Association of Fusarium Species in the Wheat Stem Rot Complex

Data from a national survey were analysed to investigate whether there was interdependence among the Fusarium species, which cause the stem rot complex of wheat. About 25 wheat stems were sampled from each of 260 sites over the main wheat growing areas in the UK. Occurrence of each Fusarium species on individual stems was determined. Fusarium culmorum, F. avenaceum and Microdochium nivale were the three dominant species, detected in 248, 185 and 239 out of the 260 sites. There were no interactions among species in the distribution of the three species over the 260 sites. Several statistical tests were used to determine whether there was interdependence among the three species on the same stem within each site. Of the three species, there was only limited evidence of competition between F. culmorum and F. avenaceum.     

快速鉴定草莓枯萎病的LAMP检测技术

[目的]建立草莓枯萎病的镰刀菌快速检测方法.[方法]利用环介导等温扩增技术(LAMP)针对镰刀菌保守区域ITS设计一套LAMP扩增引物,利用Bst DNA聚合酶完成LAMP扩增反应,对7种常见病原菌进行检测,分析LAMP引物的特异性;比较LAMP与传统PCR方法的敏感性.[结果]建立的LAMP法能够特异性的检测尖孢镰刀...     

Detection and Differentiation of Trichothecene and Enniatin-Producing Fusarium Species on Small-Grain Cereals

A large number of Fusarium species are associated with Fusarium head blight of wheat and other small-grain cereals as well as seedling blight and brown foot rot. Different Fusarium species tend to predominate under different environmental conditions and in different regions. In addition to causing disease, these fungi are of particular significance because they produce a number of mycotoxins including the trichothecenes and enniatins that contaminate infected grain. The nature and amount of the mycotoxins that accumulate will alter according to the species or even the particular isolates involved in the infection. It is highly desirable to be able to analyse such complex infections to determine which species and, preferably, which chemotypes are present, in order to understand the factors that affect the pathogenicity of each species and to evaluate the potential risk for contamination of grain with mycotoxins. This paper reports the development of molecular methods, based upon the polymerase chain reaction (PCR), for the detection of mycotoxigenic fungi. Several of the Fusarium species involved are closely related, making the development of specific assays problematic. We describe the development of primers specific to individual species and discuss how this work provides insight into fungal populations and relates to taxonomic studies. In some instances, it is desirable to detect the presence of potential mycotoxin producers rather than individual fungal species. Generic assays have been produced for several genes involved in trichothecene biosynthesis and for enniatin synthetase in order to permit the detection of species able to produce the associated mycotoxins. Additional work is under way to refine assays to enable detection related to the class of trichothecene and chemotype of isolate because of the potential risk posed to human and animal consumers by different trichothecenes.     

Identification of a Novel Fusarium Head Blight Resistance Quantitative Trait Locus on Chromosome 7A in Tetraploid Wheat

ABSTRACT Fusarium head blight (FHB) caused by Fusarium graminearum is one of the most destructive diseases of durum (Triticum turgidum sp. durum) and common wheat (T. aestivum). Promising sources of FHB resistance have been identified among common (hexaploid) wheats, but the same is not true for durum (tetraploid) wheats. A previous study indicated that chromosome 7A from T. turgidum sp. dicoccoides accession PI478742 contributed significant levels of resistance to FHB. The objectives of this research were to develop a genetic linkage map of chromosome 7A in a population of 118 recombinant inbred lines derived from a cross between the durum cv. Langdon (LDN) and a disomic LDN-T. turgidum sp. dicoccoides PI478742 chromosome 7A substitution line [LDN-DIC 7A(742)], and identify a putative FHB resistance quantitative trait locus (QTL) on chromosome 7A derived from LDN-DIC 7A(742). The population was evaluated for type II FHB resistance in three greenhouse environments. Interval regression analysis indicated that a single QTL designated Qfhs.fcu-7AL explained 19% of the phenotypic variation and spanned an interval of 39.6 cM. Comparisons between the genetic map and a previously constructed physical map of chromosome 7A indicated that Qfhs.fcu-7AL is located in the proximal region of the long arm. This is only the second FHB QTL to be identified in a tetraploid source, and it may be useful to combine it with the QTL Qfhs.ndsu-3AS in order to develop durum wheat germ plasm and cultivars with higher levels of FHB resistance.     

