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MONITORING MYCORRHIZAL FUNGI ON PLANTED WHITEBARK PINES IN THE GYE DO PLANTED PINES HAVE THE “RIGHT STUFF” ON THEIR ROOT - PowerPoint PPT Presentation


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MONITORING MYCORRHIZAL FUNGI ON PLANTED WHITEBARK PINES IN THE GYE DO PLANTED PINES HAVE THE “RIGHT STUFF” ON THEIR ROOTS FOR SURVIVAL?. Cooperators: Mary Hecktner, YNP Resource Manager, Yellowstone National Park Dan Reinhart, Resource specialist, YNP & Kay Izlar, U of M

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slide1

MONITORING MYCORRHIZAL FUNGI ON PLANTED WHITEBARK PINES IN THE GYE

DO PLANTED PINES HAVE THE “RIGHT STUFF” ON THEIR ROOTS FOR SURVIVAL?

Cooperators:

Mary Hecktner, YNP Resource Manager, Yellowstone National Park

Dan Reinhart, Resource specialist, YNP & Kay Izlar, U of M

Bob Keane, USFS Fire Ecologist, Missoula Fire Office

Julie Shea, USFS Fire Officer Gallatin National Forest, MT

Stan Cook, USFS Silviculturist, Gallatin National Forest, MT

Cyndi Smith, Parks Canada Ecologist, Waterton Lakes National Park

Joyce Lapp, Park Service Silviculturist, Glacier National Park

Cathy L Cripps, PhD, Montana State University

Paul Trusty, graduate student, MSU

Kate Mohatt, graduate student, MSU

Ben Johnson, undergraduate, MSU

Don Bachman, freelance field assistant

GOALS OF OUR RESEARCH PROGRAM

“MYCORRHIZAL FUNGI OF WHITEBARK PINE”

1. DISCOVERY - ongoing

2. MONITORING – today’s topic

WHAT ARE MYCORRHIZAL FUNGI?

DUNRAVEN PASS, YNP

FRIDLEY BURN, GALLATIN NATIONAL FOREST, MT

3. APPLICATION - the future

Seedling planted along Dunraven Pass

slide2

WHITE BARK PINE FORESTS ARE SERIOUSLY DECLINING

THREATS

  • White bark pine blister rust
  • Mountain pine beetle
  • Fire exclusion
  • Climate change

Tomback, Arno & Keane 2001

Significant restoration efforts have been ongoing for the last 15 years, yet no one has addressed mycorrhizal fungi and whitebark pine!

slide3

WHAT ARE MYCORRHIZAE?

2 MAIN TYPES

A mutualistic relationship between certain fungi and plant roots

“beneficial to both”

Benefits to fungus

get sugars from the plant i.e. Food!

Potential benefits to plant

enhanced phosphorus uptake

improved access to nitrogen

protection from

drought

soil pathogens/grazers

heavy metals

Mycorrhizal fungi can also

aggregate soil

provide links to other plants

spores

Fruiting bodies

mycelium

ectomycorrhizae

Brundrett et al. 2007 website

slide4

ARE MYCORRHIZAL FUNGI IMPORTANT IN WHITEBARK PINE SYSTEMS?

  • PINES CANNOT SURVIVE IN NATURE WITHOUT MYCORRHIZAL FUNGI
  • PINES ASSOCIATE WITH ONLY A SUBSET OF THE 6,000 SPECIES OF ECM FUNGI
  • SOME ECM FUNGI ARE SPECIFIC FOR: PINES, 5-NEEDLE PINES, OR EVEN STONE PINES
  • SOME ECM FUNGI ASSOCIATE WITH MANY TREE SPECIES (GENERALISTS)
  • TYPICALLY ONE TREE HOSTS MANY SPECIES OF ECM FUNGI
  • ECM FUNGI ARE NOT ALL THE SAME, EACH PROVIDES UNIQUE BENEFITS TO THE TREE

YES! But which ones?

ECM FUNGI ARE CRUCIAL TO ESTABLISHMENT, SURVIVAL, SUSTAINABILITY OF WHITEBARK PINE!

