马铃薯早疫病菌的分子鉴定与田间早期诊断

马铃薯早疫病由茄链格孢Alternaria solani引起,通常导致产量损失达30％以上[1-2].目前关于马铃薯早疫病菌的鉴定主要是基于形态特征进行.然而,该病菌在离体培养时不易产孢,增加了鉴定的难度.马铃薯早疫病的早期诊断在早疫病防控中占有重要地位,建立马铃薯早疫病菌的分子鉴定和早疫病的田间早期诊断技术将有助于防控措施的制定.本研究选择ITS序列作为早疫病菌分子鉴定的目标区域,通过ITS序列分析设计马铃薯早疫病菌的特异性引物,建立早疫病菌的分子鉴定技术并探索早疫病早期诊断技术.     

南疆骏枣黑斑病症状表现及病原菌鉴定

为明确南疆骏枣黑斑病的症状表现及病原菌种类,2013—2014年在新疆建设兵团第一师选取具有代表性的5个骏枣园,对枣叶、花、果等组织上黑斑病的症状进行了系统调查,采用组织分离法和回接试验分离病原菌及测定其致病性,并根据病菌形态特征、ITS序列和β-tubulin序列分析对病原菌进行了鉴定。结果表明:枣叶、花和果均可感染黑斑病菌,室内接种和田间骏枣黑斑病症状相同,叶、花和果中的病原菌可以相互侵染,并在枣果上均表现出黑斑病典型症状;不同发病组织中分离纯化得到357株菌株,形态观察表明,侵染叶、花和果的菌株均为链格孢属链格孢菌Alternaria alternata(Fr.)Keissler;通过ITS序列和β-tubulin序列分析,并结合形态学特征进一步确定引起骏枣不同组织的黑斑病病原菌为链格孢菌A.alternata。     

青海省樱桃叶斑病病原菌的分离与鉴定

为明确引起青海省樱桃叶斑病病原菌的种类,本研究从西宁市城北区、海东市乐都区和贵德县的樱桃上采集有叶斑症状的叶片,采用组织分离法分离出病原菌,通过观察病原菌的形态特征,结合rDNA-ITS、EF-1α和Alt a 1基因序列分析对病原菌的种类进行了鉴定,并用柯赫氏法则进行验证。结果表明,共获得279株病原菌菌株,分属链格孢Alternaria alternata、细极链格孢A.tenuissima和刺盘孢属Colletotrichum spp.,分离频率分别为84.95%、5.02%和10.03%。采用柯赫氏法则进行离体叶片致病性测定,接种叶片100%发病,说明引起青海省樱桃叶斑病的病原菌为链格孢、细极链格孢和刺盘孢属真菌,且链格孢为主要病原菌。该研究结果可为青海省樱桃叶斑病的综合防治提供理论依据。     

河北和山东鸭梨果实上链格孢菌鉴定

为了查清造成鸭梨果实储藏期黑斑病的链格孢Alternaria的种类,从其产孢表型、分生孢子特征和分子生物学等方面对分离的链格孢菌进行了研究.经对分离到的188支Alternaria菌株的形态学鉴定,共确定了3个种,即链格孢Alternaria alternata、细极链格孢A.tenuissima和侵染链格孢A.infectoria,比例分别为41.0%、54.8%和1.6%.分子生物学研究结果表明,Altemaria大孢子种彼此间及大孢子种与小孢子种之间可以根据ITS和gpd序列差异明确区分;而供试的多数小孢子种在ITS和gpd序列上差异很小,无法区分.     

AsSod基因调控茄链格孢致病性及对胁迫的响应

由茄链格孢Alternaria solani引起的马铃薯早疫病是马铃薯生产上的重要病害之一,可导致马铃薯大面积减产,进而造成巨大的经济损失.本研究在获得缺失AsSod基因的茄链格孢突变株(ΔsSod)的基础上,对△sSod进行了不同的胁迫处理.结果 表明,相比于野生型菌株和回复株,突变株ΔAsSod对细胞壁胁迫因子SD...     

