The use of entomogenous fungi for pest control and the role of toxins in pathogenesis

Fungi have been suggested as agents for the biological control of insects for over a century, but their use remains extremely limited. This paper examines the biology of entomogenous fungi, highlighting their need for moisture, and reviews studies on the Entomophthoraceae, Beauveria bassiana, Beauveria brongniartii, Metarhizium anisopliae and Verticillium lecanii. It is suggested that current studies should be directed towards the control of pests living in humid micro-climates and that moisture-retaining formulations should be developed to reduce the reliance of fungi on moisture. Many entomogenous fungi produce insecticidal toxins in submerged culture and these compounds are of interest as a source of new toxophores. The most extensively studied toxins are the cyclic depsipeptides from M. anisopliae, termed destruxins, and their possible role in pathogenesis is discussed. Beauveria bassiana also produces the cyclic peptides termed beauvericin, beauverolides, and bassianolide. Toxins produced by V. lecanii, and the Entomophthoraceae are also described.     

Growth and pathogenicity of isolates of the fungus Metarhizium anisopliae against the parasitic mite, Psoroptes ovis: effects of temperature and formulation

Psoroptes mites (Acari: Psoroptidae) are important ectoparasites of mammals, and are of particular economic significance as the agents of mange in sheep. To be effective against mites, putative fungal biocontrol agents must be able to operate at the relatively high temperatures and humidities found at the sheep skin surface. To consider this, the growth rates of different isolates of the entomopathogenic fungus Metarhizium anisopliae (Metschnikoff) Sorokin (Deuteromycotina: Hyphomycetes) were compared and the pathogenicity of these isolates against Psoroptes derived from rabbits (Psoroptes ovis Hering, syn P cuniculi) were evaluated at temperatures between 28 degrees C and 40 degrees C, and when formulated in either Tween 80 or silicone oil. For this study four multi-conidia, arthropod-derived, isolates of M anisopliae were used: from the USA, France, Denmark and Brazil. One single-conidia culture derived from the US isolate was also included in the investigation. Fungal growth was higher at the lower temperatures and none of the isolates grew at 40 degrees C. The growth of the US and single-conidia isolate declined markedly with temperature. In contrast, the Danish, French and Brazilian isolates grew almost as well at 32 degrees C and 35 degrees C as at 28 degrees C and 30 degrees C. The French and Brazilian isolates showed some growth at 37.5 degrees C but the Danish and US isolates did not. The number of fatal infections which resulted from exposure of mites to the fungal isolates was also strongly influenced by temperature. At 30 degrees C all isolates gave between 70 and 90% infection. The number of infections declined with increasing temperature and no infections were seen at 40 degrees C. However, the French and Danish isolates of M anisopliae gave higher numbers of infections than the other isolates at elevated temperatures. When formulated in silicone oil, significantly higher levels of infection were obtained than when formulated in Tween 80, even at the relatively high temperature of 37.5 degrees C. It is suggested that high-temperature adapted isolates of M anisopliae formulated in silicone oil offer good candidates as control agents under the conditions found at the sheep skin surface.     

温度对绿僵菌LA菌株孢子萌发、酶活力及毒力的影响

为明确温度对绿僵菌LA菌株生物学特性和毒力的影响,采用微室萌发、酶标仪测定A值以及体外涂抹法,对不同温度下LA菌株孢子萌发率、胞外蛋白酶、几丁质酶活力以及对东亚飞蝗的毒力进行了测定,结果表明:LA菌株在30℃下孢子萌发速率最快,对东亚飞蝗的致死速率也最快;38℃下培养,孢子萌发率开始受到限制,40℃下培养孢子的萌发率接近于0;高温处理转至适温培养,对孢子萌发速率产生一定影响,48℃下处理不同时间,孢子萌发速率、萌发率均显著降低,转至30℃下培养24h仅有10.6%～20.9%的孢子可萌发;LA胞外蛋白酶活力培养至第6天达到最高,为31.37U/mL,不同温度下活力测定表明37℃下胞外蛋白酶活力最高,几丁质酶在培养至第5天酶活力达到最高,为34.62U/mL,最高酶活温度为50℃。     

Characteristics of Beet Yellows Closterovirus Transmission to Sugar Beets by Aphis fabae

