南方农业学报2023，Vol.54Issue(8)：2245-2256,12.DOI:10.3969/j.issn.2095-1191.2023.08.006
抗烟草黑胫病菌生防细菌筛选、鉴定及体外拮抗机理初探
Screening,identification and antagonistic mechanism in vitro of biocontrol bacteria against tobacco black shank pathogen
摘要
Abstract
[Objective]Strains with high antagonistic activity against the pathogen of tobacco black shank were isola-ted from tobacco rhizosphere soil in the high incidence area of tobacco black shank disease in Yunnan.The taxonomic sta-tus of strains was clarified and the in vitro antagonism mechanism was preliminary studied to provide biocontrol resources and reference basis for the prevention and control of tobacco black shank disease.[Method]Strains antagonistic to tobacco black shank pathogen were isolated from tobacco rhizosphere soil using the plate standoff method.The strains with strong antagonistic activity were identified by physiological and biochemical characteristics and 16S rDNA sequence analysis.The biocontrol activity and mode of action of biocontrol bacteria against Phytophthora nicotianae in vitro were deter-mined.[Result]Six bacterial strains with antagonistic activity against P.nicotianae were isolated from the collected tobac-co rhizosphere soil samples,among which strain CD-1 showed the strongest antagonistic activity,with a radius of inhibi-tion zone of 10.12 mm.According to physiological and biochemical characteristics and 16S rDNA sequence analysis,the strain was identified as Bacillus licheniformis.Strain CD-1 had broad-spectrum antifungal properties,and both intracellu-lar and extracellular antifungal substances existed.Among them,the antifungal substance extracted using dichloromethane as bacterial fermentation liquid extractant had the best inhibitory effect.When pH was in the range of 6-8 and the tempera-ture was between 40-80℃,it showed relatively stable inhibitory activity.Strain CD-1 cultured in NB medium showed strong inhibitory activity against P.nicotianae in both culture supernatant extracts and cell extracts,with inhibition rates of 76.91%and 67.23%,respectively.The antifungal substances secreted by strain CD-1 could destroy the spore plasma membrane of P.nicotianae and inhibit spore germination.Among them,the spore membrane loss rate after treating P.ni-cotianae spores with the antifungal crude extract 0.4 mg/mL for 6 h reached 53.62%,and the spore germination was com-pletely inhibited,while obvious ion and nucleic acid leakage was detected.[Conclusion]The metabolite produced by B.li-cheniformis CD-1 has a broad antimicrobial spectrum and strong bacteriostatic activity,which has the potential to be de-veloped into antifungal agents.关键词
烟草黑胫病/生防细菌/分离鉴定/体外拮抗机理Key words
tobacco black shank/biocontrol bacteria/isolation and identification/antagonistic mechanismin vitro分类
农业科技引用本文复制引用
付克剑,苏友波,崔永和,黄智华,张继来,张立猛,邓毅书,张智浩,聂强,周洪印..抗烟草黑胫病菌生防细菌筛选、鉴定及体外拮抗机理初探[J].南方农业学报,2023,54(8):2245-2256,12.基金项目
云南省重大科技专项计划项目(202002AE320005) （202002AE320005）
中国烟草总公司云南省公司科技计划一般项目(2020530000 242025) Yunnan Major Science and Technology Special Project(202002AE320005) （2020530000 242025）
Science and Tech-nology Plan General Project of Yunnan Tobacco Company,China National Tobacco Corporation(2020530000242025) （2020530000242025）
