The effector SIX8 is required for virulence of Fusarium oxysporum f.sp. cubense tropical race 4 to Cavendish banana

Plant pathogens employ effectors as molecular weapons to manipulate host immunity and facilitate colonization. Fusarium oxysporum f. sp. cubense is the agent of wilt disease in banana plantlets and four races of the pathogen have been identified based on the cultivar specificity. A total of 9 SIX genes have been detected in the genome of Foc TR4 and 6 genes detected in Foc1. Among these SIX genes, SIX2 and SIX8 are only detected in Foc TR4, not identified in Foc1. Expression profiles analysis revealed that SIX genes of Foc TR4 are highly induced after inoculation to Cavendish banana plantlets. Virulence analysis of the SIX2 and SIX8 knock-out mutants showed that SIX8 is required for the virulence of Foc TR4 while SIX2 has no obvious functions. Over expression of SIX8-FLAG proteins in the SIX8 knock-out mutant partly restored the virulence. Western blot analysis suggested that SIX8 could be secreted into the extracellular space and a signal peptide resided the N-terminal polypeptide sequence. This study provides some clues for further research on mechanism of SIX8 in regulating virulence of Foc TR4.     

Races and virulence of Fusarium oxysporum f.sp. cubense on local banana cultivars in Kenya

Virulence of 31 Kenyan isolates of Fusarium oxysporum obtained from bananas showing symptoms of Panama disease was tested against the differential banana cvs Bluggoe, Gros Michel, Dwarf Cavendish, and two other local cvs Muraru and Wang'ae. Seventeen isolates were assigned to either race 1 or race 2 of F. oxysporum f.sp. cubense (FOC). Race 4 was not apparent in this sample of 31 isolates from Kenya as none were pathogenic to cv. Cavendish, and no wilted Cavendish have been observed in field surveys in Kenya. Races could not be assigned to 12 isolates as they were virulent on more than one differential cultivar, and two were apparently not pathogenic. All isolates assigned to races 1 and 2 belonged to the VCG bridging complex 0124/5/8/20, but some other isolates belonging to this VCG complex could not be assigned to race. All five isolates assigned to VCG 01212 could not be assigned to known races. Considerable variability thus exists within FOC isolates within this region. Local cultivars of banana showed differential resistance to the pathogen. The interaction of cultivars and isolates on the level of disease was significant. Overall, cv. Wang'ae was the most susceptible to most of the isolates tested, regardless of their race, and could therefore be used as a reference cultivar in pathogenicity tests of isolates of FOC in the East African region. Of the cultivars tested that are widely grown on smallholder farms in Kenya, Muraru was the least susceptible.     

农杆菌介导的香蕉枯萎病菌4号生理小种转化体系的优化

香蕉枯萎病是世界范围内香蕉种植区最为严重的病害之一,严重威胁和影响着香蕉产业的发展.本文针对香蕉枯萎病病原菌的4号生理小种,建立了农杆菌介导的转化体系,确定了影响转化效率主要因子的优化体系是:农杆菌在IM培养基诱导前农杆菌OD_(600)为0.15、农杆菌经IM液体培养基诱导的时间为7 h、乙酰丁香酮(AS)浓度为150 μmol/L、Focr4孢子浓度为1×10~6个/mL、共培养时间为48 h、培养温度为25℃、诱导培养基pH值为5.5.在此条件下,转化效率能达到700～800个转化子/10~6个香蕉枯萎病菌孢子.PCR验证表明外源的T-DNA已经成功随机地整合到该病原菌基因组中.目前,应用该转化体系已获得2 300多个转化子,为后续克隆相关致病基因打下了良好基础.     