Cripps 2001 Encyclopedia of Plant Pathology; Smith and Read 1998. Mycorrhizal Symbiosis

slide5

HOW DO WE STUDY ECTOMYCORRHIZAL FUNGI?

  • Identification of ectomycorrhizal fruiting bodies in pure whitebark pine stands (& DNA analysis)
  • 2)Sampling ectomycorrhizae on roots (DNA analysis of ITS region)
  • Matching DNA to that of sporocarps or Genbank Library for identification

ITS-1

DNA

Amanita alpina

Courtesy CSIRO

Fruiting bodies

Ectomycorrhizae on roots

TO IDENTIFY FUNGI ON ROOTS & CONFIRM ASSOCIATION

Gardes & Bruns 1993. ITS primers with enhanced specificity for basidiomycetes-application for the identification of mycorrhizae and rusts. Mol. Ecol. 2: 113-118.

slide6

1. DISCOVERY

WHICH MYCORRHIZAL FUNGI ASSOCIATE WITH WHITE BARK PINE IN THE GREATER YELLOWSTONE ECOSYSTEM & THE SURROUNDING REGION?

New World District, Gravelly Mountains, Sacajawea Peak, Big Sky Ski Area, Golden Trout Lakes, Dunraven Pass, Waterton Park, Glacier Park

Glacier National Park

Waterton Lakes National Park

Yellowstone National Park

32 species of ECM fungi confirmed with whitebark pine by fruiting bodies or ectomycorrhizae on roots

BASIDIOMYCOTA

AMANITACEAE

Amanita "alpina"

HYGROPHORACEAE

Hygrophorus gliocyclus

Hygrophorus marzuolus (Fr.)

Hygrophorus olivaceoalbus

Hygrophorus subalpinus

TRICHOLOMATACEAE

Leucopaxillus paradoxis

Tricholoma moseri

CORTINARIACEAE

Cortinarius clandestinus

Cortinarius duracinus

Cortinarius “flavobasalis”

Cortinarius “flavoroseus”

Cortinarius aff. fulminoides

Cortinarius subolivescens

Cortinarius sp.

Dermocybe crocea (Schff.) Mos.

Inocybe sp.

RUSSULALES

Lactarius deliciosus

Russula cf tortulosa /queletii

Russula sp. 2, Russula sp. 3

Unculturable, shared with other conifers

OLDER TREES

BOLETALES

Boletus edulis

Chroogomphus sp. nov.

Rhizopogon cf milleri

Rhizopogon cf evadens

Rhizopogon spp.

Suillus subalpinus

Suillus sibiricus

Suillus tomentosus var. discolor

Suillus sp.

PHALLALES - GOMPHALES

Hysterangium separabile

THELEPHORALES

Tomentelloid type 1

Tomentelloid type 2

ASCOMYCOTA

Cenococcum geophilum

SUILLOIDS – mostly specific for pines, 5-needle pines & stone pines

SEEDLINGS, YOUNGER TREES, & OLDER TREES

Unculturable, some may be specific for pines or shared with other conifers

MOSTLY OLDER TREES

Generalist, important in dry conditions for relations

SEEDLINGS UNDER CANOPY & OLDER TREES

Cripps & Mohatt 2005 (Nutcracker Notes), Mohatt 2006 (MSU Thesis)

Mohatt, Cripps & Lavin (in ed) Ectomycorrhizal fungi of whitebark pine (a tree in peril) revealed by sporocarps and molecular analysis of mycorrhizae from treeline forests in the Greater Yellowstone Ecosystem. Can. J. Bot. (coming soon!)

slide7

Suilloids–show host specificity

Cripps photos

Moser 2004. In Cripps: Fungi in Forest Ecosystems, NYBG Press. (fungi with 5-needle pines in Alps, Altai, Rocky Mts)

Suillus subalpinus – 5-needle pine/stone pines

Suillus sibericus – stone pines

Chroogomphus sp nov (new species)- whitebark pine

Rhizopogon milleri & R. evadens (pine/5-needle/stone pine)

slide8

UNDERGROUND MYCORRHIZAL FUNGI COMMON IN THE SYSTEM

Rhizopogon

“POGIES”

pine

pine

These fungi are eaten by squirrels, deer, elk & bears which spread the spores

Gautieria

pine-spruce-fir

Peaco photo, YNP

Czares & Trappe 1994. Spore dispersal of ectomycorrhizal fungi by mammal mycophagy. Mycologia 86

slide9

Spore dispersal by deer

DUNRAVEN PASS, YNP

“POGIES”

Ashkannejhad & Horton 2006. Ectomycorrhizal ecology on coastal dunes: interactions involving Pinus contorta, Suilloid fungi and deer. New Phytol. 169:345-354.

