Relationship of inoculum ofVerticillium dahliae to disease in tomato plants

Summary Tomato (Lycopersicon esculentum Mill.) seedlings were inoculated by dipping roots in suspensions of conidia ofVerticillium dahliae Kleb. There was a linear relationship between inoculum concentration and infection at 21 days after inoculation in steam-disinfested soil. Thereafter the number of infections per plant increased. At higher concentrations data indicated a synergistic interaction between conidia. Synergism was more pronounced and was detected earlier in soil not disinfested before inoculation. When conidia from 3-, 7-, and 14-day-old cultures were used, the greatest response was from 3-day-old cultures. A lower total response and lower rate of response to older inoculum indicated a decrease in aggressiveness of conidia with age. Older plants were less affected by the pathogen when plants inoculated at the fourth and sixth leaf stages with minimum root disturbances were compared. There was also an increase in infection with an increase in the volume of root zone infested. In field experiments infection reduced stand when roots were disturbed at inoculation. With minimum root disturbance yields were reduced without a reduction in stand. Part of thesis to be submitted by the senior author to the University of Stellenbosch in partial fulfilment of the Ph.D. (Agric.) degree.     

Resistance of the termite, <Emphasis Type="Italic">Coptotermes formosanus</Emphasis> Shiraki to <Emphasis Type="Italic">Metarhizium anisopliae</Emphasis> due to grooming

Termites, Coptotermes formosanus, reared individually, were highly susceptible to the entomopathogenic fungus, Metarhizium anisopliae, while termites reared in␣groups were highly resistant. When reared in groups, the termites treated with M.␣anisopliae conidia on the body surface were groomed by their nestmates and more than 80% of the conidia were removed from the cuticle within 3 h. However, there was not a significant reduction in the numbers of conidia on the body surfaces of termites reared individually. For the termites maintained in groups, conidia were found in foreguts, midguts and hindguts, but very few conidia were detected in the guts of termites reared individually. Conidia in the alimentary tracts did not germinate, but some of were alive. As a result, it seems that the removal of foreign bodies, such as fungal conidia, from the␣cuticle is one function of termite mutual grooming behavior and that conidia removed from the cuticle are eliminated through alimentary tracts. This study indicates that mutual grooming behavior is very effective in protecting these termites from M.␣anisopliae infection.     

Effect ofAspergillus parasiticus soil inoculum on invasion of peanut seeds

Environmental control plots adjusted to late season drought and elevated soil temperatures where inoculated at peanut planting with low and high levels of conidia, sclerotia, and mycelium from a brown conidial mutant ofAspergillus parasiticus. Percentage infection of peanut seeds from undamaged pods was greatest for the subplot containing the high sclerotial inoculum (15/cm² soil surface). Sclerotia did not germinate sporogenically and may have invaded seeds through mycelium. In contrast, the mycelial inoculum (colonized peanut seed particles) released large numbers of conidia into soil. Soil conidial populations of brownA. parasiticus from treatments with conidia and mycelium were positively correlated with the incidence of seed infection in undamaged pods. The ratio ofA. flavus to wild-typeA. parasiticus in soil shifted from 7:3 to 1:1 in the uninoculated subplot after instigation of drought, whereas in all subplots treated with brownA. parasiticus, the ratio of the two species became approximately 8:2. Despite high levels of brownA. parasiticus populations in soil, nativeA. flavus often dominated peanut seeds, suggesting that it is a more aggressive species. Sclerotia of wild-typeA. parasiticus formed infrequently on preharvest peanut seeds from insect-damaged pods.     

A larval hunger signal in the bumblebee Bombus terrestris

Larvae of Bombus terrestris, a pollen-storing bumblebee, are dependent on progressive provisioning by workers. We test the hypothesis that larval cuticular chemicals can act as a hunger signal. We first show with a new classical conditioning experiment, using a Y-shaped tube, that workers can be trained to prefer the extracts of normally fed larvae over those of starved larvae. This proves the ability of workers to discriminate between larval extracts. Second, we show in a bioassay that workers also use these perceived differences to feed larvae according to their nutritional status. Larval broods sprayed with the extracts of the starved larvae were fed significantly more than larval broods sprayed with the extracts of normally fed larvae or with the solvent (n-pentane) only. We therefore conclude that B. terrestris larvae signal their need for food via their cuticular chemicals, and discuss the extent to which this form of communication could give larvae some control over their development. Received 2 September 2005; revised 11 April; accepted 24 April 2006.     

