French vineyards provide information that opens ways for effective resistance management of Botrytis cinerea (grey mould)

Resistance to fungicides is an evolutionary process resulting from the selection of advantageous genotypes in naturally diverse populations. Seven fungicide modes of action are authorised to control grey mould caused by Botrytis cinerea on grapevine in France, and five of them have encountered specific resistance, with variable frequencies in populations and possible consequences for field fungicide efficacy. Moreover, multidrug resistance is caused by fungicide efflux and allows a weak resistance towards six unrelated modes of action. Here, a review is given of the fungicide resistance status of B. cinerea in France, particularly in the vineyards of Champagne, which are the most affected. Recently developed resistance and recent findings concerning the associated resistance mechanisms are focused upon in particular. Finally, antiresistance strategies are presented, and examples of managed resistance are discussed in a more general manner with the aim of extending this knowledge to other crops and countries undergoing similar resistance problems. © 2013 Society of Chemical Industry     

4种杀菌剂及其复配剂对番茄灰霉病菌的毒力

采用菌丝生长速率法测定了咯菌腈、氟啶胺、啶酰菌胺和苯醚甲环唑4种杀菌剂及其两元复配剂对番茄灰霉病菌的毒力。结果显示,咯菌腈、氟啶胺、啶酰菌胺和苯醚甲环唑对番茄灰霉病菌的有效抑制中浓度(EC50)分别为:0.018 0、0.018 1、1.896 8和2.087 4μg/mL。复配剂啶酰菌胺∶苯醚甲环唑1∶5、1∶3和1∶1,咯菌腈∶苯醚甲环唑1∶5增效作用最明显;复配剂咯菌腈∶苯醚甲环唑1∶3,咯菌腈∶氟啶胺1∶3,啶酰菌胺∶咯菌腈5∶1,啶酰菌胺∶苯醚甲环唑3∶1具有增效作用,SR值范围为1.5~4.05,其中以复配剂啶酰菌胺∶苯醚甲环唑1∶5增效作用最好,其SR值为4.05;其余不同配比的各组合复配剂具有相加作用,其SR值范围为0.5~1.46。表明咯菌腈、氟啶胺、啶酰菌胺和苯醚甲环唑4种不同作用机制的杀菌剂可以交替或复配使用,以阻止或延缓灰霉病菌抗药性的进一步加剧,为灰霉病的综合防控和抗药性治理提供理论依据。     

Multiple resistance of Botrytis cinerea from kiwifruit to SDHIs,QoIs and fungicides of other chemical groups

BACKGROUND: Botrytis cinerea Pers.: Fr. is a high‐risk pathogen for fungicide resistance development that has caused resistance problems on many crops throughout the world. This study investigated the fungicide sensitivity profile of isolates from kiwifruits originating from three Greek locations with different fungicide use histories. Sensitivity was measured by in vitro fungitoxicity tests on artificial nutrient media. RESULTS: Seventy‐six single‐spore isolates were tested for sensitivity to the SDHI fungicide boscalid, the QoI pyraclostrobin, the anilinopyrimidine cyprodinil, the hydroxyanilide fenhexamid, the phenylpyrrole fludioxonil, the dicarboxamide iprodione and the benzimidazole carbendazim. All isolates from Thessaloniki showed resistance to both boscalid and pyraclostrobin, while in the other two locations the fungal population was sensitive to these two fungicides. Sensitive isolates showed EC₅₀ values to boscalid and pyraclostrobin ranging from 0.9 to 5.2 and from 0.04 to 0.14 mg L^?1 respectively, while the resistant isolates showed EC₅₀ values higher than 50 mg L^?1 for boscalid and from 16 to > 50 mg L^?1 for pyraclostrobin. All QoI‐resistant isolates carried the G143A mutation in cytb. Sensitivity determinations to the remaining fungicides revealed in total eight resistance phenotypes. No isolates were resistant to the fungicides fenhexamid and fludioxonil. CONCLUSION: This is the first report of B. cinerea field isolates with resistance to both boscalid and pyraclostrobin, and it strongly suggests that there may be a major problem in controlling this important pathogen on kiwifruit. Copyright © 2010 Society of Chemical Industry     

