RESUMO
The promotion of sugarcane growth by the endophytic Pantoea agglomerans strain 33.1 was studied under gnotobiotic and greenhouse conditions. The green fluorescent protein (GFP)-tagged strain P. agglomerans 33.1::pNKGFP was monitored in vitro in sugarcane plants by microscopy, reisolation, and quantitative PCR (qPCR). Using qPCR and reisolation 4 and 15 days after inoculation, we observed that GFP-tagged strains reached similar density levels both in the rhizosphere and inside the roots and aerial plant tissues. Microscopic analysis was performed at 5, 10, and 18 days after inoculation. Under greenhouse conditions, P. agglomerans 33.1-inoculated sugarcane plants presented more dry mass 30 days after inoculation. Cross-colonization was confirmed by reisolation of the GFP-tagged strain. These data demonstrate that 33.1::pNKGFP is a superior colonizer of sugarcane due to its ability to colonize a number of different plant parts. The growth promotion observed in colonized plants may be related to the ability of P. agglomerans 33.1 to synthesize indoleacetic acid and solubilize phosphate. Additionally, this strain may trigger chitinase and cellulase production by plant roots, suggesting the induction of a plant defense system. However, levels of indigenous bacterial colonization did not vary between inoculated and noninoculated sugarcane plants under greenhouse conditions, suggesting that the presence of P. agglomerans 33.1 has no effect on these communities. In this study, different techniques were used to monitor 33.1::pNKGFP during sugarcane cross-colonization, and our results suggested that this plant growth promoter could be used with other crops. The interaction between sugarcane and P. agglomerans 33.1 has important benefits that promote the plant's growth and fitness.
Assuntos
Endófitos/crescimento & desenvolvimento , Pantoea/crescimento & desenvolvimento , Saccharum/crescimento & desenvolvimento , Saccharum/microbiologia , Biofilmes , Celulase/biossíntese , Quitinases/biossíntese , Endófitos/metabolismo , Proteínas de Fluorescência Verde , Ácidos Indolacéticos/metabolismo , Pantoea/genética , Pantoea/isolamento & purificação , Pantoea/metabolismo , Fosfatos/metabolismo , Raízes de Plantas/microbiologia , Rizosfera , Saccharum/metabolismoRESUMO
The herbicide propanil has long been used in rice production in southern Brazil. Bacteria isolated from contaminated soils in Massaranduba, Santa Catarina, Brazil, were found to be able to grow in the presence of propanil, using this compound as a carbon source. Thirty strains were identified as Pseudomonas (86.7%), Serratia (10.0%), and Acinetobacter (3.3%), based on phylogenetic analysis of 16S rDNA. Little genetic diversity was found within species, more than 95% homology, suggesting that there is selective pressure to metabolize propanil in the microbial community. Two strains of Pseudomonas (AF7 and AF1) were selected in bioreactor containing chemotactic growth medium, with the highest degradation activity of propanil exhibited by strain AF7, followed by AF1 (60 and 40%, respectively). These strains when encapsulated in alginate exhibited a high survival rate and were able to colonize the rice root surfaces. Inoculation with Pseudomonas strains AF7 and AF1 significantly improved the plant height of rice. Most of the Pseudomonas strains produced indoleacetic acid, soluble mineral phosphate, and fixed nitrogen. These bacterial strains could potentially be used for the bioremediation of propanil-contaminated soils and the promotion of plant growth.
