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Stachybotrys elegans is a mycoparasite of the soilborne plant pathogen fungus Rhizoctonia solani. The mycoparasitic activity of S. elegans is correlated with the production of cell wall-degrading enzymes such as chitinases. This study details the cloning and characterization of the cDNA, sechi44, that encodes an extracellular endochitinase. The expression regulation of sechi44 was altered when S. elegans was in interaction with its host, R. solani, and also when the mycoparasite was grown on minimal media amended with different carbon and nitrogen sources. Direct contact with R. solani significantly upregulated sechi44 expression which followed a cyclical pattern suggesting that this gene has a role not only in mycoparasitism, but also in linear growth of the mycoparasite. The addition of high concentrations of glucose and ammonium triggered a decrease of sechi44 expression suggesting that sechi44 is subject to glucose and ammonium repression. In a separate study, several genes (1016 clones) whose transcription was substantially up-regulated during the mycoparasitic interaction were identified using SSH and microarray analysis. Twenty-five percent (261 clones) of these were sequenced and assigned to putative functions. Among them, 15 expressed sequence tags (ESTs) were identified in R. solani whose functions were related to defense while the majority of ESTs were identified in S. elegans and assigned functions related to toxin metabolism, pathogenic process, stress response., multidrug resistance, apoptosis, transport, ATP synthesis, replication, transcription and DNA repair, translation, transduction, protein degradation, and ribosomal protein. The overexpression of 13 selected genes of S. elegans was validated and confirmed using quantitative reverse transcription polymerase chain reaction (QRT-PCR). The temporal gene expression of nine genes was monitored when the mycoparasite was grown on R. solani (host) and Sclerotinia sclerotiorum (non-host) mycelia and sclerotia. Some genes such as seglu, selec, and se151 were completely inhibited by the presence of non-host hyphae suggesting that these genes play an important role during mycoparasitism. Also, the absence of these corresponding transcripts suggests that the non-host produces transcription inhibitors. As expected, gene expression of cytochrome P450 was highly up-regulated early after germination of S. elegans conidia. This is in agreement with our finding in the EST data mining study, in which a role in toxin production was assigned to cytochrome P450.

The mycoparasite Stachybotrys elegans represents a potential biocontrol agent of Rhizoctonia solani, the causal organism of potato stem canker. The differential expression of two mycoparasitism-induced genes was monitored in S. elegans during interactions with its host, Rhizoctonia solani, and a non-host, Sclerotinia sclerotiorum. Using real-time reverse-transcription polymerase chain reaction (QRT-PCR), comparative analyses demonstrated that hyphal and sclerotial forms triggered different patterns of gene expression in the mycoparasite, as did the presence of the host or non-host. The calmodulin gene did not appear to be involved in conidial germination or appressoria formation of S. elegans. Potential roles of calmodulin during mycoparasitism are suggested, but further studies are required. The expression of the endochitinase-encoding gene, sechi44, was susbstantial only at later stages of interaction with living host sclerotia. Host defense mechanisms probably play a role in regulating sechi44 expression. Knowledge of the genetic mechanisms underlying this mycoparasitic relationship will further our knowledge on the potential use of S. elegans in biocontrol strategies. / Le mycoparasite Stachybotrys elegans est un agent potentiel de lutte biologiquepour le contrôle de Rhizoctonia solani, un phytopathogène causant le chancre dela tige et des stolons chez la pomme de terre (Solanum tuberosum). L'expressionde deux gènes induits pendant le mycoparasitisme a été étudiée chez S. elegans,alors que le mycoparasite était en interaction avec son hôte, R. solani, et un nonhôte,Sclerotinia sclerotiorum en utilisant la PCR quantitative en temps réel. Desanalyses comparatives ont démontré que les différentes formes d'hyphes et desc1érotes, ainsi que la présence de l'hôte ou du non-hôte, induisent différentspatrons d'expression. Le gène codant pour la calmodulin (calmodulin) ne semblepas être impliqué dans la germination des conidia ou dans la formation desappressoria chez S. elegans. Des rôles possibles de calmodulin sont suggérés,mais des études plus poussées demeurent nécessaires. L'expression du gènesechi44, codant pour une endochitinase, est importante dans des stades plusavancés du mycoparasitisme sur les sc1érotes vivants de 1 'hôte. Des mécanismesde défense de l'hôte jouent probablement un rôle important dans la régulation del'expression de sechi44. Une meilleure connaissance de la régulation génétiquelors du mycoparasitisme pourrait nous aider à évaluer le potentiel de S. elegansdans des stratégies de biocontrôle.

No description available.

Stachybotrys elegans (Pidopl.) W. Gams is a destructive mycoparasite of the soilborne plant pathogen Rhizoctonia solani. It colonizes effectively all types of cells of R. solani, and is considered as an effective biological control agent (BCA). Monitoring the presence of this mycoparasite in the field trials requires the development of a reliable and sensitive diagnostic assay that is able to detect and differentiate the BCA from their target host. To achieve this, designed SCAR (sequenced characterized amplified regions) primers designated as SE-13F and SE-13R were generated from informative RAPD markers. They were tested in conventional PCR assays alone or in conjunction with the recently developed SCAR primers (SBU-177/336) designed for Rhizoctonia solani (Kuhn) on several types of DNA. These included DNA extracted from pure cultures, co-cultures of the BCA and the pathogen, plant tissue and several types of soils inoculated with both the BCA and the pathogen. Irrespective of the type of the biological samples from which the DNA was extracted, the primers SE-13F/SE-13R successfully amplified only S. elegans. No cross-reaction was observed when the primers were used to amplify DNA of other fungi, bacteria and plant tissues. Likewise, the primer pair SBU-177/336 detected only its target organism, i.e., R. solani. The detection limit using these primers on amplified DNA was as little as 1 pg DNA extracted from pure cultures of S. elegans, 100 pg DNA extracted from greenhouse soil and 33 pg DNA extracted from natural soil. This work is the first report on the development of SCAR markers for the BCA, S. elegans. These molecular markers offer not only an alternative diagnostic assay to conventional detection methods, but also the possibility of being used in ecological studies.

No description available.