马铃薯晚疫病菌单孢分离物生物学特性的初步研究*

研究了马铃薯晚疫病菌单游动孢子分离的方法,并对其单游动孢子分离物菌落生长直径、产孢量以及对甲霜灵的敏感性进行了初步研究。结果表明同一菌株的不同单游动孢子菌落生长直径和产孢量有显著差异,但不同游动孢子分离物对甲霜灵敏感性没有显著差异。     

马铃薯晚疫病菌单孢分离物生物学特性的初步研究

研究了马铃薯晚疫病菌单游动孢子分离的方法,并对其单游动孢子分离物菌落生长直径、产孢量以及对甲霜灵的敏感性进行了初步研究。结果表明同一菌株的不同单游动孢子菌落生长直径和产孢量有显著差异,但不同游动孢子分离物对甲霜灵敏感性没有显著差异。     

双孢蘑菇遗传多样性分析

应用AFLP指纹技术对双孢蘑菇的20个野生菌株和5个栽培品种的遗传多样性进行了研究。AFLP指纹揭示出20个野生异核体菌株所固的基因型。5个商业品种表现出比较一致的AFLP指纹,但也显示出它们之间的一些差别。由单孢分离获得的同核体菌株携带着部分异核体菌株的AFLP指纹;由同一子实体分离得到的大部分单孢菌株是异核体菌株,它们具有与其亲本一致的AFLP指纹。UPGMA分析揭示出2个与地理分布（美洲、欧洲）和相对应的组。研究结果表明：（1）在野生菌株之间存在着明显的遗传差异;（2）大多数单孢分离的菌株具有与母本一致的遗传物质;（3）野生菌株间的遗传变异大于栽培品种间的变异;（4）在双孢蘑菇的遗传多样性分析中,AFLP技术是非常有效的工具。     

胶孢镰刀菌产生串珠镰刀菌素的不稳定性

从陕北克山病病区分离到的两株串珠镰刀菌素产生菌株——胶孢镰刀菌(Fusarium subglutinans Wollenw.et Reinking)陕-6号和2-17号进行单孢分离,分别得到23和19个单孢分离株。这些单孢菌株可分为两种培养型:一种形态上与原始菌株相似,产生串珠镰刀菌素,产色素,具有大、小分生孢子,转管八次产毒量有下降;另一种则不产串珠镰刀菌素和孢子,无色素,后者在二株菌的单孢分离菌中所占比例分别为60.9％和15.8％。由此可见,胶孢镰刀菌产毒稳定性受异核体和该菌单核变异性共同影响。     

胶孢镰刀菌产生串珠镰刀菌素的不稳定性

从陕北克山病病区分离到的两株串珠镰刀菌素产生菌株－胶孢镰刀菌陕－６号和２－１７号进行单孢分离,分别得到２３和１９个单孢分离株。这些单孢菌株可为两种培养型：一种形态上与原始菌株相似,产生串珠镰刀菌素,产色素,具有大、小分生孢子,转管八次产毒量有下降,另一种则不产串珠镰刀菌素和孢子,无色素,后者在二株菌的单孢分离菌中所占比例分别为６０．９％和１５．８％。由此可见,胶孢镰刀菌产毒稳定性受异核体和该菌单     

分离自黄瓜的多主棒孢霉不同表型菌株对杀菌剂的敏感性

【目的】通过对分离自黄瓜的多主棒孢霉不同表型菌株适宜生长温度、产孢量等表型特征和对8种杀菌剂的敏感性研究,为多主棒孢霉侵染引起的黄瓜叶斑病和茎疫病的化学防治提供技术支持。【方法】采用温度梯度法测定病菌适宜生长温度;采用PDA培养基25°C黑暗培养5 d和21 d,计测不同表型菌株单位面积产孢量;采用含药平板法测定不同表型菌株对8种杀菌剂的敏感性。【结果】分离自黄瓜的多主棒孢霉不同表型菌株适宜生长温度为25-30°C;在PDA平板25°C黑暗培养5 d,气生菌丝稀少型菌株cu-4、cu-5即大量产孢,产孢量明显多于气生菌丝丰茂型菌株;在试验浓度下,5个试验菌株对8种杀菌剂的敏感性依次为:代森锰锌氟硅唑戊唑醇苯醚甲环唑速克灵百菌清嘧菌酯多菌灵。【结论】分离自黄瓜的多主棒孢霉不同表型菌株在适宜生长温度、菌丝生长速度、产孢量及对杀菌剂的敏感性等方面均存在差异。在试验浓度下,供试菌株对多菌灵和嘧菌酯的敏感性极低(抑制率40%),这2种杀菌剂已失去对该地区黄瓜褐斑病的防控作用。     

