花生白绢病研究进展

由齐整小核菌Sclerotium rolfsii Sacc. 侵染引起的土传真菌性病害——花生白绢病,严重威胁花生的生产。为挖掘对花生白绢病有较好防治效果的拮抗细菌,利用稀释涂布法从感染S. rolfsii的植株根际土壤中分离得到570株细菌,采用平板对峙法筛选出32株S. rolfsii拮抗细菌,结合16S rRNA基因序列、形态学及生理生化特征,确定细菌的分类学地位。通过S. rolfsii与拮抗细菌的发酵液共培养和盆栽试验结果表明：在所筛选的32株拮抗细菌中,4株细菌HNZZ23、HNKF5、HNXX14和HNXX169对S. rolfsii的抑制时间长,抑制效果好,其发酵液对S. rolfsii菌丝生长的抑制率分别为85.30%、85.88%、85.16%和91.00%,对S. rolfsii菌核萌发的最大抑制率分别为100%、85.33%、82.67%和93.33%。菌株HNZZ23和HNXX14的发酵液对盆栽花生白绢病的防治效果分别为30.50%和25.73%,且菌株HNZZ23对花生植株的生长具有显著的促进作用。经分类学鉴定,菌株HNZZ23、HNKF5、HNXX14和HNXX169分别为萎缩芽孢杆菌Bacillus atrophaeuse、短小芽孢杆菌B. pumilus、铜绿假单胞菌Pseudomonas aeruginosa和枯草芽孢杆菌B. subtilis。本研究结果表明,感病花生根际细菌HNZZ23、HNKF5、HNXX14和HNXX169对花生白绢病原菌S. rolfsii具有良好的拮抗作用和生防潜力,可为花生白绢病的生物防治提供优质菌种资源。

     

白术两种主要土传病害的分离、鉴定及杀菌剂的室内活性筛选

立枯病和白绢病是白术生产种植中的两种主要土传病害,在苗期和生长期都有发生,危害严重。本研究从浙江省磐安县采集具有典型病症的白术植株,对病原进行了分离、纯化和致病性测定。综合形态学特征及rDNA-ITS序列分析表明,白术上的病害是由立枯丝核菌Rhizoctonia solani Kühn引起的立枯病和由齐整小核菌Sclerotium rolfsii Sacc.引起的白绢病。室内毒力测定结果表明,10种供试杀菌剂对立枯病菌的毒力从大到小为:噻呋酰胺咯菌腈戊唑醇四氟醚唑吡唑醚菌酯丙硫菌唑嘧菌酯啶酰菌胺苯醚甲环唑咪鲜胺;对白绢病菌的室内毒力从大到小为:吡唑醚菌酯噻呋酰胺嘧菌酯戊唑醇咯菌腈苯醚甲环唑丙硫菌唑啶酰菌胺四氟醚唑咪鲜胺,其中噻呋酰胺对两种病菌都具有很高的活性,EC_(50)分别为0.06和0.03mg/L,可用于两种病害的防治。     

稻曲病菌拮抗菌的筛选及拮抗活性测定

从对稻曲病菌具有拮抗能力的18株芽孢杆菌中选出拮抗作用较强的2个菌株A1、H-51,分别对稻曲病菌进行抑菌率、致畸性、孢子萌发影响以及对水稻促生性试验。结果表明,拮抗菌2个菌株的发酵液对稻曲病菌最大抑制率分别为67.66％和62.87％,对病菌分生孢子的萌发抑制率分别为77.37％和59.74％,但对水稻促生作用不明显。     

花生白绢病互作研究进展

花生白绢病是由齐整小核菌Sclerotium rolfsii Sacc.引起的发生在花生茎基部的真菌病害,严重制约花生的品质与产量。本研究利用稀释涂布法从怀牛膝根际土壤中分离纯化了116株放线菌,并从中筛选鉴定了能够防控花生白绢病的菌株。通过平板对峙试验筛选得到两株抑菌活性较好且稳定的菌株Soil-1-5和Soil-3-28,它们对齐整小核菌的抑制率分别为80.43%和92.34%。根据形态学观察、生理生化试验及16S rRNA基因序列分析鉴定,两株菌分别被鉴定为疮痂链霉菌Streptomyces scabiei和藤黄灰链霉菌Streptomyces luteogriseus。采用菌丝生长速率法测定了拮抗菌株无菌发酵滤液对植物病原菌的抑制作用。结果表明,菌株Soil-1-5和Soil-3-28的无菌发酵滤液稀释5倍后对齐整小核菌的抑制率分别为73.67%和57.11%,且对禾谷镰孢菌等6种植物病原菌均有不同程度的抑制作用。此外,两菌株的无菌发酵滤液对齐整小核菌菌核萌发和菌核形成也有较好拮抗作用。盆栽试验结果表明,菌株Soil-1-5和Soil-3-28对花生白绢病的防治效果分别为51.92%和31.74%,其中菌株Soil-3-28对花生生长有促进作用。综上,菌株Soil-1-5和Soil-3-28对花生白绢病有较好的防治效果,具有潜在应用价值。

