Silencing of early auxin responsive genes MdGH3-2/12 reduces the resistance to Fusarium solani in apple

Apple replant disease(ARD)has led to severe yield and quality reduction in the apple industry.Fusarium solani(F.solani)has been identified as one of the main microbial pathogens responsible for ARD.Auxin(indole-3-acetic acid,IAA),an endogenous hormone in plants,is involved in almost all plant growth and development processes and plays a role in plant immunity against pathogens.Gretchen Hagen3(GH3)is one of the early/primary auxin response genes.The aim of this study was to evaluate the function of MdGH3-2 and MdGH3-12 in the defense response of F.solani by treating MdGH3-2/12 RNAi plants with F.solani.The results show that under F.solani infection,RNAi of MdGH3-2/12 inhibited plant biomass accumulation and exacerbated root damage.After inoculation with F.solani,MdGH3-2/12 RNAi inhibited the biosynthesis of acid-amido synthetase.This led to the inhibition of free IAA combining with amino acids,resulting in excessive free IAA accumulation.This excessive free IAA altered plant tissue structure,accelerated fungal hyphal invasion,reduced the activity of antioxidant enzymes(SOD,POD and CAT),increased the reactive oxygen species(ROS)level,and reduced total chlorophyll content and photosynthetic ability,while regulating the expression of PR-related genes including PR1,PR4,PR5 and PR8.It also changed the contents of plant hormones and amino acids,and ultimately reduced the resistance to F.solani.In conclusion,these results demonstrate that MdGH3-2 and MdGH3-12 play an important role in apple tolerance to F.solani and ARD.

Host-Induced Gene Silencing of Effector AGLIP1 Enhanced Resistance of Rice to Rhizoctonia solani AG1-IA

Rice sheath blight, caused by Rhizoctonia solani AG1-IA, is a major disease in rice-growing areas worldwide. Effectors of phytopathogenic fungi play important roles during the infection process of fungal pathogens onto their host plants. However, the molecular mechanisms by which R. solani effectors regulate rice immunity are not well understood. Through prediction, 78 candidate effector molecules were identified. Using the tobacco rattle virus-host induced gene silencing(TRV-HIGS) system, 45 RNAi constructs of effector genes were infiltrated into Nicotiana benthamiana leaves. The results revealed that eight of these constructs resulted in a significant reduction in necrosis caused by infection with the AG1-IA strain GD-118. Additionally, stable rice transformants carrying the double-stranded RNA construct for one of the effector genes, AGLIP1, were generated to further verify the function of this gene. The suppression of the AGLIP1 gene increased the resistance of both N. benthamiana and rice against GD-118, and also affected the growth rate of GD-118, indicating that AGLIP1 is a key pathogenic factor. Small RNA sequencing showed that the HIGS vectors were processed into si RNAs within the plants and then translocated to the fungi, leading to the silencing of the target genes. As a result, AGLIP1 might be an excellent candidate for HIGS, thereby enhancing crop resistance against the pathogen and contributing to the control of R. solani infection.

接触辉光放电等离子体对Fusarium solani的杀菌作用机制及动力学模型研究

为研究接触辉光放电等离子体(CGDP)对茄病镰刀菌(Fusarium solani)的杀菌作用及机制,本试验以Fusarium solani为试材,考察CGDP处理过程中电压、处理时间、抗坏血酸浓度、孢子初始浓度等因素对杀菌效果的影响;并对Fusarium solani生长、孢子形态、细胞膜完整性及过程中所产生的活性粒子进行分析。通过Linear、Weibull和Log-Logistic 3种数学模型分析不同电压下CGDP杀菌动力学特性,以相关系数(R^(2))、精确因子(Af)、偏差因子(Bf)和均方根方差(RMSE)4个参数评价拟合效果。结果表明,CGDP对Fusarium solani孢子有着明显的杀菌效果;升高电压、延长处理时间、降低抗坏血酸浓度和孢子初始浓度均能有效抑制孢子生长,使杀菌效率提高(P<0.05);CGDP处理30 min,活性物质(·OH、H_(2)O_(2)、NO_(3)^(-))浓度分别增加至1.57 mg·L^(-1)、73.89 mmol·L^(-1)和12.72 mg·L^(-1),pH值由7.07降至4.66;孢内核酸和蛋白质的渗漏量以及PI染色和扫描电镜结果表明,CGDP对Fusarium solani的杀菌作用可能是通过破坏孢子形态和细胞膜完整性后被活性物质攻击而引起的;模型拟合结果表明,Weibull模型拟合度最高(R^(2)=0.9242),并且RMSE较小(0.6367),可以用于描述和预测CGDP对Fusarium solani的杀菌动力学特征。本研究结果为低温等离子体杀菌技术和杀菌机制研究提供了理论指导。

