分蘖洋葱鳞茎粗提物对西瓜枯萎病病原菌(Fusarium oxysporum f.sp.niveum)的影响

采用分蘖洋葱鳞茎粗提物对西瓜枯萎病病原菌(Fusarium oxysporum f.sp.niveum)菌丝生长、孢子萌发、病原菌生物量及产孢量等抑制作用进行研究。结果表明,分蘖洋葱鳞茎粗提物对西瓜枯萎病病原菌的菌丝生长、孢子萌发率、病原菌生物量及产孢量均有一定的抑制作用。当分蘖洋葱鳞茎粗提物浓度为1 000 mg.mL-1时,对西瓜枯萎病病原菌的抑制作用最大,抑菌率达到69.21%。探索了分蘖洋葱鳞茎粗提物防治西瓜枯萎病的可行性,为今后指导西瓜无公害生产提供理论依据。

Biochemical and Physiological Changes of Three Watermelon Cultivars Infested with Fusarium oxysporum f.sp.niveum

The dynamic changes in membrane permeability, activities of disease-related enzymes, and contents of pathogenesis-relevant chemical compounds and photosynthetic pigments in root cells of three watermelon cultivars were studied after inoculation with Fusarium oxysporum f. sp. niveum at seedling stage. The results showed that the capacity of self-regulating and returning to normal status of cultivar Kelunsheng (resistant) was greater than that of a susceptible cultivar, Zaohua, in terms of malonaldehyde (MDA) content, relative conductivity, and activity of superoxide dismutase (SOD) and catalase (CAT). The resistant cultivar maintained a higher activity of dehydrogenase, higher content of vitamin C(Vc), and relatively lower content of soluble sugar than the susceptible cultivar. The content of soluble protein was higher in the resistant cultivar than that in the susceptible one at day 1 after inoculation. The capacities of the resistant cultivar to inhibit chlorophyll deterioration and maintain a higher carotenoid content were significantly stronger than those of the susceptible cultivar.

棉花枯萎病菌(Fusarium oxysporum f.sp.Vasinfectum(ATK)Sndyder & Hansln的抑菌土初步研究

1987～1988年在河南新乡用相同接菌量种植感病品种,对棉花抗枯萎病品种86—1连作10年以上田块土样进行盆栽和田间小区试验。结果表明.枯萎病平均病指分别为29.4和4.4,与对照果园土比较抑菌效果分别为39.9％和85.8％。上述田块中86—1棉株根围5cm内土样,盆栽枯萎病平均病指19.5,抑菌效果60.5％。连作10年以上枯萎病圃土样,病指分别为27.2和6.8,抑菌效果为43.9％和76.2％。1990～1991年在辽宁省经作所试验结果,连作10年以上枯萎病圃田间抑菌效果56.2％,连作5年病圃抑菌效果17.6％。上述结果证明,枯萎病圃衰退的主要原因是棉花枯萎病菌抑菌土的产生和存在。土样内枯萎菌菌量测定结果表明,3个抑菌土土样内菌量比果园导菌土减少48.6％～59.2％。抑菌土经高压灭菌后抑菌效果完全消失,初步证明抑菌因子可能以生物因子为主。

应用巢式PCR检测黄瓜尖镰孢菌(Fusarium oxysporum f．sp．cucumbrum)

文章应用BIK2/BIK3和BIK1/BIK4两对引物,采用巢式PCR(Nested-PCR)技术对镰孢菌属26个菌株(代表了6个镰孢菌种)以及从黄瓜根际分离的20株真菌、6株细菌和7株放线菌共计59个菌株进行了扩增。结果显示,只有3个黄瓜尖镰孢菌的致病菌株能产生一条长度大约为800bp的片断。对接种黄瓜尖镰孢菌的黄瓜植株进行检测结果表明,巢式PCR能从接种5d后的黄瓜病样中特异性地检测出病原菌,而症状的出现需要10～13d时间。该项技术具有较高的灵敏度,适用于黄瓜枯萎病早期侵染检测研究。

