Peptide-derived broad-reacting antisera against tospovirus NSs-protein.
Sequence alignments of tospovirus species of serogroup I to IV revealed a stretch of 24 amino acids at the C terminus of the non-structural protein NSs with a highly conserved sequence. Based on this sequence the 24 amino acids peptide YFLSKTLEVLPKNLQTMSYLDSIQC was synthesized and used to raise antisera in two rabbits. The specificity of the antisera against NSs from infected plants was confirmed with Western blots and by immunogold labelling and electron microscopy. These antisera detected tospovirus isolates of serogroup I to III in antigen-coated plate ELISA and Western blots but failed to detect isolates of serogroup IV. Epitope scanning using overlapping octopeptides composing the peptide suggested that the antisera contained antibodies against two different epitopes. Strongly reacting peptides were found at the C-terminus of the original peptide sequence when probing with one of the antisera. In this part the sequence was homologous to serogroup I, II and III, with all deviations from serogroup IV located here. Additional octopeptides, based on this region, synthesized with sequence modifications back to the serogroup IV sequence in all possible combinations, had low reactivity. However two of the modified peptides with partly restored serogroup IV sequences revealed promising reactivity and could be suitable to raise an antiserum with broader reactivity, including serogroup IV.
Heinze
C
C
Institut für Angewandte Botanik, Universität Hamburg, Marseiller Str. 7, D-20355, Hamburg, Germany.
Roggero
P
P
Sohn
M
M
Vaira
A M
AM
Masenga
V
V
Adam
G
G
D000595:Amino Acid Sequence; D000818:Animals; D000914:Antibodies, Viral / Q000096:biosynthesis*; D015153:Blotting, Western; D017124:Conserved Sequence; D004797:Enzyme-Linked Immunosorbent Assay; D018604:Epitope Mapping; D007106:Immune Sera / Q000096:biosynthesis*; D007150:Immunohistochemistry; D016253:Microscopy, Immunoelectron; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry / Q000276:immunology*; D010947:Plants, Toxic; D011817:Rabbits; D016415:Sequence Alignment; D014026:Nicotiana / Q000821:virology; D017865:Tospovirus / Q000737:chemistry / Q000276:immunology* / Q000648:ultrastructure; D017361:Viral Nonstructural Proteins / Q000737:chemistry / Q000276:immunology*
D000914:Antibodies, Viral; D007106:Immune Sera; D010455:Peptides; D017361:Viral Nonstructural Proteins