Review on safety of the entomopathogenic fungi Beauveria bassiana and Beauveria brongniartii

The commercial use of entomopathogenic fungi and their products as mycoinsecticides necessitates their registration. Worldwide, several registration guidelines are available, however, most of them focus on similar or even the same safety issues. With respect to the two entomopathogenic fungi, Beauveria bassiana (Bals.-Criv.) Vuill. and Beauveria brongniartii (Sacc.) Petch, many commercial products have been developed, and numerous papers on different biological, environmental, toxicological and other safety aspects have been published during the past 30-40 years. The aim of the present review is to summarise these data. The following safety issues are presented: (1) identity of Beauveria spp.; (2) biological properties of Beauveria spp. (history, natural occurrence and geographical distribution, host range, mode of action, production of metabolites/toxins, effect of environmental factors); (3) analytical methods to determine and quantify residues; (4) fate and behaviour in the environment (mobility and persistence in air, water and soil); (5) effects on non-target organisms (non-target microorganisms, plants, soil organisms, aquatic organisms, predators, parasitoids, honey bees, earth worms and nontarget arthropods); (6) effects on vertebrates (fish, amphibia, reptiles and birds); and (7) effects on mammals and human health. Based on the present knowledge it is concluded that both Beauveria species are considered to be safe.     

Biological control of Strophosoma spp. (Coleoptera: Curculionidae) in greenery (Abies procera) plantations using Hyphomycetes

In Abies procera plantations Strophosoma melanogrammum and S. capitatum cause economic damage due to the adult stage feeding on the needles. No chemical treatments of these weevils are allowed in Denmark, so the potential of biological control was evaluated. We studied pathogenicity of thirteen isolates of entomopathogenic fungi and the field effect of soil application of Metarhizium anisopliae against Strophosoma spp. All of the tested isolates were capable of causing infections under laboratory conditions and average survival time at 20°C ranged between 13 and 23 days for S. melanogrammum and between 23 and 28 days for S. capitatum when dipped in a fungal suspension adjusted to 1×10⁷ conidia mL^-1. Under field conditions up to 90% of the living collected individuals died due to M. anisopliae infection in the treated plots, whereas less than 1% died of M. anisopliae infection in control plots. In accordance, the population of Strophosoma spp. was reduced in plots where the fungus was applied compared to control plots.     

Persistence of Conidia of Metarhizium anisopliae in Sugarcane Fields: Effect of isolate and formulation on persistence over 3.5 years

Metarhizium anisopliae is being used in Australia as a biopesticide for control of sugarcane whitegrubs in soil. The field persistence in sugarcane soil of two isolates of M. anisopliae each in four formulations was tested by mixing the formulation with soil which was then placed in PVC rings and buried in sugarcane fields. The two isolates used were FI-1045, M. anisopliae var. anisopliae, the active ingredient in BioCane™ currently used for greyback canegrub control, Dermolepida albohirtum (Coleoptera: Scarabaeidae: Melolonthinae), and FI-147, M. anisopliae var. lepidiotum, being tested as a biopesticide for Lepidiota spp. (Coleoptera: Scarabaeidae: Melolonthinae) and other species of canegrub. The four formulations were rice granules (as in BioCane™), a wettable powder derived from conidia screened from the rice granules, conidia off rice suspended in water and conidia produced on agar plates, dried, and then mixed with water for adding to soil. FI-1045 was tested at three different sites in north Queensland with a range of soil types and climatic conditions while FI-147 was tested at three similarly diverse sites in southern Queensland. The PVC rings were destructively sampled every 6 months for 3.5 years and the number of viable conidia remaining determined by plating onto a selective medium. The exponential decay was determined. Monthly decay rates ranged from 0.0309 to 0.0835 (mean 0.0512). A small proportion of conidia survived the 3.5 years at all sites and all formulations. Overall, isolate FI-147 persisted better than FI-1045, but was used at the more Southerly sites. Rainfall and soil type had negligible effects on persistence. The agar-produced FI-147 conidia declined most slowly, while the two rice-produced but water-formulated conidia gave similar results. Isolate FI-1045 survived best as the BioCane™ formulation and this rice granule formulation was almost as persistent as the agar conidia with FI-147. A small proportion of conidia, in some formulations and at some sites, were recovered from immediately below the rings. This movement was thought to be due to activity of earthworms or mites. The results suggest that 3 years would be the maximum period for a BioCane™ formulation to provide some level of infection in the target pest unless augmented by conidia from infected grubs. The effectiveness of these new conidia may be reduced due to their highly aggregated distribution.     