牛蒡寡糖-钕配合物对西瓜枯萎病菌(Fusarium oxysporum f. sp. niveum)的影响

[目的]确定牛蒡寡糖－钕配合物对病原菌的最佳作用浓度。[方法] 采用平板培养和液体培养法，研究了不同浓度牛蒡寡糖－钕配合物对尖镰孢菌菌落直径、果胶酶活性、菌丝生长量和菌丝细胞膜透性的影响。[结果] 在平板培养试验中，2～180 mg/L的牛蒡寡糖－钕配合物促进菌落扩展，但菌丝稀疏，200～400 mg/L 的牛蒡寡糖－钕配合物抑制菌落扩展，菌丝稀而薄。在液体培养试验中，2～400 mg/L的牛蒡寡糖－钕配合物抑制菌丝生长，抑制率为4.3%～64.4%，随着牛蒡寡糖－钕配合物浓度的升高，其抑制作用加强；2～400 mg/L 的牛蒡寡糖－钕配合物降低了果胶酶活性；当测定时间为15、30、60、90、120和180 min 时，2～400 mg/L的牛蒡寡糖－钕配合物增加了菌丝细胞膜透性。[结论] 该研究为牛蒡寡糖-钕配合物在植物上的合理应用提供了理论依据。

从土壤中分离棉枯萎病菌(Fusarium oxysporum f.sp.vasinfetum (Atk) Synder & Hansen)选择性培养基研究

本文介绍一种用于分离检测土壤中棉枯萎病菌的选择性培养基—植选2号。其成分为:KH_2PO_41g,MgSO_4·7H_2O 0.5g,K_2S_2O_50.2g,KCl0.6g,NH_4NO_3 0.5g,蛋白胨5g,山梨糖10g,蔗糖5g,琼脂20g,蒸馏水1000ml,PCNB 620mg,Oxgall1g,硫酸链霉素300mg,盐酸金霉素75mg。根据棉枯萎病在此培养基上的形态特征,能较容易地识别和检测该病菌。

棉花枯萎病菌(Fusarium oxysporum f. sp. Vasinfectum)突变体的诱发与鉴定

通过对32个棉花枯萎病菌菌株进行的突变体诱发与鉴定实践及技术方法探索,进一步验证了可以采用KClO3培养基来诱发突变体,并利用各突变体对不同N源的利用能力差异而采用MM,MH,MA,MO2培养基对各突变体类型进行鉴定。实验结果显示绝大多数菌株(93.7%)在KClO3培养基上能诱发出突变体,但多数(85%)突变体类型不全,且不同菌株诱得突变体的难易不同,个别菌株很难诱发到突变体。各突变体的诱发率高低依次为nit1>nit3>nit8>nitM,与国内外相关报道结果基本一致。

一株Fusarium oxysporum f. sp. niveum拮抗菌的筛选、鉴定及其抑菌特性

为筛选西瓜枯萎病拮抗菌株并探究其抑菌特性,以齐齐哈尔市青昕蔬菜基地根际土为材料,采用平板对峙法筛选到1株能高效抑制西瓜专化型尖孢镰刀菌( Fusarium oxysporum f.sp.niveum,Fon)的拮抗菌株,通过形态特征、16S rDNA、 gyrA 和 gyr B序列分析鉴定其菌种;通过盆栽试验验证拮抗菌株对西瓜枯萎病的抑制作用;采用生长速率法研究拮抗菌株不同生长时期的无菌发酵滤液及其不同贮存条件下的抑菌能力;利用扫描电子显微镜(SEM)和共聚焦激光扫描显微镜(CLSM)观察并分析无菌发酵滤液对Fon孢子形态和膜完整性的影响。结果表明,拮抗菌株WD为 Bacillus amyloliquefaciens,能有效抑制西瓜枯萎病的发生,抑制率达到57.57%,其无菌发酵滤液在衰亡期有较高的抑菌能力,对Fon菌丝生长抑制率为54.32%;无菌发酵滤液对温度、pH具有较高的耐受性,30 ℃、pH 7时抑菌活性最强,抑菌率可达58.36%;在4 ℃贮存至少45 d,且紫外光照120 min内对其抑菌活性没有影响;Fon孢子暴露在WD无菌发酵滤液下,孢子表面皱缩凹陷,膜完整性被破坏。综上所述,Bacillus amyloliquefaciens WD是1株高效的拮抗菌株,为西瓜枯萎病生防菌剂的研发与利用奠定了基础。

Isolation and identification of Fusarium oxysporum f. sp. cubense in Fujian Province, China

Fusarium wilt, caused by Fusarium oxyporum f. sp. cubense(Foc), is the most serious disease affecting banana production.To clarify the distribution of the Foc races in Fujian Province of China, 79 soil samples were collected from four regions of Zhangzhou City, the primary banana production area in Fujian. We isolated and identified 12 Foc strains based on internal transcribed spacer(ITS) sequence analysis, PCR amplification by using Foc-specific primers and pathogenicity assays.Our analysis indicated that 11 isolates belong to Foc race 1, and 1 isolate belongs to the Foc tropical species race 4(TR4).Although TR4 has previously been reported to occur in primary banana-producing provinces, such as Hainan, Guangxi,and Guangdong of China, this is the first report of TR4 isolated from the soil in Fujian Province. Monitoring the presence of Foc, in particular, the TR4 strains in the soil, is the basic strategy to prevent and control Fusarium wilt.

