桑天牛成虫交配后血淋巴和生殖系统内可溶性总糖和蛋白质含量的变化

以生理状况相似的未交配桑天牛Apriona germari Hope雌、雄成虫为供试昆虫, 观察、记录交配活动;采集血淋巴,解剖生殖系统并绘图;用蒽酮比色法和福林-酚法检测交配前、后成虫血淋巴和生殖系统内可溶性总糖和蛋白质含量变化。结果表明：交配后1 h,桑天牛雄虫血淋巴内的可溶性总糖含量增加21.38%,蛋白质含量降低22.66%;雄虫生殖系统内的可溶性总糖和蛋白质含量都明显升高;雄性附腺作为某些特异性蛋白质的合成场所其内的可溶性总糖和蛋白质含量分别降低81.76%和63.76%,雌虫血淋巴和卵巢内的可溶性总糖和蛋白质含量都升高。交配后, 雄虫发生陪伴行为最短历时为4 h, 可能是其重要的生殖策略之一。     

微孢子虫和狄斯瓦螨分别侵染后的意蜂血淋巴蛋白质含量变化

微孢子虫Nosema apis和狄斯瓦螨微孢子虫Nosema apis和狄斯瓦螨 Varroa destructor (Acari: Varroidae)均为危害意蜂Apis mellifera的重要寄生虫,该文对其危害后意蜂血淋巴蛋白质含量的变化进行了研究。用考马斯亮蓝法测定了意蜂受侵染后血淋巴的蛋白质总量,并用高压超薄层等电点聚焦法进行血淋巴蛋白质分类。结果显示,病蜂血淋巴蛋白质总量,在人工感染微孢子虫后1～10天呈微孢子虫Nosema apis和狄斯瓦螨 Varroa destructor (Acari: Varroidae)均为危害意蜂Apis mellifera的重要寄生虫,该文对其危害后意蜂血淋巴蛋白质含量的变化进行了研究。用考马斯亮蓝法测定了意蜂受侵染后血淋巴的蛋白质总量,并用高压超薄层等电点聚焦法进行血淋巴蛋白质分类。结果显示,病蜂血淋巴蛋白质总量,在人工感染微孢子虫后1～10天呈上升趋势,然后逐渐下降,感染后12～27天保持在感染前意蜂血淋巴总蛋白质含量水平以下。螨侵染后意蜂血淋巴蛋白质含量明显增高,与健康意蜂相比差异极显著。高压超薄层等电点聚焦分析表明：狄斯瓦螨自然侵染意蜂后,意蜂血淋巴蛋白质组分与健康对照组相比发生了明显改变。这些结果提示,意蜂对于微孢子虫或狄斯瓦螨的侵染产生了一定的免疫反应。     

唐古特瑞香提取物对菜粉蝶幼虫的毒杀作用

生测表明,唐古特瑞香Daphne tangutica的甲醇抽提物具有明显的拒食和胃毒活性,活性跟踪得到流分9。流分9导致菜粉蝶Pieris rapae幼虫体小,中肠组织细胞受到破坏,脂肪体呈消融状;对试虫生理指标的影响主要表现在降低血淋巴蛋白质含量和抑制中肠酯酶活力。     

刀豆氨酸对亚洲玉米螟生长发育的影响

以不同浓度刀豆氨酸注射和饲喂亚洲玉米螟`Ostrinia furnacalis`` 5龄幼虫,影响其以后各虫态的生长发育,有引起畸蛹和畸蛾的明显作用,发育成的雌蛾性腺发育受到抑制,卵管变短,卵细胞数减少。不同交配实验表明对发育成的雄蛾影响更为严重,能使其不育。供试幼虫48小时后取血样,表明血淋巴蛋白质区带与对照组不同,认为刀豆氨酸对亚洲玉米螟的上述作用可能与刀豆氨酸取代精氨酸参入蛋白质的结果有关。     

刀豆氨酸对亚洲玉米螟的生理生化效应

本文报道亚洲玉米螟Ostrinia furnacalis五龄幼虫注射0.25-1.0毫克/克刀豆氨酸48小时后有关生理生化的变化,其中血淋巴蛋白质电泳区带数目减少,浓度降低,而游离氨基酸总量明显增加,其中脯氨酸和谷氨酸的变化最为明显。虫体酸性磷酸酯酶的活性显著升高,而精氨酸酶的活性却有所降低。刀豆氨酸可引起虫体内尿素含昆下降,而对尿酸含量影响不大。经刀豆氨酸处理后发育成长的成虫其雌虫卵巢和雄虫附腺的蛋白质合成均显著下降,并对上述结果做了简要的讨论。     

