Screening And Action Mode Of Biocontrol Bacterium Against Potato Black Scurf

To develop the micro-fungicide for suppressing against potato black scurf,which caused by Rhizoctonia solani,the bacteria with biocontrol ability against potato black scurf were screened and identified,and the mode for inhibiting the growth of R.solani were clarified in this study.A total of 2106 soil bacteria were firstly tested for inhibiting the growth of R.solani in vitro,and then 61 bacteria with stronger antifungal abilities were evaluated for suppressing of potato black scurf under greenhouse condition.Strain HMB33604 was selected for its excellent biocontrol ability.Strain HMB33604 was identified as Bacillus amyloliquefaciens by Biolog microbiological identification system as well as multiple genes sequence alignment.The optimal fermentation medium for strain HMB33604 was firstly determined from 11 industrial media.The medium composition was further optimized for strain HMB33604,and concentration of 1.43 × 109 CFU/mL could be obtained.The fermentation broth,cell free supernatant and bacterial cell suspension of strain HMB33604 were evaluated for suppressing of potato black scurf,respectively.Both the fermentation broth and cell free supernatant showed significant biocontrol effects against potato black scurf with 52.16%and 66.36%respectively.However,the control effect of the cell suspension was only 16.90%.Meantime,the population of R.solani in the potato rhizosphere was quantified by Real-time PCR.Results showed that both the treatments of fermentation broth and cell free supernatant significantly decreased the population of R.solani compared to that of the cell suspension and blank control.Therefore,the antifungal active compounds produced by strain HMB33604 played an important role in suppression of the potato black scurf.The lipopeptide was extracted from the culture of strain HMB33604 by HCI precipitation,and fengycin,surfactin and iturin could be detected from the lipopeptide extracts by UPLC-Triple TOF-MS/MS.The genes encoding the lipopeptide antibiotics were cloned from the genome of strain HMB33604,and the sequences of PCR fragment also confirmed the production of fengycin,surfactin and iturin A by strain HMB33604.Fengycin,surfactin and iturin were separated by HPLC,and only the fengycin and iturin showed strong inhibition abilities against the growth of R.solani.The synergistic effects between strain HMB33604 and biocontrol Trichoderma strain LTR-2,T11W,21990 and strain 7 were evaluated in greenhouse for suppression of potato black scurf,respectively.Results showed Trichoderma strain LTR-2 could increase the biocontrol ability of strain HMB33604 for potato black scurf.In addition,strain HMB33604 also showed promising biocontrol effects aginst both cotton verticillium wilt and rhizoctonia damping-off.