烟草黑胫病拮抗根际芽胞杆菌FB-16的筛选鉴定及其抑菌活性

为获得对烟草黑胫病有较强生防效果的拮抗细菌,从健康烟草根际采集30份土壤样品,分离纯化得到347株细菌,经病原真菌定向筛选后得到1株对烟草黑胫病菌等病原真菌拮抗活性较好的细菌FB-16,其对烟草黑胫病菌的抑菌带宽为23mm。采用菌丝生长速率法、作用机制试验、温室防病试验测定FB-16的抑菌作用,并通过形态学特征、生理生化特征及16S rDNA序列分析对菌株进行鉴定。结果显示,菌株FB-16发酵液对烟草黑胫病菌菌丝生长抑制率为95.96%;其代谢产物对烟草黑胫病菌菌丝有致畸作用;菌株FB-16为解淀粉芽胞杆菌Bacillus amyloliauefaciens(Gen-Bank登录号为JN245982);饱和度25%硫酸铵获得的抑菌物质对烟草黑胫病菌的抑菌活性较高,抑菌圈直径达42.00 mm;FB-16活性产物处理的烟草植株在接种烟草黑胫病菌7天后的防治效果为69.96%。表明菌株FB-16在烟草黑胫病生物防治中具有潜在的利用价值。     

内生枯草芽胞杆菌Itb57对烟草疫霉的抑制及生理生化影响

为明确内生枯草芽胞杆菌Itb57菌株抑制烟草疫霉生长的机理,分别采用电镜扫描法、生化测定法和电导法研究了其对烟草疫霉菌丝形态及生理生化指标的影响。结果显示,经Itb57菌株发酵滤液处理的疫霉菌丝畸形膨大、分枝短粗,孢子囊的形成及萌发受到显著抑制,当发酵滤液浓度为50%时,抑制率分别为79.69%和70.85%;且与对照相比,处理组疫霉菌细胞壁主要成分碱溶性、碱不溶性和水溶性的β-1,3-葡聚糖含量分别降低了51.18%、42.56%和39.42%,β-1,3-葡聚糖合成酶活性降低了72.45%,DNA含量降低了14.64%;线粒体复合酶活性显著低于对照,其中复合酶II活性降低了79.25%,菌丝处理液电导率升高。表明枯草芽胞杆菌Itb57菌株发酵滤液能减少疫霉细胞壁主要成分β-1,3-葡聚糖含量,破坏细胞膜的完整性,导致菌丝生长受阻或产生畸形,对烟草黑胫病的控制具有潜在的生防价值。     

几丁寡糖对烟草黑胫病的控制效应及其机制

在室内条件下,采用菌丝生长速率法测定了几丁寡糖对烟草黑胫病菌的抑制作用,继而在温室盆栽和人工接种条件下,分别测定几丁寡糖、木霉、几丁质、几丁寡糖+木霉、几丁寡糖+木霉+几丁质等5种处理对烟草黑胫病的防治效果。结果表明,几丁寡糖在离体条件下对烟草黑胫病菌菌丝生长无抑制作用,但在盆栽试验中对烟草黑胫病具有显著防治效果。5种处理中,几丁寡糖单独处理对烟草黑胫病的防治效果最高,为57.81%,其次为几丁寡糖+木霉+几丁质和几丁寡糖+木霉处理,分别为53.49%和52.49%。在温室条件下对烟草植株施用几丁寡糖及人工接种黑胫病菌后10天内,几丁寡糖能显著提高烟草体内的SOD、POD、PPO和几丁质酶活性,且活性峰值分别比对照提高96.30%、136.36%、10.34%和9.10%。     

一株烟草黑胫病高效拮抗菌的筛选、鉴定及生防潜力评价

采用稀释涂布平板法、对峙培养法和牛津杯法从烟草黑胫病土壤中分离和筛选出21株对烟草黑胫病有拮抗作用的细菌,其中,XCS007菌株拮抗作用最明显,运用微观形态学、生理生化特征及16Sr DNA基因序列鉴定其为一株铜绿假单胞菌(Pseudomonas aeruginosa)。菌株XCS007菌液及其粗提物对烟草黑胫病病菌菌丝的抑制率分别是91.80%和97.11%;盆栽试验结果显示,其防治效果分别是83.18%和83.55%;大田试验结果显示,其防治效果分别达到78.64%和79.54%。     

