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Framework programme: 5
Project number:
EC contribution: € 1 050 000
Duration: 36 months
Type: RS
Starting date: 1 December 2002
Graphic element Characterisation of Mycobacterium Tuberculosis Populations During Infection: A  Longitudinal Study on Drug Resistance Development
Keywords: Tuberculosis; resistance; multidrug resistance; molecular typing; MDR-TB; SNP; single nucleotide polymorphisms


Drug resistance arises in Mycobacterium tuberculosis (MTB) generally by mutation of chromosomal genes, but despite combined drug therapy, the spread of multidrug-resistant (MDR) strains is increasing alarmingly. The LONG-DRUG project is a longitudinal study, carried out in an area with high MDR-TB prevalence, which aims to document the generation of multidrug resistant MTB strains through the use of novel molecular techniques. The combined use of a variety of techniques on longitudinally obtained samples and primary cultures will enable clinically significant knowledge to be generated on the evolution of multidrug resistance in Mycobacterium.

MTB positive clinical samples will be collected in the central TB Hospital of Abkhazia (former Soviet Union), an area of increased MDR-TB prevalence, by Médecins Sans Frontières (MSF). An already ongoing medical aid programme of MSF is a guarantee for longitudinal sample collection. After sample shipment and routine microbiological examination, the minimal inhibitory concentration to primary and secondary anti-tuberculosis drugs will be determined. The determination of the resistance phenotype will target the qualitative molecular analysis to resistance-related genes. Real time PCR and RFLP-PCR will be used to screen for heterogeneous MTB populations. Quantitative real time PCR using molecular beacons on sequenced single nucleotide polymorphisms will finally elucidate prevalence of single resistant clones in bacterial populations. The genetic relationship of all significant strains will be analysed by IS-RFLP and spoligotyping. Centralised data management and correlation of clinical, bacteriological and molecular data should enable elucidation of the contribution of resistant subpopulations to the development of MDR-TB and thus the clinical significance of drug-resistant TB clones in mixed populations.


High prevalence of drug resistance in TB in various geographical settings suggests that MDR-TB may become a more significant problem in the future, even for industrialised countries. The fact that no significant novel first line anti-TB drugs have been developed over the last 40 years does contribute to the significance of the problem.


The first aim of the LONG-DRUG consortium is the generation of a collection of longitudinally collected samples from patients with MDR-TB. Diverse molecular tools will be generated and validated for an in-depth characterisation of these samples. The primary objectives are;

1) the molecular characterisation of multiple drug resistance development in MTB populations over time
2) the clarification of the epidemiological relationship of the strains and sub-clones identified in the study population
3) the elucidation of the generation of resistant sub-populations of MTB are of clinical relevance.

Expected results:

Generate knowledge on (i) the resistance development in a bacterial population infecting the human host over time, (ii) the fate of the different resistant subpopulations generated during this process, and (iii) the clinical relevance of these microbial populations.

Achieved results:

Over 300 samples have been collected to which the minimal inhibitory concentration MIC for the classical first line anti TB drugs is being determined. About 200 of these are resistant to at least one drug, and the determination of the MIC for second line anti TB drugs is in progress. The collection of samples already contains an extensive follow-up from over 15 MDR-TB patients.

Potential applications:

Novel molecular tools will be prepared and validated during the project and will be available to diagnostic laboratories promptly after publication. It is possible that the careful analysis of drug resistance development in MTB will develop novel recommendations for use of molecular technologies in drug resistance detection. The partners also envisage that the molecular tools being developed, exploiting the SNPs generated by the selective pressure of drugs, may enable the consortium to develop more general considerations on the characterisation of the progression and outcome of human pulmonary TB.


Marco R. Oggioni
LAMMB, Dept. Biol Mol.
Università di Siena
LAMMB, Policlinico Le Scotte 1S
Viale Bracci
53100 Siena
Tel: +39 0577 233101
Fax: +39 0577 233334


Official Address Other Information
2Francesco ChecchiEPICENTRE
Rue Saint Sabin 8
FR-75011 Paris
Tel: +33 1 40 21 29 04
Fax: +33 1 40 21 28 03
3Graziella OreficiLaboratory of Bacteriology and Medical Mycology
Istituto Superiore di Sanità
Viale Regina Elena 299
IT-00161 Rome
Tel: +39 06 4990 2333
Fax: +39 06 4938 7112
4Peter AndrewDepartment of Microbiology and Immunology
University of Leicester
PO Box 138
University Rd
UK-LE1 9HN Leicester
United Kingdom
Tel: +44 116 252 2941
Fax: +44 116 252 5030
Website: Kingdom/
5Heinz RinderDepartment of Infectious Diseases and Tropical Medicine
Universität München
Leopoldstr. 5
DE-80802 München
Tel: +49 89 2180 3618
Fax: +49 89 336112
6Sabine Rüsch-GerdesNational Reference Centre for Mycobacteria
Research Centre Borstel
Parkallee 18
DE-23845 Borstel
Tel: +49 4537 188213
Fax: +49 4537 188311
7Germano OrrùDip. Scienze Odontostomatologiche
Università di Cagliari
Via Binaghi 4
IT-9121 Cagliari
Tel: +39 070 537420
8Thierry Jarosz3Es (Essai clinique Evaluation Epidemiologie Statistiques)
49 rue A Lancon
FR-75013 Paris
Tel: +33 1 53 80 34 95
9Francis VaraineMédecins Sans Frontières
Rue Saint Sabin 8
FR-75011 Paris
Tel: +33 1 40 21 29 35

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