MEDLINE Abstracts: Use of Polymerase Chain Reaction to Diagnose Mycobacterium tuberculosis in the Ur
MEDLINE Abstracts: Use of Polymerase Chain Reaction to Diagnose Mycobacterium tuberculosis in the Urine
Should PCR be used to detect M tuberculosis in the urine? Find out in this easy-to-navigate collection of recent MEDLINE abstracts compiled by the editors at Medscape Transplantation.
Mycobacterium tuberculosis by PCR Analysis.
Portillo-Gomez L, Morris SL, Panduro A
Int J Tuberc Lung Dis. 2000;4:361-370
Setting: The diagnosis of extra-pulmonary tuberculosis (EPTB) remains an important clinical problem, primarily because of the inadequate sensitivity of conventional bacteriologic methods for detecting Mycobacterium tuberculosis in extra-pulmonary specimens.
Objective: To evaluate whether a IS6110-based polymerase chain reaction (PCR) method can be utilized to detect M. tuberculosis in non-pulmonary specimens.
Design: Specimens from 286 Mexican patients with a presumptive clinical diagnosis of EPTB were prospectively examined by Ziehl-Neelsen staining, mycobacterial culture on Lowenstein-Jensen slants, and by PCR. The DNA for PCR was extracted by the buffer lysis method and phenol-guanidine thiocyanate-chloroform. Primers that amplify a 200 bp fragment from the insertion-like M. tuberculosis sequence element IS6110 were utilized.
Results: Our results demonstrate that this PCR method is highly specific (100%) for identifying M. tuberculosis from a variety of specimens including cerebrospinal fluid (CSF), pleural fluid, ascitic fluid, pericardial fluid, urine, and lymph node exudate. Moreover, the sensitivity of PCR for detecting M. tuberculosis in CSF (94%), pleural fluid (94%), ascitic fluid and other extrapulmonary specimens (93%) greatly exceeds the sensitivity of conventional smear and culture methods.
Conclusion: These results demonstrate that PCR can be a highly specific and sensitive aid in the detection of M. tuberculosis from extra-pulmonary specimens.
Moussa OM, Eraky I, El-Far MA, Osman HG, Ghoneim MA
J Urol. 2000;164:584-548
Objective: To establish a polymerase chain reaction (PCR) assay for the rapid detection and identification of mycobacteria in urine, and to assess the value of such assay in routine laboratory diagnosis of genitourinary tuberculosis.
Materials And Methods: Urine specimens from 1000 patients with clinical suspicion of urinary tuberculosis were examined. Two assays for the detection and identification of Mycobacterium tuberculosis (M. tuberculosis) complex and mycobacteria other than tuberculosis (MOTT) by non-radioactive DNA hybridization of PCR-product were applied. The first assay used PCR primers and probe derived from M. tuberculosis species-specific DNA insertion sequence, IS6110. The second utilized mycobacterium genus-specific sequence encoding ribosomal ribonucleic acid (16S rRNA). The results obtained by PCR were compared with those obtained by standard microbiological methods, acid-fast bacilli (AFB) stain and culture.
Results: Compared with cultures, the sensitivity of AFB staining was 52.07% and the specificity was 96.7%. In comparison to the results of culture, the overall sensitivity and specificity of the IS6110-PCR assay was 95.59% and 98.12% respectively. While the corresponding results for the 16S rRNA gene-PCR were 87.05% and 98. 9%.
Conclusion: The high sensitivity and specificity in addition to the potential for rapid detection of mycobacteria, makes this test a useful tool in the clinical management of mycobacterial infection in urine. Urine specimens may contain M. tuberculosis and/or other mycobacteria; therefore, there are advantages in using genus-specific primers in parallel with species-specific primers in PCR assay.