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。     

新杀菌剂对番茄灰霉病菌的室内毒力及田间防效

室内采用菌丝生长速率法、孢子萌发法和黄瓜子叶法测定了咯菌腈、抑霉唑、福美双、吡唑醚菌酯和嘧霉胺对番茄灰霉病菌( Botrytis cinerea)的抑制作用。结果表明,菌丝生长速率法和孢子萌发法的测定均以咯菌腈毒力最高,EC 50分别为0.005 2 μg/mL和0.087 6 μg/mL;黄瓜子叶法测定以抑霉唑毒力最高,其EC 50为0.675 3 μg/mL。田间药效试验结果表明：50%咯菌腈可湿性粉剂90 g/ hm2对番茄灰霉病的田间防效达到90%以上,显著高于对照药剂50%啶酰菌胺水分散粒剂300 g/hm2和40%嘧霉胺悬浮剂 480 g/hm2的防效。     

山西省蔬菜灰霉病菌对嘧霉胺的抗药性检测

为明确山西省蔬菜灰霉病菌对嘧霉胺的抗药性,2010-2011年用单孢分离法从山西省数十个县、市蔬菜主产区采集、分离到171株灰霉病病原菌(Botrytis cinerea),并采用最低浓度抑制法测定各菌株对嘧霉胺的敏感性。结果表明,山西省灰霉病菌对嘧霉胺的平均抗性频率为50.29%,平均抗性倍数为1 423.4。在山西省不同地区灰霉病菌对嘧霉胺的抗性存在差异,其中嘧霉胺使用较多的朔州地区抗药性频率达到100%。     

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%。     

草莓灰霉病菌对嘧菌酯的抗性检测及抗性菌株的生物学特性研究

由灰葡萄孢(Botrytis cinerea Pers.)引起的草莓灰霉病是严重危害草莓的病害之一。为了解草莓灰霉病菌对杀菌剂嘧菌酯的抗性情况,本研究针对2012年从国内10省市采集和分离的134株草莓灰霉病菌菌株,利用特异性引物BcAR-F/BcAR-R扩增其cytb基因的部分序列,进行了嘧菌酯抗性的分子检测;对检测出的抗性菌株在嘧菌酯浓度分别为22.4、112和224μg·mL~(-1)的含药培养基上进行抗性验证;此外,还比较了抗性菌株与敏感菌株主要生物学性状的差异。结果表明,有54个菌株对嘧菌酯表现抗性,均属于中等抗性水平,占检测总数的40.3%,分布于北京市、湖北、江苏、河北、辽宁和四川各省;对15个抗性菌株cytb基因的序列分析表明,其第143位发生突变,由甘氨酸变为丙氨酸;对随机选取的4株抗性菌株和2株敏感菌株的适合度测定结果发现,抗性菌株和敏感菌株在最适生长温度、菌落生长速度、孢子萌发率方面没有表现出规律性差异,但抗性菌株的产孢量和致病力均显著低于敏感菌株。     

灰葡萄孢抗二甲酰亚胺类杀菌剂研究进展

综述了灰葡萄孢对二甲酰亚胺类杀菌剂抗性的产生及现状,抗性菌株的生物学特性、生理生化特性以及抗药性机制,并就灰葡萄孢对二甲酰亚胺类杀菌剂抗性研究进行了展望。     

In vitro and In vivo Activity of Cyprodinil and Pyrimethanil on Botrytis cinerea Isolates Resistant to other Botryticides and Selection for Resistance to Pyrimethanil in a Greenhouse Population in Greece

No cross-resistance was observed between pyrimethanil or cyprodinil and the fungicides benomyl, iprodione or carbendazim + diethofencarb. In vitro, both anilinopyrimidine fungicides were effective against strains of Botrytis cinerea resistant to benzimidazoles and/or dicarboximides and against a wild type strain insensitive to diethofencarb (EC₅₀ values ranged between 0.03–0.19 and 0.006–0.054gml^–1 for pyrimethanil and cyprodinil, respectively). Preventive applications of anilinopyrimidines completely protected young cucumber plants and fruits that were inoculated with all strains of B. cinerea. The effectiveness of pyrimethanil against grey mould was studied in greenhouse grown tomatoes in relation to (a) the type of infection and the progress of the disease on different plant parts and (b) the response of the naturally occurring B. cinerea population to the selection pressure caused by eight successive applications of this fungicide. Pyrimethanil effectively controlled grey mould on leaves, fruits and stems but did not significantly reduce the number of dead plants and fruits with 'ghost spot' symptoms. The selection pressure caused by the consecutive applications of pyrimethanil resulted in reduction of its effectiveness on leaves that became apparent after the sixth application. This was correlated with a shift of the B. cinerea population (not previously exposed to anilinopyrimidines) towards reduced sensitivity, probably due to the development of a low level of resistance (R _L = 7.7). Pyrimethanil delayed the onset of the disease but it did not reduce the infection rate.     

