蜡样芽孢杆菌B3-7在大田小麦根部的定殖动态及其对小麦纹枯病的防治效果

Abstract

为了阐明蜡样芽孢杆菌B3-7在大田条件下的生态适应性以及对于小麦纹枯病的生防效果，通过利用绿色荧光蛋白编码基因gfp标记生防菌株B3-7，室内比较了GFP标记菌株和原始出发菌株在菌落形态、生长特性，生物薄膜产生以及在小麦根部定殖等方面的特性，结果发现GFP标记菌株和出发菌株在上述特性方面无明显差别。在此基础上，大田条件下测定了GFP标记菌株在小麦根部的定殖动态和对于小麦纹枯病的生防效果。结果发现，GFP标记菌株在小麦根部能够长期定殖，其存在量在小麦分蘖期最大，每克根重达到10⁵ CFU，拔节期后，该细菌数量一直维持在10⁴ CFU之上。同时发现，生防菌株能够有效降低小麦纹枯病的严重度和提高罹病小麦的产量。小麦分蘖期、孕穗期和灌浆期生防菌对于小麦纹枯病的防治效果分别达到60%、34%，34%，小麦成熟后产量提高13%-15%。结果表明，B3-7在大田条件下具有较好的生态适应性和防治小麦纹枯病的能力。;Sharp eyespot of wheat is caused by the fungus <em>Rhizoctonia cerealis</em> and is an important soil-Borne disease of wheat worldwide. At present, no commercially cultivated disease resistant varieties are available to efficiently protect wheat against infection by this pathogen. Control of the plant disease sharp eyespot currently relies on application of pesticides such as Triadimefon and Validamycin; however, pesticides are expensive and pose serious health and environmental hazards. Persistent application of pesticides can cause selection of pesticide resistant pathogenic fungi. Biological control is an environmentally sound and effective means of reducing or mitigating plant diseases and the effects of plant pathogens through the use of natural microorganisms. In the rhizosphere (on the plant root or in close vicinity to the root) bacteria are abundant, most often organized in microcolonies. Bacteria in the rhizosphere, known as rhizobacteria, not only benefit from the nutrients secreted by the plant root but also beneficially influence the plant in a direct or indirect way, resulting in stimulation of plant growth. Among the beneficial microorganisms isolated from the rhizosphere,<em> Bacillus</em>spp. offer several advantages over fluorescent pseudomonads and other Gram-negative bacteria as seed inoculants for protection against root pathogens. <em>Bacillus</em> spp. have a long shelf life because of their ability to form endospores and their unique mechanisms of action as they can produce broad-spectrum antibiotics and compete for ecological niches against plant pathogenic fungi.<br><em>Bacillus cereus</em> B3-7, a biological control strain isolated from the wheat rhizosphere, shows potential for control of sharp eyespot and for efficient yield increase in wheat. To elucidate the ecological adaptability of B3-7 and its efficacy for biological control of sharp eyespot of wheat under field conditions, the green fluorescent protein (<em>gfp</em>) gene was inserted into the B3-7 genome and a GFP-tagged B3-7 strain was constructed. Colony shape, growth rate, biofilm formation and colonization dynamics on wheat roots were analyzed with B3-7 and the GFP-tagged strain in our laboratory. No significant differences were obtained for any of the characteristics tested between the two strains. Colonization ability of the B3-7 GFP-tagged strain on wheat roots was assessed under field conditions. The results indicated that the GFP-tagged strains could persistently colonize the roots of wheat. The number of colonizing bacteria was highest during the tillering period at 10⁵ CFU/g roots. The colonization number decreased to 10⁴ CFU/g roots in the heading period, and persisted through the milking period. Efficacy of B3-7 and the GFP-tagged strain as biological control agents against sharp eyespot of wheat was also evaluated under field conditions. The results revealed that the two strains could reduce disease severity and effectively increase the yield of wheat. The disease control efficacy of the two strains for sharp eyespot of wheat reached 60% to 62%, 34% to 39%, and 34% to 38% during the tillering, heading, and milking periods, respectively. The yield of wheat treated with the biological control strain increased 13% to 15% compared with that of the untreated control. The results indicated that <em>Bacillus cereus</em> B3-7 has the capacity for environmental adaptability and the potential to act as a biological control agent against sharp eyespot of wheat under field conditions.