Genetic diversity of Australian Fusarium graminearum and F. pseudograminearum

Genotypic diversity in Fusarium pseudograminearum and F. graminearum from Australia and the relationship between diversity and pathogen aggressiveness for head blight and/or crown rot of wheat were examined. Amplified fragment length polymorphism (AFLP) analysis revealed a high level of genotypic diversity within each species. Sixty-three of the 149 AFLP loci were significantly different between the two species and 70 of 72 F. pseudograminearum and 56 of 59 F. graminearum isolates had distinct haplotypes. When head blight and crown rot severity data from a recently published work on isolates representing the entire range of aggressiveness were used, only the genotypic diversity of F. pseudograminearum was significantly associated with its aggressiveness for the two diseases. Cluster analyses clearly demonstrated the polyphyletic structures that exist in both pathogen populations. The spatial diversity within F. graminearum was high within a single field, while frequent gene flow ( N _m ∼ 14) and a low fixation index ( G _st = 0·03) were recorded among F. pseudograminearum isolates from the adjacent states of New South Wales and Queensland. The differences in population structure between the heterothallic F. pseudograminearum (teleomorph G. coronicola ) and the homothallic F. graminearum (teleomorph G. zeae ) were not as pronounced as expected given their contrasting mating systems. Neither species was panmictic or strictly clonal. This points to sexual recombination in F. pseudograminearum , suggesting that ascospores of G. coronicola may also play a role in its biology and epidemiology.     

Spatial aggregation in Fusarium pseudograminearum populations from the Australian grain belt

Previous studies have evaluated the overall structure of populations of Fusarium pseudograminearum (teleomorph, Gibberella coronicola ), causal agent of cereal crown rot, but there is no information available on spatial relationships of genetic variation in field populations. Three 1-m-row sections in crown-rot-affected wheat fields in the Australian grain belt were intensively sampled to estimate population genetic parameters and the spatial aggregation, or clustering, of disease aggregates and genotypes. Estimates of population genetic parameters based on amplified fragment length polymorphisms (AFLPs) indicated that the genetic diversity in isolates from the 1-m-row populations described a significant portion of the diversity recorded for corresponding field and regional populations. In point pattern analysis, there was physical clustering and aggregation of F. pseudograminearum isolates from two of the three sites. Analysis of the spatial distribution of clonal haplotypes (DICE similarity ≥ 97%) indicated significant aggregation of clones in all three 1-m-row populations. Based on matrix comparison tests, both mating types and genetic distances had significant spatial aggregation for at least two of the three 1-m-row populations. This is consistent with the presence of non-random spatial genetic structure due to clonal aggregation. High levels of genetic diversity and spatial structuring of disease and genotypes in at least two of the three 1-m-row populations is consistent with the hypothesis that stubble is a primary inoculum source in no-tillage farming systems, resulting in aggregated patterns of disease and allowing for haplotypes to be maintained in the field over a number of annual cropping cycles.     

Mycelial compatibility reactions of Australian Fusarium graminearum and F. pseudograminearum isolates compared with AFLP groupings

Using the mycelial reactions of 435 combinations of 14 Fusarium pseudograminearum and 15 F. graminearum isolates, it was demonstrated for the first time that mycelial reactions/barrage formation cannot be clearly used to distinguish F. graminearum and F. pseudograminearum. Mutually compatible isolates produced very different patterns of compatibility with other isolates. However, about 60% of pairings between F. graminearum and F. pseudograminearum isolates were compatible, indicating common ancestry. The Mantel tests used to determine any possible associations between mycelial compatibility reactions and AFLP genotypic diversity data revealed no association between the two systems in either species. In addition, no association was found between mycelial compatibility reactions and sexual reproduction in the two species. Implications of the higher frequency of mycelial compatibility reactions observed in F. pseudograminearum than in F. graminearum are discussed.     

