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Molecular Plant Pathology

Mutagenic Analysis and Localization of a Highly Conserved Epitope Near the Amino Terminal End of the Citrus Tristeza Closterovirus Capsid Protein. H. R. Pappu, Plant Pathology Department, P. O. Box 110680, University of Florida, Gainesville, 32611-0680, Present address: Department of Plant Pathology, University of Georgia, Coastal Plain Experiment Station, P.O. Box 748, Tifton 31793-0748; S. S. Pappu(2), T. Kano(3), M. Koizumi(4), M. Cambra(5), P. Moreno(6), H.-J. Su(7), S. M. Garnsey(8), R. F. Lee(9), and C. L. Niblett(10). (2)(10) Plant Pathology Department, P. O. Box 110680, University of Florida, Gainesville, 32611-0680; (3)(4)Division of Plant Protection, Fruit Tree Research Station, 2-1 Fujimoto, Tsukuba, Japan; Fruit Tree Research Station, Okitsu, Shimizu, Shizouka 424-02, Japan; (5)(6)IVIA, Moncada-Valencia, Spain; (7)Department of Plant Pathology and Entomology, National Taiwan University, Republic of China; (8)USDA-ARS, Horticultural Research Laboratory, 2120 Camden Road, Orlando, FL 32803; (9)Citrus Research and Education Center, Lake Alfred, FL 33850. Phytopathology 85:1311-1315. Accepted for publication 20 June 1995. Copyright 1995 The American Phytopathological Society. DOI: 10.1094/Phyto-85-1311.

The monoclonal antibody (MAb) 3DF1 is the first commercially available citrus tristeza closterovirus (CTV)-specific MAb. It detects a broad spectrum of CTV isolates from various parts of the world. To precisely map the antigenic determinant recognized by 3DF1, the capsid protein (CP) genes of four 3DF1-nonreactive isolates were cloned as complementary DNA and their nucleotide sequences determined. Comparison of the deduced CP sequences of the four nonreactive isolates with those of previously sequenced 3DF1-reactive isolates revealed differences at three positions near their amino terminal ends. The amino acids Asp-2, Lys-13, and Phe-28 were conserved in all the 3DF1-reactive isolates, but they were replaced by Gly, Thr/Asp, and Tyr, respectively, in the CPs of the nonreactive isolates. Site-specific mutations were introduced into the cloned CP genes of the 3DF1-nonreactive isolate B215 and the 3DF1-reactive isolate T36. The serological reactivities of the wild-type and mutant CPs of B215 and T36 expressed as recombinant fusion proteins in Escherichia coli were evaluated by Western blot analysis. A point mutation (A?G) resulting in an Asp?Gly change at amino acid position 2 of the CP of isolate T36 abolished the reactivity with the MAb, whereas a reverse mutation resulting in a Gly?Asp change at the same position conferred reactivity on the CP of the nonreactive B215 isolate. The implications of the observed antigenic diversity on virus detection are discussed.