PCR amplification of the spliced leader gene for the diagnosis of trypanosomatid parasites of plants and insects in methanol-fixed smears.
A PCR-based method was adapted for the amplification of DNA from methanol-fixed smears of insects and plants parasitized by trypanosomatids. The PCR target was the multicopy spliced leader (SL) gene. Amplicons were hybridized with an oligonucleotide probe (SL3') specific for Phytomonas. The method has the advantage of dispensing with the cultivation of parasites, many of which are very fastidious or non-cultivable. The technique was applied to archival glass slides and to newly collected material. It proved to specific for Phytomonas spp., enabling their detection in plants and insects. Sequence comparison of the amplicons obtained revealed the existence of different strains/species of Phytomonas circulating among diseased palsms and fruit.
Department of Parasitology, University of São Paulo, Brazil.
M G
MG
Serrano
M
M
Campaner
G A
GA
Buck
M M
MM
Teixeira
E P
EP
Camargo
D000818:Animals; D001483:Base Sequence; D004274:DNA, Recombinant / Q000235:genetics; D017125:Genes, Protozoan / Q000235:genetics*; D007313:Insecta / Q000469:parasitology*; D008969:Molecular Sequence Data; D010944:Plants / Q000469:parasitology*; D016133:Polymerase Chain Reaction / Q000379:methods; D020038:RNA, Spliced Leader / Q000235:genetics*; D013045:Species Specificity; D014351:Trypanosomatina / Q000235:genetics*
D004274:DNA, Recombinant; D020038:RNA, Spliced Leader