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RIF assay and detection of false-positive rifampicin resistance in Mycobacterium tuberculosis
Diagnostic Microbiology and Infectious Disease 74 (2012) 207–209
Contents lists available at SciVerse ScienceDirect
Diagnostic Microbiology and Infectious Disease j o u r n a l h o m e p a g e : w w w. e l s e v i e r. c o m / l o c a t e / d i a g m i c r o b i o
An evaluation of the Xpert MTB/RIF assay and detection of false-positive rifampicin resistance in Mycobacterium tuberculosis☆,☆☆ Deborah A. Williamson ⁎, Indira Basu, James Bower, Joshua T. Freeman, Gillian Henderson, Sally A. Roberts Department of Clinical Microbiology, Auckland District Health Board, Auckland, New Zealand
a r t i c l e
i n f o
Article history: Received 16 April 2012 Accepted 16 June 2012 Available online 20 July 2012 Keywords: Mycobacterium tuberculosis Resistance Rifampicin Xpert MTB/RIF assay
a b s t r a c t Recent reports suggest that false-positive rifampicin resistance may be assigned by the Xpert MTB/RIF assay. We analysed 169 specimens using the MTB/RIF assay. Using culture as the gold standard, we found that the assay had 100% sensitivity and specificity for detecting M. tuberculosis. However, we found that the assay incorrectly assigned rifampicin resistance in 4/13 (31%) of cases. © 2012 Elsevier Inc. All rights reserved.
Multidrug-resistant tuberculosis (MDR-TB) is a significant global health threat. An important goal of control programmes for MDR-TB is a reduction in the time taken for laboratory diagnosis. Recent advances in molecular diagnostics based on detection of mutations in an 81-base pair region of the rpoB gene have led to the development of rapid molecular assays for rifampicin resistance, which is frequently used as a surrogate marker for MDR-TB (Telenti et al., 1993). In particular, the Cepheid Xpert® MTB/RIF assay (Cepheid, Sunnyvale, CA, USA) has demonstrated high sensitivity and specificity for detection of both M. tuberculosis and rifampicin resistance (Boehme et al., 2010, 2011). The World Health Organization (WHO) has endorsed the MTB/RIF assay as the initial diagnostic test for patients suspected of having MDR-TB or co-infection with human immunodeficiency virus (HIV) (Trébucq et al., 2011). As a consequence, it seems likely that the number of TB cases diagnosed by the MTB/RIF assay will increase over the forthcoming years (Trébucq et al., 2011). In addition, increased use of the MTB/RIF assay is likely to result in increased diagnosis of MDR-TB and subsequent use of second-line antituberculous agents. Here, we describe an evaluation of the of the MTB/RIF assay, with particular attention given to the specificity of the assay for detecting rifampicin resistance. Between December 2009 and November 2011, all acid-fast bacilli (AFB) smear-positive respiratory specimens sent to the Mycobacteriology Laboratory, Auckland City Hospital, New Zealand, were processed and analysed as previously described using the Xpert® MTB/RIF assay (version 3.0) (Boehme et al., 2010). In patients for whom there was a ☆ Funding: This study was supported by internal funding. ☆☆ Conflicts of interests: None for all authors. ⁎ Corresponding author. Tel.: + 64-9-307-4949x23260; fax: + 64-9-307-4922. E-mail address: [email protected] (D.A. Williamson). 0732-8893/$ – see front matter © 2012 Elsevier Inc. All rights reserved. doi:10.1016/j.diagmicrobio.2012.06.013
possible clinical indication of MDR-TB (e.g., immigration from a highprevalence area; previous treatment for TB), smear-positive extrapulmonary specimens were also tested using the MTB/RIF assay. Nonsterile specimens underwent processing using the 4% sodium hydroxide decontamination method, and all specimens were cultured in the BACTEC Mycobacterial Growth Indicator Tube (MGIT) 960 system
Table 1 Results of MTB/RIF assay and Mycobacterium tuberculosis (MTB) culture. Specimen type
No. of specimens tested by MTB/RIF assay
Respiratory (AFB-positive) Sputum 80 Bronchoalveolar 2 lavage fluid Bronchial washings 6 Tracheal aspirate 1 Total 89 Extrapulmonary (AFB-positive) Lymph node 5 Tissue 3 Middle ear washings 1 Total 9 Positive MGIT culture vials 71 Total 169 a b c
Culture results No. of specimens Positive for Negative for positive MTB MTB but for MTB positive by MTB/RIF for other assay mycobacterial species 62 2
62 2
18a
2 1 67
2 1 67
4b 22
5 3 1 9 65 141
5 3 1 9 65 141
6c 28
Two Mycobacterium avium; 3 M. kansasii; 2 M. abscessus; 1 M. xenopii. Four M. avium. Two M. avium; 2 M. gordonae; 1 M. fortuitum; 1 M. chelonae.