Pathogenicity and In Planta Mycotoxin Accumulation Among Members of the Fusarium graminearum Species Complex on Wheat and Rice

ABSTRACT Fusarium head blight (FHB), or scab, is a destructive disease of small grains caused by members of the Fusarium graminearum species complex, comprised of at least nine distinct, cryptic species. Members of this complex are known to produce mycotoxins including the trichothecenes deoxynivalenol (DON) along with its acetylated derivatives and nivalenol (NIV). In this study, 31 strains, belonging to eight species of this complex and originating from diverse hosts or substrates, were tested for differences in aggressiveness and mycotoxin production. Large variation among strains, both in terms of their aggressiveness and the ability to produce trichothecenes on a susceptible cultivar of wheat was found; variation appears to be a strain-specific rather than species-specific characteristic. While pathogenicity was not influenced by the type of mycotoxin produced, a significant correlation was observed between the amount of the dominant trichothecene (DON and its acetylated forms or NIV) produced by each strain and its level of aggressiveness on wheat. Some isolates also were tested for their ability to infect rice cv. M201, commonly grown in the United States. While tested strains were capable of infecting rice under greenhouse conditions and causing significant amount of disease, no trichothecenes could be detected from the infected rice florets.     

基于TaqMan MGB探针的花生矮化病毒检测研究

花生矮化病毒(Peanut stunt virus,PSV)是我国进境检疫性有害生物。本研究根据该病毒不同分离株外壳蛋白基因(coat protein,CP)的保守序列,设计了特异性引物与TaqMan MGB荧光探针,建立了PSV的实时荧光RT-PCR检测方法。方法特异性研究表明,针对PSV的2个不同株系PSV-E和PSV-W,均能够得到典型扩增曲线,Ct值分别为20.10和21.22;而对于黄瓜花叶病毒、番茄不孕病毒、马铃薯Y病毒、菜豆荚斑驳病毒以及烟草环斑病毒等其他毒株则没有典型扩增曲线,也无Ct值。灵敏度比较发现,该方法比普通RT-PCR检测方法的灵敏度提高100倍,具有快速、灵敏和高特异性的优点,适合对PSV的检测。     

利用TaqMan探针实时荧光PCR方法检测香石竹细菌性萎蔫病菌

 根据香石竹细菌性萎蔫病菌基因组16S-23S rRNA保守序列,设计并合成了一对特异性引物和一条具有稳定点突变特异性探针,建立了对香石竹细菌性萎蔫病菌的TaqMan实时荧光PCR检测方法。除香石竹细菌性萎蔫病菌外,还对其他7种病原细菌菌株进行了荧光PCR检测。结果表明,只有香石竹细菌性萎蔫病菌产生荧光,其他病原细菌均没有荧光产生。与常规PCR相比,实时荧光PCR检测特异性强,灵敏度高,能检测到浓度为0.4 pg/μL的DNA,且能直接用于苗木等样品的检测,适合病害的快速诊断和口岸检验检疫应用。      

Antifungal Activity Toward Fusarium culmorum in Soluble Wheat Extracts

ABSTRACT This study investigated antifungal activity in soluble extracts from seed of a range of wheat cultivars differing in susceptibility to Fusarium head blight. Antifungal activity was assessed in terms of beta-D-glucuronidase (GUS) activity of a Fusarium culmorum GUS transformant using a sensitive laboratory assay. Significant antifungal activity was detected in seed extracts from WEK0609, CM 820036, and Arina. Initial characterization of the Arina seed extract indicated that it contained antifungal proteinaceous compounds. The Arina extract yielded two (60 and 80%) ammonium sulfate fractions containing inhibitory compounds. Gel filtration chromatography and subsequent sodium dodecyl sulfate-polyacrylamide gel electrophoresis of antifungal fractions showed that the antifungal activities detected in the Arina 60 and 80% ammonium sulfate fractions were associated with putative proteinaceous compounds with apparent molecular masses of approximately 60 and 28 kDa, respectively.     

