Peptide-derived broad-reacting antisera against tospovirus NSs-protein

Sequence alignments of tospovirus species of serogroup I to IV revealed a s tretch of 24 amino acids at the C terminus of the non-structural protein NS s with a highly conserved sequence. Based on this sequence the 24 amino aci ds peptide YFLSKTLEVLPKNLQTMSYLDSIQC was synthesized and used to raise anti sera in two rabbits. The specificity of the antisera against NSs from infec ted plants was confirmed with Western blots and by immunogold labelling and electron microscopy. These antisera detected tospovirus isolates of serogr oup I to III in antigen-coated plate ELISA and Western blots but failed to detect isolates of serogroup IV. Epitope scanning using overlapping octopep tides composing the peptide suggested that the antisera contained antibodie s against two different epitopes. Strongly reacting peptides were found at the C-terminus of the original peptide sequence when probing with one of th e antisera. in this part the sequence was homologous to serogroup I, II and III, with all deviations from serogroup IV located here. Additional octope ptides, based on this region, synthesized with sequence modifications back to the serogroup IV sequence in all possible combinations, had low reactivi ty. However two of the modified peptides with partly restored serogroup IV sequences revealed promising reactivity and could be suitable to raise an a ntiserum with broader reactivity, including serogroup IV. (C) 2000 Elsevier Science B.V. All rights reserved.