Stefan Niemann

Publications of Stefan Niemann

• Special Issue on Molecular evolution, epidemiology and pathogenesis of Mycobacterium tuberculosis and other mycobacteria.

  Authors: Igor Mokrousov, Stefan Niemann, Nalin Rastogi

  Infection, genetics and evolution : journal of molecular epidemiology and evolutionary genetics in infectious diseases. 03/2012;

• rplC T460C identified as a dominant mutation in linezolid resistant Mycobacterium tuberculosis strains.

  Authors: Patrick Beckert, Doris Hillemann, Thomas A Kohl, Jörn Kalinowski, Elvira Richter, Stefan Niemann, Silke Feuerriegel

  Antimicrobial agents and chemotherapy. 02/2012;

  The ribosomal L3 protein was identified as novel target in linezolid (LZD) resistant Mycobacterium tuberculosis strains. Next-generation-sequencing confirmed rplC T460C as sole mutation in an LZDThe ribosomal L3 protein was identified as novel target in linezolid (LZD) resistant Mycobacterium tuberculosis strains. Next-generation-sequencing confirmed rplC T460C as sole mutation in an LZD resistant M. tuberculosis H37Rv strain selected in vitro. Sequencing analysis revealed the rplC T460C mutation in eight further LZD resistant isolates (three in vitro selected mutants, five patient isolates including isolates from three different patients developing LZD resistance during treatment), but in none of the susceptible control strains (n=84).

• Online tools for polyphasic analysis of Mycobacterium tuberculosis complex genotyping data: Now and next.

  Authors: Thomas Weniger, Justina Krawczyk, Philip Supply, Dag Harmsen, Stefan Niemann

  Infection, genetics and evolution : journal of molecular epidemiology and evolutionary genetics in infectious diseases. 02/2012;

  Molecular diagnostics and genotyping of pathogens have become indispensable tools in clinical microbiology and disease surveillance. For isolates of the Mycobacterium tuberculosis complex (MTBC,Molecular diagnostics and genotyping of pathogens have become indispensable tools in clinical microbiology and disease surveillance. For isolates of the Mycobacterium tuberculosis complex (MTBC, causative agents of tuberculosis), multilocus variable number tandem repeat analysis (MLVA) targeting mycobacterial interspersed repetitive units (MIRU) has been internationally adopted as the new standard, portable, reproducible, and discriminatory typing method. Here, we review new sets of specialized web based bioinformatics tools that have become available for analyzing MLVA data especially in combination with other, complementary genotyping markers (polyphasic analysis). Currently, there are only two databases available that are not restricted to store one kind of genotyping data only, namely SITVIT/SpolDB4 and MIRU-VNTRplus. SITVIT/SpolDB4 (http://www.pasteur-guadeloupe.fr:8081/SITVITDemo) contains spoligotyping data from a large number of strains of diverse origin. However, besides options to query the data, the actual version of SITVIT/SpolDB4 offers no functionality for more complex analysis e.g. tree-based analysis. In comparison, the MIRU-VNTRplus web application (http://www.miru-vntrplus.org), represents a freely accessible service that enables users to analyze genotyping data of their strains alone or in comparison with a currently limited but well characterized reference database of strains representing the major MTBC lineages. Data (MLVA-, spoligotype-, large sequence polymorphism, and single nucleotide polymorphism) can be visualized and analyzed using just one genotyping method or a weighted combination of several markers. A variety of analysis tools are available such as creation of phylogenetic and minimum spanning trees, semi-automated phylogenetic lineage identification based on comparison with the reference database and mapping of geographic information. To facilitate scientific communication, a universal, expanding genotype nomenclature (MLVA MtbC15-9 type) service that can be queried via a web- or a SOAP-interface has been implemented. An extensive documentation guides users through all application functions. Perspectives for future development, including generalization to other bacterial species, are presented.

• Whole-genome sequencing of rifampicin-resistant Mycobacterium tuberculosis strains identifies compensatory mutations in RNA polymerase genes.

