Clinical Features of Mycobacterium canettii Infection: A Retrospective Study of 20 Cases Among French Soldiers and Relatives.

BACKGROUND: Mycobacterium canettii forms part of the Mycobacterium tuberculosis complex. Mycobacterium canettii infections are mainly described in the Horn of Africa. The permanent presence of French soldiers in Djibouti raises the question of the risk of being infected with M. canettii. Here, we describe M. canettii infections among French military and their families between 1998 and 2015. METHODS: This retrospective study relied on 3 sources of data: the reference center for mycobacteria in the Biology Department at Percy Military Hospital in Paris, the French Military Center for Epidemiology and Public Health, and the scientific literature. After an exhaustive census of the strains, we studied the epidemiological data on 20 cases among French soldiers and their families. RESULTS: Twenty cases of M. canettii infections are reported, including 5 unpublished cases. Adenitis predominates (n = 15), especially in the cervico facial area and among children; 1 case was observed 1 month after dental care in Djibouti. The pulmonary forms were less frequent (n = 6), and 3 atypical forms are described. All patients had stayed in Djibouti. CONCLUSIONS: Cases of M. canettii infection among the French military consisted mainly of adenitis; disseminated forms were possible with immunodeficiency. Their evolution under specific treatments was comparable to that of tuberculosis. The presumed origin of the infection seemed to be environmental, possibly a water reservoir, and not due to human-to-human contagion.

Evolution of smooth tubercle Bacilli PE and PE_PGRS genes: evidence for a prominent role of recombination and imprint of positive selection.

BACKGROUND: PE and PE_PGRS are two mycobateria-restricted multigene families encoding membrane associated and secreted proteins that have expanded mainly in the pathogenic species, notably the Mycobacterium tuberculosis complex (MTBC). Several lines of evidence attribute to PE and PE_PGRS genes critical roles in mycobacterial pathogenicity. To get more insight into the nature of these genes, we sought to address their evolutionary trajectories in the group of smooth tubercle bacilli (STB), the putative ancestor of the clonal MTBC. METHODOLOGY/PRINCIPAL FINDINGS: By focussing on six polymorphic STB PE/PE_PGRS genes, we demonstrate significant incongruence among single gene genealogies and detect strong signals of recombination using various approaches. Coalescent-based estimation of population recombination and mutation rates (ρ and θ, respectively) indicates that the two mechanisms are of roughly equal importance in generating diversity (ρ/θ = 1.457), a finding in a marked contrast to house keeping genes (HKG) whose evolution is chiefly brought about by mutation (ρ/θ = 0.012). In comparison to HKG, we found 15 times higher mean rate of nonsynonymous substitutions, with strong evidence of positive selection acting on PE_PGRS62 (dN/dS = 1.42), a gene that has previously been shown to be essential for mycobacterial survival in macrophages and granulomas. Imprint of positive selection operating on specific amino acid residues or along branches of PE_PGRS62 phylogenetic tree was further demonstrated using maximum likelihood- and covarion-based approaches, respectively. Strikingly, PE_PGR62 proved highly conserved in present-day MTBC strains. CONCLUSIONS/SIGNIFICANCE: Overall the data indicate that, in STB, PE/PE_PGRS genes have undergone a strong diversification process that is speeded up by recombination, with evidence of positive selection. The finding that positive selection involved an essential PE_PGRS gene whose sequence appears to be driven to fixation in present-day MTBC strains lends further support to the critical role of PE/PE_PGRS genes in the evolution of mycobacterial pathogenicity.

Genomic analysis of smooth tubercle bacilli provides insights into ancestry and pathoadaptation of Mycobacterium tuberculosis.

Global spread and limited genetic variation are hallmarks of M. tuberculosis, the agent of human tuberculosis. In contrast, Mycobacterium canettii and related tubercle bacilli that also cause human tuberculosis and exhibit unusual smooth colony morphology are restricted to East Africa. Here, we sequenced and analyzed the whole genomes of five representative strains of smooth tubercle bacilli (STB) using Sanger (4-5× coverage), 454/Roche (13-18× coverage) and/or Illumina DNA sequencing (45-105× coverage). We show that STB isolates are highly recombinogenic and evolutionarily early branching, with larger genome sizes, higher rates of genetic variation, fewer molecular scars and distinct CRISPR-Cas systems relative to M. tuberculosis. Despite the differences, all tuberculosis-causing mycobacteria share a highly conserved core genome. Mouse infection experiments showed that STB strains are less persistent and virulent than M. tuberculosis. We conclude that M. tuberculosis emerged from an ancestral STB-like pool of mycobacteria by gain of persistence and virulence mechanisms, and we provide insights into the molecular events involved.