Honey bees can disseminate a microbial control agent to more than one inflorescence pest of oilseed rape

Pollen beetles (Meligethes aeneus) and cabbage seed weevils (Ceutorhynchus assimilis) are major pests of oilseed rape (Brassica napus) throughout Europe. In field cage experiments in both winter and spring rape, honey bees (Apis mellifera) effectively transported the entomopathogenic fungus Metarhizium anisopliae to the flowers, causing infection and mortality of both adult and larval pollen beetles, as well as of adult seed weevils. External conidiation was observed on many of the dead pest insects. Although some external conidiation also occurred on dead honey bees, reduction in honey bee colony size during the experiments appeared unrelated to the fungus. The potential of this technique for integration into pest management strategies for the crop, particularly in association with a trap crop, is discussed.     

Evaluating the importance of residual effects from previous years' treatment on the efficiency of different strategies for control of Sahelian grasshoppers with Metarhizium anisopliae var. acridum

A model for the residual effect of previous years' treatment with the mycoinsecticide Metarhizium anisopliae var. acridum has been constructed based on field data from treatment of two different grasshopper species in east Niger. This model was incorporated in an earlier developed simulation model to evaluate control strategies for the grasshopper Oedaleus senegalensis in West Africa and the model system was used to assess the potential importance of second year's residual effects for the efficiency of different treatment strategies. It has earlier been hypothesized that the persistence of M. anisopliae and the ability of this agent to impose long-term control on grasshopper populations through secondary cycling in some cases might render the use of M. anisopliae more efficient than the use of chemical insecticides like fenitrothion. Results show that this effect is possible if M. anisopliae is used in a repetitive treatment strategy where control operations are concentrated in the main millet production areas.     

Prospects of a fungus-contamination device for the control of tsetse fly Glossina fuscipes fuscipes

The prospects of the fungus Metarhizium anisopliae (Metsch.) Sorok. applied in contamination devices (Cds) to control tsetse fly Glossina fuscipes fuscipes Newstead was tested in a field experiment in Lake Victoria from 2 March 1999 to 31 August 2000. One hundred and sixty pyramidal traps mounted with Cds were deployed along the lakeshore and rivers on Mfangano Island. Contamination devices were loaded with 1.5-2.0 g of dry conidia/Cd. On the second island, Nzenze Island, four pyramidal traps fitted with plastic bags were deployed and served as the conventional 'trap and kill' population suppression method. A third island, Ngodhe Island, remained untreated and served as a control. Cds were recharged monthly with fresh conidia; plastic bags were also changed monthly. The apparent changes in population density were monitored weekly using biconical traps set at random on the three islands. To assess the incidence of M. anisopliae in tsetse flies on Mfangano Island, flies captured during monitoring were maintained in the laboratory and their mortality recorded. Fly population was reduced to 82.4 and 95.8% relative to untreated control on Mfangano and Nzenze islands, respectively, during the experimental period. Compared to the fungus-treated island, the number of flies caught in monitoring traps increased considerably in 'trap kill' treatment at 5 months after the treatments were removed. The incidence of M. anisopliae in fly populations was low during the 12 weeks following the initiation of the experiment but increased afterward until termination of the treatment. M. anisopliae could still be recovered from fly populations at 3 months after termination of the treatment, although the incidence was low. The results of this study have shown that application of M. anisopliae in a contamination device can suppress the population of G. fuscipes fuscipes comparable to the 'trap and kill' technology.     

