Insecticidal activity of Bacillus thuringiensis towards Agrotis exclamationis larvae-A widespread and underestimated pest of the Palearctic zone.

acillus thuringiensis is an entomopathogenic bacterium commonly used as a bioinsecticide against numerous invertebrate pests. However, the efficacy of this microbe has not yet been determined towards Agrotis exclamationis-a lepidopteran, polyphagous pest, widespread throughout the Palearctic zone. In this work we have detected very low susceptibility of A. exclamationis to B. thuringiensis commercial strains, used as microbial formulations in pest control. To investigate this matter, the biological activity of six selected (Cry1Aa, Cry1Ca, Cry1Ia, Cry2Ab, Cry9Ea and Vip3Aa), heterogously-expressed Bacillus thuringiensis insecticidal proteins has been assessed towards A. exclamationis. Only Cry9Ea and Vip3Aa caused significant mortality in the tested pest species, with LC50 values of 950 and 140 ng/cm2, respectively. The histopathological effects of Cry9Ea and Vip3Aa on A. exclamationis were determined. On the other hand, Cry1- and Cry2-type toxins, which are the main active molecules of the majority of currently-used B. thuringiensis-based biocontrol agents (including the commercial strains tested in this work), did not cause mortality in target insect, but only different levels of growth inhibition. Moreover, in the case of Cry1Ca and Cry1Ia hormesis has been observed-a phenomenon that may be disadvantageous in implementation of these proteins in pest management. The obtained results broaden the existing knowledge regarding B. thuringiensis insecticidal protein target range and depict variable susceptibility of A. exclamationis to different groups of Cry/Vip toxins. This work indicates Cry9Ea and Vip3Aa as good candidates for efficient biological control of A. exclamationis and possibly other Agrotinae and discusses the potential use of Vip3-type and Cry9-type insecticidal proteins as successful bioinsecticides.

A novel Bacillus thuringiensis Cry9Ea-like protein with high insecticidal activity towards Cydia pomonella larvae.

BACKGROUND: The host specificity of known Bacillus thuringiensis Cry and Vip pesticidal proteins still needs extensive investigation and the proteins currently used in crop protection are not effective against many pest species. Cydia pomonella (L.) is a widespread and economically important pest of apples, very difficult to control, since the larvae spend most of their life inside a fruit. Currently, large amounts of broad-spectrum, detrimental synthetic agents are used to combat this herbivore and therefore biopesticides with high activity against C. pomonella are very much needed. RESULTS: The toxicity of B. thuringiensis Cry9Ea along with five distinct pesticidal proteins (Cry1Aa, Cry1Ca, Cry1Ia, Cry2Ab and Vip3Aa) has been determined towards the first-instar larvae of C. pomonella. Cry9Ea has much higher activity than the remaining tested proteins (30-1200-fold lower LC50 ) and possibly is the most potent B. thuringiensis pesticidal protein bioassayed against C. pomonella so far. In contrast, Cry9Ea is not toxic towards Spodoptera exigua (Hübn.), indicating a potentially narrow spectrum of activity. Both insect species show high variability in susceptibility to the remaining Cry/Vip proteins. CONCLUSIONS: The obtained results extend the existing knowledge regarding B. thuringiensis pesticidal protein host range and indicate Cry9Ea as a promising candidate for successful biological control of C. pomonella. © 2020 Society of Chemical Industry.

TOXiTAXi: a web resource for toxicity of Bacillus thuringiensis protein compositions towards species of various taxonomic groups.

Bioinsecticides consisting of different sets of Bacillus thuringiensis (Bt) Cry, Cyt and Vip toxins are broadly used in pest control. Possible interactions (synergistic, additive or antagonistic) between these proteins can not only influence the overall efficacy of certain Bt-based bioinsecticide, but also raise questions regarding environmental safety. Here, we assemble, summarize and analyze the outcomes of experiments published over 30 years, investigating combinatorial effects among Bt Cry, Cyt and Vip toxins. We collected the results on 118 various two-to-five-component combinations that have been bioassayed against 38 invertebrate species. Synergism, additive effect and antagonism was indicated in 54%, 32% and 14% of experiments, respectively. Synergism was noted most frequently for Cry/Cyt combinations, followed by Cyt/Vip and Cry/Cry. In Cry/Vip combinations, antagonism is more frequent and higher in magnitude compared to other categories. Despite a significant number of tested Bt toxin combinations, most of them have been bioassayed only against one pest species. To aid the research on Bt pesticidal protein activity, we present TOXiTAXi ( http://www.combio.pl/toxitaxi/ ), a universal database and a dedicated web tool to conveniently gather and analyze the existing and future bioassay results on biocidal activity of toxins against various taxonomic groups.