A new technique for monoconidial culture of the most aggressive isolate in a given population of Bipolaris sorokiniana, cause of foliar spot blotch in wheat and barley

We developed a new technique for monoconidial culture of the most aggressive isolate in a given population of Bipolaris sorokiniana, to facilitate the evaluation of spot blotch resistance in wheat and barley. Blotched portions of infected barley leaves were placed on a glass slide in a moist chamber for production of conidia by associated fungal hyphae. Conidia were collected separately and grown on water agar discs. Individual water agar discs having conidium growth were inoculated on barley leaves. The conidium producing the earliest symptom with the largest lesion was considered most aggressive. This lesion was incubated in a moist chamber and the conidial offspring were tested for pathogenicity. When a uniform infection was observed, a small piece of the lesion was cut using a sterilized scalpel, surface sterilized with NaOCl, and inoculated in the centre of Petri dishes containing potato dextrose agar medium. The inoculated Petri dishes were incubated at 25 ± 1 °C to yield monoconidial cultures of the most aggressive isolate. Variability in symptom expression caused by the most aggressive isolate of a given population was much less than variability in symptom expression caused by all isolates collectively. The techniques will be useful for plant pathologists and breeders in screening for spot blotch resistance in wheat and barley.     

Susceptibility of Larvae of Galleria mellonella to Infection by Aspergillus fumigatus is Dependent upon Stage of Conidial Germination

The ability of conidia of the human pathogenic fungus Aspergillus fumigatus to kill larvae of the insect Galleria mellonella was investigated. Conidia at different stages of the germination process displayed variations in their virulence as measured using the Galleria infection model. Non-germinating (‘resting’) conidia were avirulent except when an inoculation density of 1 × 10⁷ conidia per insect was used. Conidia that had been induced to commence the germination process by pre-culturing in growth medium for 3 h were capable of killing larvae at densities of 1 × 10⁶ and 1 × 10⁷ per insect. An inoculation density of 1 × 10⁵ conidia per insect remained avirulent. Conidia in the outgrowth phase of germination (characterised as the formation of a germ tube) were the most virulent and were capable of killing 100% of larvae after 5 or 24 h when 1 × 10⁷ or 1 × 10⁶ conidia, that had been allowed to germinate for 24 h, were used. Examination of the response of insect haemocytes to conidia at different stages of the germination process established that haemocytes could engulf non-germinating conidia and those in the early stages of the germination process but that conidia, which had reached the outgrowth stages of germination were not phagocytosed. The results presented here indicate that haemocytes of G. mellonella are capable of phagocytosing A. fumigatus conidia less than 3.0 μm in diameter but that conidia greater than this are too large to be engulfed. The virulence of A. fumigatus in G. mellonella larvae can be ascertained within 60–90 h if infection densities of 1 × 10⁶ or 1 × 10⁷ activated conidia (pre-incubated for 2–3 h) per insect are employed.     

Invasion of the pathogenic fungus Metarhizium anisopliae through the guts of germfree desert locusts,Schistocerca gregaria

Less than 1% of an ingested inoculum of the pathogenic fungus Metarhizium anisopliae was retained for long enough (ca. 24 h) in the gut of the desert locust, Schistocerca gregaria, for germination and penetration to have occurred. The residual inoculum did not initiate an infection in guts of fed conventional or axenic locusts. However, symptoms of mycosis (hyphal bodies in the haemolymph, fungal penetration of the hindgut intima and epithelium, tetanic paralysis) were consistently observed in axenic but not conventional locusts which were starved post-inoculation.It is concluded that the antifungal toxin produced by the gut bacteria defends the desert locust against gut invasion by Metarhizium anisopliae during periods of starvation when the physical defences, prominent in fed insects, are less apparent.     

