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Applied and Environmental Microbiology, August 2006, p. 5318-5324, Vol. 72, No. 8
0099-2240/06/$08.00+0     doi:10.1128/AEM.00219-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

Analyses of Cry1Ab Binding in Resistant and Susceptible Strains of the European Corn Borer, Ostrinia nubilalis (Hübner) (Lepidoptera: Crambidae)

Herbert A. A. Siqueira,¹ Joel González-Cabrera,² Juan Ferré,² Ronald Flannagan,³ and Blair D. Siegfried^1*

Department of Entomology, 202 Plant Industry Bldg., University of Nebraska, Lincoln, Nebraska 68583-0816,¹ Departamento de Genética, Facultad de CC Biológicas, Universidad de Valencia, 46100-Burjassot, Valencia, Spain,² Pioneer Hi-Bred International Inc., 7301 NW 62nd Ave., Johnston, Iowa 50131³

Received 27 January 2006/ Accepted 30 May 2006

Cry1Ab toxin binding analysis was performed to determine whether resistance in laboratory-selected Ostrinia nubilalis strains is associated with target site alteration. Brush border membrane vesicles were prepared using dissected midguts from late instars of susceptible and resistant strains (Europe-R and RSTT) of O. nubilalis. Immunoblot analysis indicated that three different proteins bound to Cry1Ab toxin and were recognized by an anticadherin serum. In a comparison of resistant and susceptible strains, reduced Cry1Ab binding was apparent for all three bands corresponding to cadherin-like proteins in the Europe-R strain, while reduced binding was apparent in only one band for the RSTT strain. Real-time analysis of Cry1Ab binding to gut receptors using surface plasmon resonance suggested slight differences in affinity in both resistant strains. Additional binding analysis was conducted using ¹²⁵I-labeled Cry1Ab, Cry1Ac, and Cry1Aa. Slight differences were again observed between the resistant and susceptible strains for Cry1Ab binding. However, when binding of ¹²⁵I-labeled Cry1Aa was tested, a 10-fold reduction in the concentration of binding sites was observed in the Europe-R strain. Expression of the O. nubilalis cadherin gene was similar in both the resistant and susceptible strains and did not account for differences in binding. In combination, the results of the present work suggest that differences in susceptibility to Cry1A toxins in the Europe-R strain of O. nubilalis are associated with altered receptor binding, although the precise nature of this mechanism is still uncertain.

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* Corresponding author. Mailing address: Department of Entomology, University of Nebraska, 202 Plant Industry Bldg., Lincoln, NE 68583. Phone: (402) 472-8714. Fax: (402) 472-4687. E-mail: bsiegfried1{at}unl.edu.

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Applied and Environmental Microbiology, August 2006, p. 5318-5324, Vol. 72, No. 8
0099-2240/06/$08.00+0     doi:10.1128/AEM.00219-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

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