2009 Inoculation of WBP with Native Mycorrhizal Fungi in the Nursery

Project: Inoculation of Whitebark Pine with Native Mycorrhizal Fungi in the Nursery

AttachmentsMSU_Cripps_2009_Full REPORT             MSU_Cripps_2009_ Phase 1

Agency/Forest or Park/District: MSU Plant Growth Center, MSU. Montana

Project coordinator: Cathy L. Cripps

Contact  Cathy L. Cripps  –  Plant Sciences & Plant Pathology Dept.,  Montana State University, Bozeman, MT.  59717  406-994-5226    ccripps@montana.edu


Kent Eggleston of the Coeur d´Alene Forest Nursery (& indirectly John Schwandt) providing seedlings. Mary Hekter (Yellowstone National Park), Bob Keane (USDS Fire Ecologist), Kay Izlar (USFS Botanist), Cyndi Smith (Waterton Lakes National Park), Joyce Lapp (Glacier Park Nursery), Tara Carolin (Glacier Park Ecologist), Stan Cooke (USFS silvicuturist) and Julie Shea (USFS Fire Officer).

Source of funding /amount

FHP: $15,800 Supplemental funding: $3,900 in kind Cripps, Coeur d´Alene Nursery donation of seedlings and Greenhouse material donated by MSU.

Dates of restoration efforts:  Year 2009


To transfer and apply the European technology for inoculating stone pines to whitebark pine seedlings grown in US nurseries for out-planting on restoration sites. Specifically:

–          Apply Moser´s method of growing native fungal inoculum for use in US nurseries

–          Apply Moser´s method of inoculating stone pines to whitebark pine

–          Evaluate various potting media to optimize mycorrhization at transplant stage

–          Evaluate bio-containers vs status quo containers for mycorrhizal colonization of transplants

–          Assess treatments for most effective (timely/vigorous) colonization of mycorrhizal fungi

Acres/ha treated  N/A


Seedlings: we have 400 seedlings, including those that were previously inoculated, un-inoculated controls, extras and those grown from germinating seed.

Fungi: we will further test about 4-6 of our native mycorrhizal strains which are already growing.

1-     Grow (bulk up) selected mycorrhizal fungi in soil, liquid media and grain (Moser 1956, Göbl 1974, Weisleitner 2008, pers comm.).

2-     Use resulting fungal inoculum on seedling being transplanted into various media types from the Styrofoam blocks.

3-     Transplant inoculated seedlings into both bio-containers and plastic containers in the greenhouse. Bio-containers are of a new carbon material and are used in Austria.

Planting? If so, source of seedlings? Resistance?  No


Phase 1 is complete. Phase 2 is a continuing two year project and is not complete. We gained additional isolates of native mycorrhizal fungi to test this summer. Isolates were selected on past performance of the species in Phase 1 trials. Experiments to examine fertilizer and soil effects are set up, seedlings are inoculated with selected native fungi, and are being maintained in the MSU Plant Growth Center (see Appendix 1 for experimental design). We will subject seedlings to cold treatment over the winter and reinoculate them; we expect to assess data in the spring. This winter we are working on developing an inoculum to be “added at planting”. We will also inoculate seedlings (provided by Melissa Jenkins) to be grown in the Coeur D’Alene Nursery—inoculation will take place at the nursery itself as a move towards testing FS nursery conditions as amenable to inoculation. We hope that inoculation can take place before seedlings go into cold treatment, and possibly again afterwards. These seedlings will be planted by the Forest Service (Melissa Jenkins, as per her other seedlings) in various treatments (including burns) and later assessed for survival. We do not expect results from this part of the project for 1 or 2 years. This summer we will continue to collect sporocarps for spore slurries and to test shelf life of current spore slurries, and do additional green house experiments. We are amenable to additional studies as fits the needs of J. Schwandt.

Collection of Sporocarps of Native ECM Fungi

We collected sporocarps of native fungi from whitebark pine forests to develop into spore slurries. We made several field trips for this purpose and were highly successful due to high precipitation this field season. A surprise was to see the widespread distribution of Suillus sibiricus in many whitebark pine forests. Several isolates of native fungi were selected (from Phase 1) for further testing in the greenhouse as an inoculum for whitebark pine seedlings. Our current list of isolates of native ectomycorrhizal fungi collected from whitebark pine forests is shown in Table 1. Many isolates were grown out on MMN (some with antibiotics) for isolation and are maintained in the lab on MMN media in petri dishes and are now in tubes on agar. Spore slurries were developed from sporocarps when possible.

Development of spore slurries

The 2009 field season had high precipitation and produced lots of sporocarps in whitebark pine forests. We took advantage of this and gathered sporocarps from several sites (especially suilloids) and processed them into spore slurries either directly or dried them for later use (Table 2). For inoculum development, sporocarps were stripped of mature hymenium or gleba, and these portions were then either a) developed directly into spore slurries b) dehydrated or c) frozen. For spore slurries, the tissue was blended and then some were filtered; all are kept refrigerated until ready to be used. The shelf life of slurries is currently being assessed. Several native ectomycorrhizal fungi identified as good root colonizers in Phase 1 were used to inoculate one and two year old seedlings for Phase 2. Extra seedlings from Phase 1 plus additional new germinants from the Coeur D’Alene nursery were used in Phase 2.

Experiments currently underway in MSU Plant Growth Center

Experiment 1: Examination of the effects of fertilizer on mycorrhizal colonization of whitebark pine seedlings with native fungi (CLC 2440, Suillus sibiricus). Several fertilizer regimes including one similar to that used in the Coeur D’Alene nursery were selected for assessment to determine if fertilizer deters mycorrhizal colonization (and at what level). See experimental design in Appendix 1, Table 3.

Experiment 2: Examination of the effects of soil type on mycorrhizal colonization of whitebark pine seedlings for various isolates of native fungi. Three soil types (two used by the Coeur D’Alene nursery were selected to be tested. This includes a peat:vermiculite mix, a peat:bark mix and soil mix 2 (peat:MSU mix:vermiculite). This test will help determine if the type of substrate can affect the mycorrhizal colonization process. See experimental design in Appendix 1, Table 4. We are awaiting results on these experiments and expect to assess them after cold treatment in the spring.

Summary of results to date:

1. Inoculum: spore slurries are developed and ready for use. We are testing shelf-life.

2. Inoculum: to be used ‘at planting’ is being developed, but studies show it is preferable to inoculate seedlings in the greenhouse.

3. Whitebark pine seedlings are inoculated for various tests are we are awaiting results for    a) tests for fertilization effects

b) tests for soil/substrate effects

4. Inoculum: is ready for Melissa Jenkins seedlings and will be applied in spring

Monitoring since completion of the project N/A


            Plans for future monitoring?  N/A

Will outcome meet goals?  Yes

Future actions/follow up?  Project still ongoing