Influence of Dimilin on a microsporidian infection in the gypsy moth Lymantria dispar (L.) (Lepidoptera: Lymantriidae)

Bioassay studies were conducted to investigate the influence of Dimilin (diflubenzuron), a chitinsynthetase inhibitor used for insecticidal control of the gypsy moth, Lymantria dispar, on the development and viability of a microsporidian pathogen of L. dispar. Before or after an infection with a Nosema species, L. dispar larvae were fed Dimilin in sublethal dosages. Dimilin fed to L. dispar larvae at 0.65 ng/cm² diet surface resulted in a total larval mortality of 53%. Although the microsporidian infection alone did not cause high mortality rates (9%), mortality increased to 96% when L. dispar larvae were inoculated with both Dimilin and Nosema spores. When Dimilin was fed to the larvae 24 h before or 6 days after inoculation with the microsporidium, the number of mature spores produced was significantly reduced. When Dimilin was fed to the larvae 24 h after microsporidian inoculation, the number of spores produced was not significantly reduced. Spores that were produced in larvae after Dimilin had been ingested with the diet were less infectious than spores produced in control larvae; the experimental infection rate decreased from 94% when spores obtained from control larvae were used, to 48 or 10% when spores obtained from larvae fed Dimilin 24 h or 6 days after Nosema inoculation, respectively, were used. Mature microsporidian spores washed in Dimilin solution prior to oral inoculation, however, were as infectious as spores stored in liquid nitrogen. We have shown that Dimilin interferes with the establishment of the parasite in its host. In addition, when Nosema sp. succeeds in infecting the L. dispar host despite treatment with Dimilin, the microsporidium does not develop optimally and spore production is reduced.     

Microhabitat selection as an antipredator strategy in the aquatic insect Pachydiplax longipennis Burmeister (Odonata: Libellulidae)

Summary An investigation of the larval dragonfly fauna associated with the plant, Sagittaria platyphylla, was conducted in a small pond. Despite the presence of several larval anisopteran species in the pond, only Pachydiplax longipennis larvae were found on Sagittaria plants. A study of the microspatial distribution of P. longipennis larvae on S. platyphylla indicated that larvae use the various regions of a plant in a highly non-random fashion. Larvae show a strong preference for the leaf axil area. A generalized predator, the bluegill sunfish (Lepomis macrochirus), was allowed to selectively eat either of two larvae placed in various plant regions. This experiment indicated that larvae in a leaf axil area were significantly less susceptible to bluegill predation than larvae positioned in other plant regions. The microspatial distribution of starved larvae revealed that larvae with high hunger levels occupied the leaf axil area significantly less than well fed larvae, suggesting 1) larvae do not use these regions as feeding sites, and 2) high hunger levels may induce a behavioral shift in habitat use, with starved larvae forced into areas of high predation risk by the need to fulfill nutritional requirements.     

Location of ecdysteroid 22-O-acyltransferase in the larvae ofHeliothis virescens

Larvae of the tobacco budworm,Heliothis virescens, are resistant to high levels of ingested 20-hydroxyecdysone which could cause potential inhibition to the development of many other lepidopteran species. This resistance is attributed to the ability of the larvae to metabolize this molting hormone to its 22-acyl ester forms. When tobacco budworm larvae were fed large quantities of 20-hydroxyecdyone, the hormonal metabolites were found in gut and fat body tissues. When incubated with 20-hydroxyecdysone gut tissue converted 20-hydroxyecdysone into its 22-acyl ester metabolites. Lumen site of the midgut was found to be the major location of this bio-transformation. In contrast, fat body tissue failed to convert 20-hydroxyecdysone to 22-acyl ester metabolitesin vitro. After the oral injection of³H-ecdysone, the major metabolites formed were ecdysone 22-acyl esters whereas the majority of³H-ecdysone was transformed to polar metabolites after it was injected into the hemocoel of the larvae. Similar distributions of ecdysteroid 22-O-acyltransferase and alkaline phosphatase activity in subcellular fractions demonstrates the co-localization of these enzymes in plasma membrane of the gut epithelial cells. These results suggest that gut brush border membrane is the major site of ecdysteroid 22-acyl ester formation inH. virescens larvae.     

