Genomic and metabolic features of Bacillus cereus, inhibiting the growth of Sclerotinia sclerotiorum by synthesizing secondary metabolites

Abstract

We investigated the biocontrol mechanism of Bacillus cereus CF4-51 to find powerful microbes that effectively control Sclerotinia sclerotiorum. To assess its inhibitory effect on fungal growth, the plant pathogen (S. sclerotiorum) was co-cultured with Bacillus cereus. Scanning electron microscope (SEM) was used to study the morphology of S. sclerotiorum treated with CF4-51 biofumigant. The expression of sclerotium formation-related genes was analyzed by qRT-PCR. We performed whole genome sequencing of CF4-51 by PacBio Sequel platform. Lipopeptides were extracted from strain CF4-51 according to the method of hydrochloric acid precipitation and methanol dissolution. The volatiles CF4-51 were identified using gas chromatography–mass spectrometry (GC–MS). We found that the volatile organic compounds (VOCs) released by CF4-51 damaged the S. sclerotiorum hyphae and inhibited the formation of sclerotia. The qRT-PCR data revealed the down-regulated expression of the genes involved in sclerotial formation. Moreover, we analyzed the B. cereus CF4-51 genome and metabolites. The genome consisted of 5.35 Mb, with a GC content of 35.74%. An examination of the genome revealed the presence of several gene clusters for the biosynthesis of antibiotics, siderophores, and various other bioactive compounds, including those belonging to the NRPS-like, LAP, RIPP-like, NRPS, betalactone, CDPS, terpene, ladderane, ranthipeptide, and lanthipeptide (class II) categories. A gas chromatography–tandem mass spectrometry analysis identified 45 VOCs produced by strain CF4-51. Among these, technical grade formulations of five were chosen for further study: 2-Pentadecanone, 6,10,14-trimethyl-,1,2-Benzenedicarboxylic acid, bis(2-methylpropyl) ester, Dibutyl phthalate, Cyclododecane, Heptadecane. the five major constituents play important roles in the antifungal activity of the VOCs CF4-51 on the growth of S. sclerotiorum. The secondary metabolites produced by strain CF4-51are critical for the inhibition of S. sclerotiorum hyphal growth and sclerotial formation.

Bacillus VOCs in the Context of Biological Control

“…Additionally, the result was 77.1% and 50% disease suppression in fumigation treatment with cultivation broth of these two strains, respectively [ 65 ]. Similar thin or gapped structures of the uneven cell wall surface presenting a retracted protoplasm were also observed in B. cereus CF4-51 VOCs-treated mycelia of Sclerotinia sclerotiorum [ 66 ]. Acetic acid (20.68%), propanoic acid (33.30%), butanoic acid (26.87%), valeric acid (43.71%), and isovaleric acid (53.10%) produced by Bacillus sp.…”

“…A higher minimal inhibitory concentration of the VOCs produced by B. megaterium USB2103 was required to inhibit sclerotia germination compared to mycelial growth of Sclerotinia sclerotiorum , except in the case of dimethyl trisulfide whose minimal inhibitory concentration has shown similar results either on mycelium growth or sclerotia germination [ 49 ]. B. cereus CF4-51 VOCs (2-pentadecanone, 6,10,14-trimethyl-, 1,2-benzenedicarboxylic acid bis(2-methylpropyl) ester, dibutyl phthalate, cyclododecane, and heptadecane) have shown supreme inhibition of Sclerotinia sclerotiorum (65.4%) in comparison with the cell free supernatant (18.2–42.4%) and lipopeptides (55.6%) by altering the expression of four genes related to sclerotia formation ( Ss-sl2 , SOP1 , SsAMS2 , and SsSac1 ) [ 66 ]. B. velezensis VM11, B. velezensis VM10, and B. amyloliquefaciens VM42 VOCs exhibited fungistatic effects toward Sclerotinia sclerotiorum by inhibiting sclerotia production by 62.9%, 54.7%, and 72.5%, respectively, while healthy harvested sclerotia lost their germination ability after being incubated together with 5 μL of Bacillus endophyte cultures [ 57 ].…”

Identification of bioactive compounds of Bacillus velezensis HNA3 that contribute to its dual effects as plant growth promoter and biocontrol against post-harvested fungi

“…Bacillus cereus synthesizes 2-benzenedicarboxylic acid, which exhibits inhibitory activity against Sclerotinia sclerotiorum ( 78 ). Besides, it was identified in Bacillus atrophaeus XEGI50 extract that was used as a biocontrol agent against verticillium wilt disease in cotton plants ( 79 ).…”

Computation-aided studies related to the induction of specialized metabolite biosynthesis in microbial co-cultures: An introductory overview

“…So, the product of the cluster depended on the accompanying microorganism [98] . Hu et al [99] investigated the inhibitory effects exerted by Bacillus cereus CF4–51 on the plant pathogen Sclerotinia sclerotiorum . The co-cultivation experiments revealed that the volatile organic compounds (VOCs) generated by B. cereus CF4–51 disturbed the development and damaged the hyphal structures of S. sclerotiorum .…”