208
D.A. Williamson et al. / Diagnostic Microbiology and Infectious Disease 74 (2012) 207–209
Table 2 Specimens with rifampicin resistance as detected by the MTB/RIF assay. Specimen
Type
1 2 3 4 5a
Culture Culture Culture Culture Culture
6 7 8a 9 10 11 12 13 14 a
Basis of rifampicin resistance as detected by the MTB/RIF assay
Rifampicin phenotypic susceptibility result (1.0 mg/L)
Mutation(s) detected by rpoB gene amplification and sequencing
Probe E did not bind Probe E did not bind Probe E did not bind Probe D did not bind Probes D and E did not bind
R R R R S
Middle ear washings Sputum Culture isolate
Probe E did not bind Probe D did not bind Probe B did not bind
R S S
Sputum Sputum Neck aspirate Culture isolate Tissue Culture isolate
Probe D did not bind Probe B did not bind CT max 5.6 Probes A, D, and E did not bind Probe A did not bind Probes D and E did not bind
R R S S S S
Ser 531 → Leu (TCG → TTG) Ser 531 → Leu (TCG→TTG) Ser 531 → Leu (TCG→TTG) His 526 → Leu (CAC → TTA) His 526 → Asn (CAC → AAC) Ala 532 → Val (GCG → GTG) Ser 531 → Leu (TCG→TTG) No mutation detected Leu 511 → Pro (CTG → CCG) Met 515 → Ile (ATG →ATA) His 526 → Tyr (CAC → TAC) Asp 516 → Val (GAC → GTC) No mutation detected No mutation detected Gln 510 → Gln (CAG → CAA) No mutation detected
isolate isolate isolate isolate isolate
Clinical significance of these mutations described in Williamson et al. (2012).
(BD, Franklin Lakes, NJ, USA) and on Lowenstein-Jensen (LJ) solid media (Fort Richard, Auckland, New Zealand). In addition, samples with a positive growth reading in the MGIT 960 system and AFB subsequently detected in the liquid culture media were also tested by the MTB/RIF assay if there was a clinical possibility of MDR-TB. An in-house DNA extraction method was used to extract genomic DNA from positive MGIT culture vials, as previously described (Williamson et al., 2012). All M. tuberculosis isolates had standard first-line drug susceptibility testing (DST) performed in the MGIT 960 system, according to the manufacturer's instructions (Siddiqi and Rüsch-Gerdes, 2006). Rifampicin susceptibility was tested at the standard critical concentration of 1 mg/L (Siddiqi and Rüsch-Gerdes, 2006). All isolates that had rifampicin resistance detected by phenotypic methods and/or the MTB/RIF assay underwent confirmatory rpoB gene amplification and DNA sequencing as previously described (Williamson et al., 2012). A total of 169 specimens (89 smear-positive respiratory specimens; 9 smear-positive extra-pulmonary specimens and 71 positive MGIT liquid culture vials) from 169 patients were analysed (Table 1). Culture results were available for all clinical specimens (Table 1). With the use of culture as the “gold standard”, the overall sensitivity and specificity of the MTB/RIF assay for the detection of M. tuberculosis were 100% (141/141) and 100% (28/28), respectively. The MTB/RIF assay detected rifampicin resistance in 13/169 (7.7%) specimens. However, using standard phenotypic methods, rifampicin resistance was detected in only 7/13 (54%) isolates (Table 2). In 2 of the remaining 6 isolates, amplification and sequencing of the rpoB gene revealed mutations associated with increased but low-level rifampicin resistance. The clinical significance of such mutations has been previously described by our group (Williamson et al., 2012). In 3 of the remaining 4 isolates, despite failure of 1 or more of the rpoB target probes to hybridise adequately, sequencing of the rpoB gene revealed no mutations (Table 2). In the fourth isolate, rpoB sequencing revealed a silent CAG (Gln) to CAA (Gln) mutation at codon 510. This mutation resulted in failure of the corresponding probe (probe A) to hybridise and an interpretation of rifampicin resistance by the MTB/RIF assay. Similar to previous studies, we have found the MTB/RIF assay highly sensitive and specific for the detection of M. tuberculosis, when used for both smear-positive pulmonary and extrapulmonary specimens as well as for isolates in liquid culture media. However, we found that the assay was less reliable for the detection of rifampicin resistance, producing false-positive results in 4/13 (31%) specimens. False-positive rifampicin resistance results obtained by the MTB/RIF assay have been previously described and were resolved in 1 study by raising the amplification cycle threshold maximum (ΔCT max) value from 3.5 to 5.0, following changes made to the assay software in