取食聚苯乙烯塑料对黄粉虫幼虫肠道微生物群落的影响北大核心

随着塑料在人类社会中的普及,越来越多的废弃塑料及其前体物质被遗留在环境中,且其在自然环境中的降解速度十分缓慢,为此寻找有效的降解途径成为亟待解决的科学问题。【目的】探究利用黄粉虫(Tenebrio molitor)幼虫取食聚苯乙烯对其肠道微生物种群及其代谢路径的响应,以期通过食物诱导寻找一条生物降解和利用聚苯乙烯的有效途径。【方法】以聚苯乙烯为唯一食物来源喂饲黄粉虫幼虫,通过测量幼虫存活率和个体的体重来测定其生长发育情况;通过对其肠道内容物进行16S rRNA基因测序,分析其肠道菌群结构的变化;采用京都基因与基因组百科全书(Kyoto encyclopedia of genes and genomes,KEGG)分析法来预测相关功能基因。【结果】取食聚苯乙烯黄粉虫幼虫存活率和体重均下降,聚苯乙烯塑料明显减少;取食聚苯乙烯的黄粉虫幼虫肠道菌群丰度与多样性明显减少,在门水平,取食聚苯乙烯的黄粉虫幼虫肠道的优势菌为变形菌门(Proteobacteria)、软壁菌门(Tenericutes)和厚壁菌门(Firmicutes);在属水平,取食聚苯乙烯的黄粉虫幼虫肠道优势菌为螺旋体菌属(Spiroplasma)、肠杆菌(Enterobacillus)和大肠埃氏-志贺氏菌(Escherichia-Shigella);通过KEGG功能预测,找到与芳香类和烷烃降解功能相关的基因共18种,取食聚苯乙烯组黄粉虫幼虫肠道菌群降解聚苯乙烯相关通路丰度升高,相关基因表达增强。【结论】聚苯乙烯可以为黄粉虫幼虫生长发育提供一定的物质和能量,且能够使其完成一个世代过程;幼虫长时间取食单一食物后,其肠道菌群结构会发生目标性变化,利用KEGG预测能够找到与聚苯乙烯代谢相关的基因,为后续研究工作提供了有价值的依据。     

Investigations on the parasitism ofOtiorrhynchus salicicola andO. Sulcatus [Col.: Curculionidae] byHeterorhabditis sp. [Nematoda]

Experiments were conducted to study the efficacy of the insect parasitic nematodeHeterorhabditis sp. (HW79) as a biological control agent ofOtiorrhynchus salicicola. This weevil species is reported as a pest of ornamental plants in Switzerland and Italy. Dipping plastic boxes containing heavily infested cuttings of laurel (Prunus laurocerasus) in a nematode suspension resulted in approximately 100% parasitisation of full-grown larvae, pupae and non-emerged young adults. The average dose resulting from dipping varied between 56,000 and 62,000 nematodes per liter soil. This experiment was run under natural outdoor conditions. In a further outdoor experiment, pottedLigustrum plants were inoculated with eggs ofO. salicicola and later 20,000 infective juvenile nematodes per liter soil were added to the soil surface. The resulting weevil mortality in the treated pots was 78%. In seven greenhouse tests using the same nematode dose in pots with horticultural soil to which weevil larvae had been added, weevil mortality varied between 76% and 100%, the arithmetic average being 90%. These results indicate that Heterorhabditid nematodes may provide an effective means of controllingO. salicicola. In an other experiment usingO. sulcatus larvae, the influence of application time on nematode efficacy was investigated. When nematodes were added a few days before weevil larvae had hatched from the eggs, no parasitic effect was obtained. Nematode applications done shortly after larval hatching however, resulted in complete weevil control. These results are of significance in timing nematode applications in practice.      

