黑腹果蝇抗真菌肽基因Drs和Drs-lC在原核中的表达及抗菌活性的初步测定

对黑腹果蝇(Drosophila m elanogaster)抗真菌肽基因Drs和它的同系物基因Drs-lC在pET-21d载体中与T7.Tag标签序列进行了融合表达,并对融合表达产物进行Xa因子切割前后的抗菌活性测定。首先通过长引物PCR获得5′端带有Xa因子识别切割序列的Drs和Drs-lC,分别克隆至pET-21d表达载体上,然后转化E.coliO rigam i(DE3),筛选得到阳性克隆。以IPTG诱导目的基因与载体上的T7.Tag标签序列融合表达,将表达产物进行初步纯化后,以Xa因子进行切割处理,然后测定处理前后样品的抗菌活性。结果显示与T7.Tag标签序列融合表达的D rs、D rs-lC样品和经Xa因子切割后的样品对供试真菌黄色镰刀菌(Fusarium culm orum)、尖孢镰孢菌(Fusarium oxs-porum)和粗糙脉孢菌(Neuropora crassa)均有明显的抑制作用。     

28种药用植物提取液对6种植物病原真菌抑菌活性的测定

以瓜果腐霉（Pythium aphanidermatum）、辣椒疫霉（Phytophthora capsici）、尖镰孢菌（Fusarium oxysporium）、茄镰孢菌（Fusarium solani）、灰葡萄孢（Botrytis cinerea）、禾谷镰孢菌（Fusarium graminearum）等6种植物病原真菌为供试菌株,采用菌丝生长法对28种药用植物提取液的抑菌活性进行了测定。结果表明：丁香（Eugenia caryophyllata）对6种植物病原真菌的抑制率均达100%,乌梅（Armeniaca mume）、百部（Stemona sessilifolia）和木香（Aucklandia lappa）对5种植物病原真菌的抑制率均在70%以上。采用孢子萌发法对抑菌效果较好的5种药用植物进行进一步研究,结果表明：这5种药用植物对灰葡萄孢孢子萌发抑制率均在90%以上。     

A new antifungal coumarin from Clausena excavata

A new γ-lactone coumarin, named as excavarin-A, showing antifungal activity was isolated from the leaves of Clausena excavata by bioassay guided fractionation method. The structure was elucidated by spectroscopic data analysis and identified as 7((2E)-4(4,5-dihydro-3-methylene-2-oxo-5-furanyl)-3-methylbut-2-enyloxy) coumarin. Minimum inhibitory concentration (MIC) was determined against fifteen fungal strains pathogenic against plants and human. The least MIC was recorded against the human pathogen, Candida tropicalis and the plant pathogens Rhizoctonia solani and Sclerotinia sclerotiorum. Antifungal activities against the human pathogens, Aspergillus fumigatus and Mucor circinelloides and plant pathogens, Colletotrichum gloeosporioides, Lasiodiplodia theobromae, Fusarium oxysporum and Rhizopus stolonifer were stronger than that of the standard antimicrobials.     

A zoospore inhibition technique to evaluate the activity of antifungal compounds against Batrachochytrium dendrobatidis and unsuccessful treatment of experimentally infected green tree frogs (Litoria caerulea) by fluconazole and benzalkonium chloride

Effective and safe treatments of chytridiomycosis in amphibians, caused by Batrachochytrium dendrobatidis, are needed to help prevent mortality in captive programs for threatened species, to reduce the risk of spread, and to better manage the disease in threatened populations. We describe a simple method to determine minimum inhibitory concentrations (MICs) of antifungal agents that involves adding zoospores to various drug concentrations in 96 well plates and microscopic observation after four days. We report results from testing 10 commercially available antifungal compounds: benzalkonium chloride (<0.78 μg/ml), povidone iodine (312.5 μg/ml), amphotericin B (3.125 μg/ml), fluconazole (<1.56 μg/ml), itraconazole (<1.56 μg/ml), enilconazole (<1.56 μg/ml), mercurochrome (6.25 μg/ml), sodium chloride (12.5 mg/ml), methylene blue (<1.56 μg/ml) and Virkon (3.125 μg/ml). For treatment trials of juvenile Litoria caerulea, baths of benzalkonium chloride at 1 mg/L and fluconazole at 25 mg/L were used on 18 experimentally infected frogs per treatment. Although these treatments resulted in longer survival times (mean 43.7 ± 11.3 days) than in the untreated controls (37.9 ± 9.3 days), the mortality rate was still 100%. Higher doses of fluconazole are suggested for further animal trials.     

Proportion of individuals with anti-Batrachochytrium dendrobatidis skin bacteria is associated with population persistence in the frog Rana muscosa

The emergence of the infectious disease chytridiomycosis is a major factor responsible for amphibian extinctions in pristine habitats. However, some populations coexist with Batrachochytrium dendrobatidis (Bd), the agent of chytridiomycosis, and others go extinct when Bd arrives. Variation of pathogen severity among populations may be explained by differences of antimicrobial skin peptides and anti-Bd skin bacteria. Previous work showed that a population of the frog Rana sierrae had a high proportion of individuals with at least one species of cultured anti-Bd skin bacteria and was coexisting with Bd for more than 6 years. A population of the closely related sister species Rana muscosa had a significantly lower proportion of individuals with anti-Bd bacteria, and the following year, it went extinct due to Bd. We extended previous work to include another sampling of the R. sierrae population coexisting with Bd and found that, although the anti-Bd bacterial community somewhat differed, both populations had a high proportion of individuals with antifungal bacteria. We also included a population of R. muscosa that was naïve of Bd and predicted to go extinct once Bd emerged since that was the fate of neighboring populations. However, the naïve R. muscosa population had a high proportion of antifungal bacteria, which suggested that the population might survive if Bd emerged. Two years later, the population was surviving with Bd endemic. Variation in the proportion of individuals with anti-Bd skin bacteria appears to explain why some populations persist when Bd emerges in a population.     