version 4.0 (Lawn et al., 2011; Scott et al., 2011). In our series, however, raising the ΔCT max to 5.0 would not have reclassified any of our false-positive results as rifampicin-susceptible. Notably, in 1 isolate, the presence of a silent mutation at Gln510 also resulted in failure of the corresponding probe to hybridise. Previous authors have described this phenomenon in relation to a TTC (Phe)-to-TTT (Phe) mutation at codon 514 (Alonso et al., 2011; Moure et al., 2011). Given the inherent redundancy in the genetic code, it is possible that additional silent mutations within the rpoB gene may also prevent hybridisation of rpoB probes. Importantly, only 1 of the 4 falsepositive results we obtained was from a pulmonary specimen. Recent studies have highlighted the utility of the MTB/RIF assay in detecting M. tuberculosis from a range of extrapulmonary specimens (Alonso et al., 2011). Further work is therefore required to determine whether false-positive rifampicin resistance results are more likely to occur when the assay is used on pulmonary or extrapulmonary specimens. In our setting, as in others, the MTB/RIF assay has become a valuable first-line assay for the detection of M. tuberculosis. However, we found that the assay incorrectly assigned rifampicin resistance in one-third of cases. Importantly, one of the features of the MTB/RIF assay is its potential utility in resource-poor settings. In such settings, there may be limited, if any, access to confirmatory phenotypic or genotypic drug susceptibility testing. Without confirmatory testing, it is possible that MDR-TB cases may be overdiagnosed, resulting in suboptimal treatment regimens. Further work is therefore required to evaluate the performance of the MTB/RIF assay for the detection of rifampicin resistance in a range of clinical settings and on a range of specimen types. Acknowledgment The authors would like to thank Dr Ivan Bastian for kindly reviewing the manuscript. References Alonso M, Palacios JJ, Herranz M, et al. Isolation of Mycobacterium tuberculosis strains with a silent mutation in rpoB leading to potential misassignment of resistance category. J Clin Microbiol 2011;49:2688–90. Boehme CC, Nabeta P, Hillemann D, et al. Rapid molecular detection of tuberculosis and rifampin resistance. N Engl J Med 2010;363:1005–15. Boehme CC, Nicol MP, Nabeta P, et al. Feasibility, diagnostic accuracy, and effectiveness of decentralised use of the Xpert MTB/RIF test for diagnosis of tuberculosis and multidrug resistance: a multicentre implementation study. Lancet 2011;377: 1495–505. Lawn SD, Brooks SV, Kranzer K, et al. Screening for HIV-associated tuberculosis and rifampicin resistance before antiretroviral therapy using the Xpert MTB/RIF assay: a prospective study. PLoS Med 2011;8:e1001067.
D.A. Williamson et al. / Diagnostic Microbiology and Infectious Disease 74 (2012) 207–209 Moure R, Martín R, Alcaide F. Silent mutation in rpoB detected from clinical samples with rifampin-susceptible Mycobacterium tuberculosis. J Clin Microbiol 2011;49: 3722. Scott LE, McCarthy K, Gous N, et al. Comparison of Xpert MTB/RIF with other nucleic acid technologies for diagnosing pulmonary tuberculosis in a high HIV prevalence setting: a prospective study. PLoS Med 2011;8:e1001061. Siddiqi SH, Rüsch-Gerdes S. MGIT Procedure manual. Geneva, Switzerland: Foundation for Innovative New Diagnostics; 2006.
209
Telenti A, Imboden P, Marchesi F, et al. Detection of rifampicin-resistance mutations in Mycobacterium tuberculosis. Lancet 1993;341:647–50. Trébucq A, Enarson DA, Chiang CY, et al. Xpert® MTB/RIF for national tuberculosis programmes in low-income countries: when, where and how? Int J Tuberc Lung Dis 2011;15:1567–72. Williamson DA, Roberts SA, Bower JE, et al. Clinical failures associated with rpoB mutations in phenotypically occult multidrug-resistant Mycobacterium tuberculosis. Int J Tuberc Lung Dis 2012;16:216–20.