Isolation,identification and artificial inoculation of Rhizoctonia solani on pear during storage

Pear is a fruit crop of worldwide importance and cold storage is an integral part of the production and distribution of pears.An uncharacterized fungal disease has been observed on‘Huangguan’pear fruit during cold storage in Hebei Province.The fungus was consistently isolated from diseased fruit by routine tissue separation method,and shown to be the causal agent according to Koch postulates.Based on its morphology,molecular characteristics,pathogenicity and ITS sequence,the fungus was identified as Rhizoctonia solani.This study recorded postharvest fruit rot caused by Rhizoctonia solani on pear fruit in China.

Secondary pulmonary infection by Fusarium solani and Aspergillus niger during systemic steroid treatment for COVID-19:A case report

BACKGROUND Coronavirus disease 2019(COVID-19)-associated invasive pulmonary aspergillosis presents a diagnostic challenge due to its non-specific clinical/imaging features,as well as the fact that the proposed clinically diagnostic algorithms do not necessarily apply to COVID-19 patients.In addition,Fusarium spp.is a rare cause of opportunistic life-threatening fungal infections.Disseminated Fusarium infection in an immunocompromised host is intractable,with a high likelihood of resulting mortality.To our knowledge,this is the first case of secondary pulmonary infection by Fusarium solani(F.solani)and Aspergillus niger(A.niger)during systemic steroid treatment for COVID-19.CASE SUMMARY A 62-year-old male was transported to our hospital by ambulance with a complaint of fever and dyspnea.We established a diagnosis of pneumococcal pneumonia,complicated with COVID-19 and septic shock,together with acute renal failure.He was admitted to the intensive care unit,to be treated with piperacillin/tazobactam,vancomycin,and 6.6 mg per day of dexamethasone sodium phosphate,along with noradrenaline as a vasopressor,ventilator management,and continuous hemodiafiltration.His condition improved,and we finished the vasopressor on the fifth hospital day.We administered dexamethasone for ten days,and finished the course of treatment.On the eleventh day,patient respiratory deterioration was observed,and a computed tomography scan showed an exacerbation of bilateral ground-glass-opacity-like consolidation,together with newly appeared cavitary lesions in the lung.we changed antibiotics to meropenem plus vancomycin.In addition,a fungal infection was considered as a possibility based on microscopic findings of sputum,and we began coadministration of voriconazole.However,the pneumonia worsened,and the patient died on the seventeenth day of illness.Later,F.solani and A.niger were identified from sputum collected on the twelfth day.It was believed that he developed a cell-mediated immune deficiency during COVID-19 treatment,which led to the complication of pneumonia caused by the above-mentioned fungi,contributing to his death.CONCLUSION Because early initiation of intense antifungal therapy offers the best chance for survival in pulmonary fusariosis,computed tomography scans and appropriate microbiologic investigations should be obtained for severely immunocompromised patients.

一株多粘类芽孢杆菌的鉴定及其对花椒根腐病菌Fusarium solani的抑菌作用

腐皮镰刀菌(Fusarium solani)是引起花椒根腐病的主要病原菌之一,为开发高效防治花椒根腐病的绿色生防菌,实验从健康花椒根部土壤中分离筛选出一株腐皮镰刀菌较高拮抗作用的细菌KS-16,并对其进行形态学、生理生化性质和分子鉴定,研究了该菌对三株供试菌株大肠杆菌、金黄色葡萄球菌、枯草芽孢杆菌及腐皮镰刀菌的抑菌作用.结果表明,菌株KS-16被鉴定为多粘类芽孢杆菌(Paenibacillus polymyxa).活性测定发现,该菌对腐皮镰刀菌的菌丝生长抑制率达到53.78%,对三株供试菌株也有较高的抑菌活性.以PDA培养基作为发酵培养基优于LB培养基,作用10 d其抑制效果仍保持在43.33%.