Mycobacterium tuberculosis by Polymerase Chain Reaction in a Selected Population in Northwestern Mexico
Moran Moguel MC, Hernandez DA, Pena Montes de Oca PM, et al
Rev Panam Salud Publica. 2000;7:389-394
This study compares the detection of Mycobacterium tuberculosis through bacilloscopy (Ziehl-Neelsen stain), growth in Lowenstein-Jensen medium, and polymerase chain reaction (PCR) carried out with DNA taken directly from various types of samples. A total of 252 samples were analyzed (114 sputum, 96 urine, 15 cerebrospinal fluid, and 27 of other types) from 160 patients with any form of suspected tuberculosis who came to the Clinical Pathology Laboratory of the Specialties Hospital of the Western National Medical Center of the Mexican Social Security Institute. In all cases Ziehl-Neelsen stains were done, as were also cultures with Lowenstein-Jensen medium and PCR amplification of a segment of 285 base pairs specific to the M. tuberculosis complex. Of the 252 samples, with the culture, 18 were positive for nontuberculous mycobacteria. Of the 234 others, 12 (5.1%) were positive with the PCR and the culture, 174 (74.4%) negative in both tests, 47 (20.1%) positive with the PCR and negative with the culture, and 1 (0.4%) negative with the PCR and positive with the culture. Using the culture as the reference test, the PCR provided a sensitivity of 92.3%, a specificity of 78.7%, a positive predictive value of 20.3%, and a negative predictive value of 99.4%. The PCR detection limit with DNA taken from culture was 10 fg, equivalent to four or five mycobacteria. Also in comparison with the culture, the PCR correctly identified the totality of the mycobacteria of the M. tuberculosis complex. Taking the culture as the reference test, when analyzing just the sputum samples, the direct PCR provided a sensitivity of 90.9%, a specificity of 89.5%, a positive predictive value of 52.6%, and a negative predictive value of 98.7%. The PCR is a sensitive and specific technique for detecting the M. tuberculosis complex in both positive and negative bacilloscopy samples. A controlled PCR procedure makes it possible to establish or to exclude the diagnosis of tuberculosis in a time that is reduced from more than three weeks to just 24 to 48 hours. This is particularly useful when an early diagnosis is needed to establish a patient's prognosis or in organ transplant cases.
Hemal AK, Gupta NP, Rajeev TP, Kumar R, Dar L, Seth P
Urology. 2000;56:570-574
Objectives: To evaluate the role of urinary polymerase chain reaction (PCR) in the detection of Mycobacterium tuberculosis (MTb) in patients with a clinical suspicion of genitourinary tuberculosis (GUTB) and to compare its sensitivity with intravenous urography (IVU), bladder biopsy, and urine culture for acid fast bacilli (AFB).
Methods: The study was carried out between September 1997 and December 1998 in 42 patients with a clinical suspicion of GUTB. Their clinical features, organ involvement, and investigation results were studied. The diagnostic yield of urinary PCR for MTb and its sensitivity in comparison with routine urine AFB culture, bladder biopsy, and IVU were assessed.
Results: There were 25 male and 17 female patients, with a mean age of 31.04 years. Patients suspected of having GUTB most often presented with irritative voiding symptoms. Two patients had abnormal renal parameters. Of the 42 patients clinically suspected of having GUTB, radiologic abnormalities suggestive of GUTB were found in 37 (88.09%); MTb was isolated in the urine AFB culture in 13 (30.95%); bladder biopsy was positive in 11 (45.83%); and urinary PCR for MTb was positive in 34 cases (80.95%). Of 35 cases of proven GUTB, IVU was suggestive of the diagnosis in 32 (91.42%) and MTb was isolated in the urine AFB culture in 13 cases (37.14%). Bladder biopsy was positive in 11 (45. 83%) of 24 patients in whom biopsy was taken, and urinary PCR for MTb was positive in 33 (94.29%).
Conclusions: A high index of suspicion is necessary for a diagnosis of GUTB. In clinically suspected cases, IVU may be suggestive of GUTB, but it is not specific. In the present study, IVU was suggestive in 88.09% of patients. MTb was isolated in the urine AFB culture in only 37.14% of patients, and bladder biopsy was positive in 45.83%. Urinary PCR for MTb was the most sensitive indicator and was positive in 94.29% of patients. It is evident from this series that PCR provides a much faster diagnosis of urinary MTb. It is a rapid, sensitive, and specific diagnostic method and avoids a delay in starting treatment.
