Additional file 1: of Cost analysis of rapid diagnostics for drug-resistant tuberculosis

Table S1 Clinical and Laboratory Characteristics of the Patients. Table S2 Agreement between three rapid tests and MGIT for detection of resistance for isoniazid (INH), rifampin (RIF), amikacin (AMK), capreomycin (CAP), kanamycin (KAN), moxifloxacin (MOX), and ofloxacin (OFX). Table S3 Proportion of total assay runs that produced interpretable results from three diagnostic platforms (LPA, PSQ and MODS) with the ability to detect resistance to isoniazid (INH), rifampin (RIF), amikacin (AMK), capreomycin (CAP), kanamycin (KAN), moxifloxacin (MOX), and ofloxacin (OFX). (DOCX 35 kb

Drug resistance mutation pattern in a convenience sample of 41 MDR Beijing isolates from the Western Cape.

<p>No data was available for the streptomycin resistance determining region in <i>rrs</i> (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0070919#pone-0070919-t001" target="_blank">Table 1</a>). For more information see figure legend of <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0070919#pone-0070919-g003" target="_blank">Figure 3</a>.</p

Strain population structure of drug-sensitive (DS), mono-/poly-resistant (DR), <i>sensu stricto</i> multidrug-resistant (MDR <i>s.s.</i>; excluding identified pre-XDR and XDR isolates), pre-extensively drug-resistant (pre-XDR) and extensively drug resistant (XDR) isolates in three provinces of South Africa.

<p>The R220, R86 and F15/LAM4/KZN genotypes, respectively, represent a subgroup of the typical Beijing, “atypical” Beijing and LAM4 family <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0070919#pone.0070919-Strauss1" target="_blank">[14]</a>–<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0070919#pone.0070919-Pillay1" target="_blank">[16]</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0070919#pone.0070919-Muller2" target="_blank">[22]</a>–<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0070919#pone.0070919-Gandhi2" target="_blank">[24]</a>. Based on similar IS<i>6110</i> RFLP patterns and whole genome sequencing data it was previously shown that “atypical” Beijing strains in the Western and Eastern Cape, unlike in other parts of the world, represent one single genotype herein referred to as R86 <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0070919#pone.0070919-Chihota1" target="_blank">[23]</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0070919#pone.0070919-Ioerger1" target="_blank">[25]</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0070919#pone.0070919-Klopper1" target="_blank">[27]</a>. The specific presence of R220 and F15/LAM4/KZN genotypes was only assessed in the Western Cape and KwaZulu-Natal, respectively, where these genotypes were known to be frequent among XDR-TB cases <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0070919#pone.0070919-Muller2" target="_blank">[22]</a>.</p

Selection of study population.

<p>Grey boxes indicate sample sets used to analyze the strain population structures in the three South African provinces. Boxes with striped pattern indicate sample sets used to characterize drug resistance mutation patterns among XDR-TB associated genotypes. ^a) Computer-based random sampling was applied. ^b) Review of an extensive collection of data generated within multiple previous studies.</p

Geographical distribution of selected clusters of isolates.

<p>EC: Eastern Cape Province.</p><p>WC: Western Cape Province.</p><p>N_isolate: Number of isolates of a cluster detected in the municipal district indicated.</p><p>%: Proportion of isolates of a cluster detected in the municipal district indicated.</p

Drug resistance-associated genetic regions analyzed.

*<p>Genetic region covered by PCR with respect to nucleotide positions in H37Rv.</p><p>H: Isoniazid.</p><p>Eto: Ethionamide.</p><p>R: Rifampicin.</p><p>E: Ethambutol.</p><p>Z: Pyrazinamid.</p><p>S: Streptomycin.</p><p>Km: Kanamycin.</p><p>Am: Amikacin.</p><p>Cm: Capreomycin.</p><p>FQ: Fluoroquinolone.</p><p>Ofx: Ofloxacin.</p

Flowchart of patient enrollment, clinical sample processing for Line Probe Assay (LPA), Microscopic Observation of Drug Susceptibility (MODS), and Pyrosequencing (PSQ) for seven anti-TB drugs: isoniazid (INH), rifampin (RIF), moxifloxacin (MOX), ofloxacin (OFX), amikacin (AMK), kanamycin (KAN), and capreomycin (CAP).

<p>Includes one null PSQ sample result (denoted as 'a'). R: Resistant, S: Susceptible, and I: Indeterminate.</p

Performance Comparison of Three Rapid Tests for the Diagnosis of Drug-Resistant Tuberculosis

<div><p>Background</p><p>The aim of this study was to compare the performance of several recently developed assays for the detection of multi- and extensively drug-resistant tuberculosis (M/XDR-TB) in a large, multinational field trial.</p><p>Methods</p><p>Samples from 1,128 M/XDR-TB suspects were examined by Line Probe Assay (LPA), Pyrosequencing (PSQ), and Microscopic Observation of Drug Susceptibility (MODS) and compared to the BACTEC MGIT960 reference standard to detect M/XDR-TB directly from patient sputum samples collected at TB clinics in India, Moldova, and South Africa.</p><p>Results</p><p>Specificity for all three assays was excellent: 97–100% for isoniazid (INH), rifampin (RIF), moxifloxacin (MOX) and ofloxacin (OFX) and 99–100% for amikacin (AMK), capreomycin (CAP) and kanamycin (KAN) resistance. Sensitivities were lower, but still very good: 94–100% for INH, RIF, MOX and OFX, and 84–90% for AMK and CAP, but only 48–62% for KAN. In terms of agreement, statistically significant differences were only found for detection of RIF (MODS outperformed PSQ) and KAN (MODS outperformed LPA and PSQ) resistance. Mean time-to-result was 1.1 days for LPA and PSQ, 14.3 days for MODS, and 24.7 days for MGIT.</p><p>Conclusions</p><p>All three rapid assays evaluated provide clinicians with timely detection of resistance to the drugs tested; with molecular results available one day following laboratory receipt of samples. In particular, the very high specificity seen for detection of drug resistance means that clinicians can use the results of these rapid tests to avoid the use of toxic drugs to which the infecting organism is resistant and develop treatment regiments that have a higher likelihood of yielding a successful outcome.</p></div

Model for the evolution of XDR-TB associated strain families.

<p>Model for the evolution of XDR-TB associated strain families.</p

Drug resistance mutation pattern in a random selection of 193 MDR R86 isolates from the Eastern Cape.

<p>Different colours indicate different drug resistance associated genes. The area of the circles is proportional to the number of isolates (indicated in the centre of each circle) harbouring an identical drug resistance mutation for the respective resistance gene as well as all circles connected to the left. Principal branches of the tree were defined by resistance mutations in <i>pncA</i>. Other first-line drug resistance mutations were connected by logical deduction to maximize clustering and were followed by second-line resistance mutations. However, the order of acquisition of resistance mutations may remain debatable in some cases.</p