data ...................................................20 4 Neighbor-joining tree constructed using 1000 bootstrap pseudo-replicates. Bootstrap support greater than 70% indicated. ....................................................21 5 Bayesian cluster analysis results including individual admixture proportions (Q) (top) and mean ∆K and L(K) (±SD) values from 1 through 8 genetically distinct clusters (K) (bottom) as determined over 20 independent runs for (a.) all sample populations, (b.) historical and comparative collections and (c.) contemporary and comparative collections. The number of genetically distinct clusters (K) are indicated by asterisks and correspond to the number of unique colors in the individual