利用TaqMan探针检测水稻细菌性谷枯病菌

利用TaqMan探针建立了水稻细菌性谷枯病菌(Burkholderia glumae)实时荧光PCR检测方法。根据水稻细菌性谷枯病菌gyrB基因,设计并合成特异性引物和探针,对8株不同来源水稻细菌性谷枯病菌和其他同属或同寄主的参试菌株进行了检测。结果显示,该方法检测的特异性强,灵敏度可达菌悬液浓度102cfu/mL,该方法快速、简便、准确,适用于出入境检验检疫及种子健康检测领域。利用该方法对国内采集的83份水稻材料进行了检查,未发现阳性结果。     

New Leaf Spot Disease of Cymbidium Species Caused by Fusarium subglutinans and Fusarium proliferatum

Fusarium species were consistently isolated from yellow, swollen spots with reddishbrown centers and small black spots on leaves of Cymbidium plants in the greenhouse. Fusarium subglutinans caused the yellow spots and Fusarium proliferatum caused either the yellow or the black spots. We propose the name “yellow spot” for the new disease. To denote differences in their pathogenicity to orchid plants, we designate the population causing yellow spot as race Y and that causing black spot as race B of F. proliferatum. Received 29 October 1999/ Accepted in revised form 10 March 2000     

Effect of Trichothecenes Produced by Fusarium graminearum during Fusarium Head Blight Development in Six Cereal Species

Fusarium head blight (FHB) is a complex cereal disease associated with trichothecene production; these mycotoxins are factors of aggressiveness in wheat. Six species (bread and durum wheat, triticale, rye, barley and oats) were submitted to point inoculations with two isogenic strains of Fusarium graminearum; a wild strain (Tri5 +) produced trichothecenes and the mutated strain (Tri5 –) did not. The trichothecene-producing strain was generally more aggressive than the non-producing strain, but this varied according to crop species. The difference in aggressiveness was less pronounced in rye, a very resistant species. High resistance levels were observed in oats due to the large spacing between florets. In six-row barley, despite the existence of a moderate Type II resistance, the fungus was often observed to move externally from one floret to another within the dense spike, without penetrating the rachis. Bread wheat had low resistance to the trichothecene-producing strain and good resistance to the non-producing strain. Triticale responded to the strains in a similar way but was somewhat more resistant to both: symptoms on the spikelets and rachis of the triticales were restricted to below the point of inoculation. Durum wheat was susceptible to the trichothecene-producing strain and only moderately resistant to the non-producing strain, which was able to cause serious damage only to this species. Our study confirmed that the role of trichothecenes in FHB pathogenesis differs among species. The failure of the trichothecene non-producing F. graminearum strain to spread within the inflorescence of wheat, triticale, rye and barley, and the significant reduction of spread in the durum wheat spike strongly suggested that trichothecenes are a major determinant of fungal spread and disease development in Triticeae.     

Detection of Fusarium langsethiae on wheat in Belgium

In 2010, the populations of Fusarium sp. and Microdochium sp. were monitored in Belgium and 16 strains were identified as Fusarium langsethiae on wheat in Belgium. The other species identified from the sampling were F. poae, F. tritinctum, F. graminearum, F. avenaceum and Microdochium nivale. The pathogenicity potential of the F. langsethiae strains was assessed via an in vitro coleoptile growth rate test on wheat seedlings and compared with strains of F. poae, F. tritinctum, F. graminearum and F. avenaceum known to cause Fusarium head blight. The results showed the ability of F. langsethiae to cause retardation in the wheat coleoptile growth rate, but at a lower rate than F. graminearum, F. avenaceum, F. poae and F. tricinctum. A test for mycotoxin production in vitro showed the ability of the four strains tested to produce T-2 and HT-2 toxins at a rate of up to 290 mg kg^?1. This is the first report on the potential pathogenicity of F. langsethiae on wheat in Belgium, a species known to produce T-2 and HT-2 toxins, which are highly toxic for humans and animals.     