Laboratory investigations on the potential of entomopathogenic fungi for biocontrol of Helicoverpa armigera (Lepidoptera: Noctuidae) larvae and pupae

The susceptibility of third instar Helicoverpa armigera to seven strains of three entomopathogenic fungal species, i.e. Metarhizium anisopliae, Beauveria bassiana and Paecilomyces fumosoroseus, was tested under laboratory conditions using the larval immersion method. High efficacies ranging from 68 to 100% corrected mortality were recorded with more profound effects in treatments with B. bassiana and P. fumosoroseus strains. The median lethal concentration (LC₅₀) for L3 was 6.0×10⁵ in M. anisopliae 79, 1.5×10⁵ in B. bassiana 124 and 4.2×10⁴ in P. fumosoroseus 14. These three strains were further used to characterize the age-dependent mortality of different larval stages (L2-L5) and the effect against pupae of H. armigera. Larval stages did not differ in their mortality but differed i in median lethal time, with shorter values recorded in the second instar. Tested fungi also caused a high reduction between 74.4 and 100% in the emergence of pupae using the soil inoculation method and the pupal immersion technique. All three fungal species, especially P. fumosoroseus, have a high potential for biocontrol of H. armigera larvae and also as a soil treatment targeting the pupae.     

Environmental contamination with Metarhizium anisopliae from fungal bands for control of the Asian longhorned beetle, Anoplophora glabripennis (Coleoptera: Cerambycidae)

Our objective was to determine whether conidia from non-woven fiber bands impregnated with Metarhizium anisopliae F 52 could be spread to other parts of the environment, and whether Anoplophora glabripennis could become infected with conidia dispersed from fungal bands. In the field, bands containing M. anisopliae were hung at 3 m height on 15 trees. Bark samples were taken 10, 20, and 30 cm above the bands and 10, 30, and 60 cm below the bands 2, 5, and 9 days after band placement to quantify conidial densities. There were more conidia in samples taken below bands compared with samples taken above bands. A significant positive correlation was found between rainfall and the occurrence of conidia on any of the bark samples. A laboratory study was conducted to determine whether conidia from M. anisopliae fungal bands could be spread to other parts of the environment by A. glabripennis adults and whether A. glabripennis could become infected by conidia dispersed from fungal bands. One or five adult A. glabripennis were used to contaminate artificial environments with conidia. All adults subsequently exposed to contaminated environments were killed by fungal infection. Beetles exposed to environments that had been contaminated by five beetles died in fewer days compared with environments contaminated by one beetle. Beetles in both density treatments died in fewer days than beetles exposed to environments without M. anisopliae conidia. Our results indicate that environmental contamination with conidia from fungal bands can occur, and that adult A. glabripennis can acquire infection from a contaminated environment.     

Evaluation of the Entomopathogenic Fungi Metarhizium anisopliae and Paecilomyces fumosoroseus (Deuteromycotina: Hyphomycetes) for Control of the Green Leafhopper Empoasca decipiens (Homoptera: Cicadellidae) and Potential Side Effects on the Egg Parasitoid Anagrus atomus (Hymenoptera: Mymaridae)

The efficacy of Metarhizium anisopliae strain Ma43 and Paecilomyces fumosoroseus strain Pfr12 (both Deuteromycotina: Hyphomycetes) against adults of Empoasca decipiens (Homoptera: Cicadellidae) and potential side effects on the egg parasitoid Anagrus atomus (Hymenoptera: Mymaridae), were investigated in greenhouse cage and laboratory experiments. Treating leafhopper-infested faba bean plants at a dose rate of 1×10⁷ conidia mL^-1 resulted in up to 97% mortality 7 days after application and a 100% infection rate. Experiments on the residual effects revealed a significant decrease in adult E. decipiens mortality with increasing time from application to insect release. The decrease in mortality over time corresponded well with data from conidia germination tests. The germination of conidia on agar medium after washing them from the surafce of sprayed plants declined significantly from 95 and 96% immediately after application for M. anisopliae Ma43 and P. fumosoroseus Pfr12, respectively, to 29 and 27% 5 days later. Experiments on potential side effects of the entomopathogenic fungi on A. atomus showed that the tested isolates had no influence on adult emergence and longevity; however, the rates of parasitism were significantly reduced by the fungal treatments. The latter might be due to either density effects and/or could indicate that A. atomus avoids fungal-treated plants. However, the parasitoid is substantially less susceptible to the fungal strains tested than the host itself.     