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。     

Characterisation of QoI‐resistant field isolates of Botrytis cinerea from citrus and strawberry

BACKGROUND: In 2004, field isolates of Botrytis cinerea Pers. ex Fr., resistant to strobilurin fungicides (QoIs), were first found in commercial citrus orchards in Wakayama Prefecture, Japan. Subsequently, QoI‐resistant isolates of this fungus were also detected in plastic strawberry greenhouses in Saga, Ibaraki and Chiba prefectures, Japan. Biological and molecular characterisation of resistant isolates was conducted in this study. RESULTS: QoI‐resistant isolates of B. cinerea grew well on PDA plates containing kresoxim‐methyl or azoxystrobin at 1 mg L^?1, supplemented with 1 mM of n‐propyl gallate, an inhibitor of alternative oxidase, whereas the growth of sensitive isolates was strongly suppressed. Results from this in vitro test were in good agreement with those of fungus inoculation tests in vivo. In resistant isolates, the mutation at amino acid position 143 of the cytochrome b gene, known to be the cause of high QoI resistance in various fungal pathogens, was found, but only occasionally. The heteroplasmy of cytochrome b gene was confirmed, and the wild‐type sequence often present in the majority of resistant isolates, indicating that the proportion of mutated cytochrome b gene was very low. CONCLUSION: The conventional RFLP and sequence analyses of PCR‐amplified cytochrome b gene are insufficient for molecular identification of QoI resistance in B. cinerea. Copyright © 2009 Society of Chemical Industry     

8种杀菌剂对番茄灰霉病菌的毒力及田间番茄灰霉病菌对咯菌腈的敏感性

为了解决番茄灰霉病菌的抗性问题,筛选出理想的替代药剂,采用菌丝生长速率法测定了山东寿光和河北徐水两地番茄灰霉病菌对不同类型杀菌剂的敏感性。两地的番茄灰霉病菌对咯菌腈、氟啶胺均高度敏感,对其他不同作用方式的杀菌剂表现出不同的敏感性。用同样方法检测了不同地区的106株番茄灰霉菌对咯菌腈的敏感性,结果显示其EC50值分布在0.003 9～0.044 9μg/mL之间,平均EC50值为(0.014 2±0.008 1)μg/mL。咯菌腈可作为防治番茄灰霉病的候选药剂。     

Colonization of red raspberry flowers and fruit by Botrytis cinerea under commercial production conditions in northwestern Washington,USA

Colonization of red raspberry flowers and fruit by Botrytis cinerea was determined during 2017–2018 growing seasons under commercial fungicide application programmes used for grey mould management in northwestern Washington, USA. Colonization of flowers and fruit was assessed qualitatively (incidence, %) and quantitatively (abundance, number of colonies) by recovering B. cinerea from surface-disinfested samples. Both incidence and abundance of flower colonization were significantly lower than fruit colonization in both untreated and fungicide-treated plots. Incidence of flower colonization did not differ significantly between untreated and fungicide-treated plots (43% vs. 45%, respectively). In contrast, significantly greater colonization incidence was detected at green fruit stage in untreated compared to fungicide-treated plots (96% vs. 77%, respectively). Ripe fruit had the greatest colonization incidence among the three stages sampled and colonization was not significantly different between untreated and fungicide-treated plots (100% vs. 92%, respectively). Similarly, colonization abundance of flowers did not differ significantly between untreated and fungicide-treated plots (1.0 colonies per colonized flower in both treatments), but colonization abundance of green and ripe fruit was decreased 2.3- and 2.1-fold, respectively, in fungicide-treated plots. DNA fingerprinting analysis of the pathogen revealed that different multilocus genotypes colonized flowers and fruit within the same inflorescence and that genotypic diversity increased through time, suggesting independent infection events. Overall, our results demonstrate that under current environmental conditions, raspberry flowers may not be the exclusive or major route of infection for grey mould of red raspberry in northwestern Washington. Implications of current findings for management and further research are discussed.     