Pathogenicity, formulation and storage of insect pathogenic hyphomycetous fungi tested against Diabrotica speciosa

Studies were conducted tosearch for fungal strains with potentialpathogenicity against Diabrotica speciosa(Germar) (Coleoptera: Chrysomelidae).Among sixteen fungal isolates screenedthe most virulent was a Beauveria bassiana(Balsamo) Vuillemin isolate (FHD13) thatcaused 70% mortality of D. speciosathird instar larvae. The LC₅₀ value ofB. bassiana isolate FHD13 was3.48 × 10¹⁰ conidia/ml.Different temperatures (4, 17 and 26 °C)and vegetable oils (corn, sunflower and canola)used for storage did not significantly affectviability of conidia. A pathogenicity trialagainst D. speciosa larvae performed withthe corn oil formulation (1 × 10⁸ conidia/mlof oil) caused 65% of mortality.     

饲料类型对茶尺蠖幼虫肠道细菌多样性及幼虫存活率的影响

【目的】茶尺蠖是茶园中的重要害虫。研究茶尺蠖寄主食物-肠道菌群-茶尺蠖生长发育三者之间的关系对于茶尺蠖的防治具有重要的理论指导价值。【方法】分析不添加茶叶因子的纯人工饲料和茶树鲜叶对茶尺蠖幼虫的存活影响;用高通量测序技术分析不同饲料饲喂的茶尺蠖幼虫的肠道菌群异同。【结果】取食人工饲料的幼虫死亡率远远高于取食茶树鲜叶的幼虫;取食人工饲料的幼虫肠道细菌多样性和丰富度高于取食茶树鲜叶的幼虫;茶尺蠖幼虫肠道中存在很多促进宿主生长的细菌。【结论】饲料类型影响茶尺蠖幼虫的存活;饲料类型影响茶尺蠖幼虫肠道菌群结构。     

Bioevaluation of Subabul (Leucaena leucocephala) proteinase inhibitors on Helicoverpa armigera

Helicoverpa armigera, a highly polyphagous pest, has a broad host spectrum, causes significant levels of yield loss in many agriculturally important crops. Serine primarily responsible for most of the proteolytic activity in the larval gut of lepidopteron insects. Neonate larvae were reared on artificial diet and chickpea seeds smeared with Subabul Trypsin Inhibitor. Larvae fed with artificial diet showed reduction in larval weight up to 21% (HSTI) and 43% (LSTI). However, larvae fed on seeds showed significant reduction in weight, 52.4% (HSTI) and 60.3% (LSTI), suggesting that the diet also plays a vital role on the effectiveness of the inhibitors on larval growth and development. HSTI and LSTI inhibited the gut proteinases from larvae fed on artificial diet significantly (41.40% and 64.36%) compared to the gut proteinases (27.80% and 38.90%) from larvae fed on chickpea seeds. Seeds smeared with 10,000 TIU resulted in complete mortality of larvae while there was no mortality observed in artificial diet. The results reveal that LSTI is a stronger inhibitor of insect gut proteinases and for larvae fed with poor nutrition in the natural ecosystems, low level expression of inhibitor would be enough to affect the growth and development. Handling editor: Chen-Zhu Wang     

Respiratory metabolism of the soft tick,Ornithodoros turicata (Dugès)

The rate of oxygen consumption was investigated in fed larval, nymphal and adult Ornithodoros turicata ticks and in starved nymphal and adult ticks. Oxygen consumption rate of fed adult ticks increased with increasing temperature. The metabolic rate of adult ticks was affected by starvation whereby starved adult ticks showed a significantly lower oxygen consumption than their fed counterparts. The oxygen consumption rate of fed female ticks was significantly higher than that of fed males but, there was no significant difference between the oxygen consumption rates of starved female versus starved male ticks. Oxygen consumption of fed larvae was significantly greater than those of fed first through third instar nymphs. Fed and starved nymphal ticks as well as fed adult ticks ventilated continuously. In contrast, starved adults ventilated discontinuously. The ability to reduce metabolic rate, plus the capability to ventilate discontinuously allow O. turicata adults to cope with prolonged starvation.     

Physiological effect of the host plant on the suitability ofTetranychus urticae as prey forPhytoseiulus persimilis [Acari: Tetranychidae,Phytoseiidae]

Weight gain of adult femalesPhytoseiulus persimilis Athias-Henriot was determined after they were fed eggs or adult females ofTetranychus urticae Koch reared on either Lima bean (Phaseolus vulgaris L.) or nightshade (Solanum douglasii Dunal). No significant difference was found when the predator was fed eggs ofT. urticae reared on either host. Predators were heavier when fed adult femaleT. urticae reared on bean. Apparently, a factor determining the prey's suitability is incorporated into its hemolymph or tissues and is not restricted to its gut content.      

