1种辣椒新炭疽病的初步鉴定及室内药剂筛选

炭疽病是辣椒上的重要病害,影响了辣椒的产量与品质。生产中,在“辛香15号”辣椒上发现了1种新炭疽病,仅在辣椒果实上发生,典型病斑为椭圆形,周围水渍状,中间呈黑色同心轮纹,无小黑点出现,对常用杀菌剂不敏感;保湿后镜检发现,该病菌产生分生孢子盘,无刚毛,分生孢子近长椭圆形,无色,单孢,大小为12．5μm×3．75μm,有油球,一端稍尖;进一步分离培养病原菌并通过柯赫氏法则验证后,初步将其鉴定为尖孢炭疽菌（Collectotrichum acutatum Simmonds）。为了获得对该病效果好的化学药剂,采用牛津杯法测定了8种化学药剂对病菌的抑制效果,结果表明：250g/L丙环唑乳油3000倍液抑制效果最好且药效持续时间长,可作为该病的候选药剂。本试验结果可为防治尖孢炭疽菌引起的辣椒炭疽病提供理论依据和用药参考。     

桑里白粉病菌重寄生真菌的分离与鉴定

为了解桑里白粉病菌的重寄生真菌种类,通过分离和寄生性验证,获得2个桑里白粉病菌重寄生真菌菌株HP8和HP9,这两个菌株寄生闭囊壳后,子囊及子囊孢子溃解。菌株HP8菌落绿色,菌丝体无色至褐色,菌丝有隔膜,分生孢子梗结节状膨大,分生孢子椭圆形、单胞、褐色或无色,ITS序列（MT463536）与尖孢枝孢Cladosporium oxysporum（MF135506）同源性为100%,系统进化树分析发现菌株HP8与尖孢枝孢（MF135506）聚在一起;菌株HP9菌落白色,菌丝无色,分生孢子月牙形、单胞、无色,ITS序列（MT463537）与刀孢蜡蚧菌Lecanicillium psalliotae（KC881072）同源性为99%,在系统进化树中,菌株HP9与刀孢蜡蚧菌（KC881072）聚在一起。结合形态特征和ITS序列分析,鉴定出菌株HP8和HP9分别为尖孢枝孢和刀孢蜡蚧菌。高通量测序结果表明,病桑叶在室内保存4个月后,桑里白粉病菌的重寄生真菌为刀孢蜡蚧菌。     

云南葡萄产区葡萄炭疽病病原鉴定及致病力分析

为了明确引起云南葡萄产区炭疽病的病原种类,利用形态鉴定和特异性引物分子检测相结合的方法对从云南省主要葡萄产区采集的60株炭疽病菌菌株进行了鉴定。葡萄炭疽病菌菌株的菌落形态和生长速率与对照菌株尖孢炭疽菌Colletotrichum acutatum差异不明显,但其分生孢子大小显著小于尖孢炭疽菌,附着胞深褐色,球形或不规则形。胶孢炭疽菌Colletotrichum gloeosporioides特异性引物CgInt/ITS4从供试葡萄炭疽病菌菌株基因组DNA中扩增出1条约500 bp的特异性条带,而尖孢炭疽菌特异性引物CaInt2/ITS4对葡萄炭疽病菌无扩增条带。研究表明,引起云南葡萄主产区炭疽病的病原为胶孢炭疽菌;供试胶孢炭疽菌对红提葡萄均有致病力,但菌株致病力差异较大,对番茄和草莓存在交叉侵染的能力,且对多菌灵的敏感性较尖孢炭疽菌高。     

一种优化的辣椒尖孢炭疽菌遗传转化体系

由辣椒尖孢炭疽菌(Colletotrichum acutatum)侵染引起的炭疽病是辣椒生产中最具有破坏性的真菌病害,严重影响辣椒的品质和产量。本研究以辣椒尖孢炭疽病菌HHDL02为对象,采用1 mol·L^-1 NH₄Cl 2%的酶裂解液裂解分生孢子萌发2 h的芽管,裂解2～3 h可以高效制备原生质体,结合PEG介导转化法成功将GFP基因导入,其转化菌株菌落形态、菌丝生长速率、分生孢子形态、孢子萌发、附着胞形成、产孢量和致病性与野生型菌株无明显差异,且后代荧光信号遗传稳定。PEG介导的原生质体转化适合辣椒尖孢炭疽菌遗传操作,有助于其致病机理的研究。     

