The Application of Phytohormones as Biostimulants in Corn Smut Infected Hungarian Sweet and Fodder Corn Hybrids

The main goal of this research was to investigate the effects of corn smut (Ustilago maydis DC. Corda) infection on the morphological (plant height, and stem diameter), and biochemical parameters of Zea mays L. plants. The biochemical parameters included changes in the relative chlorophyll, malondialdehyde (MDA), and photosynthesis pigments’ contents, as well as the activities of antioxidant enzymes—ascorbate peroxidase (APX), guaiacol peroxidase (POD), and superoxide dismutase (SOD). The second aim of this study was to evaluate the impact of phytohormones (auxin, cytokinin, gibberellin, and ethylene) on corn smut-infected plants. The parameters were measured 7 and 11 days after corn smut infection (DACSI). Two hybrids were grown in a greenhouse, one fodder (Armagnac) and one a sweet corn (Desszert 73). The relative and the absolute amount of photosynthetic pigments were significantly lower in the infected plants in both hybrids 11 DACSI. Activities of the antioxidant enzymes and MDA content were higher in both infected hybrids. Auxin, cytokinin, and gibberellin application diminished the negative effects of the corn smut infection (CSI) in the sweet corn hybrid. Phytohormones i.e., auxin, gibberellin, and cytokinin can be a new method in protection against corn smut.

An Unconventional Melanin Biosynthetic Pathway in Ustilago maydis

Ustilago maydis is a phytopathogenic fungus responsible for corn smut disease. Although it is a very well established model organism for the study of plant-microbe interactions, its potential to produce specialized metabolites, which might contribute to this interaction, has not been studied in detail. By analyzing the U. maydis genome, we identified a biosynthetic gene cluster whose activation led to the production of a black melanin pigment. Single deletion mutants of the cluster genes revealed that five encoded enzymes are required for the accumulation of the black pigment, including three polyketide synthases (pks3, pks4 and pks5), a cytochrome P450 monooxygenase (cyp4) and a protein with similarity to versicolorin B synthase (vbs1). Metabolic profiles of deletion mutants in this gene cluster suggested that Pks3 and Pks4 act in concert as heterodimer to generate orsellinic acid (OA) which is reduced to the corresponding aldehyde by Pks5. The OA-aldehyde can then react with triacetic acid lactone (TAL) also derived from Pks3/Pks4 heterodimers to form larger molecules including novel coumarin derivatives. Our findings suggest that U. maydis synthesizes a novel type of melanin based on coumarin and pyran-2-one intermediates, while most fungal melanins are derived from 1,8-dihydroxynaphthalene (DHN) or L-3,4-dihydroxyphenylalanine (L-DOPA). Along with these observations, this work also provides an insight into the mechanisms of polyketide synthases in this filamentous fungus. IMPORTANCE The fungus Ustilago maydis represents one of the major threats for maize plants since it is responsible for corn smut disease, which generates considerable economical losses around the world. Therefore, contributing to a better understanding of the biochemistry of defense mechanisms used by U. maydis to protect itself against harsh environments, as the synthesis of melanin, could provide improved biological tools for tackling the problem and protect the crops. In addition, the fact that this fungus synthesizes melanin in an unconventional way, requiring more than one polyketide synthase for producing melanin precursors, gives a different perspective on the complexity of these multimodular enzymes and their evolution in the fungal kingdom.

Incompatibility between proliferation and plant invasion is mediated by a regulator of appressorium formation in the corn smut fungusUstilago maydis

Plant pathogenic fungi often developed specialized infection structures to breach the outer surface of a host plant. These structures, called appressoria, lead the invasion of the plant by the fungal hyphae. Studies in different phytopathogenic fungi showed that appressorium formation seems to be subordinated to the cell cycle. This subordination ensures the loading in the invading hypha of the correct genetic information to proceed with plant infection. However, how the cell cycle transmits its condition to the genetic program controlling appressorium formation and promoting the plant’s invasion is unknown. Our results have uncovered how this process occurs for the appressorium ofUstilago maydis, the agent responsible for corn smut disease. Here, we described that the complex Clb2-cyclin-dependent kinase (Cdk)1, one of the master regulators of G2/M cell cycle progression inU. maydis, interacts and controls the subcellular localization of Biz1, a transcriptional factor required for the activation of the appressorium formation. Besides, Biz1 can arrest the cell cycle by down-regulation of the gene encoding a second b-cyclin Clb1 also required for the G2/M transition. These results revealed a negative feedback loop between appressorium formation and cell cycle progression inU. maydis, which serves as a “toggle switch” to control the fungal decision between infecting the plant or proliferating out of the plant.