Soluble and wall-bound phenolics and phenolic polymers in Musa acuminata roots exposed to elicitors from Fusarium oxysporum f.sp. cubense

The accumulation of soluble and wall-bound phenolics and phenolic polymers in Musa acuminata roots exposed to cell wall-derived elicitor from the pathogen, Fusarium oxysporum, f.sp. cubense, race four, was investigated. The root tissue from the banana cultivar "Goldfinger" was found to respond strongly and rapidly towards the elicitor through the increased synthesis of phenolic compounds. Following elicitation, the conjugated and non-conjugated phenolic metabolites in the induced root tissue were extracted and quantified. Induced phenolic synthesis was rapid and reached near maximum values after 16 h. High-performance liquid chromatography revealed both compositional and quantitative differences between induced phenolics (p-coumaric, ferulic, and sinapic acids) and those constitutively present (p-coumaric- and ferulic acid). In addition, vanillic acid was found in the ester-bound fraction and protocatechuic acid in the cell-wall bound fraction of elicited tissue. The deposition and accumulation kinetics of polymerized phenolic monomers as lignin and lignin-like polymers was quantified over a time period of 0-36 h and found to reach maximum values after 24 h. Ionization difference UV spectra of lignin indicated compositional differences in the newly synthesized lignin fraction and correlated with increased concentrations of ferulic acid and sinapic acids esters. The results show that the increased flux through the phenylpropanoid pathway resulted in the synthesis of cinnamic acid and benzoic acid derivatives that were esterified and incorporated into the cell wall fraction as part of the anti-microbial defenses activated in the root tissue.     

pH值对香蕉枯萎病菌4号生理小种生长的影响

【背景】香蕉枯萎病菌4号生理小种(镰刀菌)是香蕉产业的致命威胁。已有研究表明土壤pH值越高,香蕉枯萎病发病率越低,但是现有pH值对镰刀菌影响的研究大都是用强酸强碱调节pH值,pH值没有缓冲体系保护,而且尚未检测试验终点时介质的pH值。此外,关于pH值对香蕉枯萎病菌4号生理小种(Foc4)影响的研究尚不系统,难以用于指导生产实践。【目的】为系统地了解土壤酸碱度对Foc4生长的影响。【方法】在pH 3.0-11.0之间设定9个pH值梯度,模拟酸性到碱性土壤pH值条件,于室内培养条件下系统研究pH值对Foc4生长、产孢、孢子萌发的影响及其生长过程对环境pH值的影响。【结果】弱酸性至中性环境(pH 5.0-7.0)最适宜于香蕉枯萎病菌的生长、产孢和孢子萌发。弱碱性处理(pH8.0和pH9.0)孢子平均萌发率较弱酸性环境处理(pH5.0和pH6.0)下降了73.1%。与pH 6.0酸性处理相比,pH 8.0和pH 9.0处理的产孢量分别下降了52.3%和68.1%。【结论】香蕉枯萎病菌Foc4生长和萌发过程会产酸,但是在缓冲体系液体培养基中,除了pH 9.0和pH10.0处理终点培养液pH值分别下降了0.34和0.27个单位外,其它处理起始和终点的pH值无差异。说明在缓冲体系液体培养基中的研究结果可以反映环境pH值对Foc4生长和萌发的影响。在作物可以生长的pH值范围内(pH5.0-9.0),碱性和微碱性条件(pH8.0-9.0)能明显抑制Foc4生长、产孢和孢子萌发。     

Genetics of resistance to Fusarium oxysporum f.sp. cucumerinum races 1 and 2 in cucumber line Wisconsin-2757

The mode of inheritance of resistance to Fusarium oxysporum f.sp. cucumerinum races 1 and 2 in Wisconsin-2757 (WI-2757), a gynoecious cucumber (Cucumis sativus L.), was determined by analysing segregation of F₁, F₂ and BC₁ populations of crosses with susceptible cultivar Straight-8. Resistance to either race 1 or race 2 in WI-2757 was conferred by a single dominant gene. In allelism tests, resistance to either race in WI-2757 was determined by the gene Fcu-1, which also confers resistance in line SMR-18.     

Wild Narcissus species as a source of resistance to Fusarium oxysporum f.sp. narcissi

Thirty three species and varieties of Narcissus plus seventeen cultivars with recent species parentage were screened for resistance to Fusarium oxysporum f.sp. narcissi. Accessions were either wholly resistant (bulbs remaining uninfected), partially resistant (a variable portion of the bulbs infected) or susceptible (all of the bulbs infected). Although a pattern of resistance was found among the species tested the genetic basis of the resistance is not known.     