Assuntos
Bactérias/genética , Bactérias/isolamento & purificação , Variação Genética , Oryza/crescimento & desenvolvimento , Oryza/microbiologia , Propanil/metabolismo , Rizosfera , Alginatos , Bactérias/metabolismo , Bactérias/ultraestrutura , Sequência de Bases , Biodegradação Ambiental/efeitos dos fármacos , Células Imobilizadas/citologia , Células Imobilizadas/efeitos dos fármacos , Células Imobilizadas/ultraestrutura , DNA Ribossômico/genética , Ácido Glucurônico , Ácidos Hexurônicos , Microesferas , Oryza/efeitos dos fármacos , Filogenia , Propanil/farmacologia , Pseudomonas/efeitos dos fármacos , Pseudomonas/genética , Pseudomonas/crescimento & desenvolvimento , Pseudomonas/ultraestrutura , Transformação Genética/efeitos dos fármacosRESUMO
AIMS: Biological sources for the control of plant pathogenic fungi remain an important objective for sustainable agricultural practices. Actinomycetes are used extensively in the pharmaceutical industry and agriculture owing to their great diversity in enzyme production. In the present study, therefore, we evaluated chitinase production by endophytic actinomycetes and the potential of this for control of phytopathogenic fungi. METHODS AND RESULTS: Endophytic Streptomyces were grown on minimum medium supplemented with chitin, and chitinase production was quantified. The strains were screened for any activity towards phytopathogenic fungi and oomycetes by a dual-culture in vitro assay. The correlation between chitinase production and pathogen inhibition was calculated and further confirmed on Colletotrichum sublineolum cell walls by scanning electron microscopy. CONCLUSIONS: This paper reports a genetic correlation between chitinase production and the biocontrol potential of endophytic actinomycetes in an antagonistic interaction with different phytopathogens, suggesting that this control could occur inside the host plant. SIGNIFICANCE AND IMPACT OF THE STUDY: A genetic correlation between chitinase production and pathogen inhibition was demonstrated. Our results provide an enhanced understanding of endophytic Streptomyces and its potential as a biocontrol agent. The implications and applications of these data for biocontrol are discussed.
Assuntos
Antibiose , Proteínas de Bactérias/metabolismo , Quitinases/metabolismo , Fungos/fisiologia , Doenças das Plantas/microbiologia , Plantas/microbiologia , Streptomyces/fisiologia , Proteínas de Bactérias/genética , Parede Celular/metabolismo , Quitina/metabolismo , Quitinases/genética , Streptomyces/enzimologia , Streptomyces/genéticaRESUMO
The transcripts of structurally related cellobiohydrolase genes in Phanerochaete chrysosporium-colonized wood chips were quantified. The transcript patterns obtained were dramatically different from the transcript patterns obtained previously in defined media. Cellobiose dehydrogenase transcripts were also detected, which is consistent with the hypothesis that such transcripts play an important role in cellulose degradation.
Assuntos
Basidiomycota/enzimologia , Desidrogenases de Carboidrato/genética , Celulase/genética , RNA Mensageiro/análise , Madeira , Sequência de Bases , Celulose/metabolismo , Celulose 1,4-beta-Celobiosidase , Dados de Sequência MolecularRESUMO
Mitotically unstable Aspergillus nidulans argB+ transformants obtained by the biolistic process were studied in the present work. Hybridization signals from undigested DNA and pulsed-field chromosomal bands of the transformants suggested the introduced plasmid occurred as free concatenated molecules. Fifteen vigorous growth sectors released from the transformants were analysed in order to understand the mechanisms involved in their formation. All sectors showed the integration of exogenous genes into the fungal genome by homologous or heterologous recombinant events.
Assuntos
Aspergillus nidulans/genética , Biolística , Transformação Genética , Arginina/metabolismo , DNA Fúngico/genética , Eletroforese em Gel de Campo Pulsado , Genes Fúngicos , Cariotipagem , Mitose , Recombinação GenéticaRESUMO
Amplicons of SSU and ITSI + 2rDNA of 13 strains of Fonsecaea pedrosoi and three strains of F. compacta were digested with seven restriction enzymes. In addition, the ITS1 region of strains was sequenced. With both methods significant variation was found which, however, did not coincide with established species limits based on morphology
Assuntos
Humanos , DNA , Cromoblastomicose/fisiopatologia , Cromoblastomicose/genética , Cromoblastomicose/imunologia , Polimorfismo de Fragmento de Restrição/genética , Polimorfismo de Fragmento de Restrição/imunologiaRESUMO
In order to obtain yeast strains with important characteristics for the wine industry, protoplast fusions were performed. Attention was driven to two important traits: flocculation and of H2S production. Because both are traits occuring at low frequency in wine yeast (1% each), their natural occurence is unlike. Crossing was made through protoplast fusion because strains were sexualy incompatible. Initially, auxotrophic contrasting mutants were obtained, their marks were tested for reversion and finally, their growth rates were determined. Protoplasts were obtained after treatment with Novozym 234 PEG and CaCl2. The rate of fusion was 2,7x10-4, and 54,6% of the resulting colonies were able to grow on minimal media. After several transfers to YEPD media, the flocculant and H2S- productive strains were selected.
