Effects of Cry1Ac toxin of Bacillus thuringiensis and nuclear polyhedrosis virus of Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae) on larval mortality and pupation

In the laboratory, the percentage mortality and pupation of Helicoverpa armigera (Hübner) were investigated when larvae were exposed to Cry1Ac of Bacillus thuringiensis Berliner, nuclear polyhedrosis virus of H. armigera (HaNPV) or Cry1Ac and HaNPV together. The results revealed that interactions between Cry1Ac and HaNPV varied with bioassay method and concentration of the suspension. When larvae were infected using a suspension containing both HaNPV and Cry1Ac, most combinations of Cry1Ac (62.5, 125 and 250 microg mL(-1)) and HaNPV (1.2 x 10(6), 6.0 x 10(6) and 3.0 x 10(7) PIB mL(-1)) showed an antagonistic effect. In the bioassay procedure where larvae were force fed diet containing Cry1Ac 48 h after being infected by HaNPV, interaction between Cry1Ac (0.5, 1, 2, 4 and 8 microg mL(-1)) and HaNPV (6.0 x 10(6) and 3.0 x 10(7) PIB mL(-1)) showed an additive effect, while combinations of Cry1Ac (0.5, 1, 2 and 4 microg mL(-1)) and HaNPV (1.2 x 10(6) PIB mL(-1)) showed an antagonistic effect. In the bioassay procedure where larvae being infected by HaNPV were fed on Cry1Ac diet from neonate to death or pupation, the results suggested that Cry1Ac and HaNPV showed an additive interaction. The percentage mortality was lower in the treatment of larvae infected by transgenic Bt cotton leaf discs containing HaNPV suspension than in the treatment of larvae by conventional cotton leaf discs containing HaNPV, while the pupation rate was higher. The combination of Bt cotton and HaNPV showed antagonism. The present results showed that a combination of Cry1Ac and HaNPV usually resulted in mortality levels greater than in the case of Cry1Ac but not greater than with the virus alone.     

基于形态学与ITS序列对冠突散囊菌多样性研究

为研究茯砖茶中冠突散囊菌多样性的问题,以来自中国3个省份的5种茯砖茶为样品,从中分离出优势微生物金花菌4、16、B1、B3、B7,依托形态学与和分子生物学技术对菌株进行鉴定。结果发现,这5株菌株均属冠突散囊菌,并且存在着遗传多样性,其中16号菌株的电镜观察结果与其他4株菌株差异明显,且菌株子囊孢子的“赤道”周围存在较多小孔;在ITS序列构建系统发育树中4、B1、B3、B7聚类为一组,16号聚类成另一组,说明冠突散囊菌可能存在亚种,这与形态学结果一致。     