[Update on Mycobacterium bovis infections in France: 4 cases reports]. / Actualités de l'infection à Mycobacterium bovis en France : à propos de 4 cas.

Mycobacterium bovis (M. bovis) is a cause of zoonosis. It is rare in developed countries since cattle control. We report four cases of M. bovis infection in people aged more 60 years. They were probably infected during infancy, consuming unpasteurized milk. It is the main transmission mode in developing countries where veterinary controls aren't made. M. bovis infections clinical aspects are varied and treatment is complicated by natural pyrazinamide resistance. Recent diagnostic methods using molecular biology are quick and specific and facilitate identification.

Significance of the identification in the Horn of Africa of an exceptionally deep branching Mycobacterium tuberculosis clade.

Molecular and phylogeographic studies have led to the definition within the Mycobacterium tuberculosis complex (MTBC) of a number of geotypes and ecotypes showing a preferential geographic location or host preference. The MTBC is thought to have emerged in Africa, most likely the Horn of Africa, and to have spread worldwide with human migrations. Under this assumption, there is a possibility that unknown deep branching lineages are present in this region. We genotyped by spoligotyping and multiple locus variable number of tandem repeats (VNTR) analysis (MLVA) 435 MTBC isolates recovered from patients. Four hundred and eleven isolates were collected in the Republic of Djibouti over a 12 year period, with the other 24 isolates originating from neighbouring countries. All major M. tuberculosis lineages were identified, with only two M. africanum and one M. bovis isolates. Upon comparison with typing data of worldwide origin we observed that several isolates showed clustering characteristics compatible with new deep branching. Whole genome sequencing (WGS) of seven isolates and comparison with available WGS data from 38 genomes distributed in the different lineages confirms the identification of ancestral nodes for several clades and most importantly of one new lineage, here referred to as lineage 7. Investigation of specific deletions confirms the novelty of this lineage, and analysis of its precise phylogenetic position indicates that the other three superlineages constituting the MTBC emerged independently but within a relatively short timeframe from the Horn of Africa. The availability of such strains compared to the predominant lineages and sharing very ancient ancestry will open new avenues for identifying some of the genetic factors responsible for the success of the modern lineages. Additional deep branching lineages may be readily and efficiently identified by large-scale MLVA screening of isolates from sub-Saharan African countries followed by WGS analysis of a few selected isolates.

Fazer de sua vida uma obra / Faire de sa vie une œuvre / To make of one's life a work of art

A expressão "fazer de sua vida uma obra" tornou-se um slogan que necessita ser questionado. Com efeito, se lembrarmos a distinção aristotélica entre ação e produção, práxis e poiese, não se trataria de uma confusão de categorias? Podemos conceber a vida como um modo de produção? Se for de fato uma produção artística, o que significa então pensar a vida sob uma modalidade estética? Não seríamos levados a explicitar as tensões que não deixam de se manifestar entre estética, ética e moral? Enfim, "fazer uma obra de sua vida" supõe antes de tudo desdobrá-la sob a forma de narrativa para poder avaliar o que nos conduz a explorar - na direção de Paul Ricoeur (1984; 1985) - os dilemas da escrita de si.

L'expression «faire de sa vie une œuvre¼ est devenue un slogan qu'il faut interroger. En effet, si l'on se souvient de la distinction aristotélicienne entre action et production, praxis et poiesis, ne s'agit-il pas d'une confusion de catégories ? Peut-on concevoir la vie sur le mode de la production ? S'il s'agit bien d'une production artistique, que signifie ici penser la vie sous une modalité esthétique ? N'est-on pas emmené à expliciter les tensions qui ne manquent pas de se manifester entre esthétique, éthique et morale ? Enfin, « faire oeuvre de sa vie ¼ suppose d'abord de la déployer sous forme de récit pour pouvoir l'évaluer ce qui nous conduit à explorer - à la suite de Paul Ricoeur (1984, 1985) - les dilemmes de l'écriture de soi.