Isolation and Identification of Shoot-Tip Associated Endophytic Bacteria from Banana cv. Grand Naine and Testing for Antagonistic Activity against <i>Fusarium oxysporum</i>f. sp. <i>cubense</i>

Endophytic bacteria colonizing the shoot-tips of banana cv. Grand Naine were isolated and tested for the antagonistic activity against the Panama wilt pathogen Fusarium oxysporum f. sp. cubense (Foc). Pre-isolation, the suckers were given extensive disinfection treatments and the homogenate from the excised shoot-tip portion was plated on nutrient agar (NA) and trypticase soy agar (TSA). This yielded altogether 47 isolates: 26 on NA and 21 on TSA, respectively, from the 10 suckers collected during August to February. The number of bacterial isolates obtained per sucker varied from one to 15 based on colony characteristics registering up to 10 distinct species per shoot-tip based on 16S rRNA sequence analysis. The 47 isolates belonged to 19 genera and 25 species under the phylogenetic classes of Actinobacteria, α- and γ-Proteobacteria and Firmicutes. Actinobacteria constituted the predominant phylum (55% isolates) with the constituent genera of Arthrobacter, Brevibacterium, Corynebacterium, Curtobacterium, Kocuria, Kytococcus, Micrococcus, Naumanella, Rothia and Tessaracoccus spp. and an unidentified isolate belonging to the family Frankiaceae. Proteobacteria constituted the second major phylum (Brevundimonas, Enterobacter, Klebsiella, Pseudomonas, Serratia and Sphingomonas spp.) followed by Firmicutes (Bacillus and Staphylococcus spp.). Antagonistic activity of the endophytes against Foc was tested through agar plate assays (pit and spot applications on fungal lawn) employing potato dextrose agar and NA. Endophytic Pseudomonas aeruginosa (isolate GNS.13.2a) which was associated with a single sucker showed significant growth inhibition effect on Foc while Klebsiella variicola (GNS.13.3a) and Enterobacter cloacae (GNS13.4a) exhibited moderate inhibition. The study brings out considerable sucker to sucker variation in the associated cultivable endophytic bacteria in “Grand Naine” banana and identifies a few bacterial endophytes with biocontrol potential against the devastating Foc pathogen.

Biological Characteristics of Fusarium oxysporum f. sp.niveum Isolated from Watermelon Cultivated in Plastic Greenhouse and Indoor Toxicity Determination of Different Fungicides in Hainan Province of China

[Objective]The paper was to study biological characteristics of Fusarium oxysporum f.sp.niveum in watermelon cultivated in plastic greenhouse in Hainan Province of China and to determine indoor toxicity of fungicides.[Method]Effects of various factors on mycelial growth and spore germination were studied,and the lethal temperature of spores and indoor toxicity of fungicides were also determined.[Result]The fungal mycelium had a better growth rate on watermelon juice medium than the others.The optimum temperature for mycelial growth and spore germination were 28℃and 30℃,and the optimum pH values were7-8 and 7-9,respectively.Light-dark alternation was conducive for mycelial growth.The lethal temperature of spore was 60℃for 5 min.The fungal mycelium grew better while taking fructose and glucose as carbon sources and yeast extract and peptone as nitrogen sources.Toxicity determination showed that 50%prochloraz-manganese chloride complex WP had the greatest inhibition effect on mycelial growth,with the EC50value of 0.730 9μg/mL;followed by 32.5%benzoic azoxystrobin SC,30%benzoylate·propiconazol EC,10%difenoconazole WG,25%bromothalonil·carbendazim WP and 25%bromothalonil WP,with the EC50values from 1.884 7μg/mL to 8.161 0μg/mL.[Conclusion]The study provided basic data for field control against F.oxysporum f.sp.niveum on watermelon cultivated in plastic greenhouse in Hainan Province.