黏质沙雷氏菌次生代谢物对TMV的抑制机理

【目的】分离、纯化和鉴定黏质沙雷氏菌2A2次生代谢物中具有抑制TMV侵染力的活性成分,明确其对TMV的抑制机理。【方法】通过病毒生物学测定,确定黏质沙雷氏菌2A2菌株对TMV的抑制效率。为明确黏质沙雷氏菌2A2发酵液次生代谢物中抑制TMV侵染力的活性成分,通过TLC和硅胶柱层析对其发酵液进行了分离纯化,获得主要的次生代谢物,通过局部枯斑法测定各代谢物的生物学活性,筛选可显著抑制TMV侵染活性的物质,经NMR分析,确定其成分结构,进而确定物质组分。为进一步揭示代谢物对TMV的抑制机理,利用透射电子显微镜探索代谢物对TMV粒体形态的影响,TMV粒体与代谢物甲醇溶液混合30 min后,于透射电子显微镜下观察TMV粒体形态。同时,用代谢物甲醇溶液喷施处理寄主下部3片叶,共设5个处理：处理Ⅰ,喷施代谢物24 h后,接种TMV;处理Ⅱ,代谢物与等体积TMV汁液混合30 min后,接种叶片;处理Ⅲ,先接种TMV,24 h后喷施代谢物;处理Ⅳ,宁南霉素（50 μL•mL-1）与等体积TMV汁液混合30 min后,接种下部叶3片做阳性对照;处理Ⅴ,无菌水与等体积TMV汁液混合30 min后,接种下部叶3片做空白对照。分别在次生代谢物诱导和TMV侵染后1、3、5、7、9 d,取未经任何处理的上部叶,TRIZOL法快速提取总RNA并反转录cDNA,利用real-time RT-PCR分析样品PR基因和TMV的RNA相对表达量,进而明确代谢物处理寄主对寄主PR基因表达的影响和对TMV在寄主体内复制增殖的抑制作用。【结果】黏质沙雷氏菌2A2菌株对TMV具有显著的抑制作用。对其发酵液进行分离纯化,获得3种主要代谢物,通过局部枯斑法测定,编号为BJH-2的代谢物可显著抑制TMV的侵染活性,经NMR分析,确定了该代谢物为灵菌红素（prodigiosin）。透射电子显微镜结果显示灵菌红素可显著破坏TMV粒体,使TMV典型的杆状病毒粒体降解断裂成排列紊乱的短杆状。real-time RT-PCR结果表明,灵菌红素处理植株后,处理Ⅰ、处理Ⅱ、处理Ⅲ可诱导寄主体内PR基因表达随着时间逐渐上调,第7 天左右,三者处理的寄主体内PR1、PR2、PR4、PR5表达上调至高峰,显著高于空白对照,亦高于处理Ⅳ的阳性对照,从而提高系统抗性;灵菌红素处理植株后,处理Ⅰ、处理Ⅱ、处理Ⅲ寄主体内TMV RNA的表达量随着时间上调减缓,在接种TMV后的第9天,三者处理的寄主体内的TMV RNA含量分别是空白对照的11.98%、5.23%、15.90%,显著低于空白对照,亦低于处理Ⅳ的阳性对照。【结论】灵菌红素既对TMV在寄主体内的复制增殖具有抑制作用,又可诱导寄主产生系统抗性,整体效果高于宁南霉素,可作为防治植物病毒新的生物制剂。     

产玫瑰红色素粘质沙雷氏菌的分离鉴定及抑菌活性

旨在分离鉴定产玫瑰红色素菌株,及其分离菌株和色素对病原菌的生物拮抗作用。经形态观察、生理生化试验和16S rRNA序列分析对其进行鉴定,通过控制变量法确定最佳产色素的温度及光照条件,破裂菌体提取色素分析此菌株产色素的量和红色素的性质,采用滤纸片法研究对病原菌的拮抗作用。结果表明：（1）分离细菌Dse\|01菌株最终鉴定为粘质沙雷氏菌（Serratia marcescens）;（2）该菌在25～35 ℃条件下培养色素形成较早且颜色较深,30 ℃温度下产色素最佳,光照对其色素形成时间没有明显影响;（3）30 ℃ 150 r·min-1培养24 h,提取获得色素粗品确定为灵菌红素,产量达（567.90±7.77）mg·L-1;（4）分离菌株的全菌液、上清液、菌体和提取色素对病原性大肠杆菌、沙门氏菌均无拮抗作用,而全菌液、菌体和提取色素对金黄色葡萄球菌具有明显的抑制作用。     

不同生态环境下野生大豆根解剖结构演化研究

以不同生态环境下的野生大豆,即:野生大豆2013-001-01(盐生环境)和野生大豆2013-001-02(中生环境)为试材,基于结构植物学角度,应用石蜡切片和光学显微技术对供试的野生大豆根解剖结构进行比较研究,旨在为我国大豆属植物颉颃盐逆境研究提供解剖学依据。结果表明:2013-001-02根的维管柱为五原型,2013-001-01根的维管柱已经演化为四原型,相对于2013-001-02,2013-001-01根的皮层较完整,韧皮纤维发达且形成"韧皮屏障",次生木质部导管数量较多且存在侵填体,导管的傍管薄壁细胞较大且数量较多,射线径向长度相对较短。研究证明:野生大豆2013-001-01植物的根已经演化出颉颃盐逆境解剖结构。     

