Potential of Lecanicillium spp. for management of insects, nematodes and plant diseases

Fungi in the genus Lecanicillium (formerly classified as the single species Verticillium lecanii) are important pathogens of insects and some have been developed as commercial biopesticides. Some isolates are also active against phytoparasitic nematodes or fungi. Lecanicillium spp. use both mechanical forces and hydrolytic enzymes to directly penetrate the insect integument and the cell wall of the fungal plant pathogen. In addition to mycoparasitism of the plant pathogen, the mode of action is linked to colonization of host plant tissues, triggering an induced systemic resistance. Recently it was demonstrated that development of Lecanicillium hybrids through protoplast fusion may result in strains that inherit parental attributes, thereby allowing development of hybrid strains with broader host range and other increased benefits, such as increased viability. Such hybrids have demonstrated increased virulence against aphids, whiteflies and the soybean cyst nematode. Three naturally occurring species of Lecanicillium, L. attenuatum, L. longisporum, and an isolate that could not be linked to any presently described species based on rDNA sequences have been shown to have potential to control aphids as well as suppress the growth and spore production of Sphaerotheca fuliginea, the causal agent of cucumber powdery mildew. These results suggest that strains of Lecanicillium spp. may have potential for development as a single microbial control agent effective against several plant diseases, pest insects and plant parasitic nematodes due to its antagonistic, parasitic and disease resistance inducing characteristics. However, to our knowledge, no Lecanicillium spp. have been developed for control of phytopathogens or phytoparasitic nematodes.     

Probiotic effects of beta-glucuronidase on the peach-potato aphid Myzus persicae (Aphididae)

beta-glucuronidase (GUS) is a reporter protein commonly expressed in transgenic plants allowing the visualization of the transformed individuals. In our recent work, we showed that consumption of transformed potato plants expressing this GUS enzyme improves performance of the phloem feeding aphid Myzus persicae. Those results led us to the conclusion that the expression of GUS in potato plants might be responsible for the probiotic effect measured in feeding aphids. In the present paper, artificial diets were used to provide active GUS (10 and 500 microg ml(-1)), inactivated heated GUS (500 microg ml(-1)), glucuronic acid (10, 100 and 500 microg ml(-1)), and bovine serum albumin (500 microg ml(-1)) to M. persicae. Our results reveal that these chemicals provided as food intake might influence the biological parameters of this aphid. Experiments showed a probiotic effect of 500 microg ml(-1) GUS diet, resulting in reduced larval mortality, and increased adult reproduction period and fecundity, which led to an increased population growth potential (r(m)=0.17+/-0.01 versus r(m)=0.12+/-0.03 for aphids fed on control diet). A lower amount of added GUS led to fewer variations, biological parameters being only slightly altered (r(m)=0.14+/-0.03). Statistically similar alterations of the biological parameters were obtained when comparing aphids fed on the diet added with inactivated GUS or the non-structural bovine serum albumin protein (r(m)=0.15+/-0.02 and 0.14+/-0.03, respectively). Feeding assays conducted with glucuronic acid supplemented diets enhanced longevity and nymph production of the adult aphids and reduced larval mortality, resulting in r(m)=0.15+/-0.02 for the highest dose (500 microg ml(-1)). Although 100 microg ml(-1) glucuronate diet did not induce any effect on M. persicae (r(m)=0.12+/-0.03), aphids fed on 10 microg ml(-1) glucuronate diet exhibited unexpected reduced demographic parameters (r(m)=0.10+/-0.03). Immuno-histological analysis showed GUS labeling along the whole digestive epithelium of adults and in various tissues including embryos and bacteriocytes. These results suggest that GUS crosses through the digestive tract. Western blots performed with protein extracts of transformed potato plants expressing the gus gene showed a unique band of molecular weight 76 kDa. On the contrary, in extracts from aphids fed on transgenic potato plants or bred on GUS 500 microg ml(-1) artificial diet, several proteins of lower molecular weight were hybridized, revealing proteolysis of ingested GUS. It is concluded that GUS protein, and more precisely GUS activity, is responsible for the probiotic effects on aphid feeding. The possible pathways of induction of such physiological alterations by GUS are discussed.     