DNA methylation patterns of banana leaves in response to Fusarium oxysporum f. sp. cubense tropical race 4

Fusarium wilt of banana, which is caused by Fusarium oxysporum f. sp. cubense tropical race 4(Foc TR4), is a serious soil-borne fungal disease. Now, the epigenetic molecular pathogenic basis is elusive. In this study, with methylation-sensitive amplification polymorphism(MSAP) technique, DNA methylation was compared between the leaves inoculated with Foc TR4 and the mock-inoculated leaves at different pathogenic stages. With 25 pairs of primers, 1 144 and 1 255 fragments were amplified from the infected and mock-inoculated leaves, respectively. DNA methylation was both changed and the average methylated CCGG sequences were 34.81 and 29.26% for the infected and the mock-inoculated leaves. And DNA hypermethylation and hypomethylation were induced by pathogen infection during all pathogenic stages. Further, 69 polymorphic fragments were sequenced and 29 of them showed sequence similarity to genes with known functions. And RT-PCR results of four genes indicated that their expression patterns were consistent with their methylation patterns. Our results suggest that DNA methylation plays important roles in pathogenic response to Foc TR4 for banana.

Tolerance of New Introgressive Hybrid and Backcross Forms Pathogenic Micromitisms (<i>Verticillium dahliae</i>Kleb and Fusarium oxysporum f.sp. vasinfectum)

The article is based on the use of experimental polyploidy method, with the introduction of new introgressive hybrid forms combining several species genotype with pathogenic Verticillium dahliae Kleb. and the effects of mycotoxins separated from the Fusarium oxysporum f.sp. vasinfectum micromicette on the yield of plant seeds. New artificial complex hypertension forms based on experimental polyploidy Verticillium dahliae Kleb. and Fusarium oxysporum f.sp. vasinfectum combine the potential of resistance to mycotoxins separated from microcrystals, making a tremendous contribution to the selection of new varieties and to the effectiveness of selection as a result of the use of genetic selective research as genetic-selective genetic-selector studies.

Construction of PEG-mediated Genetic Transformation and Gene Knockout System in Fusarium oxysporum f. sp.cubense Tropic Race 4

Fusarium wilt of banana, caused by Fusarium oxysporum f. sp. cubense tropic race 4(Foc TR4), is a typical vascular and soil-borne disease which has significantly threatened the sustainable development of banana industry. In order to reveal the infection process and pathogenesis of Foc TR4, the young mycelia(66.7 mg/ml) of wild-type strain of Foc TR4(WT-Foc TR4) cultured for 18-20 h were lysed with enzyme mixture for protoplast formation, which consisted of 25 mg/ml driselase, 0.4 mg/ml chitinase, 15 mg/ml lysing enzyme and 1.2 mol/L potassium chloride. The resulted protoplasts of 2×10~7 cells/ml were used to test the efficiency of transformation mediated by polyethylene glycol, and up to 9 transformants per microgram of DNA were obtained. AmCyan, RFP and YFP genes were stably transferred into the WT-Foc TR4, separately, using the protoplast transformation system. The gene FoOCH1 encoding α-1, 6-mannosyltransferase in the WT-Foc TR4 was knocked out using the split-marker recombination technology. The genetic transformation and gene knockout system in this pathogen lays a foundation for the study of functional genomics and plant-pathogen interactions.

Characterization of Fusarium Oxysporum Isolates Obtained from Wax Gourd and Chieh-qua in China by Pathogenicity, RAMs and Sequence Analysis of the rDNA Internal Transcribed Spacers (ITS1 and ITS2)

Wax gourd (Benincasa hispida Thumb. Cogn) is called white gourd, winter melon, Chinese preserving melon, Chinese squash, and don kwa. It has been cultivated in China for over 2 300 years. It probably

Comparative study of <i>Fusarium oxysporum</i>f sp. <i>lycopersici</i>and <i>Meloidogyne incognita</i>race-2 on plant growth parameters of tomato

Many species of soil-inhabiting fungus Fusarium, cause severe yield loss in many crops. Experiments were conducted in net house condition with complete randomized block design to determine the individual effect of different in-oculum levels of root-knot nematode, Meloidogyne incognita, Race-2 and Fusarium oxysporum f sp. lycopersici on plant growth parameters viz., Plant length, fresh and dry weight and number of fruits of tomato var. P21. The experimental results showed that both the pathogens cause significant reduction in plant growth parameters. However, the fungus was not much effective on plant growth parameters in comparison to root-knot nematode. Greatest reduction in plant growth parameters was recorded in plants inoculated with 8000 J2/Kg soil of Meloidogyne in-cognita race 2. The threshold level of root-knot nematode was 1000 J2/kg soil while threshold level of Fusarium was @ 1 g/Kg soil. Inoculum level of Fusarium oxysporum f sp. lycopersici and Meloidogyne in-cognita race-2 was pathogenic and caused significant reduction at and above 1 g/kg soil and 1000 J2/Kg soil respectively.