Interaction Between the Entomopathogens Beauveria bassiana, Metarhizium anisopliae and Paecilomyces fumosoroseus and the Mycoparasites Clonostachys spp., Trichoderma harzianum and Lecanicillium lecanii

Biocontrol agents of numerous insect pests and fungal pathogens exist but virtually nothing is known about their interaction if used simultaneously. Our objective was to investigate the compatibility of the entomopathogens Beauveria bassiana, Metarhizium anisopliae and Paecilomyces fumosoroseus, and the broad host-range mycoparasites Clonostachys spp., Trichoderma harzianum and Lecanicillium lecanii. In vitro host-range tests revealed that M. anisopliae was highly susceptible to all mycoparasites tested. B. bassiana was attacked by Clonostachys rosea, and P. fumosoroseus. was resistant to mycoparasites. M. anisopliae but not P. fumosoroseus killed nymphs of Bemisia tabaci in bioassays. B. bassiana and M. anisopliae proved lethal to Cosmopolites sordidus, Diatraea saccharalis and Sitophilus oryzae. Coapplication of mycoparasites with entomopathogens did not affect their biocontrol efficacy in vivo, although the reisolation success of entomopathogens could be significantly reduced, especially from smaller insect species. Trichoderma spp. were reisolated from mycoparasite-treated insects more frequently than C. rosea. The coapplication of the highly susceptible M. anisopliae generally enhanced mycoparasite recovery. Mycofungicide preparations caused some insect mortality but less than a copper hydroxide fungicide which is still permissible in organic agriculture. We concluded that the tested entomopathogens and mycoparasites are compatible elements of integrated pest management.     

Selection of an Isolate of the Insect Pathogenic Fungus Metarhizium anisopliae Virulent to the Lettuce Root Aphid, Pemphigus bursarius

The virulence of hyphomycete entomopathogenic fungi was measured in laboratory bioassays against the lettuce root aphid, Pemphigus bursarius, a serious pest of field lettuce grown in the UK. Of 25 isolates of fungi examined, only one isolate, Metarhizium anisopliae 391.93, killed lettuce root aphids consistently. This fungus was isolated originally from the closely related saltmarsh aphid, P. trehernei. The median lethal concentration of conidia at 10 days post6- 1 inoculation estimated from five independent bioassays was 2.45 × 10⁶ conidia ml^-1. The fungus had no significant effect on the mean number of offspring/aphid produced, but it sporulated 6 profusely on host cadavers, producing approximately 4 × 10⁶ conidia/cadaver 14 days after treatment, and diseased aphids died attached to plant roots. It thus has the potential to spread through densely packed colonies of P. bursarius feeding on the roots of susceptible or partially resistant plants.     

Effects of chemical and botanical insecticides used for locust control on Metarhizium anisopliae var. acridum conidia after short- to medium-term storage at 30°C

The short- to medium-term viability and growth of Metarhizium anisopliae var. acridum conidia were investigated when combined with six insecticides, at three different concentrations. All of the insecticides used in this study were suitable for immediate spraying with M. anisopliae var. acridum conidia except for fenitrothion. Fipronil, teflubenzuron, and fenitrothion formulations significantly reduced conidial viability over time. The 10% teflubenzuron treatment caused loss of viability relatively quickly with 9.9% germination after 28 days. Mycelial growth was affected by all the treatments except fenitrothion.     