Virulence of Hypocreales fungi to pecan aphids (Hemiptera: Aphididae) in the laboratory

There is need for efficacious biocontrol agents for aphids in commercial orchards. As a preliminary step to this end we determined the virulence of several Hypocreales fungi to pecan aphids. In the first experiment we tested the virulence of Isaria fumosorosea (ARSEF 3581) blastospores to three pecan aphids Monellia caryella, Melanocallis caryaefoliae, and Monelliopsis pecanis under laboratory conditions. Rates of 1 × 10⁷ or 1 × 10⁸ spores per ml were applied in 2 ml via a spray tower to 90 mm Petri dishes containing 10 aphids each. Mortality and mycosis were determined after 24, 48 and 72 h. Treatment effects were observed by 48 h post-application, and by 72 h the higher application rate caused >90% mortality and mycosis in M. caryella and M. caryaefoliae, whereas <70% was observed in M. pecanis.We conducted two subsequent experiments (Experiments 2 and 3), using the same methodology, to compare the virulence of several Hypocreales species and strains against the aphid of primary economic concern to most pecan growers, M. caryaefoliae. In Experiment 2, we compared blastospores and conidia of two I. fumosorosea strains (ARSEF 3581 and ATCC 20874 [= strain 97]). The blastospores of ARSEF 3581 and conidia of ATCC 20874 showed higher virulence than other treatments and thus were included in Experiment 3, which also compared the virulence of conidia of Beauveria bassiana (GHA strain) and Metarhizium anisopliae (F52 strain). Results in Experiment 3 indicated the highest virulence in I. fumosorosea 3581 blastospores and M. anisopliae (F52) followed by I. fumosorosea (20874) conidia. The detection of pathogenicity to pecan aphids establishes the potential for commercial usage and additional study. Results reported here will narrow treatments to test in future greenhouse and field trials.     

Pandora neoaphidis transmission and aphid foraging behaviour

Pandora neoaphidis is an aphid-specific entomopathogen that produces infective conidia. As aphid movement increases, so does the likelihood of contact with conidia. Volatile distress signals released in response to aphid infestation as an indirect defence against herbivory may affect aphid foraging and, therefore, the fungus-aphid interaction. In this study, two different methods were used to investigate the effect of plant volatiles and P. neoaphidis-sporulating cadavers on (1) the colonisation of Vicia faba plants by Acyrthosiphon pisum and (2) P. neoaphidis transmission. This study indicates that A. pisum does not avoid bean plants containing P. neoaphidis and that transmission of conidia occurs during plant colonisation and, to a lesser extent, during in situ feeding. Although significantly more aphids were recovered from damaged plants compared to undamaged plants, the likelihood of infection was not affected by previous infestation by aphids.     

微波辐射对桃蚜生长发育和繁殖的影响

【目的】研究筛选对桃蚜Myzus persicae有致死作用的安全微波频率和照射时长,以为探究新型物理防蚜技术,弥补化学防治上的缺陷提供参考依据。【方法】在暗箱中,应用微波发射仪分别发射1375, 2 750, 5 500和11 000 MHz 4个不同频率的微波照射桃蚜1日龄无翅成蚜,每个频率的照射时长分别为15, 30, 60和120 s;照射后在人工气候箱中饲养,分别于照射后8, 24, 48和72 h观察其生长发育及繁殖状况,统计桃蚜死亡率、繁殖力(累计产蚜量)及子代有翅蚜率。【结果】4个不同频率的微波分别在4个不同照射时长下,对桃蚜1日龄无翅成蚜的死亡率、繁殖力和子代翅型分化都有不同程度的影响。照射后72 h, 5 500 MHz微波照射时间为15 s时对桃蚜1日龄无翅成蚜的致死作用最强,死亡率达到55.00%,在照射时间为30和120 s时可抑制子代桃蚜向有翅蚜的分化。2 750 MHz微波照射30和60 s时促进桃蚜1日龄成蚜繁殖,照射30 s时繁殖力最强,而照射15和120 s时却表现为抑制繁殖,且2 750 MHz微波照射30 s能抑制子代桃蚜向有翅蚜分化。【结论】微波辐射能够影响桃蚜1日龄成蚜的存活、繁殖和子代翅型分化。本研究初步筛选出了对桃蚜1日龄无翅成蚜有致死作用的微波频率和照射时长。     