  Authors: Iñaki Comas, Sonia Borrell, Andreas Roetzer, Graham Rose, Bijaya Malla, Midori Kato-Maeda, James Galagan, Stefan Niemann, Sebastien Gagneux

  Nature genetics. 12/2011; 44(1):106-10.

  Epidemics of drug-resistant bacteria emerge worldwide, even as resistant strains frequently have reduced fitness compared to their drug-susceptible counterparts. Data from model systems suggest thatEpidemics of drug-resistant bacteria emerge worldwide, even as resistant strains frequently have reduced fitness compared to their drug-susceptible counterparts. Data from model systems suggest that the fitness cost of antimicrobial resistance can be reduced by compensatory mutations; however, there is limited evidence that compensatory evolution has any significant role in the success of drug-resistant bacteria in human populations. Here we describe a set of compensatory mutations in the RNA polymerase genes of rifampicin-resistant M. tuberculosis, the etiologic agent of human tuberculosis (TB). M. tuberculosis strains harboring these compensatory mutations showed a high competitive fitness in vitro. Moreover, these mutations were associated with high fitness in vivo, as determined by examining their relative clinical frequency across patient populations. Of note, in countries with the world's highest incidence of multidrug-resistant (MDR) TB, more than 30% of MDR clinical isolates had this form of mutation. Our findings support a role for compensatory evolution in the global epidemics of MDR TB.

• Evaluation of Mycobacterium tuberculosis typing methods in a 4-year study in Schleswig-Holstein, Northern Germany.

  Authors: Andreas Roetzer, Sieglinde Schuback, Roland Diel, Frauke Gasau, Tanja Ubben, Alessia di Nauta, Elvira Richter, Sabine Rüsch-Gerdes, Stefan Niemann

  Journal of clinical microbiology. 12/2011; 49(12):4173-8.

  In order to evaluate the discriminatory power of different methods for genotyping of Mycobacterium tuberculosis complex (MTBC) isolates, we compared the performance of (i) IS6110 DNA fingerprintIn order to evaluate the discriminatory power of different methods for genotyping of Mycobacterium tuberculosis complex (MTBC) isolates, we compared the performance of (i) IS6110 DNA fingerprint typing, (ii) spoligotyping, and (iii) 24-loci mycobacterial interspersed repetitive units-variable number of tandem repeats (MIRU-VNTR) typing in a long-term study on the epidemiology of tuberculosis (TB) in Schleswig-Holstein, the northernmost federal state of Germany. In total, we analyzed 277 MTBC isolates collected from patients between the years 2006 and 2010. The collection comprised a broad spectrum of 13 different genotypes, among which strains of the Haarlem genotype (31%) were most prominent, followed by strains belonging to the Delhi and Beijing lineages (7% and 6%, respectively). On the basis of IS6110 restriction fragment length polymorphism (RFLP) and spoligotyping analyses, 211 isolates had unique patterns (76%) and 66 isolates (24%) were in 20 clusters. MIRU-VNTR combined with spoligotyping analyses revealed 202 isolates with unique patterns (73%) and 75 isolates in 18 clusters (27%). Overall, there was 93.1% concordance between the typing results obtained; 198 strains were identified as unique, and 60 isolates were clustered by both typing combinations (including all 31 isolates with confirmed epidemiological links). Of the remaining 19 isolates with discrepant results, 15 were falsely clustered by MIRU-VNTR (six Beijing genotype strains) and four were clustered by IS6110 RFLP (low IS6110 copy number) only. In conclusion, in the study population investigated, a minority of isolates, especially of the Beijing genotype, clustered by standard 24-loci MIRU-VNTR and without an obvious epidemiological link may require second-line typing by IS6110 RFLP or hypervariable MIRU-VNTR loci.

• Impact of Fgd1 and ddn diversity in Mycobacterium tuberculosis complex on in vitro susceptibility to PA-824.

