Progress 01/01/05 to 12/31/05
Outputs
We performed In vitro studies on rhizosphere competence. Since we were the first to document rhizosphere competence in an entomopathogenic fungus, we investigated the generality of rhizosphere competence in other fungi commonly regarded as insect pathogens. We used an agarose root model to study behavior and a seedling-soil model to study the dispersal of fungal propagules during root growth. We also used a petri dish-soil model that enabled the easy and repeatable study of different combinations of soil-related variables on gene expression. Using these models we found that many strains of M. anisopliae with a broad host range (e.g., ARSEF2575) germinated at low concentrations of root exudates. However, germination frequencies were reduced by high concentrations of exudates, suggesting they contain mixtures of inducers and repressors of fungal growth. Strains of M. anisopliae specialized to particular hosts showed decreased rhizosphere competence e.g., ARSEF 4600 .
Other species, including the endophytic Beauveria bassiana strain ARSEF 3113, were much less responsive to root exudates than M. anisopliae 2575. Even a commercially available rhizosphere competent strain of T. harzianum (T-22), still required higher concentrations of root exudates to germinate than 2575. My group mostly uses strain 2575 that successfully infects locusts, caterpillars, flies, crickets and beetles, amongst others. Likewise it is promiscuous with respect to the plant roots it colonizes, at least in the seedling-soil model studies. We employed genomic approaches to address the mechanisms of rhizosphere competence. Because our original libraries were made from fungi growing on insect tissues our database only represented insect-response genes. We therefore indexed root exudate induced transcripts and arrayed them with insect response genes to assess expressed genes for critical purposes (e.g., fungal responses to soil, soil microbes, root exudates and other soil
amendments). These continuing studies are providing a vivid picture of differences, overlaps and networking in secreted products (enzymes/toxins etc) and physiological parameters (protein phosphorylation events, transcriptional regulatory factors and physiological cues etc) that define 2575s life as a pathogen and as a saprophyte. To date, we have identified 1918, 1581 and 592 genes that show > 2-fold upregulation on cuticle, hemolymph and root exudates, respectively. Although many genes showed much higher levels of expression in exudate than other media (including an adhesin. We have developed a strategy of using the specific promoters of these genes as a resource for improving biocontrol potential either by expressing crucial transgenes targeted to the rhizosphere or, alternatively, modulating habitat range by specific production of an anti-sense to a crucial regulatory gene.
Impacts
New environment friendly genetically-modified microbial inoculates are being produced commercially and used to protect plants from disease and to promote plant growth. These new products should lead to a reduction in the use of biocides and chemical fertilizers. Nethertheless, biological safety issues must be considered prior to the release of these transgenics into the environment. Mycorrhizial interactions and interactions between bacteria and plants have received the most attention. However, there is an extra dimension in the quality of the interactions with fungi, as unlike bacteria they can potentially grow and spread through the soil and rhizosphere as hyphal growth. Unfortunately, the current predictive data base for risk assessment issues regarding future releases of genetically engineered fungi remains small and very little is known concerning the survival of individual genotypes in the field. Nor has it been demonstrated that we have the capacity to detect
differences in behavior and ecology between parent and transgenic organisms that would allow us to better interpret changes in the ecosystem level. We are remedying these deficiencies by providing the knowledge required to predict and improve fungal responses to various environmental stimuli and to determine side-effects of genetic alterations on functionality in insecticidal products and survival in soil.
Publications
- Wang, C and St. Leger, R.J. 2005. Developmental and transcriptional responses to host and non host cuticles by the specific locust pathogen Metarhizium anisopliae sf. acridum. Eukaryotic Cell 4: 937-947
- Wang, C., Hu, G and St. Leger, R.J. 2005. Differential gene expression by Metarhizium anisopliae growing in root exudate and host (Manduca sexta) cuticle or hemolymph reveals mechanisms of physiological adaptation. Fungal Genetics and Biology 42: 704-718.
- Wang, C., Butt, T.M. and St. Leger, R.J. 2005. Colony sectorization of Metarhizium anisopliae is a sign of ageing. Microbiology 151: 3223-3236
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Progress 01/01/04 to 12/30/04
Outputs
Presumably fungi have different subsets of genes responsive to different growth conditions and so identifying these would illuminate the mechanisms of adaptation. In this study we sampled cDNA libraries constructed during fungal growth in bean root exudate and indexed the core-set of of exudate -induced transcripts that allow saprophytic growth in the rhizosphere. We then arrayed these expressed sequence tags and performed microarray analysis to demonstrate physiological adaptations by Metarhizium anisopliae to growth in the rhizosphere. Critical differences in transcriptional control between pathogenic growth on insects and saprophytic growth on root exudate include perception mechanisms, carbon metabolism, proteolysis, cell surface properties, and synthesis of secondary metabolites. These differences suggested previously unsuspected strategems of adaptation to roots that are being tested experimentally.
Impacts
This project will provide new insight into the intimate relationship between genes, organisms and the environment
Publications
- Wang, C., Hu, G and St. Leger, R.J. 2005. Differential gene expression by Metarhizium anisopliae growing in root exudates and host (Manduca sexta) cuticle or hemolymph reveals mechanisms of physiological adaptation. Fungal Genetics and Biology. In press.
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