Mattson et al. 2002. Consumption of sporocarps by Yellowstone grizzly bears. Ursus 13:95-103.

Cripps photos

slide10

2.MONITORING

Project 1: Whitebark Pine Restoration, Dunraven Pass Yellowstone National Park: Monitoring the mycorrhizal status of planted whitebark pine seedlings.

  • Sept. 11-13 2006 whitebark pine seedlings planted along Dunraven Pass
  • Sept. 11-13 2006 Kay Izlar (U of M) set up 10 plots along the pass
  • Sept 14: We selected 10 of nursery seedlings from this batch to check roots for nursery fungi
  • June 2006 Izlar measured % survival on 10 plots
  • June 2007 We sampled roots of seedlings near/on each plot (after 9 months)

10 sites where whitebark pine seedlings were planted & monitored

(Izlar data)

Greater Yellowstone Coordinating Committee & Rocky Mountain Research Station funding. Cooperators: Mary Hecktner Resource manager YNP, Dan Reinhart YNP, and Kay Izlar, U of M.

slide11

OBJECTIVES: To determine

  • if planted seedlings are effectively colonized by native fungi [ON-SITE MONITORING]
  • inoculum potential of replaced soil [GREENHOUSE BIOASSAY]
  • if imported nursery fungi persist on planted seedlings [MOLECULAR TOOLS]

WHAT IS THE MYCORRHIZAL CONDITION OF NURSERY SEEDLINGS BEFORE OUT- PLANTING?

  • 98% non-mycorrhizal roots for sample of 10 nursery seedlings from Coeur D'Alene nursery
  • 2% nursery fungi on roots : Thelephora & E-strain
  • exotic non-native fungi (benefit or concern?)
  • do they persist in soil after out-planting and spread?
  • evidence of fungal root pathogens

FUNGI ON NURSERY SEEDLINGS? BENEFIT OR CONCERN?

Root hairs = non-mycorrhizal

E-strain fungus – typical in nurseries

Thelephora sp – typical in nurseries

Cripps photos

slide12

Number of mycorrhizal root tips per seedling sampled on each site

Izlar data

Izlar notes

  • No (little) mycorrhizal colonization on compromised seedlings
  • Native fungi were colonizing roots on sites 1, 6, 7, 9 and 11 (50% of sites). Suilloids
  • No colonization of sites 4, 5, 8 (S & SE aspect).
  • Only nursery fungi were present on sites 2 and 3.
slide13

Percent mycorrhizal colonization of root tips and depth of ectomycorrhizae on cone-tainerized roots systems

  • Most seedlings were not well-colonized after 9 months
  • A majority of mycorrhizae were 4-12 cm deep.
slide14

Kay Izlar’s “star performer”: seedling #1, site 6 Collected below a mature whitebark pine

Mycorrhizal colonization of cone-tainerized roots

Cripps photos

slide15

NATIVE ECTOMYCORRHIZAE ON WHITEBARK PINE SEEDLINGS PLANTED ALONG DUNRAVEN PASS (sampled 9 months after planting)

Suilloid type

Rhizopogon

Piloderma

Cenococcum & sclerotia

Cripps photos

A DIVERSITY OF NATIVE SUILLOIDS WERE PRESENT, ALONG WITH OTHER NATIVE ECTOMYCORRHIZAL FUNGI (Rhizopogon species, Suillus species).

Molecular identifications by Paul Trusty

slide16

ARE MYCORRHIZAL FUNGI FOUND IN REPLACED SOIL ON DUNRAVEN PASS AND AVAILABLE TO SEEDLINGS?