Suppression of Meloidogyne hapla populations by Hirsutella minnesotensis

The effects of the endoparasitic fungus Hirsutella minnesotensis on populations of Meloidogyne hapla from Michigan (MI), Rhode Island (RI), Connecticut (CT), Lyndonville, New York (NYL), Geneva, New York (NYG), and Wisconsin (WI) were studied in the greenhouse. Twenty-day-old tomato (cv. Rutgers) seedlings were inoculated with either 0 or 600 eggs of each nematode population mixed with either 0, 0.02, or 0.1 g of fresh H. minnesotensis mycelium 0.1 L^-1 of soil in pots containing 0.5 L of soil, and maintained at 25±2°C for 2 months. No effect of the fungal treatments and nematode treatments on tomato plant heights and shoot dry weights was observed. While all M. hapla populations were suppressed by H. minnesotensis, the degree to which each population was affected varied slightly. Across fungal treatments and nematode populations, the fungus reduced total number of nematodes in roots by 61-98%, with the highest for NYG and RI, intermediate for NYL and CT, and lowest for MI and WI populations. The study demonstrated that H. minnesotensis may be used as a potential suppressor of M. hapla in vegetable production systems in the Great Lakes Region.     

摩西管柄囊霉Funneliformis mosseae对连作大豆分枝期根系AM真菌群落结构的影响

本文研究摩西管柄囊霉Funneliformis mosseae对短期连作3个大豆品种分枝期根系AM真菌群落结构的影响,旨在探索不同大豆品种与AM真菌在连作土壤中的互作效应。通过向黑农44（HN44）、黑农48（HN48）和恳丰16（KF16）3个大豆品种盆栽土壤中接种F. mosseae,对接菌连作0年（未连作对照组）、连作1年、连作2年土壤中的大豆根系,采用Nested-Program Control Register-DGGE技术分析。结果表明,接种F. mosseae后,不同大豆品种根系AM真菌多样性指数和丰度值表现为连作2年的土壤>连作1年的土壤>未连作的土壤;其中,球囊霉属Glomus和F. mosseae为3个大豆品种根系中AM真菌的优势菌群。接种F. mosseae对连作1年、连作2年3个大豆品种根系AM真菌群落结构具有显著影响。     

粘盖乳牛肝菌在低磷、酸铝胁迫下的生长和代谢响应

低磷和酸铝胁迫是酸性土壤限制植物生长的主要因素。有研究指出外生菌根(ectomycorrhiza,ECM)可提高宿主植物对铝毒害和低磷胁迫的适应性。然而,目前有关ECM真菌自身对低磷和酸铝环境的适应机理还不清楚。基于此,本研究以我国南方酸性土壤广泛分布的ECM真菌——粘盖乳牛肝菌Suillus bovinus为研究对象,在纯培养条件下研究了低磷、酸铝胁迫对其生长、营养吸收及菌丝分泌物的影响。结果表明,粘盖乳牛肝菌是一种耐铝型真菌,酸铝胁迫(1 mmol/L)不影响其菌丝生长,而低磷胁迫(20 μmol/L)则显著限制其菌丝生长(P<0.05)。值得注意的是,低磷胁迫的抑制效应可被酸铝胁迫逆转。低磷胁迫显著降低了粘盖乳牛肝菌对磷的吸收(P<0.05),而酸铝胁迫则对菌丝钾的吸收有促进作用。低磷、酸铝胁迫同样改变了菌丝分泌物组成。在低磷胁迫下,大量酚酸类、有机酸及脂质代谢物的积累量下调;而酸铝胁迫下则有大量酚酸类物质上调,有机酸和脂质中上调代谢物数量也高于下调数量;低磷酸铝复合胁迫下酚酸和有机酸类代谢物积累量均显著上调。另外,吲哚-3-乙酸(IAA)在各胁迫下均显著上调。以上结果可在一定程度上解释粘盖乳牛菌对低磷、酸铝环境的适应机理,并对后续进一步阐明ECM的共生适应机理有一定指导意义。     