香榧果实褐斑病病原菌鉴定及防治药剂筛选

为明确近年来在香榧Torreya grandis果实上出现的一种新病害——褐斑病的病原菌,通过分离物致病性测定、形态学观察、rDNA-ITS序列分析及病原菌生物学特性进行鉴定,并对苯醚甲环唑、嘧菌酯、甲基硫菌灵、咪鲜胺、肟菌酯及吡唑醚菌酯6种防治药剂进行了初步筛选。结果表明,从病样中分离到的菌株XF01在回接试验出现的症状与田间自然感病症状一致,证明菌株XF01为香榧果实褐斑病的致病菌。该病原菌菌丝初为灰白色,后为深灰色或青褐色;分生孢子梗单生或簇生,无分枝,淡褐色;分生孢子多倒梨形或卵形,大小平均为19.258μm×9.048μm,横、纵隔膜分别为1~7个和0~3个,无喙或具柱状短喙,形态特征与链格孢菌Alternaria alternata一致。该菌ITS序列(Gen Bank登录号为KU525533)与链格孢菌株ITS序列(Gen Bank登录号为KR864893.1)同源性为100%,并聚为一簇,分子鉴定结果与形态学鉴定结果一致,确定引起该病害的病原菌为链格孢菌。该病菌菌丝生长、产孢和孢子萌发的最佳温度均为28℃,最适生长pH均为7。室内毒力测定结果显示,咪鲜胺对该菌菌丝生长的抑制效果最好,EC50为0.065μg/mL;吡唑醚菌酯对其孢子萌发的抑制效果最好,EC50为0.002μg/mL,表明这2种药剂可用于香榧果实褐斑病的田间防治。     

引起葡萄叶斑病的链格孢种类的初步鉴定

为明确北京市大兴区、顺义区和山西省襄汾县葡萄树上叶斑病的病原菌,利用组织分离法对病样进行分离纯化,采用多基因系统发育学分析(ITS,Alt a 1和gpd)和形态学对纯化后菌株进行鉴定,并结合柯赫氏法则对其进行验证。结果表明,共获得37株链格孢属真菌菌株,分属链格孢Alternaria alternata、葡萄链格孢A. viniferae和乔木链格孢A. arborescens,柯赫氏法则检测结果显示接种菌株对所接叶片与果穗均具致病性,说明这3种链格孢种均为葡萄叶斑病的病原菌;葡萄链格孢为主要的病原菌,有28株,故将该类病害命名为葡萄链格孢叶斑病;此外,该类菌株在2～40℃之间均能生长,25℃为最适生长温度。     

阿尔泰藜芦提取物抗真菌活性研究初报

采用超声辅助法用乙醇对阿尔泰藜芦(Veratrum lobelianum Bernh.)根和根茎部浸提;浸提液经浓缩后,依次用石油醚、氯仿、乙酸乙酯、水萃取得到相应的萃取物;采用菌丝生长抑制法测定了阿尔泰藜芦提取物对灰葡萄孢菌(Botrytis cinerea)、茄链格孢菌(Alternaria solani)、尖孢镰刀菌(Fusarium oxysporum)及链格孢菌(Alternaria alternata)的抑菌活性。结果表明,4种萃取物对灰葡萄孢菌、茄链格孢菌、尖孢镰刀菌及链格孢菌4种供试病原菌均有一定的抑制作用。4种萃取物(石油醚、氯仿、乙酸乙酯、水)对链格孢菌的抑制率均最高,EC_(50)分别为1 683.00、929.33、766.00、358.67μg/mL。水相萃取物对4种病原菌的抑制效果最佳,其对灰葡萄孢菌、茄链格孢菌、尖孢镰刀菌及链格孢菌的EC_(50)分别为939.33、554.00、1 294.33μg/mL和358.67μg/mL。     

