河北鸭梨黑斑病病原菌的鉴定

鸭梨黑斑病菌株 ,在PCA平板上 ,菌落直径 69 2～ 73 5mm ,有明显浅灰与灰褐色交替的同心轮纹。分生孢子梗单生 ,浅褐色 ,有隔 2～ 7个 ,长 2 0 5～ 59 0 μm ,合轴延生或不延生。以孔生方式产生分生孢子 ,孢子串生 ,孢子链多次分枝 ,孢子链 5～ 1 1个孢子。孢子倒棍棒形 ,长卵形 ,黄褐色至黑褐色 ,有横隔 3～ 6个 ,纵隔 0～ 3个 ,孢子尺度 1 2 5～ 42 0 μm× 5 5～ 1 3 0 (- 1 4 5) μm ,喙长 0～ 8 0 μm。根据病菌形态特征 ,黑斑病菌鉴定为链格孢Alternariaalternata(Fr .)Keissl.     

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.     

黄骅冬枣细极链格孢的分离鉴定

从黄骅冬枣发病叶片上分离到一株真菌L5-4-2,对其进行了菌落形态、分生孢子形态特征观察和致病性测试。ITS序列分析结果表明,该分离物L5-4-2与链格孢属(Alternaria sp.)(JN689942.1、JN545807.1、JF694748.1、MN115554.1)序列的相似性均在99.25%以上;ACT序列分析表明,分离物L5-4-2与细极链格孢(Alternaria tenuissima)在同一个分支,且Bootstrap支持率为100%。结合菌落形态特征、分生孢子形态特征及致病性测试结果 ,将其鉴定为细极链格孢[Alternaria tenuissima (Fr.) Wiltshire]。     

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。     

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.     

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。     

用于虫生真菌遗传转化的苯菌灵抗性菌株筛选

发展虫生真菌的抗药性是协调生防与化防,稳定真菌杀虫剂应用效果的一个重要策略。本研究从代表性地区采集植物病原样本,分离得到35株菌株,鉴定为3属5种,即灰葡萄孢霉Botrytis cinerea、扩展青霉Penicillium expansum、指状青霉Penicillium digitatum、长柄链格孢菌Alternaria longipes和茄链格孢菌Alternaria solani。应用菌落直径法测定其对苯菌灵的抗性,结果表明：供试的11株灰葡萄孢霉菌株中,最高EC50为936.2670μg·mL^-1;12株青霉属菌株中,最高EC50为0.0384μg·mL^-1;12株链格孢菌株中,最高EC50为703.5557μg·mL^-1。其中灰霉和链格孢属菌株中有4株EC50大于600μg·mL^-1的高抗菌株,即灰葡萄孢霉的菌株SD2和SD1以及茄链格孢的菌株CC2-2和LF1-2,这4株菌株可作为遗传转化抗性基因克隆的基础材料。     

云南省荞麦叶枯病病原菌鉴定及其生物学特性

为明确云南省荞麦叶枯病病原菌种类,采用常规组织分离法获得病原菌菌株LW2015.3,通过形态学特征及分子生物学技术对菌株LW2015.3进行鉴定,并研究其生物学特性。结果表明,荞麦叶枯病病原菌的分生孢子呈倒棍棒状或倒梨状,褐色,具3～8个横隔膜,0～4个纵斜隔膜,大小为16.5～45.0μm×5.0～13.5μm,厚垣孢子呈球形,直径为6.0～12.0μm;该菌株ITS序列系统发育进化分析结果表明,菌株LW2015.3与链格孢Alternaria alternata(登录号:MG195995.1)的同源性为100%,结合形态学特征与分子鉴定结果确定云南省荞麦叶枯病病原菌为链格孢A. alternata(登录号:KT362732.1)。该病原菌菌丝生长适宜温度为20～30℃,25℃为最适温度;当p H为6～9时菌丝生长速率加快,pH 7最适菌丝生长;PDA培养基和PSA培养基最适菌丝生长;该病原菌对以麦芽糖为碳源和以硝酸钠为氮源时的利用率最高;菌丝的致死温度为50℃,10 min;不同光照条件对菌丝生长的影响有显著差异,连续光照最有利于菌丝生长。     