The phrase "to make of one's life a work of art" has become a slogan. It is time to put it into question. If one goes back to Aristotle's distinction between action and production, praxis and poiesis, then this phrase will sound like a category mix up. Can we think of life in terms of production? And if the production implied is the artistic one, what does it mean to think of life from an aesthetic point of view? Isn't one led to examine the tensions arising between aesthetics ethics and moral? Finally, "to make of one's life a work of art" entails that life should be first made the object of a narrative form in order to be evaluated: this is the reason why we are led - following here Paul Ricoeur (1984-1985)- to examine the dilemmas raised by the writing of the self.

Assessment of the SD Bioline Ag MPT64 Rapid™ and the MGIT™ TBc identification tests for the diagnosis of tuberculosis.

Successful control of tuberculosis relies on the rapid detection of Mycobacterium tuberculosis. Few chromatographic lateral flow assays for the discrimination of the M. tuberculosis complex were developed from culture media. We compared the values of 2 assays to assess their place in diagnosis of tuberculosis. We conclude of their efficiency and relevance to supplant the conventional methods.

Molecular characteristics of "Mycobacterium canettii" the smooth Mycobacterium tuberculosis bacilli.

Since the first discovery of the smooth tubercle (SmTB) bacilli "Mycobacterium canettii" less than 60 isolates have been reported, all but one originating from a limited geographical location, the Horn of Africa. In spite of its rarity, the SmTB lineage deserves special attention. Previous investigations suggested that SmTB isolates represent an ancestral lineage of the Mycobacterium tuberculosis complex (MTBC) and that consequently they might provide essential clues on the origin and evolution of the MTBC. There is evidence that unlike the rest of the MTBC, SmTB strains recombine chromosomal sequences with a yet unknown Mycobacterium species. This behavior contributes to the much larger genetic heterogeneity observed in the SmTB isolates compared to the other members of the MTBC. We have collected 59 SmTB isolates of which 14 were newly recovered since previous reports, and performed extensive phenotypical and genotypical characterization. We take advantage of these investigations to review the current knowledge of "M. canettii". Their characteristics and the apparent lack of human to human transmission are consistent with the previously proposed existence of non-human sources of infection. SmTB strains show remarkably common features together with secondary and taxonomically minor genetic differences such as the presence or absence of the CRISPR (Clustered Regularly Interspersed Palindromic Repeat) locus (usually called Direct Repeat or DR region) or number of IS sequences. Multiple Locus Variable number of tandem repeat Analysis (MLVA) and DR region analyses reveal one predominant clone, one minor clone and a number of more distantly related strains. This suggests that the two most frequent clones may represent successfully emerging lineages.

[Tuberculous meningitis: diagnosis and therapeutic difficulties]. / Méningites tuberculeuses: difficultés diagnostiques et thérapeutiques.

Tuberculosis remains a public-health problem in 2010 with 9 millions cases and 1,7 million deaths worldwide each year. Tuberculosis meningitis is rare (0.5 to 1%) but is associated with high mortality and disability among survivors. An early starting of treatment is crucial. Despite molecular biology methods, microbiological diagnosis remains a challenge for the biologist. We report here 2 cases of tuberculous meningitis with different clinical and biological presentations, which underline diagnosis and therapeutic difficulties encountered in the management of this disease. The first one occurred in an HIV infected patient and the second one was caused by a multidrug-resistant strain. Clinical issues were severe with important neurological residual disability and death. Biological methods available for tuberculous meningitis diagnosis are exposed.

Comparison of two commercial assays for the characterization of rpoB mutations in Mycobacterium tuberculosis and description of new mutations conferring weak resistance to rifampicin.

OBJECTIVES: The aim of this study was to compare the efficiency of two commercial assays, INNO-LiPA Rif.TB and MTBDRplus, for the identification of mutations in the rpoB hot-spot region and to assess the efficiency of these mutations in conferring resistance to rifampicin. METHODS: A collection of 126 rifampicin-resistant Mycobacterium tuberculosis and Mycobacterium africanum isolates and 18 rifampicin-susceptible isolates from different countries were analysed using the two hybridization assays. RESULTS: For 60 strains the hot-spot region of the rpoB gene was sequenced, confirming the results of the hybridization assays and allowing the identification of new mutations. In total, 17 mutations involving 10 codons were observed, two of which are newly described (D516Y and E562G/P564L). Mutations L533P, H526L, D516Y and L511P and the double mutation E562G/P564L conferred a low level of resistance. CONCLUSIONS: The assays INNO-LiPA Rif.TB and MTBDRplus identified rpoB mutations in 98.4% of cases. MTBDRplus provided additional information due to the overlapping hybridization probes and in addition allowed the investigation of katG mutations for isoniazid resistance.

