41st Annual Meeting of the Society for Invertebrate Pathology
and 9th International Conference on Bacillus thuringiensis

Venom from the endoparasitic wasp, Pimpla hypochondriaca, contains factors with anti-haemocyte and immunosuppressive properties. The gene for one such factor (vpb) has been cloned and recombinant protein produced. Bio-assays utilising VPB were performed and indicated that introduction of this immunosuppressive protein into the haemocoel of two lepidopteran pests, increases their susceptibility to the biological control agent, Bacillus thuringiensis. The potential for improving the efficacy of Bt through suppression of pest immune responses is discussed.

Becker Underwood produce and market a number of micro- and macro-biological control products. Our micro-biological products are based on entomopathogenic fungi (EPF) and beneficial bacteria. Our 13 macro-biological products contain entomopathogenic nematodes (EPN) and are manufactured in the worlds largest nematode production facility in Littlehampton, UK. Green Guard (Metarhizium anisopliae), a micro-biological product and is sold throughout Australia for the control of locusts and grasshoppers. Nemaslug (Phasmarhabditis hermaphrodita) is a macro-biological, this species of nematode was originally isolated from Rothamsted research station, UK and is now widely used throughout Europe to control slugs. This paper will detail how these products have gone from discovery, through laboratory screening and to successful commercialisation. Selling products into over 65 countries, Becker Underwood faces a variety of different registration requirements. Registration requirements are arguably the greatest challenge to companies for the commercialisation of biological control products. Other important issues including mass production, quality control, shipping and applicator training will also be discussed as these can be more demanding than chemical-based plant protection products.

Nematicides have been used to control plant parasitic nematodes, but over the last decade legislative measures have restricted their use as they are amongst the most toxic compounds used in agriculture. Therefore alternative approaches are being explored. These range from the development of resistant varieties and genetic engineering to the development of biological control agents. The life-cycle of plant parasitic nematodes includes two levels of trophic interaction, one between the plant and the parasitic nematode, and another between the nematode and any microbial pathogens present in the soil. Therefore the cuticle is an organ that provides a barrier between the nematode and its environment. The cuticles of plant parasitic nematodes have exhibit inter and intra specific variability with respect to the nematode hyperparasite Pasteuria penetrans. Endospores of this Gram positive obligate bacterium can adhere to and infect one strain of nematode but not another. This variation appears to be as great in parthenogenetically reproducing plant parasitic nematodes as in amphimictic reproducing groups. The implications of this variation for the population dynamics of the hyperparasite will be discussed.

The microsporidium Vairimorpha invictae and parasitic flies in the genus Pseudacteon have established or are being considered for release in the USA for the biological control of fire ants, Solenopsis invicta. Pseudacteon flies oviposit into adult fire ants, where maggots that eclose from eggs migrate to the ants’ head, pupate, and eventually decapitate the host. The compatibility of these biocontrol agents was examined by determining if the parasitic fly, P. obtusus would become infected if it developed in the microsporidia-infected fire ants. P. obtusus were allowed to oviposit and develop in V. invictae infected S. invicta in the laboratory. There was no evidence of microsporidian infection in P. obtusus adults that developed in heads that were matched to infected bodies (n=39). S. invicta bodies that could not be matched with their decapitated heads had an estimated infection rate of 87%. V. invictae was not detected in any of the P. obtusus that emerged from unmatched heads (N=318). These results further defined the host specificity of V. invictae and indicated that V. invictae will not directly interfere with P. obtusus parasitism.

The efficacy of different treatment applications with P. formosoroseus blastospore formulation (Pfr strain 3581) was assessed for potent to manage the citrus weevil, Diaprepes abbreviatus, DRW. Pfr was applied topically on the larvae and adults at a rate of 107 blastospores/mL and incubated in original rearing cups in the dark at 25˚C for 2 – 3 weeks. Another technique assessed feeding DRW detached citrus leaves previously sprayed with blastospores, after which DRW was incubated in Petri dish chambers at 25˚C for 2 – 3 weeks under 16 hr photophase. In the greenhouse, soil drench experiments were conducted to assess the effectiveness in killing the larvae. Each pot containing a citrus seedling drenched with 100 mL suspensions of Pfr at 107 blastospores/mL and then drenched with 400, 900 and 1,400 mL of water. Larval mortality due to the infection of Pfr was assessed after 2 weeks. Pfr may provide an added management tool against these difficult to control citrus pests.