块状耳霉的分离、鉴定、培养和寄主范围

1981年自感染真菌的扁豆蚜(Aphis craccivora Koch)虫尸上分离到一株虫毒菌,经鉴定是耳霉属的块状耳霉(couidiobolus thromboides Drechsler)。该菌易于分离和培养,在培养基上可形成大量休眠孢子,休眠孢子容易萌发,六天中萌发率达53%。经试验该种虫霉可以感染多种蚜虫,是一种有希望制成杀蚜菌剂的真菌。     

蝗虫微孢子虫Nosema locustae对红胫戟纹蝗Dociostaurus kraussi kraussi致病性及呼吸代谢的影响

【目的】研究蝗虫微孢子虫Nosema locustae对红胫戟纹蝗Dociostaurus kraussi kraussi致病性及呼吸代谢的影响,为筛选蝗虫微孢子虫适宜的新疆本地的活体增殖寄主提供实验依据。【方法】采用逐头口服法感染红胫戟纹蝗,以镜检法检测蝗虫感染情况,并用呼吸仪测量试虫的呼吸代谢。【结果】在6.5×105个孢子/头的感染剂量下,红胫戟纹蝗的感染率和死亡率分别为41.70%和72.22%。蝗虫微孢子虫的感染剂量与其毒力高度相关(r2=0.961),LD50为1.7088×104个孢子/头;随孢子浓度增加和感染时间延长,试虫的CO2释放率显著降低(P<0.05),但耗氧率没有显著变化。【结论】红胫戟纹蝗被微孢子虫感染后表现出典型症状,高剂量组感染下死亡率达到70%以上,由此推断红胫戟纹蝗可作为微孢子虫在新疆本地的潜在增殖寄主。     

鹿儿岛被毛孢分生孢子胞外黏液蛋白提取与鉴定

【背景】分生孢子表层黏液在虫生真菌与昆虫的互作中扮演着重要角色。鹿儿岛被毛孢是寄主专一、孢子表面具黏液层的虫生真菌,但其胞外黏液成分尚不清楚。【目的】优化鹿儿岛被毛孢分生孢子黏液蛋白的提取方法,对黏液蛋白组分进行质谱鉴定,探究分生孢子胞外黏液的蛋白成分。【方法】用低浓度还原剂二硫苏糖醇(Dithiothreitol,DTT)对胞外黏液进行温和洗脱并优化提取条件,聚丙烯酰胺凝胶电泳(Sodium Dodecyl Sulfate Polyacrylamide Gel Electrophoresis,SDS-PAGE)检测蛋白提取浓度,用碘化丙啶(Propidium Iodide,PI)染色法检测孢子细胞膜选择通透性,通过肽段酶解、LC-MS/MS数据采集及蛋白质比对分析鉴定蛋白质。【结果】鹿儿岛被毛孢分生孢子黏液蛋白的最佳提取条件:孢子浓度为5×107 cells/mL,DTT浓度为2 mmol/L,4℃提取24 h;黏液蛋白提取后不影响孢子细胞膜通透性。提取的黏液蛋白共鉴定到蛋白质77个,蛋白丰度排前10的蛋白多为分泌性小分子蛋白,包括未注释的蛋白、几丁质酶、脂酶或是与昆虫体壁降解相关的...     

莱氏绿僵菌对斜纹夜蛾的致病力及生理效应

【目的】测定莱氏绿僵菌Metarhizium rileyi Nr5772菌株对斜纹夜蛾Spodoptera litura幼虫及蛹的致病能力,研究莱氏绿僵菌侵染后在寄主体内的发育及对寄主的生理效应,探讨莱氏绿僵菌的致病机制。【方法】采用浸渍法测定莱氏绿僵菌孢子对斜纹夜蛾3-6龄幼虫及蛹的致死中浓度(LC_(50))和致死中时(LT_(50))。采用微量注射法接种莱氏绿僵菌虫菌体,在不同时间后采集斜纹夜蛾幼虫血淋巴,在显微镜下检查虫菌体的数量、形态及寄主血细胞数量,并用酶标仪测定寄主血淋巴酚氧化酶(Phenoloxidase,PO)的活性。【结果】M.rileyi孢子对3龄斜纹夜蛾幼虫毒力最强,10 d后LC_(50)=3.12×10~6个孢子/mL,龄期越大,致病力越低;孢子浓度为5×10~9个/mL时,对3龄幼虫的致死速度最快,LT_(50)=4.55 d,致死速度随龄期的增大和浓度的降低逐渐减缓;M.rileyi孢子对蛹的致病力远低于对幼虫的致病力。注射接种虫菌体后,64 h内,虫菌体数量在寄主血腔中以幂函数的形式增长,寄主的血细胞数量没有明显的变化;在侵染初期(接种后44 h内),血淋巴PO活性正常;在侵染后期,虫菌体数量不再增加(55-64 h后),逐渐转化为菌丝体,并快速杀死寄主,PO活性受到抑制。【结论】莱氏绿僵菌Nr5772菌株对斜纹夜蛾幼虫有较强的致病力,应在害虫低龄期应用;莱氏绿僵菌在侵染初期对寄主血细胞和血淋巴PO无影响,后期则完全抑制PO活性。     

