Characterization of Clonal Complexity in Tuberculosis by Mycobacterial Interspersed Repetitive Unit-Variable-Number Tandem Repeat Typing

Viedma, Darío García de; Alonso-Rodriguez, Noelia; Andrés, Sandra; Serrano, Manuel Nistal Martín; Bouza, Emilio

doi:10.1128/jcm.43.11.5660-5664.2005

Cited by 50 publications

(38 citation statements)

References 21 publications

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“…limit of mixed templates for our PCR assays was estimated to be 1:26 to 1:40, compared with a reported 1:99 detection limit for MIRU-VNTR typing (11).…”

Section: Discussionmentioning

confidence: 67%

“…However, it is possible that, had we the means to investigate all samples in this study by also using MIRU-VNTR typing, we may have identified more mixed infections. Of the 3 most commonly used techniques for genotyping M. tuberculosis isolates (IS6110-RFLP, spoligotyping, and MIRU-VNTR typing), it is MIRU-VNTR typing that has the greatest capacity to detect the presence of multiple strains, as has been previously demonstrated (11,(22)(23)(24).…”

Section: Discussionmentioning

confidence: 99%

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Molecular Detection of Mixed Infections of Mycobacterium tuberculosis Strains in Sputum Samples from Patients in Karonga District, Malawi

Mallard¹,

McNerney²,

Crampin

et al. 2010

J Clin Microbiol

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The occurrence of mixed infections of Mycobacterium tuberculosis is no longer disputed. However, their frequency, and the impact they may have on our understanding of tuberculosis (TB) pathogenesis and epidemiology, remains undetermined. Most previous studies of frequency applied genotyping techniques to cultured M. tuberculosis isolates and found mixed infections to be rare. PCR-based techniques may be more sensitive for detecting multiple M. tuberculosis strains and can be applied to sputum. To date, one study in South Africa has used a PCR approach and suggested that mixed infection could be common. We investigated mixed infections in northern Malawi using two lineage-specific PCR assays targeting the Latin AmericanMediterranean (LAM) and Beijing lineages. Compared with spoligotyping, the specificity and sensitivity of both assays was 100%. From 160 culture-positive sputa, mixed LAM and non-LAM strains were detected in 4 sputa belonging to 2 (2.8%) patients. Both patients were HIV positive, with no history of TB. Cultured isolates from both patients showed only LAM by PCR and spoligotyping. In a set of 377 cultured isolates, 4 were mixed LAM and non-LAM. Only one showed evidence of more than one M. tuberculosis strain using IS6110-based restriction fragment length polymorphism (IS6110-RFLP) and spoligotyping analyses. Corresponding sputa for the 4 isolates were unavailable. Mixed Beijing and non-Beijing strains were not detected in this study. Mixed infections appear to be rare in our setting and are unlikely to affect findings based on DNA fingerprinting data. Molecular methods, which avoid the selective nature of culture and target distinct strains, are well suited to detection of mixed infections.It used to be thought that tuberculosis in the human host was caused by infection with a single Mycobacterium tuberculosis strain (25). This belief was first challenged when phage typing identified the presence of more than one M. tuberculosis strain in isolates obtained from a single patient (2,17). With the development of DNA fingerprinting (28) and other PCR-based genotyping methods (15, 27) to differentiate M. tuberculosis strains, the presence of multiple strains within a single patient has been demonstrated during routine genotyping studies (5,19,20,24,33). The results from these and further studies designed to specifically examine mixed infections (1,10,20,22,23,29,32) have shown that they occur in different geographical settings among both HIV-negative and HIV-positive tuberculosis (TB) patients.While the occurrence of mixed infections of M. tuberculosis is no longer questioned, the rate at which they occur is unknown. Yet this is of fundamental importance, as much of our understanding of tuberculosis pathogenesis and epidemiology has assumed that mixed infection is rare. The frequency is likely to differ according to the level of M. tuberculosis transmission in different settings. If mixed infections are common, it would necessitate reinterpretation of studies on recent infection, the importance of reinfe...

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“…limit of mixed templates for our PCR assays was estimated to be 1:26 to 1:40, compared with a reported 1:99 detection limit for MIRU-VNTR typing (11).…”

Section: Discussionmentioning

confidence: 67%

Section: Discussionmentioning

confidence: 99%

Molecular Detection of Mixed Infections of Mycobacterium tuberculosis Strains in Sputum Samples from Patients in Karonga District, Malawi

Mallard¹,

McNerney²,

Crampin

et al. 2010

J Clin Microbiol

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“…However, we cannot completely rule out the possibility of undetected polyclonal infection, since we did not genotype more than one isolate per episode in all cases. It is also possible that mixed infection remained undetected by MIRU; however, such a scenario seems improbable, since the percentage of one of the strains would have to be lower than 1% (9). Despite these limitations, to the best of our knowledge, this is the first study to experimentally address the hypothesis of possible misassignment of reinfections due to undetected polyclonal infection.…”

Section: Discussionmentioning

confidence: 95%

“…The new PCR-based technique MIRU-VNTR has proven to be a faster, simpler, and more efficient approach for the identification of clonal complexity (9). Combination of MIRU-VNTR with analysis of several serial isolates per patient has been recommended to improve detection of clonal heterogeneity and to minimize underestimation of clonally complex infections (19).…”