     

土传黄瓜立枯病高效拮抗菌的筛选鉴定及其生物效应

采用平板对峙法从黄瓜根际土壤中分离出的400余株细菌菌株中筛选出16株对立枯丝核菌Rhizoctonia solani具有拮抗效果的菌株,抑菌带直径在0.81～1.93 cm之间。并从中选出3株抑菌带直径在1.6cm以上的菌株N33、N35和N43,结合形态、生理生化特性及16S rDNA序列比对分析,鉴定N33菌株为假单胞菌属Pseudomonas sp.,N35和N43菌株为芽胞杆菌属Bacillus sp.。通过在黄瓜育苗基质中添加选育的高效拮抗菌株,观测其对黄瓜苗生长的促进作用以及对立枯病的防治作用,3株菌株均具有促进黄瓜苗期生长和防治苗期立枯病的作用,其中N43菌株促生及防病效果均最显著,地上部鲜重比对照处理增加62.16%,防治效果达62%。     

棉花黄萎病拮抗细菌H14的筛选鉴定及其拮抗机理分析

为筛选对大丽轮枝菌Verticillium dahliae Kleb.具有拮抗活性的根际细菌,以大丽轮枝菌为靶菌,分离获得棉花根际细菌,利用平板对峙法和琼脂扩散法筛选具有较高拮抗活性的菌株,采用室内盆栽法测定筛选所得菌株对棉花黄萎病的防效,并通过形态特征、生理生化特性和16S r DNA基因序列分析确立其分类地位,采用底物降解法和抗菌肽基因克隆法检测其产生水解酶和抗菌肽的能力。结果显示,试验共分离获得182株棉花根际细菌,筛选得到3株对大丽轮枝菌抑菌率大于50.00%且抑菌圈直径大于15 mm的菌株,其中菌株H14的抑菌率和抑菌圈直径最大,分别为54.25%和18.10 mm。该菌能在0～9%NaCl和pH 4.5～9.0的NB培养基上生长;经形态特征观察、生理生化试验及16S rDNA基因序列分析,最终将其鉴定为莫哈韦芽胞杆菌Bacillus mojavensis;菌株H14对大丽轮枝菌孢子萌发的抑制率和对棉花黄萎病的盆栽防效分别为89.55%和74.57%;菌株H14能合成蛋白酶,含有srfAA、fenD、bacA脂肽类抗生素合成基因。表明莫哈韦芽胞杆菌菌株H14能够合成蛋白酶和脂肽类拮抗物质,具有良好的抑菌和防病能力。     

加工型辣椒品种对细菌性叶斑病的抗性鉴定

由黄褐假单胞Pseudomonas fulva侵染引起的辣椒细菌性叶斑病是近期在新疆北疆加工辣椒生产区发现的一种新病害。为评估辣椒品种对细菌性叶斑病的抗性水平,采用苗期人工接种结合田间调查,鉴定了生产中13个主栽辣椒品种对细菌性叶斑病的抗性。结果表明,供试材料对细菌性叶斑病均无免疫性,但不同辣椒品种对辣椒叶斑病抗性差异较大,‘改良佳线1号’和‘丰力红冠’为抗病品种;‘佳线4号’、‘新丰6号’、‘秦椒王’、‘陕早红’、‘宝丰2号’为耐病品种;‘红安6号’、‘朝天椒’为感病品种,高感品种为‘金塔’、‘铁皮椒’、‘红龙13’和‘超级韩国甜椒’。研究结果为辣椒的抗病育种和病害防治提供了基础。     

