In Vitro Efficacy Assessment of Botanical Extracts against Botrytis gladiolorum Causing Gladiolus Leaf Blight

An experiment was conducted at the Mycology laboratory, Department of Plant Pathology, Sher-e-Bangla Agricultural University, Dhaka, Bangladesh to find out the efficacy of selected botanical extracts against Botrytis gladiolorum (Bg) causing gladiolus leaf blight. Infected leaves samples were collected from gladiolus farmers’ field and brought to the laboratory for study. Ten selected botanicals were used against the colony growth of (Bg). The botanicals were Mehendi, Chrysanthemum, Basil (Tulsi), Onion, Neem, Bael, Arjuna, Garlic, Aloevera (Ghritkumary) and Turmeric. Botanical extracts were applied at the rate of 5%, 10% and 20%. The radial mycelia growth was found minimum (11.60 mm) in garlic extract treated plate at the dose of 5% at 5 DAI, which was statistically similar with turmeric extract treated (14.00 mm) plate and the inhibition of growth was 60% and 51.72%, respectively. Similar trend was found at 10 DAI and 15 DAI. At 15 DAI, garlic and turmeric extract gave the best result against Bg, which was statistically similar with onion (50.07% inhibition) and mehendi (49.93%). All botanicals showed significantly different results over control and found effective in reducing the mycelial growth at the dose of 10%. At 5 DAI, no radial mycelia growth was found in garlic treated plate, which was statistically similar with onion treated plate, means that the inhibition of growth was 100%. Similar trend was also found at 10 DAI, and 15 DAI but at 15 DAI, onion (30.20 mm) gave the statistically similar results with Garlic (30.10 mm) and the mycelia growth inhibition was (57.70%) and (57.84%), respectively. In case of 20% dose, garlic extract showed the best result at 5 DAI and the radial mycelia growth was found minimum (00.00 mm) which was statistically similar with onion extract treated (00.00 mm) plate and turmeric extract treated plate. The inhibition of growth was 100%. At 15 DAI, the inhibition of fungal growth was found (73.74%), (71.23%) and (66.90%), respectively with treated by turmeric (18.80 mm), garlic (20.60 mm) and onion (23.70 mm).

Quality assurance of postharvest grapes against Botrytis cinerea by terbinafine

Worldwide,fruit is an indispensable treasure house of nutrition for human beings,occupying a vital position of human diet.Postharvest fruit storage requires efficient antifungal agents to control Botrytis cinerea,which is a vital postharvest disease affecting fruit and leading to enormous losses.However,with the enormous abuse of existing antifungal drugs,the problem of drug-resistant fungi is imminent,making the controlling diseases caused by pathogenic fungi even more challenging.Drug repurposing is an efficient alternative method,we evaluated a well-known antifungal chemical,terbinafine,against the agricultural pathogen,B.cinerea in vitro,as a result,terbinafine showed strong anti-fungal activity.Furthermore,the in vivo antifungal activity of terbinafine was evaluated,the results showed that terbinafine could reduce the decay area on grapes.Terbinafine could disrupt the cell membrane integrity,increase cell membrane permeability,and eventual cell death of B.cinerea.In addition,terbinafine reduced decay incidence,and weight loss and maintained the soluble solids,titratable acidity,ascorbic acid,total phenolic,and malondialdehyde content during the storage period of grapes.Overall,terbinafine could be an antifungal preservative for postharvest table grapes fresh-keeping.

Identification of Resistance to Pathogenesis Related Protein GmPR1L in Tobacco Botrytis cinerea Infection

Soybean(Glycine max(Linn.)Merr.)annual leguminous crop is cultivated all over the world.The occurrence of diseases has a great impact on the yield and quality of soybean.In this study,based on the RNA-seq of soybean variety M18,a complete CDS(Coding sequence)GmPR1L of the pathogenesis-related protein 1 family was obtained,which has the ability to resist fungal diseases.The overexpression vector and interference expression vector were transferred into tobacco NC89,and the resistance of transgenic tobacco(Nicotiana tabacum L.)to Botrytis cinerea infection was identified.The results show that:Compared with the control,the activities of related defense enzymes SOD(Superoxide dismutase),POD(Peroxidase),PAL(L-phenylalanine ammonia-lyase)and PPO(Polyphenol oxidase)in the over-expressed transgenic tobacco OEA1 and OEA2 increased to different degrees,and increased significantly at different infection time points.The activities of defense enzymes in the interfering strains IEA1 and IEA2 were significantly lower than those in the control strains.The results of resistance level identification showed that the disease spot rate of OEA1 was significantly lower than that of the control line,and the disease spot rate of OEA2 was significantly lower than that of the control line.The plaque rate of the interfering expression line IEA1-IEA2 was significantly higher than that of the control line.It is preliminarily believed that the process related protein GmPR1L can improve the resistance of tobacco to B.cinerea.