烟草黑胫病拮抗菌的筛选、鉴定及发酵条件优化

为获得对烟草黑胫病（tobacco black shank）具有较好拮抗作用的菌株,从食腐昆虫美洲大蠊Periplaneta americana肠道分离纯化得到1株对烟草黑胫病具有较强抑制作用的D5-8菌株,拟进一步明确该菌株的分类地位,优化其发酵条件并测定其对烟草黑胫病的防治效果。利用形态特征、生理生化特性观察及16S rRNA基因序列分析对D5-8菌株进行鉴定;以细菌发酵液OD₆₀₀值及对病原菌的抑制率为指标,对其发酵培养基和发酵条件进行单因素试验及正交试验;通过室内盆栽试验,明确菌株D5-8对烟草黑胫病的防治效果。结果表明,经平板对峙试验发现菌株D5-8对烟草寄生疫霉的抑制率达66.80%;通过形态特征和生理生化特征初步确定菌株D5-8为芽胞杆菌属Bacillus,结合16S rRNA基因序列分析鉴定菌株D5-8为特基拉芽胞杆菌Bacillus tequilensis;其最适培养基：牛肉浸膏8 g,酵母浸粉5 g,麦芽糖10 g,蒸馏水1000 mL;最佳的发酵条件为温度28℃、初始pH 6、转速210 r/min、装液量30 mL、接种量2%、培养时间60 h、光照时间4 h;盆栽试验结果表明,发酵条件优化后D5-8发酵液（10⁸cfu/mL）对烟草黑胫病的保护防效和治疗防效可达66.89%和53.72%,均高于原始发酵液的61.84%和51.83%。研究结果为特基拉芽胞杆菌D5-8生防菌剂的开发及其对烟草黑胫病的防治应用提供了理论依据。     

撕裂蜡孔菌对烟草黑胫病的防治效果及对烤烟生长的影响

为探讨撕裂蜡孔菌Ceriporia lacerata菌株HG2011对烟草黑胫病的防治效果和对烤烟的促生作用,利用该菌株制备发酵液和固体菌剂,通过拮抗和盆栽试验研究其对烟草黑胫病病原菌烟草疫霉Phytophthora nicotianae生长的抑制作用、对黑胫病的防治效果及对烟苗生长和养分吸收的影响。结果表明:撕裂蜡孔菌菌株HG2011能显著抑制烟草疫霉生长;在带毒平板试验中,含20%发酵液的培养基对烟草疫霉菌丝生长的抑制率达54.50%;在平板对峙试验中,菌株HG2011能使烟草疫霉菌丝扭曲、畸形和空泡化,最终完全侵入并覆盖烟草疫霉菌落。在盆栽试验中,施用撕裂蜡孔菌固体菌剂可降低发病烟苗叶片中的丙二醛含量,并能维持正常的细胞膜透性,使烟苗黑胫病发病率比单独接种烟草疫霉处理显著降低了15.55%～37.77%,且预防效果71.16%优于治疗效果17.98%。在化肥配施高量菌剂处理中烟苗的株高、最大叶面积和生物量均最高,比单施化肥处理显著增加9.08%、36.28%和11.84%;烟苗氮、磷、钾的吸收量比单施化肥处理显著增加11.29%、38.73%和12.79%。表明撕裂蜡孔菌菌株HG2011能有效防治烟草黑胫病,刺激烟苗生长、促进养分吸收,表现出良好的防病促生效应。     

Pseudomonas sp.20#-5防治烟草黑胫病田间试验初报

20^#-5菌株是本实验室从土壤中分离保存的1株假单胞菌,2006年对其发酵液防治烟草黑胫病进行了田间防效试验。结果显示,不同发病时期20^#-5菌发酵液的防治效果在64.39%～79.70%,高于对照药剂,防治效果较好。不同施药时期调查结果表明,发病前施用20^#-5菌发酵液的防治效果要好于发病后施用,说明20^#-5菌发酵液对烟草黑胫病具有有效的预防作用。     

