Detection and quantification of Mycobacterium leprae in tissue samples by real-time PCR.
Real-time PCR technology has improved molecular diagnostics of many pathogens, but no such test is available for Mycobacterium leprae. In this report we describe the establishment and the pre-clinical evaluation of such an assay. The test achieved a theoretical analytical sensitivity limit of 194 M. leprae cells per skin biopsy specimen and facilitated quantification of mycobacteria in tissue over a range of 54-54,000,000 cells per sample. In punch skin biopsies from 39 untreated Ugandan patients with newly diagnosed leprosy, the clinical diagnosis could be confirmed in 88.9% of multibacillary and 33.3% of paucibacillary (microscopically negative) patients. Real-time detection thus did not increase the clinical sensitivity of PCR as compared to conventional protocols, in spite of its evidently high analytical sensitivity. On the other hand, as still no culture system exists for M. leprae, the assay appears to be a robust tool for detection of the bacterium in selected clinical situations, as well as for quantitation in experimental settings.
Kramme
Stefanie
S
National Reference Center for Tropical Infections, Bernhard Nocht Institute of Tropical Medicine, Bernhard-Nocht Str. 74, 20359 Hamburg, Germany.
Bretzel
Gisela
G
Panning
Marcus
M
Kawuma
Joseph
J
Drosten
Christian
C
D000942:Antigens, Bacterial / Q000235:genetics; D001426:Bacterial Proteins / Q000235:genetics; D001706:Biopsy; D004269:DNA, Bacterial / Q000737:chemistry / Q000302:isolation & purification; D006801:Humans; D007918:Leprosy / Q000175:diagnosis* / Q000382:microbiology; D008969:Molecular Sequence Data; D009166:Mycobacterium leprae / Q000235:genetics / Q000302:isolation & purification*; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D012867:Skin / Q000382:microbiology; D014454:Uganda
D000942:Antigens, Bacterial; D001426:Bacterial Proteins; D004269:DNA, Bacterial