禾谷丝核菌拮抗细菌的鉴定及其拮抗产物分析

 小麦纹枯病是长江中下游麦区重要的土传病害。明确小麦根际拮抗细菌种类和拮抗机理对小麦纹枯病生物防治具有重要意义。从江苏省姜堰市小麦根际分离到拮抗细菌A6、A12、B37、B40、B41和B42。通过生理生化测定和16SrDNA同源序列比对分析,这些菌株都为荧光假单胞杆菌。在温室条件下进行种子处理,这些菌株对小麦纹枯菌都有一定的控制作用,其中菌株A6、B41和B42的控病作用明显,菌株B42对小麦纹枯病的控制效果最好,温室防效达71.67%。基因检测和次生代谢物分析的结果表明:6个菌株都不产生2,4-二乙酰基间苯三酚和吩嗪,菌株A12、B37和B42含有与吡咯菌素合成相关的基因PrnD,在菌株B37中还检测到合成基因PrnC;菌株A12、B37和B42产生嗜铁素;除了菌株A6和B37,其余菌株都产生蛋白酶;所有菌株都不产生几丁质酶。     

梨炭疽菌拮抗细菌CL01的鉴定及其抗菌活性

为挖掘可用于防治梨炭疽病的生防资源,从江苏句容梨园根际土壤中分离筛选到1株对梨炭疽菌具有较强拮抗效果的菌株CL01.基于菌落菌体形态、生理生化特征、16S rDNA和rpoB基因序列,该菌株被鉴定为奥托威类芽胞杆菌Paenibacillus ottowii.CL01能够抑制梨炭疽菌菌丝的生长,受抑制的真菌菌落边缘菌丝变...     

香蕉枯萎病拮抗菌的筛选及其作用机制研究

通过分离和筛选,从香蕉园或者其他果园的土壤中分离获得13株对香蕉枯萎病菌(Fusarium oxysporum f.sp.cubense)具有抑制作用的拮抗菌,并对部分拮抗菌抑制病菌菌丝生长和孢子萌发进行了试验。结果表明,拮抗菌株d4、d5、B3和p发酵液对香蕉枯萎病菌生长具有显著的抑制作用,在平板上产生的抑菌圈直径为21.75～34.75 mm,抑菌效果具有持续稳定性,对孢子萌发的抑制率为90.49%～97.18%;拮抗菌对病菌的作用表现为对菌丝的消融、菌丝细胞的泡囊化、抑制病菌分生孢子的萌发、孢子芽管的扭曲。     

木霉对烟草黑胫病菌的拮抗机制

采用平板对峙法、水解酶平板活性测定、圆孔滤液法与游动孢子萌发检测、挥发性抑菌物检测等方法探讨木霉生防菌株对烟草疫霉Phytophthora nicotianae Brada de Hann的拮抗作用机制。结果表明,TR13、TR14、TR17对烟草疫霉有竞争、重寄生和抗生作用,但不同木霉菌株对烟草疫霉的拮抗作用有所不同,TR13、TR14的竞争作用较强;水解酶平板活性测定表明,3个木霉菌株均产生β-1,3-葡聚糖酶、纤维素酶,两种酶均参与烟草疫霉细胞壁的消解,以TR13与TR14酶的活性较高;3个菌株均产生非挥发性抗生素抑制烟草疫霉菌孢子萌发,但对疫霉菌菌丝的生长影响不大;TR13、TR14几乎不产生挥发性抗生素,TR17则产生挥发性抗生素,明显抑制疫霉菌生长。     

Sterol content and other characteristics of pimaricin-resistant mutants of Aspergillus nidulans

Pimaricin-resistant mutants of Aspergillus nidulans were selected on a medium containing the polyene-antibiotic. Some resistant mutants contained markedly reduced amounts of ergosterol, but others contained almost normal levels of this sterol. Most resistant mutants which lacked ergosterol had a biochemical lesion in sterol C-22 desaturation. Analysis of sterols in one of these isolates showed the presence of 5,7-ergostadienol, 5,7,24(28)-ergostatrienol, and 5,8-ergostadienol. The sterol C-14 demethylation inhibitor, fenarimol, was more toxic to this mutant than to the wild type. On the other hand, mutants inactive in sterol C-22 desaturation were resistant to oligomycin but showed wild type sensitivity to carboxin. Attempts to select sterol C-14-demethylation-deficient mutants of Aspergillus nidulans, Monilinia fructicola, and Pyricularia oryzae on polyene-containing media were unsuccessful. Apparently C-14-methyl sterols do not support growth of these filamentous fungi.     

