Inhibition of fungal and bacterial plant pathogens by synthetic peptides: in vitro growth inhibition, interaction between peptides and inhibition of disease progression

Four synthetic cationic peptides, pep6, pep7, pep11 and pep20, were tested alone and in combinations for their antimicrobial activities against economically important plant pathogenic fungi (Phytophthora infestans and Alternaria solani) and bacteria (Erwinia carotovora subsp. carotovora and E. carotovora subsp. atroseptica). In in vitro studies, P. infestans and A. solani were inhibited by all four peptides, while E. carotovora subsp. carotovora and E. carotovora subsp. atroseptica were inhibited only by pep11 and pep20. All peptides completely inhibited P. infestans and A. solani on potato leaves and P. infestans on tubers at concentrations comparable to the in vitro IC50 (effective concentration for 50% growth inhibition) values, suggesting that these peptides are more potent in preventing infection than in inhibiting hyphal growth in vitro. Microscopic observations of P. infestans and A. solani when treated with these peptides revealed hyphal anomalies. In tuber-infectivity assays, pep11 and pep20 reduced bacterial softrot symptoms by 50% at 2.0 to 2.30 microM and by 100% at 20 microM. In assays involving two-way combinations of these peptides, growth inhibitions of fungi and bacteria by the combinations were no more than the sum of growth inhibitions by each peptide when used alone, indicating that they act additively. pep11 and pep20 are not phytotoxic to potato plants at 200 microM. With strong and broad-spectrum antimicrobial activities of pep11 and pep20 against fungi and bacteria, and with no antagonistic activities, the expression of these peptides in transgenic potato plants could lead to enhanced disease resistance against these pathogens.     

Antifungal activities of N-arylbenzenesulfonamides against phytopathogens and control efficacy on wheat leaf rust and cabbage club root diseases

A set of N-arylbenzenesulfonamides with various substituents at the arylamine and benzenesulfonyl positions were prepared, and their antifungal properties were measured in vitro against such plant pathogenic fungi as Pythium ultimum, Phytophthora capsici, Rhizoctonia solani, and Botrytis cinerea. Compounds 3, 4, 8, 9, 10, 14, 16, 18, 20, 21, 24 and 27 had antifungal activity over a broad spectrum of the phytopathogenic fungi tested, where 50% of inhibition (ED50) was in the range of 3-15 microg/ml. Based on the in vitro activity, six derivatives (3, 4, 10, 18, 21 and 27) were selected and tested further for their fungicidal efficacy in vivo. The fungicidal efficacy of 10, 21 and 27 had a disease control value of over 85% at 50 microg/ml against wheat leaf rust, while that of 4 was selective against cabbage club root disease.     

Isolation and anti-oomycete activity of nyasol from Anemarrhena asphodeloides rhizomes

The methanol extract of Anemarrhena asphodeloides rhizomes exhibited strong antifungal activity against the plant pathogenic fungi Magnaphothe grisea, Rhizoctonia solani, and the plant pathogenic oomycete Phytophthora capsici. The antifungal substance isolated from the rhizomes of A. asphodeloides was identified to be nyasol, (Z)-1,3-bis(4-hydroxyphenyl)-1,4-pentadiene by NMR and mass spectral analysis. Nyasol effectively inhibited the mycelial growth of Colletotrichum orbiculare, P. capsici, Pythium ultimum, R. solani, and Cladosporium cucumerinum in a range of 1-50 mug/ml, but did not affect the growth of bacteria and yeast. In a greenhouse test, treatment with the antifungal compound nyasol was significantly effective in suppressing the Phytophthora blight on pepper plants.     

Stereoselective synthesis and fungicidal activities of (E)-alpha-(methoxyimino)-benzeneacetate derivatives containing 1,3,4-oxadiazole ring

Fifteen novel (E)-alpha-(methoxyimino)-benzeneacetate derivatives, the analogues of strobilurins, which contain two pharmacophoric substructures of (E)-methyl methoxyiminoacetate moiety and 1,3,4-oxadiazole ring were stereoselectively synthesized. It was first found that the coupling reaction could give stereoselectively the key intermediate (E) and (Z)-methyl 2-(hydroxyimino)-2-o-tolylacetate 2 with a ratio of 14:1. The preliminary bioassays indicated that all the compounds 1 showed potent fungicidal activity against Rhizoctonia solani, Botrytis cinereapers, Gibberella zeae, Physalospora piricola and Bipolaris mayclis, and all of the tested compounds 1a-1o had more potent fungicidal activities against R. solani than Kresoxim-methyl.     