花椰菜(Brassica oleracea var.botrytis)抗黑腐病差异表达cDNA片段的克隆及功能的初步研究

采用花椰菜抗黑腐病近等基因系C731（感病系）和C712（抗病系）作为材料，利用cDNA—AFLP技术，研究了花椰菜黑腐病抗性在黑腐病菌侵染和非侵染条件下基因表达的情况．获得了两个阳性克隆M2和M6．Northern杂交和点杂交进一步证明M2是组成型表达的cDNA片段，只是在病菌侵染与非侵染条件下表达丰度不同．而M6是差异表达的cDNA片段．Southern杂交表明M6 cDNA片段在花椰菜基因组中是单拷贝序列．随后的序列分析发现M6cDNA片段与拟南芥1号染色体的BACF19P19中66665～66813bp有84％同源性，该片段编码拟南芥的2A6蛋白的部分序列．推测的氨基酸序列与拟南芥的2A6蛋白部分序列有91％同源性．用H2O2作为外源分子胁迫处理的初步功能分析发现：M6cDNA片段受H2O2诱导，在诱导早期16～24h高度表达，其在叶片中的积累呈逐渐上升趋势．根据序列分析和初步的功能分析的结果推测：M6cDNA片段可能就是编码花椰菜中类2A6蛋白的部分基因片段．是参与花椰菜抗病反应信号途径的相关调控基因片段．

花椰菜(Brassica oleracea var.botrytis)黑腐病抗性基因同源序列分离及克隆的研究

通过从 NBS 保守序列设计简并引物 PCR 的方法,以花椰菜(Brassica oleracea vat.botrytis)抗、感黑腐病的近等基因系为材料,分离得到 NBS-LRR 型抗性基因同源序列,并获得1个克隆,命名为 RGA330-7.Southern 杂交表明,该片段在近等基因系中存在明显的多态性,且该片段在抗黑腐病基因位点至少存在3个以上类似 RGA330-7的同源拷贝.序列分析结果认为该克隆与 NBS-LRR 型抗性基因的部分 CDSs 有很高的同源性,说明该片段属于 NBS-LRR 型.系统进化分析该序列与甘蓝型油菜的2个抗病同源序列归为一类,很可能这3个不同来源的抗性基因同源序列同属于一种抗性基因家族.因此推测该序列与花椰菜抗黑腐病基因紧密相关,为进一步克隆花椰菜抗黑腐病基因提供了可靠的候选基因,对分子标记辅助抗性育种具有重要意义。

Baseline Sensitivity of Botrytis cinerea from Liaoning Province to Boscalid

[Objective] This study aimed to avoid and delay the generation of resis- tance in Botrytis cinerea to boscalid. [Method] The sensitivity of Botrytis cinerea to boscalid was tested by the mycelial growth rate method. The distribution of sensitivity variation of B. cinerea to boscalid in different regions was cleared, and the sen- sitivity baseline of B. cinerea from Liaoning Province to boscalid was established. [Result] The ECso values of B. cinerea strains from Liaoning Province ranged from 0.080 0 to 7.787 2μg/ml, and the highest ECho value was 97.34 times higher than the minimum. The average EC50 value （1.973μg/ml） of the 158 strains was treated as the baseline sensitivity of B. cinerea from Liaoning Province to boscalid. [Conclu- sion] It is essential to conduct a risk assessment of drug resistance in B. cinerea to boscalid.

Tomato Resistance to Botrytis cinerea Induced by Chitosan

[Objective] The aim was to explore the effect of tomato＇s resistance to Botrytis cinerea induced by exogenous Chitosan.[Method] The leaf spraying method was used to determine the induced resistance effect of Chitosan to tomato B.cinerea and the chlorophyll content,soluble protein,soluble sugar,proline influence,the activities of peroxidase（POD）,malondialdehyde（MDA）and the Proline content in leaves of tomato.[Result] The induced resistance of Chitosan reached 58.26% at 14th d after treatment;Compared with the B.cinerea stress group,the chlorophyll content in leaves of tomato seedlings of B.cinerea stress ＋ Chitosan group had increased by 34.63%,while that of soluble protein content,soluble sugar content,Pro content and POD activity was 5.30%,10.83%,16.21% and 16.88%,respectively（except for the MDA content,which was decreased by 16.54%）.[Conclusion] Exogenous Chitosan could improve photosynthetic efficiency and protect enzymes activity to improve the resistance of tomato seedlings to B.cinerea.