利迪链霉菌E12发酵液及其粗提物的抑菌活性初探

利迪链霉菌E12的发酵液对多种植物病原真菌具有较强的抑制活性。采用琼脂稀释法考察了淀粉和葡萄糖对利迪链霉菌发酵液抑菌活性的影响;采用大孔树脂吸附层析和浓缩沉淀提取发酵液中的抑菌活性物质;利用聚丙烯酰胺凝胶(SDS-PAGE)与含β-1,3-葡聚糖的琼脂糖凝胶直接反应测定粗提物的分子质量及β-1,3-葡聚糖酶活性并采用琼脂扩散法测定其抑菌活性。结果表明:当以淀粉作为惟一碳源时发酵液的抑菌率为89.2%,当以葡萄糖作为惟一碳源时发酵液无抑菌活性;在SDS-PAGE上仅出现1条大小为3 kDa左右的条带,且该条带具有抑菌活性,与其位置对应的琼脂糖凝胶上出现透明带。以上结果表明,利迪链霉菌E12可能产生一种具有β-1,3-葡聚糖酶活性的抑菌活性物质。     

黄柄曲霉ASD次生代谢产物的鉴定及抑菌活性

为明确土壤来源的黄柄曲霉ASD次生代谢产物中的抑菌物质及其生物活性,利用硅胶柱层析、凝胶柱层析和半制备高效液相色谱对ASD发酵液的二氯甲烷粗提物进行纯化,运用核磁共振及高分辨质谱技术,结合文献报道数据对化合物的结构进行鉴定,利用X-射线单晶衍射验证鉴定结果;采用含药稀释平板法及菌丝生长速率法测定化合物对辣椒疫霉菌、大斑凸脐蠕孢菌和禾生刺盘孢菌等9种病原真菌的抑菌活性。结果表明,从ASD发酵液中分离获得已知活性抑菌物质为sulochrin和4,5-二甲基间苯二酚。其中,sulochrin对辣椒疫霉菌、大斑凸脐蠕孢菌和禾生刺盘孢菌有较好的抑制作用,4,5-二甲基间苯二酚只对辣椒疫霉菌具有较好的抑制作用,两者对灰葡萄孢菌、玉蜀黍尾孢菌和新月弯孢菌等6株致菌抑菌效果不明显或无抑制作用。     

Immunohistochemical investigation of cotton carpel tissue exposed to xylanolytic hydrolases of Aspergillus flavus

Cotton carpel tissue (35–45 days post-anthesis) that had been treated with a mixture of xylanolytic hydrolases derived from Aspergillus flavus was subjected to immunocytochemical analysis. Microscopic examination of treated tissues revealed severe degradation of the secondary wall structure. Control tissue cells revealed the presence of high concentrations of xylans/arabinoxylans throughout the cell wall, as well as significant concentrations of arabinogalactan proteins in secondary wall structure. Carpel cells treated with a mixture of A. flavus-produced xylanolytic hydrolases showed a much reduced presence of labeling by xylan-specific antibodies on the inner wall surface, suggesting a severe loss of these plant polysaccharides in the secondary wall structure. Carpel exposure to a purified 14 kD endoxylanase from A. flavus also resulted in a severe reduction of xylans from secondary wall structure, although penetration of the tissue was not as dramatic. Arabinogalactan proteins were not as severely affected by the xylanolytic hydrolases. Comparison of control tissue with hydrolase-treated tissue stained with toluidine blue revealed an apparent reduction in wall thickness, supporting the conclusion of secondary wall structure degradation. Interestingly, the pectins could only be detected in the samples treated with xylanolytic enzymes, indicating that the pectins were being masked by xylans. These results are consistent with the conclusion that the xylanolytic hydrolase complex of A. flavus is a critical factor for host cell wall maceration and may represent another important fungal virulence factor, in addition to pectolytic hydrolase activities.     

羌活提取物微乳剂制备及体外抗真菌活性

采用乙醇溶液热萃取方法提取羌活根茎活性成分,通过稳定性等物理性能测试进行评价并确定微乳剂最优配方。微乳剂由质量分数 (下同)为15%的羌活提取物 (含羌活醇0.34%)、9%的环己酮、16%的HMK-2100、10%的乙醇、49.9%的水和0.1%的AF1506组成。理化性质测试结果表明,所制备的羌活提取物微乳剂各项指标均符合国家标准。为了比较研究其抑菌活性,同时还制备了5%羌活提取物 (含羌活醇0.11%)微乳剂和30%羌活提取物 (含羌活醇0.68%)乳油。采用菌丝生长速率法研究了羌活提取物不同剂型产品对小麦纹枯病菌Rhizoctonia cerealis、辣椒疫霉Phytophthora capsici、水稻稻瘟病菌Pyricularia grisea和水稻纹枯病菌Rhizoctonia solani的抑制作用。结果显示,15%羌活提取物微乳剂的抑菌效果显著,在用水稀释500倍 (羌活提取物质量浓度为300 mg/L,羌活醇质量浓度为6.75 mg/L)后施用,其对水稻稻瘟病菌和水稻纹枯病菌菌丝生长的抑制率均在90%以上。应用性能测试结果表明,羌活提取物微乳剂在绿萝叶片表面具有优异的铺展和润湿能力。     

