Analysis of factors associated with MDR.

<p>Analysis of factors associated with MDR.</p

Demographic and clinical characteristics of study population.

<p>Demographic and clinical characteristics of study population.</p

Proportion of Culture Positive and MDR-TB Inpatients by hospital proximity to Church of Scotland Hospital.

CoSHa<p>refers to Church of Scotland Hospital at Tugela Ferry. Adjacent to CoSH corresponds to 8 hospitals in sub-districts neighbouring CoSH. Not adjacent to CoSH corresponds to 10 hospitals at locations not neighbouring CoSH. Culture positive is the number and percentage of sampled individuals with culture positive tuberculosis.</p>MDRb<p>is the number and percentage of sampled individuals with rifampicin and isoniazid resistance including XDR-TB.</p

Anti TB drug resistance profile of TB patients with Drug Susceptibility results.

a<p>11 patients had unknown previous treatment.</p

Microbiology results for two phases of the study for all sampled inpatients.

<p>The symbols N^a and %^b correspond to the total number and percentage of sampled patients with the indicated result. Culture^c is a composite result of MGIT and 7H11 agar. Culture Positive refers to growth of <i>M. tuberculosis</i> on either media, and culture other to negative on both media or no result. MDR^d is resitance to both rifampicin and isoniazid and is inclusive of XDR.</p

Proportion of Symptomatic Inpatients with Culture confirmed Tuberculosis and MDR-TB by hospital and District of KwaZulu-Natal.

<p>Patients were defined as symptomatic if they were coughing on the day of sampling. The diameter of the circles representing hospitals is proportional to the proportion of all culture positive patients with MDR-TB. The shading of the circles shows whether the hospitals were adjacent to Church of Scotland Hospital (CoSH) at Tugela Ferry. The figures in brackets at each hospital indicate the proportion (%) of all sampled patients who were culture positive and the proportion who had confirmed MDR-TB.</p

Diverse strains contribute to drug resistance in KwaZulu-Natal.

<p>(A) Midpoint rooted maximum-likelihood phylogeny of 340 <i>M</i>. <i>tuberculosis</i> isolates. Four of the seven known <i>M</i>. <i>tuberculosis</i> lineages were identified: CAS (<i>Lin1</i>), Beijing (<i>Lin2</i>), EAI (<i>Lin 3</i>), and Euro-American (<i>Lin4</i>). Digital spoligotyping identified 17 unique spoligotypes in the dataset; spoligotypes are shown on this figure if they are represented by three or more strains. Corresponding spoligotypes and phenotypes are reported for all strains in <a href="http://www.plosmedicine.org/article/info:doi/10.1371/journal.pmed.1001880#pmed.1001880.s009" target="_blank">S4 Table</a>. Phenotypic XDR, MDR, poly- and monodrug resistance (labeled “Drug-resistant other”), and pansusceptible strains are indicated by colored tick marks at the tip of each leaf node. (B) Histogram of pairwise SNP distances between strains. The number of pairs within each SNP distance range is plotted. The peaks correspond to the distance between major lineages. The peak at the far left of the figure corresponds to the distance between pairs of strains within a clone.</p

Demographic characteristics of participants and phenotypic drug susceptibility of strains.

<p>* HIV-positive and HIV-negative individuals were compared using Fisher’s exact test for categorical variables and nonparametric Mann-Whitney test for continuous variables.</p><p>^† Data in this row are from the XDR data, not the total dataset.</p><p>Results having a <i>p</i>-value < 0.05 were considered statistically significant.</p><p>Data are <i>n</i> (%) or mean ± standard deviation (SD).</p

Isoniazid resistance is the first step towards drug resistance.

<p>Acquisition of <i>katG</i> S315 mutations precedes all other resistance mutations, including rifampicin, in all instances in which the order of acquisition can be disambiguated. For the 214 strains with genotypic resistance to two or more MDR or XDR defining drugs, and in which the order of acquisition of these mutations could be disambiguated, we quantified the number of evolutions in which resistance to one drug was gained before resistance to a second drug. Isoniazid resistance was divided into mutations conferred by the <i>katG</i>S315 codon versus “Other INH” mutations (defined as loss-of-function mutations in <i>katG</i> that do not involve codon 315 or mutations in the <i>inhA</i> promoter). Reported numbers represent the number of independent evolutionary events (not the number of strains) in which the drug resistance indicated by the row labeled “first resistance” was acquired before the drug resistance indicated by the column labeled “second resistance.” The background color is shaded to indicate the fraction of unambiguous evolutionary events in which the “first resistance” was acquired before the “second resistance” for that given drug pair.</p

Molecular evolution and dating of drug resistance emergence within the Tugela Ferry XDR Clone.

<p>Midpoint rooted maximum-likelihood phylogeny of 107 <i>M</i>. <i>tuberculosis</i> isolates of the LAM4 spoligotype. The gray shaded box identifies the Tugela Ferry XDR Clone. KZN605, the historical XDR strain collected in Tugela Ferry during the outbreak, is a member of this clone. Two additional historical isolates, KZN1435 and KZN4207, are not members of the Tugela Ferry XDR Clone. Each evolutionary gain of a drug resistance mutation was assigned to its position on the phylogenetic tree by parsimony (colored circles). A–E traces the stepwise order of drug resistance acquisition in the Tugela Ferry XDR Clone and estimates the year when each mutation was gained. Gray bars indicate the 95% highest posterior density (HPD) intervals. (A) <i>katG</i> S315T (isoniazid); <i>gidB</i> 130 bp deletion (streptomycin); 1957 (95% HPD: 1937–1971); (B) <i>inhA</i> promoter -8 (isoniazid and ethionamide); 1964 (95% HPD: 1948–1976); (C) <i>embB</i> M306V (ethambutol); 1967 (95% HPD: 1950–1978); (D) <i>rpoB</i> L452P (rifampicin); <i>pncA</i> 1bp insertion (pyrazinamide); 1984 (95% HPD: 1974–1992); and (E) <i>rpoB</i> D435G (rifampicin); <i>rrs</i> 1400 (kanamycin); <i>gyrA</i> A90V (ofloxacin); 1995 (95% HPD: 1988–1999). The accumulation of individual drug-resistant mutations within a strain is denoted to the right of the phylogenetic tree. The dates of drug discovery are displayed at the bottom of the figure [<a href="http://www.plosmedicine.org/article/info:doi/10.1371/journal.pmed.1001880#pmed.1001880.ref053" target="_blank">53</a>]. Four additional LAM4 strains on a distant branch were not included in this figure because of size constraints. Bootstrap values are provided for lettered nodes, and bootstrap values for all nodes are shown in S5 Fig.</p