Section: Discussionmentioning

confidence: 99%

Evaluation of the Inaccurate Assignment of Mixed Infections by Mycobacterium tuberculosis as Exogenous Reinfection and Analysis of the Potential Role of Bacterial Factors in Reinfection

et al. 2011

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Molecular analysis of recurrent tuberculosis has revealed that a second episode may be caused by a strain of Mycobacterium tuberculosis other than that involved in the first infection, thus indicating that exogenous reinfection plays a role in recurrence. We focused on two aspects of reinfection that have received little attention. First, we evaluated whether a lack of methodological refinement could lead to inaccurate assignment of mixed infections as exogenous reinfection, in which a differential selection of each of the coinfecting strains occurred over time. We used the mycobacterial interspersed repetitive-unit-variable-number tandem-repeat (MIRU-VNTR) method to genotype 122 isolates from 40 patients with recurrent tuberculosis. We identified 11/40 (27.5%) cases with genotypic differences between the isolates involved in the sequential episodes. Major genotypic differences were found in 8/11 cases, suggesting exogenous reinfection; in the remaining 3 cases, subtle genotypic differences were observed, probably indicating microevolution from a parental strain. In all cases, only a single strain was detected for the isolate(s) from each episode, thus ruling out the possibility that reinfection could correspond to undetected mixed infection. Second, we analyzed the infectivity of a selection of 12 strains from six cases with genotypically different strains between episodes. No main differences were observed in an ex vivo model of infection between the strains involved in the first episodes and those involved in the recurrent episodes. In our setting, our results suggest the following: (i) the possibility of misassignment of mixed infection as exogenous reinfection is improbable, and (ii) bacterial infectivity does not seem to play a role in exogenous reinfection. The traditional assumptions in the analysis of infection byMycobacterium tuberculosis are that each episode is caused by a single M. tuberculosis strain and that recurrence is considered to result from an endogenous reactivation of the strain involved in the first episode. In recent years, molecular analysis of M. tuberculosis isolates has revealed that a recurrence may also be caused by a strain other than that isolated in the first episode, suggesting that exogenous reinfection plays a role in recurrence of tuberculosis (5,8,26). More complex situations have also been observed, including simultaneous infection by two M. tuberculosis strains (5, 28) and infection by the same strain at different anatomical sites (13) or at different sites within the lung (14).Exogenous reinfection has mainly been observed in immunocompromised patients (8, 21) and in settings with a high prevalence of tuberculosis (8, 26) or a high risk for transmission (5), thus suggesting that mainly socioepidemiological factors determine the occurrence of these phenomena. However, detection of exogenous reinfection in settings with moderate and low prevalence of tuberculosis and in both HIV-positive and HIV-negative patients (2, 4, 12) could indicate that specific bacterial factors m...

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“…If XDR-TB becomes increasingly prevalent within nosocomial settings in Tomsk, time spent in these facilities could, in the future, become associated with risk of infection or reinfection with XDR-TB, as has been observed in other epidemic locations (22). In addition to reinfection during MDR treatment, it is also possible that heteroresistance (clonal heterogeneity with subpopulations of MDR and XDR variants) (23) or mixed infection with different MDR and XDR strains were present at baseline (24)(25)(26)(27). If the XDR strains were relatively rare within an individual, standard drug resistance testing may not have detected XDR that was present at the time of treatment initiation for MDR disease.…”

Section: Discussionmentioning

confidence: 99%

Development of Extensively Drug-resistant Tuberculosis during Multidrug-resistant Tuberculosis Treatment

Shin¹,

Keshavjee²,

Gelmanova³

et al. 2010

Am J Respir Crit Care Med

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Rationale: Extensively drug-resistant (XDR) tuberculosis (TB) may arise in individuals on treatment for multidrug-resistant (MDR) TB. Preventing this amplification of resistance will likely improve clinical outcomes and delay the secondary spread of XDR-TB. Objectives: To measure the proportion of individuals that develops XDR-TB during the course of MDR-TB treatment, and to identify those factors associated with the development of XDR. Methods: We performed a retrospective analysis of 608 consecutive patients with documented MDR-TB who were started on MDR-TB treatment between September 10, 2000 and November 1, 2004 in the Tomsk Oblast TB Treatment Services in Western Siberia, Russian Federation. Measurements and Main Results: A total of 6% of patients were observed to develop XDR-TB while on MDR-TB treatment. These patients were significantly less likely to be cured or to complete treatment. Using Cox proportional hazard models, we found that the presence of bilateral and cavitary lesions was associated with a greater than threefold increase in hazard (adjusted hazard ratio [HR], 3.47; 95% confidence interval [CI], 1.32-9.14). Prior exposure to a secondline injectable antibiotic was associated with a greater than threefold increase in hazard (adjusted HR, 3.65; 95% CI, 1.81-7.37), and each additional month in which a patient failed to take at least 80% of their prescribed drugs was associated with nearly an additional 20% hazard of developing XDR-TB (adjusted HR, 1.17; 95% CI, 1.01-1.35). Conclusions: Early and rapid diagnosis, timely initiation of appropriate therapy, and programmatic efforts to optimize treatment adherence during MDR-TB therapy are crucial to avoiding the generation of excess XDR-TB in MDR-TB treatment programs.

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Characterization of Clonal Complexity in Tuberculosis by Mycobacterial Interspersed Repetitive Unit-Variable-Number Tandem Repeat Typing

Cited by 50 publications

References 21 publications

Molecular Detection of Mixed Infections of Mycobacterium tuberculosis Strains in Sputum Samples from Patients in Karonga District, Malawi

Molecular Detection of Mixed Infections of Mycobacterium tuberculosis Strains in Sputum Samples from Patients in Karonga District, Malawi

Evaluation of the Inaccurate Assignment of Mixed Infections by Mycobacterium tuberculosis as Exogenous Reinfection and Analysis of the Potential Role of Bacterial Factors in Reinfection

Development of Extensively Drug-resistant Tuberculosis during Multidrug-resistant Tuberculosis Treatment

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