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Title: LOCALIZATION OF GLUCOSE OXIDASE IN THE BIOCONTROL FUNGUS TALAROMYCES FLAVUSWITH IMMUNOCTYOCHEMISTRY

Author
item STOSZ, SARAH - FORMER BPDL, ARS
item ROY, STEPHAN - FORMER NL, ARS
item Murphy, Charles - Charlie
item Wergin, William
item Fravel, Deborah

Submitted to: Phytopathology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 3/17/1998
Publication Date: N/A
Citation: N/A

Interpretive Summary: The soil-inhabiting plant pathogen Verticillium dahliae causes a wilt disease in many vegetable, ornamental, field, and tree crops resulting in severe economic losses. The beneficial fungus Talaromyces flavus kills the overwintering structures of the pathogen by producing the enzyme glucose oxidase. The action of glucose oxidase produces hydrogen peroxide which kills the pathogen. This work was undertaken to find where glucose oxidas is in and around cells of the the beneficial fungus. An antibody specific for the enzyme was made and linked to submicroscopic gold particles. The antibody was incubated with T. flavus and then examined with a transmission electron microscope (TEM). The gold particles appear black in TEM. Glucose oxidase was found in both young and old cells, although the amount decreased significantly with age. Active transport of the enzyme from young cells into the surrounding medium was observed. The enzyme was also slowly released into the medium as cells aged. The enzyme was extremely stable and retained activity after storage for two weeks at room temperature. Understanding the roles that the enzyme associated with the cell, as well as that away from the cell, will help to optimize production of the enzyme to improve biocontrol. This information will be used by scientists developing practical biocontrol measures for use against Verticillium wilt.

Technical Abstract: Production of the enzyme glucose oxidase has been shown to be involved in the biological control of Verticillium wilt by Talaromyces flavus. In this study, the location of glucose oxidase was determined in T. flavus by immunocytochemistry using glucose oxidase-specific polyclonal antibody. Immunostaining revealed the glucose oxidase was both intracellular and extracellular. Old as well as young hyphal cells contained glucose oxidase, but labeling of the cell-wall associated enzyme decreased as cells aged. Glucose oxidase was released from cells through exocytosis and not by cell lysis. Enzyme stability studies confirmed that the glucose oxidase of T. flavus is an extremely stable enzyme, retaining 13% of its original activity after two weeks at 25 deg C and retaining activity for several days at temperatures up to 50 deg C.