黑龙江省高粱靶斑病病原菌的鉴定及其生物学特性

为明确黑龙江省高粱靶斑病病原菌种类及其生物学特性，采用组织分离法对采集自黑龙江省8个地区的75份高粱病叶进行病原菌分离，利用形态学特征观察及分子生物学技术对选取的代表菌株进行鉴定，并分析其生物学特性。结果表明，共分离得到42株病原菌菌株，按采集地选择的8株代表菌株的菌丝生长速率不同，但形态学特征较一致，分生孢子为棕黄色，呈圆柱形，中部较宽，稍弯曲，大小为29~96 μm×9~15 μm，具3~9个隔膜，脐点明显，基部平脐；8 株代表菌株的ITS序列与高梁生双极蠕孢菌Bipolaris sorghicola（GenBank登录号为AF071332.1）的同源性为99%~100%；结合形态学特征和ITS序列分析确定黑龙江省高粱靶斑病病原菌为高梁生双极蠕孢菌B. sorghicola。8株代表菌株在燕麦片琼脂培养基上生长最好，但不同菌株的菌丝生长速率存在差异；在10~35℃范围内均能生长，但不同菌株的最适生长温度不同；光照对菌丝生长有促进作用；最适生长的pH为8；不同菌株对碳、氮源的利用差异较大，表明高粱生双极蠕孢菌菌株间存在生物学特性差异。     

进境美国苹果星裂壳孢果腐病菌的首次截获-病原真菌的鉴定及风险分析

 广东口岸从美国华盛顿输华苹果（Malus domestica）中截获可疑腐烂病果, 症状表现为果梗凹腐或萼凹腐,病部果皮暗褐色至黑色,病健交界处纹带褐色,分生孢子器黑色、颗粒状,直径0.3~0.8 mm,部分埋生或近表生, 显微镜检可见无色单胞的分生孢子。分离物生长温度-3~25℃, 最适为20℃, 经柯赫氏法则验证确认为截获病果的病原菌。Blast分析表明从分离物基因组中扩增到的ITS基因与GenBank中已知的Phacidiopycnis washingtonensis菌株ITS序列同源性达100%。经形态特征、培养性状及ITS系统发育分析, 将病原菌鉴定为苹果星裂壳孢果腐病菌（Ph. washingtonensis Xiao& J.D.Ro-gers,2005）,在分类上隶属于半知菌亚门球壳孢目星裂壳孢属(Phacidiopycnis),此截获鉴定属我国首次。迄今为止我国尚未有该病菌发生为害的报道,本文就此病菌对我国苹果和梨产业的潜在风险进行了评估分析。     

江西瑞昌山药炭疽病菌的形态及分子鉴定

为了明确江西瑞昌山药炭疽病病原菌种类归属,本文从当地采集呈典型症状的炭疽病叶片进行了病原菌分离鉴定.通过组织分离获得8个在培养性状和分生孢子形态大小均一致且均具有致病性的分离株,8个分离株在PDA平板上菌落初为白色,后变为灰色至深灰色,菌落中央产生橘红色黏质分生孢子团.分生孢子无色,长椭圆形至纺锤形,单胞,大小为(15.6~18.0)μm×(3.6~6.0)μm.对其中之一的分离株YRRC-1进行rDNA-ITS区段扩增和序列测定,获得长度为536 bp的rDNA-ITS序列,该序列与胶孢炭疽菌(Colletotrichum gloeosporioides)的对应序列同源性达100%.根据分离病菌的培养特征、形态大小和序列鉴定结果,认为瑞昌山药炭疽病菌属于胶孢炭疽菌(C. gloeosporioides).     

