Simultaneous detection and differentiation of pathogenic and nonpathogenic Leptospira spp. by multiplex real-time PCR (TaqMan) assay

• Autorzy: Bedir O. , Kilic A. , Atabek E. , Kuskucu A.M. , Turhan V. , Basustaoglu A.C.
• Języki publikacji: EN

Abstrakty

EN

Leptospirosis, caused by pathogenic Leptospira, is one of the most important zoonoses in the world. Several molecular techniques have been developed for detection and differentiation between pathogenic and saprophytic Leptospira spp. The aim of this study was to develop a rapid and simple assay for specific detection and differentiation of pathogenic Leptospira spp. by multiplex real-time PCR (TaqMan) assay using primers and probes targeting Leptospira genus specific 16S ribosomal RNA gene, the pathogen specific ligA/B genes and non-pathogen Leptospira biflexa specific 23S ribosomal RNA gene. Sixteen reference strains of Leptospira spp. including pathogenic and nonpathogenic and ten other negative control bacterial strains were used in the study. While the 16S primers amplified target from both pathogenic and non-pathogenic leptospires, the ligA/B and the 23S primers amplified target DNA from pathogenic and non-pathogenic leptospires, respectively. The multiplex real-time PCR (TaqMan) assay detection limit, that is, the sensitivity was found approximately 1 x 10² cells/ml for ligA/B gene and 23S ribosomal RNA gene, and 10 cells/ml 16S ribosomal RNA. The reaction efficiencies were 83-105% with decision coefficients of more than 0.99 in all multiplex assays. The multiplex real-time PCR (TaqMan) assay yielded negative results with the ten other control bacteria. In conclusion, the developed multiplex real-time PCR (TaqMan) assay is highly useful for early diagnosis and differentiation between pathogenic and non-pathogenic leptospires in a reaction tube as having high sensitivity and specificity.

Słowa kluczowe

EN

human disease; zoonosis; leptospirosis; simultaneous detection; differentiation; pathogenic bacteria; non-pathogenic microorganism; Leptospira; multiplex real-time polymerase chain reaction

• Wydawca: -
• Czasopismo: Polish Journal of Microbiology
• Rocznik: 2010
• Tom: 59
• Numer: 3
• Opis fizyczny: p.167-173,fig.,ref.

Twórcy

autor

Bedir O.

• Department of Microbiology and Clinical Microbiology, Gulhane Military Medical Academy and School of Medicine, 06018, Ankara, Turkey

autor

Kilic A.

• Department of Microbiology and Clinical Microbiology, Gulhane Military Medical Academy and School of Medicine, 06018, Ankara, Turkey

autor

Atabek E.

• Central Veterinary Control and Research Institute, Etlik, Ankara, Turkey

autor

Kuskucu A.M.

• Department of Microbiology and Clinical Microbiology, Gulhane Military Medical Academy and School of Medicine, 06018, Ankara, Turkey

autor

Turhan V.

• Department of Infectious Diseases, Gulhane Military Medical Academy and School of Medicine, Haydarpasa Training Hospital, Istanbul, Turkey

autor

Basustaoglu A.C.

• Department of Microbiology and Clinical Microbiology, Gulhane Military Medical Academy and School of Medicine, 06018, Ankara, Turkey

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