Comparative analysis of genetic structures and aggressiveness of Fusarium pseudograminearum populations from two surveys undertaken in 2008 and 2015 at two sites in the wheat belt of Western Australia

Fusarium crown rot (FCR), caused predominantly by Fusarium pseudograminearum (Fp) in Australia, is an important fungal disease of wheat and barley. FCR causes significant yield losses and reduced grain quality worldwide. This study investigated the population dynamics of FCR-causing F. pseudograminearum isolates from Western Australia (WA), a major wheat-growing region. Wheat samples were collected from a total of seven different sites in 2008 and 2015. Two sites, Tammin and Karlgarin, with moderate to high FCR incidence, were intensively sampled in both years. The results revealed significant increase in Fp isolation frequency between 2008 and 2015. Over 86% of 1100 Fusarium isolates were Fp in 2015 compared with 59% of 639 isolates from 2008. Mating type idiomorphs, toxin chemotypes and population genetic structures were determined for a subset of 279 Fp isolates (132 isolates from 2008 and 165 from 2015). Mating type analysis revealed differences in MAT1-1 and MAT1-2 distributions between Tammin and Karlgarin for both years. Results also showed that 97.6% of Fp isolates analysed had the 3-ADON trichothecene chemotype. Additionally, for the first time in Australia, the 15-ADON chemotype was identified in 2.3% and 2.4% of Fp isolates from 2008 and 2015, respectively. The genetic structure of Fp population determined using 21 cleaved amplified polymorphic sequence (CAPS) markers revealed a high level of genetic variation within and between populations. In addition, 2015 isolates from Tammin and Karlgarin were significantly more aggressive (P < 0.0001) than 2008 isolates. This finding may have implications in managing this significant fungal disease.     

Fusarium pseudograminearum as an emerging pathogen of crown rot of wheat in eastern China

Fusarium crown rot of wheat has been spreading in the Huanghuai wheat-growing area in China since 2010, leading to a potential yield loss. To investigate the pathogens associated with this disease in Jiangsu and Shandong provinces in recent years, 617 Fusarium isolates were isolated from nine sites in these two provinces between 2014 and 2016. Of these isolates, 372 were identified as Fusarium pseudograminearum, and the remaining isolates were identified as F. asiaticum and F. graminearum, suggesting that F. pseudograminearum is becoming a predominant causative pathogen of crown rot of wheat in eastern China. Trichothecene gene detection and chemical analyses of trichothecenes indicated that the F. pseudograminearum isolates belonged to the 3-ADON or 15-ADON chemotype, and one isolate had the NIV genotype but produced no detectable NIV. 3-ADON isolates were predominant in Jiangsu, whereas 15-ADON isolates were prevalent in Shandong. The mating type of the F. pseudograminearum isolates were identified. MAT-1 and MAT-2 existed, but in most collections, particularly those in Jiangsu, the ratios of the two mating types deviated significantly from an expected 1:1 ratio. The reason for the occurrence of F. pseudograminearum is hypothesized, and the chemotype and mating type distribution of this species in these two provinces are analysed.     

Colonization of soft wheat following infection of the stem base by Fusarium culmorum and translocation of deoxynivalenol to the head

Following inoculation of the base of soft wheat seedlings with Fusarium culmorum, disease symptoms typical of crown rot developed at the stem base and extended up to the third node by plant maturity. Fungus was isolated from all tissues exhibiting symptoms but not from symptomless tissues. Histopathological analysis revealed that the fungus was present mainly in the parenchymatic cells of the stem base and colonized the tissues via apoplastic and symplastic pathways. Host response in advance of pathogen colonization was observed. At maturity, plants were divided into sections from the inoculated area to the head. Heads were also separated into grain, rachis and chaff components. Colonization by the fungus was assessed by isolation from surface‐sterilized segments and quantified by real‐time PCR. Disease symptoms and the fungus were detected up to the third node, while deoxynivalenol (DON) was present in all stem segments and heads. Within the head, the DON concentration was higher in the rachis than in the chaff and grain components. These results demonstrate that F. culmorum can extensively colonize stem tissues but not reach the head by the time of plant maturity. In contrast, DON was detected in tissues beyond those colonized by the fungus, translocating to the head where, although accumulating mainly in the rachis, significant quantities accumulated in the grain. These findings indicate that there is a potential threat of contamination of grain with DON where severe crown rot is present in a crop.     