茄链格孢Alternaria solani DT-15除草活性及生物学特性初步研究

为了筛选获得高效除草生防菌株,本研究以青海省大通县野薄荷病样中分离到的DT-15菌株作为研究对象,对其进行离体致病性测定、活体除草效果评价、安全性评价、菌株分类鉴定及菌株最适碳氮源筛选研究。离体条件下DT-15菌株对3种杂草的致病性由强到弱依次为密花香薷>冬葵>藜。盆栽条件下藜、冬葵发病率高达91%以上,鲜重防效达94%以上,且对主栽作物蚕豆、小麦、豌豆、油菜表现为相对安全。菌株DT-15鉴定为茄链格孢Alternaria solani。对菌落直径、孢子悬浮液的OD值进行了测定,结果表明:小麦麸皮是最适的碳源,硝酸钠是最适的氮源。DT-15菌株具有蚕豆、小麦、豌豆、油菜田防除部分阔叶杂草的微生物除草剂开发潜力。

外源硅对纹枯病菌(Rhizoctonia solani)侵染下水稻叶片光合功能的改善

为了进一步探讨外源加硅增强水稻对纹枯病的抗性作用,以抗病品种91SP和感病品种Lemont为材料,研究了人工接种纹枯病菌条件下外源硅对水稻叶片叶绿素含量、光合作用、叶绿素荧光特性和MDA含量的影响。结果表明:(1)外源加硅能降低抗病品种91SP的纹枯病病级和病情指数,显著降低感病品种Lemont的病级和病情指数;(2)接种纹枯病菌后,水稻叶片叶绿素含量、净光合速率(Pn)、气孔导度(Gs)均明显降低,胞间CO2浓度(Ci)增大,而加硅处理的水稻叶片叶绿素含量、Pn、Gs不同程度增加,Ci有所降低;(3)接种纹枯病菌后,两个品种PSⅡ最大光化学效率(Fv/Fm)、PSⅡ有效光化学效率(Fv′/Fm′)、PSⅡ实际光化学效率(ΦPSⅡ)、光化学猝灭系数(qP)和表观光合电子传递速率(ETR)均降低,非光化学猝灭系数(qNP)增大,而对于加硅处理的水稻叶片,上述荧光参数在纹枯病菌侵染条件下的变化均受到不同程度的抑制。(4)外源硅可不同程度地减缓纹枯病菌侵染引起的丙二醛(MDA)含量的增加,对感病品种Lemont的缓解作用要大于抗病品种91SP。可见,外源硅处理可以不同程度地缓解纹枯病菌侵染条件下非气孔因素引起的水稻叶片光合速率的下降以及对光合机构的破坏作用,提高光化学效率,改善叶片的光合功能,减轻叶片膜脂过氧化程度,增强水稻对纹枯病的抗性。

镧对Rhizoctonia solani的毒力及其致病酶活性的影响

采用琼脂平板生长速率法及液体培养基培养测定了La对立枯丝核病菌(Rhizoctoniasolani)的抑制作用和毒力,并测定了其对病菌胞外的果胶酶、蛋白酶和纤维素酶等几种致病酶的活性的影响。结果表明,随着La浓度升高,对病菌菌丝生长的抑制作用增强,固体培养上所测定的对病菌的EC50和EC95分别为171.9和667.7mg·L-1;在液体培养基中所测定的EC50和EC95分别为111 4和500 7mg·L-1。在一定浓度范围内,La提高了单位量菌丝所产生3种致病酶的活性,但由于对菌丝生长量的强烈抑制,使病菌胞外3种致病酶的总量或总活性受到抑制,减低了病菌的致病力。

桉树枯萎病菌Fusarium solani分子检测技术研究

桉树枯萎病是桉树人工林中的重要枝干病害,一旦发生则对桉树人工林造成严重危害,Fusarium solani是其主要病原之一。本试验通过RAPD标记技术,找到一条致病菌特异片段,对该片段进行克隆、测序,根据测序结果设计出特异性引物F9/R9,进一步优化扩增条件,并检测其灵敏度。结果表明,特异性引物F9/R9能从桉树枯萎病菌F.solani扩增出一条大小为200 bp左右的特异条带,而其它8种参试菌株和对照均无条带产生。该引物最低可以检测到4×10-7 ng·μL^-1的病原菌基因组DNA,也能检测到单个孢子的存在。同时可从发病桉树枝干和带菌土壤中特异性检测到枯萎病菌F.solani。因此,利用特异性引物F9/R9对桉树枯萎病菌F.solani快速检测技术操作简单,特异性强,可用于该病害早期诊断及检疫,对指导林农及时进行药剂防治,进而限制病害扩展具有重要意义。