北京地区桃黑斑病新症状的发现与诊断

2019年-2020年在北京地区桃园病害调研普查时发现一种桃树病害的新发症状-新叶坏死,主要症状表现为当年新生枝条顶端新叶和生长点腐烂坏死。为明确这种新叶坏死症状是否由病原菌侵染引起,从北京平谷2个乡镇3个桃园采集35份病梢样品,经病样组织分离和菌株纯化,选取12个代表性菌株进行形态学鉴定、系统进化分析以及致病性测定。结果显示,通过形态学鉴定和多基因(ITS,Alt a1,gpd)系统发育分析,以上分离株均鉴定为链格孢Alternaria alternata;3株代表性菌株的致病性测定验证了A.alternata是引起新叶坏死症状的病原菌,并且该病原菌也能导致桃枝条和果实发病,产生黑斑病症状。这是首次报道A.alternata除了引起桃果实黑斑症状之外,还可以引起桃树新叶坏死症状,结果进一步丰富了链格孢侵染桃树的症状多样性,为链格孢引起的病害的整体防控提供了理论基础。     

我国梨和部分国外梨果实上链格孢菌的鉴定研究

本文从链格孢的产孢表型、分生孢子特征和分子生物学方面研究了从河北鸭梨、新疆库尔勒香梨、美国和日本以及智利的梨上分离的链格孢菌的种类.经对115支Alternaria菌株的形态学鉴定,共确定了3个种,即Alternaria alternata、A.tenuissima和A.infectoria.ITS系统发育分析研究表明,Alternaria大孢子种彼此间及大孢子种与小孢子种间已发生明显分化,可以根据ITS序列差异明确区分.而供试的多数小孢子种在ITS序列上差异很小,因而不能根据ITS序列差异来区分.     

吡唑醚菌酯与苯醚甲环唑混合物对茄链格孢的联合毒力及其对马铃薯产量的影响

为明确吡唑醚菌酯与苯醚甲环唑混合物对茄链格孢Alternaria solani毒力的增效作用,采用菌丝生长速率法和孢子萌发法测定了吡唑醚菌酯、苯醚甲环唑及其不同比例混合物对茄链格孢的毒力,以Wadley公式评价其联合毒力,并通过田间试验评价了混合物对马铃薯早疫病的防治效果及其对马铃薯产量的影响。结果表明:吡唑醚菌酯与苯醚甲环唑质量比为1∶1、2∶1、3∶1、4∶1、5∶1、1∶2和1∶5的混合物对抑制菌丝生长表现为增效,其中配比1∶2的增效最明显,增效系数(SR)为2.05,但该配比混合物对抑制分生孢子萌发仅表现为相加作用(SR 1.29)。在田间药效试验中,配比为1∶2的混合物对马铃薯早疫病的防效显著高于相同剂量的单剂及其他常用药剂的防效,增产率达46.8%,亦显著高于对照药剂。     

马铃薯早疫病菌对咯菌腈的敏感基线及其对不同药剂的交互抗性

采用菌丝生长速率法测定了河北省马铃薯主产区的122株早疫病菌Alternaria alternata对咯菌腈的敏感性,并测定了紫外诱导的15个抗咯菌腈突变菌株及其3个敏感亲本菌株对咯菌腈、异菌脲、腐霉利、苯醚甲环唑、吡唑醚菌酯的敏感性。田间马铃薯早疫病菌群体对咯菌腈保持高度敏感,122株早疫病菌对咯菌腈的EC₅₀值在0.0025~0.0857 μg/mL之间,平均值为0.0404±0.0171 μg/mL,相差34.87倍,呈近似正态分布,可作为早疫病菌对咯菌腈的敏感基线。此外,咯菌腈与吡唑醚菌酯和苯醚甲环唑之间无交互抗性关系,而与异菌脲和腐霉利之间存在正交互抗性关系。在田间药效试验中,50%咯菌腈WP施药量为105、115 g/hm²时,对马铃薯早疫病防效达80%以上,显著高于常规药剂80%代森锰锌WP 1 350 g/hm²和25%丙环唑EC 37.5 g/hm²的防效。表明咯菌腈可作为吡唑醚菌酯和苯醚甲环唑的替代药剂用于马铃薯早疫病的防治。     

Identification of Alternaria spp. on wheat by pathogenicity assays and sequencing

