Limited vegetative compatibility as a cause of somatic recombination in Trichoderma pseudokoningii

With the aim of a better characterization of the somatic recombination process in Trichoderma pseudokoningii, a progeny from crossings between T. pseudokoningii strains contrasting for auxotroph markers was characterized by RAPD markers and PFGE (electrophoretic karyotype). Cytological studies of the conidia, conidiogenesis and heterokaryotic colonies were also performed. The genotypes of the majority of the recombinant strains analyzed were similar to only one of the parental strains and the low frequency of polymorphic RAPD bands suggested that the nuclear fusions may not occur into the heterokaryon. In some heterokaryotic regions the existence of intensely staining hyphae might be related to cell death. We proposed that a mechanism of somatic recombination other than parasexuality might occur, being related to limited vegetative compatibility after postfusion events, as described for other Trichoderma species.

Pareamento de fungos cultivados por diferentes espécies de formigas cortadeiras no Rio Grande do Sul / Fusion of fungi cultivated by different species of cutter ants in Rio Grande do Sul, Brazil

Este trabalho teve por objetivo estudar a possível ocorrência de heterocariose por meio do pareamento de fungos provenientes de diferentes espécies de formigas cortadeiras coletados em diferentes regiões do Estado do Rio Grande do Sul, utilizando a técnica AFLP. Dez grupos de pareamentos foram realizados, sendo que em dois não ocorreu a fusão das colônias. Após ocorrerem os pareamentos, foram coletados discos de micélio da área de intersecção das colônias e das áreas extremas de pareamento, para a realização da extração do DNA e padronização. Para as reações de PCR, utilizaram-se 21 combinações de primers AFLP, dos quais foram selecionados três que apresentaram maior polimorfismo e consistência. Os dados moleculares foram transformados em matriz binária e analisados pelo programa estatístico NTSYS v. 2.1. As três combinações de primers utilizadas geraram 245 bandas polimórficas. A similaridade média dos genótipos variou de 0,03 a 0,18 em todos os pareamentos. A grande distância genética encontrada entre os organismos possivelmente ocorreu por três fatores: a possível ocorrência de heterocariose entre os fungos, a utilização de fungos oriundos de diferentes espécies de formigas e de diferentes regiões. O trabalho sugere que a heterocariose pode representar um importante papel no ciclo de vida deste fungo, uma vez que sua multiplicação é clonal e há ausência de frutificação.

This research work had as objective to study the possible heterokaryosis occurrence through the fusion of mushrooms originating from nests of different species of cutter ants collected in different areas of the State of Rio Grande do Sul, using the AFLP technique. Ten fusion groups were accomplished, from which two did not perform a colony coalition. After the fusion occurred, disks of mycelium of the colonies intersection area and fusion external areas were collected, the extraction of DNA and standardization was performed. PCR reactions were tested using 21 AFLP primer combinations, from which three were selected because they presented larger polymorphism and consistence. The molecular data were transformed at binary head office and analyzed by the statistical program NTSYS v. 2.1. The three combinations of primers generated 245 polymorphic bands. The average similarity of the genotypes varied from 0.03 to 0.18 in all the fusions. The large genetic distance found among the organisms possibly happened due to three factors: the possible heterokaryosis occurrence among the mushrooms; the use of mushrooms originating from different species of ants and; from different areas. This work suggests that the heterokaryosis can represent an important role in the life cycle of this mushroom, since it multiplies by clonal propagation and due to the absence of fructification.

Vegetative compatibility and molecular characterization of Fusarium graminearum isolates from the State of Paraná, Brazil

Fusarium graminearum isolates causing Fusarium head blight in wheat were collected in Brazil and analyzed by random amplified polymorphic DNA (RAPD) markers and vegetative compatibility grouping (VCG). Nitrate non-utilizing mutants (nit) from each isolate were paired to verify heterokaryon formation. Three VCGs were identified among F. graminearum isolates: VCG1 included F-2, F-3 and F-4 isolates; VCG2 included F-1, F-6 and F-9 isolates; VCG3 included F-5, F-7 and F-8 isolates. Based on PCR amplification with eight different primers, the isolates showed great genetic similarity among themselves. Dendrogram analysis demonstrated two RAPD groups: Group A, consisting of isolates F-2 and F-9, and Group B, composed of the remaining isolates. Results suggest the clonal origin of F. graminearum isolates.

Isolados de Fusarium graminearum, obtidos de espigas de trigo com sintomas de Giberela, foram analisados pela técnica do Polimorfismo de DNA Amplificado ao Acaso (RAPD) e pelos Grupos de Compatibilidade Vegetativa (GCV). Mutantes auxotróficos (nit) de cada isolado foram pareados em todas as combinações possíveis, para a formação de heterocários. Três GCVs foram identificados: GCV1, incluindo os isolados F-2, F-3 e F-4; GCV2, incluindo os isolados F-1, F-6 e F-9; e GCV3, formado pelos isolados F-5, F-7 e F-8. Dois grupos foram identificados com base nos marcadores de RAPD: o grupo A, formado pelos isolados F-2 e F-9, e o grupo B, composto pelos demais isolados, os quais apresentaram grande similaridade entre si. Os resultados sugerem a origem clonal dos isolados de F. graminearum analisados.

SELECTION AND IDENTIFICATION OF NITRATE NON-UTILIZING MUTANTS OF COLLECTOTRICHUM GLOEOSPORIOIDES / 微生物学通报

Eight isolates of Colletotrichum gloeosporioides isolated from host plants Cunninghamia lanceolata lanceolata and Euonymus japonichum, respectively, were cultured on MMC medium containing KCIO3 to select chlorate-resistant and nitrate non-utilizing mutants (Nit). All the Nit mutants obtained by this way belong to one of 3 kinds of the following phenotypes: the nitrate reductase structural locus (nit1), the nitrate-assimilation pathway-specific regulatory locus(nit3), and the molybdenum-containing cofactor locus(nitM). The higher mutation frequency on MMC amended with increasing concentration of KCIO3 was induced, and various nitrogen sources were able to influence production of the phenotypic classes. Seven of the 8 isolates tested were self-compatibility in which the two mutants with different phenotypes from the same isolate could genetically complement. The two isolates frail C. lanceolata belong to the same vegetative compatibility group (VCG), the other six isolates belong to distinctive VCGs.