Inhibitory Effects of Original Nano-Cu_2O Drug and Nano-Cu_2O Suspension on Snake Melon Botrytis cinerea

The drug-containing culture medium method for the test of toxicity was adopted to compare inhibitive effects of original nano-Cu2O drug and nano-Cu2O suspension, and nano-Cu2O drug has better inhibitive effects on snake melon Botry- tis cinerea than original nano-Cu2O drug with the same mass concentration, and inhibitory effects are positively correlated with concentration. Correlation coefficients of the toxicity regression equation are 0.892 2 and 0.996 1, effective concentration EC50 of original nano-Cu2O drug and that of nano-Cu2O suspension are 3 948.9 and 167.9 mg/kg. Original nano-Cu2O drug has an inhibitive effect on snake melon Botrytis cinerea, but the inhibition of nano-Cu2O suspension is more obvious.

Effects of Chitosan and Organosilicon on Antioxidant Enzyme Activity of Tomato Seedlings Diseased with Botrytis cinerea

[Objective] The aim was to explore effects of chitosan and organosilicon on tomato resistance against Botrytis cinerea. [Method] With leaf spraying method adopted, four groups were set in the test, including control group, groups treated with Botrytis cinerea, with chitosan and Botrytis cinerea, and with chitosan, organosilicon, and Botrytis cinerea, in order to study on effects of chitosan and organosilicon on antioxidant enzyme activities of tomato. [Result] Antioxidant enzyme activities of tomato seedlings were improved in groups with chitosan, organosilicon, and Botrytis cinerea and with chitosan and Botrytis cinerea compared with the group treated with Botrytis cinerea. On the 6th d, activities of PAL, SOD, POD, PPO and CAT in groups with chitosan and Botrytis cinerea, and with chitosan, organosilicon, and Botrytis cinerea increased by 27.36%, 52.07%, 43.55%, 82.02% and 71.82%, and 18.91%, 30.22%, 57.14%, 38.09% and 53.64%, respectively. [Conclusion] Antioxidant enzyme activities of tomato seedlings could be improved by chitosan and organosilicon.

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.

Biological Characteristics of Ginseng Botrytis cinerea Pers

[ Objective ] The study systematically studied the biological characteristics of ginseng Botrytis cinerea Pers. [ Method ] The pure pathogenic fungus was isolated from ginseng B. cinerea collected in the field by tissue segregation and purification cultivation. Subsequently, using PDA medium plate culture method, the effect of various culture conditions on mycelium growth and sporulation of ginseng B. cinerea was detected. [ Result] The optimum temperature for mycelium growth and sporulation of B. cinerea was 25 ℃. The appropriate temperature for conidia germination ranged from 20 to 25 ℃. And the optimum pH value for mycelium growth and conidia germination was 6.0. The optimum carbon source was sucrose, followed by glucose and fructose. The optimum nitrogen source was peptone, fol- lowed by beef extract, yeast extract, alanine and ammonium nitrate. Among the media, the growth of mycelium cultured on PDA medium was the fastest with the production of gray mycelium and dense colonies. Lethal temperatures for sclerotia, mycelium and conidia were 60, 55 and 50 ℃, respectively. [ Conclusion] The study provided the scientific basis for the research on the incidence law of B. cinerea and its control.

HDA19调控拟南芥对真菌Botrytis cineara的抗性反应

HDACs(Histone deacetylase)家族蛋白质负责组蛋白H3K4和H4K19脱乙酰化,并参与植物生长和应激反应的信号转导过程。茉莉酮酸酯Jasmonates(JA)是一种重要的天然植物激素,不仅调节植物生长和发育而且还参与植物对多种逆境胁迫响应的信号转导和调控过程。但是,HDACs在植物中参与JA信号转导的具体机制目前还不是很清楚。该研究以HDA19(Histone deacetylase 19)为对象,探讨了HDACs在植物JA信号转导中的功能和作用。结果表明:HDA19的T-DNA插入纯合突变体在JA处理条件下没有出现明显的JA根长反应。在相同处理条件下,hda19的不同突变体株系与相同生态型背景的野生型植株(WT)花色素苷含量无显著差异,但下游JAZ1、VSP1等JA信号通路的标记基因都显著上调表达。同时,hda19相比于WT对真菌Botrytis cineara的抗性显著增强,且hda19中下游基础防御标记基因PDF1.2、Thi2.1、ERF1等的表达水平显著高于WT。基于上述研究结果,该研究认为HAD19通过JA信号通路负调控拟南芥对真菌B.cineara的防御反应。

