氨水熏蒸对高发枯萎病蕉园土壤微生物区系及发病率的影响

采用涂布计数及荧光定量PCR和PCR-DGGE技术,研究了氨水熏蒸对土壤微生物区系的影响,并研究氨水熏蒸对高发枯萎病香蕉园枯萎病的防控效果和对香蕉产量的影响。结果表明:与熏蒸前相比,高发枯萎病香蕉园土壤施用130 L/667 m²的氨水熏蒸后,尖孢镰刀菌的数量下降了1个log单位;与对照处理相比,下降了0.5个log单位。氨水熏蒸后土壤可培养细菌数量与对照相比没有显著差异,但显著低于熏蒸前,可培养真菌的数量显著低于熏蒸前和对照;可培养细菌真菌的比值与熏蒸前相比没有显著差异,但显著高于对照。与对照及熏蒸前相比,氨水熏蒸后土壤中总细菌及总真菌的数量显著下降;熏蒸前后及对照处理总细菌真菌的比值没有明显差异。各土壤样品总细菌及真菌群落结构的PCR-DGGE分析结果表明,氨水熏蒸后土壤微生物在聚类上和条带组成上均与熏蒸前及对照有显著性差异。与对照相比,氨水熏蒸处理使香蕉枯萎病的发病率降低了20%,每666.7m²增产1.25t。以上结果表明,利用合适浓度的氨水对高发枯萎病香蕉园的土壤熏蒸,能够有效改良土壤微生物区系和降低枯萎病的发生。

Effects of ammonia fumigation on soil microflora and banana production in an orchard with serious Fusarium wilt disease

Banana Fusarium wilt disease, caused by the fungus Fusarium oxysporum f. sp. cubense race 4, has been reported to be the most limiting factor in Cavendish-banana production worldwide since 1996. Furthermore, the Cavendish-banana production comprised about 90% of the banana growing areas in south China, and more than 200,000 ha banana orchards in this area have been attacked. Among the managements for controlling Fusarium wilt disease, pre-planted fumigation of soil is one of the effective and stable means for this disease prevention. However, methyl bromide, one widely used soil fumigant, will be banned internationally in 2015 due to the destruction of the ozone layer. In this study, ammonia, to our knowledge, has been first applied as soil fumigant to control this soil borne disease as a chemical alternative to methyl bromide in banana planting field. Traditional plate counting, Real-Time PCR and PCR-DGGE methods were used to determine the effects of ammonia fumigation on soil microflora, disease control efficiency and banana yield in a banana orchard with serious Fusarium wilt disease. Through the estimation by plate counting method, the number of Fusarium in the soil collected after fumigation with 130 L/667 m² of ammonia decreased 1 and 0.5 log unit, respectively, when compared with pre-fumigation (BF) and the non-fumigation control (CK). The numbers of soil culturable bacteria in the soils from fumigation treatment (AMO) and CK were significantly lower than that from BF treatment, while there was no significant difference could be detected among AMO and CK. CK and BF treatments showed significantly higher numbers of soil culturable fungi than AMO, while no significant difference could be detected among BF and CK treatments. The ratio of bacteria to fungi (B/F) in AMO was significantly higher than that in CK, but no significant difference was observed among pre-fumigating treatment (BF) and ammonia fumigating treatment (AMO). Through estimation by Real-Time PCR, the numbers of total bacteria and fungi in AMO were significantly lower than that in the BF and CK, while no significant difference of total B/F was found. Based on fungi PCR-DGGE fingerprints, the intensities of bands A1, A2 and A3 strengthened and bands of B1, B2 and B3 weakened in the AMO treatment whereas band F related to Fusarium sp. was obviously weakened in AMO treatment compared to BF and CK treatments. Soil bacteria PCR-DGGE fingerprints also revealed that the intensities of bands a1, a2 and a3 strengthened in AMO treatment and bands b1, b2, b3 and b4 weakened in the AMO treatment. However, in soil bacteria or fungi community, the three-replicate cluster of AMO treatment distinguished separately to the other clusters of BF and CK treatments while the clusters of BF and CK grouped together. Based on DGGE results, the soil bacterial and fungal structure for AMO treatment was obviously altered after fumigation compared to that of BF and CK. Field experiment showed that the Fusarium wilt disease incidence in AMO decreased about 20% compared to that in the CK and the banana yield per 667 m² for AMO increased 1.25 t due to the disease incidence decrease. In conclusion, all the results indicated that using ammonia fumigation could effectively improve soil microflora and control the banana Fusarium wilt disease.