Insights into stress responses in mandarins triggered by Bacillus subtilis cyclic lipopeptides and exogenous plant hormones upon Penicillium digitatum infection.

KEY MESSAGE: Bacillus subtilis CLP extract activates defense gene expression and increases the unique protein production involving in pathways of ISR, SAR, ubiquitin-proteasome system, and glycolysis for stress responses in flavedo tissues. Cyclic lipopeptides (CLPs) of Bacillus subtilis ABS-S14 had ability to activate plant defensive pathways, increase resistance and control green mold rot caused by Penicillium digitatum in mandarin fruit. The current study investigated transcriptional and proteomic data to highlight the unique induction effect of CLPs produced by B. subtilis ABS-S14 on the defense mechanism of mandarins in response to P. digitatum attack, and their differences from those following the exogenous plant hormone application. The proteomic patterns of the flavedo tissues as affected by Bacillus CLP extract, salicylic acid (SA), methyl jasmonate (MeJA), and ethephon (Et) were explored. qPCR analysis revealed the great effects of CLP extract in enhancing the transcription of PAL, ACS1, GLU, POD, and PR1. Tryptic peptides by LC-MS analysis between treatments with and without fungal infection were compared. B. subtilis CLP extract empowered the plant's immune response to wound stress by the significant production of calmodulin-binding receptor-like cytoplasmic kinase 2, molybdenum cofactor sulfurase, and NAD+-dependent glyceraldehyde-3-phosphate dehydrogenase. Ubiquitin carrier protein abundance was developed only in the treated flavedo with CLP extract coupled with P. digitatum infection. The gene expression and overall proteome findings involving pathways of ubiquitin proteasome system, ISR, SAR, and energy production provide a new insight into the molecular mechanisms of the antagonist B. subtilis ABS-S14 inducing resistance against green mold in mandarins.

Targeted transcriptional and proteomic studies explicate specific roles of Bacillus subtilis iturin A, fengycin, and surfactin on elicitation of defensive systems in mandarin fruit during stress.

Application of Bacillus cyclic lipopeptides (CLPs); fengycin, iturin A and surfactin has shown a great potential in controlling the spread of green mold pathogen invasion (Penicillium digitatum) in wounded mandarin fruit during postharvest period. The limited defensive protein profiles followed specific expression of pivotal genes relating to plant hormone mediating signaling pathways of the CLPs' action on stimulating host plant resistance have been exhibited. The present study aimed to elucidate the specific effect of individual CLP obtained from Bacillus subtilis ABS-S14 as elicitor role on activation of plant defensive system at transcriptional and proteomic levels with and without P. digitatum co-application in mandarin fruit. Fengycin and iturin A elevated the gene expression of PAL, ACS1, ACO, CHI, and GLU while significantly stimulating plant POD transcription was only detected in the treatments of surfactin both with and without following P. digitatum. An increase of LOX and PR1 gene transcripts was determined in the treatments of individual CLP with fungal pathogen co-application. Fengycin activated production of unique defensive proteins such as protein involved in ubiquinone biosynthetic process in treated flavedo without P. digitatum infection. Proteins involved in the auxin modulating pathway were present in the iturin A and surfactin treatments. CLP-protein binding assay following proteome analysis reveals that iturin A attached to 12-oxophytodienoate reductase 2 involved in the oxylipin biosynthetic process required for jasmonic acid production which is implicated in induced systemic resistance (ISR). This study suggests specific elicitor action of individual CLP, particularly iturin A showed the most powerful in stimulating the ISR system in response to stresses in postharvest mandarins.

Cyclic LIPopeptides from Bacillus subtilis ABS-S14 elicit defense-related gene expression in citrus fruit.

Effects of cyclic lipopeptides (CLPs) obtained from Bacillus subtilis ABS-S14 on eliciting defense-related gene transcription and activity of defense-related enzymes; glucanase (GLU), chitinase (CHI), peroxidase (POX) and lipoxygenase (LOX) in Citrus sinensis cv. Valencia fruit were determined. The maximum level of GLU transcripts induced in fruit treated with fengycin was significantly greatest among treatments at 48 h. Surfactin enhanced the LOX and POX transcripts. In parallel, corresponding enzyme activities were correlated with changes in gene expression observed in fruit inoculated with Penicillium digitatum following treatment with individual CLPs. Synergistic effects of fengycin and iturin A, fengycin and surfactin were shown in gene transcript of GLU and CHI, respectively, and surfactin induced POX and LOX gene expression of citrus flavedo without pathogen infection. These results suggest that fengycin and surfactin act as elicitors of defense-related gene expression in "Valencia" fruit following infection.