Pathogenicity of hyphomycet fungi to aphids Aphis gossypii Glover and Myzus persicae (Sulzer) (Hemiptera: Aphididae)

The aphids Aphis gossypii and Myzus persicae are cosmopolitan, poliphagous and damage cultivated plants. The effects of the entomopathogenic fungi Beauveria bassiana (isolate IBCB 66), Metarhizium anisopliae (isolate IBCB 121), Paecilomyces fumosoroseus (isolate IBCB 141) and Lecanicillium (= Verticillium) lecanii (isolate JAB 02) on third instar nymphs of A. gossypii and M. persicae were evaluated in the laboratory at 25 degrees C, 70 +/- 10% RH and 12h photophase. The aphids were transferred to petri dishes with a foliar disk (cotton or pepper) with a layer of 1 cm tick of agar-water. The fungi were applied in a suspension containing 1.0 x 106 to 1.0 x 108 conidia/ml. In the control treatment 1 ml of sterilized water was added to the foliar disks. The mortality of aphids was evaluated daily. B. bassiana and M. anisopliae caused 100% mortality at the seventh day after inoculation, for both species. L. lecanii was the fungus that provided mortality later in the aphids and M. persicae was more susceptible to both fungi than A. gossypii.     

Maternal reproductive decisions are independent of feeding in the black bean aphid,Aphis fabae

Aphids are phloem feeders and an important assumption has been that reproduction is initiated only after phloem ingestion. Here we investigate the plant tissue location of parturition cues in winged and wingless, summer virginoparae and autumn migrants (gynoparae) of the black bean aphid, Aphis fabae. These seasonal forms have different host preferences. Using electrical penetration graph (EPG: to observe activity of the mouthparts) and video-monitoring procedures we demonstrate that the time to first parturition after host-plant contact is significantly shorter than the time to first registered phloem contact in the summer winged form. In gynoparae, the time to first parturition does not significantly differ from time to first phloem contact but is shorter than time to first phloem ingestion. Times to first parturition, first registered phloem contact and first phloem ingestion do not differ significantly in the summer wingless form. Simultaneous EPG and video recording procedures show that a high proportion of individuals of all morphs (45-70%) initiate reproduction before sustained phloem activities (salivation/ingestion). The only behaviours that all individuals demonstrate before first parturition are ‘non-penetration’ (aphid on plant surface with mouthparts outside plant) and stylet ‘pathway activity’, including secretion of gelling saliva and penetration of the non-vascular (epidermis and mesophyll) cells. A short period of penetration of the peripheral plant tissues (five cell punctures per individual) by aphids tethered and monitored by EPG decreases the time to first parturition of the winged summer form when subsequently placed on a Parafilm sachet containing 15% sucrose solution. This treatment also significantly increases the incidence of reproduction and individual reproductive output of gynoparae over a 24-h period. No detectable effects of tissue penetration on subsequent reproductive output are observed in the wingless summer form. Additionally, EPGs reveal that a number of aphids of all morphs display xylem ingestion, which occurs predominantly before initiation of phloem feeding but is not necessary to initiate parturition. It is concluded that aphids are likely to detect parturition cues during stylet punctures of cells within peripheral tissue layers, before reaching the phloem vessels.     

