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当前位置: 首页 > 文章 > 吡咯伯克霍尔德菌WY6-5的溶磷、抑菌与促玉米生长作用研究 中国农业科学 2019 (9) 1574-1586
Position: Home > Articles > 吡咯伯克霍尔德菌WY6-5的溶磷、抑菌与促玉米生长作用研究 Scientia Agricultura Sinica 2019 (9) 1574-1586

吡咯伯克霍尔德菌WY6-5的溶磷、抑菌与促玉米生长作用研究

作  者：
宫安东;朱梓钰;路亚南;万海燕;吴楠楠;Cheelo Dimuna;龚双军;文淑婷;侯晓
单  位：
信阳师范学院生命科学学院/河南省茶树生物学重点实验室
关键词：
溶磷;气体抑菌;吡咯伯克霍尔德菌;二甲基二硫;微生物肥料;玉米
摘  要：
[目的]筛选兼具高效溶磷和抑菌作用的微生物,检测其溶磷效果和抑菌活性,鉴定抑菌代谢产物,并分析筛选微生物对植物生长的作用,为新型多功能抑菌微生物菌肥的研发提供材料.[方法]采集信阳毛尖茶车云山茶厂百年龄茶树根际土壤,稀释后涂布难溶性无机磷或难溶性有机磷培养基表面,培养后检测溶磷活性,测定溶磷圈直径,筛选具有高效溶磷作用的微生物,进行后续溶磷效果分析.高效溶磷菌WY6-5接种于培养液和土壤中,检测不同培养时间下,可溶性磷含量的变化规律,分析菌株WY6-5的溶磷活性;玉米盆栽土壤中接种菌株WY6-5菌液,种植27 d后分析玉米植株长势,检测溶磷菌WY6-5对苗期玉米生长的影响;采用双皿对扣培养法,验证菌株WY6-5产挥发性物质的抑菌作用,检测其对不同病原真菌的广谱抑菌效果,气相色谱串接质谱(GC-MS/MS)分析挥发性代谢物质,鉴定主效抑菌成分.[结果]筛选到3个兼具有降解难溶性无机磷和有机磷作用的微生物菌株,尤以菌株WY6-5溶磷效果最优.培养基培养条件下,对难溶性无机磷的溶解直径达2.3 cm,溶磷圈直径与菌落直径比为4.6;对难溶性有机磷溶解直径3.6 cm,溶磷圈直径与菌落直径比达7.2.表型观察、生理生化鉴定和系统发育树分析表明,菌株WY6-5为乳白色细菌,16S rRNA序列与Burkholderia pyrrocinia CIP 105874和Burkholderia stabilis CIP 106845两个菌株的同源性最高,进化树中聚成独立一支.另外,WY6-5与Burkholderia pyrrocinia具有高度相同的生理生化反应结果.因此,本研究将WY6-5鉴定为吡咯伯克霍尔德菌(Burkholderia pyrrocinia).WY6-5在液体培养和土壤中均具有较好的溶磷活性,20 d培养时间内,液体培养液中磷含量最高达520.4 mg·L-1,为对照组176倍;土壤试验3-20 d期间,WY6-5处理组可溶性磷含量均高于对照组,且在盆栽试验中,能高效促进苗期玉米植株的生长,处理组叶长、叶宽、叶片数、茎粗、株高、鲜重等指标显著优于对照组;同时,菌株WY6-5还可产生挥发性抑菌物质,高效广谱抑制8种重要病原真菌的生长,抑菌率最高达100%,经GC-MS/MS检测发现,挥发性物质含有一种主效抑菌物,相对丰度达97%以上,鉴定为二甲基二硫.[结论]吡咯伯克霍尔德菌(Burkholderia pyrrocinia)WY6-5分离自茶树根际土壤,在培养基、培养液和土壤环境下,均具有高效的溶磷效果,将难溶性的无机磷转化为植物可吸收的可溶性磷,并促进苗期玉米植株的生长;同时该菌还可产生挥发性抑菌物质二甲基二硫,高效抑制8种重要植物病原真菌的生长,抑制率最高达100%.菌株WY6-5兼具有提升土壤磷肥力、促进植物生长和和抑制真菌病害等多种重要作用,具有较好的生物学功能.
作  者：
GONG AnDong;ZHU ZiYu;LU YaNan;WAN HaiYan;WU NanNan;Cheelo Dimuna;GONG ShuangJun;WEN ShuTing;HOU Xiao;College of Life Science,Xinyang Normal University/Henan Key Laboratory of Tea Plant Biology;
单  位：
GONG AnDong%ZHU ZiYu%LU YaNan%WAN HaiYan%WU NanNan%Cheelo Dimuna%GONG ShuangJun%WEN ShuTing%HOU Xiao%College of Life Science,Xinyang Normal University/Henan Key Laboratory of Tea Plant Biology
关键词：
phosphate solubilizing;;volatile antifungal activity;;Burkholderia pyrrocinia;;dimethyl disulfide;;microbial fertilizer;;maize
摘  要：
【Objective】 The study was carried out to screen microbe with phosphate solubilizing(P-solubilizing) and antagonistic activity, to evaluate their efficacies for P-solubilizing, fungal inhibition and plant growth promoting, and to identify antifungal compounds, so as to provide new resources for the development of microbial fertilizers. 