北京地区草坪草夏季斑枯病菌对嘧菌酯的敏感性及其Cytb序列分析
投稿时间:2017-11-16      点此下载全文 HTML
引用本文:胡健,杨静雅,李婕,杨高文,任海彦,等.北京地区草坪草夏季斑枯病菌对嘧菌酯的敏感性及其Cytb序列分析[J].农药学学报,2018,20(1):33-40.
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作者单位E-mail
胡健 南京农业大学 草业学院, 南京 210095  
杨静雅 南京农业大学 草业学院, 南京 210095  
李婕 南京农业大学 草业学院, 南京 210095  
杨高文 南京农业大学 草业学院, 南京 210095  
任海彦 南京农业大学 草业学院, 南京 210095 hren@njau.edu.cn 
基金项目:国家自然科学基金(31701810);江苏省自然科学基金(BK20160725)
中文摘要:采用菌丝生长速率法,测定了水杨肟酸(SHAM)对草坪草夏季斑枯病菌Magnaporthe poae菌丝生长及其对嘧菌酯敏感性的影响,并在此基础上测定了采自北京地区的114株夏季斑枯病菌对嘧菌酯的敏感性;扩增并分析了夏季斑枯病菌细胞色素b基因(Cytb)部分序列,以期探寻该病原菌对嘧菌酯敏感性下降的分子机制。结果显示:不同浓度SHAM对菌丝生长的抑制作用差异显著(F = 20.812,P < 0.000 1);添加SHAM(40 μg/mL)与否,嘧菌酯对菌丝生长的抑制中浓度(EC50)无显著差异(F = 0.041 0,P = 0.842 3)。嘧菌酯对供试114株菌株的EC50值范围为0.001 0~0.146 5 μg/mL,平均值为(0.017 4 ± 0.026 1)μg/mL。供试夏季斑枯病菌对嘧菌酯的敏感性频率不符合正态性分布(W = 0.499 7, P < 0.05),表明群体中已出现对嘧菌酯敏感性下降的亚群体;连续单峰频次分布为70.18%的菌株(W = 0.970 8,P = 0.064 0 > 0.05)的平均EC50值为(0.007 8 ± 0.003 7)μg/mL,该值可作为夏季斑枯病菌对嘧菌酯的相对敏感性基线。比对分析获得夏季斑枯病菌Cytb的部分序列(8 639 bp),进一步扩增发现对嘧菌酯敏感性存在差异的12株菌株均在该序列上紧临143位密码子处插入了一段长1 956 bp的内含子,且在Cytb的129、137和143位密码子上未出现与抗性相关的点突变。上述结果可为草坪草夏季斑枯病的有效防治提供依据,同时可为深入探寻病原菌对甲氧基丙烯酸酯类(QoIs)杀菌剂敏感性下降的分子机制提供参考。
中文关键词:草坪草  夏季斑枯病  夏季斑枯病菌  嘧菌酯  敏感性  抗性  细胞色素b
 
Azoxystrobin sensitivity in Magnaporthe poae populations collected from turfgrass in Beijing and analysis of its cytochrome b (Cytb) gene sequence
Abstract:The effect of salicylhydroxamic acid (SHAM) on mycelial growth of Magnaporthe poae from turfgrass and in vitro azoxystrobin sensitivity were determined by mycelial growth rate method. Thereafter, the sensitivities of 114 M. poae isolates to azoxystrobin were determined by the same method. The partial sequence of M. poae cytochrome b (Cytb) gene was cloned and analyzed to investigate the molecular mechanism of the decreased sensitivity to azoxystrobin in M. poae. The results showed that the inhibition of mycelial growth by SHAM solution of different concentrations was significantly different (F = 20.812, P < 0.000 1). Sensitivity of M. poae to azoxystrobin amended with SHAM (40 μg/mL) was not significantly different from that without SHAM (F = 0.041 0, P = 0.842 3). The effective concentration for 50% inhibition of mycelial growth (EC50) values of the tested isolates ranged from 0.001 0-0.146 5 μg/mL, and the average of EC50 values was (0.017 4 ± 0.026 1) μg/mL. The frequency of azoxystrobin sensitivity in M. poae was not normally distributed (W = 0.499 7, P < 0.05), suggesting that sub-populations of decreased sensitivity to azoxystrobin in M. poae emerged. The majority of M. poae isolates (70.18%) have the frequency of EC50 values with unimodal curve distribution (W = 0.970 8, P = 0.064 0 > 0.05), and the mean EC50 value of these isolates was (0.007 8 ± 0.003 7) μg/mL, which can be used as a relative sensitivity baseline. A partial sequence of M. poae Cytb gene (8 639 bp) was obtained by aligning to the reference mitochondrial genome of M. poae. Sequence analysis revealed the presence of a 1 956 bp intron immediately downstream of codon 143. The intron was identified in all 12 isolates of M. poae Cytb with different sensitivity to azoxystrobin, and no point mutations were observed at codons 129, 137 and 143, which were responsible for azoxystrobin resistance reported previously. The results will lay a scientific basis for the effective management of summer patch disease and provide a useful reference for further investigating the molecular mechanism of the decreased sensitivity to QoIs in M. poae.
Key words:turfgrass  summer patch  Magnaporthe poae  azoxystrobin  sensitivity  resistance  cytochrome b
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