线虫生防真菌淡紫紫孢菌PLHN鉴定及其生物学特性

从为害海南省茄子的根结线虫(Meloidogyne sp.)雌虫体内分离获得一株对根结线虫具致病性的真菌菌株PLHN,研究了该菌株的培养特性和形态特征。同时,克隆分析其r DNA-ITS区序列,并测定该菌株的生物学特性。结果表明,菌株PLHN在PDA上菌落呈淡粉紫色,富生粉质状分生孢子,菌落背面中心呈炒米黄色;分生孢子梗上轮生2～6个瓶梗,大小为(2.81±0.43)μm×(6.27±1.18)μm;分生孢子近圆形,无色,大小为(2.60±0.40)μm×(2.63±0.38)μm。该菌株rDNA-ITS区序列与Genbank中多个淡紫紫孢菌(淡紫拟青霉)菌株的对应序列相似性均达99%～100%。因此,将PLHN菌株鉴定为淡紫紫孢菌(Purpureocillium lilacinum)。该菌株最适培养基为PDA,菌落生长、产孢和孢子萌发的最适pH均为7.0,最适培养温度均为28℃,黑暗有利于该菌生长、产孢及孢子萌发。     

进口美国苹果上拟盘多毛孢菌的鉴定

从进口的美国华盛顿州红帅(reddelicious)苹果中分离到一种引起苹果果实腐烂的病原真菌,根据该真菌的菌落及分生孢子的形态特征,以及该真菌DNA的ITS序列,将其鉴定为拟盘多毛孢菌(Pestaloti-opsissp.)。该真菌菌丝生长和孢子萌发的最适温度是25℃。分生孢子长梭形直立或稍弯曲,15·6~22·2(18·8)μm×4·8~7·1(5·7)μm,5个细胞,4个隔膜均为真隔膜,中间3个细胞暗褐色,两端细胞淡色,3个有色细胞中第1、2个有色细胞颜色较深,茶褐色;第3个有色细胞淡褐色;分隔处稍缢缩;孢子顶端有2~3根不分支的附属丝,长7·0~19·5(11·9)μm,基细胞具短柄,长0·4~3·2(1·4)μm。分生孢子梗较长,呈淡色,不分支,无隔膜。用ITS区通用引物PM1/PM4对该真菌DNA进行PCR扩增和测序,获得了525bp的ITS区序列。     

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.     

壶瓶枣褐斑病病原菌的鉴定

 近几年来,山西红枣发生了1种严重的果实病害,症状表现为果顶或果肩部位形成红褐色的病斑。本研究以壶瓶枣为材料,对病菌进行分离。通过室内和田间致病性测定以及人工接种后再分离病菌,证明编号为CN535的真菌菌株为该病的致病菌。该病菌在PDA上7d菌落直径达69.2~73.5mm,基内菌丝和气生菌丝均发达,具明显的浅灰与墨绿色的同心轮纹;分生孢子单生或短链生,具纵横隔膜和短喙,大小为(22.5~40.0)μm×(8.0~13.5)μm,为典型的Alternaria属真菌特征。其rDNAITS序列分析结果表明该菌与A. alternata、A. tenuissima、A. longipes、A. mali和A. citri的同源性均为100%。用2对链格孢菌的专用引物AAF₂/AAR3和Aalt-F/Aalt-R分别扩增出相对应的341和450bp的片段。综合形态特征和分子分析结果,确定壶瓶枣褐斑病的病原菌为A. alternata (Fries) Keissler。     

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.     

Citrus Black Rot is Caused by Phylogenetically Distinct Lineages of Alternaria alternata

ABSTRACT Phylogenetic analysis revealed that isolates of Alternaria alternata causing black rot of citrus were associated with six well-supported evolutionary lineages. Isolates recovered from brown spot lesions on Minneola tangelo, leaf spot lesions on rough lemon, and healthy citrus tissue and noncitrus hosts were related closely to isolates from black-rotted fruit. Phylogenies estimated independently from DNA sequence data from an endopolygalacturonase gene (endoPG) and two anonymous regions of the genome (OPA1-3 and OPA2-1) had similar topologies, and phylogenetic analysis was performed on the combined data set. In the combined phylogeny, isolates from diverse ecological niches on citrus and noncitrus hosts were distributed in eight clades. Isolates from all lineages, regardless of ecological or host association, caused black rot in fruit inoculation assays, demonstrating that small-spored Alternaria isolates associated with different ecological niches on citrus and other plant hosts are potential black rot pathogens. These data also indicated that the fungi associated with black-rotted fruit do not form a natural evolutionary group distinct from other Alternaria pathogens and saprophytes associated with citrus. The use of the name A. citri to describe fungi associated with citrus black rot is not justified and it is proposed that citrus black rot fungi be referred to as A. alternata.     

