Abstract:
A polyclonal antiserum was prepared against a purified microsomal
chitinase isolated from the fungus Choanephora cucurbitarum. Indirect immunofluorescence
was used to localize chitinase at various developmental stages of
five zygomycetous fungi and during abiotrophic mycoparasite interaction with a
susceptible and resistant host. This was compared to localization of oligomers
of N-acetylglucosamine with the lectin wheat germ agglutinin (WGA). Dotimmunoblot
and Western blot techniques revealed that the anti-serum reacted
strongly with the antigen from which it was derived. Cross reactivity of the
antiserum was found with WGA and another chitin binding lectin, Phyto/acca
americana agglutinin (PAA). Immuno-fluorescence results showed the direct
involvement of chitinase in spore swelling, germination, sporangium
development and response during mechanical injury. There appeared to be no
involvement of chitinase during apical hyphal growth or new branch initiation in
any of the fungi tested despite mild proteolysis and permeabilization of the cell
surface prior to labelling. Binding with WGA revealed similar patterns of
fluorescence to that of chitinase localization but differed by showing
fluorescence and therefore chitin localization at the apex and new branch
initiation when tested at different developmental stages. There was no
difference between chitinase localization and binding with WGA in a
susceptible host and resistant host challenged with the mycoparasite,
Piptocephalis virginiana. Differences in binding ability of antichitinase and lectin
WGA suggests that the latter is not a suitable indicator for indirect localization of
the lytic enzyme, chitinase.