Analysis of Host Species Specificity of Magnaporthe grisea Toward Wheat Using a Genetic Cross Between Isolates from Wheat and Foxtail Millet

ABSTRACT A genetic cross was performed between a Setaria isolate (pathogenic on foxtail millet) and a Triticum isolate (pathogenic on wheat) of Magnaporthe grisea to elucidate genetic mechanisms of its specific parasitism toward wheat. A total of 80 F(1) progenies were obtained from 10 mature asci containing 8 ascospores. Lesions on wheat leaves produced by the F(1) progenies were classified into four types, which segregated in a 1:1:1:1 ratio. This result suggested that the pathogenicity of the F(1) population on wheat was controlled by two genes located at different loci. This idea was supported by backcross analyses. We designated these loci as Pwt1 and Pwt2. Cytological analyses revealed that Pwt1 and Pwt2 were mainly associated with the hypersensitive reaction and papilla formation, respectively.     

Molecular Detection of Phytophthora ramorum by Real-Time Polymerase Chain Reaction Using TaqMan, SYBR Green, and Molecular Beacons

ABSTRACT Sudden oak death, caused by Phytophthora ramorum, is a severe disease that affects many species of trees and shrubs. This pathogen is spreading rapidly and quarantine measures are currently in place to prevent dissemination to areas that were previously free of the pathogen. Molecular assays that rapidly detect and identify P. ramorum frequently fail to reliably distinguish between P. ramorum and closely related species. To overcome this problem and to provide additional assays to increase confidence, internal transcribed spacer (ITS), beta-tubulin, and elicitin gene regions were sequenced and searched for polymorphisms in a collection of Phytophthora spp. Three different reporter technologies were compared: molecular beacons, TaqMan, and SYBR Green. The assays differentiated P. ramorum from the 65 species of Phytophthora tested. The assays developed were also used with DNA extracts from 48 infected and uninfected plant samples. All environmental samples from which P. ramorum was isolated by PARP-V8 were detected using all three real-time PCR assays. However, 24% of the samples yielded positive real-time PCR assays but no P. ramorum cultures, but sequence analysis of the coxI and II spacer region confirmed the presence of the pathogen in most samples. The assays based on detection of the ITS and elicitin regions using TaqMan tended to have lower cycle threshold values than those using beta-tubulin and seemed to be more sensitive.     

Occurrence and Pathogenicity of Fusarium Species in Cereals in Finland1

Several Fusarium species occurred abundantly in spring cereal seed samples studied in 1966, 1968 and 1972 in Finland. A total of 17 species of Fusarium were isolated and identified. The most frequently isolated species were F. avenaceum (Fr.) Sacc., F. culmorum (W. G. Sm.) Sacc., F. poae (Peck) Wr. and F. tricinctum (Cda) Sacc. These species were found in more than 40% of the seed samples from 1972. The same 4 Fusarium species in addition to F. graminearum Schwabe were used in the pathogenicity test. Artificially inoculated spring wheat and barley seeds were grown in the field during the growing season of 1973. Of these 5 species F. culmorum was clearly the most pathogenic. This species significantly lowered the shooting percentage and the yield of both cereals. Foot rot assessment also revealed F. culmorum as the most damaging species. Natural soil-borne infection obviously increased the foot and root rot, and thus the soil-borne F. culmorum strongly affected all treatments. In the harvested seed F. poae dominated in all treatments, but F. culmorum still occurred most abundantly in the seed lot harvested from plots originally inoculated with this species.     

Major Changes in Fusarium spp. in Wheat in the Netherlands

The re-emergence of fusarium head blight throughout the world and especially in Western Europe prompted a survey of the situation in the Netherlands. To allow for a high throughput screening of large numbers of samples, a diagnostic PCR method was developed to detect the most common species of Fusarium occurring on wheat. Seven primer pairs were tested for their ability to identify isolates of Fusarium avenaceum, F. culmorum, F. graminearum, F. poae, F. proliferatum and Microdochium nivale var. majus and M. nivale var. nivale. Each primer pair only generated a PCR product with the corresponding Fusarium species and all PCR fragments had different molecular sizes. This allowed the generation of these amplicons using a mixture of all seven primer pairs. The robustness of this multiplex PCR encouraged us to screen a large series of isolates collected in 2000 and 2001. In both years 40 fields were sampled leading to a collection of 209 isolates from 2000 and 145 isolates from 2001. The results of the multiplex PCR demonstrated that F. graminearum was the most abundant species in the Fusarium complex on wheat in both years. This is in sharp contrast to reports from the 1980s and early 1990s, which found F. culmorum as the predominant species. Primers derived from the tri7 and tri13 genes, which are implicated in the acetylation and oxygenation of the C-4 atom of the backbone of the trichothecene molecule, were used to discriminate between deoxynivalenol and nivalenol (NIV) producers. The populations of F. culmorum and F. graminearum both showed a slight increase in NIV-producers in 2001.     