1株油茶病害拮抗真菌的鉴定、生物学特性及拮抗作用研究

为了给油茶病害的生物防治提供科学依据,从油茶根部土壤分离获得1株能拮抗油茶主要病害的真菌F217-3。依据该菌株的形态特征,并结合全自动微生物鉴定系统及r DNA-ITS序列分析,将其鉴定为草酸青霉(Penicilium oxalicum)。对其生物学特性研究结果表明:该菌株在查氏培养基上生长最佳,碳源和氮源分别以蔗糖和尿素最适宜,生长适温为28℃,致死温度为65℃,在p H值为3~10条件下均生长良好,其中最适p H值是4~7,通气与否对菌丝生长无明显影响,但光照24 h条件下菌丝生长较差。平板对峙试验结果表明,F217-3对油茶炭疽病菌(C.nicotianae)、油茶软腐病菌(A.camellia)、黑斑病菌(A.alternate)和溃疡病菌(B.dothidea)的抑菌率均达到90%以上,可抑制油茶胶孢炭疽病菌、黑斑病菌分生孢子产生,抑制或致畸油茶炭疽病菌、溃疡病菌和叶斑病菌菌丝,是一株具有广谱拮抗作用的生防菌株。     

香蕉枯萎病生防细菌的筛选、鉴定及其抑菌作用

采用平板对峙法,从香蕉根部土样中分离到1株枯萎病拮抗芽孢杆菌XY-10。根据形态特征及16S rDNA分析将其鉴定为多粘类芽孢杆菌Paenibacillus polymyxa。拮抗试验表明菌株XY-10不同浓度的发酵滤液均可抑制病原菌孢子萌发和生长,当发酵滤液浓度分别为10%、30%和50%时,其在高度抑制水平上的抑制率分别为7%、23%和40%。经发酵滤液处理,病原菌菌丝末端膨大成球状、菌丝破裂、消解,菌丝体分枝增多。     

长白山地区不同海拔环境拮抗放线菌的筛选

利用稀释平板倾注法从吉林长白山地区不同海拔高度的不同环境中共分离到各种放线菌菌株104株.以24种农业常见植物致病真菌作为目标供试菌,采用抑制菌丝生长速率法对104株放线菌进行抗菌活性筛选,优选抗菌活性较佳的12株.将最佳放线菌No.6菌株进行液体发酵,并测定其发酵液滤液的抑菌活性.结果表明,No.6放线菌菌株对香蕉灰纹菌、辣椒疫霉菌、黄瓜炭疽菌、番茄灰霉菌、黄瓜黑星菌、番茄叶霉菌、番茄炭疽菌、棉花黄萎菌、黄瓜枯萎菌、小麦赤霉菌、稻曲病菌、水稻恶苗病菌、小麦纹枯病菌、水稻稻瘟病菌、水稻纹枯菌、油菜菌核菌均具有优异的防治效果,发酵滤液稀释10倍后对黄瓜黑星菌、番茄炭疽菌、稻曲病菌、水稻稻瘟病菌、水稻纹枯菌的抑制率分别达90.0%、93.3%、92.0%、93.3%、92.0%.     

Efficacy of insecticide mixtures against larvae of Culex quinquefasciatus (Say) (Diptera: Culicidae) resistant to pyrethroids and carbamates

The efficacy of insecticide mixtures of permethrin (pyrethroid) and propoxur (carbamate) was tested by larval bioassays on two strains of Culex quinquefasciatus (Say), one resistant to pyrethroids and the other resistant to carbamates. The method consisted in combining one insecticide at the highest concentration causing no mortality (LC0) with increasing concentrations of the second one. The concentration-mortality regression lines were determined for permethrin and propoxur alone and in combination, and synergism ratios (SR) were calculated in order to determine the magnitude of an increase or decrease in efficacy with use of the mixtures. With the pyrethroid-resistant strain (BK-PER), the results showed that propoxur at LC0 significantly enhanced the insecticidal activity of permethrin (SR50 = 1.54), especially on the upper range of the concentration-mortality regression. Conversely, when permethrin at LC0 was tested with propoxur against the carbamate resistant strain (R-LAB), an antagonistic effect was observed (SR50 = 0.67). With the BK-PER strain, an increased oxidative detoxification (MFO) appeared to be the main mechanism responsible for the synergistic interaction. Nevertheless, antagonism in the R-LAB strain is probably due to a physiological perturbation implying different target sites for pyrethroid (ie sodium channel) and carbamate insecticides [ie acetylcholinesterase (EC 3.3.3.7) and choline acetyltransferase (EC 2.3.1.6)].     