辣椒炭疽病生防菌株的筛选、鉴定及其抑菌机理

为有效防控由胶孢炭疽菌Colletorichum gloeosporioides引起的辣椒炭疽病,自辣椒上分离得到内生细菌,通过平板拮抗和辣椒离体生防试验筛选对胶孢炭疽菌有抑制作用的拮抗菌株,通过形态学特征、生理生化特征以及分子生物学技术对其进行鉴定,并于室内测定其对胶孢炭疽菌菌丝生长的影响、对辣椒炭疽病的防效及接种后辣椒内抗病活性物质含量以及防御酶活性。结果显示,从辣椒上共分离纯化获得46株细菌,其中菌株SQ-6对胶孢炭疽菌有明显的抑制作用,抑制率为61.11%,显著高于其他45株。结合菌株SQ-6的形态学特征、生理生化特征以及分子生物学特征,将该菌株鉴定为解淀粉芽胞杆菌Bacillus amyloliquefaciens。SQ-6菌株的50%无细胞滤液可引起胶孢炭疽菌菌丝畸形、断裂等,对其抑制率为57.87%。SQ-6菌株的10%、50%发酵液和10%、50%无细胞滤液均能显著降低由胶孢炭疽菌引起的辣椒炭疽病的发病率和病情指数,其中50%无细胞滤液的防效最好。SQ-6菌株能够提高辣椒内Vc、酚类和黄酮类物质含量,诱导辣椒内过氧化物酶（peroxidase,POD）、丙氨酸解氨酶（p...     

法国向日葵种子中向日葵黑茎病菌的首次截获与检测

从法国进境向日葵种子中分离到3株疑似向日葵黑茎病的菌株,对所有菌株进行形态学、致病性测定和分子序列比对分析。分离菌菌落乳白色或象牙色至灰白色,有大量黑褐色分生孢子器产生,分生孢子器球形至扁球形,内含无色单胞、卵圆形分生孢子,有明显或不明显油球;针刺接种4片真叶向日葵幼苗的下胚轴,7～9d后茎部产生典型黑茎病黑色椭圆形病斑,病斑上着生黑色分生孢子器;菌丝DNA用ActF1/R1和ITS1/ITS4扩增,序列与NCBI基因库中P.macdonaldii序列相似性为98%～100%。形态学、分子生物学及致病性检测结果显示,截获的3株菌均为向日葵黑茎病菌。     

辽宁省辣椒土传病害镰刀菌鉴定及rDNA-ITS序列分析

［目的］ 对分离的辣椒镰刀菌形态、基因序列进行研究,以确定病原菌的种类及这些菌的同源性,为进一步开展该病害诊断和防治研究奠定基础。［方法］ 对采自辽宁省各地辣椒土传病害的16株病菌进行形态观察及rDNA-ITS序列分析。［结果］ 引起辽宁省辣椒土传病害的主要镰刀菌有尖镰孢、茄镰孢、锐顶镰孢和串珠镰孢等4种,根据菌株分生孢子形态和rDNA ITS序列同源性,将ZW13、LY3、PJLJ、HC5、CY2、CY5、TLLJ、LY5、LY1菌株聚类为尖镰孢;FSLJ2和FSLJ3菌株聚类为茄镰孢;ZW和ZW18菌株聚类为锐顶镰孢;KPLJ、LZLJ、FSLJ1菌株聚类为串珠镰孢。［结论］ 从分子系统树和菌株间的遗传距离看出镰刀菌同种同源性都很高,不同种之间同源性相对较低,种间遗传差异较大。     

Anthracnose of sweet pepper caused by Colletotrichum scovillei in Japan

Severe fruit rot of sweet pepper was found in Shimane, Hyogo, Chiba, Toyama, and Nagano prefectures, Japan from 2005 to 2011. Dark, sunken spots with concentric rings of orange conidial masses appeared on fruits. Pathogenic isolates from diseased fruits in the prefectures were identified as Colletotrichum scovillei. This species was added to the pathogens of sweet pepper anthracnose in Japan. The representative isolate was pathogenic to sweet pepper, tomato and chili pepper fruits, kidney bean pod, azuki bean, pea and strawberry leaves, but a caused no symptoms on cucumber or carrot in inoculation tests.     

