[Rapid diagnosis of tuberculosis by the polymerase chain reaction (PCR)].
The polymerase chain reaction (PCR) was used for the detection of Mycobacterium tuberculosis DNA. More than 2000 different clinical specimens were analyzed by this assay. The efficiency of two different methods for processing the DNA from biological material was analyzed. DNA amplification was done according to standard protocols by amplifying a segment of 402 bp of the 65 kD mycobacterial gene, electrophoretic separation of the amplification product followed by Southern transfer and hybridization with a Mycobacterium tuberculosis-specific probe or by a semi-nested amplification procedure in which the initial amplification product was reamplified by a second round with a Mycobacterium tuberculosis-specific primer. The specificity of primers and probe for mycobacterial DNA was proven by testing 70 of class-I microorganisms, as well as 20 different strains and own isolates of Mycobacterium tuberculosis and 67 strains of 25 different MOTTs. Some of the amplification products were sequenced. The clinical relevance of the results and the sensitivity of the PCR method were confirmed by simultaneous quantitative bacterial culture from the same clinical specimens. The results of conventional culture method received after 8 to 10 weeks culture time correlated with the results from PCR obtained within 12 hours in 95.4% in the semi-nested amplification procedure. The discrepancy of 4.6% was caused by positive results of PCR and negative cultures which might be due to the higher sensitivity of PCR compared to culture technique. The results show that PCR may be used for detection of Mycobacterium tuberculosis in clinical specimens. The specificity can be regarded as largely proven, advantages are velocity and sensitivity.(ABSTRACT TRUNCATED AT 250 WORDS)
Seelig
R
R
Privates Institut für Immunologie und Molekulargenetik GmbH, Karlsruhe.
Renz
M
M
Bottner
C
C
Stockinger
K
K
Czichos
J
J
Schulz
V
V
Seelig
H P
HP
D001483:Base Sequence; D004269:DNA, Bacterial / Q000032:analysis; D005798:Genes, Bacterial; D006360:Heat-Shock Proteins / Q000235:genetics; D006801:Humans; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides / Q000737:chemistry; D016133:Polymerase Chain Reaction; D014376:Tuberculosis / Q000175:diagnosis*
D004269:DNA, Bacterial; D006360:Heat-Shock Proteins; D009838:Oligodeoxyribonucleotides
Tuberkulose-Schnelldiagnostik mit der Polymerase-Ketten-Reaktion (PCR).