Aceti A, Zanetti S, Mura MS, et al
Thorax. 1999;54:145-146
Background: Despite the increased dissemination of tuberculosis among HIV infected patients, the diagnosis is difficult to establish. Traditional microbiological methods lack satisfactory sensitivity. We have developed a highly sensitive and specific nested polymerase chain reaction (PCR) capable of detecting Mycobacterium tuberculosis DNA in urine specimens and have used this test to examine urine specimens from HIV patients with active pulmonary tuberculosis.
Methods: Urine specimens from 13 HIV infected patients with microbiologically proven active pulmonary tuberculosis, 10 AIDS patients with non-tuberculous mycobacterial infection (documented by blood culture), 53 AIDS patients with no evidence of mycobacterial disease, and 80 healthy subjects (25 with positive skin test to purified protein derivative) were tested for M tuberculosis using PCR, acid fast staining (AFS), and culture.
Results: Of the urine specimens from patients with active tuberculosis, all tested positive by PCR, two by culture, and none by AFS. No reactivity was observed in urine specimens from patients with non-tuberculous mycobacterial infection. Of the 53 AIDS patients without mycobacterial infection, one had a positive urine PCR. Normal subjects were all negative.
Conclusions: Urine based nested PCR for M tuberculosis may be a useful test for identifying HIV patients with pulmonary tuberculosis.
Ginesu F, Pirina P, Sechi LA, et al
J Chemother. 1998;10:295-300
The aim of our study was to evaluate the diagnostic value of various methods widely used in microbiological diagnosis of tuberculosis: direct smear examination for acid-fast bacilli, cultural identification in Lowestein-Jensen (L-J) medium, the radiometric BACTEC 460 system, and Polymerase Chain Reaction (PCR). Three hundred and ninety-three clinical samples of sputum (375), gastric aspirate (3), pleural fluid (12) and urine (3) were taken from 125 patients hospitalized at our Institute for suspected pulmonary tuberculosis, between January 1995 and June 1997. On completion of diagnosis, 35 were found to be affected by active tuberculosis (30 pulmonary, 4 pleural and 1 urinary) and 90 by other non-tubercular diseases (pneumonia, lung cancer, non-tubercular pleural effusion, etc.). In our study, direct smear examination for acid-smear bacilli gave diagnostic value results of 88% and positive predictive value of 91.67%. Cultural identification in L-J and BACTEC 460 TB radiometric system media resulted in diagnostic values of 96.80% and 94.40%, respectively, and positive predictive values of 100% for both of them. Finally, One-Tube Nested-PCR, a variant which uses specific primers for the IS6110 insertion sequence specific for Mycobacterium tuberculosis, gave us 88.80% (91.43% sensitivity and 87.78% specificity) diagnostic value results, and 74.42% (11 false-positives) positive predictive value. On the basis of our results, we can affirm that PCR is a good method for microbiological diagnosis of tuberculosis, given its high sensitivity and specificity and unparalleled rapidity. However, the high number of false-positives that we found suggests that results obtained should be confirmed with BACTEC, which considerably reduces the time required for identification, and makes it possible to carry out an antibiotic assay rapidly.
M. tuberculosis
Eing BR, Becker A, Sohns A, Ringelmann R
J Clin Microbiol. 1998;36:2023-2029
The new Roche Cobas Amplicor Mycobacterium tuberculosis assay, which is a semiautomated version of the manually performed Roche Amplicor M. tuberculosis test, was compared to culture and an IS6110-based in-house PCR protocol. A total of 1,681 specimens from 833 patients, including specimen types other than sputum, were tested in parallel by both the in-house PCR and the Cobas Amplicor M. tuberculosis assay. After we resolved discrepant PCR results, the sensitivity, specificity, and positive and negative predictive values for the Cobas Amplicor M. tuberculosis assay were 66.33, 99.71, 94.36, and 97.66%, respectively. The corresponding values for the in-house PCR were 91.08, 99.85, 97.87, and 99.37%, respectively. For culture- and smear-positive specimens, the sensitivity of the Cobas Amplicor M. tuberculosis test was 96.42% (in-house PCR, 100%). If only smear-negative sputum specimens were considered, the Cobas Amplicor M. tuberculosis assay exhibited a sensitivity of 45.45% (in-house PCR, 63.63%) relative to that of culture. With a modified protocol for DNA extraction (washing of samples plus ultrasonication), both PCR methods performed better with gastric aspirates than with sputum samples (sensitivity of the Cobas Amplicor M. tuberculosis assay with smear-negative gastric aspirates, 70.00%; sensitivity of in-house PCR, 90.00%). With dithiothreitol being used for liquefaction of specimens in this study, the Cobas Amplicor M. tuberculosis assay exhibited an inhibition rate of 9.16%. In our view, the new Cobas Amplicor M. tuberculosis test (i) is well suited for typing of smear-positive specimens, (ii) may also be applied to gastric aspirates and other types of specimens if DNA extraction methods are modified appropriately, and (iii) exhibits a sensitivity with smear-negative sputum specimens which makes it recommendable that a minimum of three samples from the same patient be tested.