The Herbicidal Activity of Mutant Isolates from Botrytis cinerea

Fifteen mutant isolates were obtained by ultraviolet mutation from parent isolate Botrytis cinerea BC-4. Among them three mutant isolates, BC4-1, BC4-2, and BC4-15, showed strong herbicidal activity. BC4-1 showed maximum herbicidal activity for inhibition of germination and growth of Digitaria sanguinalis L. and Arnaranthus retroflexus L. The results also showed that herbicidal activity was influenced by differing pH of PD media, with pH value of 4.0 being the optimum. The crude toxin was extracted using chloroform, petroleum ether, and ethyl acetate, respectively, and the ethyl acetate extracts showed the strongest inhibitory activity on the germination and growth of D. sanguinalis L. and A. retroflexus L. Using HPLC, one fraction with an absorption peak at 271 nm was separated from the crude toxin. This fraction could strongly inhibit the growth of D. sanguinalis L. at a concentration of 100 mg L^-1 and could completely inhibit the seed germination of D. sanguinalis L. and A. retroflexus L. at a concentration of 50 mg L^-1 .

Hot air treatment activates defense responses and induces resistance against Botrytis cinerea in strawberry fruit

The effect of hot air（HA, 45°C, 3.5 h） treatment on reducing gray mold caused by Botrytis cinerea in strawberry fruit and the possible mechanisms were investigated. The results showed that HA treatment significantly reduced lesion diameter and enhanced activities of chitinase（CHI）, β-1,3-glucanase and phenylalanine ammonia-lyase（PAL） in strawberry fruit. Total phenolic contents were also increased by HA treatment. The activities of antioxidant enzymes including superoxide dismutase（SOD）, catalase（CAT） and ascorbate peroxidase（APX） were higher in HA treated strawberry fruit than those in control. Expression of three defense related genes such as CAT, CCR-1 allele and PLA6 was greatly induced in HA treated strawberry fruit with or without inoculation by B. cinerea. In addition, the in vitro experiment showed that HA treatment inhibited spore germination and tube growth of B. cinerea. These results suggested that HA treatment directly activated disease resistance against B. cinerea in strawberry fruit without priming response and directly inhibiting growth of B. cinerea.

Effect of plant extracts on activity of some defense enzymes of apple fruit in interaction with Botrytis cinerea

The efficacy of seven plant extracts(neem,fennel,lavender,thyme,pennyroyal,salvia and asafetida) in controlling postharvest of apple(caused by Botrytis cinerea) was evaluated in vitro and in vivo.In vitro,all plant extracts treatments inhibited spore germination.Inhibitory rates of pore germination was 17.41 and 20.83% for neem extract treatment(methanolic and aqueous extracts,respectively) with significant difference compared to control(73.6 and 85.33%) for aqueous and methanol extracts.In the storage conditions,the application of aqueous extract of neem(at concentration of 25%) resulted in 89.11% reduction of disease severity compared with the untreated control.Results of enzymes activity showed the plant extracts can increase the activity of peroxidase,phenylalanine ammonia-lyase,β-1,3-glucanase and polyphenol oxidase in the presence of pathogens,in apple fruits.However,the results of this research revealed that application of neem extracts was more effective than the application of other plant extracts.According to this study,it could be concluded that plant extracts may be useful to control postharvest disease as a safe alternative option to chemical fungicides.

Effects of different LED light wavelengths on the resistance of tomato against Botrytis cinerea and the corresponding physiological mechanisms

New types of light-emitting diode （LED） sources were applied to irradiate Botrytis cinerea mycelium and tomato leaves that were inoculated with B. cinerea to assess the effect of different LED light wavelengths on the infection of tomato with B. cinerea, to determine the optimum light wavelengths to control B. cinerea, and to explore the mechanism of LED influ- ence on the development of gray mold. The results showed that purple light and blue light irradiation significantly inhibited the growth of B. cinerea mycelium, and the inhibition rates were 22.3 and 15.16%, respectively, and purple light exhibited a better inhibitory effect than blue light. The lesion development of B. cinerea on tomato leaves was significantly inhibited upon irradiation with red and purple light with inhibition rates of 32.08 and 36.74%, respectively. Irradiation with red light inhibited the oxidative burst of superoxide anion （O2-） that was caused by infection with B. cinerea, and red light regulated the H2O2 content in the tomato leaf, which increased and rapidly returned to a lower level. In addition, red light irradiation improved the activity of superoxide dismutase （SOD）, catalase （CAT） and peroxidase （POD） in tomato leaves. However, purple light irradiation did not make tomato leaves exhibit this kind of physiological response. Therefore, red light and purple light can suppress gray mold in tomatoes, but the disease suppression mechanisms of these two types of LED light are different. In general, red light suppresses gray mold primarily by regulating the tomato defense mechanism for disease, whereas the suppression of tomato gray mold by purple light can be partially explained by the photo-inhibition of B. cinerea.