  Authors: Silke Feuerriegel, Claudio U Köser, Davide Baù, Sabine Rüsch-Gerdes, David K Summers, John A C Archer, Marc A Marti-Renom, Stefan Niemann

  Antimicrobial agents and chemotherapy. 09/2011; 55(12):5718-22.

  PA-824 is a promising drug candidate for the treatment of tuberculosis (TB). It is in phase II clinical trials as part of the first newly designed regimen containing multiple novel antituberculosisPA-824 is a promising drug candidate for the treatment of tuberculosis (TB). It is in phase II clinical trials as part of the first newly designed regimen containing multiple novel antituberculosis drugs (PA-824 in combination with moxifloxacin and pyrazinamide). However, given that the genes involved in resistance against PA-824 are not fully conserved in the Mycobacterium tuberculosis complex (MTBC), this regimen might not be equally effective against different MTBC genotypes. To investigate this question, we sequenced two PA-824 resistance genes (fgd1 [Rv0407] and ddn [Rv3547]) in 65 MTBC strains representing major phylogenetic lineages. The MICs of representative strains were determined using the modified proportion method in the Bactec MGIT 960 system. Our analysis revealed single-nucleotide polymorphisms in both genes that were specific either for several genotypes or for individual strains, yet none of these mutations significantly affected the PA-824 MICs (≤ 0.25 μg/ml). These results were supported by in silico modeling of the mutations identified in Fgd1. In contrast, "Mycobacterium canettii" strains displayed a higher MIC of 8 μg/ml. In conclusion, we found a large genetic diversity in PA-824 resistance genes that did not lead to elevated PA-824 MICs. In contrast, M. canettii strains had MICs that were above the plasma concentrations of PA-824 documented so far in clinical trials. As M. canettii is also intrinsically resistant against pyrazinamide, new regimens containing PA-824 and pyrazinamide might not be effective in treating M. canettii infections. This finding has implications for the design of multiple ongoing clinical trials.

• Polymorphisms in isoniazid and prothionamide resistance genes of the Mycobacterium tuberculosis complex.

  Authors: Michaela Projahn, Claudio U Köser, Susanne Homolka, David K Summers, John A C Archer, Stefan Niemann

  Antimicrobial agents and chemotherapy. 06/2011; 55(9):4408-11.

  Sequence analyses of 74 strains that encompassed major phylogenetic lineages of the Mycobacterium tuberculosis complex revealed 10 polymorphisms in mshA (Rv0486) and four polymorphisms in inhASequence analyses of 74 strains that encompassed major phylogenetic lineages of the Mycobacterium tuberculosis complex revealed 10 polymorphisms in mshA (Rv0486) and four polymorphisms in inhA (Rv1484) that were not responsible for isoniazid or prothionamide resistance. Instead, some of these mutations were phylogenetically informative. This genetic diversity must be taken into consideration for drug development and for the design of molecular tests for drug resistance.

• Overview of errors in the reference sequence and annotation of Mycobacterium tuberculosis H37Rv, and variation amongst its isolates.

  Authors: Claudio U Köser, Stefan Niemann, David K Summers, John A C Archer

  Infection, genetics and evolution : journal of molecular epidemiology and evolutionary genetics in infectious diseases. 06/2011;

  Since its publication in 1998, the genome sequence of the Mycobacterium tuberculosis H37Rv laboratory strain has acted as the cornerstone for the study of tuberculosis. In this review we address someSince its publication in 1998, the genome sequence of the Mycobacterium tuberculosis H37Rv laboratory strain has acted as the cornerstone for the study of tuberculosis. In this review we address some of the practical aspects that have come to light relating to the use of H37Rv throughout the past decade which are of relevance for the ongoing genomic and laboratory studies of this pathogen. These include errors in the genome reference sequence and its annotation, as well as the recently detected variation amongst isolates of H37Rv from different laboratories.

• European 1: a globally important clonal complex of Mycobacterium bovis.