ABSTRACT Laboratory studies were conducted on the characteristics of beet yellows closterovirus (BYV) transmission to sugar beets by the bean aphid, Aphis fabae. The relative transmission efficiency and the retention, acquisition, and inoculation thresholds were evaluated using clonal aphid colonies. Transmission efficiency was studied between different apterous aphid species (A. fabae and Myzus persicae ), between alate and apterous morphs of A. fabae and M. persicae, and among five clonal aphid colonies of A. fabae. Results indicated, on average, that apterous M. persicae transmitted twice as efficiently as apterous A. fabae (60 and 34%, respectively), apterous forms of A. fabae were equally efficient in transmitting BYV as compared with alate A. fabae (28 and 29%, respectively), and the five clonal colonies of A. fabae ranged from 28 to 40% transmission efficiency with an average of 34%. The maximum retention time recorded for BYV by A. fabae was between 24 and 48 h. After a 1-h acquisition access, a transmission efficiency of 10% was obtained, with maximum transmission efficiency (the highest average percent transmission that is statistically different from all other previous averages) of 39% reached after 6 h of acquisition access. Inoculation thresholds occurred within 1 h, with maximum transmission efficiency (42%) reached after a 6-h inoculation access. Using an electronic monitoring system, a range of 9 to 38 min (20.5 mean) was recorded for the stylets of A. fabae to reach the phloem sieve elements. This was consistent with our threshold of a 20-min acquisition access recorded through direct observation.     

松墨天牛病原菌及其致病性研究

对华东地区松材线虫病疫区和松墨天牛主要发生的13个地区随机抽样调查,共获松墨天牛病原真菌9属12种,病原细菌2属2种。其中,我国首次报道的松墨天牛病原菌新种类有：金龟子绿僵菌小孢变种Metarhizium anisopliae var.anisopliae、粉拟青霉Paecilomyces farinosus、尖镰孢Fusarium oxysporum、蜡蚧轮枝霉Verticillum lecanii、青霉Penicillium spp.、黄曲霉Aspergillus flavus、曲霉Aspergillus spp.、木霉Trichoderma spp.和粘质沙雷氏菌Serratia marcescens。经致病性测定表明,球孢白僵菌、金龟子绿僵菌小孢变种、粉拟青霉对松墨天牛有较强的致病力,处理后16d的致死率分别为100%、100%和70%。     

Effect of temperature on conidial germination, mycelial growth and aggressiveness of the defoliating and nondefoliating pathotypes of Verticillium dahliae from cotton in China

Three defoliating (D) pathotype isolates and five nondefoliating (ND) pathotype isolates of Verticillium dahliae from cotton in central China were tested for adaptation to various temperatures in conidial germination on water agar (10?C33°C), mycelial growth on potato dextrose agar (10?C33°C) and infection of cotton seedlings (Gossypium hirsutum cvs. ??E Mian 24?? and ??Yin Rui 361??) (25?C33°C). Results showed that the D-pathotype isolates adapted better than the ND-pathotype isolates to 30°C for conidial germination and mycelial growth, although the isolates of the two pathotypes had the same optimum temperature of 25°C. Under day/night temperatures of 25/25, 27/27 and 30/25°C for 20 and 25?days, the D-pathotype isolates induced the defoliation syndrome on seedlings of the two upland cotton cultivars, whereas the ND-pathotype isolates did not induce defoliation syndrome. The values for the areas under the disease progress curve (AUDPC) and the vascular discoloration index (VDI) were higher for the D-pathotype isolates than those for the ND-pathotype isolates in each temperature treatment. Under 30/30 or 33/27°C, at least two of the three D-pathotype isolates still had higher AUDPC values and/or VDI values than all the ND-pathotype isolates on E Mian 24. Therefore, the D-pathotype isolates appear more aggressive than the ND-pathotype isolates in infection of cotton. Results also showed that the cotton cultivar Yin Rui 361 was more tolerant than the cotton cultivar E Mian 24 to infection by both pathotypes of V. dahliae. This study suggests that the D-pathotype isolates can well adapt to high temperature and heavily infect cotton under 25?C30°C, and these features might be responsible for the rapid spread of this pathotype in central China.     

影响蜡蚧轮枝菌发酵产孢量和孢子活力的基本因素分析

不同接种浓度、发酵时间、菌株和基质对蜡蚧轮枝菌固体发酵的产孢量都有显著影响，而对孢子活力的影响不明显，萌发率均在90％以上。在5至7天内，随发酵时间的延长产孢量有明显增长；7至10天则变化不大；15天时产孢量和孢子活力均显著降低。随接种孢子浓度每增加1倍，菌株（VLFNL 95－01）发酵的产孢量平均提高15.6%，浓度加大至32倍则提高71.9%。3个菌株比较，VLFNL95－01产孢量明显高得多。不同固体基质发酵的结果有显著差异，产孢量最高的（正交3号配方）比其次的（黄豆饼粉＋玉米碎粒＋磷酸盐）就提高了95.1%。     