Japanese pear black spot and apple alternaria blotch

The pathogenicity and taxonomy of 15 isolates of Alternaria spp. from pear and apple were compared. Only isolates from Asian pear ( Pyrus pyrifolia ) from Italy and Korea were virulent on leaves and young fruits of the susceptible Asian pear cv. Nijisseiki. Their conidial morphology was typical of A. gaisen (= A. kikuchiana ). Only isolates of A. mali from USA were virulent on susceptible American apple cvs Indo and Red Gold. No virulence was demonstrated in any isolate/host combination among isolates from stem infections of Asian and European pear ( Pyrus communis ), leaf spots of European apple, and ripe fruit rots of Chinese pear ( Pyrus ussuriensis ), European pear and apple. These non-virulent isolates could be readily distinguished from A. gaisen and A. mali by their pattern of branching of conidial chains, the branching associated with A. alternata sensu stricto being most common among non-virulent isolates. This limited survey implies that A. gaisen is only virulent to Asian pear and the toxigenic form of A. mali to certain American apple cultivars; also that A. gaisen is not established outside eastern Asia or the toxigenic form of A. mali outside eastern Asia and parts of USA.     

中国柑橘黑腐病和褐斑病病原菌的系统发育分析

 近些年链格孢属真菌的分类有了长足的进展,但柑橘褐斑病和黑腐病的病原却还是存在着一些争议。本研究从中国柑橘果实上收集了49株黑腐病菌菌株,并选取了8株从柑橘叶片上分离的具代表性的褐斑病菌菌株,基于Alta1 、endoPG、LSU、OPA10-2、OPAl-3和OPA2-1 等6个基因位点串联序列构建系统发育树。结果表明,柑橘黑腐病和褐斑病都可由不止一种链格孢菌引起,均以Alternaria alternata为主。两种病害的病原菌之间不能通过该系统发育树区分,但在致病性上存在差异,且能通过扩增ACT毒素合成基因进行区分。为了使两种病害的病原更加方便阐述,作者建议以它们的主要类群对其进行命名,柑橘褐斑病病原学名还是遵循前人的称呼,为the tangerine pathotype of A. alternata,即链格孢菌橘致病型,而柑橘黑腐病病原应为A. alternata,即链格孢菌。     

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。     

云南、广西三七黑斑病病原链格孢菌的鉴定

正黑斑病是三七栽培生产中常见的一大病害,叶片受害产生近圆形或不规则水浸状病斑,常导致成株落叶、幼苗生长点及茎秆顶端腐烂枯死。其病原一般认为是链格孢属真菌人参链格Alternaria panax Whetzel~([1,2]),也有相关研究证明黑斑病病原为细链格孢Alternaria tenuis Nees~([3]),后定名为链格孢Alternaria alternata Keissl~([4])。本研究利用ITS序列和histone 3部分编码序列的PCR鉴定,结合形态学鉴定,分析三七主产区黑斑病菌的组成和分布情况及几种病原菌的致病力差异,以期为三七黑斑病防治提供理论依据。     

中国柑橘黑腐病和褐斑病病原菌的系统发育分析

 近些年链格孢属真菌的分类有了长足的进展,但柑橘褐斑病和黑腐病的病原却还是存在着一些争议。本研究从中国柑橘果实上收集了49株黑腐病菌菌株,并选取了8株从柑橘叶片上分离的具代表性的褐斑病菌菌株,基于Alta1 、endoPG、LSU、OPA10-2、OPAl-3和OPA2-1 等6个基因位点串联序列构建系统发育树。结果表明,柑橘黑腐病和褐斑病都可由不止一种链格孢菌引起,均以Alternaria alternata为主。两种病害的病原菌之间不能通过该系统发育树区分,但在致病性上存在差异,且能通过扩增ACT毒素合成基因进行区分。为了使两种病害的病原更加方便阐述,作者建议以它们的主要类群对其进行命名,柑橘褐斑病病原学名还是遵循前人的称呼,为the tangerine pathotype of A. alternata,即链格孢菌橘致病型,而柑橘黑腐病病原应为A. alternata,即链格孢菌。