河北省小麦主栽品种对茎基腐病抗性鉴定及评价指标相关性分析

为明确河北省小麦主栽品种对假禾谷镰刀菌Fusarium pseudograminearum引起的小麦茎基腐病的抗性水平, 采用温室苗期与田间接种鉴定相结合的方法, 评价20份河北省小麦主栽品种的抗病性。结果显示, ‘衡0628’苗期表现为抗病, ‘山农22’等6个品种苗期表现为中抗, ‘冀麦585’等5个品种成株期表现为中抗, ‘石麦15’和‘藁优5766’ 2个品种在苗期和成株期均表现为中抗。抗性指标相关性分析结果显示, 苗期病情指数与茎基部组织中病原菌DNA含量有极强的相关性, 成株期病情指数与白穗率有显著相关性。本研究结果表明, ‘石麦15’和‘藁优5766’两个品种的抗性水平较高, 且抗性稳定, 可利用苗期茎基部组织中病原菌DNA含量及成株期白穗率分别作为评价小麦茎基腐病苗期及成株期抗性的指标之一。     

Geographic distribution and genetic diversity of Fusarium graminearum and F. asiaticum on wheat spikes throughout China

A large number of Fusarium graminearum and F. asiaticum isolates were collected from wheat spikes from all regions in China with a history of fusarium head blight (FHB) epidemics. Isolates were analysed to investigate their genetic diversity and geographic distribution. Sequence characterized amplified region (SCAR) analyses of 437 isolates resolved both species, with 21% being F. graminearum (SCAR type 1) and 79% being F. asiaticum (SCAR type 5). AFLP profiles clearly resolved two groups, A and B, that were completely congruent with both species. However, more diversity was detected by AFLP, revealing several subgroups within each group. In many cases, even for isolates from the same district, AFLP haplotypes differed markedly. Phylogenetic analyses of multilocus DNA sequence data indicated that all isolates of SCAR type 1, AFLP group A were F. graminearum , whilst isolates of SCAR type 5, AFLP group B were F. asiaticum , demonstrating that it is an efficient method for differentiating these two species. Both species seem to have different geographic distributions within China. Fusarium graminearum was mainly obtained from wheat growing in the cooler regions where the annual average temperature was 15°C or lower. In contrast, the vast majority of F. asiaticum isolates were collected from wheat growing in the warmer regions where the annual average temperature is above 15°C and where FHB epidemics occur most frequently. This is the first report of the distribution of, and genetic diversity within, F. graminearum and F. asiaticum on wheat spikes throughout China.     

Aetiology and toxigenicity of Fusarium graminearum and F. pseudograminearum causing crown rot and head blight in Australia under natural and artificial infection

Globally fusarium head blight (FHB) of wheat is predominantly caused by Fusarium graminearum (FG) and crown rot (FCR) by F. pseudograminearum (FP). While both FG and FP can cause FHB in Australia, the reasons why the morphologically and culturally similar FG is not a major FCR pathogen has remained elusive. Using aetiology and toxigenicity, this study clarifies the contrasting roles of FG and FP in FCR and FHB in Australia. Naturally infected wheat from 42 sites during 2010 FHB epidemics, and wheat inoculated with either pathogen to induce FCR or FHB at three field plantings in 2011, were used to determine pathogen prevalence and deoxynivalenol (DON) content of the crown, stem base, stem top, rachis and grain. As the primary aetiological agent, FP prevalence in the crown correlated with FCR severity while FG in grain and/or the rachis correlated with FHB severity. FG was an effective colonizer of the crown and stem base but colonization was symptomless. DON content was linked to FG biomass in all tissues except the crown, where FP biomass was the main contributor. Of the 30 measures derived to analyse pathogen fitness in 2011, 10 described the superior fitness of FG for FHB; six defined FP fitness for FHB including inoculum dispersal; and eight defined FCR fitness of both FP and FG. FG had superior FHB fitness but weak saprophytic survival may have undermined its FCR fitness.     