球孢白僵菌对桃蚜的毒力测定

球孢白僵菌（Ｂｅａｕｗｅｒｉａｂａｓｓｉａｎａ）的６个菌株（ｂｂｓｇ８７０２、ｂｂｓｇ８７０１、ｂｂ－ｓｇ８８０１、ｂｂｄｎｕ８８０３、ｂｂｄｎ８８０４和ｂｂｄｎ８８０６）对桃蚜（Ｍｙｚｕｓｐｅｒｓｉｃａｅ）的生物测定证明,所测朱对桃蚜具有毒力。运用时间－剂量－死亡率模分析死亡率随时间和剂量的变化趋势,菌株ｂｂｓｇ８７０２对桃蚜的毒力最强,其次是ｂｂｓｇ８８０１、ｂｂｄｎ８８０６、ｂｂｄｎ８８０４     

蜡蚧轮枝菌对月季长管蚜的毒力测定

研究结果表明蜡蚧轮枝菌对月季长管蚜有较强的致病性,孢子浓度2.0×105个/mL时可造成大量月季长管蚜感染死亡,其LT50=（6.62±0.14）d; 而高浓度2.0×108个/mL对月季长管蚜感染致死的LT50=（4.69±0.16）d,第8天的死亡率达93.6%,致死中时随浓度增加而缩短。第5天的致死中浓度为（1.72±0.18）×107个/mL,第8天为（1.53±0.15）×104个/mL,致死中浓度随时间增加而减小。     

Plum pox in north america: identification of aphid vectors and a potential role for fruit in virus spread

ABSTRACT Thirteen aphid species were tested for their ability to transmit Pennsylvania isolates of Plum pox virus (PPV) collected in Columbia (PENN-3), Franklin (PENN-4), and York (PENN-7) Counties, PA. Four species, Aphis fabae, A. spiraecola, Brachycaudus persicae, and Myzus persicae, consistently transmitted PPV in preliminary transmission tests. Two species, Metopolophium dirhodum and Rhopalosiphum padi, were occasional inefficient vectors. Toxoptera citricida, from Florida, also was an effective vector but it does not occur in major stone-fruit-growing states. Species not transmitting PPV in parallel tests included Acyrthosiphon pisum, Aphis glycines, Aulacorthum solani, Macrosiphum euphorbiae, Rhopalosiphum maidis, and Sitobion avenae. When given a 3-day probing access period simultaneously on PPV-infected peach seedlings and healthy peach seedlings, Myzus persicae, Aphis spiraecola, A. fabae, and B. persicae transmitted PPV to 63, 31, 38, and 32% of the healthy peach seedlings, respectively. When given a similar probing period on PPV-infected peach fruit and healthy peach seedlings, the same aphid species transmitted PPV to 50, 35, 0, and 0% of seedlings, respectively. Results support the hypothesis of secondary PPV spread by indigenous aphids in Pennsylvania, and suggest that PPV-infected fruit has the potential to function as a virus source for long-distance dispersal.     

Ribotyping of EntomopathogenicVerticittium lecanii in Japan

To estimate the genetic diversity in 30 isolates ofVerticillium lecanii from aphids, whiteflies, mite and black pine in Japan, including two commercialized strains (Mycotal and Vertalec), DNA polymorphisms in ribosomal DNA of those isolates were analyzed using polymerase chain reaction (PCR). The internal transcribed spacer (ITS) and intergenic spacer (IGS) regions of the nuclear ribosomal RNA gene of each isolate were analyzed by PCR-RFLP (restriction fragment length polymorphism). The size of the PCR product from the ITS region was ~ 580 bp in 27 of the isolates. A 600 bp ITS product was detected in Mycotal and Vertalec. One Japanese isolate produced both the 580 bp and 600 bp products. Enzymatic digestion of the ITS region with Sau3A I,Msp I,Hae III andRsa I revealed RFLPs that consisted of eight haplotypes. Mycotal and Vertalec were specific haplotypes that differed from other isolates. The Japanese isolates had a complex relationship with the original host, but we identified several specific haplotypes common to an aphid origin. Ten distinct IGS haplotypes were detected in the IGS region, some of which were associated with aphid and whitefly origins. These results suggest that the haplotype of rDNA RFLP analysis can be used for studying genetic diversity inV. lecanii.     