拮抗细菌CE抑菌物质对桃褐腐病菌的抑制作用及其机制

采用十字交叉法和血球计数板法测定草莓枯萎病菌的生物学特性。结果表明,PSA培养基最适合草莓枯萎病菌的生长和产孢,菌落生长的适温为5～40℃（最适温度为28℃）,适宜pH值为5,菌落产孢的适宜pH值为6,且光暗交替可促进菌落的生长和产孢,同时明确了不同碳氮源对草莓菌落生长及产孢的影响。采用菌落生长速率法,测定草莓枯萎病菌对7种杀菌剂的敏感性,烯唑醇和戊唑醇对菌落生长的抑制毒力最强,EC50分别为0.2072μg·mL-1和0.2130μg·mL-1,分别是多菌灵的238.85倍和232.35倍;腈菌唑和苯醚甲环唑的毒力次之,EC50分别为0.7929μg·mL-1和0.8513μg·mL-1,分别是多菌灵的62.42倍和58.13倍;三唑酮和霜霉威的毒力最小     

Degradation of acylanilide pesticides by Aspergillus niger

Aspergillus niger converts the herbicide 3′-chloro-2-methyl-p-valerotoluidide (solan) to 3′-chloro-4′-methylacetanilide and the fungicide 2,5-dimethylfuran-3-carboxanilide to acetanilide. The metabolites were formed by hydrolysis with an aryl acylamidase, followed by subsequent acetylation resulting in the corresponding acetanilides. Their structures were elucidated by mass spectrometric analysis and confirmed by comparison with synthetic compounds.     

Inhibitory and stimulatory effects of essential oils and individual monoterpenoids on growth and sporulation of four soil-borne fungal isolates of Aspergillus terreus, Fusarium oxysporum, Penicillium expansum, and Verticillium dahliae

The effect of essential oils and individual monoterpenoids on soil-borne fungi, in pure and mixed cultures, in growth media and in the soil environment, was investigated. Essential oils were extracted from lavender (Lavandula stoechas), oregano (Origanum vulgare subsp. hirtum), sage (Salvia fruticosa) and spearmint (Mentha spicata). The monoterpenoids tested were fenchone, carvacrol, 1,8-cineole, carvone, α-pinene and terpinen-4-ol. Their effect was examined on growth and sporulation of Aspergillus terreus, Fusarium oxysporum, Penicillium expansum and Verticillium dahliae isolated from an organic cultivation of tomato. All tested essential oils and individual monoterpenoids inhibited mycelial growth in all fungi and conidial production in most fungi. The strongest inhibitory activity on mycelial growth was exhibited by oregano and spearmint oils and by carvacrol and carvone, respectively their main constituents. The inhibitory activity was clearly fungistatic in A. terreus and F. oxysporum but fungicidal in V. dahliae. On sporulation, clearly stimulatory effects were observed alongside inhibitory ones. Conidial production was always promoted by α-pinene in P. expansum and by sage oil in F. oxysporum. At certain dosages it was promoted by cineole and carvone in F. oxysporum, and by lavender oil in A. terreus and V. dahliae. Experiments with carvone and carvacrol against mixed fungal cultures in a soil environment showed that V. dahliae was the most sensitive and A. terreus the most tolerant of the four fungi. Our results demonstrate strong but divergent effects and selectivity of action of the lower terpenoids on fungal strains that can become serious pests of tomato. Of special importance is the complete inhibition of growth and conidial production of V. dahliae, a pathogen otherwise very resistant to chemical control.     

甜菜夜蛾抗药性及其机理

采用FAO推荐的点滴法,测定了甜菜夜蛾山东泰安种群对顺式氯氰菊酯等5种药剂的抗性,以及5种增效剂对5种药剂的增效作用。用浸渍法测定了2种渗透剂对药剂的增效作用。结果表明,泰安种群与敏感种群相比,对顺式氯氰菊酯、灭多威、丙溴磷、氟啶脲和硫丹的抗性分别达1536.0,532.1,193.5,25.0及11.9倍,且抗性谱广、发生程度严重。5种增效剂和2种渗透剂与不同药剂混用,均表现不同程度的增效作用,表明该种群为多重抗性。其抗性与多功能氧化酶、水解酶、谷胱甘肽-S-转移酶的活性提高有关,也涉及抗击倒基因敏感度和表皮穿透性的改变。