The pathogenicity of Alternaria spp. isolated from wheat leaves collected in regions where alternaria leaf blight has been reported was compared with that of IMI reference isolates of A. triticina and A. alternata using two durum and two bread wheat genotypes. To identify isolates putatively corresponding to A. triticina , morphological and DNA sequence analyses based on ribosomal DNA from the internal transcribed spacer (ITS) region (ITS1, 5·8S rRNA gene, ITS2) and toxicity bioassays of culture filtrate were combined. Glasshouse inoculations provided reliable information to assess the pathogenicity of A. triticina isolates on wheat. Alternaria leaf blight symptoms were produced by the A. triticina isolates only on durum wheat cv. Bansi, while A. alternata , A. tenuissima and A. arborescens isolates were found to be nonpathogenic on the wheat cultivars tested. Alternaria triticina isolates were distinguished from other Alternaria species by Simmons and Roberts' sporulation pattern 6 and two to three conidia per sporulation unit associated with primary conidia bearing long (> 7  µ m) apical secondary conidiophores. Phylogenetic analysis also proved effective at discriminating wheat-pathogenic A. triticina from other nonpathogenic Alternaria species. Alternaria triticina isolates yielded longer ITS sequences than A. alternata , A. tenuissima and A. arborescens isolates, leading to clear-cut differences as visualized with agarose gel electrophoresis. Additionally, only culture filtrates of A. triticina isolates caused nonspecific necrotic lesions on leaves of 3-week-old wheat plants.     

花椒枯穗病病原鉴定

Ten Zanthoxylum bungeanum ear blight diseased samples were collected from the Research Base of Forestry Research Institute of Hebei Province of China during 2009. Three isolates were obtained from diseased samples and no distinct difference could be observed among them. The fungus E-1 was determined to be responsible for this disease by the Koch’s Postulation. Morphological and the phylogenetic tree constructed based on ITS sequences showed that the morphological characters were the same as Alternaria alternata and the homology between E-1 and A. alternata was up to 100 percent. It indicated that E-1 was A. alternate（JQ973810）.     

Morphological, pathogenic, and molecular characterization of alternaria isolates associated with alternaria late blight of pistachio

ABSTRACT Alternaria isolates were obtained from various pistachio tissues collected in five orchards in California. For all isolates, morphological characteristics of the colony and sporulation apparatus were determined and compared with those of representative isolates of A. alternata, A. tenuissima, A. arborescens, and A. infectoria. A selection of the pistachio isolates and the representative Alternaria isolates were evaluated for pathogenicity to pistachio. Molecular characteristics of these isolates were determined using random amplified polymorphism DNA (RAPD) analysis, polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP) analysis of nuclear intergenic spacer rDNA, and sequence analysis of nuclear internal transcribed spacer (ITS) rDNA. Based on morphological characteristics, the pistachio isolates were grouped as identical or very similar to either A. alternata, A. tenuissima, A. arborescens, or A. infectoria. Isolates from the alternata, tenuissima, and arborescens species-groups were pathogenic to pistachio and no significant differences in pathogenicity were observed. Isolates from the infectoria species-group were only weakly pathogenic to pistachio. Based on cluster analysis of RAPD and PCR-RFLP data, three distinct clusters were evident; the infectoria cluster, the arborescens cluster, and a combined alternata/tenuissima cluster. Based on analysis of ITS sequence data, the infectoria species-group was phylogenetically distinct from the other species-groups. Isolates of the alternata, tenuissima, and arborescens species-groups comprised a monophyletic clade in which the three species-groups could not be further resolved.     

Wirkung verschiedener Fungizide auf den Befall der D��rrfleckenkrankheit (Alternaria spp.) sowie auf den Ertrag der Kartoffel

Potato early blight caused by Alternaria solani and Alternaria alternata is a major disease of potatoes and other Solanaceae. By now, Alternaria species have established themselves as destructive pathogens. Due to considerable yield losses early blight is getting a serious problem in many German potato growing areas. Up to now, the management of this disease is achieved mainly through protective fungicide applications. By this, fungicides with different mode of actions have a considerable influence on the course of disease progress. For this, field trials were carried out in 2007 and 2008 and the efficacy of various active ingredients was determined. Marked differences were observed in the control of early blight disease due to the effectiveness of applied active ingredients. The application of potent fungicides, which are highly efficient, resulted in prolonged photosynthesis. As a result of this potato yield increased significantly. Our investigation show, that disease control can be better achieved with strobilurines like Azoxystrobin or Boscalid + Pyraclostrobin than with Mancozeb, Chlorthalonil or Fenamidon.     