  Authors: Noel H Smith, Stefan Berg, James Dale, Adrian Allen, Sabrina Rodriguez, Beatriz Romero, Filipa Matos, Solomon Ghebremichael, Claudine Karoui, Chiara Donati [......] Alicia Aranaz, Stephen V Gordon, Bo-Young Jeon, Gunilla Källenius, Stefan Niemann, M Beatrice Boniotti, Paul D van Helden, Beth Harris, Martín José Zumárraga, Kristin Kremer

  Infection, genetics and evolution : journal of molecular epidemiology and evolutionary genetics in infectious diseases. 05/2011; 11(6):1340-51.

  We have identified a globally important clonal complex of Mycobacterium bovis by deletion analysis of over one thousand strains from over 30 countries. We initially show that over 99% of the strainsWe have identified a globally important clonal complex of Mycobacterium bovis by deletion analysis of over one thousand strains from over 30 countries. We initially show that over 99% of the strains of M. bovis, the cause of bovine tuberculosis, isolated from cattle in the Republic of Ireland and the UK are closely related and are members of a single clonal complex marked by the deletion of chromosomal region RDEu1 and we named this clonal complex European 1 (Eu1). Eu1 strains were present at less than 14% of French, Portuguese and Spanish isolates of M. bovis but are rare in other mainland European countries and Iran. However, strains of the Eu1 clonal complex were found at high frequency in former trading partners of the UK (USA, South Africa, New Zealand, Australia and Canada). The Americas, with the exception of Brazil, are dominated by the Eu1 clonal complex which was at high frequency in Argentina, Chile, Ecuador and Mexico as well as North America. Eu1 was rare or absent in the African countries surveyed except South Africa. A small sample of strains from Taiwan were non-Eu1 but, surprisingly, isolates from Korea and Kazakhstan were members of the Eu1 clonal complex. The simplest explanation for much of the current distribution of the Eu1 clonal complex is that it was spread in infected cattle, such as Herefords, from the UK to former trading partners, although there is evidence of secondary dispersion since. This is the first identification of a globally dispersed clonal complex M. bovis and indicates that much of the current global distribution of this important veterinary pathogen has resulted from relatively recent International trade in cattle.

• On the mutation rates of spoligotypes and variable numbers of tandem repeat loci of Mycobacterium tuberculosis: continued-When tuning matters.

  Authors: Philip Supply, Stefan Niemann, Thierry Wirth

  Infection, genetics and evolution : journal of molecular epidemiology and evolutionary genetics in infectious diseases. 03/2011;

• Mycobacterium tuberculosis embB codon 306 mutations confer moderately increased resistance to ethambutol in vitro and in vivo.

  Authors: Claudia Plinke, Kerstin Walter, Sahar Aly, Stefan Ehlers, Stefan Niemann

  Antimicrobial agents and chemotherapy. 03/2011; 55(6):2891-6.

  Ethambutol (EMB) is a major component of the first-line therapy of tuberculosis. Mutations in codon 306 of embB (embB306) were suggested as a major resistance mechanism in clinical isolates. ToEthambutol (EMB) is a major component of the first-line therapy of tuberculosis. Mutations in codon 306 of embB (embB306) were suggested as a major resistance mechanism in clinical isolates. To directly analyze the impact of individual embB306 mutations on EMB resistance, we used allelic exchange experiments to generate embB306 mutants of M. tuberculosis H37Rv. The level of EMB resistance conferred by particular mutations was measured in vitro and in vivo after EMB therapy by daily gavage in a mouse model of aerogenic tuberculosis. The wild-type embB306 ATG codon was replaced by embB306 ATC, ATA, or GTG, respectively. All of the obtained embB306 mutants exhibited a 2- to 4-fold increase in EMB MIC compared to the wild-type H37Rv. In vivo, the one selected embB306 GTG mutant required a higher dose of ethambutol to restrict its growth in the lung compared to wild-type H37Rv. These experiments demonstrate that embB306 point mutations enhance the EMB MIC in vitro to a moderate, but significant extent, and reduce the efficacy of EMB treatment in the animal model. We propose that conventional EMB susceptibility testing, in combination with embB306 genotyping, may guide dose adjustment to avoid clinical treatment failure in these low-level resistant strains.