烟蚜茧蜂寄生对烟蚜生长发育和繁殖的影响

【目的】为探究烟蚜茧蜂Aphidius gifuensis对烟蚜Myzus persicae寄生后烟蚜生长发育和繁殖影响及其在烟蚜防控中的作用。【方法】采用室内叶片培养皿法,测定了不同龄期烟蚜(1-4龄若蚜及成蚜)被烟蚜茧蜂寄生后烟蚜当代(F₀)及后代(F₁)的发育历期、寿命、产仔量。【结果】烟蚜1龄若蚜被烟蚜茧蜂寄生后,1龄若蚜历期受到显著影响,较未被寄生对照缩短了0.65 d (33.00%),而2-4龄若蚜期和成蚜历期未受显著影响;2龄若蚜被寄生后,4龄若蚜和成蚜历期均受到显著影响,较未被寄生的对照分别缩短了1.50 d (100.00%)和0.83 d (100.00%),而2和3龄若蚜期未受显著影响;3龄若蚜被寄生后,4龄若蚜和成蚜期均受到显著影响,较未被寄生对照分别缩短了1.50 d (100.00%)和0.83 d (100.00%),而3龄若蚜期未受显著影响。2龄若蚜被寄生后烟蚜的寿命和产仔量均受到显著影响,寿命较未被寄生对照缩短了8.39 d(54.00%),每雌产仔量较未被寄生对照减少了48.67头(100.00%)。此外,2-3龄若蚜被寄生后其后代(F₁)1龄若蚜历期受影响最大,较未被寄生烟蚜后代1龄若蚜期缩短了0.80 d(40.00%),其F₁代平均寿命较未被寄生烟蚜后代平均寿命延长了5.72 d(41.20%),但其产仔量与对照差异不显著。【结论】不同龄期烟蚜被烟蚜茧蜂寄生后,其发育历期和寿命比未被寄生的缩短,产仔量比被未寄生的少,但其F₁代平均寿命比未被寄生后代烟蚜的长且其产仔量未受显著影响。当代2龄若蚜期是控制烟蚜增长的关键时期。这些结果对确定烟蚜茧蜂防治烟蚜适期及认识其机理具有重要参考意义。     

Plant waxy bloom on peas affects infection of pea aphids by Pandora neoaphidis

This study examined the effects of the surface wax bloom of pea plants, Pisum sativum, on infection of pea aphids, Acyrthosiphon pisum, by the fungal pathogen Pandora neoaphidis. In prior field surveys, a higher proportion of P. neoaphidis-killed pea aphids (cadavers) had been observed on a pea line with reduced wax bloom, as compared with a sister line with normal surface wax bloom. Laboratory bioassays were conducted in order to examine the mechanisms. After plants of each line infested with aphids were exposed to similar densities of conidia, the rate of accumulation of cadavers on the reduced wax line was significantly greater than on the normal wax bloom line; at the end of the experiment (13d), the proportion of aphid cadavers on the reduced wax line was approximately four times that on the normal wax bloom line. When plants were exposed to conidia first and then infested with aphids, the rate of accumulation of cadavers was slightly but significantly greater on the reduced wax line, and infection at the end of the experiment (16d) did not differ between the lines. When aphids were exposed first and then released onto the plants, no differences in the proportion of aphid cadavers were observed between the pea lines. Greater infection of pea aphid on reduced wax peas appears to depend upon plants being exposed to inoculum while aphids are settled in typical feeding positions on the plant. Additional experiments demonstrated increased adhesion and germination by P. neoaphidis conidia to leaf surfaces of the reduced wax line as compared with normal wax line, and this could help explain the higher infection rate by P. neoaphidis on the reduced wax line. In bioassays using surface waxes extracted from the two lines, there was no effect of wax source on germination of P. neoaphidis conidia.     

