Screening, identification and biological control effect of antagonistic fungus against fusarium wilt of Saposhnikovia divaricata
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摘要:目的
从防风根际土中分离筛选出对防风枯萎病具有较强防治效果的生防真菌。
方法采用对峙培养法从根际土壤中筛选出1株可拮抗尖孢镰刀菌的真菌菌株MR-97，并测定其抑菌谱；根据真菌菌落特征、显微特征，结合ITS序列分析等方法对拮抗菌株进行鉴定；通过显微观察MR-97对尖孢镰刀菌菌丝生长的影响；采用抗生素标记法测定拮抗菌株在土壤中的定殖能力；进行田间盆栽试验，检验MR-97对防风枯萎病的防治效果。
结果本研究筛选出1株对尖孢镰刀菌抑菌效果为64.44%的拮抗菌株MR-97，其对木贼镰刀菌、灰葡萄孢等8种常见病原菌具有较强的抑菌效果，经鉴定菌株MR-97为土曲霉Aspergillus terreus。MR-97与尖孢镰刀菌对峙培养可使病原菌菌丝产生膨大、畸形、菌丝破损、内含物凝集等现象；在土壤中具有良好的定殖效果，土壤含菌量最高为9.8×106 CFU/g。盆栽试验中，MR-97对防风枯萎病防效为67.86%，防治效果较好。
结论土曲霉MR-97可有效抑制尖孢镰刀菌等多种病原菌菌丝生长，可在土壤中较快定殖并发挥生防效力。作为优质的生防菌源，MR-97具有一定的开发应用潜力。
Abstract:ObjectiveThe present study aimed to screen and isolate the antagonistic fungal strains with strong control effect against fusarium wilt from rhizosphere soil ofSaposhnikovia divaricata.
MethodThe strain MR-97, which was antagonistic to Fusarium oxysporum, was isolated from the rhizosphere soil of S. divaricata by confrontation culture method, and its bacteriostatic spectrum was determined. The strain was identified by fungal colony characteristics, microscopic characteristics and ITS sequence analysis. The effect of strain MR-97 on hyphae growth of pathogenic fungus were recorded by microscopic examination. The colonization ability of strain MR-97 in the soil was evaluated by antibiotic marker method. The control effect of strain MR-97 on fusarium wilt of S. divaricata were evaluated by pot experiment in the field.
ResultThe antagonistic strain MR-97 was screened with antibacterial rate of 64.44% againstF. oxysporum. It also had strong antibacterial effect against eight common pathogens including F. equiseti and Botrytis cinerea, etc. The strain MR-97 was identified asAspergillus terreus. The hyphal of pathogen enlargement, malformation, damage and agglutinated with inclusions, etc. when the strain MR-97 and F. oxysporum were cultured in confrontation. It had a good colonization effect in the soil, and the highest content of soil bacteria was 9.8×106 CFU/g. The biocontrol effect of MR-97 on fusarium wilt of S. divaricata was 67.86% in pot experiment, which had good control efficiency.
ConclusionAspergillus terrestris MR-97 can effectively inhibit the mycelia growth of F. oxysporum and other pathogens, can colonize quickly in soil and has strong biocontrol effect. MR-97 strain, as a good source of biocontrol bacteria, has certain potentials of development and utilization.
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图 1 菌株MR-97菌落(a)及显微形态特征(b)
Figure 1. Colony (a) and micromorphological characteristics (b) of strain MR-97
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图 2 菌株MR-97基于 rDNA-ITS建立系统发育树
Figure 2. The phylogenetic tree of strain MR-47 based on rDNA-ITS sequence
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图 3 菌株MR-97对尖孢镰刀菌菌丝生长的影响
a：尖孢镰刀菌正常生长状态；b：粗细不均，表面粗糙；c：膨大畸形； d: 干瘪、皱缩、扭曲、折叠
Figure 3. Antibacterial effect of strain MR-97 on mycelium growth ofFusarium oxysporum
a: Normal growth state; b: Uneven thickness and rough surface; c: Swollen malformation; d: Wizened, shrunken, twisted and folded
表 1 菌株MR-97对盆栽防风枯萎病防病效果1)
Table 1 Control effect of strain MR-97 on Fusarium wilt of potted Saposhnikovia divaricata
处理 Treatment 病情指数 Disease index 防效/% Control effect 清水对照 Water control 50.59±4.29a 多菌灵500倍液 Carbendazim 500× liquid 16.00±4.23b 68.37±8.36 枯草芽孢杆菌可湿性粉剂 Bacillus subtilis wettable powder 22.18±3.72b 56.15±7.35 哈茨木霉可湿性粉剂 Trichoderma harzianum wettable powder 19.59±2.28b 61.27±4.51 MR-97 16.25±4.47b 67.86±8.84 1)表中数据为平均值±标准差；同列数据后不同小写字母表示差异显著(P<0.05，Duncan’s法) 1) Data are means ± standard deviations; Different lowercase letters in the same column indicate significant difference(P<0.05, Duncan’s method) 
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表 2 不同处理对盆栽防风的促生效果1)
Table 2 Promotion effect of different treatments on growth of potted Saposhnikovia divaricata
处理 Treatment 株高/cm Plant height 根长/cm Root length 全株鲜质量/g Whole fresh biomass 根鲜质量/g Root fresh biomass 全株干质量/g Whole dry biomass 根干质量/g Root dry biomass 清水对照 Water control 51.86±2.24a 28.24±1.99a 7.52±0.72b 2.01±0.35b 2.24±0.63a 0.45±0.13b 多菌灵500倍液 Carbendazim 500× liquid 53.44±3.24a 30.93±3.8a 10.63±2.07b 3.86±0.83ab 2.73±0.08a 1.02±0.39ab 枯草芽孢杆菌可湿性粉剂 Bacillus subtilis wettable powder 50.63±2.33a 29±0.38a 7.67±1.03b 2.32±0.58b 1.94±0.78a 0.52±0.03b 哈茨木霉可湿性粉剂 Trichoderma harzianum wettable powder 48.95±1.92a 27.88±2.57a 7.43±1.87b 3.44±1.57b 2.09±0.53a 1.02±0.3ab MR-97 55.21±10.95a 32.71±4.34a 18.11±2.5a 6.69±2.92a 4.55±2.8a 1.55±0.63a 1)表中数据为平均值±标准差；同列数据后不同小写字母表示差异显著(P<0.05，Duncan’s法) 1) Data are means ± standard deviations; Different lowercase letters in the same column indicate significant difference (P<0.05, Duncan’s method)
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