Real-time PCR法定量检测柑橘绿霉病菌对抑霉唑的抗性频率

 抑霉唑被广泛用来防治由指状青霉菌(Penicillium digitatum)引起的柑橘绿霉病。已有研究表明,柑橘绿霉病菌对抑霉唑的抗性由CYP51基因启动子区5个126-bp转录增强子的简单串联重复和126-bp转录增强子上199-bp的特异性片段插入所引起。基于这2种抗性分子机制,通过设计特异引物和优化条件,建立real-time PCR高通量分子检测技术,用于快速检测柑橘包装贮藏库中绿霉病菌群体对抑霉唑的抗性频率F_R,指导科学用药。     

抑霉唑、咪鲜胺与指状青霉CYP51的结合模式及交互抗性机制分析

为解析指状青霉Penicillium digitatum对抑霉唑和咪鲜胺的抗性机制和交互抗性机制,基于指状青霉甾醇14α-去甲基化酶（sterol 14α-demethylases,CYP51）CYP51A和CYP51B的氨基酸序列,采用同源建模的方法构建其三维结构,根据已报道的可能与抑霉唑抗性相关的突变位点CYP51A-Y308H及与咪鲜胺抗性相关的突变位点CYP51B-Y136H、CYP51B-Q309H和CYP51BF506I构建突变模型,采用分子对接法分析其突变前后与抑霉唑、咪鲜胺的结合模式,并进行氨基酸序列保守性分析。结果显示,抑霉唑和咪鲜胺与指状青霉CYP51的结合无特异性,两者均能与其CYP51A和CYP51B结合。指状青霉的CYP51A-Y308氨基酸残基及CYP51A-Y308H突变体氨基酸残基均不能与抑霉唑和咪鲜胺形成相互作用力,说明该突变与指状青霉对抑霉唑和咪鲜胺的抗性无关。指状青霉的CYP51B-Y136H突变导致其与咪鲜胺和抑霉唑的亲和力下降;指状青霉的CYP51B-Y136氨基酸位于CYP51B蛋白的保守区域,其产生的与咪鲜胺和抑霉唑抗性相关的随机突变在药剂...     

山东蓬莱葡萄灰霉菌对7种杀菌剂的抗药性检测

为了明确葡萄灰霉菌对啶酰菌胺、多菌灵、咯菌腈、异菌脲、腐霉利、嘧霉胺的抗药性和对抑霉唑的敏感性,本试验采用菌丝生长速率法和孢子萌发法检测了采自山东蓬莱地区的69株葡萄灰霉菌对上述前6种杀菌剂的抗药性、对抑霉唑的敏感性及抑霉唑与其他6种杀菌剂的交互抗性关系。结果表明,抑霉唑对这69株葡萄灰霉菌的EC₅₀分布在0.403～28.76μg/mL之间,平均值为(9.34±10.34)μg/mL;葡萄灰霉菌菌株中抗啶酰菌胺(BosR)、多菌灵(CarR)、咯菌腈(FluR)、异菌脲(IprR)、嘧霉胺(PyrR)、腐霉利(ProR)的比例分别为100%、100%、9.47%、97.18%、100%、89.20%,测试菌株的抗药性均为多抗类型,没有单抗菌株,其中对3种杀菌剂(啶酰菌胺、多菌灵、嘧霉胺)、对4种杀菌剂(啶酰菌胺、多菌灵、异菌脲、嘧霉胺)、对5种杀菌剂(啶酰菌胺、多菌灵、异菌脲、嘧霉胺、腐霉利或啶酰菌胺、多菌灵、咯菌腈、异菌脲、嘧霉胺)和对6种杀菌剂(啶酰菌胺、多菌灵、咯菌腈、异菌脲、嘧霉胺、腐霉利)的抗性频率分别为2.33%、9.30%、79.07%、2.33%、6.97%,表明啶酰菌胺、多菌灵、嘧霉胺对测试葡萄菌株完全丧失防效,建议在该葡萄产区停止使用这些药剂,测试菌株对腐霉利、异菌脲的抗性频率高,建议采取限制使用、禁止单独使用等措施,测试菌株对咯菌腈的抗性频率较低,可以继续使用但需按照科学使用规则进行。抑霉唑与其他6种杀菌剂间不存在交互抗性关系,说明其可以和其他药剂同时使用但建议减少使用。     

