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Molecular Pathogens 2012, Vol.3, No.4, 19
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22
al., 1994). In reaction mixture, by adding chitinolytic
as well as glucanolytic enzymes, showed synergistic
effect in the enhancement of antifungal responses of
five toxins of fungi against
B. cinerea
. These toxins were
benomyl, miconazole, gliotoxin, captan and flusilazole.
Dose response curves were made for each enzyme-toxin
combination and observed the ED values of the
mixtures were substantially lower than ED values of the
two compounds used alone in all combination. For
example, the addition of endochitinase at a
concentration of 10 pg/mL cause 86 fold reduction in
the ED value of toxin. The level of synergism
appeared to be higher when enzymes were combined
with toxins having primary sites of action associated
with membrane structure, compared with pesticides
having multiple or cytoplasmic sites of action. Among
enzymes tested, the maximum levels of synergism
with synthetic fungicides were observed for the
endochitinase from
T. harzianum
strain PI, which,
when used alone, was the most effective chitinolytic
enzyme against phytopathogenic fungi of those
tested. The utilization of hydrolytic enzymes to
synergistically improve the antifungal potential of
fungitoxic compounds may decrease the effect of
pesticides on animals and plants.
The majority of pathogenic fungi have cell wall
with chitin and callose (Bartnicki-Garcia, 1968), and
dissolution of these structural polymers has adverse
impact upon the growth and development of fungi
(Poulose, 1992). Biocontrol microbes (
Gliocladizmz
and
Trichoderma
) produce enzyme for degrading cell
wall especially chitinolytic, to control pathogenic
fungi of plants (Broglie et al., 1991; Ordentlich et al.,
1988; Di Pietro et al., 1993; Lorito et al., 1993). When
these enzymes are present together, then they have
synergistic effect (Lorito et al., 1993). Additionally
these enzymes should not have toxic effects for plants
as well as other vertebrates, since they have not any
target polymer. For the control of pathogenic fungi,
these cell wall degrading enzymes show synergistic
effects in combination with pesticides (Collins and
Pappagianis, 1974; Köller, 1992; Roberts et al., 1988;
Watanabe et al., 1988). Cell wall break down with
enzyme may improve the chemical uptake into the
target cell (Poulose, 1992).
Strain (PI and 41) of
T. harzianum
has affectivity
against
B. cinerea
and
Pbtophtbora
diseases (Tronsmo,
1991; Smith et al., 1990). Endochitinase (41 kD) and
chitobiosidase (40 kD) are from strain PI of
T.
harzianum
and both enzymes have strong inhibitory
effect on fungi, which have chitin in their cell wall,
especially, when both are used in combination (Lorito
et al., 1993 and Lorito et al., 1994). These enzymes in
combination have synergistic effect for increasing the
affectivity of biocontrol agent of bacteria (Lorito et al.,
1993). Glucosidase (78 kD), third enzyme from P1
strain
Trichoderma
, also showed antifungal potential
against various chitinous fungi, in
in vitro
assay
(Lorito et al., 1994). From strain 41 of
G. viren
, an
enzyme endochitinase, inhibits elongation of germ
tubes and germination of spores in
B. cinerea
, but at
low level than chitinase enzymes of
T. harzianum
(Di
Pietro et al., 1993).
There are several reports according to which the cell
wall breaking enzymes, act synergistically and cause
various morphological syndromes within hyphae,
even vegetative cell is burst (Lorito et al., 1993; De La
Cruz et al., 1992; Lorito et al., 1993; Mauch et al.,
1988; Nevalainen et al., 1991). Additionally plant
chitinases were synergistic with inhibitors of chitin
synthesis in fungi, such as polyoxin B and nikkomycin
(Roberts et al., 1988; Poulose, 1992). Chitinolytic
enzymes from
T. harzianum
have been shown to act
synergistically against a wide range of disease-causing
fungi in the Ascomycetes, Deuteromycetes and
Basidiomycetes (Lorito et al., 1993).
Lorito et al (1994) suggested that the utilization of cell
wall degrading enzymes of fungi in combination with
pesticides can be fruitful for various applications of
agriculture or even for veterinary and human sciences.
Davies and Pope (1978) suggested that a mixture of
chitinolytic enzymes and chemical antifungal agents
had fruitful antimycotic action in immuno-suppressed
laboratory animals. Plant-pathogen interactions
cause host-specific biochemical responses, therefore
the plant could tackle the attack from pathogens
(Dangl and Holub, 1997). Chitin and glucan are
cell-wall molecules of various pathogenic fungi and