Detection and identification of mycobacteria by amplification of a segment of the gene coding for the 32-kilodalton protein

Soini, Hanna; Skurnik, Mikael; Liippo, K.; Tala, E; Viljanen, Matti K.

doi:10.1128/jcm.30.8.2025-2028.1992

Cited by 88 publications

(46 citation statements)

References 23 publications

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“…The clinical sensitivity of the PCR procedure found in our study is in accordance with results of other groups, which are between 0.74 and 1.00 (3,5,7,9,10). However, some studies report a relatively low PCR sensitivity: e.g., Pierre et al (14) and Soini et al (24) found sensitivities for their PCR assays of 63 and 55.9%, respectively. Such low sensitivities in PCR studies may be explained by suboptimal assay conditions.…”

Section: Discussionsupporting

confidence: 91%

Clinical evaluation of a Mycobacterium tuberculosis PCR assay

et al. 1995

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On the basis of previously published PCR primer sequences, we have designed a sensitive system for detecting DNA of the Mycobacterium tuberculosis complex (MTB) in patient sputum samples which employs a fast and simplified sample preparation method appropriate for routine diagnostic testing. In order to evaluate the accuracy of the PCR assay, we performed a prospective study with 103 patients, comparing PCR results with culture results of samples obtained from a parallel culture assay as well as with subsequent culture results. Using two MTB-specific PCR primer systems, we found 48 of 49 tuberculosis (Tb) patients to be PCR positive (PCR sensitivity, 0.98). Sixteen of 54 presumably non-Tb patients showed amplifiable MTB DNA (specificity, 0.7). The study demonstrates that for diagnostic applications of MTB PCR two MTB-specific primer pairs should be used. MTB infection is extremely unlikely in cases of MTB PCR-negative samples: with our method for the exclusion of active Tb, the validity of one PCR assay seems to be equivalent to those of at least three culturing procedures. Positive PCR results do not necessarily reflect active MTB infection. It remains to be shown whether positive PCR results in Tb-negative patients mean false-positivity, an early laboratory finding which predicts a subsequent reactivation of a prior Tb infection, or whether asymptomatic patients may carry PCR-amplifiable MTB DNA without any clinical relevance. It is important to point out that the validity of PCR results in clinical studies depends on the use of contamination controls parallel to all PCR steps and the simplicity of the DNA extraction method as well as on the specificity of the PCR results.

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Section: Discussionsupporting

confidence: 91%

Clinical evaluation of a Mycobacterium tuberculosis PCR assay

et al. 1995

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“…In contrast, Pierre et al (24) and Soini et al. (27) reported disappointingly insensitive PCR assays (63% and 55.9%, respectively) although they were highly specific (91% and 100%). In subsequent evaluations using a larger number of samples, intermediate results were obtained.…”

Section: Discussionmentioning

confidence: 93%

Comparison of two different PCR detection methods. Application to the diagnosis of pulmonary tuberculosis

Rodrı́guez

Fuentes

Royo

1997

APMIS

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Comparison of two different PCR detection methods. Application to the diagnosis of pulmonary tuberculosis. APMIS 105: 612-616, 1997. The objectives are to assess the influence of the detection of the amplified DNA fragment on the sensitivity and specificity of the polymerase chain reaction (PCR). One hundred seventy-five sputum samples from 123 patients were processed. Sixty samples were taken from 60 subjects without tuberculosis, and the rest were taken from subjects with tuberculosis confirmed by culture. A fragment of the IS61 10 sequence of Mycobacterium tuberculosis, which was detected using two different methods, was amplified. The detection methods used were a digoxigenin-labeled specific probe and chemiluminescent development and reamplification (nested PCR) combined with agarose gel electrophoresis. Sensitivity with probe detection was 75.65% and specificity 100%. Using the nested PCR technique, sensitivity rose to 93.04%, but specificity decreased to 96.6%. PCR is a quick and adequate way to diagnose pulmonary tuberculosis in cases where staining is negative yet there is a clinical suspicion of tuberculosis, even though a standardization process and large scale evaluation are still needed to determine its true usefulness.

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“…The most rapid diagnostic procedures for identifying mycobacteria involve the PCR technology (2,4,12). However, direct PCR of sputum, blood, and feces is often limited by the inadequate sensitivity of current diagnostic protocols (2,12,(26)(27)(28). Recently, Mazurek et al (18) have reported that detection of M. tuberculosis organisms in cerebrospinal fluid is accelerated and enhanced following immunomagnetic enrichment.…”

Section: Discussionmentioning

confidence: 99%

“…However, the usefulness of the DNA amplification procedures is often limited by the presence of PCR inhibitors in clinical samples. In some studies, rates of occurrence of false-negative results of greater than 20% have been reported (2,20,26).…”

mentioning

confidence: 99%

Rapid detection of Mycobacterium avium in stool samples from AIDS patients by immunomagnetic PCR

Bai

Reyn

et al. 1996

J Clin Microbiol

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Direct PCR detection of bacteria in clinical samples is often hindered by the presence of compounds that inhibit the PCR. To improve and accelerate the diagnosis of Mycobacterium avium-M. intracellulare complex infections, an immunomagnetic PCR (IM-PCR) assay was developed. This IM-PCR procedure combines the separation of mycobacteria by antimycobacterial monoclonal antibody coupled to magnetic beads with an M. avium-M. intracellulare complex-specific PCR protocol based on 16S rRNA gene sequences. As few as 10 M. avium bacilli were detected in spiked human stool samples, a clinical specimen usually refractory to conventional PCR analysis, by the IM-PCR method. Moreover, M. avium organisms were detected in about 24 h in 18 of 22 culture-confirmed fecal samples from AIDS patients. This IM-PCR protocol should allow for the rapid and sensitive detection of M. avium isolates in clinical specimens.

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Detection and identification of mycobacteria by amplification of a segment of the gene coding for the 32-kilodalton protein

Cited by 88 publications

References 23 publications

Clinical evaluation of a Mycobacterium tuberculosis PCR assay

Clinical evaluation of a Mycobacterium tuberculosis PCR assay

Comparison of two different PCR detection methods. Application to the diagnosis of pulmonary tuberculosis

Rapid detection of Mycobacterium avium in stool samples from AIDS patients by immunomagnetic PCR

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