Mycobacterium tuberculosis by PCR Analysis of Urine and Other Clinical Samples from AIDS and Non-HIV-Infected Patients
Sechi LA, Pinna MP, Sanna A, et al
Mol Cell Probes. 1997;11:281-285
A number of different clinical specimens, such as sputum, cerebrospinal fluid and blood, have been reported to be good substrates for the detection of Mycobacterium tuberculosis by PCR assay. We wanted to search for the presence of mycobacteria in other body fluids, such as urine. Urine samples and other samples obtained from AIDS patients and non HIV-infected patients were analysed by PCR. The results were compared with those obtained using conventional methods (Bactec 460 TB and AFB (acid fast bacilli strain)). We analysed 412 urine samples and 210 different other samples (sputum and cerebrospinal fluid) obtained from AIDS patients by PCR; almost identical levels of PCR-positive (14-17%) results were observed in all samples analysed. The results were then compared with those obtained with the Bactec 460 TB and AFB. PCR, Bactec 460 TB and acid fast stain were also used to analyse 190 urine samples and 230 other samples from non-HIV infected patients in the consumption ward of Sassari Hospital. The number of urine samples positive by PCR (6.3%) and Bactec 460 TB (2.1%) was half that obtained from samples taken from the AIDS patients. As expected, an increase in the number of positive sputum samples was observed with all methods. The results indicate that PCR analysis of urine samples represents a valid alternative for fast and sensitive detection of M. tuberculosis. This method can be routinely used in the clinical laboratory, especially in HIV-infected patients.
Should PCR be used to detect M tuberculosis in the urine? Find out in this easy-to-navigate collection of recent MEDLINE abstracts compiled by the editors at Medscape Transplantation.
Mycobacterium tuberculosis by PCR Analysis.
Portillo-Gomez L, Morris SL, Panduro A
Int J Tuberc Lung Dis. 2000;4:361-370
Setting: The diagnosis of extra-pulmonary tuberculosis (EPTB) remains an important clinical problem, primarily because of the inadequate sensitivity of conventional bacteriologic methods for detecting Mycobacterium tuberculosis in extra-pulmonary specimens.
Objective: To evaluate whether a IS6110-based polymerase chain reaction (PCR) method can be utilized to detect M. tuberculosis in non-pulmonary specimens.
Design: Specimens from 286 Mexican patients with a presumptive clinical diagnosis of EPTB were prospectively examined by Ziehl-Neelsen staining, mycobacterial culture on Lowenstein-Jensen slants, and by PCR. The DNA for PCR was extracted by the buffer lysis method and phenol-guanidine thiocyanate-chloroform. Primers that amplify a 200 bp fragment from the insertion-like M. tuberculosis sequence element IS6110 were utilized.
Results: Our results demonstrate that this PCR method is highly specific (100%) for identifying M. tuberculosis from a variety of specimens including cerebrospinal fluid (CSF), pleural fluid, ascitic fluid, pericardial fluid, urine, and lymph node exudate. Moreover, the sensitivity of PCR for detecting M. tuberculosis in CSF (94%), pleural fluid (94%), ascitic fluid and other extrapulmonary specimens (93%) greatly exceeds the sensitivity of conventional smear and culture methods.
Conclusion: These results demonstrate that PCR can be a highly specific and sensitive aid in the detection of M. tuberculosis from extra-pulmonary specimens.