Activity of vegetative insecticidal proteins Vip3Aa58 and Vip3Aa59 of Bacillus thuringiensis against lepidopteran pests.

Vegetative insecticidal proteins (Vips) secreted by some isolates of Bacillus thuringiensis show activity against insects and are regarded as insecticides against pests. A number of B. thuringiensis strains harbouring vip3A genes were isolated from different sources and identified by using a PCR based approach. The isolates with the highest insecticidal activity were indicated in screening tests, and their vip genes were cloned and sequenced. The analysis revealed two polymorphic Vip protein forms, which were classified as Vip3Aa58 and Vip3Aa59. After expression of the vip genes, the proteins were isolated and characterized. The activity of both toxins was estimated against economically important lepidopteran pests of woodlands (Dendrolimus pini), orchards (Cydia pomonella) and field crops (Spodoptera exigua). Vip3Aa58 and Vip3Aa59 were highly toxic and their potency surpassed those of many Cry proteins used in commercial bioinsecticides. Vip3Aa59 revealed similar larvicidal activity as Vip3Aa58 against S. exigua and C. pomonella. Despite 98% similarity of amino acid sequences of both proteins, Vip3Aa59 was significantly more active against D. pini. Additionally the effect of proteolytic activation of Vip58Aa and Vip3Aa59 on toxicity of D. pini and S. exigua was studied. Both Vip3Aa proteins did not show any activity against Tenebrio molitor (Coleoptera) larvae. The results suggest that the Vip3Aa58 and Vip3Aa59 toxins might be useful for controlling populations of insect pests of crops and forests.

Persistence of the spores of B. thuringiensis subsp. kurstaki from Foray bioinsecticide in gleysol soil and on leaves.

The aim of this study was to determine how long the spores of Bacillus thuringiensis subsp. kurstaki HD-1 from Foray bioinsecticide persist in soil and on leaf surface after application of the bioinsecticide in an oak forest. Foray 04 UL was sprayed over a 195-hectare oak forest on the Krotoszyn Plateau in Poland. B. thuringiensis was isolated from soil samples and tree leaves taken from randomly chosen sites. B. thuringiensis subsp. kurstaki HD-1 in the samples was identified upon clonal analysis of the cultured isolates by using the RAPD method. One month after Foray spraying, the number of B. thuringiensis increased in soil and decreased on leaf surface comparing to the number estimated two days after the application. The reduction in the number of B. thuringiensis was noted six months after the pesticide application and the number was decreasing during the following months. No B. thuringiensis was noted on leaf surface one year after Foray spraying and in soil after one and a half years. The study showed that B. thuringiensis spores from biopesticide can survive in the forest environment; however, relatively short persistence time does not pose environmental risk.

Insecticidal activity of Bacillus thuringiensis strains isolated from soil and water.

We attempted to search novel Bacillus thuringiensis strains that produce crystals with potential utility in plant protection and with higher activity than strains already used in biopesticide production. Seven B. thuringiensis soil and water isolates were used in the research. We predicted the toxicity of their crystals by cry gene identification employing PCR method. The isolate MPU B63 with interesting, according to us, genes content was used in evaluating its crystal toxicity against Cydia pomonella caterpillars. The strain MPU B63 was cultured from water sample and had cry1Ab, cry1B, and cry15 genes. The LC50 crystals of MPU B63 were compared to LC50 of commercial bioinsecticide Foray determined against C. pomonella (codling moth). The activity of MPU B63 inclusions against codling moth larvae was approximately 24-fold higher than that of Foray. The results are a promising introduction for further study evaluating the potential usefulness of isolate MPU B63 crystals in plant protection.