Microbial Endophytes Katie Fishburn and Jared Bodecker Phytoremediation Microbial Endophytes Katie Fishburn and Jared Bodecker

Endophytes Microbes that live in plant tissue, that increase plant biomass and resistance to pathogens Of the 300,000 known species of plants, there is at least one plant-associated endophyte (Strobel G. et al) Can remediate pollutants just as good as rhizospheric microbes, in some cases better There are clear advantages of endophytes compared to the closely associated rhizospheric cousins

Known endophytes in PR Highlight the endophytes were discussing

Endophyte vs. Rhizospheric Population control is easier than soil microbes, due to lack of competition (protected inside plant tissue) Degradation of pollutants will be localized to inside the plant, being contained from leaking in the soil and air environment. Plant species-specific selection of necessary bacteria for a specific pollutant Microbial sampling is easier. (plant tissue)

Applications endophyte-phytoremediation The diagram of four major applications

Phytoremediation: Organics 2,4-D (herbicide) degradation Isolates from poplar inoculated into pea plants; planted in soils with 13 mg of 2,4-D Control: high levels of 2,4-D remain Inoculated plants: no 2,4-D remains in test soils PAH tolerance in willow and poplar Used also as carbon source by microbes TNT, RDX, HMX degradation by methylobacterium found in hybrid poplar Mineralized 60% of RDX and HMX in 2 months 2,4-D (herbicide) degradation Isolates from poplar inoculated into pea plants; planted in soils with 13 mg of 2,4-D Control: high levels of 2,4-D remain Inoculated plants: no 2,4-D remains in test soils

2, 4-D Degradation Root tissue Stem tissue Psuedomonas putida mini Tn5 (GFP)

Phytoremediation: Organics 2,4-D (herbicide) degradation Isolates from poplar inoculated into pea plants; planted in soils with 13 mg of 2,4-D Control: high levels of 2,4-D remain Inoculated plants: no 2,4-D remains in test soils PAH tolerance in willow and poplar Used also as carbon source by microbes TNT, RDX, HMX degradation by methylobacterium found in hybrid poplar Mineralized 60% of RDX and HMX in 2 months PAH tolerance in willow and poplar Used also as carbon source by microbes

Poly Aromatic Hydrocarbons Backdrop picture courtesy of NASA

Phytoremediation: Organics 2,4-D (herbicide) degradation Isolates from poplar inoculated into pea plants; planted in soils with 13 mg of 2,4-D Control: high levels of 2,4-D remain Inoculated plants: no 2,4-D remains in test soils PAH tolerance in willow and poplar Used also as carbon source by microbes TNT, RDX, HMX degradation by methylobacterium found in hybrid poplar Mineralized 60% of RDX and HMX in 2 months TNT, RDX, HMX degradation by methylobacterium found in hybrid poplar Mineralized 60% of RDX and HMX in 2 months

Methylobacterium spp. Strain BJ001

Phytoremediation: Inorganics

Brassica chinensis Brassica chinensis Mucor isolate Cd and Pb tolerance Cd and Pb more water soluble; more bioavailable

Thlaspi goesingense Thlaspi goesingense Methylobacterium and Shpingomonas Ni accumulation and tolerance siderophores and ethylene relieve

Plant Yield and Growth Promotion Enhanced nutrient/mineral availability and uptake Phosphate, nitrogen Phosphate solubilization Siderophore production Minerals made more bioavailable; solubilized by siderophore Osmotic adjustment, stomatal regulation

Plant Health and Protection Lessen/prevent effects of pathogens on plants Fungal, bacterial, viral diseases less harmful after inoculation of endophytes Endophytes trigger induced systemic resistance (ISR) Immune-like response by plants

Industrial/Medical Applications Pseudomonas, Burkholderia, Bacillus Antibiotics, anti-cancer compounds, antifungal, antiviral, insecticidal, immunosuppressant agents

Making a (6) Million $ Endophyte Engineering plant-associated microbes Horizontal Gene Transfer Improving already efficient phytoremediation By-products of degradation can even more toxic when released into the air When in planta, you have natural reduction of volatile toxins

Improving Toluene Degradation Normal plant degradation of toluene creates toxic by-products that are volatized into the environment. Burkholderia cepacia G4 (rhizoshperic) with toluene-degrading qualities from pTOM plasmid (toluene-ortho-monooxygenase) B. cepacia VM1330 (endophyte), with the pMON plasmid

Enhanced Toluene-resistance Growth Index (Fresh Weight) Control- Yellow Lupine (no inoculate) BU0072- YL endophyte strain G4- YL rhizospheric strain (pTOM) VM1330- YL transformed endophyte strain (pTOM) Total Weight (grams)

Reduced Toluene-volatization Improved degradation in VM1330 and Yellow Lupine. Which lowers phytotoxicity and evapotranspiration Sub-phytotoxic levels used in experiments (100mg/L)

Lupine endophytes in Poplar Decreased Toluene-volatization Control- Poplar (no innoculate) VM1466- P., endophyte (pTOM) BU61- P., rhizospheric (pTOM)

Horizontal Gene Transfer Opposite of vertical transfer (parental or ancestral) Genetic material is transferred from another organism to another organism without being an offspring Common routes of transfer Transformation: Uptake of endogenous DNA (nature/lab) Transduction: Delivery via vector (bacteriophage) Conjugation: a physical connection between two organisms delivers DNA from one to the other

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