Moussa OM, Eraky I, El-Far MA, Osman HG, Ghoneim MA
J Urol. 2000;164:584-548
Objective: To establish a polymerase chain reaction (PCR) assay for the rapid detection and identification of mycobacteria in urine, and to assess the value of such assay in routine laboratory diagnosis of genitourinary tuberculosis.
Materials And Methods: Urine specimens from 1000 patients with clinical suspicion of urinary tuberculosis were examined. Two assays for the detection and identification of Mycobacterium tuberculosis (M. tuberculosis) complex and mycobacteria other than tuberculosis (MOTT) by non-radioactive DNA hybridization of PCR-product were applied. The first assay used PCR primers and probe derived from M. tuberculosis species-specific DNA insertion sequence, IS6110. The second utilized mycobacterium genus-specific sequence encoding ribosomal ribonucleic acid (16S rRNA). The results obtained by PCR were compared with those obtained by standard microbiological methods, acid-fast bacilli (AFB) stain and culture.
Results: Compared with cultures, the sensitivity of AFB staining was 52.07% and the specificity was 96.7%. In comparison to the results of culture, the overall sensitivity and specificity of the IS6110-PCR assay was 95.59% and 98.12% respectively. While the corresponding results for the 16S rRNA gene-PCR were 87.05% and 98. 9%.
Conclusion: The high sensitivity and specificity in addition to the potential for rapid detection of mycobacteria, makes this test a useful tool in the clinical management of mycobacterial infection in urine. Urine specimens may contain M. tuberculosis and/or other mycobacteria; therefore, there are advantages in using genus-specific primers in parallel with species-specific primers in PCR assay.
Mycobacterium tuberculosis by Polymerase Chain Reaction in a Selected Population in Northwestern Mexico
Moran Moguel MC, Hernandez DA, Pena Montes de Oca PM, et al
Rev Panam Salud Publica. 2000;7:389-394
This study compares the detection of Mycobacterium tuberculosis through bacilloscopy (Ziehl-Neelsen stain), growth in Lowenstein-Jensen medium, and polymerase chain reaction (PCR) carried out with DNA taken directly from various types of samples. A total of 252 samples were analyzed (114 sputum, 96 urine, 15 cerebrospinal fluid, and 27 of other types) from 160 patients with any form of suspected tuberculosis who came to the Clinical Pathology Laboratory of the Specialties Hospital of the Western National Medical Center of the Mexican Social Security Institute. In all cases Ziehl-Neelsen stains were done, as were also cultures with Lowenstein-Jensen medium and PCR amplification of a segment of 285 base pairs specific to the M. tuberculosis complex. Of the 252 samples, with the culture, 18 were positive for nontuberculous mycobacteria. Of the 234 others, 12 (5.1%) were positive with the PCR and the culture, 174 (74.4%) negative in both tests, 47 (20.1%) positive with the PCR and negative with the culture, and 1 (0.4%) negative with the PCR and positive with the culture. Using the culture as the reference test, the PCR provided a sensitivity of 92.3%, a specificity of 78.7%, a positive predictive value of 20.3%, and a negative predictive value of 99.4%. The PCR detection limit with DNA taken from culture was 10 fg, equivalent to four or five mycobacteria. Also in comparison with the culture, the PCR correctly identified the totality of the mycobacteria of the M. tuberculosis complex. Taking the culture as the reference test, when analyzing just the sputum samples, the direct PCR provided a sensitivity of 90.9%, a specificity of 89.5%, a positive predictive value of 52.6%, and a negative predictive value of 98.7%. The PCR is a sensitive and specific technique for detecting the M. tuberculosis complex in both positive and negative bacilloscopy samples. A controlled PCR procedure makes it possible to establish or to exclude the diagnosis of tuberculosis in a time that is reduced from more than three weeks to just 24 to 48 hours. This is particularly useful when an early diagnosis is needed to establish a patient's prognosis or in organ transplant cases.
Hemal AK, Gupta NP, Rajeev TP, Kumar R, Dar L, Seth P
Urology. 2000;56:570-574
Objectives: To evaluate the role of urinary polymerase chain reaction (PCR) in the detection of Mycobacterium tuberculosis (MTb) in patients with a clinical suspicion of genitourinary tuberculosis (GUTB) and to compare its sensitivity with intravenous urography (IVU), bladder biopsy, and urine culture for acid fast bacilli (AFB).