METHOD: SOIL BIOASSAY IN GREENHOUSE

T1: REPLACED SOIL, UNSTERILIZED

T2: NATIVE WBP FOREST SOIL, UNSTERILIZED

T3: REPLACED SOIL, STERILIZED (CONTROL)

T4: NATIVE SOIL, STERILIZED (CONTROL)

Native fungi?

Nursery fungi?

Develop of a method to test various soils for presence of appropriate fungi---------

before out-planting

slide17

Mycorrhizal colonization of 2-year-old nursery seedlings planted in various soils from Dunraven Pass (YNP) after 1 year in greenhouse conditions

† incomplete sterilization

Molecular ID by P Trusty

  • Soil bioassay revealed native mycorrhizal fungi appropriate for colonization of whitebark pine in replaced & native soil on Dunraven Pass.
  • Results are not correlated with individual sites (soil samples were mixed)
  • Suilloid fungi present
  • Nursery fungi may have persisted on seedlings in both soils

Native fungi: all Suilloid fungi (Rhizopogon subbadius, R. sp. 1, and Suillus aff. borealis, Suillus sp., Amphinema sp.).

Nursery fungi: Wilcoxina (E-strain) confirmed on some, not confirmed molecularly on all)

slide18

CONCLUSIONS

  • Colonization by native mycorrhizal fungi initiated on 50% of sites after 9 months.
  • Most of the colonization N side of Pass
  • No colonization (1 exception) on compromised seedlings.
  • Colonization levels low on most seedlings after 9 month (one notable exception).
  • Suilloid fungi are present in soil of at least 50% of sites, also in replaced soil on Pass.

RECOMMENDATIONS

  • Second monitoring for % survival & mycorrhizal colonization (2009).
  • Planting seedlings S of pass (with controls)
    • - with native soil from whitebark pine forests (2008/2009)
    • inoculated in greenhouse with native fungi from Yellowstone Park
      • Greenhouse (2008), field (2009)
    • test retroactive methods for inoculation (2008/9)
  • a commercial inoculum of exotic mycorrhizal fungi should NOT be used in sensitive whitebark pine forests!
slide19

2.MONITORING

Assessment of mycorrhizal colonization of rust resistant white bark pine seedlings planted in Post-fire Restoration Treatments on a severe burn (Fridley Burn, Gallatin NF, MT) as a measure of sustainability.

Paul Trusty graduate student MSU

Pacific Biodiversity Institute

NASA EARTH OBSERVATORY AUG 19, 2001

  • In 2002, rust-resistant seedlings from the Coeur D’Alene nursery were planted on burn

Rocky Mountain Research Station Funding. Cooperator Bob Keane, Fire Ecologist,Missoula, MT. Julie Shea, USFS Fire Officer Gallatin NF, Stan Cook, USFS Silviculturist, Gallatin NF, MT

slide20

OBJECTIVES: To determine

  • if planted seedlings are effectively colonized on Fridley burn [ON-SITE MONITORING]
  • inoculum potential of soil from a severe burn [GREENHOUSE BIOASSAY]
  • if imported nursery fungi persist on planted seedlings [MOLECULAR TOOLS]
  • Compare mycorrhizal fungi on seedlings:

2. planted rust resistant seedlings

3. naturally regenerating seedlings

1. Naturally regenerating seedlings

Cripps photos

slide21

METHODS: Monitoring by comparison 2006

3 transects in burn and 3 in adjacent unburned whitebark pine forest, 2X samplings

Minimally destructive sampling: root samples removed from seedlings in situ

N=60 T1 = burned, planted seedlings N=60 T2 = unburned adjacent forest, naturally regenerating seedlings N=24 T3 = burned, naturally regenerating seedlings

RESEARCH PROJECT OF PAUL TRUSTY, MSU GRADUATE STUDENT

Cripps photo

slide22

PRELIMINARY RESULTS:

Mycorrhizal colonization of whitebark pine seedlings 5 years after burn

Cortinarius - unburned

  • 88-98% of root tips were mycorrhizal for all treatments
  • Range of 1-5 “morphotypes” of fungi per seedling
  • More “morphotypes” per seedling for unburned forest vs those on burns (planted/natural)

Amphinema - burn

Pseudotomentella- burn

PAUL TRUSTY DATA & PHOTOS 2006

Byssocorticium - burn

slide23

Relative Importance of Ectomycorrhizal types by “treatment”

A “shift” in mycorrhizal taxa after the burn

burned

unburned

Importance value = relative frequency & relative abundance

Species shift:1. Mycorrhizal fungi on burn different from those in unburned forests.