Effect of pO2 on antitumor drug cytotoxicity on MDR and non-MDR variants selected from the LoVo metastatic colon carcinoma cell line.

Two chemosensitive cell lines, LoVo-fusoid (LoVo-f) and LoVo-small cells (LoVo-sc) were derived from the original LoVo cell line. These two variants and the multidrug-resistant (MDR) cell line LoVo-Dox were screened for various properties. In non-permeabilized cells, only LoVo-sc showed mucin-2 staining whereas labelling was positive in all permeabilized cell lines. As shown by electron microscopy screening and by relative resistance to trypsin detachment, only LoVo-sc cells showed strong mucus secretion. All three cell lines displayed strong staining for P-glycoprotein (P-gp), multidrug resistance-associated protein (MRP) and lung-resistance-related protein (LRP) in different locations according to the drug resistance state. The three cell lines showed intracellular labelling of LRP and MRP. The sensitive cells showed P-gp in a large perinuclear ring and in the cytoplasm, but little (LoVo-sc cells) or no staining (LoVo-f cells) was shown at the plasma membrane level. For the Lovo-Dox cells, P-gp was located in the plasma membrane, in cellular anchorages and in the cytoplasm as well. Cell resistance against antineoplastic agents often results from mobilization of various factors, the modulation of which is linked to the culture conditions. As most of the protocols utilize cells growing in (air + 5-10% CO2) atmosphere e.g. 20% O2, balance of the respective participants in the MDR multi-modal mechanism may not be representative of the in vivo situation and may lead to erratic pharmacological response. Indeed, cells within solid tumours were exposed to low pO2, most of them being under hypoxic condition (0.1-5% O2). In the absence of anticancer drugs, all LoVo cell lines grew notably faster at 20% O2 than at 5% O2. Moreover, respective sensitivities of both non-MDR variants to doxorubicin were altered according the pO2. Whatever the pO2 was, virtually none of the antioxidants tested affected the cytotoxic activity of doxorubicin for the three cell lines. By contrast, trolox showed a strong inhibitory effect on doxorubicin activity. These results underline the importance of evaluating the role of hypoxia on the cytotoxic effect of chemotherapeutic agents used either as single drugs or in combination therapy.

Insights into the evolutionary history of tubercle bacilli as disclosed by genetic rearrangements within a PE_PGRS duplicated gene pair.

BACKGROUND: The highly homologous PE_PGRS (Proline-glutamic acid_polymorphic GC-rich repetitive sequence) genes are members of the PE multigene family which is found only in mycobacteria. PE genes are particularly abundant within the genomes of pathogenic mycobacteria where they seem to have expanded as a result of gene duplication events. PE_PGRS genes are characterized by their high GC content and extensive repetitive sequences, making them prone to recombination events and genetic variability. RESULTS: Comparative sequence analysis of Mycobacterium tuberculosis genes PE_PGRS17 (Rv0978c) and PE_PGRS18 (Rv0980c) revealed a striking genetic variation associated with this typical tandem duplicate. In comparison to the M. tuberculosis reference strain H37Rv, the variation (named the 12/40 polymorphism) consists of an in-frame 12-bp insertion invariably accompanied by a set of 40 single nucleotide polymorphisms (SNPs) that occurs either in PE_PGRS17 or in both genes. Sequence analysis of the paralogous genes in a representative set of worldwide distributed tubercle bacilli isolates revealed data which supported previously proposed evolutionary scenarios for the M. tuberculosis complex (MTBC) and confirmed the very ancient origin of "M. canettii" and other smooth tubercle bacilli. Strikingly, the identified polymorphism appears to be coincident with the emergence of the post-bottleneck successful clone from which the MTBC expanded. Furthermore, the findings provide direct and clear evidence for the natural occurrence of gene conversion in mycobacteria, which appears to be restricted to modern M. tuberculosis strains. CONCLUSION: This study provides a new perspective to explore the molecular events that accompanied the evolution, clonal expansion, and recent diversification of tubercle bacilli.