Comparative population structure and trichothecene mycotoxin profiling of Fusarium graminearum from corn and wheat in Ontario,central Canada

Fusarium graminearum causes fusarium head blight (FHB) of wheat and gibberella ear rot (GER) of corn in Canada and also contaminates grains with trichothecene mycotoxins. Very little is known about trichothecene diversity and population structure of the fungus from corn in Ontario, central Canada. Trichothecene genotypes of F. graminearum isolated from corn (n = 452) and wheat (n = 110) from 2010 to 2012 were identified. All the isolates were deoxynivalenol (DON) type. About 96% of corn isolates and 98% of wheat isolates were 15‐acetyl deoxynivalenol (15ADON) type. The fungal population structures from corn (n = 313) and wheat (n = 73) were compared using 10 variable number tandem repeat (VNTR) markers. The fungal populations and subpopulations categorized based on host, cultivar groups, years and geography showed high gene (H = 0.818–0.928) and genotypic (GD = 0.999–1.00) diversity. Gene flow was also high between corn and wheat population pairs (Nm = 8.212), and subpopulation pairs within corn (Nm = 7.13–23.614) or wheat (Nm = 19.483) populations. Phylogenetic analysis revealed that isolates from both hosts were F. graminearum clade 7. These findings provide baseline data on 3‐acetyl deoxynivalenol (3ADON) and 15ADON profiles of F. graminearum isolates from corn in Canada and are useful in evaluating mycotoxin contamination risks in corn and wheat grains. Understanding the fungal genetic structure will assist evaluation and development of resistant cultivars/germplasm for FHB on wheat and GER on corn.     

黄淮麦区假禾谷镰刀菌对3种杀菌剂敏感性测定

以假禾谷镰刀菌Fusarium pseudograminearum为主要病原菌引起的小麦茎基腐病已经成为黄淮麦区的主要小麦病害之一,对小麦生产安全带来严重威胁。为了解假禾谷镰刀菌对氰烯菌酯、戊唑醇和咯菌腈3种杀菌剂的敏感性,采用菌丝生长速率法对采自河南、河北、山东的108株假禾谷镰刀菌进行了室内毒力测定。试验结果表明：氰烯菌酯对假禾谷镰刀菌的EC₅₀为0.088～0.929μg/mL,EC₅₀均值为(0.471±0.181)μg/mL;敏感性分布为连续单峰曲线,经Shapiro-Wilk正态性检验符合正态分布(W=0.988,P=0.437>0.05),所以将所有菌株的EC₅₀平均值0.471μg/mL定为假禾谷镰刀菌对氰烯菌酯的敏感基线;戊唑醇对供试菌株的EC₅₀为0.015～0.961μg/mL,EC₅₀均值为(0.384±0.219)μg/mL,敏感性分布不符合连续单峰的正态分布;咯菌腈对供试菌株的EC₅₀为0.029～0.354μg/mL,E...     

Histopathological assessment of wheat seedling tissues infected by Fusarium pseudograminearum

Histopathological assessment of infection by the crown rot pathogen Fusarium pseudograminearum in wheat seedling tissues was performed using fluorescence microscopy. The coleoptiles and leaf sheaths of four host cultivars of differing susceptibility were examined. Leaf sheaths were most frequently penetrated via stomata, indicated by initial lesions forming at the guard cells. Internally, cell wall penetration was facilitated by penetration structures which appeared as hyphal swellings or septate foot‐shaped appressoria. Colonization of leaf sheaths resulted in the re‐emergence of hyphae from stomata on both surfaces of the sheath. These hyphae are hypothesized to have two major roles; first as exploratory hyphae for colonization of new tissues, and secondly as sites of profuse conidial production. The formation of conidia on the leaf sheath surface was only recorded on the most susceptible bread wheat genotype. No other major differences in host–pathogen interactions were observed among these cultivars. Almost all cell types in the leaf sheath tissues were extensively colonized, except for the vascular bundles and silica cells. This investigation provides the first comprehensive assessment of F. pseudograminearum infection structures and growth patterns during the infection of wheat seedlings.     