Transformation of insect-resistant gene into cauliflower (Brassica oleracea L.var. botrytis)

Cowpea Trypsin Inhibitor (CpTI) gene was transferred into the cotyle dons and hypocotyls of cauliflower by Agrobacterium-mediated transformation met hod. The best selective concentration of kanamycin (kan) was 15 mg L-1. The con centration of carbencillin (carb) was 500 mg L-1. 14 transgenic cauliflower pla nts were obtained. The putative transformants were assayed by PCR and Southern b lotting analysis. The results indicated that CpTI gene was transferred into caul iflower successfully.

Plastic bronchitis associated with Botrytis cinerea infection in a child:A case report

BACKGROUND Plastic bronchitis(PB)frequently occurs in children after the surgical repair of congenital cardiac defects or in the presence of inflammatory or allergic diseases of the lung.Accurate epidemiological data of this condition are still lacking.CASE SUMMARY A 5-year-old boy,with a clear medical history,presented to our hospital with persistent cough and pneumonia with segmental atelectasis on chest computerized tomography.He showed no significant improvement after 1 wk of amoxicillin-clavulanate potassium treatment.Bronchial casts were extracted using flexible bronchoscopy.Pathological examination of the dendritic cast confirmed the diagnosis of type I PB.Botrytis cinerea was detected by next-generation sequencing of the bronchoalveolar lavage fluid.After the removal of the airway obstruction and fluconazole treatment,the patient recovered and was discharged 14 d after admission without the recurrence of cough.CONCLUSION Botrytis cinerea pneumonia should be considered in children with PB who still have prolonged cough and atelectasis after a regular course of antibiotic therapy.Flexible bronchoscopy and etiological examination should be performed in a timely manner to determine the diagnosis,clear the airway obstruction,and target etiological treatment.

Occurrence of Boscalid Resistance in Botrytis cinerea from Greenhouse Cucumbers in China

Gray mold caused by Botrytis cinerea is a major disease of greenhouse cucumbers and its control is largely dependent on the succinate dehydrogenase inhibitor boscalid.In this study,gray mold-infected cucumbers were collected from greenhouses located in 11 towns in which boscalid had been used,and 163 B.cinerea strains were isolated by single-spore method.The EC50 value effective concentration that could inhibit conidia germination by 50%compared with the control for sensitive isolates ranged from 0.045μg/mL to 50.148μg/mL in the assays.Additionally,101 isolates were resistant to boscalid,and the resistance frequency ranged from 33.33%to 86.67%in these towns.Most of the resistant isolates showed a low or moderately resistant phenotype,with the resistance frequency of moderately resistant isolates being as high as 80%.The SdhB gene harbored a single mutation corresponding to His272.The fitness evaluation showed that the hyphal growth,sporulation,germinability and pathogenicity were different among the resistant and sensitive isolates.The results suggested that dual resistance to boscalid in B.cinerea populations could result in failure to control gray mold.

An ABAGS-Like Metabolite of Botrytis cinerea Isolate BC4 and Its Inhibitory Activity to Seed Germination of Weeds

Aimed at finding out natural compounds to kill weeds, a plant pathogenic fungus, Botrytis cinerea isolate BC4, was chosen as a source. A significantly polar metabolite, which was different in chromatographic behavior from ABAGE-like metabolite, was isolated from cultural filtrate of the fungus by column chromatography on silica gel, preparative LC, TLC and HPLC. The structure of the metabolite was determined by HPLC-ESI MS, g.l.c, IR, ^1H NMR and hydrolysis. The results showed that this metabolite was quite similar with ABAGS. The bioassay showed the metabolite had inhibition to seed germination of both broadleaf weeds [Amaranthus retroflerus L. and Chenopodium album L.] and gramineae weeds [Digitaria sanguinalis L. Scop and Echinochloa crusgalii L. Beauv]. It also had inhibitory activity to the seedling growth of broadleaf weeds. The bioassay, using Amaranthus retroflexus L. as an indicator, showed that the lowest concentration of the metabolite to inhibit root＋sprout growth of A. retroflexus was 0.6 μM, and the concentration for 50% inhibition was 1.3μM.