Mycelial growth inhibition of plant pathogenic fungi by extracts of South African plant species

Eleven plant species were collected from the Blyde River Canyon area, Mpumalanga, South Africa. Methanolic crude extracts of the plant samples were concentrated to dryness, dissolved in sterile distilled water and bioassayed in the laboratory for their fungicidal potential against seven plant fungal pathogens, namely Botrytis cinerea, Fusarium oxysporum, Sclerotium rolfsii, Rhizoctonia solani, Verticillium dahliae, Botryosphaeria dothidea and Pythium ultimum, at equal concentrations. A modified agar dilution method was used to determine the inhibitory effect of the plant extracts on the mycelial radial growth of the plant pathogens. Antifungal activity was confirmed in all the plants, but not in all plant parts and also not against all the test organisms. The crude extract of Eucomis autumnalis performed best of all the plant extracts tested, as it showed significant antifungal activity against all seven of the plant pathogenic test organisms and compared favourably to the inhibition of the mycelial growth by a broad spectrum synthetic fungicide (carbendazim/ difenoconazole). The crude extract of Schrebera alata came out second best, in the sense that at least one plant part inhibited the mycelial radial growth of four of the seven test organisms by at least 50%.     

几种真菌发酵液对致病疫霉的抑制作用

测定了８种真菌发酵液在５种不同浓度下对致病疫霉菌丝生长、游动孢子静止、静止胞萌发、附着胞形成和侵入丝形成等不同阶段的影响。结果表明,供试真菌不同浓度的发酵液,对致病疫霉上述各个阶段均有一定程度的抑制作用,并均随发酵液浓度增加,抑制作用逐渐增强,浓度为１００％时,抑制作用均达到最高。其中,立枯丝核菌发酵液的抑制作用最强,浓度为１００％时,对致病疫霉菌丝生长的抑制率达到９０．４％,而静止胞萌发率仅为２．４％,附着胞及侵入丝均未见形成。     

石菖蒲提取物对四种储粮害虫的生物活性研究

研究了石菖蒲4种溶剂无水乙醇、丙酮、乙酸乙酯、石油醚提取物对玉米象、谷蠹、长角扁谷盗和锯谷盗4种储粮害虫的驱避作用和触杀作用。结果表明：石菖蒲4种溶剂提取物对4种试虫均有明显的驱避作用和触杀作用,处理60 h的平均驱避等级均达到Ⅲ级以上,对4种试虫的触杀死亡率均达到41.11%以上。     

Syntheses and activities as trehalase inhibitors of N-arylglycosylamines derived from fluorinated anilines.

Twelve N-arylglycosylamines were prepared in a one-pot reaction by treatment of D-glucose, D-galactose, D-mannose or D-xylose with fluorinated anilines in the presence of small amount of hydrochloric acid. The inhibitory activity against porcine trehalase and the fungicidal activity of the title compounds toward Rhizoctonia solani have been tested.     

Sensitivity of bacterial and fungal plant pathogens to the lytic peptides,MSI-99, magainin II,and cecropin B

In vitro and leaf disk assays of bacterial and fungal plant pathogens were conducted using three cationic lytic peptides, MSI-99, magainin II (MII), and cecropin B (CB). Growth of bacterial organisms was retarded or completely inhibited by low concentrations of these lytic peptides. The peptides also significantly reduced germination of fungal spores and growth of mycelia; however, higher concentrations of peptides were needed to inhibit fungal growth compared with those needed to inhibit bacteria. The relative efficacy of the peptides depended on the microorganism tested, but CB was the most inhibitory to the majority of the bacteria and fungi assayed. MSI-99, a synthetic derivative of MII with increased positive charge, showed equal or two- to fivefold higher antibacterial activity compared to MII in the in vitro assays. MSI-99 was also superior to MII against the oomycete, Phytophthora infestans but was slightly inferior to MII in assays with the true fungi, Penicillium digitatum and Alternaria solani. In the leaf disk assays, pretreating spores of Alternaria solani and Phytophthora infestans with the peptides at concentrations as low as 10 microg per ml led to significant reductions in the size of early blight lesions and prevented development of any late blight lesions on tomato leaf disks. Our results from in vitro and leaf disk assays suggest that MSI-99 can be used as a transgene to generate tomato lines with enhanced resistance to bacterial and fungal diseases of this crop.     

Effect of a preparation from Chaetomium fungi on the growth of phytopathogenic fungi

We studied the fungicidal activity of a biological preparation from the fungi of the genus Chaetomium against soil phytopathogenic fungi Rhizoctonia solani and Fusarium oxysporum. The inhibitory effect of the preparation under study depended on its concentration, duration of storage, and growth characteristics of pure cultures of the phytopathogens. The highest (98.8%) inhibitory activity was observed on day 3 of the interaction with Rhizoctonia solani. After a 2-year storage, this preparation was capable of inhibiting the growth of the phytopathogens only at high doses. The preparation precluded the development of bare patch and increased the productivity of potato plants. The preparation may serve as an alternative to chemical fungicides for plant protection.     