镧素对镰刀菌Fusarium solani及其致病酶的影响

采用琼脂平板生长速率法及液体培养基培养测定了 L a对镰刀菌 Fusarium solani的抑制作用和毒力 ,并测定了其对病菌胞外的果胶酶、蛋白酶和纤维素酶等几种致病酶活性的影响。结果表明 ,随着 L a2 O3 浓度升高 ,对病菌菌丝生长的抑制作用增强 ,对病菌的 EC50 和 EC95分别为 2 78.2和5 5 2 .0 mg/ L ;在一定浓度范围内 ,L a提高了单位量菌丝所产生 3种致病酶的活性 ,但由于菌丝生长受到抑制 ,除蛋白酶外 ,病菌胞外致病酶果胶酶和纤维素酶的总量或总活性受到了抑制 。

不同发酵条件对Fusarium.solani ZH0101产木聚糖酶的影响

以本实验室筛选到的一株产木聚糖酶但不产纤维素酶的真菌Fusarium solani ZH0101为供试菌株,利用麦秸为诱导物,对产木聚糖酶的液体发酵培养基进行优化。对发酵时间、麦秸添加量、培养基起始pH值、磷酸盐、金属离子、无机氮源和有机氮源对产酶的影响进行研究,优化最佳的培养条件。优化后的液体发酵培养基条件为:麦秸20g/L、KH2PO4 4g/L、CH3COONH4 1g/L、酵母膏2g/L和起始pH值为6.0,发酵时间为14d。优化条件下所产无纤维素酶活力的木聚糖酶酶活力为26.85U/mL,能比未优化前的20.82U/mL提高近30%。

Cloning and Analysis of WRKY Gene of Rice Induced by Rhizoctonia solani Kuhn

[Objective] The aim was to clone the up-regulated expression gene of rice induced by Rhizoctonia solani.[Method] The EST fragment K16 obtained by suppression subtraction hybridization（SSH）was cloned and confirmed by reverse transcription-polymerase chain reaction（RT-PCR）.Then RT-PCR products were cloned into the PMD18-T vector and sequenced.The functions of the sequence were predicted with bioinformatics method.[Result] A 1 079 bp gene was obtained.The gene encoded a protein with 236 amino acids.The protein contains many motif sites,two WRKY domains and a C2H2 zinc finger motif.The gene showed high identities with WRKY8,WRKY24 and WRKY30 gene of rice.[Conclusion] The up-regulated expression gene induced by R.solani was representative WRKY family gene.The gene could play an important role on rice sheath blight resistance.

直缘乌头内生真菌Alternaria solani化学成分研究

目的:对直缘乌头内生真菌Alternaria solani的化学成分进行研究。方法:采用各种柱色谱对直缘乌头内生真菌Alternaria solani的甲醇提取物进行分离纯化,经波谱数据(1H-NMR,13C-NMR,MS)进行结构鉴定。结果:分离得到15个化合物,分别鉴定为:20-羟基麦角甾-4,6,8(14),22-四烯-3-酮(1)、(22E,24R)-麦角甾-4,6,8(14),22-四烯-3-酮(2)、(22E,24R)-麦角甾-7,22-二烯-3β-醇(3)、24-亚甲基-麦角甾-7-烯-3β-醇(4)、过氧麦角甾醇(5)、cyclo(D)-Pro-(D)-Leu(6)、cyclo-(L-Val-LPro)(7)、cyclo-(S-Pro-S-Ile)(8)、tentoxin(9)、porrotoxin(10)、7-dehydroxyl-zinniol(11)、zinniol(12)、8-zinniol methyl ether(13)、3-甲氧基-4-甲基-5-(3’,3'-二甲基烯丙氧基)-2-甲氧基甲基苯甲酸(14)、拟盘多毛孢H2-倍半萜内酯(15)。结论:从直缘乌头内生真菌Alternaria solani中分离得到15个化合物,其中化合物2~10,12~15是首次从该植物内生真菌中分离得到。

Antifungal Effect of Streptomyces 702 Antifungal Monomer Component DZP8 on Rhizoctonia solani and Magnaporthe grisea

The aim of this study was to investigate the in vitro antifungal effects of antifungal monomer component DZP8 isolated from Streptomyces 702 on the mycelium growth, sclerotium formation and germination of Rhizoctonia solani and on the mycelium growth, conidial formation, germination, appressorium formation of Magnaporthe grisea. The results showed that the antifungal monomer component DZP8 has strong antifungal effect on both the R. solani and M. grisea. The EC50 and EC90 of DZP8 were 1.81 and 3.35 μg/ml on Ft. solani respectively, and 37.01 and 136.21 μg/ml on M. grisea respectively. Under the treatment of 48.01 μg/ml DZP8, the sclerotium formation rate of R. solani was just 39.21%, the formation time delayed by 216 h and the dry weight decreased by 81.37% in comparison the con- trol; and 33.51 μg/ml DZP8 significantly inhibited the sclerotium germination. In the presence of 160.08 μg/ml DZP8, the sporulation of M. grisea was just 9.29% of control sample; 20.14 μg/ml DZP8 inhibited the conidial germination suppression rate by 95.16%, and the appressorium formation by 100%.