RFLP mapping of I1, a new locus in tomato conferring resistance against Fusarium oxysporum f. sp. lycopersici race 1

Summary The inheritance and linkage relationships of a gene for resistance to Fusarium oxysporum f. sp. lycopersici race 1 were analyzed. An interspecific hybrid between a resistant Lycopersicon pennellii and a susceptible L. esculentum was backcrossed to L. esculentum. The genotype of each backcross-1 (BC₁) plant with respect to its Fusarium response was determined by means of backcross-2 progeny tests. Resistance was controlled by a single dominant gene, I1, which was not allelic to I, the traditional gene for resistance against the same fungal pathogen that was derived from L. pimpinellifolium. Linkage analysis of 154 molecular markers that segregated in the BC₁ population placed I1 between the RFLP markers TG20 and TG128 on chromosome 7. The flanking markers were used to verify the assignment of the I1 genotype in the segregating population. The results are discussed with reference to the possibility of cloning Fusarium resistance genes in tomato.     

Gerbera jamesonii, Osteospermum sp. and Argyranthemum frutescens: New Hosts of Fusarium oxysporum f. sp. chrysanthemi

The pathogenicity of five isolates of Fusarium oxysporum obtained from infected gerbera (Gerbera jamesonii), chrysanthemum (Chrysanthemum morifolium), Paris daisy (Argyranthemum frutescens) and African daisy (Osteospermum sp.) plants was tested on some varieties of the following Compositae hosts: C. morifolium, G. jamesonii, Argyranthemum frutescens (Paris daisy) and Osteospermum sp. and compared with the host range and pathogenicity of an isolate of F. oxysporum f. sp. chrysanthemi obtained from the ATCC collection. The results indicated that isolates of F. oxysporum from G. jamesonii as well as those from A. frutescens and Osteospermum sp. belong to the forma specialischrysanthemi. The isolate from gerbera was virulent on all tested varieties of gerbera, C. morifolium, A. frutescens and Osteospermumsp. Similar results were obtained testing the isolates obtained from A. frutescens and Osteospermumsp. The strain from C. morifolium infected cultivar of gerbera, A. frutescens and Osteospermum sp. The pathogenicity of isolate of F. oxysporum f. sp. chrysanthemi obtained from the ATCC showed a different cultivar range particularly in the case of chrysanthemum and gerbera.     

基于多基因序列分析对尖孢镰孢菌古巴专化型（香蕉枯萎病菌）生理小种的鉴定

由尖孢镰孢菌古巴专化型Fusarium oxysporum f. sp. cubense, Foc引起的香蕉枯萎病是香蕉生产上的毁灭性病害,自1996年以来已对我国华南地区香蕉生产造成了严重危害。传统上香蕉枯萎病菌生理小种的鉴定主要采用人工接种鉴别寄主尔后测定病菌致病性的方法,但实验周期长,且受季节影响。以来自澳大利亚的香蕉枯萎病菌生理小种1号（BW1）、2号（Race 2）、3号（Race 3）以及亚热带4号（BW4）为对照,对分离自我国华南地区主要香蕉产区（广东、广西、海南、福建等省区）的14株香蕉枯萎病菌的单孢菌株进行致病性测定,并结合热带4号小种（TR4）和亚热带4号小种（ST4）的分子特异检测方法,确定其生理小种类型;同时,利用ITS、TEF-1α、IGS、histone H3、β-tubulin等 5个主要用于镰孢菌系统发育学研究的基因,研究不同地区不同来源的Foc菌株之间的亲缘关系及其与非病原尖孢镰孢菌的关系,并评价这5个基因在香蕉枯萎病菌生理小种鉴定上的应用价值。研究结果表明：（1）来源于我国华南地区的4号小种主要为热带4号小种;（2）TEF-1α、IGS、histone H3等3个基因片段能够将Foc中不同生理小种的菌株划分成不同的系统发育谱系,与致病性测定的结果具有对应关系,也能较好地反映尖孢镰孢菌种内菌株的亲缘关系,可用于香蕉枯萎病菌生理小种鉴定;（3）我国Foc 1号生理小种的遗传多样性高于4号生理小种,Foc 1号生理小种的菌系与来自香蕉果实上的非病原尖孢镰孢菌的亲缘关系比其与Foc 4号生理小种的菌系的亲缘关系更近。     