泸州烟草黑胫病病原生理小种组成及对甲霜灵和烯酰吗啉的敏感性

烟草黑胫病(Phytophthora nicotianae)是四川省泸州烟区最为严重的病害之一,为明确泸州烟草黑胫病生理小种组成及其对甲霜灵和烯酰吗啉的敏感性,从泸州主产烟区采集样品,分离得到24株烟草黑胫病菌株;通过鉴别寄主法结合TTZ生化反应,发现24株菌株中,0号生理小种15株,占62.50%,1号生理小种9株,占37.50%。对24株菌株进行甲霜灵和烯酰吗啉的药剂敏感性检测,发现甲霜灵对24株菌株的EC50范围在0.052~0.513μg/mL之间,敏感菌株和中抗菌株分别占41.67%和58.33%,未发现高抗菌株;烯酰吗啉对24株菌株的EC50范围在0.110~1.615μg/mL之间,敏感菌株和中抗菌株分别占37.50%和62.50%,未发现高抗菌株。上述结果表明,泸州烟区烟草黑胫病菌同时存在0号生理小种和1号小种,而且该烟区的黑胫病菌已对甲霜灵和烯酰吗啉产生不同程度的抗性。     

烟草黑胫病防治的关键施药技术研究

为了有效控制烟草黑胫病的发生和为害,选用目前市面上防治效果较好的农药68%精甲霜·锰锌对烟草黑胫病进行防治试验。结果表明,每667 m~2采用精甲霜·锰锌100 g拌底肥窝施和发病初期用68%精甲霜·锰锌1 000倍液对准烟草茎基部灌根,对黑胫病的防治效果为100%。     

贵州省烟草黑胫病菌对烯酰吗啉的敏感性

为了明确烟草黑胫病菌对烯酰吗啉的敏感性现状,从贵州省各地采集了47株烟草黑胫病菌株,分别测定了菌丝生长和游动孢子囊形成对烯酰吗啉的敏感性.结果表明,烯酰吗啉对烟草黑胫病菌菌丝生长和孢子囊形成的抑制基本一致,对菌丝EC50范围为0.792 6～4.232 7μg/mL,平均为2.476 8μg/mL;对孢子囊的抑制作用略强,EC50范围为0.394 5～4.625 3μg/mL,平均值为2.040 0 μg/mL.统计分析表明,烟草黑胫病菌47个菌株菌丝生长和孢子萌发对烯酰吗啉的敏感性符合正态分布,不同菌株对烯酰吗啉的敏感性有显著差异,而且菌株对烯酰吗啉的敏感性与其地理来源有一定的相关性.     

三种生防芽孢杆菌对5种烟田常用化合物的敏感性

芽孢杆菌是植物病害生物防治中重要的生防细菌。采用浊度测定法研究了生防细菌——枯草芽孢杆菌、解淀粉芽孢杆菌及多粘芽孢杆菌对烤烟专用复合肥[m(N)∶m(P2O5)∶m(K2O)=10∶10∶25]、生石灰、甲霜灵、代森锰锌及硫酸链霉素5种烟草生产上常用化学品的敏感性。结果表明:甲霜灵对3种芽孢杆菌的生长无影响,而烤烟专用复合肥和生石灰对3种生防细菌均有一定的促生长作用;而代森锰锌和硫酸链霉素能显著抑制枯草芽孢杆菌、解淀粉芽孢杆菌和多粘芽孢杆菌的生长,其EC50值分别为14.15、11.88、11.30μg/mL和5.20、5.69和2.64μg/mL。     

Resistance selection and mechanisms of oriental tobacco budworm (Helicoverpa assulta Guenee) to indoxacarb

The oriental tobacco budworm, Helicoverpa assulta, is one of the most destructive pests for numerous commercial crops, and these organisms are responsible for enormous economic losses in Chinese agriculture. Insect larvae often feed within host plant fruits, providing protection from many currently used insecticides and making field control of H. assulta very difficult. Owing to its novel mode of action, high insecticidal activity, and low mammalian toxicity, the nonsystemic insecticide indoxacarb has been considered a promising alternative for the control of lepidopterous pests of agricultural significance. Indoxacarb evidences an elevated insecticidal activity against H. assulta. After 13 generations of selection with indoxacarb and bifenthrin insecticides under laboratory conditions, the LC₅₀ of these compounds for H. assulta increased by 4.19-fold and 10.67-fold, respectively. The synergists diethyl maleate (DEM) and triphenyl phosphate (TPP) increased indoxacarb toxicity by 2.76-fold and 4.10-fold in resistant strains and, comparatively, 1.58-fold and 1.75-fold in susceptible strains, suggesting that carboxylesterase (CarE) and glutathione-S-transferases (GSTs) may be involved in the development of indoxacarb resistance in H. assulta. Activity and kinetic parameters observed in detoxification enzymes further demonstrated that the enhanced activity of CarE and GSTs may be critical in development of indoxacarb resistance in H. assulta. The data provides a foundation for further study of the indoxacarb resistance mechanism observed in H. assulta and the rational use of indoxacarb as a rotation insecticide with other insecticide classes for the control of H. assulta.     