Methods: The study was carried out between September 1997 and December 1998 in 42 patients with a clinical suspicion of GUTB. Their clinical features, organ involvement, and investigation results were studied. The diagnostic yield of urinary PCR for MTb and its sensitivity in comparison with routine urine AFB culture, bladder biopsy, and IVU were assessed.
Results: There were 25 male and 17 female patients, with a mean age of 31.04 years. Patients suspected of having GUTB most often presented with irritative voiding symptoms. Two patients had abnormal renal parameters. Of the 42 patients clinically suspected of having GUTB, radiologic abnormalities suggestive of GUTB were found in 37 (88.09%); MTb was isolated in the urine AFB culture in 13 (30.95%); bladder biopsy was positive in 11 (45.83%); and urinary PCR for MTb was positive in 34 cases (80.95%). Of 35 cases of proven GUTB, IVU was suggestive of the diagnosis in 32 (91.42%) and MTb was isolated in the urine AFB culture in 13 cases (37.14%). Bladder biopsy was positive in 11 (45. 83%) of 24 patients in whom biopsy was taken, and urinary PCR for MTb was positive in 33 (94.29%).
Conclusions: A high index of suspicion is necessary for a diagnosis of GUTB. In clinically suspected cases, IVU may be suggestive of GUTB, but it is not specific. In the present study, IVU was suggestive in 88.09% of patients. MTb was isolated in the urine AFB culture in only 37.14% of patients, and bladder biopsy was positive in 45.83%. Urinary PCR for MTb was the most sensitive indicator and was positive in 94.29% of patients. It is evident from this series that PCR provides a much faster diagnosis of urinary MTb. It is a rapid, sensitive, and specific diagnostic method and avoids a delay in starting treatment.
Aceti A, Zanetti S, Mura MS, et al
Thorax. 1999;54:145-146
Background: Despite the increased dissemination of tuberculosis among HIV infected patients, the diagnosis is difficult to establish. Traditional microbiological methods lack satisfactory sensitivity. We have developed a highly sensitive and specific nested polymerase chain reaction (PCR) capable of detecting Mycobacterium tuberculosis DNA in urine specimens and have used this test to examine urine specimens from HIV patients with active pulmonary tuberculosis.
Methods: Urine specimens from 13 HIV infected patients with microbiologically proven active pulmonary tuberculosis, 10 AIDS patients with non-tuberculous mycobacterial infection (documented by blood culture), 53 AIDS patients with no evidence of mycobacterial disease, and 80 healthy subjects (25 with positive skin test to purified protein derivative) were tested for M tuberculosis using PCR, acid fast staining (AFS), and culture.
Results: Of the urine specimens from patients with active tuberculosis, all tested positive by PCR, two by culture, and none by AFS. No reactivity was observed in urine specimens from patients with non-tuberculous mycobacterial infection. Of the 53 AIDS patients without mycobacterial infection, one had a positive urine PCR. Normal subjects were all negative.
Conclusions: Urine based nested PCR for M tuberculosis may be a useful test for identifying HIV patients with pulmonary tuberculosis.
Ginesu F, Pirina P, Sechi LA, et al
J Chemother. 1998;10:295-300
The aim of our study was to evaluate the diagnostic value of various methods widely used in microbiological diagnosis of tuberculosis: direct smear examination for acid-fast bacilli, cultural identification in Lowestein-Jensen (L-J) medium, the radiometric BACTEC 460 system, and Polymerase Chain Reaction (PCR). Three hundred and ninety-three clinical samples of sputum (375), gastric aspirate (3), pleural fluid (12) and urine (3) were taken from 125 patients hospitalized at our Institute for suspected pulmonary tuberculosis, between January 1995 and June 1997. On completion of diagnosis, 35 were found to be affected by active tuberculosis (30 pulmonary, 4 pleural and 1 urinary) and 90 by other non-tubercular diseases (pneumonia, lung cancer, non-tubercular pleural effusion, etc.). In our study, direct smear examination for acid-smear bacilli gave diagnostic value results of 88% and positive predictive value of 91.67%. Cultural identification in L-J and BACTEC 460 TB radiometric system media resulted in diagnostic values of 96.80% and 94.40%, respectively, and positive predictive values of 100% for both of them. Finally, One-Tube Nested-PCR, a variant which uses specific primers for the IS6110 insertion sequence specific for Mycobacterium tuberculosis, gave us 88.80% (91.43% sensitivity and 87.78% specificity) diagnostic value results, and 74.42% (11 false-positives) positive predictive value. On the basis of our results, we can affirm that PCR is a good method for microbiological diagnosis of tuberculosis, given its high sensitivity and specificity and unparalleled rapidity. However, the high number of false-positives that we found suggests that results obtained should be confirmed with BACTEC, which considerably reduces the time required for identification, and makes it possible to carry out an antibiotic assay rapidly.