• [The validity of Mycobacterium tuberculosis complex species identification. Comments on the article: "Differentiation of species within the Mycobacterium tuberculosis complex by molecular techniques"].

  Authors: Claudio U Köser, Silvia Piñero-Fernández, Stefan Niemann, David K Summers

  Enfermedades infecciosas y microbiología clínica. 02/2011; 29(4):320.

• Negative and Positive Predictive Value of a Whole-Blood Interferon-{gamma} Release Assay for Developing Active Tuberculosis: An Update.

  Authors: Roland Diel, Robert Loddenkemper, Stefan Niemann, Karen Meywald-Walter, Albert Nienhaus

  American journal of respiratory and critical care medicine. 01/2011; 183(1):88-95.

  only limited data are available on the predictive value of interferon-γ release assays for progression from latent tuberculosis infection to active tuberculosis (TB). to build on our initial studyonly limited data are available on the predictive value of interferon-γ release assays for progression from latent tuberculosis infection to active tuberculosis (TB). to build on our initial study comparing the QuantiFERON-TB Gold in-tube assay (QFT) with the tuberculin skin test (TST) in close contacts of patients with TB and evaluating progression to active TB for up to 4 years. a cohort of close contacts of smear-positive index cases established between May 2005 and April 2008 was tested with QFT and TST. Through April 2010, progressors to active TB were consecutively recorded. of the 1,414 contacts (141 children), 1,033 were still resident in Hamburg at the end of the study period, and results of both tests were available for 954. QFT, but not TST, results were associated with exposure time (P < 0.0001). For QFT, 198 of 954 (20.8%) were positive; 63.3% (604) were TST positive at greater than 5 mm and 25.4% at greater than 10 mm. Nine hundred and three contacts refused chemoprevention and 19 developed active TB. All 19 (100%) had been QFT positive with a progression rate of 12.9% (19 of 147) over the observation period. Corresponding values for the TST were significantly lower: 89.5% (17 of 19) and 3.1% (17 of 555) at greater than 5 mm, and 52.6% (10 of 19) and 4.8% (10 of 207) at greater than 10 mm, respectively. The progression rate of 28.6% (6 of 21) for QFT-positive children was significantly higher than 10.3% (13 of 126) for adults (P = 0.03). results suggest that QFT is more reliable than the TST for identifying those who will soon progress to active TB, especially in children.

• Variant G57E of mannose binding lectin associated with protection against tuberculosis caused by Mycobacterium africanum but not by M. tuberculosis.

  Authors: Thorsten Thye, Stefan Niemann, Kerstin Walter, Susanne Homolka, Christopher D Intemann, Margaret Amanua Chinbuah, Anthony Enimil, John Gyapong, Ivy Osei, Ellis Owusu-Dabo, Sabine Rüsch-Gerdes, Rolf D Horstmann, Stefan Ehlers, Christian G Meyer

  PloS one. 01/2011; 6(6):e20908.

  Structural variants of the Mannose Binding Lectin (MBL) cause quantitative and qualitative functional deficiencies, which are associated with various patterns of susceptibility to infectious diseasesStructural variants of the Mannose Binding Lectin (MBL) cause quantitative and qualitative functional deficiencies, which are associated with various patterns of susceptibility to infectious diseases and other disorders. We determined genetic MBL variants in 2010 Ghanaian patients with pulmonary tuberculosis (TB) and 2346 controls and characterized the mycobacterial isolates of the patients. Assuming a recessive mode of inheritance, we found a protective association between TB and the MBL2 G57E variant (odds ratio 0.60, confidence interval 0.4-0.9, P 0.008) and the corresponding LYQC haplotype (P(corrected) 0.007) which applied, however, only to TB caused by M. africanum but not to TB caused by M. tuberculosis. In vitro, M. africanum isolates bound recombinant human MBL more efficiently than did isolates of M. tuberculosis. We conclude that MBL binding may facilitate the uptake of M. africanum by macrophages, thereby promoting infection and that selection by TB may have favoured the spread of functional MBL deficiencies in regions endemic for M. africanum.

