河南省番茄灰霉病菌对3种杀菌剂的抗药性检测

为了明确河南省番茄灰霉病菌对苯并咪唑类杀菌剂多菌灵、二甲酰亚胺类杀菌剂腐霉利和氨基甲酸酯类杀菌剂乙霉威的抗药性状况,2013年从河南省不同保护地中采集番茄灰霉病病果或病叶,经单孢分离共获得番茄灰霉病菌菌株139株。采用最低抑制浓度法（MIC）测定了其对多菌灵、腐霉利和乙霉威的抗药性。结果显示：番茄灰霉病菌对多菌灵、腐霉利和乙霉威产生抗性菌株的频率分别为81. 29%、80.58%和93.53%; 所测菌株对3类杀菌剂的抗性类型有BenRDicSNPCS、BenRDicRNPCS、BenRDicSNPCR、BenSDicRNPCR、BenSDicSNPCR和BenRDicRNPCR 6种,所占比例分别为2. 88%、3.60%、3.60%、5.04%、13.67%和71. 22%。表明河南省番茄灰霉病菌已对多菌灵、腐霉利和乙霉威产生抗药性,迫切需要筛选新的杀菌剂防治番茄灰霉病。     

樱桃采后灰霉病菌对甲基硫菌灵、乙霉威和腐霉利的抗性

本文采用单孢分离法对四川汉源和山东烟台等地采集的樱桃果实进行了采后灰霉病的病原菌分离和鉴定;采用区分剂量法分别测定了菌株对苯并咪唑类杀菌剂甲基硫菌灵、乙霉威和二甲酰亚胺类杀菌剂腐霉利的敏感性,并进一步分析了抗药性菌株的分子机制。结果表明,分离得到的54株樱桃采后灰霉病菌均为灰葡萄孢Botrytis cinerea,对甲基硫菌灵的总抗性频率高达79.6%,其中甲基硫菌灵抗性-乙霉威敏感 (BEN R1) 菌株频率为 25.9%;甲基硫菌灵-乙霉威双重抗性菌株 (BEN R2) 频率为53.7%;检测到腐霉利抗性菌株 (DCF R) 9 株,频率为16.7%。甲基硫菌灵抗性菌株在β-tubulin基因上的突变共有2种类型: BEN R1抗性菌株中,第198位密码子发生点突变 (GAG→GCG),编码氨基酸由Glu (E)突变成缬氨酸Ala (A);在BEN R2抗性菌株中,第198位密码子发生点突变 (GAG→GTG),编码氨基酸由Glu (E)突变成缬氨酸Val (V)。DCF R菌株在BcOS1的第365位密码子由ATC突变成AAC或AGC,导致编码的氨基酸由异亮氨酸Ile (I)突变成天冬酰胺Asn (N)或丝氨酸Ser (S)。本研究表明樱桃采后灰霉病菌对甲基硫菌灵和腐霉利存在不同程度抗性,应在加强抗药性监测的同时与其他类型杀菌剂交替使用,延缓抗药性发展。     

不同抗性型灰葡萄孢菌Botrytis cinerea对不同作用机制杀菌剂的敏感性研究

不同类型杀菌剂对灰葡萄孢菌菌株的离体、活体毒力测定结果表明 ,苯并咪唑类的多菌灵、甲基硫菌灵之间 ,二甲酰亚胺类的腐霉利、异菌脲、菌核净之间表现交互抗性 ;苯并咪唑类和 N -苯氨基甲酸酯类乙霉威表现有负交互抗性趋势。离体测定中 ,对不同类型菌株以吡咯类杀菌剂咯菌腈 (EC50 200mg/ L) ,咯菌腈以及甾醇抑制剂戊唑醇、氟硅唑和丙环唑均表现出较高的活性。     