M. tuberculosis
Eing BR, Becker A, Sohns A, Ringelmann R
J Clin Microbiol. 1998;36:2023-2029
The new Roche Cobas Amplicor Mycobacterium tuberculosis assay, which is a semiautomated version of the manually performed Roche Amplicor M. tuberculosis test, was compared to culture and an IS6110-based in-house PCR protocol. A total of 1,681 specimens from 833 patients, including specimen types other than sputum, were tested in parallel by both the in-house PCR and the Cobas Amplicor M. tuberculosis assay. After we resolved discrepant PCR results, the sensitivity, specificity, and positive and negative predictive values for the Cobas Amplicor M. tuberculosis assay were 66.33, 99.71, 94.36, and 97.66%, respectively. The corresponding values for the in-house PCR were 91.08, 99.85, 97.87, and 99.37%, respectively. For culture- and smear-positive specimens, the sensitivity of the Cobas Amplicor M. tuberculosis test was 96.42% (in-house PCR, 100%). If only smear-negative sputum specimens were considered, the Cobas Amplicor M. tuberculosis assay exhibited a sensitivity of 45.45% (in-house PCR, 63.63%) relative to that of culture. With a modified protocol for DNA extraction (washing of samples plus ultrasonication), both PCR methods performed better with gastric aspirates than with sputum samples (sensitivity of the Cobas Amplicor M. tuberculosis assay with smear-negative gastric aspirates, 70.00%; sensitivity of in-house PCR, 90.00%). With dithiothreitol being used for liquefaction of specimens in this study, the Cobas Amplicor M. tuberculosis assay exhibited an inhibition rate of 9.16%. In our view, the new Cobas Amplicor M. tuberculosis test (i) is well suited for typing of smear-positive specimens, (ii) may also be applied to gastric aspirates and other types of specimens if DNA extraction methods are modified appropriately, and (iii) exhibits a sensitivity with smear-negative sputum specimens which makes it recommendable that a minimum of three samples from the same patient be tested.
Mycobacterium tuberculosis by PCR Analysis of Urine and Other Clinical Samples from AIDS and Non-HIV-Infected Patients
Sechi LA, Pinna MP, Sanna A, et al
Mol Cell Probes. 1997;11:281-285
A number of different clinical specimens, such as sputum, cerebrospinal fluid and blood, have been reported to be good substrates for the detection of Mycobacterium tuberculosis by PCR assay. We wanted to search for the presence of mycobacteria in other body fluids, such as urine. Urine samples and other samples obtained from AIDS patients and non HIV-infected patients were analysed by PCR. The results were compared with those obtained using conventional methods (Bactec 460 TB and AFB (acid fast bacilli strain)). We analysed 412 urine samples and 210 different other samples (sputum and cerebrospinal fluid) obtained from AIDS patients by PCR; almost identical levels of PCR-positive (14-17%) results were observed in all samples analysed. The results were then compared with those obtained with the Bactec 460 TB and AFB. PCR, Bactec 460 TB and acid fast stain were also used to analyse 190 urine samples and 230 other samples from non-HIV infected patients in the consumption ward of Sassari Hospital. The number of urine samples positive by PCR (6.3%) and Bactec 460 TB (2.1%) was half that obtained from samples taken from the AIDS patients. As expected, an increase in the number of positive sputum samples was observed with all methods. The results indicate that PCR analysis of urine samples represents a valid alternative for fast and sensitive detection of M. tuberculosis. This method can be routinely used in the clinical laboratory, especially in HIV-infected patients.
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