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%。     

Characterization of a new densovirus infecting the green peach aphid Myzus persicae

A new icosahedral DNA virus was isolated from aphids (Myzus persicae) that showed abnormal growth and development. The purified virus particles have a diameter of 20 nm and contain a single-stranded DNA molecule of approximately 5.7 kb. The viral particles are composed of five structural proteins (92, 85, 68, 64, and 57 kDa). As the main biophysical properties of this virus are similar to those of the members of the genus Densovirus it was tentatively named Myzus persicae densovirus (MpDNV). A PCR-based detection method and a polyclonal antiserum raised against MpDNV allowed the detection of the virus in a single-infected aphid. MpDNV is immunologically related to Junonia coenia densovirus, but not to other members of the subfamily Densovirinae. Biological assays showed that MpDNV could be both transmitted transovarially and horizontally via honeydew and saliva. MpDNV was able to infect whiteflies but not other aphid species tested.     

Glucogalactan: A polysaccharide isolated from the cell-wall of Verticillium Lecanii

A water-soluble polysaccharide was extracted with alkali from the cell wall of Verticillium lecanii (also called Lecanicillium lecanii). After freezing and thawing, the water-soluble fraction was purified by gel filtration chromatography on Sepharose CL-6B and eluted as one peak by HPSEC/RID. Monosaccharide analysis showed galactose and glucose (1.1:1), with traces of mannose (<1%). The structural characteristics were determined by spectroscopic analysis, FT-IR and 1D and 2D ¹H and ¹³C NMR, and methylation results. On the basis of the data obtained, the following structure of the polysaccharide (E₃S_IV fraction) was established:     

Effects of different Beauveria bassiana isolates on field populations of Lygus lineolaris in pigweed (Amaranthus spp.)

The tarnished plant bug, Lygus lineolaris (Palisot de Beauvois), is a pest of various fruit, vegetable, fiber, and seed crops; including cotton. Lygus spp. populations often build on alternate host plants before moving to cotton, and in the midsouthern U.S. wild host plants, such as pigweed (Amaranthus spp.), play a major role in L. lineolaris population development. Three isolates of the entomopathogenic fungus Beauveria bassiana (Balsamo) were evaluated for L. lineolaris control in redroot pigweed (Amaranthus retroflexus L.): one from L. lineolaris in Mississippi (TPB3); one from Lygus hesperus (Knight) in California (WTPB2); and one commercial isolate from Mycotrol^® (GHA). Fungal applications resulted in moderate to high mycosis in adults (33 to 80%) and moderate mycosis in nymphs (36 to 53%) that were collected from field plots at 2 days post-treatment and incubated under laboratory conditions. Although TPB3 was previously found to be more pathogenic in laboratory bioassays, there was not a consistent separation of this isolate from the other two isolates in field trials. Where differences in adult mycosis or mortality were observed, TPB3 was the most pathogenic. However, in one field trial 7 day mortality for nymphs treated with GHA was higher than those treated with TPB3 or WTPB2. Infection rates at 2, 7, and 14 days post-treatment from caged and non-caged adults suggested that movement of adults among plots occurred, which could have masked some treatment effects. Fungal treatments did not significantly reduce populations relative to controls. This may have been caused by delayed mortality rates under field conditions and/or difficulties with estimating population change under field conditions characteristic of wild host plant populations (e.g., heterogeneous populations, adult movement, and small plot size). Further work evaluating time–dose–mortality over dynamic temperatures, spring and fall field trials on this and other wild hosts, and improved methods for estimating populations on wild hosts are needed.     