• Treatment of tuberculosis in a region with high drug resistance: outcomes, drug resistance amplification and re-infection.

  Authors: Maryline Bonnet, Manuela Pardini, Francesca Meacci, Germano Orrù, Hasan Yesilkaya, Thierry Jarosz, Peter W Andrew, Mike Barer, Francesco Checchi, Heinz Rinder, Graziella Orefici, Sabine Rüsch-Gerdes, Lanfranco Fattorini, Marco Rinaldo Oggioni, Juliet Melzer, Stefan Niemann, Francis Varaine

  PloS one. 01/2011; 6(8):e23081.

  Emerging antituberculosis drug resistance is a serious threat for tuberculosis (TB) control, especially in Eastern European countries. We combined drug susceptibility results and molecular strainEmerging antituberculosis drug resistance is a serious threat for tuberculosis (TB) control, especially in Eastern European countries. We combined drug susceptibility results and molecular strain typing data with treatment outcome reports to assess the influence of drug resistance on TB treatment outcomes in a prospective cohort of patients from Abkhazia (Georgia). Patients received individualized treatment regimens based on drug susceptibility testing (DST) results. Definitions for antituberculosis drug resistance and treatment outcomes were in line with current WHO recommendations. First and second line DST, and molecular typing were performed in a supranational laboratory for Mycobacterium tuberculosis (MTB) strains from consecutive sputum smear-positive TB patients at baseline and during treatment. At baseline, MTB strains were fully drug-susceptible in 189/326 (58.0%) of patients. Resistance to at least H or R (PDR-TB) and multidrug-resistance (MDR-TB) were found in 69/326 (21.2%) and 68/326 (20.9%) of strains, respectively. Three MDR-TB strains were also extensively resistant (XDR-TB). During treatment, 3/189 (1.6%) fully susceptible patients at baseline were re-infected with a MDR-TB strain and 2/58 (3.4%) PDR-TB patients became MDR-TB due to resistance amplification. 5/47 (10.6%) MDR- patients became XDR-TB during treatment. Treatment success was observed in 161/189 (85.2%), 54/69 (78.3%) and 22/68 (32.3%) of patients with fully drug susceptible, PDR- and MDR-TB, respectively. Development of ofloxacin resistance was significantly associated with a negative treatment outcome. In Abkhazia, a region with high prevalence of drug resistant TB, the use of individualized MDR-TB treatment regimens resulted in poor treatment outcomes and XDR-TB amplification. Nosocomial transmission of MDR-TB emphasizes the importance of infection control in hospitals.

• On the mutation rates of spoligotypes and variable numbers of tandem repeat loci of Mycobacterium tuberculosis.

  Authors: Philip Supply, Stefan Niemann, Thierry Wirth

  Infection, genetics and evolution : journal of molecular epidemiology and evolutionary genetics in infectious diseases. 12/2010; 11(2):251-2.

  Reasonable estimation of mutation rates of molecular markers has implications for molecular epidemiology and evolutionary studies. A recent report (Reyes and Tanaka, 2010) suggested mutation rateReasonable estimation of mutation rates of molecular markers has implications for molecular epidemiology and evolutionary studies. A recent report (Reyes and Tanaka, 2010) suggested mutation rate estimates for variable number of tandem repeat (VNTR) loci in Mycobacterium tuberculosis that are at least an order of magnitude higher than those previously reported (Grant et al., 2008; Wirth et al., 2008). We believe that this deviation can be explained by methodological and analytical problems, including the use of a IS6110 RFLP mutation rate reference that is too indeterminate for subsequently estimating mutation rates of VNTR markers, the utilisation of an infinite allele model unsuitable for the latter markers, and the inclusion of some hypervariable VNTR loci that were excluded from the previous comparison reports. Moreover, the ensuing conclusion on potential (general) underestimation of the extent of recent transmission when using VNTR markers (i.e. because they would mutate so rapidly) conflicts with the large amount of published data demonstrating only rare changes of standard VNTR loci among isolates from clonal transmission or infection.