我国抗虫转基因杨树生态安全性研究进展

转基因树木与农作物相比, 人们更关注其长时间种植可能导致转基因扩散到周围野生近缘种。由于生长周期长, 转基因树木会增加转基因不稳定性, 对非靶标生物的影响, 靶标害虫对转基因植物产生抗性, 增加树木入侵性(杂草化), 以及由于基因漂移或基因逃逸对环境产生的负面影响或新的环境风险。过去十几年, 针对我国抗食叶害虫的两个商业化转Bt基因欧洲黑杨(Populus nigra)和转双抗虫基因741杨[P. alba× (P. davidiana + P. simonii) × P. tomentosa], 已开展了有关生态安全性方面的多项研究。本文围绕抗虫转基因树木生态安全性研究进展进行了综述。抗虫转基因杨树对节肢动物种群和群落结构产生了一定影响, 使昆虫的多样性提高, 但对土壤微生物区系未见明显影响。转基因欧洲黑杨通过花粉和种子发生的基因漂移几率很低。转基因杨树通过内生菌发生的水平转移可能会对环境造成的潜在危险也进行了评价。文章最后指出对抗虫转基因杨树农林复合生态系统开展生物安全研究的必要性。     

土壤生物与土壤污染研究前沿与展望

随着社会经济发展,人类生产活动对自然环境产生越来越广泛深刻的影响,土壤污染已成为危及生态系统稳定、农产品质量安全和人体健康的突出环境问题之一。重金属、有机污染化合物、病原菌及抗性基因等各类污染物大量进入土壤后,对土壤生物系统造成毒害作用,影响到土壤生态功能;另一方面,土壤生物如细菌、真菌、土壤动物等在一定程度上能够适应土壤污染,深刻影响着污染物在土壤中的迁移转化过程,在土壤污染修复中具有潜在重要作用。从土壤污染的生态毒理效应、土壤生物对土壤污染的响应与适应机制、污染土壤修复原理与技术等三方面综述了土壤生物与土壤污染相关研究前沿,展望了重点研究方向。     

连作土壤中施加球毛壳ND35菌肥对杨树生理特性的影响

在山东农业大学林学实验站杨树连作土壤中施加不同剂量(0.17、0.33、0.67、1.00、1.33和1.67 g·kg^-1)的球毛壳ND35菌肥,测定不同处理下一年生杨树叶片的光响应过程、叶绿素荧光以及叶黄素循环等光合生理生化指标,研究球毛壳ND35菌肥对杨树根系和光合生理性能的影响.结果表明: 随连作土壤中菌肥含量的增加,杨树叶片的叶绿素含量(Chl)呈增加趋势,电子传递速率(ETR)、净光合速率(P_n)、量子效率(Φ)、硝酸还原酶(NR)活性以及根系活力等生理指标均呈先增加后降低的趋势,依赖于叶黄素循环的光合热耗散呈降低趋势,而非光化学淬灭(NPQ)呈先降低后增加的趋势.在土壤施加菌肥剂量0.67和1.00 g·kg^-1处理时,光合机构的过剩光能减少,向光化学反应方向分配的光能增多,光能利用效率提高.在连作土壤中施加适量(0.67～1.00 g·kg^-1)的球毛壳ND35菌肥,能在一定程度上提高杨树的根系生理活性,提高杨树叶片对光能的利用效率,有利于改善杨树叶片的光合机构运转状态,提高叶片的光合作用效率.     