短时高温暴露下东亚飞蝗雌虫血淋巴蛋白表达分析及质谱鉴定

为研究短时高温对东亚飞蝗Locusta migratoria manilensis Meyen血淋巴蛋白的影响,采用Bradford法、SDSPAGE电泳和质谱等方法,对东亚飞蝗雌虫血淋巴样品进行检测。结果表明:短时高温对血淋巴蛋白含量有显著影响(P0.01),36℃-42℃范围内,随温度升高,血淋巴蛋白浓度亦升高,其中39℃、42℃处理组与对照组差异显著(P0.01);短时高温对血淋巴蛋白种类存在一定影响,对照组雌虫血淋巴中存在11种蛋白,高温处理后,4种蛋白含量逐渐增加,6种蛋白含量没有明显变化,1种蛋白消失;经质谱检测,鉴定了5种蛋白,分别为载脂蛋白前体、酚氧化酶原、2个储存蛋白和19 kDa血淋巴蛋白,另外6条蛋白未被鉴定。推测载脂蛋白前体、酚氧化酶原、储存蛋白在东亚飞蝗应对高温胁迫过程中发挥重要作用。     

Inhibition of Beauveria bassiana Proteases and Fungal Development by Inducible Protease Inhibitors in the Haemolymph of Galleria mellonella Larvae

Injection of zymosan or dead yeast cells enhanced the inhibitory activity against exocellular Beauveria bassiana proteases in the cell - free haemolymph of Galleria mellonella larvae . Pre - injected larvae exhibited no decreased mortality after subsequent injection with living B. bassiana blastospores but survived for a prolonged time before death . Increased levels of protease inhibitors in the haemolymph were also observed after injection of B. bassiana proteases . In contrast , no enhanced inhibitory activity against B. bassiana proteases was detected in infected larvae when mycosis was initiated with conidia which enabled the fungus to invade host larvae through the integument in a natural manner . B. bassiana proteases were not completely inhibited by the addition of cell - free haemolymph . Protease inhibitors obtained after heat and trichloroacetic acid precipitation of cell - free haemolymph were added to the protein medium of B. bassiana to study the effect on its growth in vitro. Enriched fractions from pre - injected larvae delayed fungal growth in comparison with fractions from untreated larvae , suggesting that delayed mortality of immunized G. mellonella larvae infected with B. bassiana is due to enhanced levels of protease inhibitors . A non - virulent form of the same strain exhibited reduced capacity to release proteases in vitro. The results strongly suggest that the capacity of insects to release inhibitors against fungal proteases influences their susceptibility against entomopathogenic fungi .     

Comparative analysis of the production of insecticidal and melanizing macromolecules by strains of Beauveria spp.: in vivo studies