设施蔬菜灰霉病菌对不同类型杀菌剂的抗性检测

为了明确设施蔬菜灰霉病菌Botryotinia fuckeliana的抗药性现状,采用菌丝生长速率法检测了20052006年采自浙江、江苏、山东和辽宁4省的144个菌株对6种常用防治药剂的敏感性。结果表明,灰霉病菌对百菌清已经产生了低水平的抗性,频率为5.56%,其对多菌灵的抗性非常严重,总的抗性频率为43.05%,高抗(HR)频率为27.08%;乙霉威的EC50值在0.137728.9μg/mL之间,平均为40.06μg/mL。其中多菌灵-乙霉威双抗频率为36.11%,且首次检测到了两种新的双抗类型。二甲酰亚胺类杀菌剂在生产上已经应用近20年,但灰霉病菌对异菌脲和腐霉利只有频率为20%左右的低水平抗性,没有检测到高抗菌株;苯胺嘧啶类杀菌剂嘧霉胺虽然只应用几年时间,但已经产生了抗性,其抗性菌株频率为4.16%。研究表明,设施蔬菜灰霉病菌对常用的6种防治药剂均产生了不同程度的抗性。     

番茄叶霉病菌异菌脲抗药性突变体的诱导与生物学性状

测定了苯并咪唑类杀菌剂敏感-乙霉威抗性(BenS-DieR)、苯并咪唑类杀菌剂抗性-乙霉威敏感(BenR-DieS)和苯并咪唑类杀菌剂抗性-乙霉威抗性(BenR-DieR)3种类型的番茄叶霉病菌Cladosporium fulvum菌株对不同类型药剂的敏感性。结果表明,蕃茄叶霉病菌对供试药剂的敏感性与其对苯并咪唑类杀菌剂及乙霉威的敏感性无关。根据药剂对3类菌株EC50值的平均值, 16种杀菌剂抑制菌丝生长的活性依次为腐霉利＞乙烯菌核利＞异菌脲＞戊唑醇＞百菌清＞嘧霉胺＞醚菌酯＞代森锰锌＞8-羟基喹啉铜＞丙环唑＞苯醚甲环唑＞嘧菌酯＞灭锈胺＞烯酰吗啉＞烟酰胺＞三唑酮;抑制孢子萌发的活性依次为醚菌酯＞腐霉利＞百菌清＞乙烯菌核利＞灭锈胺＞8-羟基喹啉铜＞异菌脲＞代森锰锌＞嘧菌酯＞烟酰胺＞嘧霉胺＞戊唑醇＞丙环唑＞苯醚甲环唑＞三唑酮＞烯酰吗啉。通过紫外诱变共获得17株抗异菌脲突变体,突变频率为4.5×10-7。其中低抗、中抗和高抗菌株分别占 17.65%、70.59%和11.75%。这些突变体对腐霉利和乙烯菌核利表现交互抗性,对苯并咪唑类、脱甲基抑制剂(DMIs)、QoIs等药剂的敏感性与亲本菌株之间没有显著性差异,与亲本菌株在生长、产孢、致病能力等方面也无显著差异,但对渗透胁迫的敏感性要显著高于亲本。     

湖南省草莓灰霉病菌对4种杀菌剂的抗药性检测

为明确湖南省草莓灰霉病菌菌株的抗药性状况,2013-2015年从湖南省不同地区草莓灰霉病病果或病叶上经单孢分离获得草莓灰霉病菌454株,采用最小抑制浓度法(MIC)检测不同地区草莓灰霉病菌株对多菌灵、腐霉利、异菌脲、嘧霉胺的抗药性。结果表明:草莓灰霉病菌对多菌灵、腐霉利、异菌脲、嘧霉胺的抗性频率分别为55.73%,77.31%、3.52%和77.31%;所测菌株对4种杀菌剂的敏感性类型有Cbz~RPcm~RIpd~RPmt~R、CbzSPcm~RIpd~RPmt~R、Cbz~RPcm~RIpdSPmt~R、Cbz~SPcm~RIpdSPmt~S、Cbz~RPcm~SIpd~SPmt~R、Cbz~SPcm~RIpd~SPmt ~R等6种,其所占比例分别为33.26%、5.07%、41.41%、4.63%、3.96%、11.67%,未发现对4种杀菌剂均敏感的类型,表明湖南省草莓灰霉病菌已对多菌灵、腐霉利和嘧霉胺产生抗药性,对异菌脲的抗药性较低。     

