Isolating and Identifying Fungal Endophytes from Roots of Rare Orchid Cypripedium Arietinum in Grand Sable Dunes, Michigan Kari Farkas-Lasich [email protected]

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6-2018 Isolating and Identifying Fungal Endophytes from Roots of Rare Orchid Cypripedium arietinum in Grand Sable Dunes, Michigan Kari Farkas-Lasich [email protected]

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Recommended Citation Farkas-Lasich, Kari, "Isolating and Identifying Fungal Endophytes from Roots of Rare Orchid Cypripedium arietinum in Grand Sable Dunes, Michigan" (2018). All NMU Master's Theses. 553. https://commons.nmu.edu/theses/553

This Open Access is brought to you for free and open access by the Student Works at NMU Commons. It has been accepted for inclusion in All NMU Master's Theses by an authorized administrator of NMU Commons. For more information, please contact [email protected],[email protected]. ISOLATING AND IDENTIFYING FUNGAL ENDOPHYTES FROM ROOTS OF RARE ORCHID CYPRIPEDIUM ARIETINUM IN GRAND SABLE DUNES, MICHIGAN

Kari L. Farkas-Lasich

THESIS

Submitted to Northern Michigan University In partial fulfillment of the requirements For the degree of

MASTER OF SCIENCE IN BIOLOGY

Office of Graduate Education and Research

June 2018

ABSTRACT

ISOLATING AND IDENTIFYING FUNGAL ENDOPHYTES FROM ROOTS OF RARE ORCHID CYPRIPEDIUM ARIETINUM IN GRAND SABLE DUNES, MICHIGAN

Kari L. Farkas-Lasich

Recently the population of Cypripedium arietinum in Grand Sable Dunes, Michigan was estimated at more than 3.5 million plants, greater than 90% of the world’s known population. There is little research on this species’ root associated fungal endophytes.

This new population estimate allows the ethical, destructive sampling required to shed light on root associated symbionts and further successful preservation and restoration of the species. Samples were collected of current year’s roots from 75 C. arietinum orchids at 75 locations within 4 jack pine forests in the Grand Sable Dunes, Pictured Rocks

National Lakeshore, Michigan. Fungal endophyte cultures were obtained from roots and subcultured for DNA extraction. DNA was PCR amplified with primers that isolate the highly conserved 5.8s rDNA and the highly variable internal transcribed spacer (ITS) regions on either side of it. DNA was sequenced and sequences were identified using

NCBI BLAST. Of the 36 DNA samples 27 were positively identified. Phialocephala fortinii and Leptodontidium orchidicola were the most prevalent identifications. These fungi both have saprotrophic abilities and may play an important role in C. arietinum’s lifecycle.

Keywords. Cypripedium arietinum, Phialocephala fortinii, Leptodontidium orchidicola,

Rhizoscyphus ericae, Grand Sable Dunes

KARI L. FARKAS-LASICH

June 2018

ACKNOWLEDGMENTS

I would like to thank my advisor Dr. Donna Becker and committee members Dr. Robert

Belton and Dr. Alan Rebertus for encouragement and support throughout this research.

Bruce Leutscher, Chief of Science and Natural Resources at Pictured Rocks National

Lakeshore provided great resources to help this project along. Thank you to Jake Rich,

Jessica Ford, Steve Caird and Kristin Denryter for braving cold and rainy fall days in the field, to Steve Schaar and Mike Peters for endless helpful lab hours and more, to

Adrienne Bozic for the many exciting conversations that led to this project and to the

NMU Biology Department faculty, staff and graduate students who are amazing resources with a wealth of knowledge and creative problem solving ideas.

Thanks to the Michigan Botanical Club, NMU Excellence in Educat