Opportunism of Conidiobolus obscurus stems from depression of infection in situ to progeny colonies of host alatae as disseminators of the aphid-pathogenic fungus

Conidiobolus (Entomophthorales: Ancylistaceae) includes common aphid pathogens but causes sporadic mycosis worldwide. This epizootiological opportunism was explored herein by examining the potential of mycosis transmission in the progeny colonies of 513 Myzus persicae alates as disseminators of C. obscurus often infecting aphids. The alates exposed to spore showers were flown for 2.05 (0.01-8.95) km on flight mills and then reared individually on cabbage at 20-23 degrees C for 14 day colonization. All truly infected alates were mycosed within 6 days and averagely left 3.9 (0-15) nymphs while those uninfected produced 11.6 (0-35) nymphs during the same period. Secondary and tertiary infections occurred only in 16.2% and 4.8% of the progeny colonies of the mycosed alates respectively, due to c. 60% of the cadavers forming resting spores. Most of the contagious infections appeared on days 4-8 after colonization and no more occurred from day 11 onwards. Trends of colony sizes (last-day averaging 51.5 aphids) and mycosis transmission (sixth-day maximum 6.3%) fit well to logistic (r2 = 0.99) and Gompertz growth models (r2 = 0.91) respectively. The results confirm that the opportunism of C. obscurus stems from depression of contagious infection after dissemination by host alates and suggest that it be overwhelmed by the prevalence of other non-resting fungal species.     

Effect of temperature,relative humidity and aphid developmental stage on the efficacy of the mycoinsecticide Mycotal® against Myzus persicae

The green peach aphid, Myzus persicae, is a major pest worldwide. An examination of the impact of temperature, relative humidity (RH) and developmental stages of M. persicae on the efficacy of the whitefly mycoinsecticide Mycotal®, based on Lecanicillium muscarium and the effects of infection on aphid fecundity was evaluated under controlled conditions. Although this fungus can be grown at a broad range of temperatures (15–30°C), the optimum temperature for control of M. persicae ranged between 20 and 30°C. L. muscarium had high efficacy as a microbial control agent against M. persicae between 55% and 90% RH. Total mortality of aphids treated with different spore dosages of L. muscarium varied according to the developmental stage: adults, fourth and third instar nymphs proved more susceptible than first instar nymphs. Although the fungus did not affect the rate of nymph production, the reproductive period of aphids significantly decreased with increasing the spore dosage. Thus, total fecundity of treated aphids was 22.6 ± 1.1 and 31.6 ± 2.4 offspring per adult at the medium (644 ± viable spore/mm²) and low (330 ± 40 viable spore/mm²) dosages, compared with 45.7 ± 4.3 offspring per untreated aphid. The results suggest that L. muscarium has the potential as a biological control agent of M. persicae.     

温度对桃蚜和马铃薯长管蚜实验种群生命表参数的影响

【目的】桃蚜Myzus persicae (Sulzer)和马铃薯长管蚜Macrosiphum euphorbiae (Thomas)是2种主要的马铃薯害虫。本研究旨在明确温度对这两种马铃薯蚜虫生长发育、存活、繁殖及种群增长的影响。【方法】在室内测定了5个温度（10, 15, 20, 25和30℃）下桃蚜和马铃薯长管蚜的生长发育、存活和繁殖指标,并组建了4个恒温条件下（10,15,20和25℃）的实验种群生命表。【结果】在10~25℃范围内,2种蚜虫的若蚜期、世代历期、成蚜寿命和产蚜期等均随温度的升高而缩短,而30℃高温抑制了其发育、存活和繁殖。2种蚜虫的平均世代历期（T）随温度的升高而缩短,桃蚜和马铃薯长管蚜分别从10℃的30.08 d和35.35 d缩短至25℃的14.28 d和12.95 d。桃蚜的净增殖率（R₀）在15℃时最高（86.00）,其次为20℃（73.75）,再次为25℃（62.49）,最低为10℃（51.00）;马铃薯长管蚜的R0在15℃最高（58.97）,其次为10℃（51.98）,再次为20℃（48.94）,最低为25℃（12.36）。桃蚜的内禀增长率（r_m）随温度的升高而增大,从最小10℃的0.1307增大到25℃的0.2896;马铃薯长管蚜的r_m在20℃时最大（0.2182）,其次为25℃（0.1942）,再次为15℃（0.1485）,最小为10℃（0.1118）。在相同的温度下,桃蚜的发育速率、净增殖率和内禀增长率均高于马铃薯长管蚜。【结论】温度对2种马铃薯蚜虫的生长发育、存活、繁殖及种群增长有显著的影响,桃蚜在马铃薯上的种群增长能力强于马铃薯长管蚜。这一结果为马铃薯蚜虫预测模型的建立奠定了必要的基础,并解释了桃蚜在马铃薯上发生数量多于马铃薯长管蚜的原因。     