灰霉病菌对三种杀菌剂的抗性表现型分布及稳定性测定

采用最低抑制浓度法(MIC)和菌落直径法,对采自山西晋中、晋南、晋东南和晋北4个地区的188个灰霉病菌株对3种杀菌剂的抗性表现型和稳定性进行了测定,旨在了解灰霉病菌的抗药性状况和预测其抗性发展趋势。结果表明:对多菌灵的抗性表现型可分为敏感(S)、低抗或中抗(RM)和高抗(R),对腐霉利和乙霉威可分为敏感(S)和抗性(R)。以对多菌灵、腐霉利和乙霉威的抗性为序,检测出8种不同类型的抗性表现型,即RRR、RRS、RSR、RSS、RMRS、RMSR、RMSS 和SSR。各表现型的分布频率随各地区用药历史和水平的不同而变化,并与抗性监测结果相一致。对55个不同抗性表现型的单孢菌株的稳定性测定结果表明:继代无药培养10代后,以RRR和RSS型表现最稳定,而其余类型菌株均不同程度发生了变异,尤其是RMSS和RMRS型菌株100%发生了变异,抗性表现型不稳定的菌株占52.7%。对多菌灵的抗性变化趋势是由敏感向低抗、再向高抗发展;对腐霉利的抗性变化主要是由抗性转变为敏感;而对乙霉威的抗性则相对比较稳定。     

蔬菜灰霉病菌对腐霉利抗药性变异及其治理

蔬菜灰霉病菌（Botrytis cinerea）分生孢子经紫外光灯照射，可获得10^-4～10^-5频率的腐霉利抗药性突变体。获得的13个抗性突变体的抗性水平平均EC50值为18.45ug/ml,EC90值为305.17ug/ml。抗性突变体的可溶性蛋白谱带比其亲本敏感菌株多2～4条，其中Rf=0.38的蛋白谱带是抗性菌株所拥有而敏感菌株所共同缺乏的。抗性菌株对腐霉利抗药性稳定遗传，具有与亲本菌株相同的致病性。苗期抗、感测定结果表明，灰霉克杀菌剂对腐霉利突变体具有明显的克抗作用。田间抗性菌治理使用灰霉克400～800倍对大棚番茄灰霉病平均防效为95.93%～75.8%,优于腐霉剂600倍的平均防效(61.32%)。     

番茄叶霉病菌对多菌灵、乙霉威及代森锰锌抗性检测

报道了番茄叶霉病菌Fulvia fulva对多菌灵、乙霉威及代森锰锌的敏感性基线,以及抗性频率和抗性水平。离体条件下,多菌灵、乙霉威及代森锰锌对番茄叶霉病菌敏感菌株的平均EC50值分别为0.101、2.475、9.067 μg/mL;最低抑制浓度(MIC)值分别为0.5、5、50 μg/mL 。山西晋南地区番茄叶霉病菌对3种杀菌剂的抗性频率最高,分别达到97.4%、70.5%、98.7%;山西吕梁地区与太原地区相对较低,但该病菌对3种杀菌剂的抗性频率也都超过了30%。辽宁沈阳、山东寿光、河北保定番茄叶霉病菌对多菌灵的抗性频率均为100%;对乙霉威的抗性频率前两地为100%,保定为10%;对代森锰锌的抗性频率都超过90%。所有抗性菌株对多菌灵均属于高抗类型,抗性指数超过5000,测不出MIC值;对乙霉威有50%的高抗菌株,抗性指数在100以上;对代森锰锌各地均未发现高抗菌株,低抗和中抗菌株所占比例较大,其抗性指数集中在50左右。对多菌灵与乙霉威具有双重抗性的菌株占测试菌株总数的49.9%,并且首次在田间发现了对3种杀菌剂都具有抗性的番茄叶霉病菌多抗菌株。     

