Identification of Bacillus thuringiensis Strains for the Management of Lepidopteran Pests

ABSTRACTBacillus thuringiensisCry1Ab and Cry1Fa toxins are environmentally safe insecticides that control important insect pests.Spodoptera frugiperdais an important maize pest that shows low susceptibility to Cry1A toxins, in contrast to Cry1Fa, which is highly active against this pest and is used in transgenic maize forS. frugiperdacontrol. The β16 region from domain III of Cry1Ab has been shown to be involved in interactions with receptors such as alkaline phosphatase (ALP) or aminopeptidase (APN) in different lepidopteran insects. Alanine-scanning mutagenesis of amino acids of Cry1Ab β16 (509STLRVN514) revealed that certain β16 mutations, such as N514A, resulted in increased toxicity of Cry1Ab forS. frugiperdawithout affecting the toxicity for other lepidopteran larvae, such asManduca sextalarvae. Exhaustive mutagenesis of N514 was performed, showing that the Cry1Ab N514F, N514H, N514K, N514L, N514Q, and N514S mutations increased the toxicity towardS. frugiperda. A corresponding mutation was constructed in Cry1Fa (N507A). Toxicity assays of wild-type and mutant toxins (Cry1Ab, Cry1AbN514A, Cry1AbN514F, Cry1Fa, and Cry1FaN507A) against fourS. frugiperdapopulations from Mexico and one from Brazil revealed that Cry1AbN514A and Cry1FaN507A consistently showed 3- to 18-fold increased toxicity against four of fiveS. frugiperdapopulations. In contrast, Cry1AbN514F showed increased toxicity in only two of theS. frugiperdapopulations analyzed. The mutants Cry1AbN514A and Cry1AbN514F showed greater stability to midgut protease treatment. In addition, binding analysis of the Cry1Ab mutants showed that the increased toxicity correlated with increased binding to brush border membrane vesicles and increased binding affinity forS. frugiperdaALP, APN, and cadherin receptors.IMPORTANCESpodoptera frugiperdais the main maize pest in South and North America and also is an invasive pest in different African countries. However, it is poorly controlled byBacillus thuringiensisCry1A toxins expressed in transgenic crops, which effectively control other lepidopteran pests. In contrast, maize expressing Cry1Fa is effective in the control ofS. frugiperda, although its effectiveness is being lost due to resistance evolution. Some of the Cry1Ab domain III mutants characterized here show enhanced toxicity forS. frugiperdawithout loss of toxicity toManduca sexta. Thus, these Cry1Ab mutants could provide useful engineered toxins that, along with other Cry toxins, would be useful for developing transgenic maize expressing stacked proteins for the effective control ofS. frugiperdaand other lepidopteran pests in the field.

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Microbial control agent containing a Sri Lankan Bacillus thuringiensis isolate for control of Lepidopteran pests

Planta Medica ◽

10.1055/s-0029-1234383 ◽

2009 ◽

Vol 75 (09) ◽

Author(s):

R Samarasekera ◽

DA Siriwardhana ◽

L HerathI ◽

D Ahangama

Keyword(s):

Bacillus Thuringiensis ◽

Microbial Control ◽

Control Agent ◽

Sri Lankan ◽

Lepidopteran Pests

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Synergy of Lepidopteran Nucleopolyhedroviruses AcMNPV and SpliNPV with Insecticides

Insects ◽

10.3390/insects11050316 ◽

2020 ◽

Vol 11 (5) ◽

pp. 316 ◽

Cited By ~ 1

Author(s):

Beatriz Dáder ◽

Eduardo Aguirre ◽

Primitivo Caballero ◽

Pilar Medina

Keyword(s):