四川猕猴桃灰霉病菌对4种杀菌剂的抗药性检测

为明确猕猴桃灰霉病菌对常见杀菌剂的抗药性状况,本试验于2016年-2018年调查统计了四川猕猴桃生产中防控灰霉病所用的药剂种类和施药频次,并从四川8个猕猴桃主产区采集病叶、病花和病果,经单孢分离获得122个灰葡萄孢Botrytis cinerea菌株,采用最低抑制浓度法(MIC)测定其对4种化学药剂嘧霉胺、咯菌腈、腐霉...     

Identification and characterization of Botrytis medusae,a novel cryptic species causing grey mould on wine grapes in Australia

In a collection of 735 Botrytis isolates sampled from Australian wine grape-growing regions, a single isolate from clade I and group I (based on Bc-hch RFLP analysis) was found. As many Botrytis species are known to live sympatrically, it was hypothesized that this isolate might be a new Botrytis species. After phenotypic and molecular assays supported this hypothesis, the species was designated B. medusae. Phylogenetic analyses using the nuclear genes G3PDH, HSP60, RPB2, NEP1 and NEP2 consistently placed B. medusae in an early-diverging clade I Botrytis spp. lineage. Botrytis medusae produced white aerial mycelium, grew faster at 30 °C and produced long-branched conidiophore extensions, compared with B. cinerea and B. pseudocinerea. Botrytis medusae was only able to infect wounded grape leaves and was significantly less virulent on wounded leaves and berries than B. cinerea. Botrytis medusae also lacked villiform appendages on the conidial surface and long conidiophores, which are defining features of B. sinoviticola and B. californica, respectively. Identification and characterization of new cryptic Botrytis species living in sympatry on grapevines could potentially provide information to assist disease management strategies for B. cinerea.     

番茄灰霉病菌拮抗放线菌的筛选、鉴定及其活性评价

从安徽省寿县农田土壤中分离筛选到1株对番茄灰霉病菌Botrytis cinerea有强拮抗作用的放线菌菌株HNU-1。根据菌体形态特征、培养特征、细胞壁组分、生理生化特性和16S rRNA基因序列分析,将该菌株鉴定为吸水链霉菌Streptomyces hygroscopicus。菌株HNU-1发酵滤液可抑制灰霉病菌菌丝生长和分生孢子萌发,且浓度越高,抑制能力越强;当发酵滤液稀释6.67倍时则完全抑制灰霉病菌菌丝生长和分生孢子萌发。盆栽试验结果表明,菌株HNU-1发酵滤液6.67倍稀释液对番茄灰霉病的预防与治疗效果分别为87.8%和77.9%,均显著高于50%多菌灵可湿性粉剂500倍稀释液。     

Heat treatment for the control of rose and carnation grey mould (Botrytis cinerea)

Thermal dip treatment for flower heads was found effective against grey mould of roses. Dipping of flowers in tap-water at 50 C for 20 40 s was more effective than at higher or lower temperatures and than for longer treatments. The treatment was effective in five out of six rose cultivars and in one carnation cultivar. A 60, reduction in disease severity following the treatment was observed in flowers naturally infected by Botrytis cinerea, whereas the treatment was not effective against artificial infection of flowers. Conditioning of flowers for 2 days at 10 C and high relative humidity before incubation under grey-mould-conducive conditions (15 C. high humidity) increased the efficacy of the thermal treatment. Moreover, the combination of heat with 3 mM catechol was additively more efficient in reducing grey mould. The surfactant Tween 20 (001 %) improved the effect of treatment at 45 C for 20 s. A combination of heat with the fungicide chlorothalonil did not improve effectiveness, whereas when polyoxin B was combined with heat it was better than either treatment alone. The temperature of petals at the edge and centre of flower reached 37 and 27 C, respectively, at the end of a 20-s incubation period in 50 C water. Conidia treated at 34-40 C for 10-20 s incited 60% less severe disease than conidia treated at 25 C in water.