【Method】 Rhizosphere soil samples of tea trees were collected from Cheyun mountain factory in Xinyang, Henan, China. Each sample was diluted, and spread onto the surface of insoluble organic and inorganic media. The diameter of P-solubilizing zone was measured after 5 days of incubation. The strain WY6-5 was chosen for further studies because it showed the highest P-solubilizing activity on insoluble phosphate. Additionally, the strain WY6-5 was inoculated in liquid medium and soil for 20 days to test P-solubilizing activity, and inoculated in maize grown soil to test plant growth promoting effects. Moreover, strain WY6-5 was co-cultured with eight different fungi to determine antagonistic activity by using a two dish face-to-face cultural method. The volatiles were characterized and identified with gas chromatography-tandem mass spectrometry(GC-MS/MS). 【Result】 Three strains with P-solubilizing activity were isolated from rhizosphere soils of tea trees, and which were capable of dissolving insoluble organic and inorganic phosphorus medium. The strain WY6-5 demonstrated the highest P-solubilizing activity with the solubilizing halo up to 2.3 cm on insoluble organic medium, and 3.6 cm on insoluble and inorganic phosphorus medium. The ratios of the P-solubilizing halo diameter to the colony diameter on both media were 4.6 and 7.2, respectively. Based on morphological characteristics, physiological, biochemical and phylogenetic analyses, the strain WY6-5 was identified to be Burkholderia pyrrocinia. P-solubilizing activity of WY6-5 was also observed in the liquid medium or in soil after 20 days of incubation. The concentration of soluble phosphate in liquid medium was up to 520.4 mg·L~(-1), which was 176 times higher than that in control treatment. During 3-20 d, the phosphate concentrations in soil under WY6-5 treatments were constantly higher than that under control treatment. In addition, the strain WY6-5 significantly promoted the growth of maize seedling in terms of the number, length, width and area of leaf as well as plant height and fresh weight. Moreover,the volatile compounds produced from the WY6-5 inhibited the growth of all eight different fungi, with the mycelium inhibition rate up to 100%. The antifungal volatile was subsequently identified as dimethyl disulfide through GC-MS/MS. 【Conclusion】 The Burkholderia pyrrocinia strain WY6-5 isolated from rhizosphere soils of a tea tree was found to be able to dissolve insoluble phosphate in both liquid medium and soil, to promote the growth of maize seedlings, and to produce volatile dimethyl disulfide with broad antifungal activity, implying an important biological functions.

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