Inheritance and genetic linkage of fusarium wilt {Fusarium oxysporum f.sp. cucumerinum race 1) and scab {Cladosporium cucumerinum) resistance genes in cucumber (Cucumis sativus)

The inheritance of resistance of the cucumber cv. SMR 18 to the race 1 of Fusarium oxysporum f.sp. cucumerinum, the linkage relationship between resistance to race 1 of F. oxysporum f.sp. cucumerinum, resistance to Cladosporium cucumerinum and fruit spine colour, and the reactions of several cucumber cultivars to inoculations with race 1 of F. oxysporum f.sp. cucumerinum and C. cucumerinum were examined. The inbred line Straight 8 (P,), which has white fruit spines and is susceptible to both fusarium wilt and scab was crossed with the inbred line SMR 18 (P₂), which has black fruit spines and resistance to both diseases. When F, F₂, F₃, BC₁P₁ BC₁P₂ and BC₁P₁ selfed progenies were inoculated at the cotyledon stage with a suspension of spores of race 1 of F. oxysporum f.sp. cucumerinum, the ratios of resistant to susceptible plants indicated that resistance was conferred by a single dominant gene, designated Fcu-1. When 171 BC^! plants were selfed and from each resulting F₂ family different groups of 15–25 seedlings each were tested for resistance to either disease, segregation data indicated that the Fcu-1 locus and the Ccu locus for C. cucumerinum resistance were completely linked. No evidence for linkage was found between the Fcu-1 (Ccu) locus and the B locus for fruit spine colour. Among the 59 cultivars tested at the seedling stage, 15 were susceptible, while the remainder were highly resistant to inoculations with both pathogens.     

Vegetative Compatibility,Pathogenicity and Virulence Diversity of Fusarium oxysporum f. sp. melongenae Recovered from Eggplant

Fusarium oxysporum (Schlechtend.: Fr.) f. sp. melongenae (Fomg) recovered from symptomatic eggplants from five eggplant‐growing areas in Turkey, including the south, west, north‐west, north and south‐east regions. The objective of this study was to investigate the genetic diversity of the Fomg isolates from different geographical location by pathogenicity and VCG tests. Three hundred and seventy‐four Fomg isolates were classified as highly virulent, virulent, moderately virulent and low virulent through pathogenicity assays. No correlation was observed between virulence of Fomg isolates and their locations. The nitrate non‐utilizing mutants (nit) were generated as nit1, nit3 and NitM, based on phenotyping of Fomg growth characteristics of the Fomg isolates on diagnostic media with various sources of nitrogen. The majority of nit mutants (39.4%) recovered were nit1 from minimal medium (MM) containing of 2.0% potassium chlorate (MMC). The most of Fomg isolates were identified as heterokaryon self‐compatible (HSC) based on their ability to form a stable heterokaryon, while four isolates were classified as heterokaryon self‐incompatible (HSI). A large amount of Fomg isolates were vegetatively compatible and assigned as members of the same VCG, whereas nit mutants of 10 Fomg isolates that did not complement with tester strains only paired by themselves (HSC), these isolates were termed vegetative incompatible (vic). The complementation of 33 isolates with tester strains was slow and quite weak, but not paired with themselves even though they are HSC. About 96.3% of the Fomg isolates were assigned to VCG 0320, while the remaining 3.7% were classified as vegetative incompatible group.     

香蕉枯萎病菌4号生理小种致病相关基因foABC1的分离

通过对香蕉枯萎病菌4号小种致病突变体B1233的进一步研究,分离了被突变的致病相关基因foABC1,同源性分析及保守结构预测该基因编码一类ABC转运蛋白,其功能可能同稻瘟病菌的ABC转运蛋白一样,负责真菌毒素的泵出,或是像其他真菌的ABC转运蛋白,在病原菌侵染寄主植物时能忍耐植物因防卫反应所释放的植保素或抗毒素类物质。     