49种植物源粗提物对黄瓜灰霉菌的生物活性筛选

黄瓜灰霉病是由灰霉菌Botrytis cinerea引起的重要病害.目前,尚无理想的抗灰霉病黄瓜品种,药剂防治仍然是控制该病害的必要措施.20世纪80年代以前防治灰霉病的杀菌剂如苯并咪唑类的多菌灵、硫代氨基甲酸酯类的甲基托布津、二羧酰亚胺类的异菌脲、腐霉利等已使用了近30年,灰霉病菌对这些药剂均产生了不同程度的抗药性.作者测定了植物源粗提物对黄瓜灰霉病菌的活性,以期研制开发出无公害植物源杀菌剂.     

Distribution and fitness of isolates of Botrytis cinerea with multiple fungicide resistance in Spanish greenhouses

Forty-nine greenhouses of vegetable crops were surveyed in southeast Spain at the beginning of the disease season in December 1992 to estimate frequencies of resistance to benzimidazoles, dicarboximides and N -phenylcarbamates (NPC) in B. cinerea . Out of 261 isolates collected, 28% were sensitive to both benzimidazoles and dicarboximides, 15% were benzimidazole-resistant and dicarboximide-sensitive, 8% were benzimidazole-sensitive and dicarboximide-resistant and 46% were benzimidazole- and dicarboximide-resistant. Resistance to benzimidazole, dicarboximide and N -phenylcarbamate was determined by measuring the ability of each isolate to grow in the presence of carbendazim, procymidone and diethofencarb fungicides respectively. Carbendazim- or procymidone- resistant isolates were found in all surveyed greenhouses. Three isolates were found with resistance to carbendazim, procymidone and diethofencarb collected in two adjacent greenhouses that were sprayed with the carbendazim and diethofencarb mixture. All other isolates were sensitive to the mixture because they were either sensitive to carbendazim and resistant to diethofencarb or vice versa. Fitness of 31 isolates of B. cinerea was determined in vivo by measuring their sporulation and lesion growth rate on leaf disks. No fitness costs were associated with resistance to iprodione (dicarboximide) and benomyl (benzimidazole). Isolates with EC₅₀ values higher than 101 mg/L for benomyl and 1.6 mg/L for iprodione were considered to be field resistant (they caused visible lesions on cucumber leaf disks treated with each fungicide).     

Detection and characterization of benzimidazole resistance of <Emphasis Type="Italic">Botrytis cinerea</Emphasis> in greenhouse vegetables

From 2003 to 2006, a total of 426 single-conidial isolates of B. cinerea collected from greenhouse vegetables in China were characterized for resistance to benzimidazole fungicides and diethofencarb according to inhibition of mycelial growth. Rapid development of double-resistance to benzimidazoles and diethofencarb was observed. Three types of benzimidazole-resistant isolates, Ben R1, Ben R2 and Ben R3 were detected. A new phenotype, Ben R3, which showed low level of resistance to benzimidazole fungicides and resistance to diethofencarb, was detected with frequencies of 6.8%, 10.0%, 13.2% and 12.4% from 2003 to 2006, respectively. Further studies indicated that Ben R3 was caused by a point mutation from GAG in sensitive(S) isolates to GTG at codon 198 in the β-tubulin gene, predicted to cause a change from glutamic acid to valine. Ben R3 isolates had comparable growth, sporulation and pathogenicity ability as isolates of other phenotypes but were more sensitive at lower temperatures.     

