Coexistence of cry9 with the vip3A Gene in an Identical Plasmid of Bacillus thuringiensis Indicates Their Synergistic Insecticidal Toxicity

Abstract

Bacillus thuringiensis (Bt) strains may express several insecticidal proteins with synergistic features, achieving high insecticidal toxicity and delaying development of resistance in insect pests. Previous work showed that Cry9Aa and Vip3Aa proteins present synergistic activity against Chilo suppressalis. In this study, genome-wide analysis of 489 Bt genomes revealed that cry9A was associated with the vip3A gene in seven Bt strains. Among all Bt genomes analyzed, not a single strain was found to have the cry9A gene alone without the presence of the vip3A gene. The complete genome sequencing of two Bt strains, 4AP1 and 4AO1, revealed that cry9A and vip3A genes were located in the same plasmid in both strains. The genome context analysis suggested a recombination mechanism responsible for the insertion of the cry9A gene into the plasmid containing vip3A. The coexistence of Cry9A with Vip3A proteins in strain 4AP1 was confirmed by liquid chromatography−tandem mass spectrometry and western blot analyses. Furthermore, another Cry9 protein codified by the gene in the identical plasmid also showed synergistic activity with the Vip3A protein. Overall, our results support that cry9 genes coexisted with vip3A and that complete genome sequencing combined with protein expression analysis may be used to identify associations of insecticidal proteins with potential synergistic toxicity.

Bacillus thuringiensis Cry9Aa Insecticidal Protein Domain I Helices α3 and α4 Are Two Core Regions Involved in Oligomerization and Toxicity

“…33 In addition, it was shown that cry9 and vip3 genes coexist in different Bt strains, indicating their coevolution for insecticidal activity. 33,35 This study on the mechanism of action of Cry9Aa will help to further improve the use of this insecticidal protein to enhance its toxicity and counter insect resistance.…”

“…xylostella strains and Cry9Cb1 toxin against Cry1Fa-resistant Ostrinia furnacalis indicated that Cry9 toxins exhibited no cross-resistance with Cry1 proteins. , Binding assays showed that Cry9Ca and Cry9E do not share receptors with Cry1Ab in different insects, suggesting that Cry9 proteins could be useful for resistance management. , On the other hand, Cry9Aa showed synergistic insecticidal activity when tested together with Cry1Ca or Vip3Aa against some lepidopteran larvae, such as Chilo suppressalis or Helicoverpa armigera. , Furthermore, the synergistic effect between Cry9Aa and Vip3Aa was shown to be based on a specific interaction between these two toxins . In addition, it was shown that cry9 and vip3 genes coexist in different Bt strains, indicating their coevolution for insecticidal activity. , This study on the mechanism of action of Cry9Aa will help to further improve the use of this insecticidal protein to enhance its toxicity and counter insect resistance.…”

Stability is essential for insecticidal activity of Vip3Aa toxin against Spodoptera exigua

“…Vegetative insecticidal proteins 3A (Vip3A), secreted from Bacillus thuringiensis (Bt) during their vegetative growth stage, are important insecticidal proteins that show toxicity against a wide variety of lepidopteran pests (Chakrabarty et al, 2020;Gupta et al, 2021). It was worth mentioning that Vip3A proteins displayed synergistic effect when coupling with Cry toxins for the control of agricultural pests (Wang et al, 2020;Lemes et al, 2014). It was also reported that Vip3A proteins were very active against insect species of Spodoptera, which showed low susceptibility to Cry proteins (Zhu et al, 2006; Baranek et al, 2015;Song et al, 2016).…”

IDOPS, a Profile HMM-Based Tool to Detect Pesticidal Sequences and Compare Their Genetic Context

“…Surrounding elements often include chaperones, crystallization domains, mobile elements, transporters, prophages, and virulence factors (Koni and Ellar, 1993 ; Shao et al, 2001 ; Elleuch et al, 2016 ; Adalat et al, 2017 ; Fayad et al, 2020 ; Lechuga et al, 2020 ). Moreover, the arrangement and distribution of such elements across genomes reveal crucial details about toxin functionality, host adaptation, diversification, and evolution of biopesticides (Khasdan et al, 2007 ; Peng et al, 2015 ; Ruffner et al, 2015 ; Fiedoruk et al, 2017 ; Zheng et al, 2017 ; Fayad et al, 2020 ; Wang et al, 2020 ). Hence, a more exhaustive approach would not only detect pesticidal sequences but also enable comparative genomics analysis of the candidate toxin in order to characterize the complete expression unit.…”