Antifungal activity of herbal extracts against plant pathogenic fungi

The present study was designed to evaluate in vitro antifungal activity of herbal extracts against three plant pathogenic fungi (viz. Rhizopus stolonifer, Botrytis cinerea and Colletotrichum coccodes). Extracts of leaves and rinds of Garcinia indica, rhizomes of Curcuma aromatica, roots of Glycyrrhiza gahliae, leaves of Nyctanthes arbour-tristis and seeds of Vernonia anthelmintica were used for screening. Screening was done using poisoned food technique. Relatively potent extracts were shortlisted from this study and were further assayed to find out their minimum fungicidal concentration (MFC). From the above studies, it was observed that ethyl acetate extract of rhizomes of C. aromatica and unripe fruit rinds of G. indica have shown the lowest MFC values amongst the other tested plant extracts. This study indicates that the potential of these plant extracts in the management of diseases caused by plant pathogenic fungi.     

Development of a Sensitive Serological Method for Specific Detection of Latent Infection of Macrophomina phaseolina in Cowpea

A double-antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA) was developed for the specific detection and quantification of Macrophomina phaseolina in plant tissue. Both polyclonal antisera produced against immunogens from mycelium and culture filtrate of M. phaseolina detected the fungus in mycelial and plant extracts, although the antibodies raised against mycelium were more sensitive. No cross-reaction occurred with Rhizopus stolonifer , Pythium ultimum , Mucor hiemalis , Fusarium oxysporum , Septoria nodorum , Rhizoctonia solani , Sclerotinia sclerotiorum , Phytophthora infestans and Aspergillus niger . In enzyme assays, activity of the endo-acting hydrolytic enzymes 1,3-β-glucanase and, less, cellulase, but not xylanase was detected in infected plants. DAS-ELISA was more sensitive than the 1,3-β-glucanase assay. In polyacrylamide gel electrophoresis (PAGE) up to 18 protein bands were observed, with four bands occurring in the 12 tested isolates deriving from various geographical origin in Niger and Nigeria. The enzyme assays and protein patterns were considered not suitable for specific M. phaseolina detection. Macrophomina phaseolina was essentially located in the roots and hypocotyls, and less in epicotyls and leaves of infected plants. The antibodies were also useful to detect latent infection and the infection of cowpea seeds.     

INVESTIGATIONS ON FUNGICIDES

Twenty (aryloxythio)trichloromethanes were examined for in vitro fungicidal activity against six fungi. All compounds showed a direct fungistatic effect and some exhibited a marked fumigant action. Good protectant action against Alternaria solani on tomato and Botrytis fabae on broad bean was obtained with certain compounds, but none was better than the three standard protectants used for comparison. When supplied to plants through their roots, eight conferred significant systemic fungicidal protection against Alternaria solani in tomato, but there was no significant protection against Botrytis fabae in broad bean. In preliminary tests the 2:4:5-trichlorophenoxythio analogue gave promising results when examined as a fumigant fungicide for eradication of downy mildew in lettuce plants, for reducing lenticel rot in stored apples and for preventing blue-mould in oranges.     

瑞香狼毒根的抑菌活性研究（Ⅰ）

以苹果干腐病菌、小麦赤霉病菌、番茄早疫病菌、南瓜枯萎病菌、玉米大斑病菌、烟草赤星病菌和辣椒疫霉病菌为供试菌,采用生物活性跟踪法、生长速率测定法和系统溶剂提取法对瑞香狼毒根中的杀菌活性物质进行了筛选和分离。结果表明,杀菌活性物质主要集中在乙酸乙酯提取物和甲醇粗提物中。从乙酸乙酯提取物中分离出两种杀菌活性物质AF1-4和AF1-5,同时还发现瑞香狼毒根中还存在增菌物质。     

Synthesis and fungicidal activity of tubulin polymerisation promoters. Part 2: pyridazines

Special tetrasubstituted pyridazines are potent fungicides by promoting the tubulin polymerisation, hereby disrupting the microtubule dynamics in the fungus. They are monocyclic analogs of similar substituted triazolopyrimidines and pyridopyrazines with the same mode of action. The fungicidal activity of these pyridazines was evaluated against the plant pathogens Botrytis cinerea (grey mould), Mycosphaerella graminicola (wheat leaf blotch) and Alternaria solani (potato and tomato early blight). Structure-activity relationship studies revealed the importance of a methyl and a chlorine substituent next to both ring nitrogen atoms and two aryl or heteroaryl groups in the other two pyridazine positions.     