Cheaper is not always worse: strongly protective isolates of a defensive symbiont are less costly to the aphid host

Defences against parasites are typically associated with costs to the host that contribute to the maintenance of variation in resistance. This also applies to the defence provided by the facultative bacterial endosymbiont Hamiltonella defensa, which protects its aphid hosts against parasitoid wasps while imposing life-history costs. To investigate the cost–benefit relationship within protected hosts, we introduced multiple isolates of H. defensa to the same genetic backgrounds of black bean aphids, Aphis fabae, and we quantified the protection against their parasitoid Lysiphlebus fabarum as well as the costs to the host (reduced lifespan and reproduction) in the absence of parasitoids. Surprisingly, we observed the opposite of a trade-off. Strongly protective isolates of H. defensa reduced lifespan and lifetime reproduction of unparasitized aphids to a lesser extent than weakly protective isolates. This finding has important implications for the evolution of defensive symbiosis and highlights the need for a better understanding of how strain variation in protective symbionts is maintained.     

Interspecific symbiont transfection confers a novel ecological trait to the recipient insect

In Japan, pea aphids Acyrthosiphon pisum mainly feed on vetch and clover, and many aphid clones produce more progeny on vetch than on clover. In this context, particular genotypes of the facultative symbiont Regiella insecticola enhance reproduction of infected pea aphids specifically on clover, thereby broadening the suitable food plant range of the insect. A species that is sympatric to A. pisum, vetch aphids Megoura crassicauda, are commonly found on vetch but not on clover. Laboratory rearing of M. crassicauda strains revealed active reproduction on vetch but substantially no reproduction on clover. Experimental transfection of Regiella from A. pisum to M. crassicauda by haemolymph injection established stable and heritable infection in the recipients, although no Regiella infection has been detected in natural populations of M. crassicauda. Different strains of Regiella-transfected M. crassicauda grew and reproduced on vetch, but exhibited lower fitness in comparison with corresponding uninfected aphid strains. Strikingly, the Regiella-transfected M. crassicauda exhibited improved survival and some reproduction on clover. These results suggest that Regiella has the potential to confer an ecological trait, adaptation to clover, on novel insect hosts, and also account for why Regiella is able to infect M. crassicauda but is scarcely found in these aphid populations.     

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.     

Effects of kaolin particle film on Myzus persicae (Hemiptera: Aphididae) behaviour and performance

The emergence of resistance mechanisms to, and revocation of, many insecticides used in the control of the polyphagus aphid pest, Myzus persicae (Sulzer), has increased the pressure to develop novel approaches for the control of the pest in many crops. Kaolin-based particle films provide a physical barrier against insect pests and show considerable potential for controlling M. persicae. We conducted a series of laboratory experiments to investigate the mode of action of kaolin against aphids. The material appeared to have no direct effect on M. persicae; spraying adult aphids with aqueous kaolin suspension had no significant impact on their subsequent survival or reproduction on untreated plants. Similarly, when aphids were placed on kaolin-treated host-plants (Brassica oleracea), their performance (survival, growth rate and reproduction) was not significantly different from aphids on untreated plants. However, when M. persicae were given a choice between kaolin-treated and untreated (or water solvent-treated) leaf areas, both adults and nymphs exhibited a significant preference for non-kaolin-treated host-plant material. Rejection of kaolin-treated plant material occurred very rapidly (within 20 min) and this behavioural effect may be related to the efficacy of kaolin in controlling aphids under field conditions.