球孢白僵菌单孢子分离株在继代培养过程中菌落局变的遗传分析

研究了球孢白僵菌Beauveria bassiana单孢株继代过程中的菌落局变现象,并通过AFLP分析了角变子与原菌株间在分子水平上的差异。结果表明,野生型出发菌株Bb13的单孢分离株13S5和13S8在继代培养过程中均发生生长速率加快和产孢量下降现象。13S5在前5代,13S8在前6代产孢量有所下降但不显著,至第10代时产孢量比第1代分别下降了81.7%和69.0%。菌株角变现象在继代培养5-6代后表现明显,而13S8角变子出现的频率更高。AFLP指纹图谱分析表明,用20个引物组扩增出的98个位点中,两个角变子与野生型菌株间有12条差异条带,变异率达12.2%。由此证明单孢子分离株在继代培养中发生菌落局变后遗传物质已产生了变异。     

链格孢霉醇和链格孢霉醇单甲醚产生菌株的筛选

本文对分离自小麦、马铃薯、番茄和茄子上链格孢霉属(Alternaria)2个种(链格孢和茄链格孢)的96个菌株,用枯草杆菌生长抑制试验筛选链格孢霉醇(AOH)和链格孢霉醇单甲醚(AME)的产生菌株,有48株产生毒性作用(占所测菌株的50％)。18株产强、中毒性菌用高效液相色谱分析,有13株产AOH和AME(占所测菌株的72.2％)。链格孢的产毒素菌株率比茄链格孢低。但产毒素含量却是前者明显高于后者。其中产AOH和AME的最高含量,链格孢菌株XA-8分别为280和5140mg/kg,而茄链格孢菌株SA-10分别为95.9和94.3mg/kg。     

板栗疫病菌cpfluG基因的功能研究

fluG基因在许多丝状真菌中对无性孢子的发育起调控作用,然而在树木病原菌板栗疫病菌(Cryphonectria parasitica)中,无性孢子的发育调控机制尚未完全明确。本研究克隆了板栗疫病菌的fluG同源基因cpfluG,采用同源重组的方法构建了fluG基因缺失菌株ΔcpfluG。相对于野生型菌株,ΔcpfluG突变株几乎完全丧失产孢能力,色素分泌显著减少,但致病性不受影响。cpfluG基因的缺失导致已知的产孢相关基因cls-31表达量显著下调,但不影响pks基因和cls-32的表达。本研究结果揭示,fluG是板栗疫病菌孢子发育调控的一个关键成员,但板栗疫病菌分生孢子发育的调控细节可能有别于已报道的丝状真菌。     

响应面法优化蝉棒束孢SLGY-2产孢条件

棒束孢菌Cordyceps sp.是一类重要的昆虫病原真菌,具有开发潜能。前期(通过生物毒力测定)筛选一株对斜纹夜蛾具有较高毒力蝉棒束孢SLGY-2菌株,为进一步提高其产孢活力,分别利用单因素方差法和中心组合设计配合响应面分析两种方法开展了本次研究。本研究以产孢量为指标,选择培养基营养成分含量、pH值、温度以及光照条件等进行单因素试验,通过响应曲面法优化最佳产孢条件。结果表明:SLGY-2菌株在实际和模拟较为吻合,模型中最佳产孢条件为培养基营养成分含量为38.3 g、pH 6.55、温度25.21℃、光照18.04 h。在该培养条件下,产孢量达到5.70×10~8个/mL,可用于孢子批量生产。     