芽孢杆菌LP-3抗菌物质的初分离及抑菌活性测定

内生芽孢杆菌LP-3对多种植物真菌具有抑制作用,为分析其分泌的抗菌物质的抑菌活性,分别采用5种有机溶剂对抑菌物质进行萃取,比较了抗菌粗提物与两种杀菌剂对梨黑斑病菌的抑菌效果,测定了粗提物对9种植物病原菌的毒力,结果表明,甲醇萃取物对梨黑斑病菌的抑制效果最佳,抑菌率可达68.4%,对病菌菌丝及孢子的EC50分别为1.18mg.L-1和3.61mg.L-1,介于腐霉利和异菌脲之间,显示出良好的抑菌活性,而且该物质抗菌谱广,对9种植物病原菌的EC50在1.18mg.L-1～67.93mg.L-1之间。     

链霉菌S417菌株发酵液的抗真菌活性及稳定性研究

以16种植物病原真菌为供试菌,采用菌丝生长速率法测定拮抗链霉菌S417发酵液的抑菌谱;以采后香蕉炭疽病菌为指示菌,管碟法测定发酵液经不同理化园子处理后的抑菌活性.结果显示:链霉菌S417发酵液对16种植物病原真菌均具有不同程度的抑菌活性,其中对香蕉炭疽病菌、玉米大斑病菌等5种真菌的抑制率在82.53％～69.63％之间,显著高于其他供试菌株.120℃处理20 min,发酵液仍有较强抑菌活性;紫外线照射25 min,对发酵液抑菌活性无显著影响;阳光照射4h,抑菌活性丧失;发酵液对酸碱稳定,在pH值6.0时活性最强.     

1,2-肼二(二硫代甲酸甲酯)的合成及抑菌活性研究

以二硫化碳、水合肼、硫酸二甲酯等为原料,合成了1,2-肼二(二硫代甲酸甲酯)。其结构经熔点、红外光谱进行了确证。采用菌丝生长速率法研究了1,2-肼二(二硫代甲酸甲酯)的抑菌活性。结果表明:1,2-肼二(二硫代甲酸甲酯)对葡萄黑痘病菌、辣椒炭疽病菌、苹果腐烂病菌、棉花枯萎病菌和苹果轮纹病菌等5种供试病原菌都有很好的抑制作用,其EC50值均小于20 mg/L。其中对苹果腐烂病菌的抑制作用最强,其EC50值为2.2 mg/L。     

The disease resistance potential of Trichoderma asperellum T-Pa2 isolated from Phellodendron amurense rhizosphere soil

To promote the application of Trichoderma,many countries have collected Trichoderma resources.In the present study,nine isolates were isolated from a rhizos-phere soil of Phellodendron amurense and were identified as three species:Trichoderma brevicompactum (one isolate),T.asperellurn (two isolates),T.atroviride (six isolates).Dual culture experiments showed that T.asperellum T-Pa2 grew fast and produced the best inhibition rates against six tested pathogens (80.25-91.65 ％) via competition and mycoparasit-ism.Populus davidiana × P.alba var.pyramidalis Louche(PdPap poplar) was treated with T-Pa2,which increased the catalase activity,nitrate reductase activity,and content of osmosis molecules significantly (P ＜ 0.05).Meanwhile,induction by T-Pa2 increased the resistance of PdPap pop-lar against Alternaria alternata via modulating the expres-sion of salicylic acid,jasmonic acid,and auxin transduction pathway genes.The results will form the basis for the collec-tion and application of biocontrol agents in forestry.     

72种中草药提取物对香蕉、杧果果实采后病害病原菌的抑菌活性

以9种分别来自香蕉、杧果采后病害的病原菌[Colletotrichum musae(Berk.et Curt.)Arx、Fusarium semitectum Berk.et Rav.、Verticillium theobromae(Turc.)Mason & Hughes、Cladosporium cladosporioides(Fre.)De Varies、Nigrospora oryzae(Berk et Br.)Petch、Colletotrichum gloeosporioides Penz、Botryodiplodia theobromae Pat.、Alternaria alternate(Fr.)Keissl.和Dothiorella dominicana Petr.& Cif.]为供试菌种,在中草药质量浓度相当于20mg/mL干药材的条件下,测定了41科72种中草药丙酮提取物对菌丝体生长的抑制作用。结果表明,供试的所有中草药提取物均有不同程度的抑菌作用,平均抑菌率为34.80%,其中,丁香对9种病原菌的抑菌率为100%,肉桂和大茴香的抑菌率大于90%,平均抑菌率大于50%的中草药提取物有16种,同一中草药对不同病原菌及不同中草药对同一病原菌的抑菌活性均存在差异。平均抑菌率大于90%的丁香、肉桂和大茴香对9种病原菌的EC50值为0.38～13.47mg/mL。