Fengycin produced by Bacillus subtilis NCD-2 plays a major role in biocontrol of cotton seedling damping-off disease.

Bacillus subtilis strain NCD-2 is strongly antagonistic toward phytopathogenic fungi, and functions as an excellent biocontrol agent for cotton soil-borne diseases. The aims of this study were to characterize the main active antifungal compound from strain NCD-2 and clarify its role in suppressing cotton damping-off disease. Strain NCD-2 and lipopeptide extract prepared from an NCD-2 culture strongly inhibited the growth of Rhizoctonia solani in vitro. The lipopeptides of strain NCD-2 were separated by fast protein liquid chromatography (FPLC) and the antifungal compound was identified as a cluster of fengycin homologs analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. A fengycin-deficient mutant was obtained by in-frame deletion of the fengycin synthetase gene in B. subtilis NCD-2. Compared with the wild-type strain, this mutant showed decreased abilities to inhibit the growth of R. solani in vitro and to suppress cotton damping-off disease in vivo. Studies showed that the population of fengycin-deficient mutant was almost same as that of the wild-type NCD-2 strain in the cotton rhizosphere. However, the population of R. solani in the cotton rhizosphere colonized by the fengycin-deficient mutant was twice that in the cotton rhizosphere colonized by the NCD-2 wild-type strain. This study demonstrated that fengycin-type lipopeptides are the main antifungal active compounds produced by B. subtilis NCD-2. These compounds play a major role in restricting the population of R. solani in the cotton rhizosphere and in suppressing cotton damping-off disease.

Guo Q ，Dong W ，Li S ，Lu X ，Wang P ，Zhang X ，Wang Y ，Ma P ... -  《-》

DegQ regulates the production of fengycins and biofilm formation of the biocontrol agent Bacillus subtilis NCD-2.

Bacillus subtilis NCD-2 is an excellent biocontrol agent for tomato gray mold and cotton soil-borne diseases. The fengycin lipopeptides serve as a major role in its biocontrol ability. A previous study revealed that insertion of degQ with the mini-Tn10 transposon decreased the antifungal activity of strain NCD-2 against the growth of Botrytis cinerea. To clarify the regulation of degQ on the production of fengycin, we deleted degQ by in-frame mutagenesis. Compared with the wild-type strain NCD-2, the degQ-null mutant had decreased extracellular protease and cellulase activities as well as antifungal ability against the growth of B. cinerea in vitro. The lipopeptides from the degQ-null mutant also had significantly decreased antifungal activity against B. cinerea in vitro and in vivo. This result was confirmed by the decreased fengycin production in the degQ-null mutant that was detected by fast protein liquid chromatography analysis. Quantitative reverse transcription PCR further demonstrated that degQ positively regulated the expression of the fengycin synthetase gene. In addition, the degQ-null mutant also had a flatter colony phenotype and significantly decreased biofilm formation ability relative to the wild-type strain. All of those characteristics from degQ-null mutant could be restored to the strain NCD-2 wild-type level by complementation of intact degQ in the mutant. Therefore, DegQ may be an important regulator of fengycin production and biofilm formation in B. subtilis NCD-2.

Wang P ，Guo Q ，Ma Y ，Li S ，Lu X ，Zhang X ，Ma P ... -  《-》

Fengycin produced by Bacillus subtilis 9407 plays a major role in the biocontrol of apple ring rot disease.

Apple ring rot, caused by Botryosphaeria dothidea, is a serious apple disease in China. Bacillus subtilis 9407 was isolated from healthy apples and showed strong antifungal activity against B. dothidea. To identify the primary antifungal compound of B. subtilis 9407 and determine its role in controlling apple ring rot, a transposon mutant library was constructed using TnYLB-1, and a mutant completely defective in antifungal activity was obtained. The gene inactivated in the antifungal activity mutant had 98.5% similarity to ppsB in B. subtilis subsp. subtilis str. 168, which encodes one of the five synthetases responsible for synthesizing fengycin. A markerless ppsB deletion mutant was constructed. Compared with the wild-type strain, lipopeptide crude extracts from ΔppsB showed almost no inhibition of B. dothidea mycelial growth. Furthermore, fengycin-like lipopeptides (retention factor 0.1-0.2) that exhibited antifungal activity against B. dothidea were observed in the wild-type strain by thin-layer chromatography (TLC)-bioautography analysis, but not in ΔppsB. Semipreparative reverse-phase high performance liquid chromatography (RP-HPLC) detection revealed that ΔppsB lost the ability to synthesize fengycin. These results suggest that ppsB is responsible for synthesizing fengycin and that fengycin is the major antifungal compound produced by B. subtilis 9407 against B. dothidea. Moreover, a biocontrol assay showed that the control efficacy of ΔppsB was reduced by half compared with the wild-type strain, indicating that fengycin plays a major role in controlling apple ring rot disease. This is the first report on the use of a B. subtilis strain as a potential biological control agent to control apple ring rot disease by the production of fengycin.

Fan H ，Ru J ，Zhang Y ，Wang Q ，Li Y ... -  《-》

Purification and identification of Bacillus subtilis SPB1 lipopeptide biosurfactant exhibiting antifungal activity against Rhizoctonia bataticola and Rhizoctonia solani.

Mnif I ，Grau-Campistany A ，Coronel-León J ，Hammami I ，Triki MA ，Manresa A ，Ghribi D ... -  《-》

Identification and Characterization of Lipopeptides from Bacillus subtilis B1 Against Sapstain Fungus of Rubberwood Through MALDI-TOF-MS and RT-PCR.

Sajitha KL ，Dev SA ，Maria Florence EJ 《-》