Direct Application of the INNO-LiPA Rif.TB Line-Probe Assay for Rapid Identification of
            <i>Mycobacterium tuberculosis</i>
            Complex Strains and Detection of Rifampin Resistance in 360 Smear-Positive Respiratory Specimens from an Area of High Incidence of Multidrug-Resistant Tuberculosis

Viveiros, Miguel; Leandro, Clara; Rodrigues, Liliana; Almeida, J. M. Das Neves; Bettencourt, Rosário; Couto, Isabel; Carrilho, Lurdes; Diogo, José; Fonseca, Ana Mafalda; Lito, Luı́s; Lopes, João Roberto Spotti; Pacheco, Teresa; Pessanha, Mariana; Quirim, Judite; Sancho, Luísa; Salfinger, Max; Amaral, Leonard

doi:10.1128/jcm.43.9.4880-4884.2005

Cited by 62 publications

(37 citation statements)

References 24 publications

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“…The test displays excellent specificity and good sensitivity, similar to those of other genetic tests for RIF resistance, such as line probe or molecular beacon tests (8,11,14,16,17). The observed discrepancies between the results of conventional DST and the TB-Biochip system (all were falsely called susceptible with the TB-Biochip system) likely result from the large number of mutations found in RIF-resistant isolates and the limited range of mutations included on the biochip.…”

Section: Discussionmentioning

confidence: 88%

Evaluation of the TB-Biochip Oligonucleotide Microarray System for Rapid Detection of Rifampin Resistance in Mycobacterium tuberculosis

et al. 2006

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The TB-Biochip oligonucleotide microarray system is a rapid system to detect mutations associated with rifampin (RIF) resistance in mycobacteria. After optimizing the system with 29 laboratory-generated rifampinresistant mutants of Mycobacterium tuberculosis, we evaluated the performance of this test using 75 clinical isolates of Mycobacterium tuberculosis. With this small sample set, the TB-Biochip system displayed a sensitivity of 80% and a specificity of 100% relative to conventional drug susceptibility testing results for RIF resistance. For these samples (ϳ50% tested positive), the positive predictive value was 100% and the negative predictive value was 85%. Four of the seven observed discrepancies were attributed to rare and new mutations not represented in the microarray, while three of the strains with discrepant results did not carry mutations in the RIF resistance-determining region. The results of this study confirm the utility of the system for rapid detection of RIF resistance and suggest approaches to increasing its sensitivity.The emergence of multidrug-resistant tuberculosis (MDR TB) presents a threat to global TB control efforts (15,19,20). MDR TB is defined as resistant to at least isoniazid and rifampin (RIF), the two most active first-line antimicrobial agents against TB (19,20). Rapid and reliable drug susceptibility testing is essential for the prompt initiation of appropriate therapy, which rapidly renders patients noninfectious and hence facilitates infection control measures to halt transmission (1).Rifampin is the mainstay of short-course chemotherapy for TB, and the loss of rifampin as an effective drug leads to a need for a longer duration of therapy and often to a lower cure rate (19,20). In Mycobacterium tuberculosis, resistance to RIF results from mutations in the ␤ subunit of RNA polymerase, which is encoded by the rpoB gene (2,15). Approximately 95% of RIF-resistant strains carry mutations within the RIF resistance-determining region (RRDR), an 81-bp region carrying codons 507 through 533 of the rpoB gene (9,10,15). In this study, the TB-Biochip oligonucleotide microarray system (10) (Engelhardt Institute of Molecular Biology, Moscow, Russia) was used to detect RIF resistance among TB clinical isolates within a 24-hour period.The TB-Biochip oligonucleotide microarray system is designed to detect and identify 29 codon substitutions and 1 codon deletion distributed over 10 codon positions within the RRDR (10) (Fig. 1). These 30 mutations are found in Ͼ90% of RIF-resistant strains that have mutations in the RRDR (3-6, 10, 15). Each element (an acrylamide gel pad) of the microarray contains an immobilized oligonucleotide whose sequence matches that of either a wild-type or mutated segment of the RRDR. The use of acrylamide gel pads reduces the cost of each microarray to less than five U.S. dollars and increases the amount of oligonucleotide target that is present on the microarray, which increases the robustness of the hybridization reaction. Hybridization of the microarray with fluo...