百香果疮痂病病原菌鉴定

To identify the pathogen causing scab disease on the fruit of Passiflora Linn. in the passion fruit yard of Cenxi and Nanning cities, Guangxi province. The scab tissues collected from diseased fruits were isolated,purified,and made pathogenicity test,morphological observation, biological and molecular identification. The representive isolate B31, obtained from the diseased fruit, was selected for morphological observation and pathogen identification, and was verified as the causal agent of the scab disease through Koch’s postulates. The result of morphological observation using light microscope showed that the isolate B31 was very similar to Alternaria in terms of hyphae and spore morphology. Phylogenetic analysis results using multiple gene loci showed that identities of sequences of ITS, Alt al, TEF 1 and endoPG genes of isolate B31 were 100% identical to those of Alternaria alternata. This study clarifies that the pathogen causing scab disease of passion fruit in Guangxi is A. alternata, and this is the first report of A. alternate causing scab on passion fruit in China.     

Genetic Analysis of Pathogenicity and Host-specific Toxin Production of Alternaria alternata Tomato Pathotype by Protoplast Fusion

Several pathotypes of Alternaria alternata are known to produce host-specific toxins (HSTs) as agents of pathogenicity or virulence. However, investigations into the genetic controls of HST biosynthesis and pathogenicity of Alternaria pathogens have been limited by the lack of a sexual stage in the life cycle of these pathogens. We report here the development of a protoplast fusion system and its use for genetic analysis of HST production and specific pathogenicity of the tomato pathotype of A. alternata that produces AAL-toxin as a HST. Drug-resistant transformants have been isolated by genetic transformation of nonpathogenic A. alternata (strain O-94) and A. alternata tomato pathotype (strain As-27) with vectors conferring resistance to hygromycin B and geneticin, respectively. Protoplasts of the respective transformants were fused by polyethylene glycol treatment or electrofusion. Fusion products were selected by culturing in the presence of both hygromycin B and geneticin, then confirmed by amplification using a polymerase chain reaction with specific primers to the transforming drug-resistance genes. Stable fusants were purified by successive subcultures on selective medium and single-spore isolation. The resultant stable fusants, probably inter-strain hybrids, had the same pathogenicity and toxin production as the wild-type strain As-27. These results suggest that protoplast fusion has potential applications for genetic analysis of A. alternata pathogens. Received 8 September 2000/ Accepted in revised form 27 October 2000     

Evaluation of the pathogenicity of microorganisms isolated from Egyptian broomrape (Orobanche aegyptiaca) in Israel

Diseased Egyptian broomrape ( Orobanche aegyptiaca ) inflorescences were collected from a heavily broomrape-infested tomato ( Lycopersicon esculentum ) field in Israel. The microorganisms that were isolated from the diseased inflorescences were passed through Koch's postulates on Egyptian broomrape-parasitizing tomato roots in a polyethylene bag system and pots under greenhouse conditions. The fungi, Alternaria alternata , Macrophomina phaseolina , Rhizoctonia solani , and Fusarium solani , and the bacterium, Bacillus sp., were newly isolated from the diseased inflorescences of Egyptian broomrape and were found to be pathogenic to the parasite. Fusarium solani damaged all of the developmental stages of broomrape and prevented the damage that Egyptian broomrape causes to tomato plants. The level of pathogenicity and the damage of M. phaseolina , A. alternata , and Bacillus sp. to Egyptian broomrape in greenhouse experiments were relatively low. All the tested microorganisms are known as pathogens of tomato, yet none caused disease symptoms on the tomato plants grown in the inoculated polyethylene bags or in the pots. Fusarium solani demonstrated the highest potential for further development as a mycoherbicide for Egyptian broomrape control in tomato.