Endophytes: biocontrol killers?

Sarah Dodd, Daniel Than,

Stanley Bellgard and Chris Winks (LCR)

Rebecca Ganley (Scion)

Trevor James (AgR)

Endophytes

• All plants are infested with microbes

– Symptomless• Epiphytes/Endophytes• Balanced status of symbiosis• = Majority

– Disease• Pathogens• Unbalanced status of symbiosis

Endophyte/Plant relationship– Endophyte gains

• shelter and nutrients

– Plant cost • nutrients and resources

– (e.g. more susceptible to pests and diseases: BCAs)

– Plant gains • Growth promotion

– (enhanced nutrient uptake)

• Increased tolerance to harsh environments – (e.g. drought tolerance)

• Induced Resistance to pests and diseases (BCAs)

Endophytes – weeds and weed biocontrol

• Harry Evan’s (2007) proposed:

1.Weediness: Endophytes influence the fitness of a plant and therefore its invasiveness

2.Biocontrol: Endophytes interact with plants and/or biocontrol agents either enhancing or reducing their activity

Microbial interactions• Pathogens

• Endophytes/epiphytes

AntibiosisParasitismCompetitionCell lysis

Microbe interactions – under the microscope

Weed / Endophyte systems under investigation in NZ

Sclerotinia on Californian thistle

Phoma on Old Man’s Beard

Wilding Ginger Privet

Pampas

Sclerotinia on Californian thistle- a mycoherbicide

– 1. Kills entire plant– 2. Partially kills plant

– Plant recovers

– 3. No effect on plant

Aim

• To identify microbes that influence the success/failure of a pathogen biocontrol agent

• To ultimately manipulate interactions to improve consistency of weed biocontrol activity

Progress: step 1determine population

variation

Roots, shoots, flowers & seeds

Methods - to identify endophytes

• Culturing – isolate & identify key endophyte fungi

● DGGE - DNA profile comparisons

- endophyte fungi and bacteria - DNA sequence to identify bands (microbes)

Culturing – isolation and identification

Molecular -DGGE

• Samples are all assessed and bands(microbes) identified

• Variability between tissues/plants/fields has been assessed

Endophytic microbes from thistle (closest GenBank match)

Culturing DGGE Known biology

Fungi - AscomycotaAlternaria sp.Aureobasidium pullulansBionectria sp.Botryosphaeria laricinaCladosporium cladosporioidesCladosporium sp.Codinaeopsis sp.Colletotrichum acutatumCryptococcus rajasthanensisCurvularia sp.Cylindrocarpon sp.Davidiella tassianaEpicoccum nigrumEudarluca caricisExophiala sp.Fusarium cortaderiae Fusarium oxysporum Fusarium solani Fusarium sp. 1.Hypocrea/TrichodermaLeptodondium orchidicolaLewia infectoriaNeonectria radicola Nigrospora oryzaePhaeococcomyces chersonesosPhoma exiguaPhoma sp.Phomopsis theicolaPhomopsis sp.Pichia fermentansPlectosphaerella cucumerinaPlectosphaerella sp.Preussia isomeraPreussia sp.Pyrenochaeta terrestrisSclerotinia sp.Stachybotrys echinataStemphylium sp.Verticillium dahliaeXylariaceae sp.

L,SSL--L,S-L,S,PSL--L--LRRL,RLL,RLRL--RLL---LL-L--S,RL,S,P

LRPS-LSR-RL--RSL,S,R,PL,SR-R----L--RL,R---LRL-L,R,PR-LSL,P-

saprobepossible plant pathogen or saprobepossible plant pathogenpossible secondary pathogensaprobesaprobesaprobepossible secondary pathogenyeast-like saprobegrass pathogenpossible plant pathogensaprobesaproberust mycoparasitesaprobepossible plant pathogenpossible plant pathogenpossible plant pathogenpossible plant pathogenPossible mycopathogensaprobesaprobepossible plant pathogensaprobesaprobeplant pathogenpossible secondary pathogen or saprobePossible secondary pathogen or saprobepossible secondary pathogen or saprobeyeastplant pathogenpossible plant pathogencoprophilous saprobecoprophilous saprobeonion path/soil saprobeplant pathogensaprobesaprobeplant pathogensaprobe

Endophytic microbes from thistle (closest GenBank match)

Culturing DGGE Known biology

Fungi - BasidiomycotaBovista plumbea Ceratobasidium sp.Cyathus stercoreusExidiopsis sp.Flammulina velutipesGloeoporus dichrousKuehneromyces rostratusLangermannia giganteanLimonomyces roseipellisMelampsora laricis-populinaMycena sp. / Nolanea sp.Panaeolus sphinctrinusPeniophora pini / aurantiacaPleurotopsis longinquaPleurotus pulmonariusPuccinia chrysanthemi / carduorum Puccinia cnici-oleraceiRogersella griseliniae / Hyphodontia crustosa Schizophyllum communeTomentellopsis submollis / bresadoliana

--------------------

RRSRRLSLPLRRL,PS,RLLLRLS

saprobepossible plant pathogensaprobesaprobesaprobesaprobesaprobesaprobesaprobeplant pathogen (rust)saprobesaprobesaprobesaprobepossible plant pathogenplant pathogen (rust)plant pathogen (rust)saprobesaprobesaprobe

BacteriaPantoea sp. L,R - unknown

Testing influence of endophytes

• Glasshouse trials – assess disease developmenta) BCA + key endophyte

b) BCA - key endophyte

Preliminary glasshouse trials• Can endophytes influence Sclerotinia

disease? 1. No influence

2. Enhanced Sclerotinia disease3. Reduced Sclerotinia disease (e.g. Colletotrichum sp.)

Summary

• We now have evidence to support the theory that endophytes DO influence the activity of pathogen biocontrol agents

The Future

• Sclerotinia on Cali thistle– Identify key bacterial endophytes– Continue glasshouse testing– Identify key endophyte modes of action– Improve pathogen biocontrol agent activity

• Other plant systems– Identify key endophytes and test in

glasshouse– Endophytes and insect BCAs

Californian thistle Phoma

Californian thistle rust