Fusarium graminearum and Fusarium pseudograminearum caused the 2010 head blight epidemics in Australia

Wheat crops in southeast Queensland (Qld) and northern New South Wales (NSW) were infected with fusarium head blight (FHB)‐like symptoms during the 2010–11 wheat growing season. Wheat crops in this region were surveyed at soft dough or early maturity stage to determine the distribution, severity, aetiology and toxigenicity of FHB. FHB was widespread on bread wheat and durum, and Fusarium graminearum and/or F. pseudograminearum were diagnosed from 42 of the 44 sites using species‐specific PCR primers directly on spikelets or from monoconidial cultures obtained from spikelets. Stem base browning due to crown rot (CR) was also evident in some samples from both states. The overall FHB and CR severity was higher for NSW than Qld. Deoxynivalenol (DON) concentration of immature grains was more than 1 mg kg^?1 in samples from 11 Qld and 14 NSW sites, but only 13 of 498 mature grain samples sourced from the affected areas had more than 1 mg kg^?1 DON. DON concentration in straw also exceeded 1 mg kg^?1 in eight Qld and all but one NSW sites but this was not linked to DON concentration of immature grains. The proportion of spikelets with positive diagnosis for F. graminearum and/or F. pseudograminearum and weather‐related factors influenced DON levels in immature grains. The average monthly rainfall for August–November during crop anthesis and maturation exceeded the long‐term monthly average by 10–150%. Weather played a critical role in FHB epidemics for Qld sites but this was not apparent for the NSW sites, as weather was generally favourable at all sites.     

Natural occurrence of fusarium head blight,mycotoxins and mycotoxin‐producing isolates of Fusarium in commercial fields of wheat in Hubei

Combined analyses of the natural occurrence of fusarium head blight (FHB), mycotoxins and mycotoxin‐producing isolates of Fusarium spp. in fields of wheat revealed FHB epidemics in 12 of 14 regions in Hubei in 2009. Mycotoxin contamination ranged from 0·59 to 15·28 μg g^?1 in grains. Of the causal agents associated with symptoms of FHB, 84% were Fusarium asiaticum and 9·5% were Fusarium graminearum, while the remaining 6·5% were other Fusarium species. Genetic chemotyping demonstrated that F. asiaticum comprised deoxynivalenol (DON), 3‐acetyldeoxynivalenol (3‐AcDON), 15‐acetyldeoxynivalenol (15‐AcDON) and nivalenol (NIV) producers, whereas F. graminearum only included DON and 15‐AcDON producers. Compared with the chemotype patterns in 1999, there appeared to be a modest shift towards 3‐AcDON chemotypes in field populations during the following decade. However, isolates genetically chemotyped as 3‐AcDON were present in all regions, whereas the chemical 3‐AcDON was only detected in three of the 14 regions where 3‐AcDON accounted for 15–20% of the DON and acetylated forms. NIV mycotoxins were detected in seven regions, six of which also yielded NIV chemotypes. The number of genetic 3‐AcDON producers was positively correlated with amounts of total mycotoxins (DON, NIV and acetylated forms) or DON in wheat grains. Chemical analyses of wheat grains and rice cultures inoculated with different isolates from the fields confirmed their genetic chemotypes and revealed a preferential biosynthesis of 3‐AcDON and 4‐AcNIV in rice. These findings suggest the importance of chemotyping coupled with species identification for improved prediction of mycotoxin contamination in wheat.     

抗氰烯菌酯假禾谷镰孢突变体的诱导及其生物学特性

为评估引起小麦茎基腐病的病原菌假禾谷镰孢Fusarium pseudograminearum对氰烯菌酯的抗性风险,对5株敏感菌株进行了室内药剂驯化,获得33株抗性突变体,突变频率为16.5%,其对氰烯菌酯的抗性水平范围为7.39～1 665.76倍,3株表现低抗,4株表现中抗,26株表现高抗;发现在myosin-5基因上存在11种抗性突变类型,其中217位的丝氨酸突变为亮氨酸(S217L)、420位的谷氨酸突变为赖氨酸(E420K)和135位的丙氨酸突变为苏氨酸(A135T)为主要突变类型,其比例分别为45.5%、15.2%和9.1%。S217L型抗性突变体的产孢量显著下降,菌丝生长速率和致病力与亲本菌株无显著差异。E420K型抗性突变体的菌丝生长速率和致病力显著下降,产孢量与亲本菌株无显著差异。A135T型抗性突变体的菌丝生长速率和产孢量与亲本菌株无显著差异。研究结果表明假禾谷镰孢在药剂选择压力下易形成氰烯菌酯的抗性群体,对氰烯菌酯存在中到高等的潜在抗性风险,其myosin-5的点突变与其对氰烯菌酯的抗性相关。     