Eleven strains of Beauveria bassiana, and a further five species of Beauveria sp., were tested by injection of 8x10(2) conidia into the haemocoel of the larvae of the lepidopteran Galleria mellonella with the aim of analysing their toxin producing activity in vivo. Although the virulent strains killed 100% of the insects at slightly different rates (4-6 days) there were significant differences in the pattern and intensity of host melanization caused by isolates. The majority of the isolates of Beauveria spp. induced a fast and intense melanization of the cuticle of the integument and of tracheal wall, which followed one of three patterns. Another small group of two B. bassiana strains, isolated from Ostrinia nubilalis, induced very weak or no melanization. Strains 618 and 101 of B. bassiana, were selected as models of "melanizing" and "non-melanizing" strains, respectively. Ultrastructural alterations of cells of hypodermal and tracheal epithelium and of haemocytes, assumed to be at least partially caused by fungal toxins, were revealed in larvae infected by both isolates. However, their effects on the fine structure of the hypodermis were different. Injection of sera obtained from haemolymph of insects infected with B. bassiana 618 showed that they have insecticidal, melanizing, and cytotoxic effects similar to those occurring during mycosis. Chromatographic studies and bioassays with fractions prepared from crude serum have allowed a partial identification of the toxic molecules secreted by the fungus in vivo. They are proteinaceous, as shown by protease treatments, thermolabile, negatively charged, and not glycosylated with alpha-d-mannose or alpha-d-glucose. If strain B. bassiana 618 produces melanizing macromolecules which are vivotoxins secreted during the mycosis, the mode of action of isolate 101 is different. Its capacity to kill the host depends on active mycelial development, and on the production of low molecular weight toxins.     

Production of an extracellular protease by Beauveria bassiana in the haemolymph of the silkworm, Bombyx mori

Protease production by Beauveria bassiana in the haemolymph of infected silkworms (Bombyx mori) were investigated by an enzyme-linked immunosorbent assay. Results showed that B. bassiana produced a substantial amount of extracellular protease in the haemolymph of infected silkworms.     

Effects of Beauveria bassiana on embryos of the inland silverside fish (Menidia beryllina).

A chemical toxicity and teratogenicity test was adapted to assess potential adverse effects of a microbial pest control agent on a nontarget fish. Developing embryos of the inland silverside, Menidia beryllina, were exposed to conidiospores of the insect-pathogenic fungus Beauveria bassiana. Embryo rupture and death were observed. Embryo rupture did not always result in death, nor was death always associated with embryo rupture. Adherence of spores to the chorion, followed by germination and penetration by the germ tube, probably caused the embryos to rupture. Statistically significant (P less than or equal to 0.05) responses were observed in tests in which conidiospore concentrations were greater than or equal to 8.3 x 10(4) or less than or equal to 1.5 x 10(6)/ml. Conidiospores treated with a dispersant (biological detergent) showed significantly less binding (P less than or equal to 0.01) to embryos than did untreated spores. Both detergent-treated and heat-killed spores failed to cause significant adverse effects.     

Effects of Beauveria bassiana on embryos of the inland silverside fish (Menidia beryllina).

A chemical toxicity and teratogenicity test was adapted to assess potential adverse effects of a microbial pest control agent on a nontarget fish. Developing embryos of the inland silverside, Menidia beryllina, were exposed to conidiospores of the insect-pathogenic fungus Beauveria bassiana. Embryo rupture and death were observed. Embryo rupture did not always result in death, nor was death always associated with embryo rupture. Adherence of spores to the chorion, followed by germination and penetration by the germ tube, probably caused the embryos to rupture. Statistically significant (P less than or equal to 0.05) responses were observed in tests in which conidiospore concentrations were greater than or equal to 8.3 x 10(4) or less than or equal to 1.5 x 10(6)/ml. Conidiospores treated with a dispersant (biological detergent) showed significantly less binding (P less than or equal to 0.01) to embryos than did untreated spores. Both detergent-treated and heat-killed spores failed to cause significant adverse effects.     

Effects of exogenous nutrients on conidial germination and virulence against the silverleaf whitefly for two hyphomycetes

Exogenous protein and sugar sources were tested for their impact on conidial germination of two silverleaf whitefly (Bemisia argentifolii) pathogens: Beauveria bassiana and Paecilomyces fumosoroseus. In liquid culture, sugars stimulated only 5-27% germination of B. bassiana and < or =11% germination of P. fumosoroseus, whereas, yeast extract or peptone stimulated 95-100% germination. In the absence of additional nutrients, agar alone stimulated approximately 50% germination. Storing spores for different periods of time did not alter their general response to exogenous nutrients. When spores were germinated before being applied to third instar B. argentifolii, mortality was as much as 2.45 times greater and occurred more rapidly than that for fresh spores. For ungerminated conidia, the mean time to death from infection was 5.45 (SE = 0.16) and 4.74 (SE = 0.08) days for application rates of 37 and 144 conidia x mm(-2), respectively. When conidia were germinated before application, infection times dropped to 4.58 (SE = 0.16) and 4.45 (SE = 0.10) days, respectively. A likely explanation for the greater pathogenicity and virulence of germinated over ungerminated B. bassiana conidia is that only a fraction of the spores applied to whitefly nymphs actually germinate on the cuticle. For some specialized applications, such as greenhouse production systems, it may be beneficial to germinate spores immediately prior to application.     

