Comparison of PCR methods for diagnosis of canine visceral leishmaniasis in conjunctival swab samples.
Four PCR assays for detection of Leishmania DNA in conjunctival swab samples were compared. All methods had two steps: a first amplification followed by hybridization or by a new amplification (nested or seminested). Two methods (kDNA PCR-hybridization and kDNA snPCR) used primers targeted to the minicircles of kinetoplast DNA (kDNA) and the other two methods to the coding (LnPCR) and intergenic noncoding regions (ITS-1 nPCR) of ribosomal rRNA genes. kDNA PCR-hybridization was positive for 22/23 dogs (95.6%) and for 40/46 samples (86.9%), considering the right and the left conjunctivas. kDNA snPCR was positive for 21/23 dogs (91.3%) and for 40/46 samples (86.9%). The ITS-1 nPCR and LnPCR were both able to detect the parasites in 17/23 dogs (73.9%) and 29/46 (63%) and 30/46 (65.2%) samples, respectively. The positivities of the kDNA based methods were significantly higher; however the choice of the best method will depend on the kind of information required with the diagnosis.
Pilatti
Marcia Maria
MM
Centro de Desenvolvimento da Tecnologia Nuclear, Cidade Universitária-Campus da UFMG, 31120-970 Belo Horizonte, MG, Brazil. marciapilatti@yahoo.com.br
Ferreira
Sidney de Almeida
Sde A
de Melo
Maria Norma
MN
de Andrade
Antero Silva Ribeiro
AS
D000818:Animals; D017931:DNA Primers; D016054:DNA, Protozoan / Q000235:genetics / Q000302:isolation & purification; D004283:Dog Diseases / Q000175:diagnosis / Q000469:parasitology*; D004285:Dogs; D005784:Gene Amplification; D017125:Genes, Protozoan / Q000235:genetics; D007891:Leishmania / Q000235:genetics; D007898:Leishmaniasis, Visceral / Q000175:diagnosis / Q000235:genetics / Q000662:veterinary*; D009693:Nucleic Acid Hybridization; D016133:Polymerase Chain Reaction / Q000379:methods; D016053:RNA, Protozoan / Q000235:genetics / Q000302:isolation & purification; D012335:RNA, Ribosomal / Q000235:genetics / Q000302:isolation & purification
D017931:DNA Primers; D016054:DNA, Protozoan; D016053:RNA, Protozoan; D012335:RNA, Ribosomal