Characterization and expression of Fusarium graminearum endo-polygalacturonases in vitro and during wheat infection

This study investigated the expression and characterization of two polygalacturonases (PG1 and PG2) of Fusarium graminearum during infection of wheat spikelets; after purification, these were demonstrated to be products of two unique endo - pg genes annotated in the genome database of F. graminearum . Both genes ( Fgpg1 and Fgpg2 ) were expressed in vitro and during spike infection. PG1 had a greater specific activity, with a maximum at pH 5–7, was largely secreted in liquid culture and clearly detectable in the infected ovary tissue. PG2 was more active at pH 7–7·8, was poorly secreted in liquid culture and faintly detectable in infected ovaries. Both PG-encoding genes were maximally expressed 24 h after wheat spikelet infection, paralleling the expression of a pectin lyase ( Fgpnl1 ) gene; they anticipated the expression of a xylanase gene ( FgxylA ) that was induced only 48 h after infection with a maximum at 96 h. These data strongly indicate F. graminearum -secreted PG activity at an early stage of wheat infection.     

Effect of temperature and duration of wetness during initial infection periods on disease development, fungal biomass and mycotoxin concentrations on wheat inoculated with single, or combinations of, Fusarium species

Experiments were conducted under controlled environment conditions to study the relationship between environmental conditions, development of fusarium head blight (FHB) and mycotoxin production. A single isolate from each of four Fusarium species ( F. avenaceum , F. culmorum , F. graminearum and F. poae ) was used to inoculate wheat ears separately. Combinations of two or three isolates were also used to inoculate ears simultaneously. Inoculated ears were subjected to various combinations of duration of wetness (6–48 h) and temperature (10–30°C). For all inoculations, both incidence of spikelets with FHB symptoms and concentration of mycotoxins generally increased with increasing length of wetness period and temperature. There were significant positive correlations among disease incidence, fungal biomass (quantified as total amount of fungal DNA) and mycotoxins. Mycotoxin production was also greatly enhanced by high temperatures (≥ 20°C) during initial infection periods. In single-isolate inoculations, F. poae was the least aggressive. There was no evidence to support synergetic interactions between fungal isolates in causing visual symptoms; rather the results suggest, in most cases, the presence of competitive interactions. Furthermore, the competition led to large reductions in fungal biomass compared to single-isolate inoculations, often > 90% reduction for the weaker isolate(s). In contrast, mycotoxin productivity increased dramatically in the co-inoculations, by as much as 1000 times, suggesting that competition resulted in greater production of trichothecene mycotoxins. The F. graminearum isolate was most competitive and isolates of the other three species were similar in their competitiveness.     

麦类赤霉病研究进展

由镰刀菌引起的麦类赤霉病是大麦和小麦最重要的病害之一。本文综述了近年来在麦类赤霉病致病病原菌种及分子鉴定、病原菌产毒及毒素化学型分子检测、大、小麦赤霉病病原菌种群结构、病原菌抗药性及新药剂研发、病害流行预测和品种抗病性研究等方面的进展。     

几种杀菌剂对小麦茎基腐病的防治效果

小麦茎基腐病是小麦上重要的真菌性茎基部病害,在黄淮海冬麦区呈广泛流行态势,对我国小麦生产安全构成严重威胁.本研究评价了不同拌种剂、拔节期喷施杀菌剂以及两者结合使用对小麦茎基腐病的防治效果.药剂拌种防治结果表明:60 g/L戊唑醇FS、25 g/L咯菌腈FS和25％氰烯菌酯SC拌种处理能显著降低苗期病株率,其病株防治效果...