假禾谷镰孢子囊壳的田间采集和子囊孢子后代室内杂交条件优化

Optimum culture conditions in the lab for sexual stage formation of ascospore isolates from perithecia collected in field for Fusarium pseudograminearum

假禾谷镰孢(Fusarium pseudograminearum,Fp)是我国黄淮麦区小麦茎基腐病(Fusarium crown rot of wheat)的优势病原菌,目前该病害呈不断蔓延和为害加重的趋势。本研究从五月中旬小麦灌浆期至玉米收获期,在河南开封和焦作田间调查采集小麦茎基腐病样,对13份小麦茎秆和小麦残茬上带子囊壳的病样在室内进行单子囊孢子分离培养,获得纯化菌株327个,用形态学和分子生物学方法鉴定均为假禾谷镰孢,交配型MAT1-1和MAT1-2各有164株和163株。对国外报道的有性态诱导方法进行了改进和优化,用不同交配型菌株在Sachs合成固体培养基配以水稻茎节对峙培养,在黑光灯和冷白光灯交替光照及22℃条件下,温箱培养60 d左右,KF86×KF73组合在水稻茎节处产生大量子囊壳和成熟子囊孢子。本研究利用假禾谷镰孢子囊孢子后代进行室内有性杂交实验,通过改进诱导子囊壳产生的培养条件,获得高育性菌株,为今后假禾谷镰孢基因功能的有性生殖阶段研究提供材料和方法。

Fusarium pseudograminearum(Fp)is one of the main pathogens of wheat Fusarium crown rot(FCR)that has been spreading and aggravating in Huanghuai winter region.In this study,from the wheat grain filling period in mid-May to the harvest of maize in Kaifeng and Jiaozuo,Henan Province,we invested and collected the sick wheat plants and 13 stubbles with perithecia in the field.The single ascospore was isolated from ascus of perithecia.Based on morphological and molecular characteristics,a total of 327 single spore cultures were identified to be F.pseudograminearumin which 164 and 163 strains belonged to MAT-1 and MAT-2,respectively.To improve sexual stage formation in the laboratory,the mating experiment was carried out using the ascospore cultures with opposite mating type to obtain the optimum conditions started from that reported abroad.The result of mating experiment showed that using Sachs synthetic medium with rice stem node,alternate black and cold white light at 22℃for about 60 days,the KF86×KF73 combination produced a large number of mature perithecia and ascospores.Collectively,we used the ascospore cultures to set up mating experiment in the laboratory and obtained the optimized conditions and high fertile strains which provide methods and material for future study on gene functions at the sexual reproduction stage.