Forecasting Sclerotinia Disease on Lettuce: A Predictive Model for Carpogenic Germination of Sclerotinia sclerotiorum Sclerotia

ABSTRACT A predictive model for production of apothecia by carpogenic germination of sclerotia is presented for Sclerotinia sclerotiorum. The model is based on the assumption that a conditioning phase must be completed before a subsequent germination phase can occur. Experiments involving transfer of sclerotia from one temperature regime to another allowed temperature-dependent rates to be derived for conditioning and germination for two S. sclerotiorum isolates. Although the response of each isolate to temperature was slightly different, sclerotia were fully conditioned after 2 to 6 days at 5 degrees C in soil but took up to 80 days at 15 degrees C. Subsequent germination took more than 200 days at 5 degrees C and 33 to 52 days at 20 degrees C. Upper temperature thresholds for conditioning and germination were 20 and 25 degrees C, respectively. A predictive model for production of apothecia derived from these data was successful in simulating the germination of multiple burials of sclerotia in the field when a soil water potential threshold of between -4.0 and -12.25 kilopascals (kPa) was imposed. The use of a germination model as part of a disease forecasting system for Sclerotinia disease in lettuce is discussed.     

球孢白僵菌与几种化学杀虫剂和除草剂的相容性

针对真菌杀虫剂杀虫较慢、致死率较低的缺点,在IPM中常将其与亚致死量的化学农药混用,为此需不断测定其与各种新上市农药的相容性。本研究测定了当前农药市场上销售的7种杀虫剂和3种除草剂对10株球孢白僵菌Beauveria bassiana孢子萌发率、生长速度和产孢量的影响。结果表明,所有供试杀虫剂和除草剂在常规使用浓度下对球孢白僵菌分生孢子萌发都有显著抑制作用,抑制作用随农药浓度的降低而减弱;8%氯氰菊酯（绿色威雷微胶囊剂CM）、30%草除灵（草除灵悬浮剂BN）和6.9%精噁唑禾草灵（精噁唑禾草灵水乳剂FE）在稀释10倍后对孢子萌发无显著抑制作用。所有杀虫剂和除草剂在常规浓度下对菌丝生长均有显著抑制作用,1%甲氨基阿维菌素苯甲酸盐（威克达乳油EB）、40%毒死蜱（毒死蜱乳油CP）、4.5%氯氰菊酯（高效氯氰菊酯乳油BC）和8%氯氰菊酯稀释10倍后的抑制作用较小,8%氯氰菊酯甚至对RCEF0894菌丝生长有促进作用。所有杀虫剂和除草剂对产孢量均有极显著的抑制作用。综合比较,在与供试农药混用时,菌株RCEF0013和RCEF0383的产孢较好,而菌株RCEF0895、RCEF0896和RCEF0897菌丝生长较快。8%氯氰菊酯、30%草除灵和6.9%精噁唑禾草灵与球孢白僵菌的相容性相对较好,是菌药混用可能选择的对象。     

Influence of Nutrient and Environmental Factors on Conidial Germination of Potebniamyces pyri

ABSTRACT Potebniamyces pyri is the causal agent of Phacidiopycnis rot, a postharvest disease of pears. Infection of fruit occurs in the orchard, and symptoms develop during storage. Conidial germination of P. pyri in response to nutrient, temperature, wetness duration, relative humidity (RH), and pH was determined in vitro. Conidia germinated by either budding or developing germ tubes in various concentrations of pear juice solutions. The mode of conidial germination was nutrient-dependent. Low nutrient levels favored budding, whereas high nutrient levels favored germ tube development. Conidia germinated at 0 to 30 degrees C but not at 35 degrees C, with optimum temperature between 20 and 25 degrees C. Wetness durations of 4 to 5 h and 6 to 8 h at optimum temperature were required for budding and developing germ tubes, respectively, and 20 to 24 h of wetness was required to reach germination peaks. Regardless of temperature, conidia germinated primarily by budding in 10% pear juice. Secondary conidia, produced by budding of conidia, initially increased their dimensions and later germinated at 0 to 25 degrees C in the same manner as mother conidia. No germination of secondary conidia occurred at 30 degrees C. Germ tubes from conidia elongated at 0 to 25 degrees C but not at 30 degrees C. No germination occurred at 相似文献   

营养和农药对蜡蚧轮枝菌生长发育的影响

在25℃条件下,测定了4种培养基和8种农药对蜡蚧轮枝菌生长发育的影响。结果表明,马铃薯葡萄糖和查氏培养基最适于菌丝生长和产生分生孢子。所有供试农药对蜡蚧轮枝菌菌丝生长均有显著的抑制作用,其中代森锰锌的抑菌率高达100%,其次为代森锌、百菌清及敌敌畏,再次为氯氰菊酯和阿维柴油,链霉素和氟虫腈的抑菌率最低。     

