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Characterization of the resistance to Vip3Aa in Helicoverpa armigera from Australia and the role of midgut processing and receptor binding.

Chakroun M, Banyuls N, Walsh T, Downes S, James B, Ferré J - Sci Rep (2016)

Bottom Line: We found that, although relatively high, the frequency of alleles for resistance to Vip3Aa in field populations of H. armigera in Australia did not increase over the past four seasons until 2014/15.Vip3Aa-resistance does not confer cross-resistance to Cry1Ac or Cry2Ab.Proteolytic conversion of Vip3Aa protoxin into the activated toxin rendered the same products, though it was significantly slower in resistant insects.

View Article: PubMed Central - PubMed

Affiliation: ERI of Biotechnology and Biomedicine (BIOTECMED), Universitat de València, 46100 Burjassot, Spain.

ABSTRACT
Crops expressing genes from Bacillus thuringiensis (Bt crops) are among the most successful technologies developed for the control of pests but the evolution of resistance to them remains a challenge. Insect resistant cotton and maize expressing the Bt Vip3Aa protein were recently commercialized, though not yet in Australia. We found that, although relatively high, the frequency of alleles for resistance to Vip3Aa in field populations of H. armigera in Australia did not increase over the past four seasons until 2014/15. Three new isofemale lines were determined to be allelic with previously isolated lines, suggesting that they belong to one common gene and this mechanism is relatively frequent. Vip3Aa-resistance does not confer cross-resistance to Cry1Ac or Cry2Ab. Vip3Aa was labeled with (125)I and used to show specific binding to H. armigera brush-border membrane vesicles (BBMV). Binding was of high affinity (Kd = 25 and 19 nM for susceptible and resistant insects, respectively) and the concentration of binding sites was high (Rt = 140 pmol/mg for both). Despite the narrow-spectrum resistance, binding of (125)I-labeled Vip3Aa to BBMV of resistant and susceptible insects was not significantly different. Proteolytic conversion of Vip3Aa protoxin into the activated toxin rendered the same products, though it was significantly slower in resistant insects.

No MeSH data available.


Related in: MedlinePlus

Kinetics of the proteolytic processing of Vip3Aa incubated with midgut juice from H. armigera larvae.Incubations were performed at 30 °C and 0.1% of midgut juice total protein referred to Vip3Aa protein. Samples from susceptible (a) and resistant (b) insects were subjected to SDS-PAGE at different time intervals and the bands of protoxin (89 kDa, solid lines) and activated protein (62 kDa, broken lines) were quantified by densitometry (c). GR (susceptible) (●) and SP85 (resistant) (○) colonies. Data points represent the mean of three replicates with the standard error indicated by error bars.
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f1: Kinetics of the proteolytic processing of Vip3Aa incubated with midgut juice from H. armigera larvae.Incubations were performed at 30 °C and 0.1% of midgut juice total protein referred to Vip3Aa protein. Samples from susceptible (a) and resistant (b) insects were subjected to SDS-PAGE at different time intervals and the bands of protoxin (89 kDa, solid lines) and activated protein (62 kDa, broken lines) were quantified by densitometry (c). GR (susceptible) (●) and SP85 (resistant) (○) colonies. Data points represent the mean of three replicates with the standard error indicated by error bars.

Mentions: Since Vip3Aa is found in the protoxin form in cotton leaves47, we searched for differences in its conversion to the activate form between the susceptible and resistant insects. When midgut juice of GR and SP85 was incubated with Vip3Aa protoxin, many proteolytic products were obtained but no difference in the band profile between the two colonies was observed (Fig. 1a,b). The major proteolysis products were the 62 and the 20 kDa fragments in both cases. The kinetic analysis of the 89 kDa activation and the 62 kDa fragment formation showed a difference in the processing rate between the susceptible and the resistant H. armigera colonies (Fig. 1c). The processing of the 89 kDa protoxin was faster in the susceptible colony. After 15 min the protoxin completely disappeared with the midgut juice from the susceptible insects, however, with SP85 there was 31% residual protoxin which was completely activated after 60 min incubation.