[Diagnosis of Mycobacterium genavense infection by INNO-LiPA MYCOBACTERIA Amplification v2]. / Infections à Mycobacterium genavense diagnostiquées par le test INNO-LiPA MYCOBACTERIA Amplification v2.

INTRODUCTION: Diagnosis of atypical mycobacterial infections can be facilitated by molecular biology techniques. CASES: Two severely immunodepressed patients were admitted for deterioration of their general health status. Bacteriological analysis of histopathologic fragments showed the presence of acid- and alcohol-resistant bacilli. INNO-LiPA MYCOBACTERIA Amplification v2 made it possible to diagnose Mycobacterium genavense infection. COMMENTS: M. genavense is a slow-growing atypical mycobacterium. The molecular biology techniques used by INNO-LiPA MYCOBACTERIA V2 permit its swift identification.

Novel genetic polymorphisms that further delineate the phylogeny of the Mycobacterium tuberculosis complex.

In a previous report, we described a PCR protocol for the differentiation of the various species of the Mycobacterium tuberculosis complex (MTC) on the basis of genomic deletions (R. C. Huard, L. C. de Oliveira Lazzarini, W. R. Butler, D. van Soolingen, and J. L. Ho, J. Clin. Microbiol. 41:1637-1650, 2003). That report also provided a broad cross-comparison of several previously identified, phylogenetically relevant, long-sequence and single-nucleotide polymorphisms (LSPs and SNPs, respectively). In the present companion report, we expand upon the previous work (i) by continuing the evaluation of known MTC phylogenetic markers in a larger collection of tubercle bacilli (n = 125), (ii) by evaluating additional recently reported MTC species-specific and interspecific polymorphisms, and (iii) by describing the identification and distribution of a number of novel LSPs and SNPs. Notably, new genomic deletions were found in various Mycobacterium tuberculosis strains, new species-specific SNPs were identified for "Mycobacterium canettii," Mycobacterium microti, and Mycobacterium pinnipedii, and, for the first time, intraspecific single-nucleotide DNA differences were discovered for the dassie bacillus, the oryx bacillus, and the two Mycobacterium africanum subtype I variants. Surprisingly, coincident polymorphisms linked one M. africanum subtype I genotype with the dassie bacillus and M. microti with M. pinnipedii, thereby suggesting closer evolutionary ties within each pair of species than had been previously thought. Overall, the presented data add to the genetic definitions of several MTC organisms as well as fine-tune current models for the evolutionary history of the MTC.

Study of the gyrB gene polymorphism as a tool to differentiate among Mycobacterium tuberculosis complex subspecies further underlines the older evolutionary age of 'Mycobacterium canettii'.

The present investigation evaluated the PCR-restriction fragment length polymorphism (RFLP) analysis of hsp65 and gyrB targets for differentiation of the species within the Mycobacterium tuberculosis complex (MTC) both by including new restriction enzymes and previously unstudied species. The hsp65 restriction analysis using HhaI resulted in a characteristic 'Mycobacterium canettii' pattern. A study of the gyrB gene polymorphism using TaqIalpha and HinfI allowed the initial division of MTC into two major groups, one consisting of M. tuberculosis and 'M. canettii' as opposed to another single group with other species. Three different patterns were observed with RsaI, the first characteristic of Mycobacterium microti, the second with Mycobacterium bovis, M. bovis BCG and Mycobacterium caprae (M. caprae was easily separated from M. bovis, and M. bovis BCG by SacII digestion), and the third with M. tuberculosis, 'M. canettii', Mycobacterium africanum, Mycobacterium pinnipedii, and the dassie bacillus. Although further discrimination within the last group was not obtained using additional restriction enzymes, the HaeIII and RsaI digestions highlighted an important gyrB polymorphism among 'M. canettii' strains. A study of the single nucleotide polymorphisms (SNP) within the gyrB by sequence analysis not only confirmed the results of the restriction analysis, but showed further differences among 'M. canettii' isolates that were not picked up using the existing battery of restriction enzymes. As many as 11 different SNPs were identified in the collection of eight 'M. canettii' isolates studied. Considering that gyrB variability among MTC member species other than 'M. canettii' is as restricted as hsp65 variability among MTC, our data corroborate a recent proposition that the 'M. canettii' group is evolutionary much older than the other MTC members. In conclusion, gyrB PCR-RFLP is a simple and rapid low-cost method that combined with phenotypic characteristics, may be helpful to differentiate most of the subspecies within the MTC.