Hyperparasites of Rhizoctonia solani in Dutch potato fields

Gliocladium roseum was found to be the most common and probably the most effective mycoparasite in potato fields in the northern parts of the Netherlands. It is able to parasitize and kill living hyphae at temperatures of 12°C and higher. Sclerotia ofR. solani are often infected and killed by this fungus under suitable conditions, i.e. at temperatures of 16°C and more. Killing of sclerotia by other antagonistic organisms was also observed. It is also shown by not parasitic fungi and is caused by toxins produced by the antagonist.The development of theG. roseum population was studied during the growth of a potato crop in two soils. In both soils its initial level was very low. In both a slightly acid sandy soil and a neutral sandy loam, suppression ofR. solani can occur;G. roseum accumulated in the former mainly under continuous potato crops,Colletotrichum coccodes was the main antagonist in the latter.Samenvatting In de meeste Nederlandse aardappelakkers komen schimmels voor dieRhizoctonia solani kunnen aantasten en doden. De meest algemene, en waarschijnlijk ook de meest belangrijke, die we tot nu toe vonden, isGliocladium roseum (Tabel 1). Het is bekend, dat deze schimmel stoffen produceert die voorR. solani giftig zijn. Met behulp hiervan kanG. roseum, evenals andere antibiotisch actieve micro-organismen, ook de sclerotiën doden (Tabel 2). Voor doding doorG. roseum is de temperatuur een factor van belang. Hyfen worden nog gedood bij een temperatuur van 12°C, waarbij de sclerotiën niet meer aangetast kunnen worden. Gedurende het winterseizoen worden sclerotiën door deze schimmel naar alle waarschijnlijkheid niet gedood.De ontwikkeling van de populatie vanG. roseum en andere antagonisten vanR. solani werd gevolgd in aardappelvelden op een licht zure zandgrond en op een neutrale zware zavel. Op de zandgrond werden twee proefplekken bemonsterd: één waarop voor het vierde achtereenvolgende jaar aardappelen werden geteeld en één met een vruchtwisselingsschema van graan, bieten en aardappelen.In de zandgrond nam in het groeiseizoen de populatie vanG. roseum toe. Op de proefplek waar voor het vierde jaar achtereen aardappelen stonden werdR. solani vanaf half augustus onderdrukt, evenwel niet volledig. Ook in het vruchtwisselingsstuk breiddeG. roseum zich flink uit, doch een onderdrukking vanR. solani werd niet bereikt.In de zware zavel nam de populatie vanG. roseum niet toe. Hier werdR. solani — uit besmet pootgoed — onderdrukt doorColletotrichum coccodes (zelf een pathogeen van stolonen) en antagonistische bacteriën. De resultaten zijn vermeld in Tabel 3.De besmetting van de geoogste knollen met sclerotiën, zoals die voorkwam op de zandgrond, is in Tabel 4 vermeld. Op de zavel leverde schoon pootgoed een bijna schone oogst (2% van de knollen was zeer licht bezet met sclerotiën). Besmet pootgoed leverde een oogst met 58% schone knollen, 35% met een zeer lichte en 7% met een iets zwaardere sclerotiënbezetting. Hoewel uit 100% besmet pootgoed een veel schonere oogst werd verkregen, was eerder toch een beschadiging van het gewas opgetreden. Pas tegen het eind van het groeiseizoen werdR. solani flink onderdrukt.