Characterization of the resistance to Vip3Aa in Helicoverpa armigera from Australia and the role of midgut processing and receptor binding.

Chakroun M, Banyuls N, Walsh T, Downes S, James B, Ferré J - Sci Rep (2016)

Kinetics of the proteolytic processing of Vip3Aa incubated with midgut juice from H. armigera larvae.Incubations were performed at 30 °C and 0.1% of midgut juice total protein referred to Vip3Aa protein. Samples from susceptible (a) and resistant (b) insects were subjected to SDS-PAGE at different time intervals and the bands of protoxin (89 kDa, solid lines) and activated protein (62 kDa, broken lines) were quantified by densitometry (c). GR (susceptible) (●) and SP85 (resistant) (○) colonies. Data points represent the mean of three replicates with the standard error indicated by error bars.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4837340&req=5

f1: Kinetics of the proteolytic processing of Vip3Aa incubated with midgut juice from H. armigera larvae.Incubations were performed at 30 °C and 0.1% of midgut juice total protein referred to Vip3Aa protein. Samples from susceptible (a) and resistant (b) insects were subjected to SDS-PAGE at different time intervals and the bands of protoxin (89 kDa, solid lines) and activated protein (62 kDa, broken lines) were quantified by densitometry (c). GR (susceptible) (●) and SP85 (resistant) (○) colonies. Data points represent the mean of three replicates with the standard error indicated by error bars.
Mentions: Since Vip3Aa is found in the protoxin form in cotton leaves47, we searched for differences in its conversion to the activate form between the susceptible and resistant insects. When midgut juice of GR and SP85 was incubated with Vip3Aa protoxin, many proteolytic products were obtained but no difference in the band profile between the two colonies was observed (Fig. 1a,b). The major proteolysis products were the 62 and the 20 kDa fragments in both cases. The kinetic analysis of the 89 kDa activation and the 62 kDa fragment formation showed a difference in the processing rate between the susceptible and the resistant H. armigera colonies (Fig. 1c). The processing of the 89 kDa protoxin was faster in the susceptible colony. After 15 min the protoxin completely disappeared with the midgut juice from the susceptible insects, however, with SP85 there was 31% residual protoxin which was completely activated after 60 min incubation.

Bottom Line: We found that, although relatively high, the frequency of alleles for resistance to Vip3Aa in field populations of H. armigera in Australia did not increase over the past four seasons until 2014/15.Vip3Aa-resistance does not confer cross-resistance to Cry1Ac or Cry2Ab.Proteolytic conversion of Vip3Aa protoxin into the activated toxin rendered the same products, though it was significantly slower in resistant insects.

View Article: PubMed Central - PubMed

Affiliation: ERI of Biotechnology and Biomedicine (BIOTECMED), Universitat de València, 46100 Burjassot, Spain.

ABSTRACT
Crops expressing genes from Bacillus thuringiensis (Bt crops) are among the most successful technologies developed for the control of pests but the evolution of resistance to them remains a challenge. Insect resistant cotton and maize expressing the Bt Vip3Aa protein were recently commercialized, though not yet in Australia. We found that, although relatively high, the frequency of alleles for resistance to Vip3Aa in field populations of H. armigera in Australia did not increase over the past four seasons until 2014/15. Three new isofemale lines were determined to be allelic with previously isolated lines, suggesting that they belong to one common gene and this mechanism is relatively frequent. Vip3Aa-resistance does not confer cross-resistance to Cry1Ac or Cry2Ab. Vip3Aa was labeled with (125)I and used to show specific binding to H. armigera brush-border membrane vesicles (BBMV). Binding was of high affinity (Kd = 25 and 19 nM for susceptible and resistant insects, respectively) and the concentration of binding sites was high (Rt = 140 pmol/mg for both). Despite the narrow-spectrum resistance, binding of (125)I-labeled Vip3Aa to BBMV of resistant and susceptible insects was not significantly different. Proteolytic conversion of Vip3Aa protoxin into the activated toxin rendered the same products, though it was significantly slower in resistant insects.

No MeSH data available.


Related in: MedlinePlus