Efficacy of Paecilomyces fumosoroseus blastospores (Deuteromycotina: Hyphomycetes) on yellow tags vs. citrus leaves to manage Asian citrus psyllids, Diaphorina citri (Psyllidae: Hemiptera) were compared and horizontal transmission by psyllids among leaves assessed using a detached leaf bioassay. Psyllids were tested individually on either 4 leaf sections or 3 leaf sections and a treated yellow plastic tag. Treatments were citrus leaf sections or yellow tag of similar size (~100 mm2) sprayed with P. fumosoroseus. Proportions of leaf sections were sprayed as follows: 25%, 50%, 75%, 100% compared to a yellow tag. Distilled water served as a control. A Fungal Development Index was used to determine infection rate once the insect had died and showed signs of mycosis. The yellow tag treatment was equally effective as the other leaf treatments in the rate of infection and spread infection more rapidly in psyllids compared to the 25% leaf treatment. Adults began to mycose at ~4-5 days post-release. As the inoculum increased for all leaf treatments the infection rate also increased. For all fungal treatments there was 100% horizontal transmission of P. fumosoroseus spores to all non-treated leaf sections. Inoculated yellow plastic tags may serve as an autodissemination technique for managing psyllid populations in citrus.

The fall webworm, Hyphantria cunea (Lepidoptera: Arctiidae) is dangerous quarantine pest damaging the agriculture crops, forest and ornamental plants, it distributed through the territory of West Georgia and Black Sea Coast. The insect mainly inhabits in the populated area – home sites, communities suburban parks and places of mass restoring where by the viewpoint of sanitation-hygiene the using chemical pesticides are prohibited. Generally the urban horticulture is under the threat of fall webworm. In this connection it is necessary the use of environmentally safe means to plant protection from this dangerous pest. The biological control potential of different means: bacterial - XenTari DF, Dipel, fungal - BotaniGard ES (Project – GRDF-GEB2-3337-TB-04, USA) and entomopathogenic nematode (EPN) introduced from Israel (CDR-CAR Project CA CA22-007) – Steinernema feltiae were tested against 2nd, 3rd instars larvae and pupae of H.cunea in laboratory and fields. Infectivity of H.cunea by the suspensions (0.7%) XenTari DF and BotaniGard ES on 7 days have caused 96-100% mortality. The mix infection at the reduced concentrations of microbial means with entomopathogenic nematodae, S.feltiae (1.500 unit/ml) on 3 days has caused 100% mortality of larvae, which may serve for cultivation of nematodes. Biological control will take the important place in IPM.

Baculoviruses can be combined with other entomopathogens to achieve improved biological control of insect pests. Understanding the population ecology of interacting pathogens and specifically the transmission dynamics in pathogen combinations will assist in predicting the outcome of integrated biological control strategies. We tested the hypothesis that the presence of Bacillus thuringiensis subsp. kurstaki (Btk)in manipulated cabbage moth (Mamestra brassicae) field populations would affect the transmission dynamics of Panolis flammea nucleopolyhedrovirus (PaflNPV). The combination of spraying Btk with PaflNPV resulted in first generation larvae being more likely to be infected with virus when Btk was present. The acquisition of baculovirus infection as exposure time increased was found to be highly non-linear. In the presence of Btk the number of insects that were able to escape NPV infection was reduced through a combination of changes in host feeding behaviour and delay in onset of host developmental resistance. Viral cadavers in the presence of Btk produced significantly lower viral yields compared to those in the absence of Btk. When second generation larvae were exposed to these viral cadavers in situ, there were significant reductions in subsequent viral mortality. The impact of these results for season long Lepidoptera control are discussed.

Indigenous isolates of the entomopathogenic fungus Metarhizium anisopliae were screened for virulence against the wireworm species Agriotes obscurus, A. lineatus and A. sputator. In 2006, thirteen isolates were tested by dipping larvae into blastospore suspensions (1x107 spores/mL). For the most virulent isolates, a maximum of 38.3%, 30.0%, and 30.4% of the A. obscurus, A. lineatus and A. sputator larvae, respectively, were infected after nine weeks. In 2008, a similar bioassay was performed using conidial (1x107 spores/mL) instead of blastospore suspensions. For the most virulent isolates infection rates were 80.0 %, 33.3%, and 40.0% for A. obscurus, A. lineatus and A. sputator larvae, respectively, after nine weeks. The results suggest that the M. anisopliae isolates may be particularly useful to control A. obscurus, while control of the other two wireworm species may be less efficient.

Cameraria ohridella Deschka et Dimic, is an important invasive pest of Aesculus hippocastanum in Europe. Present methods of its control are based on application of non-selective insecticides and composting or burning of leaf litter. Our project is aimed at entomopathogenic fungi as potential candidates for biological control of this pest. In the present work we report the first record of Beauveria bassiana (Bals.) Vuill. as an entomopathogenic fungus of C. ohridella. B. bassiana was found and isolated from hibernating pupae of C. ohridella. Samples of the horse chestnut leaves with diapausing C. ohridella pupae were collected in autumn 2007 at Ceske Budejovice, South Bohemia, the Czech Republic. The leaves were dissected and the pupae were individually placed into the 23°C and 95% R.H. Development of mycosis on pupae was frequently observed but only a few pupae were covered by mycelia with conidiophores and conidia of B. bassiana. The strains of the fungus were isolated from individual infected pupae and deposited in CCEFO (Culture Collection of Entomopathogenic Fungi Olesna) in the Czech Republic. The virulence of the isolated strains is being evaluated. This work was supported by the MSMT grant No. 2B06005.