Genetics of Norwegian kelp forests Microsatellites reveal the genetic diversity, differentiation, and structure of two foundation kelp species in Norway Ann M. Evankow MSc Thesis Centre for Ecology and Evolutionary Synthesis Department of Biosciences University of Oslo June 2015 © Ann M. Evankow 2015 Genetics of Norwegian kelp forests: Microsatellites reveal the genetic diversity, differentiation, and structure of two foundation kelp species in Norway Ann M. Evankow http://www.duo.uio.no/ Print: University Print Centre, University of Oslo II Preface Two years is gone already. And I can honesty say that I am happy that my world has revolved around kelp for the majority of that time. Thank you, Claudia, for thinking up this amazing project. Without you, I wouldn’t be submitting a thesis about kelp genetics today. And thank you for introducing me to the University of Oslo. I felt welcomed by all of the members of the shark and kelp meetings. Thank you, Hartvig & Janne, for agreeing with Claudia and collecting kelp samples. This project would have never started without you, or finished. Thank you to all the other NIVA folks you have made this project possible and helped me along the way. Thank you, Anne, for accepting me as your student, even though my ideas were far from polyploidy and plants. You have been there for me this entire time and I could not have hoped for a more caring, patient, and optimistic supervisor. Thank you, Stein, for agreeing to supervise me, even though I didn’t end up working with seagrasses. There is always more time, yes? And kelps are wonderful, even though I spent most of the time in a DNA lab, instead of out in the field. Thank you, Marit, for helping me extract and amplify kelp DNA for the first time. I may have given up early on without your support. Thank you, Mikkel, for sharing your PCR supplies and answering my questions in the lab, often at strange hours and about unusual things. And Ryan, for letting me rant about my problems. Thank you, Robin, Sandy and, Eli, for helping me attempt to create a RADseq library of kelp. Next time. I would also like to thank Tove, Anne, Jim, and all the other amazing people I met at UNIS who inspired me and kept me believing that this project could happen. Jim, I am especially grateful for your extra assistance in the lab and comments on this paper. I hope I can visit Appledore this summer or soon! At this point, I need to thank the other masters students (and PhDers). Without you I could not have been my cheery self. You’ve made this entire project possible through coffee breaks, Bunnpris runs, late nights, early computer lab Fridays and millions of other very special moments. No really. I needed you these past two years and I’m sad that it is over so quickly. Lastly, and firstly, thank you Emil for believing in me, every moment of everyday. And… I would like to thank my family. Although I am far from home, I feel your support day in and day out. And with that, I bid you, ha det bra! III Annie with Laminaria hyperborea in Bergen, Norway. Photo credit: Hildur Magnúsdóttir IV Table of Contents Abstract ..................................................................................................................................... 1 Introduction .............................................................................................................................. 3 Loss of kelp forests in Norway ......................................................................................................... 3 Exploring gene flow with genetics .................................................................................................... 4 Study questions .................................................................................................................................. 6 Materials & Methods ............................................................................................................... 7 Study system and species .................................................................................................................. 7 Sample collection and preservation ............................................................................................... 10 DNA extraction ................................................................................................................................ 11 Microsatellite analyses .................................................................................................................... 11 Data analyses ................................................................................................................................... 13 Results ..................................................................................................................................... 17 Microsatellite selection .................................................................................................................... 17 Assumptions of genetic analyses .................................................................................................... 18 Genetic diversity .............................................................................................................................. 21 Site differentiation ........................................................................................................................... 25 Genetic structure ............................................................................................................................. 29 Discussion ................................................................................................................................ 35 The Skagerrak: one region, two patterns of diversity ................................................................. 35 The North: an oasis of unique genetic diversity ........................................................................... 37 Implications for management ........................................................................................................ 38 Future investigations ....................................................................................................................... 39 Conclusions ...................................................................................................................................... 40 Acknowledgements ................................................................................................................. 42 References ............................................................................................................................... 43 Appendix ................................................................................................................................. 49 V VI Abstract Various stressors such as higher temperatures, grazing by sea urchins, and anthropogenic effects may threaten kelp forests around the world. Conservation efforts can benefit from an understanding of current and historic patterns of gene flow and population connectivity of kelp. However, relatively little is known about these patterns, especially with regards to the northern edge of the distribution of kelp species in the North Atlantic Ocean. Knowledge of kelp population genetic diversity and structure can provide crucial information about the resilience and recolonization potential for already threatened populations. This study investigated the genetic diversity, differentiation, and structure of the two foundation kelp species in Norway, Saccharina latissima and Laminaria hyperborea. Nearly 500 individuals were genotyped from 16 different sites along the Norwegian coast using microsatellite loci that cross-amplified from other species. Roughly half of the samples per species amplified and were score-able for three polymorphic markers for S. latissima and 11 for L. hyperborea. Significant genetic structure, differentiation, and variation in genetic diversity were found among sites for both species. There were at least two distinct clusters of S. latissima and four of L. hyperborea. Genetic patterns corresponding to isolation by distance were significant for both species, except within the Skagerrak region. Genetic diversity of L. hyperborea was low in the Skagerrak region and significantly increased with higher latitudes along the Norwegian coast. Genetic diversity of S. latissima was significantly different between sites, but did not vary significantly between larger regions. Overall, this study established molecular tools for future investigations and provided the first glimpse into population genetic patterns of S. latissima and L. hyperborea in Norway. 1 2 Introduction Kelps are foundation species for highly productive marine coastal ecosystems. These large brown seaweeds create three-dimensional forest-like habitats for multitudes of species, including juvenile fish important for fisheries (Norderhaug et al. 2005, Christie et al. 2009). As primary producers, kelp species contribute significant amounts of energy to coastal marine food webs in the form of particulate organic matter (Fredriksen 2003, Norderhaug et al. 2003). Some species, such as sea urchins, are capable of digesting kelp directly and preferentially feed on kelp biomass (Vadas 1977). In addition to their ecological worth, kelps are valuable economic commodities. Kelp species are cultivated and harvested for their alginates and accumulation of rare elements (Vásquez 2009, Kerrison et al. 2015). In China, Saccharina species have been cultivated for over a century (Zhang et al. 2015).