尖孢镰刀菌古巴专化型MAPK FoSlt2敲除突变体的转录组分析

尖孢镰刀菌古巴专化型Fusarium oxysporum f. sp. cubense（FOC）是威胁香蕉生产的重要土传病原真菌。丝裂原活化蛋白激酶（mitogen-activated protein kinase,MAPK）FoSlt2信号通路在调控尖孢镰刀菌古巴专化型的生长发育、细胞壁完整性和致病性方面发挥着重要作用。为了揭示FoSlt2信号通路的致病机理和寻找农药靶标,本研究利用高通量RNA-seq技术对该病菌野生型菌株和FoSlt2敲除突变体菌株的转录组进行了比较分析,结果表明差异表达基因共有2 164个,其中上调表达基因有1 184个,下调表达基因有980个。Gene Ontology（GO）功能分析结果表明,差异表达基因主要参与在结合、催化分子功能组和代谢过程、细胞过程生物学通路中。KEGG 功能富集分析结果表明,差异基因主要参与戊糖和葡糖醛酸盐转换、氨基糖和核苷酸糖、氨基葡聚糖降解、磷酸肌醇和碳类物质代谢通路,说明这些通路与尖孢镰刀菌古巴专化型的生长发育和致病性相关。该研究为尖孢镰刀菌古巴专化型致病机制的阐明奠定了理论基础。     

A differential medium for the identification of races 1 and 4 of Fusarium oxysporum f.sp. cubense

Three distinctive colony types were produced when Fusarium oxysporum f.sp. cubense (Foc ) races 1 and 4 were cultured on a defined basal medium containing an appropriate carbon source and bromothymol blue as a pH indicator. These distinctive cultural characteristics have been used as a specific and reliable method for the differentiation of races 1 and 4 of Foc from other species of Fusarium.     

Spatial distribution and association of banana (Musa spp.) Fusarium wilt (Fusarium oxysporum f.sp. cubense) epidemics with biophysical factors in southwestern Ethiopia

Abstract

Field surveys were conducted in 2014 and 2015 to determine the distribution and association of wilt incidence with biophysical factors. Spatio-temporal analysis of wilt epidemics in the farms revealed a wide distribution of the disease. Cropping year, district, cultivar, crop density, planting date, weeding, sucker and bud management, source of moisture and planting materials, presence/absence of wilt in neighboring farms, disease management practices, farmers awareness towards wilt diseases, planting date and cropping system were significantly associated with disease incidence in a multiple-variable model. A wilt incidence of ≤30% had high probability of association to good weed, sucker and male bud management practices, high level of farmers awareness towards wilt, low population density, disease-free suckers, rainfed production, absence of wilt in neighboring farms and good disease management schemes. Findings of this study showed Fusarium wilt is a major constraint and need for on-farm trainings to concerned bodies about wilt management schemes.     

尖镰孢古巴专化型4号小种T-DNA插入突变体Focr4-1562及其基因敲除子的生物学表型研究

尖镰孢古巴专化型4号小种Fusarium oxysporum f. sp. cubense race 4（Focr4）是引起毁灭性土传病害香蕉枯萎病的危害性最大的小种,至今其致病机理尚不清楚。对已获得的野生型菌株Focr4-193-6经基因T-DNA插入引起致病性严重减弱的突变体Focr4-1562及其插入失活基因的敲除子△Focr4-1562的生物学表型进行了研究。玻璃纸穿透试验、活体叶片接种及根部接种致病性测定的结果表明,插入突变体和敲除子不能穿透玻璃纸生长,叶片接种和根部接种未见有明显病斑和球茎维管     

Gerbera jamesonii, a New Host of Fusarium oxysporum f.sp. tracheiphilum

The random amplified polymorphic DNA (RAPD) technique was used to analyze the total genomic DNA of pathogenic isolates of Fusarium oxysporum on Gerbera jamesonii by comparing them to representatives of the formae speciales chrysanthemi and tracheiphilum. A close genetic relationship was observed among most of the new isolates from G. jamesonii. They shared RAPD markers with the tested representatives of the forma specialis chrysanthemi . Some isolates of those tested from diseased G. jamesonii were placed in a different cluster, which included representative isolates of forma specialis tracheiphilum . This is the first report of F. oxysporum f.sp. tracheiphilum on G. jamesonii. A rapid protocol for DNA extraction directly from fungal colonies grown on potato dextrose agar allowed complete analysis in less than 4 h.