Genetic analysis of the maximum germination distance of Striga under Fusarium oxysporum f.sp.strigae biocontrol in sorghum

Maximum germination distance(MGD) is an important component of Striga resistance in sorghum. The objective of this study was to determine gene action influencing MGD of Striga hermonthica and Striga asiatica among selected sorghum lines treated with a biocontrol agent, Fusarium oxysporum f. sp. strigae(FOS) for effective breeding with Striga resistance, and FOS compatibility. Twelve sorghum genotypes were selected based on their Striga resistance, FOS compatibility, and superior agronomic performance. Selected genotypes were crossed using a bi-parental mating design to generate six families for genetic analysis. Agar-gel assays were used to determine low haustorium initiation factor(LHF) using the 12 parental lines, their F_1 progenies, backcross derivatives, and F_2 segregants in two sets. One set had S. hermonthica seed and the other one had S. asiatica seed. Both were treated with and without FOS. Genotypes were evaluated using a split-plot design with three replications and MGD data were recorded followed by generation mean analysis. FOS reduced MGD by 1cm under both S. hermonthica and S. asiatica infestations. Additive, dominance, and epistatic gene actions were involved in the control of MGD of the two Striga species in the evaluated populations. On average, the relative contribution of additive, additive×additive and dominance×dominance genetic effects on the MGD of S. hermonthica and S. asiatica, with FOS, were 20, 33, and 36%; and 21, 32, and 35%, respectively. Breeding methods exploiting these genetic effects may provide enhanced response to selection for Striga resistance and FOS compatibility in integrated Striga management(ISM) programmes.

Study on Control Effect of Invasive Banana Fusarium Disease(Fusarium oxysporum f.sp.cubense tropical race 4)by Different Varieties and Fertilization Treatments

[Objective]The paper was to understand the field resistance performance of different banana varieties and the prevention and control ef-fect of different fertilizers,so as to provide technical reference for the prevention and control of Fusarium wilt.[Method]Field trials were set up with three treatments:shrimp peptide organic fertilizer+shrimp peptide special protection+shrimp peptide fruit Yekang(simplified as shrimp peptide organic fertilizer treatment),conventional organic fertilizer+microbial preparations(simplified as microbial treatment),and conventional or-ganic fertilizer(simplified as control).Four different banana varieties of Brazilian banana,Guijiao No.1,Nantianhuang,and Yunjiao No.1 were se-lected for the field trial.The disease incidence of Fusarium wilt and the control effects of three fertilizers were investigated during four time periods.[Result]The disease incidence of four varieties in three treatments varied.The disease incidence of Nantianhuang and Yunjiao No.1 were signifi-cantly lower than that of other two varieties.There was also significant difference in disease incidence of three treatments.The disease incidence from high to low was control>shrimp peptide organic fertilizer treatment>microbial treatment.The average monthly TR4 pathogen content in heavily infected banana plantation was more than 2000 copies,while the highest one reached 15148.9 copies.[Conclusion]Microbial agents reduced the disease incidence of Fusarium wilt to some content.Nantianhuang and Yunjiao No.1 showed the highest disease resistance compared with other varieties.However,their resistance needs to be further improved before practical application.

Efficacy of Some Fungal Seed Dressers in Controlling Fusarium oxysporum f.sp.Cicer on Chickpea under Artificial Infection Conditions

Seeds of chickpea cv.Ghab3 were treated with three fungicides;Oxycure,Topsin and Vitaflo,and infected by Fusarium Oxysporum f.sp.Ciceri at culturing under a glasshouse condition.The results showed that Oxycure(Oxychloride)caused a significant increase in seed germination compared with other treatments,but it could not provide this protection at the post-germination phase,resulting in a significant increase in seedling mortality to 42.1%at 15 days after germination,compared with the other fungicides.In addition,Topspin(Thiophanate-methyl)was an effective protectant at germination stage,and in the seedling one too,since the percent of seedling mortality did not exceed 11.11%,whereas,Vitaflo had the least effect in protecting seeds from fungal infection,but with a significant difference with the positive control,as the percentage of non-germinated seeds was 41.66%and 62.5%,respectively.