Population Analysis of Fusarium graminearum from Wheat Fields in Eastern China

ABSTRACT Wheat heads showing symptoms of Fusarium head blight were collected from four commercial fields in Zhejiang Province, China, an area where epidemics occur regularly. A total of 225 isolates were subjected to population-level analyses using restriction fragment length polymorphism (RFLP) as markers. Diagnostic RFLP markers established that all isolates belonged to Fusarium graminearum lineage 6. Nine polymorphic probes were hybridized to all isolates, resulting in 65 multilocus RFLP haplotypes (MRH). Probing with the telomeric clone pNla17, which reveals differences among isolates in the hypervariable subtelomeric region, differentiated the 65 MRH further into 144 clones. Mean gene diversity for the four field populations was similar, ranging from H = 0.306 - 0.364 over the nine RFLP loci for clone-corrected data. High levels of gene flow were inferred from a low level of population subdivision among all field populations, indicating that they were part of the same population. Pairwise linkage disequilibrium measures did not unequivocally support a random mating population, because one-third of locus pairs were significantly different from the null hypothesis of no-association between alleles. We speculate therefore that sexual recombination may not be frequent and that high levels of genotypic diversity may be maintained by relatively low selection pressure acting on a highly diverse population.     

苜蓿萎蔫病菌TaqMan探针实时荧光PCR检测方法的建立

苜蓿萎蔫病菌是我国对外检疫性二类有害生物，目前国内尚无发生6在出入境捡验检疫中主要是采用生物学和血清学方法进行检测，劳动强度大，耗费时间长。根据苜蓿萎蔫病菌与其它细菌菌株16SrDNA序列差异，设计出对苜蓿萎蔫病菌具有稳定点突交特异性探针，利用该探针对棒形杆菌属4个种及其它属细菌进行了实时荧光PCR检测实验。结果表明，只有苜蓿萎蔫病菌能检测到荧光信号，其它细菌没有荧光产生。该方法特异性强，灵敏度高，能检测到21．4fg质粒DNA，比常规PCR灵敏100倍，而且整个过程只需要2～3h。该方法可有效地应用于进出境病原菌检测之中。     

Influence of Climatic Factors on Fusarium Species Pathogenic to Cereals

Fusarium head blight of small-grain cereals, ear rot of maize, seedling blight and foot rot of cereals are important diseases throughout the world. Fusarium graminearum, F. culmorum, F. poae, F. avenaceum and Microdochium nivale (formerly known as F. nivale) predominantly cause Fusarium diseases of small-grain cereals. Maize is predominantly attacked by F. graminearum, F. moniliforme, F. proliferatum and F. subglutinans. These species differ in their climatic distribution and in the optimum climatic conditions required for their persistence. This review deals with the influence of climate on the production and dispersal of inocula, growth, competition, mycotoxin production and pathogenicity. Most species produce inocula, grow best, and are most pathogenic to cereal heads at warm temperatures and under humid conditions. However, the optimal conditions for F. moniliforme and F. proliferatum maize ear rot tend to be hot and dry and M. nivale head blight, seedling blight and foot rot of small-grain cereals tend to occur under cooler conditions. Seedling blight and foot rot caused by other species are favoured by warm dry weather. Between them, these fungi produce four important classes of mycotoxins: trichothecenes, zearalenone, fumonisins and moniliformin. Conditions favourable for in vitro growth are also generally the most favourable for mycotoxin production on cereal grains. These fungi rarely exist in isolation, but occur as a complex with each other and with other Fusaria and other fungal genera. Climatic conditions will influence competition between, and the predominance of, different fungi within this complex.