广西剑麻黑斑病病原菌鉴定及其生防菌筛选

Pathogen identification and biocontrol microbes screening of sisal black spot in Guangxi

【目的】分离、鉴定一种在广西剑麻叶片上产生圆形、近圆形或长椭圆形黑色凹陷斑块病害的病原菌,并针对该病原菌筛选具有较好防治效果的生防菌,为病害防治提供科学依据。【方法】从广西5个剑麻种植农场采集具有圆形、近圆形或长椭圆形黑色凹陷斑块的病叶,采用组织分离法分离病原菌;采用叶片针刺法接种,进行病原菌致病性测定;通过病原菌形态特征观察和分子生物学方法鉴定病原菌。采用平板对峙培养法和载玻片孢子萌发法研究哈茨木霉(Trichoderma harzianum)菌株GZ-5、深绿木霉(T.atroviride)菌株ST-1、枯草芽孢杆菌(Bacillus subtilis)菌株B11和解淀粉芽孢杆菌(B.amyloliquefaciens)菌株YZ14-3对剑麻黑斑病病原菌菌丝生长和孢子萌发的抑制效果。【结果】从病叶组织中分离出6种真菌,其中编号为JMHB1的菌株分离率最高,达96%;致病性测定结果表明菌株JMHB1为致病菌;依据形态特征和分子生物学方法,将菌株JMHB1鉴定为新暗色柱节孢(Neoscytalidium dimidiatum)。对峙培养结果显示,生防菌菌株B11和YZ14-3可显著抑制菌株JMHB1的菌丝生长(P<0.05),抑菌圈半径分别为12.14和13.22 mm,且2株生防菌株的培养滤液均可导致菌株JMHB1的菌丝隘缩、断裂;生防菌菌株ST-1和GZ-5对菌株JMHB1的拮抗系数为Ⅲ级和Ⅳ级,但其培养滤液对菌株JMHB1的菌丝无抑制作用。菌株JMHB1的孢子可在菌株YZ14-3、B11、ST-1和GZ-5的培养滤液中的萌发,萌发率分别为31.67%,32.37%,68.63%和76.63%。【结论】引起广西剑麻叶片产生圆形、近圆形或长椭圆形黑色凹陷斑块病害的病原菌为新暗色柱节孢[N.dimidiatum(Penz.)Crous&Slipper],这是我国首次报道新暗色柱节孢侵染剑麻引起黑斑病。病害名称暂定为剑麻Neoscytalidium黑斑病。剑麻生产上可选择和搭配使用解淀粉芽孢杆菌、枯草芽孢杆菌、深绿木霉及其商品制剂防治剑麻Neoscytalidium黑斑病。

【Objective】To isolate and identify the pathogen that caused round,subround or long oval hollow black spots on sisal leaves in Guangxi,and to screen out biocontrol microbes with better control effect on the pathogen,and provide scientific basis for the control of this disease.【Method】The diseased leaves with round,subround or long oval hollow black spots were collected from 5 farms in Guangxi.The pathogen was isolated by tissue isolation method.The pathogenicity of pathogen was determined by leaf acupuncture inoculation.The pathogen was identified by observing mor-phological characteristics and molecular biological methods.The inhibitory effects of Trichoderma harzianum GZ-5,T.atroviride ST-1,Bacillus subtilis B11,B.amyloliquefaciens YZ14-3 and their filtrate on mycelial growth and spores germination of the pathogen were studied by plate confrontation culture method and slide spore germination method.【Re-sult】Six kinds of fungi were isolated from diseased tissues,the isolation rate of strain JMHB1 was the highest(96%).JM-HB1 was the pathogenic bacteria according to the pathogenicity test,which was identified as Neoscytalidium dimidiatum based on morphological characteristics and molecular identification.Biocontrol strains B11 and YZ14-3 could significant-ly inhibit the mycelia growth of JMHB1(P<0.05)according to plate confrontation culture method,the radius of inhibition zone were 12.14 mm and 13.22 mm respectively,and their culture filtrates could lead to constriction and break of JMHB1 mycelia.The antagonistic coefficients of ST-1 and GZ-5 against JMHB1 wereⅢandⅣrespectively,but their culture fil-trates had no inhibition effects on JMHB1 mycelia.Spores of JMHB1 could germinate in the culture filtrates of strains YZ14-3,B11,ST-1,GZ-5,the spore germination rates were 31.67%,32.37%,68.63%,and 76.63%,respectively.【Conclusion】The pathogen that causes round,subround or long oval hollow black spots on sisal leaves in Guangxi is iden-tified as N.dimidiatum(Penz.)Crous&Slipper.This is the first report of sisal black spot caused by N.dimidiatum in Chi-na.The disease is temporarily named as Neoscytalidium black spot.B.amyloliquefaciens,B.subtilis,T.atroviride and their commercial preparations can be selected and used to control Neoscytalidium black spot disease in sisal production.