Contents lists available at SciVerse ScienceDirect
Diagnostic Microbiology and Infectious Disease j o u r n a l h o m e p a g e : w w w. e l s e v i e r. c o m / l o c a t e / d i a g m i c r o b i o
An evaluation of the Xpert MTB/RIF assay and detection of false-positive rifampicin resistance in Mycobacterium tuberculosis☆,☆☆ Deborah A. Williamson ⁎, Indira Basu, James Bower, Joshua T. Freeman, Gillian Henderson, Sally A. Roberts Department of Clinical Microbiology, Auckland District Health Board, Auckland, New Zealand
a r t i c l e
i n f o
Article history: Received 16 April 2012 Accepted 16 June 2012 Available online 20 July 2012 Keywords: Mycobacterium tuberculosis Resistance Rifampicin Xpert MTB/RIF assay
a b s t r a c t Recent reports suggest that false-positive rifampicin resistance may be assigned by the Xpert MTB/RIF assay. We analysed 169 specimens using the MTB/RIF assay. Using culture as the gold standard, we found that the assay had 100% sensitivity and specificity for detecting M. tuberculosis. However, we found that the assay incorrectly assigned rifampicin resistance in 4/13 (31%) of cases. © 2012 Elsevier Inc. All rights reserved.
Multidrug-resistant tuberculosis (MDR-TB) is a significant global health threat. An important goal of control programmes for MDR-TB is a reduction in the time taken for laboratory diagnosis. Recent advances in molecular diagnostics based on detection of mutations in an 81-base pair region of the rpoB gene have led to the development of rapid molecular assays for rifampicin resistance, which is frequently used as a surrogate marker for MDR-TB (Telenti et al., 1993). In particular, the Cepheid Xpert® MTB/RIF assay (Cepheid, Sunnyvale, CA, USA) has demonstrated high sensitivity and specificity for detection of both M. tuberculosis and rifampicin resistance (Boehme et al., 2010, 2011). The World Health Organization (WHO) has endorsed the MTB/RIF assay as the initial diagnostic test for patients suspected of having MDR-TB or co-infection with human immunodeficiency virus (HIV) (Trébucq et al., 2011). As a consequence, it seems likely that the number of TB cases diagnosed by the MTB/RIF assay will increase over the forthcoming years (Trébucq et al., 2011). In addition, increased use of the MTB/RIF assay is likely to result in increased diagnosis of MDR-TB and subsequent use of second-line antituberculous agents. Here, we describe an evaluation of the of the MTB/RIF assay, with particular attention given to the specificity of the assay for detecting rifampicin resistance. Between December 2009 and November 2011, all acid-fast bacilli (AFB) smear-positive respiratory specimens sent to the Mycobacteriology Laboratory, Auckland City Hospital, New Zealand, were processed and analysed as previously described using the Xpert® MTB/RIF assay (version 3.0) (Boehme et al., 2010). In patients for whom there was a ☆ Funding: This study was supported by internal funding. ☆☆ Conflicts of interests: None for all authors. ⁎ Corresponding author. Tel.: + 64-9-307-4949x23260; fax: + 64-9-307-4922. E-mail address: [email protected] (D.A. Williamson). 0732-8893/$ – see front matter © 2012 Elsevier Inc. All rights reserved. doi:10.1016/j.diagmicrobio.2012.06.013
possible clinical indication of MDR-TB (e.g., immigration from a highprevalence area; previous treatment for TB), smear-positive extrapulmonary specimens were also tested using the MTB/RIF assay. Nonsterile specimens underwent processing using the 4% sodium hydroxide decontamination method, and all specimens were cultured in the BACTEC Mycobacterial Growth Indicator Tube (MGIT) 960 system
Table 1 Results of MTB/RIF assay and Mycobacterium tuberculosis (MTB) culture. Specimen type
No. of specimens tested by MTB/RIF assay
Respiratory (AFB-positive) Sputum 80 Bronchoalveolar 2 lavage fluid Bronchial washings 6 Tracheal aspirate 1 Total 89 Extrapulmonary (AFB-positive) Lymph node 5 Tissue 3 Middle ear washings 1 Total 9 Positive MGIT culture vials 71 Total 169 a b c
Culture results No. of specimens Positive for Negative for positive MTB MTB but for MTB positive by MTB/RIF for other assay mycobacterial species 62 2
62 2
18a
2 1 67
2 1 67
4b 22
5 3 1 9 65 141
5 3 1 9 65 141
6c 28
Two Mycobacterium avium; 3 M. kansasii; 2 M. abscessus; 1 M. xenopii. Four M. avium. Two M. avium; 2 M. gordonae; 1 M. fortuitum; 1 M. chelonae.
