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Persistent, symptomless, systemic, and seed-borne infection of lettuce by Botrytis cinerea
Experiments are presented which show that Botrytis cinerea, the cause of gray mould disease, is often present in symptomless lettuce plants as a systemic, endophytic, infection which may arise from seed. The fungus was isolated on selective media from surface sterilized sections of roots, stem pieces and leaf discs from symptomless plants grown in a conventional glasshouse and in a spore-free air-flow provided by an isolation propagator. The presence of B. cinerea was confirmed by immuno-labelling the tissues with the Botrytis-specific monoclonal antibody BC-12.CA4. As plants grew, infection spread from the roots to stems and leaves. Surface sterilization of seeds reduced the number of infected symptomless plants. Artificial infection of seedlings with dry conidia increased the rate of infection in some experiments. Selected isolates were genetically finger-printed using microsatellite loci. This confirmed systemic spread of the inoculating isolates but showed that other isolates were also present and that single plants hosted multiple isolates. This shows that B. cinerea commonly grows in lettuce plants as an endophyte, as has already been shown for Primula. If true for other hosts, the endophytic phase may be as important a component of the species population as the aggressive necrotrophic phase
Evidence for a Common Toolbox Based on Necrotrophy in a Fungal Lineage Spanning Necrotrophs, Biotrophs, Endophytes, Host Generalists and Specialists
The Sclerotiniaceae (Ascomycotina, Leotiomycetes) is a relatively recently evolved lineage of necrotrophic host generalists, and necrotrophic or biotrophic host specialists, some latent or symptomless. We hypothesized that they inherited a basic toolbox of genes for plant symbiosis from their common ancestor. Maintenance and evolutionary diversification of symbiosis could require selection on toolbox genes or on timing and magnitude of gene expression. The genes studied were chosen because their products have been previously investigated as pathogenicity factors in the Sclerotiniaceae. They encode proteins associated with cell wall degradation: acid protease 1 (acp1), aspartyl protease (asps), and polygalacturonases (pg1, pg3, pg5, pg6), and the oxalic acid (OA) pathway: a zinc finger transcription factor (pac1), and oxaloacetate acetylhydrolase (oah), catalyst in OA production, essential for full symptom production in Sclerotinia sclerotiorum. Site-specific likelihood analyses provided evidence for purifying selection in all 8 pathogenicity-related genes. Consistent with an evolutionary arms race model, positive selection was detected in 5 of 8 genes. Only generalists produced large, proliferating disease lesions on excised Arabidopsis thaliana leaves and oxalic acid by 72 hours in vitro. In planta expression of oah was 10–300 times greater among the necrotrophic host generalists than necrotrophic and biotrophic host specialists; pac1 was not differentially expressed. Ability to amplify 6/8 pathogenicity related genes and produce oxalic acid in all genera are consistent with the common toolbox hypothesis for this gene sample. That our data did not distinguish biotrophs from necrotrophs is consistent with 1) a common toolbox based on necrotrophy and 2) the most conservative interpretation of the 3-locus housekeeping gene phylogeny – a baseline of necrotrophy from which forms of biotrophy emerged at least twice. Early oah overexpression likely expands the host range of necrotrophic generalists in the Sclerotiniaceae, while specialists and biotrophs deploy oah, or other as-yet-unknown toolbox genes, differently
Deciphering the biology of Cryptophyllachora eurasiatica gen. et sp. nov., an often cryptic pathogen of an allergenic weed, Ambrosia artemisiifolia
The Genome of Botrytis cinerea, a Ubiquitous Broad Host Range Necrotroph
Botrytis cinerea is a necrotrophic ascomycete, causing serious pre- and postharvest crop losses worldwide on a wide variety of plant species. Considerable research in recent years has unraveled a variety of molecular tools that enables the fungus to invade host tissue, including the secretion of toxic proteins and metabolites and host cell wall degrading enzymes. High-quality sequences from two strains of B. cinerea revealed an average-sized genome with a low fraction of repetitive DNA and no specific features correlated with its pathogenic lifestyle, except for a large number of genes encoding pectin-degrading enzymes that are particularly active on cell walls of dicot hosts. The genome sequences enabled genomics, transcriptomics, proteomics, and map-based cloning approaches that have resulted in the identification of genes important for the regulation of secondary metabolite synthesis, drug efflux, light-dependent processes, and pathogenicity. The recent availability of more genome sequences from different B. cinerea strains and Botrytis species and the combination of different ‘omics’ approaches will rapidly increase our understanding of the biology and diverse infection strategies of the gray mold fungus