We have studied the efficiency of analytical procedures for two phenolic compounds found in Eucalyptus globulus and Quercus robur L.: gallic and ellagic acid. Isochratic HPLC was used s a mean of quantitative analysis. Eluants for HPLC analysis were H 2 O:CH 3 OH:H3PO4 (975,5:19,5:1, v/v/v) for gallic and (449,5:449,5:1, v/v/v) for ellagic acid. The time required for each HPLC analysis was 10 and 6 min. The recuperation of gallic acid and ellagic acid was respectively 83,0% and 58,6%. We have showed that high temperature Quercus robur L. kiln drying resulted in an important augmentation in gallic and egallic acid proportions.
After long periods of storage, plant pathogen isolates lose their sporulation capacity. The objective of this study was to evaluate re-isolation methodologies for recovering sporulation of Pestalotiopsis grandis-urophylla isolates after subjection to a long period of storage. Isolates of P. grandis-urophylla were kept for 14 months on Petri dishes with PDA medium at 10°C. After this period, the isolate colonies showed reduced mycelial growth and no sporulation. The isolates were inoculated on healthy Eucalyptus grandis-urophylla leaves, and after ten days they were subjected to three re‑isolation methods: scraping of the lesions (S) removing of injured plant tissue fragments, followed by disinfestation (D) and without disinfestation (WD). Then, the purified isolates were evaluated for the recovery of its sporulation ability. The different methods for re-isolation resulted in the occurrence of differences among the isolates, showing that sporulation is an isolate-dependent feature. The three methods (S, WD and D) allowed the sporulation recovery of P. grandis-urophylla, even after these isolates have been subjected to 14 months.
Species of Pestalotiopsis have been reported to be pathogenic to eucalypt, yet few studies have addressed their real pathogenic potential or even their diagnosis. The objective of this study was to carry out the molecular, micromorphological, physiological and pathogenic characterization of four isolates of Pestalotiopsis sp. found in eucalypt leaf spots. DNA from the isolates was extracted and PCR amplified using primers for the internal transcribed spacer (ITS), partial β-tubulin (TUB) and translation elongation factor 1-alpha (EF1-α) gene regions. For morphophysiological characterization, the fungal structures were measured and isolates evaluated for mycelial growth and sporulation under different light regimes (0, 12, and 24 h). Pathogenicity tests were conducted on healthy eucalypt leaves. The results revealed that (a) the amplified ITS region is too conserved to be used for identification of Pestalotiopsis species, and thus, TUB and EF1-α sequences are recommended for this purpose; (b) based on micromorphological characteristics and DNA sequences, the four isolates were identified as the new species Pestalotiopsis grandis-urophylla; (c) P. grandis-urophylla presents faster mycelial growth when cultivated in the dark, but for mass production of inoculum the light regime does not have a strong influence; and (d) the pathogenic potential varied among the P. grandisurophylla isolates.
A mancha de alternaria, cujo agente causal é o fungo, Alternaria spp., encontra-se distribuída pelas áreas de plantio de hortaliças no Brasil onde frequentemente a doença é veiculada por sementes contaminadas. Neste sentido, a qualidade sanitária e fisiológica de sementes é fundamental para o estabelecimento uniforme das culturas em campo, sendo um dos fatores que garantem o sucesso da produção da olerícola. O objetivo deste trabalho foi caracterizar o fungo Alternaria associado a sementes de alface americana cv. ‘Astra’. A análise de incidência foi realizada pelo método blotter test, empregando-se o delineamento inteiramente casualizado (DIC), com quatro repetições (50 sementes gerbox-1). Após sete dias a 20 ºC (± 2 ºC) e foto período de 12 h, a avaliação foi realizada examinando-se individualmente todas as sementes em microscópio estereoscópio para a confecção de lâminas semipermanentes, a qual foi realizada mediante a remoção de micélio e estruturas fúngicas encontradas sobre as sementes infectadas, para caracterização e identificação da espécie do fungo incidente. Os conídios encontrados apresentaram dimensões de 13,7 - 25,7 x 6,2 - 12,4 μm (18,6 x 8,8 μm), formato de pera invertida, com coloração marrom clara e, cujos bicos apresentaram dimensões de 2,2 - 11,0 μm (4,7 μm). Estas características discriminaram o fungo como A. alternata.
Eucalyptus species are among the most important forest crops in the world and can be affected by several pathogens, mainly by fungi of the genus Pestalotiopsis, which cause leaf spots. Studies aimed at the pathogenic characterization of Pestalotiopsis spp. from lesions of eucalyptus leaves in Brazil are still limited. The objective of this work was to evaluate the pathogenic potential of Pestalotiopsis grandis-urophylla isolates. For that, healthy leaves of adult Eucalyptus grandis ‘GG 100’ plants were inoculated with mycelial suspension of different P. grandis-urophylla isolates. The leaves inoculated with the pathogen were submitted to controlled conditions in a humid chamber in transparent acrylic gerbox boxes. Disease severity assessments were performed at 4, 6, 8 and 10 days after inoculation. Isolate E 72-04 had the highest area under the disease progress curve (AUDPC). Regarding the development of lesions, isolates E-72-02 and E-72-03 fit the polynomial model of the second degree, while isolate E-72-04 fit a linear model. The methodology tested reproduced typical symptoms of Pestalotiopsis and can be used as a parameter for new pathogenicity tests with this fungal genus.
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