河北省番茄上致病疫霉Phytophthora infestans群体结构的分析

番茄晚疫病是河北省番茄生产上最具毁灭性的病害之一,对引起该病害的致病疫霉群体结构进行分析有利于病害的防治。利用对峙培养法和菌落直径法对2007-2008年采自河北省保定、沧州和唐山分离自番茄的49个致病疫霉菌株进行了交配型和甲霜灵抗性的表型测定,结果表明该群体所有菌株均为A1交配型,以甲霜灵敏感菌株为主,抗性菌株仅7株。利用聚合酶链式反应-限制性片段长度多态性(PCR-RFLP)、简单序列重复(SSR)和扩增片段长度多态性(AFLP)等分子技术对该群体的基因型进行了分析,结果表明供试菌株线粒体基因型均为Ia型,共鉴定出了Ⅰ、Ⅱ和Ⅲ3种SSR基因型,AFLP聚类分析在相似系数0.87时可以形成α、β和γ等3个不同的分支。河北省所有番茄上致病疫霉菌株均分布在α分支上,该分支又可进一步分为7个亚分支。AFLP亚分支与甲霜灵抗性和地理来源均无明显相关性,但Ⅱ型SSR与甲霜灵抗性和地理来源有明显的相关性。综合表型和基因型数据说明河北省番茄上致病疫霉群体结构比较单一,遗传多样性程度较低。     

Genetic diversity of Rhizoctonia solani AG-3 from potato and tobacco in North Carolina

Anastomosis group 3 (AG-3) of Rhizoctonia solani (teleomorph = Thanatephorus cucumeris) is frequently associated with diseases of potato (AG-3 PT) and tobacco (AG-3 TB). Although isolates of R. solani AG-3 from these two Solanaceous hosts are somatically related based on anastomosis reaction and taxonomically related based on fatty acid, isozyme and DNA characters, considerable differences are evident in their biology, ecology, and epidemiology. However, genetic diversity among field populations of R. solani AG-3 PT and TB has not been documented. In this study, the genetic diversity of field populations of R. solani AG-3 PT and AG-3 TB in North Carolina was examined using somatic compatibility and amplified fragment length polymorphism (AFLP) criteria. A sample of 32 isolates from potato and 36 isolates from tobacco were paired in all possible combinations on PDA plus activated charcoal and examined for their resulting somatic interactions. Twenty-eight and eight distinct somatic compatibility groups (SCG) were identified in the AG-3 PT and AG-3 TB samples, respectively. AFLP analyses indicated that each of the 32 AG-3 PT isolates had a distinct AFLP phenotype, whereas 28 AFLP phenotypes were found among the 36 isolates of AG-3 TB. None of the AG-3 PT isolates were somatically compatible or shared a common AFLP phenotype with any AG-3 TB isolate. Clones (i.e., cases where two or more isolates were somatically compatible and shared the same AFLP phenotype) were identified only in the AG-3 TB population. Four clones from tobacco represented 22% of the total population. All eight SCG from tobacco were associated with more than one AFLP phenotype. Compatible somatic interactions between AG-3 PT isolates occurred only between certain isolates from the same field (two isolates in each of four different fields), and when this occurred AFLP phenotypes were similar but not identical.     

Occurrence of fenhexamid resistance in Botrytis cinerea from greenhouse strawberries in China

This study assessed the fenhexamid sensitivity of 143 Botrytis cinerea isolates collected from greenhouse strawberries in five regions of China between 2012 and 2013, and identified four isolates with moderate levels of resistance: two from the Xinjiang Uygur Autonomous Region and two from Hebei Province. The baseline fenhexamid sensitivity of B. cinerea exhibited a unimodal distribution with a mean EC₅₀ value of 0.20 ± 0.10 μg/ml (SD). The EC₅₀ values of the fenhexamid‐resistant isolates ranged from 0.05 to 0.40 μg/ml. Molecular analysis of the fenhexamid target gene erg27 revealed that the resistant isolates collected from Xinjiang (163‐6 and 163‐22) contained three mutations that led to amino acid changes (V365A, E368D and A378T) known to be associated with fenhexamid resistance, but that the isolates from Hebei lacked any mutations, indicating that an alternative mechanism could be responsible for their resistance. Most of the biological characteristics of the fenhexamid‐resistant isolates, such as mycelial growth, sclerotia production and pathogenicity, did not significantly differ from those of the sensitive ones (p ≤ .05), but it was noted that some of the resistant isolates exhibited reduced rates of sporulation and spore germination. In addition, the resistant isolates exhibited lower osmotic sensitivity than the sensitive ones. The study found no evidence of cross‐resistance with other fungicides, but that there was negative cross‐resistance with procymidone, iprodione, carbendazim and pyraclostrobin, which indicates that the inclusion of these fungicides within an integrated pest management (IPM) programme could help to minimize the risk of fenhexamid resistance developing in B. cinerea.     