Antifungal efficacy of chitosan and its thiourea derivatives upon the growth of some sugar-beet pathogens

Chitosan (CS) was modified by reaction with benzoyl thiocyanate to give a thiourea derivative (TUCS). The antifungal behavior of chitosan and its thiourea derivative was investigated in vitro on the mycelial growth, sporulation and germination of conidia or sclerotia of the following sugar-beet: Beta vulgaris pathogens isolated in Egypt, Rhizoctonia solani Kühn (AG(2-2)) Sclerotium rolfsii Sacc. and Fusarium solani (Mart.) Sacc. All the prepared thiourea derivatives had a significant inhibiting effect on the different stages of development on the germination of conidia or sclerotia of all the investigated fungi in the polymer concentration range of 5-1000 microg ml(-1). In the absence of chitosan and its derivative, R. solani exhibited the fastest growth of the fungi studied. However, growth tolerance of the modified chitosan was highest for F. solani and lowest for R. solani. The most sensitive to the modified chitosan stress with regard to their germination and number produced were the sclerotia of S. rolfsii. It has been found that the TUCS is a much better fungicidal agent (about 60 times more) than the pure CS against most of the fungal strains tested. The molecular weight and the degree of deacetylation were found to have an important effect on the growth activities of the pathogens.     

Multicomponent reactions in fungicide research: the discovery of mandipropamid

Isocyanide-based multicomponent reactions of the Ugi- and Passerini-type have been valuable tools for the rapid exploration of the novel fungicidal compound classes of phenylglycinamides and mandelamides. Mandipropamid (6), which was discovered during this derivatisation, displays excellent activity against the economically important phytopathogens Phytophthora infestans (potato and tomato late blight) and Plasmopara viticola (grape downy mildew).     

捕食线虫丝孢菌的菌寄生性测定

研究了节丛孢Arthrobotrys、单顶孢Monacrosporium和隔指孢Dactylella三个捕食线虫丝孢菌属16个菌株,对水稻立枯丝核菌RhizoctoniasolaniAG1、大豆核盘菌Sclerotiniasclerotiorum、茄科镰刀菌Fusariumsolani和恶疫霉Phytophthoracactorum四种常见土壤植物病原真菌的菌寄生性。结果表明供试菌可以通过弹簧式菌丝圈缠绕、类附着胞结构吸附、简单的菌丝缠绕或者贴附寄主菌丝生长四种方式寄生病原菌。其中,绝大多数菌株对立枯丝核病菌有寄生作用,一些供试真菌对其它三种病原真菌有寄生现象。利用孢子液浸泡法测定了其中5种捕食线虫真菌对核盘菌菌核的寄生能力,显示有较高寄生率。     

Chemical composition and antifungal activity of essential oil and various extract of Silene armeria L

The aims of this study were to examine the chemical composition of the essential oil isolated from the floral parts of Silene armeria L. by hydrodistillation, and to test the efficacy of essential oil and various leaf extracts (n-hexane, chloroform, ethyl acetate and methanol) as an antifungal potential. The GC-MS analysis determined that 28 compounds, which represented 89.03% of total oil, were present in the oil containing mainly 1-butene, methylcyclopropane, 2-butene and caryophyllene oxide. The oil (1000ppm/disc) and the leaf extracts (1500ppm/disc) revealed remarkable antifungal effect against Fusarium oxysporum, Fusarium solani, Phytophthora capsici, Colletotrichum capsici, Sclerotinia sclerotiorum, Botrytis cinerea and Rhizoctonia solani, in the growth inhibition range of 39.6-67.6% and 9.3-61.3%, respectively, along with their respective MIC values ranging from 62.5 to 1000mug/ml and 125 to 2000mug/ml. The essential oil had also a strong detrimental effect on spore germination of all the tested plant pathogens along with concentration as well as time-dependent kinetic inhibition of B. cinerea. Thus, the results obtained in this study demonstrate that S. armeria essential oil and various organic extracts possess a wide range spectrum of fungicidal activity and could become an alternative to synthetic fungicides for controlling certain important plant fungal diseases.     

Volatiles from leaves and rhizomes of Fragrant Acorus spp. (Acoraceae)

Three horticultural selections of Acorus gramineus Soland were investigated to determine the chemical composition of their leaves and rhizomes. The variety 'liquorice' was found to contain methylchavicol (49%) which accounts for the unusual anisic odor of this variety, while beta-asarone was the main component of A. christophii (43%) and 'yodo-no-yuki' (20%). The results are compared with calamus oils, and the possible biosynthetic precursors of the main components methylchavicol and beta-asarone are considered.