2. Natural & planted seedlings on burn share fungal taxa.

3. Suilloid fungi coming into burned area after 5 years.

3. Unclear at this point if nursery fungi persist after 5 years.

4. Functional significance of shift is unknown at this point.

PAUL TRUSTY DATA 2007

slide24

Correspondence analysis

  • each symbol represents 1 seedling & mycorrhizal fungi on that seedling
  • Unburned seedlings are clearly separate
  • natural & planted seedlings on burn overlap

PAUL TRUSTY DATA 2007

slide25

CONDITIONS WERE OPTIMIZED FOR MYCORRHIZAL COLONIZATION IN PLANTINGS OF RUST RESISTENT SEEDLINGS ON THE FRIDLEY BURN

    • rust resistant seedlings were planted 1 year after burn.
    • the planting area was in close proximity to mature unburned whitebark pine trees as a source of mycorrhizal inoculum.
    • burned area was previously in whitebark pine, “sporebanks” of suilloid fungi may persist in the soil.
    • mammals that have potential to disperse suilloid fungi and “inoculate” seedlings are present
    • moisture conditions were sufficient

Cone-tainerized root systems on planted seedlings after 5 years

Naturally regenerating seedlings

GREENHOUSE BIOASSY:

Picked up Suilloids & E-strain on burned & unburned soil

slide26

3. APPLICATION

FLOW CHART FOR DETERMINING WHEN INOCULATION IS NECESSARY

HOW CAN WE OPTIMIZE THE SYSTEM?

Inoculation of seedlings with native ECM can increase seedling survival and fitness (Smith 1998)

Cripps, CL  2003. Native mycorrhizal fungi with aspen on smelter-impacted sites in the Northern Rocky Mountains: occurrence and potential use in reclamation. Amercian Society of Mined Land Reclamation, Lexington, KY. Pgs. 193-208.

Mahony, C 2004. Effects of native ectomycorrhizal fungi on aspen seedlings in greenhouse studies: inoculation methods, fertilizer regimes, and plant uptake of slected elements in smelter-impacted soils. M.S. Thesis (Cripps),

Brundrett et al. 1996. Working with Mycorrhizas in Agriculture. CSIRO.

slide27

FUTURE DIRECTIONS

  • Continue to determine the mycorrhizal fungi important to whitebark pine
    • currently in Yellowstone, Glacier & Waterton Parks
  • Monitor whitebark pine seedling planted under a variety of conditions
  • primarily in areas where inoculum in not likely to be available & seedlings compromised.
  • need suggestions on where to monitor
  • Capture & screen native mycorrhizal fungi for use as inoculation
  • we now have about 15 species in culture
  • Developing methods for inoculation of seedlings
  • We need cooperators & access to young whitebark seedlings, a few weeks or months old
  • Test to see if inoculation of seedlings can be retroactive.

THANKS TO ALL THOSE WHO HAVE HELPED WITH OUR WHTEBARK PROJECTS:

Mary Hecktner, YNP Resource Manager, Yellowstone National Park; Dan Reinhart, YNP & Kay Izlar, U of M; Bob Keane, USFS Fire Ecologist, Missoula Fire Office; Julie Shea, USFS Fire Officer Gallatin NF, MT; Stan Cook, USFS Silviculturist, Gallatin NF, MT; Cyndi Smith, Parks Canada Ecologist, Waterton Lakes National Park; Joyce Lapp, Park Service Silviculturist, Glacier National Park; Tara Carolin, Glacier National Park.

slide28

THE END…….

OR JUST THE BEGINNING?