2.7 Statistical analyses
All data were analyzed with IBM SPSS (Version 23). The Student’st- test (two-tailed) was performed to analyze the difference
between Mock and AR156. One-way ANOVA followed by Tukey’s post hoc test
was performed to the difference between three or more independent
groups. The data are presented as mean ± SD.
3 RESULTS 3.1B. cereus AR156 pretreatment improves broad-spectrum disease
resistance in Arabidopsis Plants are exposed to a variety of
pathogens during growth and development. To survive, plants have evolved
a series of defense mechanisms to resist these pathogens, such as
recruiting beneficial microorganisms to the rhizosphere. It is revealed
that the plant growth-promoting rhizobacterium B. cereus AR156
enhances plant resistance to Pst DC3000 by triggering the ISR inArabidopsis . In order to demonstrate that B. cereus AR156
can trigger ISR to improve broad-spectrum disease resistance inArabidopsis , A. thaliana plants, wild-type Col-0 were
inoculated with B.cinerea , Pst DC3000
and P. capsici 5 d after pretreated with B. cereus AR156
or 0.85% NaCl. Compared to the control, B. cereus AR156
treatment significantly reduced the lesion area caused by P.
capsici (Figure 1 A and B), the lesion area caused by B. cinerea(Figure 1 C and D), and bacterial growth of Pst DC3000 (Figure 1
E and F) in Arabidopsis leaves. B. cereus AR156
pretreatment reduced Arabidopsis sensitivity to P.
capsici , Pst DC3000, and B. cinerea . Taken together,
these results confirmed that B. cereus AR156 could trigger ISR
against broad-spectrum disease.3.2 B. cereus AR156
induces Arabidopsis phytoalexin biosynthesis signaling pathway To
explore the mechanisms of AR156-triggered ISR response to broad-spectrum
diseases, we performed RNA-sequencing experiments and also analyzed the
transcriptome changes initiated by AR156 pretreatment. We analyzed the
induction of phytoalexin-related genes in Arabidopsis after AR156
pretreatment as it plays a vital role in plant response to pathogens.
Compared to Mock, phytoalexin-related genes, including camalexin
synthesis and transport-related genes, accumulated in the leaves
pretreated with AR156 (Figure 2 A). The results of RT-qPCR showed that,
several P450 enzymes associated with camalexin syntheses, such asCYP79B2 , CYP79B3 , CYP71A13 and PAD3 (Figure
2 B~F), accumulated significantly at different points
after AR156 treatment compared to Mock. Meanwhile, the transcripts ofWRKY33 , PEN3 , and PDR12 were up-regulated after
AR156 treatment (Figure 2 G and H). These results were consistent with
the RNA-sequencing results. In addition, we investigated the effect of
AR156 on camalexin synthesis in Arabidopsis , the results
demonstrated that the AR156-triggered ISR could induce the accumulation
of camalexin (Figure 2 I). Taken together, AR156-triggered ISR can
induce the accumulation of phytoalexin such as camalexin synthesis and
secretion-related genes.3.3 B. cereus AR156
prime camalexin accumulation after pathogens infection To further
investigate the role of B. cereus AR156 triggered ISR in
pathogen-induced camalexin biosynthesis, we examined the camalexin
induction in wild-type plants pretreated with B. cereus AR156
after infection by P. capsici , Pst DC3000, and B.
cinerea . As shown in Figure 3 A, compared with the Mock, B.
cereus AR156-triggered ISR increased the content of Arabidopsiscamalexin at different time points of P. capsici infection. There
was still significant induction of camalexin accumulation by AR156-ISR
96 h after pathogen infection
compared to the control.
Consistently, the induction of camalexin biosynthetic genesCYP71A13 and PAD3 by P. capsici infection was also
significantly increased under pretreated with B. cereus AR156
(Figure 3 B and C). As shown in Figure 3 D, compared with the control,
AR156-triggered ISR increased the content of Arabidopsiscamalexin at different time points of B. cinerea infection, with
significant differences after 24 h of B. cinerea infection.
Notably, AR156 still enhanced camalexin synthesis in Arabidopsis96 h after B. cinerea challenge. Consistent with this, the
induction of camalexin biosynthetic genes CYP71A13 andPAD3 by B. cinerea infection were also significantly
increased under pretreated with B. cereus AR156 (Figure 3 E, and
F). As shown in Figure 3 G, compared with the Mock, AR156-triggered ISR
increased the content of Arabidopsis camalexin at different time
points of Pst DC3000 infection, with significant differences 24 h
after Pst DC3000 infection. We also found that B. cereusAR156 induces maximal synthesis of camalexin at 72 h of PstDC3000 infection. Consistent with this, the induction of camalexin
biosynthetic genes CYP71A13 and PAD3 upon PstDC3000 infection was also significantly increased under pretreated withB. cereus AR156 (Figure 3 H and I). These results suggested that
AR156-induced ISR increases the expression of camalexin synthesis genes
and the accumulation of camalexin.