河北省苹果园根际土壤中疑似致病镰孢菌种类

为了解引起河北省苹果再植病害的病原菌,在河北省10个地区苹果园中采集土壤样品,在实验室进行病原菌的诱集分离培养,根据形态和分子特征对主要病原菌进行种类鉴定。结果表明,在分离得到的293株真菌中,有116株镰孢菌,为分离频率最高的真菌。在形态学鉴定的基础上,对供试镰孢菌进行了分子鉴定。在基于核糖体基因内转录间隔区（rDNA-ITS）序列与翻译延长因子1α（EF-1α）序列片段构建的系统发育树中,代表菌株分别与GenBank登记的所属菌株聚于同一群。研究结果明确了河北省苹果再植病害的疑似致病镰孢菌,包括：尖孢镰孢Fusarium oxysporum、木贼镰孢F. equiseti、锐顶镰孢F. acuminatum、层出镰孢F. proliferatum和茄腐镰孢F. solani。     

Morphological and Molecular Characterization of Fusarium solani Isolates

Fusarium solani is a species complex (FSSC) containing isolates that cause diseases in important crops such as root and fruit rot of Cucurbita spp., root and stem rot of pea, sudden death syndrome of soybean, foot rot of bean and dry rot of potato tubers during storage. Based on host range tests, F. solani were subdivided into different formae specialis (f. sp.) and varieties, while DNA sequences of 28S rDNA, internally transcribed spacers (ITS) rDNA and elongation factor (EF-1α) distinguished the ' F. solani complex' in 50 subspecific lineages. In this study we characterized, by cultural, morphological and molecular criteria, 34 isolates of F. solani obtained from potato, other crops and soil. The 34 isolates in the FSSC showed wide variability for their cultural, morphological and molecular traits. The wide variability observed with amplified fragment-length polymorphism (AFLP) and mini-microsatellite analyses is in agreement with the polymorphism observed, in a previous study, within FSSC. Nine of 34 isolates in the FSSC, classified as F. solani var. coeruleum , were morphologically distinguishable from the other F. solani isolates but they were distributed in different clusters; moreover, the nine isolates showed instability of the coeruleum pigmentation of the colonies, supporting the ambiguity of the taxa of this variety of F. solani. Using sequence data from ITS plus 5.8S rDNA region, the isolates were classified into different clades. In particular eight isolates were classified into a well-supported clade including F. solani f. sp . pisi , nine into a clade including only isolates of F. solani f. sp . radicicola and four into a clade including F. solani f. sp . cucurbitae , but this classification could not be used if is not in agreement with host specificity. Two of the nine F. solani var. coeruleum isolates were phylogenetically distinct from all the other FSSC strains.     

河北鸭梨病果上链格孢分离系的形态及分子特性研究

A total of 123 Alternaria isolates were obtained from roting fruits of Pyrus bretschneideri "Ya Li" collected in Hebei Province, China. The isolates, according to morphological characteristics of conidia and sporulation patterns, were segregated into four groups: A. alternata group, A. tenuissima group, A. yaliinficiens group and Alternaria sp. group. Molecular characteristics of part of the isolates were determined using random amplified polymorphism DNA (RAPD) analysis. Based on cluster analysis of RAPD data, large-and small-spored Alternaria spp. were evidently distinguished, and the small-spored species can form five clusters that fundamentally paralleled the morphological groupings.     