Multiple fungicide resistance to benzimidazoles, dicarboximides and diethofencarb in field isolates of Botrytis cinerea in Israel

During the period January-March 1989, 15 greenhouses at 12 sites in Israel were surveyed for the presence of fungicide-resistant strains of Botrytis cinerea , using a fungicide-amended Botrytis -selective medium. Resistance to benzimidazoles (Ben^R) and to dicarboximides (Dic^R) was frequent in most sites. Resistance to carbendazim + diethofencarb (Ben^R NPC^R) was found in all eight sites in which a mixture of these fungicides had been used against grey mould, but not in other sites. A new phenotype of multiple fungicide resistance was found among these isolates. The new phenotype, designated Ben^R Dic^R NPC^R, combines the three previously described characteristics of resistance to benzimidazole, dicarboximide and N -phenylcarbamate fungicides. It was found only in cucumber greenhouses that had been sprayed with the fungicide mixture carbendazim + diethofencarb against grey mould. Isolates of this phenotype were pathogenic in artificial inoculation of cucumber cotyledons treated with carbendazim, iprodione or carbendazim + diethofencarb.     

Monitoring and characterization of resistance development of strawberry Phomopsis leaf blight to fungicides

From 2005 to 2009, a total of 479 single-conidial isolates of Phomopsis obscurans were collected from strawberry. The isolates were characterized for their resistance to benzimidazole fungicides, diethofencarb, and sterol demethylation inhibitors (DMIs). Low-level DMI resistant isolates (DMI-LR) and two types of benzimidazole-resistant (Ben R) isolates, Ben R1 (benzimidazole-resistant and diethofencarb -sensitive) and Ben R2 (benzimidazole-resistant and diethofencarb -resistant), were detected. Both Ben R and DMI-LR isolates exhibited comparable growth, sporulation, and pathogenicity with the sensitive isolates. No significant difference in growth at low temperature was observed between Ben R and benzimidazole-sensitive (Ben S) isolates. Ben R1 was caused by a point mutation from GAG to GTG at codon 198 in the β-tubulin gene in Ben S isolates, predicted to cause a change from glutamic acid to valine. Ben R2 was induced by a point mutation from TTC to TAC at codon 200 in the β-tubulin gene in Ben S isolates.     

安徽省番茄灰霉菌抗药性测定和治理

测定了安徽省番茄灰霉病菌对多菌灵、腐霉利、乙霉威的抗性。结果表明,田间有B_SD_SN_S、B_SD_SN_LR、B_SD_SN_HR、B_HRD_SN_S、B_MRD_LRN_S、B_HRD_LRN_S、B_HRD_LRN_LR7种抗性类型,对多菌灵、腐霉利、乙霉威具有抗性的菌株分别占试验总菌株数的28.3%、20%和73.3%。     

Negative cross resistance between benzimidazole and N-phenylcarbamate fungicides and control of Botrytis cinerea on grapes

Isolates of Botrytis cinerea resistant to benzimidazoles (Ben^R), dicarboximides (Dic^R), or both (Ben^R Dic^R) were found on table grapes in vineyards in Israel. In vineyards treated for one or two seasons with benomyl and dicarboximides, 41% of the isolates were benomyl-resistant, 18% were dicarboximide-resistant and 21 % were resistant to both fungicides. The frequency of resistant strains was very low in non-treated vineyards. Negatively correlated cross resistance (NCCR) was manifested by 46 Ben^R isolates which were sensitive to isopropyl N -(3,4-diethoxyphenyl) carbamate (NPC) and 73 benomyl-sensitive (Ben^s) isolates which were insensitive to NPC; NCCR was not influenced by sensitivity to dicarboximides. A mixture of Ben^s Dic^R plus Ben^R Dic^s isolates, inoculated on rose petals, was inhibited by mixtures of benzimidazoles plus NPC but not by benomyl, NPC or iprodione alone. Grey mould, incited on maturing grape berries by a vineyard population composed of Ben^s and Ben^R strains, was not controlled by benomyl, while the mixture of NPC plus carbendazim was effective.