Characterization and pathogenicity of Colletotrichum species associated with anthracnose on chilli (Capsicum spp.) in Thailand

Fungal isolates from chilli ( Capsicum spp.) fruits in Thailand that showed typical anthracnose symptoms were identified as Colletotrichum acutatum , C . capsici and C . gloeosporioides . Phylogenetic analyses from DNA sequence data of ITS rDNA and β-tubulin ( tub 2) gene regions revealed three major clusters representing these three species. Among the morphological characters examined, colony growth rate and conidium shape in culture were directly correlated with the phylogenetic groupings. Comparison with isolates of C . gloeosporioides from mango and C . acutatum from strawberry showed that host was not important for phylogenetic grouping. Pathogenicity tests validated that all three species isolated from chilli were causal agents for chilli anthracnose when inoculated onto fruits of the susceptible Thai elite cultivar Capsicum annuum cv. Bangchang. Cross-infection potential was shown by C . acutatum isolates originating from strawberry, which produced anthracnose on Bangchang. Interestingly, only C . acutatum isolates from chilli were able to infect and produce anthracnose on PBC 932, a resistant genotype of Capsicum chinense . This result has important implications for Thai chilli breeding programmes in which PBC 932 is being hybridized with Bangchang to incorporate anthracnose resistance into chilli cultivars.     

山东省辣椒上尖孢炭疽复合种对吡唑醚菌酯的敏感基线及吡唑醚菌酯增效配方筛选

为明确山东省辣椒炭疽病菌对吡唑醚菌酯的敏感性,延缓其抗性发展及降低防治成本,在该省主要辣椒产区采集并经单孢分离获得175株尖孢炭疽复合种,采用菌丝生长速率法和孢子萌发法测定了病原菌群体对吡唑醚菌酯的敏感性,同时测定了吡唑醚菌酯与甲基硫菌灵、多菌灵、代森锰锌、戊唑醇和咯菌腈5种杀菌剂分别混用对尖孢炭疽复合种的联合毒力。结果表明:吡唑醚菌酯对尖孢炭疽复合种菌丝生长和孢子萌发的EC50值范围分别为0.056~0.530和0.002~0.027 μg/mL,平均值为（0.273 ± 0.067）和（0.014 ± 0.019）μg/mL,其敏感性频率分布呈连续单峰曲线,符合正态分布,可作为田间抗性监测的敏感基线;不同采集地菌株对吡唑醚菌酯的敏感性存在一定差异,其中采自菏泽市的菌株敏感性最高,而采自潍坊市的菌株敏感性最低;吡唑醚菌酯与甲基硫菌灵按体积比1:3和3:1复配,对抑制尖孢炭疽复合种的菌丝生长和分生孢子萌发均表现为增效作用,其中按体积比1:3混用时增效作用最明显,增效系数分别是4.82和3.94。结果可为吡唑醚菌酯的合理使用提供理论依据。     

Characterization of three Colletotrichum acutatum isolates from Capsicum spp.

Colletotrichum acutatum causes anthracnose on peppers (Capsicum spp.), resulting in severe yield losses in Taiwan. Fungal isolates Coll-153, Coll-365 and Coll-524 collected from diseased peppers were found to differ in pathogenicity. Pathogenicity assays on various index plants revealed that Coll-524 was highly virulent and Coll-153 was moderately virulent to three commercially available pepper cultivars. Both isolates induced anthracnose lesions and produced abundant conidia. Coll-365 was only weakly virulent on pepper fruit, where it caused small lesions and hardly produced conidia on pepper fruit. However, Coll-365 was highly pathogenic to tomato fruit and mango leaves, where it caused anthracnose lesions and formed acervuli and conidia. All three isolates showed similar abilities in the attachment and germination of conidia, formation of highly branched hyphae and appressoria, penetration of cuticles, and infection of epidermal cells on chili peppers. Coll-365 accumulated less turgor pressure in appressoria but produced higher levels of cutinase and protease activity than Coll-153 and Coll-524 did. All three isolates invaded the neighbouring cells through plasmodesmata in chili peppers and showed similar pectinase or cellulase activities in culture. However, the most virulent strain Coll-524 expressed stronger laccase activity and was more resistant to capsaicin compared to Coll-153 and Coll-365. The three isolates are different in numbers and sizes of double-stranded RNAs. Depending on the cultivar genotypes, cellular resistance of chili pepper to C. acutatum might rely on the ability to restrict penetration, colonization, or conidiation of the pathogen. We conclude that the differences in pathogenicity among the three C. acutatum isolates of pepper are attributed to their ability to colonize the host plant.     