枯草芽胞杆菌对离体荔枝果实霜疫霉病、炭疽病的防治效果

为了进一步明确从土壤中筛选得到的拮抗细菌OR-1、OR-2、OR-3、ON-6的分类地位和生防效果,采用形态学观察、理化特性结合分子生物学方法,鉴定这4株细菌皆为枯草芽胞杆菌Bacillussubitilis.拮抗菌对荔枝霜疫霉菌、炭疽菌菌丝生长均具有明显抑制作用,ON-6菌株的发酵液对菌丝生长的抑制率最高,分别为92.34％和70.36％,其次是OR-1菌株.拮抗菌处理组褐变指数均小于对照组以及杀菌剂处理组,且差异达显著水平(P＜0.05),OR-1菌株防褐变效果最好.常温下,拮抗菌发酵液对离体鲜果病害防治效果优于对照组和杀菌剂处理组,其中ON-6菌株的防治效果最佳;4℃低温储藏处理40天后,拮抗菌发酵液处理组的防治效果高于对照和杀菌剂处理组,且差异显著(P＜0.05),以ON-6和ON-3菌株的防治效果最佳.表明拮抗菌发酵液对荔枝鲜果上的霜疫霉菌和炭疽病菌有一定的抑制作用.     

寡雄腐霉发酵液对烤烟生长的影响及对烟草黑胫病的防治作用

为了探索对烟草黑胫病高效、稳定的生防制剂,利用寡雄腐霉工程菌株制备发酵液,采用温室盆栽试验研究该发酵液对烟苗生长的影响和对烟草黑胫病的防治作用。寡雄腐霉发酵液可提高正常烟苗生物量达22.6%,促进烟苗氮、磷、钾吸收;带病烟苗的发病率和病情指数分别降低了61.6%和64.2%,相对防治效果达64.2%;施用发酵液明显降低带病烟苗的细胞膜透性和丙二醛含量,提高烟叶多酚氧化酶、超氧化物歧化酶和苯丙氨酸解氨酶活性。说明寡雄腐霉发酵液可诱导烟株的防御作用,并提高其抗病能力和促生作用。     

Effects of a lectin-like protein isolated from Acacia farnesiana seeds on phytopathogenic bacterial strains and root-knot nematode

Acacia farnesiana lectin-like protein (AFAL) showed bacterioestatic effects against Xanthomonas axonopodis pv. passiflorae (Gram-negative) and Clavibacter michiganensis michiganensis (Gram-positive), with the latter being more sensitive. This effect is probably due to the ability of AFAL to interact with the bacterial cell wall where we observed that AFAL induced macroscopic change. The maximum bacterial growth inhibition was approximately 78% when incubated with Gram-negative strains, and as high as 92% percent for the Gram-positive one. The antibacterial effect of flavonoids (rutin, quercetin and morin) was also observed using low concentrations against both bacterial strains. Prior incubation of both with AFAL at high concentrations increases the inhibitory effect of flavonoids on bacterial growth. The potential use of AFAL as a control agent against the root-knot nematode Meloidogyne incognita was investigated as well, showing anti-nematode properties involving both egg hatching and motility. In the juvenile second-stage, AFAL showed reduction in larval mobility when measured against a control group. The results suggest that AFAL is effective against M. incognita and could be used as a component of integrated pest management programs. These data also suggest that lectins probably play a role in plant defense not only against invertebrate phytopathogens, herbivores and fungi but also against bacteria.     

Molecular analysis of resistance in a deltamethrin-resistant strain of Musca domestica from China

We investigated the molecular basis of resistance in a strain of house fly (BJD) from Beijing, China. This strain showed 567-fold resistance to commonly used deltamethrin. Flies were 64-fold resistant to deltamethrin synergized by piperonyl butoxide (PBO). The 5′-flanking sequence of the cytochrome P450 gene CYP6D1 in BJD strain had a 15-bp insert as in the LPR strain. Two mutations (kdr, super-kdr) in the voltage sensitive sodium channel (VSSC) were also detected in the BJD strain. Our results showed that a combination of resistance alleles for CYP6D1 and VSSC existed in deltamethrin resistant house flies in China.     

