Single-tube nested PCR assay with in-house DNA extraction for Mycobacterium tuberculosis detection in blood and urine
Abstract: INTRODUCTION : Molecular analyses are auxiliary tools for detecting Koch's bacilli in clinical specimens from patients with suspected tuberculosis (TB). However, there are still no efficient diagnostic tests that combine high sensitivity and specificity and yield rapid results in the...
| Main Authors: | , , , , , , , |
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| Format: | Article |
| Language: | English |
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Sociedade Brasileira de Medicina Tropical (SBMT)
2015-12-01
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| Series: | Revista da Sociedade Brasileira de Medicina Tropical |
| Subjects: | |
| Online Access: | http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0037-86822015000600731&lng=en&tlng=en |
| _version_ | 1818218744055332864 |
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| author | Juliana Figueirêdo da Costa Lima Gabriela de Moraes Rêgo Guedes Juliana Falcão de Araújo Lima Laís Ariane de Siqueira Lira Fabiana Cristina Fulco Santos Mercia Eliane de Arruda Lílian Maria Lapa Montenegro Haiana Charifker Schindler |
| author_facet | Juliana Figueirêdo da Costa Lima Gabriela de Moraes Rêgo Guedes Juliana Falcão de Araújo Lima Laís Ariane de Siqueira Lira Fabiana Cristina Fulco Santos Mercia Eliane de Arruda Lílian Maria Lapa Montenegro Haiana Charifker Schindler |
| author_sort | Juliana Figueirêdo da Costa Lima |
| collection | DOAJ |
| description | Abstract: INTRODUCTION : Molecular analyses are auxiliary tools for detecting Koch's bacilli in clinical specimens from patients with suspected tuberculosis (TB). However, there are still no efficient diagnostic tests that combine high sensitivity and specificity and yield rapid results in the detection of TB. This study evaluated single-tube nested polymerase chain reaction (STNPCR) as a molecular diagnostic test with low risk of cross contamination for detecting Mycobacterium tuberculosis in clinical samples. METHODS: Mycobacterium tuberculosis deoxyribonucleic acid (DNA) was detected in blood and urine samples by STNPCR followed by agarose gel electrophoresis. In this system, reaction tubes were not opened between the two stages of PCR (simple and nested). RESULTS: STNPCR demonstrated good accuracy in clinical samples with no cross contamination between microtubes. Sensitivity in blood and urine, analyzed in parallel, was 35%-62% for pulmonary and 41%-72% for extrapulmonary TB. The specificity of STNPCR was 100% in most analyses, depending on the type of clinical sample (blood or urine) and clinical form of disease (pulmonary or extrapulmonary). CONCLUSIONS: STNPCR was effective in detecting TB, especially the extrapulmonary form for which sensitivity was higher, and had the advantage of less invasive sample collection from patients for whom a spontaneous sputum sample was unavailable. With low risk of cross contamination, the STNPCR can be used as an adjunct to conventional methods for diagnosing TB. |
| first_indexed | 2024-12-12T07:28:37Z |
| format | Article |
| id | doaj.art-f3f282239a814286b26d3f3b39ad4706 |
| institution | Directory Open Access Journal |
| issn | 1678-9849 |
| language | English |
| last_indexed | 2024-12-12T07:28:37Z |
| publishDate | 2015-12-01 |
| publisher | Sociedade Brasileira de Medicina Tropical (SBMT) |
| record_format | Article |
| series | Revista da Sociedade Brasileira de Medicina Tropical |
| spelling | doaj.art-f3f282239a814286b26d3f3b39ad47062022-12-22T00:33:05ZengSociedade Brasileira de Medicina Tropical (SBMT)Revista da Sociedade Brasileira de Medicina Tropical1678-98492015-12-0148673173810.1590/0037-8682-0210-2015S0037-86822015000600731Single-tube nested PCR assay with in-house DNA extraction for Mycobacterium tuberculosis detection in blood and urineJuliana Figueirêdo da Costa LimaGabriela de Moraes Rêgo GuedesJuliana Falcão de Araújo LimaLaís Ariane de Siqueira LiraFabiana Cristina Fulco SantosMercia Eliane de ArrudaLílian Maria Lapa MontenegroHaiana Charifker SchindlerAbstract: INTRODUCTION : Molecular analyses are auxiliary tools for detecting Koch's bacilli in clinical specimens from patients with suspected tuberculosis (TB). However, there are still no efficient diagnostic tests that combine high sensitivity and specificity and yield rapid results in the detection of TB. This study evaluated single-tube nested polymerase chain reaction (STNPCR) as a molecular diagnostic test with low risk of cross contamination for detecting Mycobacterium tuberculosis in clinical samples. METHODS: Mycobacterium tuberculosis deoxyribonucleic acid (DNA) was detected in blood and urine samples by STNPCR followed by agarose gel electrophoresis. In this system, reaction tubes were not opened between the two stages of PCR (simple and nested). RESULTS: STNPCR demonstrated good accuracy in clinical samples with no cross contamination between microtubes. Sensitivity in blood and urine, analyzed in parallel, was 35%-62% for pulmonary and 41%-72% for extrapulmonary TB. The specificity of STNPCR was 100% in most analyses, depending on the type of clinical sample (blood or urine) and clinical form of disease (pulmonary or extrapulmonary). CONCLUSIONS: STNPCR was effective in detecting TB, especially the extrapulmonary form for which sensitivity was higher, and had the advantage of less invasive sample collection from patients for whom a spontaneous sputum sample was unavailable. With low risk of cross contamination, the STNPCR can be used as an adjunct to conventional methods for diagnosing TB.http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0037-86822015000600731&lng=en&tlng=enMycobacterium tuberculosisMolecular diagnostic testNested polymerase chain reactionBloodUrine |
| spellingShingle | Juliana Figueirêdo da Costa Lima Gabriela de Moraes Rêgo Guedes Juliana Falcão de Araújo Lima Laís Ariane de Siqueira Lira Fabiana Cristina Fulco Santos Mercia Eliane de Arruda Lílian Maria Lapa Montenegro Haiana Charifker Schindler Single-tube nested PCR assay with in-house DNA extraction for Mycobacterium tuberculosis detection in blood and urine Revista da Sociedade Brasileira de Medicina Tropical Mycobacterium tuberculosis Molecular diagnostic test Nested polymerase chain reaction Blood Urine |
| title | Single-tube nested PCR assay with in-house DNA extraction for Mycobacterium tuberculosis detection in blood and urine |
| title_full | Single-tube nested PCR assay with in-house DNA extraction for Mycobacterium tuberculosis detection in blood and urine |
| title_fullStr | Single-tube nested PCR assay with in-house DNA extraction for Mycobacterium tuberculosis detection in blood and urine |
| title_full_unstemmed | Single-tube nested PCR assay with in-house DNA extraction for Mycobacterium tuberculosis detection in blood and urine |
| title_short | Single-tube nested PCR assay with in-house DNA extraction for Mycobacterium tuberculosis detection in blood and urine |
| title_sort | single tube nested pcr assay with in house dna extraction for mycobacterium tuberculosis detection in blood and urine |
| topic | Mycobacterium tuberculosis Molecular diagnostic test Nested polymerase chain reaction Blood Urine |
| url | http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0037-86822015000600731&lng=en&tlng=en |
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