Development of protocols for detection of Colletotrichum acutatum and monitoring of strawberry anthracnose using real-time PCR

Real-time PCR (TaqMan®) assays were developed for the specific detection and discrimination of Colletotrichum spp., C. acutatum and C. gloeosporioides causing anthracnose in strawberry using the most divergent area of the internal transcribed spacers (ITS1 and ITS2) and 5·8S ribosomal RNA (rRNA) gene region. The specificity of the new assays was tested using DNA from six species of Colletotrichum and nine fungal species commonly found associated with strawberry material, and additionally by comparing the sequences with those from databases using a blast search. The sequences only showed identity with homologous sequences from the desired target organisms. The new assays were 10–100 times more sensitive than conventional PCR methods previously published for the diagnosis of strawberry anthracnose. When real-time PCR was compared with ELISA methods, PCR improved the sensitivity of the identification by obtaining positive results for samples of strawberry plant material that tested negative with ELISA. The development of C. acutatum was monitored using artificially infected strawberry crowns from two strawberry cultivars (Camarosa and Ventana) and a real-time PCR assay specific for this species between January and June 2006. The amount of C. acutatum detected using real-time PCR varied significantly by month ( P  < 0·001), but not by cultivar ( P  = 0·394). The new assays were shown to be useful tools for rapid detection and identification of these pathogens and to allow rapid and accurate assessment of the casual agents of anthracnose in strawberry.     

Root treatment with rhizobacteria antagonistic to Phytophthora blight affects anthracnose occurrence, ripening, and yield of pepper fruit in the plastic house and field

We previously selected rhizobacterial strains CCR04, CCR80, GSE09, ISE13, and ISE14, which were antagonistic to Phytophthora blight of pepper. In this study, we investigated the effects of root treatment of rhizobacteria on anthracnose occurrence, ripening, and yield of pepper fruit in the plastic house and field in 2008 and 2009. We also examined the effects of volatiles produced by the strains on fruit ripening and on mycelial growth and spore development of Colletotrichum acutatum and Phytophthora capsici in the laboratory, identifying the volatile compounds by gas chromatography-mass spectrometry (GC-MS). In the house tests, all strains significantly (P < 0.05) reduced anthracnose incidence on pepper fruit; strains GSE09 and ISE14 consistently produced higher numbers of pepper fruit or increased the fresh weight of red fruit more than the controls in both years. In the field tests, all strains significantly (P < 0.05) reduced anthracnose occurrence on either green or red pepper fruit; strain ISE14 consistently produced higher numbers or increased fresh weights of red fruit more than the controls in both years. In the laboratory tests, volatiles produced by strains GSE09 and ISE13 only stimulated maturation of pepper fruit from green (unripe) to red (ripe) fruit; the volatiles of certain strains inhibited the growth and development of C. acutatum and P. capsici. On the other hand, GC-MS analysis of volatiles of strains GSE09 and ISE13 revealed 17 distinct compounds in both strains, including decane, dodecane, 1,3-di-tert-butylbenzene, tetradecane, 2,4-di-tert-butylphenol, and hexadecane. Among these compounds, 2,4-di-tert-butylphenol only stimulated fruit ripening and inhibited growth and development of the pathogens. Taken together, strains GSE09 and ISE14 effectively reduced anthracnose occurrence and stimulated pepper fruit ripening and yield, possibly via bacterial volatiles. Therefore, these two strains could be potential agents for controlling Phytophthora blight and anthracnose, and for increasing fruit ripening and yield. To our knowledge, this is the first report of volatiles such as 2,4-di-tert-butylphenol produced by rhizobacteria being related to both fruit ripening and pathogen inhibition.     