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Section: Discussionmentioning

confidence: 88%

Evaluation of the TB-Biochip Oligonucleotide Microarray System for Rapid Detection of Rifampin Resistance in Mycobacterium tuberculosis

et al. 2006

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“…However, the single-use cartridge design of the Xpert assay limits its use for laboratory-based high-throughput testing. Several widely used reverse blot hybridization assays, such as the INNO-LIPA Rif.TB assay (Innogenetics, Belgium) and the MTBDRplus (Hain, Germany) assay (1,5,15,19,21,23,29,34), are available for laboratory-based rifampin resistance screening; however, these assays are complicated by their open hybridization format. Open hybridization systems require a relatively cumbersome work process, including rigorous physical separation of different work areas (2,23) due to the risk of handling open PCR amplicons in a molecular diagnostic laboratory.…”

mentioning

confidence: 99%

Rapid, High-Throughput Detection of Rifampin Resistance and Heteroresistance in Mycobacterium tuberculosis by Use of Sloppy Molecular Beacon Melting Temperature Coding

et al. 2012

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M ultidrug-resistant (MDR) and extensively drug-resistant (XDR) Mycobacterium tuberculosis is increasing worldwide (9, 27, 37). Rapid methods to detect drug resistance are needed to quickly identify drug-resistant strains and to implement appropriate therapy (4,16,36). M. tuberculosis does not naturally contain plasmids, and almost all cases of clinical drug resistance are caused by single-nucleotide polymorphisms (SNPs) or small insertions/deletions in relevant genes (28). In the case of rifampin resistance, 95 to 98% of rifampin-resistant clinical strains have mutations in the 80-bp rifampin resistance determining region (RRDR) of the M. tuberculosis RNA polymerase beta (rpoB) gene (9,11,12,15,20,28). PCR and probe-based molecular genotyping assays can be used to detect these resistance-inducing mutations. Such genotypic assays are potentially more rapid than labor-intensive culture-based drug susceptibility tests. Genotypic drug susceptibility testing has shown good overall concordance with the phenotypic antibiotic susceptibility tests of MDR and XDR clinical strains (6), and in a recent study, a genotypic test actually correlated better with clinical outcome than standard phenotypic susceptibility testing (36). When combined with automated sample processing systems, such as the Xpert MTB/RIF test (14), genotypic susceptibility tests can significantly reduce testing turnaround time, increasing patient notification rates and decreasing time to treatment (4, 32).The Xpert MTB/RIF assay is one example of a genotypic test that is being increasingly used to screen for rifampin resistance (33). However, the single-use cartridge design of the Xpert assay limits its use for laboratory-based high-throughput testing. Several widely used reverse blot hybridization assays, such as the INNO-LIPA Rif.TB assay (Innogenetics, Belgium) and the MTBDRplus (Hain, Germany) assay (1,5,15,19,21,23,29,34), are available for laboratory-based rifampin resistance screening; however, these assays are complicated by their open hybridization format. Open hybridization systems require a relatively cumbersome work process, including rigorous physical separation of different work areas (2, 23) due to the risk of handling open PCR amplicons in a molecular diagnostic laboratory. Open systems also require a relatively large number of probes to test for relevant resistanceassociated mutations. This requirement complicates assay chemistry and hybridization parameters. In contrast to reverse blot hy-

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“…Because resistance to Rif is almost always accompanied by resistance to INH [10], resistance to Rif acts as a surrogate marker for identification of MDR-TB [10]. The presumptive identification of MDR-TB can be done within a single day [10] and this therefore can rapidly identify patients in the need of intensified therapy, whilst pan-susceptible infections can readily be managed with the four first line drugs recommended by the WHO. It should be stated that in some geographic areas with a high prevalence of MDR-TB, each case of tuberculosis is nowadays treated more intensively with second line drugs [11].…”

Section: General Approachmentioning

confidence: 99%

“…The determination of resistance to Rif is afforded by the identification of mutations within the beta subunit of the rpoB gene [9]. Because resistance to Rif is almost always accompanied by resistance to INH [10], resistance to Rif acts as a surrogate marker for identification of MDR-TB [10]. The presumptive identification of MDR-TB can be done within a single day [10] and this therefore can rapidly identify patients in the need of intensified therapy, whilst pan-susceptible infections can readily be managed with the four first line drugs recommended by the WHO.…”

Section: General Approachmentioning

confidence: 99%

Advances in Personalised Treatment of Multi-drug Resistant Tuberculosis

Amaral

Soolingen²

2014

Biochem Pharmacol (Los Angel)

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Direct Application of the INNO-LiPA Rif.TB Line-Probe Assay for Rapid Identification of Mycobacterium tuberculosis Complex Strains and Detection of Rifampin Resistance in 360 Smear-Positive Respiratory Specimens from an Area of High Incidence of Multidrug-Resistant Tuberculosis

Cited by 62 publications

References 24 publications

Evaluation of the TB-Biochip Oligonucleotide Microarray System for Rapid Detection of Rifampin Resistance in Mycobacterium tuberculosis

Evaluation of the TB-Biochip Oligonucleotide Microarray System for Rapid Detection of Rifampin Resistance in Mycobacterium tuberculosis

Rapid, High-Throughput Detection of Rifampin Resistance and Heteroresistance in Mycobacterium tuberculosis by Use of Sloppy Molecular Beacon Melting Temperature Coding

Advances in Personalised Treatment of Multi-drug Resistant Tuberculosis

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