Bacillus Thuringiensis ◽

Integrated Pest Management ◽

Pest Management ◽

Spodoptera Littoralis ◽

Spodoptera Exigua ◽

Autographa Californica ◽

Antagonistic Effects ◽

Lepidopteran Pests ◽

Excellent Control ◽

Autographa Californica Multiple Nucleopolyhedrovirus

The joint use of baculoviruses and synthetic insecticides for integrated pest management requires the study of the additive, synergistic or antagonistic effects among them on pest mortality. Droplet bioassays were conducted with Autographa californica multiple nucleopolyhedrovirus (AcMNPV), Spodoptera littoralis nucleopolyhedrovirus (SpliNPV) and seven insecticides (azadirachtin, Bacillus thuringiensis, cyantraniliprole, emamectin, metaflumizone, methoxyfenozide and spinetoram) on Spodoptera exigua and Spodoptera littoralis. The lethal concentrations LC50 and LC95 were calculated through probit regressions. Then, the sequential feeding of insecticides and nucleopolyhedroviruses was studied. Larvae were provided with the LC50 of one insecticide, followed by the LC50 of one nucleopolyhedrovirus 24 h later. The inverse order was also conducted. The insecticide LC50 and LC95 were higher for S. littoralis than for S. exigua. AcMNPV showed greater toxicity on S. exigua than SpliNPV on S. littoralis. Emamectin showed synergy with AcMNPV when the chemical was applied first, and metaflumizone and AcMNPV were synergistic regardless of the order of application, both from the first day of evaluation. SpliNPV was synergistic with azadirachtin and emamectin when it was applied first, but synergy was reached after 12–13 days. Excellent control is possible with the LC50 of azadirachtin, emamectin and metaflumizone in combination with nucleopolyhedroviruses, and merits further study as a means of controlling lepidopteran pests.

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Bacillus thuringiensis Cry1Ab Domain III β-22 Mutants with Enhanced Toxicity to Spodoptera frugiperda (J. E. Smith)

Applied and Environmental Microbiology ◽

10.1128/aem.01580-20 ◽

2020 ◽

Vol 86 (22) ◽

Author(s):

Isabel Gómez ◽

Josue Ocelotl ◽

Jorge Sánchez ◽

Sotero Aguilar-Medel ◽

Guadalupe Peña-Chora ◽

...

Keyword(s):

Bacillus Thuringiensis ◽

Spodoptera Frugiperda ◽

Insect Pest ◽

Transgenic Maize ◽

Crop Damage ◽

Binding Interaction ◽

Alanine Scanning Mutagenesis ◽

Content Type ◽

Lepidopteran Pests ◽

Domain Iii

ABSTRACT The fall armyworm, Spodoptera frugiperda, is an invasive maize pest that has spread from the Americas into Africa and Asia and causes severe crop damage worldwide. Most populations of S. frugiperda show low susceptibility to Bacillus thuringiensis (Bt) Cry1Ab or Cry1Ac toxins, which have been proved to be effective against several other lepidopteran pests. In addition, S. frugiperda has evolved resistance to transgenic maize expressing Cry1Fa toxin. The specificity and toxicity of Cry toxins are determined by their binding to different larval midgut proteins, such as aminopeptidase N (APN), alkaline phosphatase (ALP), and cadherin (CAD), among other proteins, by means of exposed domain II loop regions and also by the domain III β-sheets β-16 and β-22. Here, we analyzed different Cry1Ab mutants with mutations in the domain III β-22 region. Alanine-scanning mutagenesis of this region revealed that all mutants showed increased toxicity against a nonsusceptible Cry1Ab S. frugiperda population. Further analysis of the mutant toxin Cry1AbS587A (bearing a mutation of S to A at position 587) revealed that, compared to Cry1Ab, it showed significantly increased toxicity to three other S. frugiperda populations from Mexico but retained similar toxicity to Manduca sexta larvae. Cry1AbS587A bound to brush border membrane vesicles (BBMV), and its higher toxicity correlated with higher binding affinities to APN, ALP, and CAD recombinant proteins. Furthermore, silencing the expression of APN1 and CAD receptors in S. frugiperda larvae by RNA interference (RNAi) showed that Cry1AbS587A toxicity relied on CAD expression, in contrast to Cry1Ab. These data support the idea that the increased toxicity of Cry1AbS587A to S. frugiperda is in part due to an improved binding interaction with the CAD receptor. IMPORTANCE Spodoptera frugiperda is an important worldwide pest of maize and rice crops that has evolved resistance to Cry1Fa-expressing maize in different countries. Therefore, identification of additional toxins with different modes of action is needed to provide alternative tools to control this insect pest. Bacillus thuringiensis (Bt) Cry1Ab and Cry1Ac toxins are highly active against several important lepidopteran pests but show varying and low levels of toxicity against different S. frugiperda populations. Thus, the identification of Cry1A mutants that gain toxicity to S. frugiperda and retain toxicity to other pests could be of great value to produce transgenic crops that resist a broader spectrum of lepidopteran pests. Here, we characterized Cry1Ab domain III β-22 mutants, and we found that a Cry1AbS587A mutant displayed increased toxicity against different S. frugiperda populations. Thus, Cry1AbS587A could be a good toxin candidate to produce transgenic maize with broader efficacy against this important insect pest in the field.

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