pThe physiological role of the mannitol cycle in the wheat pathogen iStagonospora nodorum/i (glume blotch) has been investigated by reverse genetics and metabolite profiling. A putative mannitol 2-dehydrogenase gene (iMdh1/i) was cloned by degenerate PCR and disrupted. The resulting mutated imdh1/i strains lacked all detectable NADPH-dependent mannitol dehydrogenase activity. The imdh1/i strains were unaffected for mannitol production but, surprisingly, were still able to utilize mannitol as a sole carbon source, suggesting a hitherto unknown mechanism for mannitol catabolism. The mutant strains were not compromised in their ability to cause disease or sporulate. To further our understanding of mannitol metabolism, a previously developed mannitol-1-phosphate dehydrogenase (gene impd1/i) disruption construct [Solomon, Tan and Oliver (2005) Mol. Plant–Microbe Interact. b18/b, 110–115] was introduced into the mutated imdh1/i background, resulting in a strain lacking both enzyme activities. The impd1mdh1/i strains were unable to grow on mannitol and produced only trace levels of mannitol. The double-mutant strains were unable to sporulate iin vitro/i when grown on minimal medium for extended periods. Deficiency in sporulation was correlated with the depletion of intracellular mannitol pools. Significantly sporulation could be restored with the addition of mannitol. Pathogenicity of the double mutant was not compromised, although, like the previously characterized impd1/i mutants, the strains were unable to sporulate iin planta/i. These findings not only question the currently hypothesized pathways of mannitol metabolism, but also identify for the first time that mannitol is required for sporulation of a filamentous fungus./p
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