Immunoelectron microscopy using the anti-pectin monoclonal
antibody JIM 7 confirmed earlier observations that
pectin degradation is a primary event in the process of host
cell wall breakdown during the development of
chocolate spot disease (causal agent: Botrytis fabae (Sard.))
of
broad bean. Close examination of infected and non-infected Vicia faba
L.
leaves indicated a loss of JIM 7-labelling, and therefore, methyl-esterified
pectin, from
swollen walls of infected and contiguous epidermal cells. Modified
mesophyll walls also possessed less methyl-esterified pectin
than healthy walls. Enzymes which attack methyl-esterified
pectin appeared to be most active in
regions of host tissue close to sites of fungal infection.
Ultrastructural studies using the enzyme, cellobiohydrolase conjugated
to gold (CBH1-Au) revealed that the
cellulose microfibrils of outer epidermal walls of non-infected
V. faba leaf tissue were heavily masked by other
components of the plant cell wall. Such material was most probably
pectin because the cellulose microfibrils of
swollen epidermal and modified mesophyll walls of infected host
tissue were heavily labelled with CBH1-Au.
These results were confirmed by double-labelling studies using JIM 7
and CBH1-Au. At early stages of the
infection process, limited cellulose degradation was observed in infected
leaf tissue.
Double-labelling experiments using the monoclonal antibody BC-KH4
directed against Botrytis matrices and
a marker for the plant cell wall (JIM 7 or CBH1-Au) confirmed previous
observations that the fungal matrices
extended through modified host walls and degenerate cytoplasm. It is
suggested that the wall-modifying action of
the pectin-degrading enzymes produced during the infection process
might facilitate pervasion of matrix material
associated with the infection hyphae through host cell walls.
Possible role(s) of such matrix material during the
post-penetration processes of the V. faba–B. fabae
relationship are discussed.