The Molecular Basis for the Antigenic Diversity of Citrus Tristeza Virus: Implications for Virus Detection

H. R. Pappu, S. S. Pappu, R. F. Lee, M. Cambra, P. Moreno, S. M. Garnsey

Abstract


Citrus tristeza virus (CTV) displays a wide range of seroiogical diversity. Strains have been reported from various countries that do not react with some of the CTV-specific monoclonal antibodies (MAb). We investigated the molecular basis for the emergence of these serotypes of CTV. While CTV-specific MAb MCA13 reacts predominantly with severe strains that cause quick decline and/or stem pitting, 3DF1 is a broad spec trum MAb that reacts with a majority of the CTV strains. To map the antigenic determinants recognized by these MAbs, capsid protein (CP) genes of several serologically and biologically di verse strains from different parts of the world were cloned and sequenced. Sequence comparisons showed that phenylalanine (Phe) at position 124 of the CP was conserved among all the MCA13-reactive strains. This was replaced by a tyrosine (Tyr) in MCA13-nonreactive strains. Sequence comparisons between 3DF1-reactive and nonreactive strains showed that amino acids Aspartic acid (Asp), Lysine, and Phe were conserved at positions 2,13 and 28 in the CPs of all the 3DF1-re active strains, respectively. These were replaced by Glycine, Threonine/Asp, and Tyr in 3DF1-nonreactive strains. The ami-no acid differences in both cases were due to single nucleotide changes in their respective codons. Mutations were intro duced that altered the codons in the cloned CP genes of selected strains and their effect on the reactivity of the two MAbs was evaluated by Western blot analysis. For MCA13, its reactivity dependent on the presence of phenylalanine residue at position 124, for 3DF1, the second amino acid, asp was found to be critical for its reactivity. Our data indicate that point mutations in the CP gene resulting in single amino acid substitutions can result in the loss of a viral epitope. This can drastically alter or eliminate the reactivity of a CTV-specific MAb. This provides the molecular basis for the existence/emergence of new sero types of CTV and indicates the necessity of using a combination of MAbs of differing specificities or a polyclonal antibody for the effective detection of diverse strains of CTV.

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Proc. Fla. State Hort. Soc.     ISSN 0886-7283