Exploring the influence of soil types underneath the canopy in winter wheat leaf area index remote estimating

Aims Remote sensing is an effective and nondestructive way to retrieve leaf area index (LAI) from plot, regional and global range. Soil background is one of the confounding factors limiting remotely estimating LAI. And soil type contains a large proportion of soil background information, which can influence the optical properties of vegetation canopy and soil. However, our knowledge on the effects stemmed from soil types underneath the canopy on LAI remote estimating have been in shortage. Thus, this study aims to explore the influences of soil types underneath the canopy on winter wheat LAI remote estimating. Methods We analyzed the sensitivity variation of eight spectral indices, named normalized difference vegetation index (NDVI), modified soil-adjusted vegetation index (MSAVI), modified chlorophyll absorption ratio index 2 (MCARI₂), red edge inflection point (REIP), red edge amplitude (Dr), red edge area (SDr), red edge symmetry (RES), normalized difference spectral index (NDSI), to LAI in different soil types, and then we identified some spectral intervals or parameters that were insensitive to soil type variations underneath the canopy. We also compared the accuracy of two commonly used regression models, partial least squares regression (PLSR) and random forest regression (RFR), in estimating LAI for different soil types. We also explored the problems arising from applying the regression model developed in single soil type area to complex soil types area in retrieving LAI. Important findings This paper demonstrates the effects of soil types underneath the canopy on LAI retrieving. 1) The sensitivity of spectral indices to LAI is significantly different due to the soil type variation, but REIP has the least effects from soil type variation among the eight spectral indices. Meanwhile, the band selection algorithm of lambda-by-lambda not only chooses the most sensitive spectral interval for LAI, but also provides a feasible way to construct the spectral index that exhibits strong resistances to the effects of soil types underneath the canopy. 2) The accuracy of LAI estimation by regression models differs under soil type considered or not. So we suggest that in small scale researches, especially in a field scale, the ability of regression models in explaining variables is the priority consideration, while the PLSR is superior to RFR in this respect. Under the premise of unknown priori knowledge of land surfaces, the RFR is more suitable for retrieving LAI than PLSR, but land surface priori knowledge is still necessary. These findings provide the theoretical basis and methods for developing remotely sensing estimating LAI models adapted to various land surfaces. Further analysis is needed in applying the findings in more crop types, cultivars and growth stages.     

Laboratory and glasshouse evaluation of entomopathogenic fungi against the two-spotted spider mite, Tetranychus urticae (Acari: Tetranychidae), on tomato, Lycopersicon esculentum

Forty isolates of entomopathogenic fungi from six genera were assessed against the two-spotted spider mite, Tetranychus urticae, in a single dose, direct application laboratory bioassay on tomato leaflets. Only three isolates caused greater mortality than the control: these were Metarhizium anisopliae 442.99, Hirsutella spp. 457.99, and Verticillium lecanii 450.99. These isolates were assessed in a multiple dose bioassay, together with three isolates cultured from commercial biopesticides as follows: Beauveria bassiana 432.99 (cultured from 'Naturalis-L', Troy Biosciences, Phoenix, TX, USA); Hirsutella thompsonii 463.99 (cultured from 'Mycar', Abbott Laboratories USA); and V. lecanii 19.79 (used in 'Mycotal' Koppert BV, The Netherlands). Beauveria bassiana 432.99, H. thompsonii 463.99, M. anisopliae 442.99, and V. lecanii 450.99 were all pathogenic to T. urticae in this bioassay. In addition, it was found that the mortality caused by B. bassiana 432.99 and Naturalis-L was increased when the mites were exposed to tomato leaflets sprayed previously with conidia suspensions, compared to spraying the mites directly. In a glasshouse experiment, sprays of B. bassiana 432.99, H. thompsonii 463.99, M. anisopliae 442.99, V. lecanii 450.99 and Naturalis-L reduced T. urticae populations in a tomato crop grown according to commercial practice. Naturalis-L reduced T. urticae numbers by up to 97%. In a second glasshouse experiment, single sprays of Naturalis-L and the acaricide fenbutatin oxide (Torq) were compared as supplementary treatments to release of the predatory mite, Phytoseiulus persimilis. Supplementary sprays of fenbutatin oxide reduced the numbers of T. urticae nymphs (80% reduction), while Naturalis-L reduced numbers of T. urticae adults, nymphs and eggs (98% reduction in all three cases). It is concluded that Naturalis-L has the potential to be used against T. urticae on glasshouse tomato crops.