• Unequal distribution of resistance-conferring mutations among Mycobacterium tuberculosis and Mycobacterium africanum strains from Ghana.

  Authors: Susanne Homolka, Christian G Meyer, Doris Hillemann, Ellis Owusu-Dabo, Ohene Adjei, Rolf D Horstmann, Edmund N L Browne, Amanua Chinbuah, Ivy Osei, John Gyapong, Tanja Kubica, Sabine Ruesch-Gerdes, Stefan Niemann

  International journal of medical microbiology : IJMM. 11/2010; 300(7):489-95.

  Isoniazid (INH) and rifampicin (RMP) resistance in Mycobacterium tuberculosis complex (MTC) isolates are mainly based on mutations in a limited number of genes. However, mutation frequencies vary inIsoniazid (INH) and rifampicin (RMP) resistance in Mycobacterium tuberculosis complex (MTC) isolates are mainly based on mutations in a limited number of genes. However, mutation frequencies vary in different mycobacterial populations. In this work, we analyzed the distribution of resistance-associated mutations in M. tuberculosis and M. africanum strains from Ghana, West Africa. The distribution of mutations in katG, fabG1-inhA, ahpC, and rpoB was determined by DNA sequencing in 217 INH-resistant (INH(r)) and 45 multidrug-resistant (MDR) MTC strains isolated in Ghana from 2001 to 2004. A total of 247 out of 262 strains investigated (94.3%) carried a mutation in katG (72.5%), fabG1-inhA (25.1%), or ahpC (6.5%), respectively. M. tuberculosis strains mainly had katG 315 mutations (80.1%), whereas this proportion was significantly lower in M. africanum West-African 1 (WA1) strains (43.1%; p<0.05). In contrast, WA1 strains showed more mutations in the fabG1-inhA region (39.2%, p<0.05) compared to M. tuberculosis strains (20.9%). In 44 of 45 MDR strains (97.8%) mutations in the 81-bp core region of the rpoB gene could be verified. Additionally, DNA sequencing revealed that 5 RMP-susceptible strains also showed mutations in the rpoB hotspot region. In conclusion, although principally the same genes were affected in INH(r)M. tuberculosis and M. africanum strains, disequilibrium in the distribution of mutations conferring resistance was verified that might influence the efficiency of molecular tests for determination of resistance.

• Thr202Ala in thyA is a marker for the Latin American Mediterranean lineage of the Mycobacterium tuberculosis complex rather than para-aminosalicylic acid resistance.

  Authors: Silke Feuerriegel, Claudio Köser, Leona Trübe, John Archer, Sabine Rüsch Gerdes, Elvira Richter, Stefan Niemann

  Antimicrobial agents and chemotherapy. 11/2010; 54(11):4794-8.

  Single nucleotide polymorphisms (SNPs) involved in the development of resistance represent powerful markers for the rapid detection of first- and second-line resistance in clinical MycobacteriumSingle nucleotide polymorphisms (SNPs) involved in the development of resistance represent powerful markers for the rapid detection of first- and second-line resistance in clinical Mycobacterium tuberculosis complex (MTBC) isolates. However, the association between particular mutations and phenotypic resistance is not always clear-cut, and phylogenetic SNPs have been misclassified as resistance markers in the past. In the present study, we investigated the utility of a specific polymorphism in thyA (Thr202Ala) as a marker for resistance to para-aminosalicyclic acid (PAS). Sixty-three PAS-susceptible MTBC strains comprising all major phylogenetic lineages, reference strain H37Rv, and 135 multidrug-resistant (MDR) strains from Germany (comprising 8 PAS-resistant isolates) were investigated for the presence of Thr202Ala. In both strain collections, the Thr202Ala SNP was found exclusively in strains of the Latin American Mediterranean (LAM) lineage irrespective of PAS resistance. Furthermore, PAS MICs (0.5 mg/liter) for selected LAM strains (all containing the SNP) and non-LAM strains (not containing the SNP), as well as the results of growth curve analyses performed in liquid 7H9 medium in the presence of increasing PAS concentrations (0 to 2.0 mg/liter), were identical. In conclusion, our data demonstrate that the Thr202Ala polymorphism in thyA is not a valid marker for PAS resistance but, instead, represents a phylogenetic marker for the LAM lineage of the M. tuberculosis complex. These findings challenge some of the previous understanding of PAS resistance and, as a consequence, warrant further in-depth investigations of the genetic variation in PAS-resistant clinical isolates and spontaneous mutants.