马铃薯早疫病菌分生孢子萌发条件的研究

为了明确马铃薯早疫病菌分生孢子萌发的条件,研究了温度、湿度、pH、光照、营养条件等对马铃薯早疫病菌分生孢子萌发的影响。结果表明,分生孢子在水滴中于30℃1 h即可萌发,8 h后达到萌发高峰,萌发率为98.4%;萌发的最适温度为30℃;相对湿度85%以上均可萌发,水滴中萌发率最高;pH6~10的条件利于分生孢子萌发;紫外线对分生孢子萌发有抑制作用;无机氮源中的铵态氮和尿素对孢子萌发有抑制作用,有机氮源中半胱氨酸和谷氨酸对孢子萌发有抑制作用;山梨糖不利于分生孢子萌发;光照对分生孢子萌发也有抑制作用;分生孢子致死温度是57℃。     

Stability of the aphid transmission phenotype in cucumber mosaic virus

The stability of the aphid transmission phenotype in seven field isolates of cucumber mosaic virus (CMV) was studied, using aphids Aphis gossypii and Myzus persicae . Field isolates, obtained from four vegetable crops, were propagated in squash and Nicotiana glutinosa , and passaged by either aphid transmission or mechanical transfer. All seven isolates were transmissible by both aphids and this aphid transmission phenotype was stable after 20–24 mechanical passages. Upon further mechanical passaging, one of the seven isolates, CMV-2 A1-MT 60x, lost its transmissibility by Myzus persicae but was still transmissible by Aphis gossypii , although at a reduced rate. Isolates maintained by both aphid transmission and mechanical transfer were transmitted more efficiently by Aphis gossypii than by Myzus persicae . A comparison of the RNA profiles showed no major differences among the CMV isolates before and after mechanical passage.     

氰烯菌酯对禾谷镰孢菌分生孢子萌发及菌丝生长的影响

氰烯菌酯(2-氰基-3-氨基-3-苯基丙烯酸乙酯,JS399-19) 是一种对禾谷镰孢菌Fusarium graminearum具有专化活性的新型杀菌剂。离体条件下,采用菌丝生长速率法测定了该药剂对禾谷镰孢菌抗性菌株和敏感菌株的生长抑制活性;同时采用孢子萌发法测定了其对禾谷镰孢菌分生孢子萌发的影响。结果表明,氰烯菌酯能够强烈地抑制禾谷镰孢菌敏感菌株菌丝的生长,EC50值分布在0.092~0.141 μg/mL之间;并可降低敏感菌株分生孢子的萌发速率,以及影响其萌发的方式,使芽管从分生孢子基部和中间细胞萌发的比率增加;同时氰烯菌酯使敏感菌株分生孢子膨大、畸形,并使其芽管肿胀、扭曲,明显抑制其芽管的伸长生长;但对抗性菌株的抑制作用和致畸作用不明显。     

球孢白僵菌与一些常用市售复合肥料的相容性

为了解5种常用市售复合肥料对球孢白僵菌孢子萌发的影响，调查混有不同浓度肥料的平板培养基上的菌落形成单位和摇瓶培养中的孢子发芽率。结果表明，在2种培养条件下“大洋生态有机肥”（F2）在高、中、低3种剂量下与对照的差异均不显著，“绿营高”（F3）仅在低剂量下与对照无显著差异，而其余剂量都极显著地抑制孢子萌发。其他3种肥料在3种剂量下对孢子萌发都有显著抑制作用。这表明只有F2及低剂量的F3可以与白僵菌分生孢子制剂混合使用。     

Virulence for faba bean and production of ascochitine by Ascochyta fabae

The production of ascochitine by seven isolates of Ascochyta fabae accounted for the toxicity of culture filtrates of the fungus to cells isolated from leaves of Vicia faba. The LD₅₀ value for cells from cultivars that were susceptible, tolerant or resistant to the fungus was similar i.e. 3·0 × 10⁻⁵ m , 3·8 × 10⁻⁵ m and 2·9 × 10⁻⁵ M, respectively. Ascochitine affected neither the germination of seeds nor the growth of mature plants at 5·17 × 10⁻⁴ m but caused necrosis and wilting of plant cuttings at 2·5 × 10⁻⁴ m and 5·10⁻⁴ m . There was no association between virulence of 16 isolates of A. fabae for three cultivars of V.faba and the production of ascochitine in vitro. One isolate produced no ascochitine in vitro and yet was the most virulent for two of three cultivars. The toxin could not be extracted from infected plants.