208
D.A. Williamson et al. / Diagnostic Microbiology and Infectious Disease 74 (2012) 207–209
Table 2 Specimens with rifampicin resistance as detected by the MTB/RIF assay. Specimen
Type
1 2 3 4 5a
Culture Culture Culture Culture Culture
6 7 8a 9 10 11 12 13 14 a
Basis of rifampicin resistance as detected by the MTB/RIF assay
Rifampicin phenotypic susceptibility result (1.0 mg/L)
Mutation(s) detected by rpoB gene amplification and sequencing
Probe E did not bind Probe E did not bind Probe E did not bind Probe D did not bind Probes D and E did not bind
R R R R S
Middle ear washings Sputum Culture isolate
Probe E did not bind Probe D did not bind Probe B did not bind
R S S
Sputum Sputum Neck aspirate Culture isolate Tissue Culture isolate
Probe D did not bind Probe B did not bind CT max 5.6 Probes A, D, and E did not bind Probe A did not bind Probes D and E did not bind
R R S S S S
Ser 531 → Leu (TCG → TTG) Ser 531 → Leu (TCG→TTG) Ser 531 → Leu (TCG→TTG) His 526 → Leu (CAC → TTA) His 526 → Asn (CAC → AAC) Ala 532 → Val (GCG → GTG) Ser 531 → Leu (TCG→TTG) No mutation detected Leu 511 → Pro (CTG → CCG) Met 515 → Ile (ATG →ATA) His 526 → Tyr (CAC → TAC) Asp 516 → Val (GAC → GTC) No mutation detected No mutation detected Gln 510 → Gln (CAG → CAA) No mutation detected
isolate isolate isolate isolate isolate
Clinical significance of these mutations described in Williamson et al. (2012).
(BD, Franklin Lakes, NJ, USA) and on Lowenstein-Jensen (LJ) solid media (Fort Richard, Auckland, New Zealand). In addition, samples with a positive growth reading in the MGIT 960 system and AFB subsequently detected in the liquid culture media were also tested by the MTB/RIF assay if there was a clinical possibility of MDR-TB. An in-house DNA extraction method was used to extract genomic DNA from positive MGIT culture vials, as previously described (Williamson et al., 2012). All M. tuberculosis isolates had standard first-line drug susceptibility testing (DST) performed in the MGIT 960 system, according to the manufacturer's instructions (Siddiqi and Rüsch-Gerdes, 2006). Rifampicin susceptibility was tested at the standard critical concentration of 1 mg/L (Siddiqi and Rüsch-Gerdes, 2006). All isolates that had rifampicin resistance detected by phenotypic methods and/or the MTB/RIF assay underwent confirmatory rpoB gene amplification and DNA sequencing as previously described (Williamson et al., 2012). A total of 169 specimens (89 smear-positive respiratory specimens; 9 smear-positive extra-pulmonary specimens and 71 positive MGIT liquid culture vials) from 169 patients were analysed (Table 1). Culture results were available for all clinical specimens (Table 1). With the use of culture as the “gold standard”, the overall sensitivity and specificity of the MTB/RIF assay for the detection of M. tuberculosis were 100% (141/141) and 100% (28/28), respectively. The MTB/RIF assay detected rifampicin resistance in 13/169 (7.7%) specimens. However, using standard phenotypic methods, rifampicin resistance was detected in only 7/13 (54%) isolates (Table 2). In 2 of the remaining 6 isolates, amplification and sequencing of the rpoB gene revealed mutations associated with increased but low-level rifampicin resistance. The clinical significance of such mutations has been previously described by our group (Williamson et al., 2012). In 3 of the remaining 4 isolates, despite failure of 1 or more of the rpoB target probes to hybridise adequately, sequencing of the rpoB gene revealed no mutations (Table 2). In the fourth isolate, rpoB sequencing revealed a silent CAG (Gln) to CAA (Gln) mutation at codon 510. This mutation resulted in failure of the corresponding probe (probe A) to hybridise and an interpretation of rifampicin resistance by the MTB/RIF assay. Similar to previous studies, we have found the MTB/RIF assay highly sensitive and specific for the detection of M. tuberculosis, when used for both smear-positive pulmonary and extrapulmonary specimens as well as for isolates in liquid culture media. However, we found that the assay was less reliable for the detection of rifampicin resistance, producing false-positive results in 4/13 (31%) specimens. False-positive rifampicin resistance results obtained by the MTB/RIF assay have been previously described and were resolved in 1 study by raising the amplification cycle threshold maximum (ΔCT max) value from 3.5 to 5.0, following changes made to the assay software in
