3. Discussion and
conclusions
Plant bioactive substances mainly target the cell membrane of fungi by
altering membrane stability and causing damage to the membrane structure
and extravasation of inclusions, ultimately resulting in fungicidal or
fungistatic effect (Zhou et al., 2014). Previous studies have shown that
glycoalkaloids predominantly combine with sterols in the fungal cell
membrane, forming a complex that destroys membrane integrity and causes
loss of normal membrane function (Sun, 2014). In the present study,
transmission electron microscopy showed that F. solani morphology
was distorted after potato glycoalkaloid treatment, and that some cell
walls were blurred or even lost. Additionally, structure of the cell and
vacuolar membranes was destroyed and the organelles distorted. These
observations indicated that potato glycoalkaloids can affect the surface
morphology of F. solani , leading to incomplete membrane structure
and causing serious damage to cytoplasm and mitochondria, consistent
with the effect of pyrolin on Monilinia fructicola (Wu et al.,
2009), ethyl acetate extract of amaranth on Xanthomonas citri(Liao et al., 2017), and water-soluble chitosan on the ultrastructure ofFusarium (Jia et al., 2016).
Electrical conductivity can indirectly reflect cell membrane
permeability, and a higher electrical conductivity of a culture broth
signifies enhanced electrolyte leakage and increased damage to cell
membranes (Shang, 2017). Peng et al. (2017) showed that extract ofCynanchum atratum could enhance the cell membrane permeability of
Italian Penicillium mycelium, while Zhang et al. (2008) revealed
that the extract of Xanthium sibiricum induced changes in
membrane permeability of Botrytis cinerea , resulting in increased
conductivity of the culture broth. Furthermore, Liu et al. (2018) found
that the total saponins and total ginsenoside from ginseng stem and leaf
can induce changes in the permeability of the mycelium membrane ofFusarium pediculae and F. solani , respectively, leading to
increased conductivity of the culture broth. Liu et al. (2018) showed
that limonene can increase the permeability of P. aeruginosa cell
membrane and destroy its cell morphology and integrity, thus effectively
inhibiting its growth. Similarly, in the present study, the cell
membrane permeability of F. solani increased after treatment with
potato glycoalkaloids, and transmission electron microscopy revealed
leakage of intracellular contents and destruction of cell membrane
structure. These results showed that F. solani cell membrane and
integrity were destroyed by potato glycoalkaloids, which directly
affected the physiological functions of the cell membrane, such as
exchange of intracellular and extracellular substances and regulation of
cell growth, leading to disturbance in fungal metabolism.
It must be noted that plant bioactive compounds also affect the
morphology and structure of fungal mycelia, causing deformity, kinking,
swelling, and lysis. As a result, the mycelial soluble protein and
soluble sugar can leak into the culture medium (Fan et al., 2015; Zhang
et al., 2016; Zhou et al., 2011). In fungi, soluble protein mainly
comprises various enzymes involved in metabolism. During growth, the
fungi secrete proteins that penetrate the cell membrane into the thallus
fluid through osmosis; thus, changes in the content of these proteins
reflect alteration in the total cellular metabolism (Liu et al., 2016).
Sugar catabolism provides the energy needed for fungal growth, and
inhibition of the absorption and utilization of sugar could lead to lack
of energy required by the fungi, affecting growth and propagation of the
thallus (Liu et al., 2016). The total lipid content in the fungal cell
membrane affects cell membrane fluidity, and a decrease in the total
lipid content may lead to a reduction in cell membrane fluidity (Shang,
2017). Thus, one approach to achieve fungicidal or fungistatic effect is
to inhibit fungal metabolism (Zhou et al., 2014). In the present study,
the contents of total sugar, protein, and fat inF.
solani initially increased and then decreased with time after treatment
with potato glycoalkaloids. However, the content of reducing sugar
decreased in F. solani treated with potato glycoalkaloids, but
significantly increased in control cells without glycoalkaloid
treatment. Similar findings were also reported by Wu (2008), who showed
that the contents of total sugar, reducing sugar, protein, and fat inB. cinerea treated with propamidine were significantly higher
than those in control B. cinerea without treatment, indicating
that plant bioactive compounds inhibited catabolism in fungi. Biological
catabolic systems are complex, and disturbance in a certain catabolic
link can obstruct the entire catabolic process, threatening life of the
organism. However, self-remedial mechanisms can overcome the blocked
metabolic processes to continue life activities (Zeng, 2012). Therefore,
it is possible that self-remedial mechanisms of F. solani allowed
the fungal cells to thrive after potato glycoalkaloid treatment,
resulting in an initial increase and subsequent decrease in contents of
total sugar, protein, and fat with treatment duration.
Furthermore, potato glycoalkaloids significantly affected the
morphological structure of F. solani . Treatment with potato
glycoalkaloids resulted in bubbly and undulated mycelial cell walls,
incomplete outer structure, discontinuous cell membrane, disordered
structure of mitochondria and other organelles, and visible
extracellular contents. The material metabolism analysis demonstrated
that potato glycoalkaloids destroyed the selective permeability of
fungal cell membranes and causing extravasation of large quantities of
internal lipids, proteins, and sugars. This hindered the hydrolysis of
reducing sugars, affecting the absorption and utilization of nutrients,
and ultimately inhibiting catabolism in fungi. However, knowledge of the
specific antifungal mechanism of potato glycoalkaloids is limited.
Therefore, further research on the effects of potato glycoalkaloids on
fungal respiratory metabolism (e.g. related enzymes activities) and
energy metabolism (e.g. inhibition of electron transport and oxidative
phosphorylation, and information expression of nucleic acids and other
molecular substances) is necessary for a better understanding of the
antifungal mechanism of these compounds and for acquiring comprehensive
and systematic theoretical support for the development and utilization
of botanical pesticides.