Ancient origin and gene mosaicism of the progenitor of Mycobacterium tuberculosis.

The highly successful human pathogen Mycobacterium tuberculosis has an extremely low level of genetic variation, which suggests that the entire population resulted from clonal expansion following an evolutionary bottleneck around 35,000 y ago. Here, we show that this population constitutes just the visible tip of a much broader progenitor species, whose extant representatives are human isolates of tubercle bacilli from East Africa. In these isolates, we detected incongruence among gene phylogenies as well as mosaic gene sequences, whose individual elements are retrieved in classical M. tuberculosis. Therefore, despite its apparent homogeneity, the M. tuberculosis genome appears to be a composite assembly resulting from horizontal gene transfer events predating clonal expansion. The amount of synonymous nucleotide variation in housekeeping genes suggests that tubercle bacilli were contemporaneous with early hominids in East Africa, and have thus been coevolving with their human host much longer than previously thought. These results open novel perspectives for unraveling the molecular bases of M. tuberculosis evolutionary success.

Rapid identification of Mycobacterium genavense with a new commercially available molecular test, INNO-LiPA MYCOBACTERIA v2.

We report a rare mesenteric localized Mycobacterium genavense infection in a severely immunocompromised human immunodeficiency virus-infected patient. An INNO-LiPA MYCOBACTERIA v2 test was performed directly on biopsy samples. This new molecular tool could be used for simultaneous identification of mycobacterium species from human specimens, but other studies are needed to validate our first results.

High genetic diversity revealed by variable-number tandem repeat genotyping and analysis of hsp65 gene polymorphism in a large collection of "Mycobacterium canettii" strains indicates that the M. tuberculosis complex is a recently emerged clone of "M. canettii".

We have analyzed, using complementary molecular methods, the diversity of 43 strains of "Mycobacterium canettii" originating from the Republic of Djibouti, on the Horn of Africa, from 1998 to 2003. Genotyping by multiple-locus variable-number tandem repeat analysis shows that all the strains belong to a single but very distant group when compared to strains of the Mycobacterium tuberculosis complex (MTBC). Thirty-one strains cluster into one large group with little variability and five strains form another group, whereas the other seven are more diverged. In total, 14 genotypes are observed. The DR locus analysis reveals additional variability, some strains being devoid of a direct repeat locus and others having unique spacers. The hsp65 gene polymorphism was investigated by restriction enzyme analysis and sequencing of PCR amplicons. Four new single nucleotide polymorphisms were discovered. One strain was characterized by three nucleotide changes in 441 bp, creating new restriction enzyme polymorphisms. As no sequence variability was found for hsp65 in the whole MTBC, and as a single point mutation separates M. tuberculosis from the closest "M. canettii" strains, this diversity within "M. canettii" subspecies strongly suggests that it is the most probable source species of the MTBC rather than just another branch of the MTBC.

High resolution, on-line identification of strains from the Mycobacterium tuberculosis complex based on tandem repeat typing.

BACKGROUND: Currently available reference methods for the molecular epidemiology of the Mycobacterium tuberculosis complex either lack sensitivity or are still too tedious and slow for routine application. Recently, tandem repeat typing has emerged as a potential alternative. This report contributes to the development of tandem repeat typing for M. tuberculosis by summarising the existing data, developing additional markers, and setting up a freely accessible, fast, and easy to use, internet-based service for strain identification. RESULTS: A collection of 21 VNTRs incorporating 13 previously described loci and 8 newly evaluated markers was used to genotype 90 strains from the M. tuberculosis complex (M. tuberculosis (64 strains), M. bovis (9 strains including 4 BCG representatives), M. africanum (17 strains)). Eighty-four different genotypes are defined. Clustering analysis shows that the M. africanum strains fall into three main groups, one of which is closer to the M. tuberculosis strains, and an other one is closer to the M. bovis strains. The resulting data has been made freely accessible over the internet http://bacterial-genotyping.igmors.u-psud.fr/bnserver to allow direct strain identification queries. CONCLUSIONS: Tandem-repeat typing is a PCR-based assay which may prove to be a powerful complement to the existing epidemiological tools for the M. tuberculosis complex. The number of markers to type depends on the identification precision which is required, so that identification can be achieved quickly at low cost in terms of consumables, technical expertise and equipment.