version 4.0 (Lawn et al., 2011; Scott et al., 2011). In our series, however, raising the ΔCT max to 5.0 would not have reclassified any of our false-positive results as rifampicin-susceptible. Notably, in 1 isolate, the presence of a silent mutation at Gln510 also resulted in failure of the corresponding probe to hybridise. Previous authors have described this phenomenon in relation to a TTC (Phe)-to-TTT (Phe) mutation at codon 514 (Alonso et al., 2011; Moure et al., 2011). Given the inherent redundancy in the genetic code, it is possible that additional silent mutations within the rpoB gene may also prevent hybridisation of rpoB probes. Importantly, only 1 of the 4 falsepositive results we obtained was from a pulmonary specimen. Recent studies have highlighted the utility of the MTB/RIF assay in detecting M. tuberculosis from a range of extrapulmonary specimens (Alonso et al., 2011). Further work is therefore required to determine whether false-positive rifampicin resistance results are more likely to occur when the assay is used on pulmonary or extrapulmonary specimens. In our setting, as in others, the MTB/RIF assay has become a valuable first-line assay for the detection of M. tuberculosis. However, we found that the assay incorrectly assigned rifampicin resistance in one-third of cases. Importantly, one of the features of the MTB/RIF assay is its potential utility in resource-poor settings. In such settings, there may be limited, if any, access to confirmatory phenotypic or genotypic drug susceptibility testing. Without confirmatory testing, it is possible that MDR-TB cases may be overdiagnosed, resulting in suboptimal treatment regimens. Further work is therefore required to evaluate the performance of the MTB/RIF assay for the detection of rifampicin resistance in a range of clinical settings and on a range of specimen types. Acknowledgment The authors would like to thank Dr Ivan Bastian for kindly reviewing the manuscript. References Alonso M, Palacios JJ, Herranz M, et al. Isolation of Mycobacterium tuberculosis strains with a silent mutation in rpoB leading to potential misassignment of resistance category. J Clin Microbiol 2011;49:2688–90. Boehme CC, Nabeta P, Hillemann D, et al. Rapid molecular detection of tuberculosis and rifampin resistance. N Engl J Med 2010;363:1005–15. Boehme CC, Nicol MP, Nabeta P, et al. Feasibility, diagnostic accuracy, and effectiveness of decentralised use of the Xpert MTB/RIF test for diagnosis of tuberculosis and multidrug resistance: a multicentre implementation study. Lancet 2011;377: 1495–505. Lawn SD, Brooks SV, Kranzer K, et al. Screening for HIV-associated tuberculosis and rifampicin resistance before antiretroviral therapy using the Xpert MTB/RIF assay: a prospective study. PLoS Med 2011;8:e1001067.
D.A. Williamson et al. / Diagnostic Microbiology and Infectious Disease 74 (2012) 207–209 Moure R, Martín R, Alcaide F. Silent mutation in rpoB detected from clinical samples with rifampin-susceptible Mycobacterium tuberculosis. J Clin Microbiol 2011;49: 3722. Scott LE, McCarthy K, Gous N, et al. Comparison of Xpert MTB/RIF with other nucleic acid technologies for diagnosing pulmonary tuberculosis in a high HIV prevalence setting: a prospective study. PLoS Med 2011;8:e1001061. Siddiqi SH, Rüsch-Gerdes S. MGIT Procedure manual. Geneva, Switzerland: Foundation for Innovative New Diagnostics; 2006.
209
Telenti A, Imboden P, Marchesi F, et al. Detection of rifampicin-resistance mutations in Mycobacterium tuberculosis. Lancet 1993;341:647–50. Trébucq A, Enarson DA, Chiang CY, et al. Xpert® MTB/RIF for national tuberculosis programmes in low-income countries: when, where and how? Int J Tuberc Lung Dis 2011;15:1567–72. Williamson DA, Roberts SA, Bower JE, et al. Clinical failures associated with rpoB mutations in phenotypically occult multidrug-resistant Mycobacterium tuberculosis. Int J Tuberc Lung Dis 2012;16:216–20.