4 │ Discussion
Early and elevated levels of the expression of various defense enzymes
are an important feature of plant disease resistance to different
pathogens (Andersen et al., 2018). Although defense-related enzymes
constitute an important protective system for plants against pathogen
invasion but the underlying mechanism of defense reactions and their
relationships with damping-off diseases in cucumber remain unclear.
Using uninoculated cucumber genotypes and time points before inoculation
as controls, this study analyzed the changes in the activities of PPO,
PAL, SOD, POX, and CAT in the roots of six cucumber genotypes with
different susceptibilities to damping-off to identify indices that allow
relevant information on damping-off resistance, which suggests that
diverse mechanisms contribute to the specific levels of resistance toP. melonis in genotypes. Although oxidative stress caused byP. melonis led to increased activities of defence‐related
enzymes, the overall enzyme activity patterns were distinct by enzyme
and genotype.
Generally, POX activity in plant tissues are induced by pathogen
infection and a greater increase was recorded in resistant plants
compared to the susceptible ones (Mydlarz & Harvell 2006; Bharathi et
al., 2019). Similar increase in POX activity, as observed in the
resistant genotypes in this study was observed in castor infected withFusarium oxysporum (Bharathi et al. 2019), cucumber infected byPythium aphanidermatum (Sabbaghi et al., 2018), and zucchini and
cucumber infected by mosaic virus (Riedle-Bauer, 2000). The increased
activity of POX in the resistant genotypes after inoculation at 14 DAI
was correlated with damping-off susceptibility in the cucumber genotypes
and related to plant disease resistance. Su et al. (2016) found that the
POX activity levels could potentially be used as a genetic marker for
resistant evaluation during the early phase of infection. Our results
support this indication that POX may also play an important role in
cucumber resistance to damping-off. POX and POD is widely distributed in
higher plants and protects cells against damaging effects of H2O2 by
catalyzing its decomposition (Fernandes et al., 2006).
After inoculation with P. melonis , the SOD activity increased in
the resistant genotypes. Although the SOD activity also increased in the
highly susceptible genotypes Mini 6-23 and Yalda, the response time and
range of increase were far lower than those in the resistant genotypes.
Therefore, the differences in the activity of SOD between the six
genotypes may be due to the complex activity patterns of SOD and
appeared to be an important physiological basis for resistance to
disease. However, the SOD activities did not correlate with damping-off
susceptibility. Other researchers have indicated an increase in the
activity of SOD after inoculation in cucumber (Nostar et al., 2013;
Moradi et al., 2016) and other plants in a way similar to our study (Su
et al., 2019).
The CAT enzyme activity displayed different patterns in different
genotypes under stress and mainly affects plant disease resistance
through two physiological pathways (Jiang et al., 2019). This seems to
be an important H202-scavenging enzyme in plants and be localized in the
peroxisomes. Catalase can be induced and then be consumed as a result of
oxidative stress (Garg & Manchanda, 2009). In this study, there was a
difference in the CAT activity between the resistant and susceptible
genotypes, which means the alternative of CAT showed intraspecific
genotypic variation in response to P. melonis inoculation and
suggested that altered CAT activity also had an important effect on
damping-off resistance. However, the physiological pathway by which CAT
affects resistance to P. melonis remains unclear and further
investigations are needed. This result is consistent with other studies
for Lilium hybrids (Su et al., 2019), walnut (Jiang et al., 2019) andBrassica juncea (Pandey et al., 2017).
Due to the importance of PAL in the phenylpropanoid pathway and the
relationship between PAL and plant disease resistance, it has always
been a hot research topic. Several previous studies using different
plant species have shown that the PAL activity is increased after fungal
infection and played an important role in plant defense against
pathogenic fungi (Saunders and O’neill, 2004; Zhang et al., 2017). In
this study, significant changes in the PAL activity occurred after
inoculation with P. melonis in resistant and moderately resistant
genotypes. There were also some differences among the genotypes.
Numerous studies also revealed that the activation of PAL and subsequent
increase in phenolic content in plants is a general response associated
with disease resistance. In black rice, PAL contributes to the
resistance mechanism against Xanthomonas oryzae pv. oryzae(Solekha et al., 2019). In tomato, PAL activity was enhanced in roots by
a biotic elicitor Fusarium mycelium extract (Mandal and Mitra,
2007). In cucumber roots, high levels of PAL were induced after
inoculation with P. aphanidermatum (Chen et al., 2000). In
transgenic soybean, overexpression of GmPAL2.1 increases resistance toPhytophthora sojae (Zhang et al., 2017). Therefore, increase in
level of PAL in the infected root tissue of cucumber genotype showed
that phenyl propanoid pathway accumulate phenolics might have prevented
the pathogen invasion, and thus, the activity maintained at higher
levels during the infection period.
Our findings show that, although the PPO activity increased in roots of
infected plants in comparison to the control in the resistant and
moderately resistant genotypes, but the activities varied among the
three genotypes. Several studies have indicated induction of PPO in
plants in response to infection by different pathogens (Vanitha et al.,
2009; Khodadadi et al., 2020). In present study, systemic induction of
PPO expression in the resistant and moderately resistant genotypes in
response to P. melonis might provide an additional line of
defense to protect cucumber plants against further attack by pathogens.
The PPO appear to play a role in resistance to P. melonis , since
this compound was present in considerably higher levels in roots of
inoculated plants of resistant genotypes which is consistent with
observations made for cucumber, walnut and common bean plants in which
PPO was demonstrated to be induced by Fusarium oxysporum,Xanthomonas arboricola and arbuscular mycorrhizal fungi,
respectively (Moghbeli et al., 2017; Jiang et al., 2019; Abdel-Fattah et
al., 2011). Additionally, a significant correlation between the
antioxidant enzymes indicate the role of antioxidant enzymes system
under P. melonis infection. The role of antioxidant enzymes in
eliciting systemic resistance against Fusarium oxysporum ,Pyricularia oryzae , Alternaria sp. and Sclerotiumsp. has been reported earlier (Rais et al., 2017). These findings
indicated the complex interaction between both oxidative-burst and
response to damping- off. Tt seems that the oxidativeburst and H2O2
accumulation also happened in resistant genotypes as an adaptive
response to P. melonis infection, and resistant genotypes may
handle stress better than susceptible genotypes during the infection due
to other mechanisms. However, the results of the changes in defensive
enzyme activity are far from sufficient to explain damping-off
resistance in cucumber. To understand cucumber resistance physiology
more clearly, other physiological indicators should be considered.
The results of this study indicated that the disease defense response is
more pronounced in the resistance genotypes as demonstrated by an
earlier defense response through increasing the content of
defense-related enzymes. In addition, the results suggest the PAL and
POX have active roles in disease resistance against damping-off;
however, there was no direct connection between damping-off resistance
and PPO, SOD and CAT activities. Meanwhile, the varied response of
defense-related enzymes suggests that the oxidative-burst response after
inoculation by P. melonis , and the response may have differed
depending on the genotype and inoculation period. These findings can
help us to understand the resistance physiology of damping-off and
provide indicators for cucumber breeding. However, diverse mechanisms
contribute to the specific levels of resistance to P. melonis in
genotypes and the mechanisms by which defense-related enzymes
accumulation contribute to resistance in cucumber remain to be explored
in future studies.