Season long intraplant epizootics of entomopathogens,Beauveria bassiana andNosema pyrausta,in a corn agroecosystem

Intraplant epizootics of entomopathogens,Beauveria bassiana (Balsamo) Vuillemin, andNosema pyrausta (Paillot) were studied in a corn,Zea mays L., agroecosystem. Egg masses of the European corn borer,Ostrinia nubilalis (Hübner), infected withN. pyrausta were placed on midwhorl-stage corn plants. Conidia ofB. bassiana were applied in an aqueous suspension. Frass from the initial insects remaining within the plants was contaminated with sufficientN. pyrausta spores to infest 80% of the filial generation in each year of a 2-yr study. Viability ofN. pyrausta within the frass was monitored throughout the winter. Potential for impact of this inoculum on the filial generation is discussed. Conidia from cadavers of the European corn borer that were killed by the initial inoculum ofB. bassiana and/or conidia from the initial inoculum ofB. bassiana significantly reduced tunneling by the filial generation of the European corn borer. However, neither the concentration ofB. bassiana nor the age of the larvae exposed toB. bassiana, had any significant (P<.05) effect on tunneling by the 2nd-generation larvae. This paper reports the results of research only. Mention of a pesticide in this paper does not constitute a recommendation for use by the U.S. Department of Agriculture or cooperators, nor does it imply registration under FIFRA as amended.     

响应面法优化爪哇虫草菌的培养条件及其对斜纹夜蛾的毒力和保护酶活性影响

【目的】优化爪哇虫草菌Bd01的固态发酵培养条件,测定分生孢子对斜纹夜蛾3龄幼虫的毒力,研究被爪哇虫草菌侵染后寄主体内的保护酶活性变化。【方法】采用单因素试验确定爪哇虫草菌Bd01最佳的固态培养基及培养条件,利用Box-Behnken响应面法优化发酵参数,采用浸渍法测定分生孢子对斜纹夜蛾3龄幼虫的毒力,同时利用分光光度计法测定斜纹夜蛾3龄幼虫体内酶活性变化。【结果】以产孢量为指标,通过响应曲面法优化的爪哇虫草菌Bd01最佳产孢条件为：培养基营养成分含量为30.24g/L,pH值为7.55,光照时长为12.06h,在该条件下,培养基的产孢量为2.78×10⁸孢子/mL。浓度为1×10⁸孢子/mL的爪哇虫草菌孢子液对斜纹夜蛾3龄幼虫具有一定毒力,处理7 d时致死中浓度(LT₅₀)为3.11 d,致死中时(LC₅₀)为4.68×10⁵孢子/mL,校正死亡率为88.68%。处理后未死亡的斜纹夜蛾幼虫体内保护酶活性与对照组相比发生显著变化。【结论】优化后的培养基能够显著增加爪哇虫草菌的产...     

Influence of toxic and non-toxic phytoplankton on feeding and survival of Dreissena polymorpha (Pallas) larvae

Grazing and survival of larvae of the zebra mussel, Dreissena polymorpha, on a green alga and cyanobacteria were studied in laboratory experiments. Clearance rates of the larvae were determined for Chlamydomonas reinhardtii (green alga), two non-toxic and two toxic Microcystis aeruginosa strains (Cyanobacteria). Clearance rates of larvae on non-toxic Microcystis were significantly higher than on toxic Microcystis. The clearance rate on Chlamydomonas reinhardtii was in between the clearance rates on toxic and non-toxic Microcystis strains and not significantly different from them. Effects of toxicity of Microcystis on the survival of zebra mussel larvae was investigated in a short-term experiment. Survival of larvae fed toxic Microcystis was lower than that of larvae fed non-toxic Microcystis, but higher than that of starved larvae. This may imply that, for survival of zebra mussel larvae, it is better to have bad quality (toxic) food than no food.