Com o objetivo de obter linhagens de leveduras com características de importância para a indústria vinícola, empregou-se a técnica da fusão de protoplastos. A atenção foi dirigida a duas características: floculação e não produção de H2S. Uma vez que cada característica é de baixa frequência (1% cada) em leveduras vinícolas, torna-se pouco provável a ocorrência natural de uma linhagem com ambas as características. O cruzamento foi realizado via fusão de protoplastos pois as linhagens apresentaram incompatibilidade sexual. Inicialmente foram obtidos os mutantes auxotróficos contrastantes e posteriormente suas marcas foram avaliadas quanto à reversão e, em seguida foram determinadas suas curvas de crescimento. Obteve-se a protoplastização por tratamento com Novozym 234 e a fusão foi mediada por PEG 40% e CaCl2 1,2 M. A taxa de fusão de protoplastos foi de 2,7x10-4, sendo que das colônias resultantes, 54,6% continuaram o crescimento após transferência para MM, "minimal media". Após várias transferências em meio YEPD foram selecionadas as linhagens floculantes e H2S-.
RESUMO
In order to obtain yeast strains with important characteristics for the wine industry, protoplast fusions were performed. Attention was driven to two important traits: flocculation and of H2S production. Because both are traits occuring at low frequency in wine yeast (1% each), their natural occurence is unlike. Crossing was made through protoplast fusion because strains were sexualy incompatible. Initially, auxotrophic contrasting mutants were obtained, their marks were tested for reversion and finally, their growth rates were determined. Protoplasts were obtained after treatment with Novozym 234 PEG and CaCl2. The rate of fusion was 2,7x10-4, and 54,6% of the resulting colonies were able to grow on minimal media. After several transfers to YEPD media, the flocculant and H2S- productive strains were selected.
Com o objetivo de obter linhagens de leveduras com características de importância para a indústria vinícola, empregou-se a técnica da fusão de protoplastos. A atenção foi dirigida a duas características: floculação e não produção de H2S. Uma vez que cada característica é de baixa frequência (1% cada) em leveduras vinícolas, torna-se pouco provável a ocorrência natural de uma linhagem com ambas as características. O cruzamento foi realizado via fusão de protoplastos pois as linhagens apresentaram incompatibilidade sexual. Inicialmente foram obtidos os mutantes auxotróficos contrastantes e posteriormente suas marcas foram avaliadas quanto à reversão e, em seguida foram determinadas suas curvas de crescimento. Obteve-se a protoplastização por tratamento com Novozym 234 e a fusão foi mediada por PEG 40% e CaCl2 1,2 M. A taxa de fusão de protoplastos foi de 2,7x10-4, sendo que das colônias resultantes, 54,6% continuaram o crescimento após transferência para MM, "minimal media". Após várias transferências em meio YEPD foram selecionadas as linhagens floculantes e H2S-.
RESUMO
This paper describes transformation of intact conidia of Aspergillus nidulans, auxotrophic for arginine, by using the biolistic process. The plasmid employed was pFB39, carrying the argB gene. The transformation frequency obtained was 81 transformants/microgram of DNA. Classical genetics and molecular analysis were conducted to analyse transformants and to determine in which chromosome integration took place.
Assuntos
Aspergillus nidulans/genética , Genes Fúngicos , Plasmídeos , Transformação Genética , Arginina/metabolismo , Aspergillus nidulans/fisiologia , Cromossomos Fúngicos , DNA Fúngico/isolamento & purificação , Técnicas de Transferência de Genes , Técnicas Genéticas , CariotipagemRESUMO
Aflatoxin production by strains recently isolated of Aspergillus flavus was studied, after different storage times understand the mechanisms of possible variations in aflatoxin production. Three A. flavus aflatoxin producing strains were utilized, classified as: a) high producer; b) medium producer and c) low producer. The experiment lasted 280 days and the moulds were studied by two preservation methods: oil covered slants on Czapeck's medium and periodic transfer on Czapeck's modified medium. Quantification of the aflatoxin produced was made at 30 day intervals, on thin-layer chromatography and visual determination by the dilution-to-extinction technique. The production of aflatoxin by all strains varied but they did not lose their initial characteristics. Microscopic examinations revealed thickened zones and hifal enlargements over some globous structures that may be related to aflatoxin production sites.