• Mycobacterium tuberculosis Beijing lineage favors the spread of multidrug-resistant tuberculosis in the Republic of Georgia.

  Authors: Stefan Niemann, Roland Diel, George Khechinashvili, Medea Gegia, Nino Mdivani, Yi-Wei Tang

  Journal of clinical microbiology. 10/2010; 48(10):3544-50.

  High rates and transmission of multidrug-resistant (MDR) tuberculosis (TB) have been associated with the Mycobacterium tuberculosis complex (MTBC) Beijing lineage, pointing to the importance ofHigh rates and transmission of multidrug-resistant (MDR) tuberculosis (TB) have been associated with the Mycobacterium tuberculosis complex (MTBC) Beijing lineage, pointing to the importance of pathogen genetic factors for the modulation of infection outcome and epidemiology. We present here an in-depth analysis of the population structure of MTBC strains from the Republic of Georgia, a high-incidence setting at the Black Sea Coast. Phylogenetic lineages were identified based on 24-locus MIRU-VNTR (for mycobacterial interspersed repetitive unit-variable number tandem repeat) and spoligotyping analysis. Clusters of strains with identical genotyping profiles were determined as an indicator for the rate of recent transmission. Among the 183 M. tuberculosis isolates investigated, the most prominent lineage found was Beijing (26%), followed by the LAM (18%), Ural (12%), and Haarlem (5%) strains. A closely related previously undefined phylogenetic group (62 strains) showed a genotyping pattern similar to laboratory strain H37RV and was denominated as "Georgia-H37RV-like." Although isoniazid resistance was found among strains of different lineages, MDR TB was nearly completely restricted to Beijing strains (P < 0.0001). Approximately 50% of the isolates were grouped in clusters, indicating a high rate of recent transmission. Our data indicate that, in addition to the confirmation of the importance of Beijing genotype strains for the TB epidemiology in former Soviet Union countries, a high-population diversity with strains of the LAM, Ural, Haarlem, and a previously undefined lineage represents nearly two-thirds of the strains found in Georgia. Higher rates among previously treated and MDR TB patients point to a higher potential of lineage Beijing to escape therapy and develop MDR TB.

• Mycobacterium tuberculosis wears what it eats.

  Authors: David G Russell, Brian C VanderVen, Wonsik Lee, Robert B Abramovitch, Mi-jeong Kim, Susanne Homolka, Stefan Niemann, Kyle H Rohde

  Cell host & microbe. 07/2010; 8(1):68-76.

  Mycobacterium tuberculosis remains one of the most pernicious of human pathogens. Current vaccines are ineffective, and drugs, although efficacious, require prolonged treatment with constant medicalMycobacterium tuberculosis remains one of the most pernicious of human pathogens. Current vaccines are ineffective, and drugs, although efficacious, require prolonged treatment with constant medical oversight. Overcoming these problems requires a greater appreciation of M. tuberculosis in the context of its host. Upon infection of either macrophages in culture or animal models, the bacterium realigns its metabolism in response to the new environments it encounters. Understanding these environments, and the stresses that they place on M. tuberculosis, should provide insights invaluable for the development of new chemo- and immunotherapeutic strategies.