V.S. Gruzdev. Maize smut (Ustilago maydis), as a means of enhancing labor contractions of the uterus. (Doctor, 1894, No. 19)

The weakness of pushing, which is encountered so often in the practical activity of obstetricians and leading not very often to serious surgical intervention, which does not always end happily for the mother and the baby, made the more innocent people look for ways to fight it, than the operation. All currently available means of enhancing labor pains are not always effective, therefore it is completely understandable and desirable to search for new more effective means. The author, wanting to try any of the newly proposed remedies with the weakness of the labor pains, settled on a corn smut.

An Unconventional Melanin Biosynthetic Pathway in Ustilago maydis

ABSTRACTUstilago maydis is a phytopathogenic fungus responsible for corn smut disease. Although it is a very well established model organism for the study of plant-microbe interactions, its biosynthetic potential has not been totally explored. By analyzing U. maydis genome, we identified a biosynthetic gene cluster whose activation led to the production of a black melanin pigment. Single deletion mutants of the cluster genes revealed that five encoded enzymes are required for the accumulation of the black pigment, including three polyketide synthases (pks3, pks4 and pks5), a cytochrome P450 monooxygenase (cyp4) and a protein with similarity to versicolorin B-synthase (vbs1). Moreover, metabolic profiles of the mutants defective for pks3 and pks4 indicated that the products of these genes catalyze together the first step in the melanin biosynthetic pathway since none of the mutants accumulated any melanin or intermediate products. Mutants deleted for pks5 produced orsellinic acid (OA) and triacetic acid lactone (TAL), suggesting that both products are produced by Pks3 and Pks4. It might thus demonstrate that Pks5 plays a role in a reaction downstream of that catalyzed by Pks3 and Pks4. OA and TAL were also found in extracts of a cyp4 deletion mutant along with several heterodimers of TAL and Pks5-derived orsellinic aldehyde compounds. According to their phenotypes and the intermediate products isolated from these strains, Cyp4 and Vbs1 seem to be involved in reactions downstream of Pks5. Our findings suggest that U. maydis synthesizes a new melanin based on coumarin and pyran-2-one intermediates, while most fungal melanins are derived from 1,8-dihydroxynaphthalene (DHN) or L-3,4-dihydroxyphenylalanine (L-DOPA). Along with these observations, this work also provides an insight into the mechanisms of polyketide synthases in this filamentous fungus.IMPORTANCEUstilago maydis represents one of the major threats for maize plants since it is responsible for corn smut disease, which generates considerable economical losses around the world. Therefore, contributing to a better understanding of the biochemistry of defense mechanisms used by U. maydis to protect itself against harsh environments, as the synthesis of melanin, could provide improved biological tools for tackling the problem and protect the crops. In addition, the fact that this fungus synthesizes melanin in a very unique way, requiring more than one polyketide synthase for producing this secondary metabolite, gives a different perspective on the complexity of these multimodular enzymes and their evolution in the fungal kingdom.

Cycling in synchrony

The corn smut fungus uses two different mechanisms to control its cell cycle when it is infecting plants.

Ustilago maydis yeast stage found on imported sweet corn

During the inspection of imported sweet corn (Zea mays), a specimen with dry rot symptoms was detected by Ministry for Primary Industries quarantine officers. A sample was sent to the MPI Plant Health and Environment Laboratory for diagnostics, and initial examination found a layer of yeast cells on the surface of the symptomatic tissue. The fungus was cultured and identified based on DNA sequences as Ustilago maydis. While the corn-smut pathogen U. maydis is well known to cause tumour like galls on corn kernels, it is a less recognised fact that this fungus can also grow as a yeast. To determine if the yeast stage could have been associated with the dry-rot symptoms observed on the specimen, healthy material was inoculated with the isolated U. maydis strain. No symptoms developed on inoculated material, indicating that the yeast cells were likely multiplying on the surface of the cut corn ear as saprobes. To our knowledge, this is the first report of U. maydis yeast stage on corn ears and indicates a previously unconsidered pathway for the organism. For the yeast stage to cause disease, mating with a compatible mating type on the surface of a living host plant would be required.