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Appl Environ Microbiol, January 1998, p. 325-332, Vol. 64, No. 1
0099-2240/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.

Electron and Fluorescence Microscopy of Extracellular Glucan and Aryl-Alcohol Oxidase during Wheat-Straw Degradation by Pleurotus eryngii

J. M. Barrasa,1,* A. Gutiérrez,2,dagger V. Escaso,1 F. Guillén,2 M. J. Martínez,2 and A. T. Martínez2

Departamento de Biología Vegetal, Universidad de Alcalá, E-28871 Alcalá de Henares, Madrid,1 and Centro de Investigaciones Biológicas, CSIC, E-28006 Madrid,2 Spain

Received 12 June 1997/Accepted 2 October 1997

The ligninolytic fungus Pleurotus eryngii grown in liquid medium secreted extracellular polysaccharide (87% glucose) and the H2O2-producing enzyme aryl-alcohol oxidase (AAO). The production of both was stimulated by wheat-straw. Polyclonal antibodies against purified AAO were obtained, and a complex of glucanase and colloidal gold was prepared. With these tools, the localization of AAO and extracellular glucan in mycelium from liquid medium and straw degraded under solid-state fermentation conditions was investigated by transmission electron microscopy (TEM) and fluorescence microscopy. These studies revealed that P. eryngii produces a hyphal sheath consisting of a thin glucan layer. This sheath appeared to be involved in both mycelial adhesion to the straw cell wall during degradation and AAO immobilization on hyphal surfaces, with the latter evidenced by double labeling. AAO distribution during differential degradation of straw tissues was observed by immunofluorescence microscopy. Finally, TEM immunogold studies confirmed that AAO penetrates the plant cell wall during P. eryngii degradation of wheat straw.

* Corresponding author. Mailing address: Departamento de Biología Vegetal, Universidad de Alcalá, E-28871 Alcalá de Henares, Madrid, Spain. Phone: 341 8854943. Fax: 341 8855066. E-mail: bvjmbg{at}bioveg.alcala.es.

dagger Present address: Instituto Recursos Naturales y Agrobiología, CSIC, E-41080 Seville, Spain.




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