采用表面增强拉曼光谱技术快速检测脐橙果皮中抑霉唑残留

由于采后处理过程中脐橙保鲜剂抑霉唑易通过果皮渗进果肉中残留,不慎食用后会对人体产生危害。因此,本研究探索一种基于表面增强拉曼光谱技术（Surface-Enhanced Raman Spectroscopy,SERS）的脐橙果皮中抑霉唑残留的快速检测方法。首先对SERS检测条件进行优化,分别确定了最优的检测条件为反应时间2 min,金胶加入量400 μL,NaBr作为电解质溶液且加入量为25 μL。基于以上最优检测条件,以自适应迭代惩罚最小二乘法（Adaptive Iterative Reweighted Penalized Least Squares,air PLS）、air PLS+归一化、air PLS+基线校正、air PLS+一阶导数、air PLS+标准正态变量（Standard Normal Distribution,SNV）和air PLS+多元散射校正（Multiplicative Scatter Correction,MSC）处理后的6组光谱数据为研究对象,分别采用这6种光谱预处理法建立支持向量回归（Support Vector Regression,SVR）模型并对预测性能进行比较后发现,air PLS方法所建立模型的预测集相关系数（Coefficient of the Determinant for the Prediction Set,R_P）最大,预测集均方根误差（Root-Mean-Square Error of Prediction,RMSEP）最小。对光谱数据进行主成分分析（Principal Component Analysis,PCA）特征提取,选择前7个主成分得分作为SVR预测模型的输入值。采用SVR、多元线性回归（Multiple Linear Regression,MLR）和偏最小二乘回归（Partial Least Squares Regression,PLSR）三种建模方法分析比较其对应的预测性能,其中SVR模型的预测集R_P可高达0.9156,预测集RMSEP为4.8407 mg/kg,相对标准偏差（Relative Standard Deviation,RPD）为2.3103,表明基于SVR算法对脐橙表面抑霉唑残留的预测值越接近实测值,越能有效提高模型预测准确性。试验结果表明,利用SERS结合PCA及SVR建模,可实现对脐橙果皮中抑霉唑残留的快速检测。     

Stepwise development of laboratory resistance to DMI-fungicides in Penicillium italicum

Isolates ofPenicillium italicum with differential levels of resistance to imazalil were obtained via step-wise mass selection of conidia of the fenarimol-resistant isolate E300-3 on imazalilamended PDA. Three out of five selection steps were successful. The resistance level to imazalil of isolates acquired after the two last selection steps was on average 122 and 197. The differential level of resistance was also apparent in decay control on oranges by imazalil inoculated with the various isolates. The isolates showed a similar cross-resistance pattern to other fungicides which inhibit C-14 demethylation of sterols (DMIs), although the level of resistance to these fungicides was significantly higher. All isolates displayed negatively-correlated cross-resistance to tridemorph and dodine. Most isolates had a normal virulence on oranges. In competition experiments with mixed-inocula of the wild-type and a resistant isolate, the proportion of the wild-type increased in successive generations on untreated oranges and the proportion of the resistant isolate increased on imazalil-treated oranges. The lower competitive ability of the resistant isolate on untreated oranges may be due to a decrease in spore production as compared with the wild-type.Since isolate E300-3 was obtained in two selection steps on fenarimol-amended PDA, the isolates obtained in the last selection steps on imazalil-amended PDA may have at least five different genes for resistance to DMIs. This is consistent with resistance to DMIs being under polygenic control, with the genes involved having an additive interaction, although this is not the only possible explanation of the results.     

1-MCP复合杀菌剂处理对“金红宝”甜瓜常温贮藏品质的影响

为探究不同处理对甜瓜常温贮藏效果的影响,以“金红宝”厚皮甜瓜为试材,研究1-甲基环丙烯(1-MCP)、1-MCP复合杀菌剂抑霉唑处理对甜瓜采后常温贮藏保鲜效果的影响。结果表明,1-MCP和1-MCP复合杀菌剂处理的保鲜效果均显著优于对照处理。其中,在延缓抗坏血酸含量和可滴定酸含量的下降,降低腐烂率、失重率等方面,1-MCP复合杀菌剂处理的保鲜效果优于1-MCP处理。综上所述,相较其他处理,1-MCP复合杀菌剂处理的保鲜效果最好,可以有效保持甜瓜的品质。     

丙硫菌唑与抑霉唑混剂对稻恶苗病菌的联合毒力及田间防效

为探明丙硫菌唑与抑霉唑混剂对稻恶苗病菌的联合毒力及田间防效,开发防治稻恶苗病的新药剂,本研究以藤仓镰孢菌为试验对象,采用菌丝生长速率法分别检测了丙硫菌唑、抑霉唑及其7种配比混剂对藤仓镰孢菌的毒力,并进行了最佳配比制剂的田间药效试验.结果 表明,丙硫菌唑与抑霉唑配比为1∶1时增效系数为1.9778,增效作用最显著.田间药...