Control Effect of Mixture of Endophytic Strain B196 and Jinggangmycin against Rhizoctonia solani

[Objective] The study was to provide the theoretical guidance for the control of rice sheath blight using the mixture of endophytic Bacillus megaterium strain B196 and fungicide. [Method] The toxicity of the mixture of endophytic strain B196 and jinggangmycin against Rhizoctonia solani was determined by inhibition rate method,and the effect of the mixture on R. solani was also tested. [Result] The mixture of B196 strain and 5% Jinggangmycin water agent with the mass ratio of 1∶68.47 had synergy effect,co-toxicity coefficient （CTC） was 209,the control effect of 200 μg/ml mixture against R. solani was 75.57% in field,which was 17.68% higher than that of single endogenous bacillus strain B196 treatments. [Conclusion] The mixture of endophytic strain B196 and Jinggangmycin had good control effect against R. solani.

Purification of Extracellular Protease Produced by Rhizoctonia solani and Its Partial Characterization

[Objective] The aim of the study was to provide the basis for researching the pathogenicity mechanism of Rhizoctonia solani.[Method] The extracellular protease was purified after ammonium sulfate precipitation through DEAE-Sephrase Fast Flow,Phenyl-Sepharose Fast Flow and Sephadex G-75 ch rom atography. [Result] The extracellular protease with molecular weight of 49.5 ku was obtained from fermentation liquid of R. solani. The optimal temperature and pH value for its activity were 6.4 and 30 ℃ respectively. Zn^2＋,Fe^3＋,Cu^2＋had inhibition on enzyme activity,while Mg^2＋,Mn^2＋had no effect on enzyme activity,and Ca^2＋ could activate enzymatic activity in low concentration.[Conclusion] R. solani could secrete extracellular protease,but the relationship between the extracellular protease and the pathogenicity of R. solani required further study.

Antifungal Activity of Botanical Extracts against Botrytis cinerea and Alternaria solani

[Objective] The antifungal activity of the extracts from,Atractylodes macracephal Koidz and Pulsatilla chinensis Bunge Regel,against Botrytis cinerea and Alternaria solani were studied under the condition of laboratory,in order to develop and utilize these two plants.[Method] The mycelium growth rate test was applied to measure the antifungal activities of extracts against fungi.[Result] the extracts of all the two plants showed strong antifungal activity against the target pathogenic fungi,especially the antifungal activity of the extract from Pulsatilla chinensis Bunge Regel was stronger and more stable.The inhibition rate to the mycelium growth of Botrytis cinerea was 80.25%.At the same concentration,the extract from Atractylodes macracephal Koidz showed little inhibition to Botrytis cinerea and Alternaria solani.The petrolelum ether extract of Atractylodes macracephal Koidz showed stronger antifungal activities and the EC50 was 5.31 mg/ml,and the n-butanol extract of Pulsatilla chinensis Bunge Regel showed stronger antifungal activities and the EC50 was 2.93 mg/ml.[Conclusion] The extracts from Pulsatilla chinensis Bunge Regel showed the stronger antifungal activity against Botrytis cinerea and Alternaria solani.

冬虫夏草内生菌 Fusarium solani 化学成分研究

目的研究冬虫夏草内生菌Fusarium solani化学成分。方法Fusarium solani乙酸乙酯提取物采用硅胶、Sephadex LH-20、制备HPLC进行分离纯化,根据理化性质及波谱数据鉴定所得化合物的结构。结果从中分离鉴定出25个化合物。结论化合物1~17、19~24为首次从该真菌的次生代谢产物中发现。

Rhizoctonia solani的一种选择性培养基

通过对多种农药、抗菌素、化学试剂和基础培养基的筛选测定,选出了Rhizoctoniasolani的一种选择性培养基。该培养基可以有效地运用于自然土壤中植物残体上R.solani的分离。经比较,该培养基的选择性能要优于国外已报道的同类培养基。