苏云金杆菌亚致死浓度对美国白蛾及其寄生蜂生长发育的影响

为阐明苏云金杆菌Bacillus thuringiensis(Bt)与白蛾周氏啮小蜂Chouioia cunea Yang的协同作用,分别用103~108孢子/m L的6个Bt浓度处理美国白蛾Hyphantria cunea 2龄幼虫后,通过室内试验测定了Bt对其致死、亚致死效应,及亚致死浓度Bt对寄生蜂子代出蜂的影响。结果表明,浓度106孢子/m L处理组美国白蛾均未能化蛹;而106孢子/m L处理组均能化蛹并羽化,且幼虫发育历期随Bt浓度的增加而延长,平均蛹重在74.92~92.18 mg之间,均高于对照的68.66 mg,但蛹重不呈线性规律变化。当Bt对美国白蛾平均校正死亡率低于17.17%时,幼虫发育历期延迟不明显,但平均蛹重为101.11~124.34 mg,均高于对照;接蜂后能够正常出蜂,出蜂率为33.13%~51.25%,每蛹出蜂量为117.43~143.42头,均显著高于对照。表明用平均校正死亡率低于20%的Bt处理对白蛾周氏啮小蜂的寄生有利,2种生防措施协同作用存在增效潜力。     

溴氰虫酰胺和氯虫苯甲酰胺对三种鳞翅目害虫的毒力作用比较

为比较溴氰虫酰胺和氯虫苯甲酰胺对甜菜夜蛾、玉米螟和小地老虎的作用差异，采用浸卵法、浸叶法、饲喂毒营养液法等测定了溴氰虫酰胺和氯虫苯甲酰胺对3种害虫的卵、3龄幼虫和成虫的毒力，以及对其3龄幼虫的拒食活性。结果表明，溴氰虫酰胺和氯虫苯甲酰胺均无杀卵作用，但能降低初孵幼虫存活率。溴氰虫酰胺对甜菜夜蛾、玉米螟和小地老虎3龄幼虫的LC₅₀值分别为0.11、0.05和0.13 mg/L，相对毒力分别是氯虫苯甲酰胺的2.8、2.7和5.1倍。溴氰虫酰胺对3种害虫成虫的LC₅₀值分别为0.11、0.09和0.22 mg/L，相对毒力分别是氯虫苯甲酰胺的3.9、3.8和2.7倍。溴氰虫酰胺在亚致死浓度0.10、0.05和0.14 mg/L的剂量下分别对甜菜夜蛾、玉米螟和小地老虎3龄幼虫表现出较强的拒食作用，处理48 h后达到最高，拒食率分别为85.58%、81.91%和86.11%。药剂处理试虫虫体皱缩，体节缩短，体重增加率明显低于对照处理。表明溴氰虫酰胺对3种鳞翅目害虫的毒力作用比氯虫苯甲酰胺好，可作为氯虫苯甲酰胺的替代药品。     

椰心叶甲人工饲料的研制及应用效果评价

为了获得入侵害虫椰心叶甲Brontispa longissima(Gestro)的人工饲料,在分析其天然寄主叶片水分和营养成分基础上,借鉴其它鞘翅目昆虫人工饲料配方,进行椰心叶甲人工饲料的配制和筛选,并评价人工饲料饲养的椰心叶甲对其寄生蜂椰甲截脉姬小蜂Asecodes hispinarum和椰心叶甲啮小蜂Tetrastichus brontispae的适合性。人工饲料配方的成分:蔗糖4%、椰子粉2%、大豆粉2%、椰树叶粉10%、酵母2%、维生素E 0.3%、抗坏血酸0.2%、对羟基苯甲酸甲酯0.2%、琼脂4%、链霉素0.03%和水75%。用所配制的人工饲料饲养椰心叶甲初孵幼虫,其蛹成活率可达36%,蛹羽化率与对照无显著差异,均在90%以上;雌成虫占52.4%,与对照无显著差异。表明人工饲料饲养所得的椰心叶甲可用于其幼虫寄生蜂椰甲截脉姬小蜂和蛹寄生蜂椰心叶甲啮小蜂的扩繁。