O presente trabalho fui realizado com a finalidade de se estudar a produção de aflatoxinas por linhagens de A. flavus, recém isoladas, em diferentes tempos de manutenção, a fim de contribuir para um melhor entendimento do mecanismo de variação na produção de aflatoxinas. Para isso, foram utilizadas três linhagens de A. flavus produtoras de aflatoxinas, classificadas como: a) grande produtora; b) média produtora e c) baixa produtora. Neste experimento, que se estendeu por 280 dias, os fungos foram estudados em dois métodos de preservação: mantido em óleo mineral, no meio Czapeck, e repicado periodicamente em meio Czapeck modificado. A análise da produção de aflatoxinas foi efetuada de 30 em 30 dias. A quantificação da toxina foi feita por cromatografía em camada delgada, pela técnica de avaliação visual, de diluição até extinção. Foi constatada uma variação na produção de toxina em todas as linhagens, contudo elas não perderam suas características originais.
RESUMO
Aflatoxin production by strains recently isolated of Aspergillus flavus was studied, after different storage times understand the mechanisms of possible variations in aflatoxin production. Three A. flavus aflatoxin producing strains were utilized, classified as: a) high producer; b) medium producer and c) low producer. The experiment lasted 280 days and the moulds were studied by two preservation methods: oil covered slants on Czapeck's medium and periodic transfer on Czapeck's modified medium. Quantification of the aflatoxin produced was made at 30 day intervals, on thin-layer chromatography and visual determination by the dilution-to-extinction technique. The production of aflatoxin by all strains varied but they did not lose their initial characteristics. Microscopic examinations revealed thickened zones and hifal enlargements over some globous structures that may be related to aflatoxin production sites.
O presente trabalho fui realizado com a finalidade de se estudar a produção de aflatoxinas por linhagens de A. flavus, recém isoladas, em diferentes tempos de manutenção, a fim de contribuir para um melhor entendimento do mecanismo de variação na produção de aflatoxinas. Para isso, foram utilizadas três linhagens de A. flavus produtoras de aflatoxinas, classificadas como: a) grande produtora; b) média produtora e c) baixa produtora. Neste experimento, que se estendeu por 280 dias, os fungos foram estudados em dois métodos de preservação: mantido em óleo mineral, no meio Czapeck, e repicado periodicamente em meio Czapeck modificado. A análise da produção de aflatoxinas foi efetuada de 30 em 30 dias. A quantificação da toxina foi feita por cromatografía em camada delgada, pela técnica de avaliação visual, de diluição até extinção. Foi constatada uma variação na produção de toxina em todas as linhagens, contudo elas não perderam suas características originais.
RESUMO
Double auxotrophic and morphological mutants of Trichoderma pseudokoningii Rifai were fused by anastomosis and by protoplast fusion. The recovery of recombinants from heterokaryons on different selective media and from heterokaryotic colonies indicated the occurrence of parasexual events. Prototrophic colonies growing on minimal medium produced binucleate spores, green in colour, revealing a non-autonomous system for conidial pigmentation. Recombinants were obtained from these dikaryotic colonies suggesting the occurrence of a highly unstable diploid phase.
Assuntos
Hibridização Genética , Protoplastos/fisiologia , Trichoderma/genética , Celulase/metabolismo , Protoplastos/enzimologia , Recombinação Genética , Especificidade da Espécie , Trichoderma/enzimologia , Trichoderma/crescimento & desenvolvimentoRESUMO
Resumo: O objetivo desse estudo foi descrever aspectos do comportamento nuclear durante a divisäo miótica em conídios e células miceliais do fungo celulolítico Humicola so. A sequência dos eventos mitóticos foi similar aquela observada em outros fungos filamentosos. O número de cromossomos observados no final da metáfase näo pode ser determinado com segurança, mas pareceu estar entre 6 e 8. As culturas de Humicola sp frequentemente mostraram anastomose de hifas e a presença de núcleos na ponte hifal, o que permitiria a ocorrência de heterocariose,e indica que o ciclo parasexual pode ocorrer nessa espécie (au)