Antagonism between Beauveria bassiana and imidacloprid when combined for Bemisia argentifolii (Homoptera: Aleyrodidae) control

Imidacloprid and the entomopathogenic fungus Beauveria bassiana (Balsamo) Vuillemin are both used to control the whitefly Bemisia argentifolii Bellows & Perring. We tested whether the two control strategies acted additively, synergistically, or antagonistically when combined for whitefly control. We found antagonism in that B. bassiana inhibited the effectiveness of imidacloprid. When B. bassiana was combined with imidacloprid, insect response was either less than or similar to (depending on B. bassiana rates) that when imidacloprid was used alone. Adding imidacloprid to B. bassiana treatments always increased mortality, but the increase was less than additive. Beauveria bassiana spore germination and colony formation were not inhibited by imidacloprid in vitro, and B. bassiana did not adsorb or degrade imidacloprid in a tank mix. We hypothesize that B. bassiana caused a behavioral response that reduced insect feeding and uptake of imidacloprid.     

Impact of two treatments of a formulation of Beauveria bassiana (Deuteromycota: Hyphomycetes) conidia on Varroa mites (Acari: Varroidae) and on honeybee (Hymenoptera: Apidae) colony health

Bee colonies in southern France were treated with conidia (asexual spores) from two strains of Beauveria bassiana, an entomopathogenic fungus. One strain was commercial (GHA) and the other had been isolated from Varroa mites in the region (Bb05002). Objectives were to evaluate treatment effect on colony weight, adult bee mass, capped brood, and on Varroa fall onto sticky boards. Treatments included conidia formulated with either carnauba or candelilla wax powder, candelilla wax powder alone, or control; in two treatment groups formulation was applied a second time after one week. Treatment did not affect colony health. Colonies treated twice with Bb05002 conidia and carnauba wax powder had significantly higher mite fall compared to colonies treated with blank candelilla wax powder. The proportion of fallen mites that were infected in both conidia treatments was higher than controls for 18 days after the second treatment. The number of fungal propagules on the bees themselves remained elevated for about 14 days after the second treatment. These results were compared to published results from previous experiments with regard to infection duration.     

琅琊山虫生真菌物种多样性和季节变化

在皖东琅琊山自然保护区设置的20个样方中共采集672份虫生真菌标本,分属于4科8属20种.优势种依次为球孢白僵菌、粉拟青霉、环链拟青霉、布氏白僵菌和玫烟色拟青霉,其中球孢白僵菌占绝对优势,相对多度高达73.8%.选用物种多度、Shannon-Wiener指数H′和Pielou均匀度E对琅琊山虫生真菌物种多样性和季节分布进行研究发现,虫生真菌数量丰富但物种多样性指数较低;夏季虫生真菌菌株最为丰富,均匀度相对较低,随着气温及降雨量的下降,菌株数量急剧下降,均匀度相对较高;不同季节白僵菌属的数量波动大,但拟青霉属波动较小.     

球孢白僵菌对烟蚜茧蜂生命参数及控害效果的影响

在室内评价了球孢白僵菌对烟蚜茧蜂生命参数及控害效果的影响。分别在烟蚜茧蜂寄生桃蚜后不同时间进行高剂量(1900孢子/mm2)接菌,检测蚜虫感病率和寄生蜂形成的僵蚜率及僵蚜出蜂率。结果表明,球孢白僵菌对僵蚜率和僵蚜出蜂率的影响随接菌时间不同而变化。在烟蚜茧蜂寄生前1d、寄生当天和寄生后3d接菌,蚜虫感病率分别为59.6%、56.2%和34.8%;与对照相比,僵蚜率分别下降94%、59%和47%,僵蚜出蜂率分别减少83%、54%和49%。在寄生后5d或7d接菌,僵蚜率和僵蚜出蜂率不受明显影响,但蚜虫感病率降低到8.2%以下。对蚜尸内白僵菌菌体含量检测表明,随着烟蚜茧蜂寄生后接菌时间的推移,菌体数量迅速下降。寄生蜂寄生后5d或7d接菌,蚜尸内几乎检测不到菌体。直接喷雾接菌烟蚜茧蜂,成蜂寿命缩短4d左右,且81.8%的蜂尸受白僵菌感染。接菌后的寄生蜂对蚜虫寄生率几乎无影响,但寄生蜂在蚜虫体内的存活时间缩短了27.8%。