[Primers design and optimization of PCR and nested-PCR assays for the specific detection of Tritrichomonas foetus].
Tritrichomonas foetus is a pathogenic protozoan that causes a venereal disease in cattle known as bovine genital tricomonosis. In spite of the efficacy to recognize the target genomic DNA, the protocols so far developed for the diagnosis of this organism by PCR promote some inespecific amplifications or they are unable to discriminate T. foetus against other species within the genus. The objective of this study was to assess and optimize PCR and nested-PCR assays for the specific diagnosis of T. foetus, using novel primers selected from the alignment of sequences of the genes 18S rRNA, 5.8S rRNA, 28S rRNA and of the internal transcribed spacers of the rDNA (ITS1 and ITS2). A pair of primers was constructed for the genus-specific amplification of a 648 bp fragment and two others to amplify T. foetus species-specific fragments of 343 and 429 bp. No cross amplification was observed against Bos taurus genomic DNA neither against the DNA of usual bovine genital pathogens. Both, single and nested-PCR assays, presented analytical sensitivity to detect at least two T. foetus organisms.
Fernandes
Paula Rogério
PR
Programa de Pós-Graduação em Ciência Animal, Universidade Federal de Goiás, Goiânia, GO, Brasil. paularogeriof@gmail.com
Da Silva
Andréa Caetano
AC
Gambarini
Maria Lúcia
ML
Linhares
Guido Fontgalland C
GF
D000818:Animals; D017931:DNA Primers; D016133:Polymerase Chain Reaction / Q000379:methods*; D016835:Tritrichomonas foetus / Q000235:genetics* / Q000302:isolation & purification*
D017931:DNA Primers