Chili anthracnose: Colletotrichum taxonomy and pathogenicity

Chili anthracnose is caused by Colletotrichum species mostly associated with the acutatum, truncatum and gloeosporioides complexes. Since 2009 the Colletotrichum taxonomy has been extensively revised based on multigene phylogenetics, which has had a large impact on the number of species known to cause anthracnose disease of chili. This review discusses (i) the taxonomy of Colletotrichum spp. infecting chili, and (ii) the impact of Colletotrichum pathotypes on breeding for resistance to anthracnose. To date, 24 Colletotrichum species have been identified as pathogens of chili anthracnose, with the three main pathogens being C. scovillei, C. truncatum and C. siamense. Identification of several pathotypes within these three Colletotrichum species, particularly pathotypes that can overcome resistance in the related Capsicum species, Ca. chinense and Ca. baccatum, will be of major concern to plant breeders as they develop resistant chili genotypes from the transfer of resistance genes from these Capsicum species into Ca. annuum. Accurate identification of the Colletotrichum species causing anthracnose and improved understanding of the biology of the Colletotrichum species and their interaction with the host will enable the application of improved integrated disease management techniques.     

Identification of Subpopulations of Colletotrichum acutatum and Epidemiology of Almond Anthracnose in California

ABSTRACT In recent years, almond anthracnose has developed into a major problem for the California almond industry. The identification of the causal pathogen as Colletotrichum acutatum was confirmed using species-specific primers and restriction fragment length polymorphisms of ribosomal DNA in comparative studies with isolates of C. acutatum from strawberry and C. gloeosporioides from citrus. Two distinct clonal subpopulations among the almond isolates of C. acutatum were identified. These two subpopulations differed in their colony appearance (pink versus gray cultures), conidial morphology, virulence in laboratory inoculation studies, temperature relationships for growth, and molecular fingerprints using random and simple-repeat primers in polymerase chain reactions. Both subpopulations were commonly isolated from the same orchard or even the same fruit. In other orchards, one subpopulation predominated over the other subpopulation. Using random, simple-repeat, and species-specific primers, isolates of the almond anthracnose pathogen from Israel were very similar to the California isolates that produce gray colonies. In addition to fruit, the pathogen was isolated from blighted blossoms, water-soaked or necrotic leaf lesions, symptomless peduncles, and spurs and wood from branches showing dieback symptoms, indicating that the amount of tissue that may be infected is more extensive than previously considered. Overwintering fruit mummies were identified as inoculum sources for early spring infections. Growth studies using almond kernels with different moisture contents indicated that postharvest damage of stored kernels likely originates from preharvest field infections.     

Management and cross-infectivity potential of Colletotrichum acutatum causing anthracnose on bell pepper in Florida

Early anthracnose caused by Colletotrichum acutatum has become an increasingly serious disease on green, unripe bell pepper fruit in Florida. This contrasts with earlier reports of anthracnose occurring on bell pepper primarily as a ripe-rot disease of mature, colored pepper fruit caused by Colletotrichum gloeosporioides. Management of anthracnose on green bell pepper fruit using fungicides and a commercial inducer of systemic acquired resistance, acibenzolar-S-methyl (ASM), was evaluated during three seasons. In two of the three trials, all the fungicides tested including azoxystrobin, fludioxonil + cyprodinil, mancozeb, famoxadone + cymoxanil, copper hydroxide, and ASM significantly increased the number of marketable fruit compared with control plants. These trials identified fungicides that could contribute to a successful pest management program on pepper for controlling anthracnose caused by C. acutatum. The cross-infectivity potential of C. acutatum was investigated on tomato and strawberry by in vitro and field inoculation. Anthracnose lesions formed readily on wound-inoculated detached fruits of all hosts in in vitro assays. Under field conditions, after inoculation, anthracnose lesions occurred on pepper fruit but no lesions of anthracnose were found on either ripe or unripe tomato or strawberry fruit in adjacent plots.