Characterization and Pathogenic Relationships of Rhizoctonia solani Isolates in a Potato–Rice System and their Sensitivity to Fungicides

Potato is planted after rice in several parts of Punjab in India and both crops are attacked by Rhizoctonia solani Kühn. Potato tubers showing black scurf and rice plants affected by sheath blight were collected from different regions of the state and the isolates of R. solani so obtained were studied to determine their variability and to ascertain their cross-infectivity and response to fungicides. Potato isolates of R. solani did not infect rice plants but some rice isolates were weakly pathogenic on potato, the sclerotia being less firmly attached on tuber surface, indicating a possible unsuccessful attempt of rice isolates to infect potato. Rice isolates (66.6%) grew faster (>20 mm colony growth per 24 h) than those of the potato isolates (15–20 mm growth rate per 24 h). Hyphal width of isolates from both hosts varied from 7.2 to 12.1 μm. Colony growth of most potato isolates (61.2%) was appressed, whereas that of most rice isolates (53.3%) was fluffy. Rice isolates (73.3%) formed larger sclerotia (1.5–2.0 mm in diameter) than those of the potato isolates (0.5–1.0 mm in diameter). Anastomosis studies indicated that potato isolates belonged to AG-3 and AG-5 groups while rice isolates belonged to the AG-1-1-A group. Representative R. solani isolates from the two hosts showed significant variation in response to fungicides (i.e. carbendazim, carboxin, pencycuron, propiconazole and validamycin) based on their ED₅₀ and ED₉₀ values.     

Phytophthora infestans isolates from Northern China show high virulence diversity but low genotypic diversity

Phenotypic and genotypic characteristics of 48 Phytophthora infestans isolates , collected in five provinces in Northern China between 1997 and 2003, were determined and compared with reference isolates. Characterisation included mating type, virulence, mitochondrial DNA (mtDNA) haplotype and DNA fingerprinting patterns based on simple sequence repeats (SSR) and amplified fragment length polymorphisms (AFLP). All isolates had the A1 mating type, mtDNA haplotype IIa and an identical SSR genotype (designated as SG-01-01) that differed from SSR genotypes found in the reference isolates, including those representing the 'old' US-1 lineage that dominated the P. infestans population worldwide prior to 1980. In contrast, the virulence spectra were highly variable and virulence to all resistance genes present in the standard differential set ( R1 to R11 ) was found. AFLP analysis revealed some diversity; eight different AFLP genotypes were found that could be grouped into two major clusters. This study shows that there is very little genotypic diversity in the P. infestans population in Northern China. The occurrence of many different races within this rather uniform population is discussed in the framework of recent insights into the molecular determinants of avirulence in potato– P. infestans 'gene-for-gene' interactions.     

Study of the aggressiveness of Rhizoctonia solani isolates

This paper presents an in vitro test to screen the pathogenicity of different Rhizoctonia solani isolates on a host range. The level of aggressivity of the different isolates was different for several host plants tested. There were significant differences between the crops and the isolates tested. In general, the disease level was higher on beans, lettuce and cabbage. In carrot and rye grass the level of infection was lower for the isolates of R. solani tested. The potato isolates of R. solani were less aggressive than the isolates coming from maize, fodder beet and sugar beet. The R. solani isolates were also biochemically characterized by pectic zymograms: the isolates Rs0401 (from maize) and Rs0504 (from sugar beet) belong both to the anastomosis group AG2-2.     

ERIC and REP-PCR Banding Patterns and Sequence Analysis of the Internal Transcribed Spacer of rDNA of Stemphylium solani Isolates from Cotton

The genetic diversity of the Stemphylium solani isolates from cotton was assessed by Enterobacterial Repetitive Intergenic Consensus (ERIC) and Repetitive Extragenic Palindromes (REP)-PCR fingerprinting. Twenty eight monosporic isolates of S. solani from cotton were used along with five isolates from tomato and one isolate of Alternaria macrospora from cotton for comparison. The dendrogram obtained revealed clear differences between the cotton and tomato isolates as well as between the tomato isolates from different geographic regions. The genetic relationships among S. solani isolates were also analyzed by sequencing the internal transcribed spacer (ITS) region of four isolates representing the three ERIC and REP groups. The tomato isolate from the State of S?o Paulo showed a distinct ITS sequence from that of the cotton isolates and tomato isolate from the State of Goiás, giving evidence that it belongs to a different genotype of S. solani. This is the first report of the entire sequence of the ITS1-5.8S-ITS2 regions of S. solani.