Persister populations of Mycobacterium tuberculosis in sputum that grow in liquid but not on solid culture media

Dhillon, Jasvir; Fourie, P. Bernard; Mitchison, D.A.

doi:10.1093/jac/dkt357

Cited by 58 publications

(50 citation statements)

References 11 publications

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“…The right panel of the figure shows bar charts of estimated median log 10 (total CFU), represented by the total bar length, with the viable CFU in blue above the line and the elusive CFU in red below the line. This graphic is comparable to the proverbial 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 61 62 63 64 and they do not grow in conventional culture media [15,18,19,38]. Consequently, antibiotic treatments aimed at the entire bacterial population are evaluated only on the population that can grow in culture, in the hope that activity measured on bacteria viable in culture will translate into activity against all populations.…”

Section: Discussionsupporting

confidence: 71%

“…It is well described in vitro that M. tuberculosis develops a reversible transition to a persistent state of general metabolic downregulation and activation of alternative pathways that stop replication, decrease ATP production and shift energy sources from carbohydrates to fatty acids [5e12]. These persisters are tolerant to commonly used antituberculosis drugs [6,13,14] and unable to form colonies on agar plates but may grow in liquid media [15], or when treated with recombinant or supernatant-derived resuscitationpromoting factors [16,17]. Such cells lack a proper cell membrane, are no longer acid-fast and fail to retain enough Auramine-O stain 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 for detection with routine fluorescent microscopy, whereas accumulated intracellular lipids can be stained with fluorescent NileRed stain [6,13,18,19].…”

Section: Introductionmentioning

confidence: 99%

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Simultaneous staining of sputum smears for acid-fast and lipid-containing Myobacterium tuberculosis can enhance the clinical evaluation of antituberculosis treatments

et al. 2015

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Section: Discussionsupporting

confidence: 71%

Section: Introductionmentioning

confidence: 99%

Simultaneous staining of sputum smears for acid-fast and lipid-containing Myobacterium tuberculosis can enhance the clinical evaluation of antituberculosis treatments

et al. 2015

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“…Higher doses of rifampicin seem to more effectively kill this subpopulation in experimental models4, 24 and in human beings 27, 28. Since liquid media, unlike solid media, detect such bacilli, 29 we believe that the higher efficacy of RIF 35 HZE seen in liquid media represents increased activity of this regimen on this bacterial phenotype. The relevance of the liquid media endpoint is further supported by the fact that patients with treatment failure or recurrent disease had later culture conversion on liquid media, but not on solid media (although numbers were small).…”

Section: Discussionmentioning

confidence: 94%

High-dose rifampicin, moxifloxacin, and SQ109 for treating tuberculosis: a multi-arm, multi-stage randomised controlled trial

Boeree

Heinrich

Aarnoutse

et al. 2017

The Lancet Infectious Diseases

322

285

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SummaryBackgroundTuberculosis is the world's leading infectious disease killer. We aimed to identify shorter, safer drug regimens for the treatment of tuberculosis.MethodsWe did a randomised controlled, open-label trial with a multi-arm, multi-stage design. The trial was done in seven sites in South Africa and Tanzania, including hospitals, health centres, and clinical trial centres. Patients with newly diagnosed, rifampicin-sensitive, previously untreated pulmonary tuberculosis were randomly assigned in a 1:1:1:1:2 ratio to receive (all orally) either 35 mg/kg rifampicin per day with 15–20 mg/kg ethambutol, 20 mg/kg rifampicin per day with 400 mg moxifloxacin, 20 mg/kg rifampicin per day with 300 mg SQ109, 10 mg/kg rifampicin per day with 300 mg SQ109, or a daily standard control regimen (10 mg/kg rifampicin, 5 mg/kg isoniazid, 25 mg/kg pyrazinamide, and 15–20 mg/kg ethambutol). Experimental treatments were given with oral 5 mg/kg isoniazid and 25 mg/kg pyrazinamide per day for 12 weeks, followed by 14 weeks of 5 mg/kg isoniazid and 10 mg/kg rifampicin per day. Because of the orange discoloration of body fluids with higher doses of rifampicin it was not possible to mask patients and clinicians to treatment allocation. The primary endpoint was time to culture conversion in liquid media within 12 weeks. Patients without evidence of rifampicin resistance on phenotypic test who took at least one dose of study treatment and had one positive culture on liquid or solid media before or within the first 2 weeks of treatment were included in the primary analysis (modified intention to treat). Time-to-event data were analysed using a Cox proportional-hazards regression model and adjusted for minimisation variables. The proportional hazard assumption was tested using Schoelfeld residuals, with threshold p<0·05 for non-proportionality. The trial is registered with ClinicalTrials.gov (NCT01785186).FindingsBetween May 7, 2013, and March 25, 2014, we enrolled and randomly assigned 365 patients to different treatment arms (63 to rifampicin 35 mg/kg, isoniazid, pyrazinamide, and ethambutol; 59 to rifampicin 10 mg/kg, isoniazid, pyrazinamide, SQ109; 57 to rifampicin 20 mg/kg, isoniazid, pyrazinamide, and SQ109; 63 to rifampicin 10 mg/kg, isoniazid, pyrazinamide, and moxifloxacin; and 123 to the control arm). Recruitment was stopped early in the arms containing SQ109 since prespecified efficacy thresholds were not met at the planned interim analysis. Time to stable culture conversion in liquid media was faster in the 35 mg/kg rifampicin group than in the control group (median 48 days vs 62 days, adjusted hazard ratio 1·78; 95% CI 1·22–2·58, p=0·003), but not in other experimental arms. There was no difference in any of the groups in time to culture conversion on solid media. 11 patients had treatment failure or recurrent disease during post-treatment follow-up: one in the 35 mg/kg rifampicin arm and none in the moxifloxacin arm. 45 (12%) of 365 patients reported grade 3–5 adverse events, with similar proportions in each arm.Interpr...

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“…The model predicted high numbers of unquantifiable bacteria (i.e., nonmultiplying state), as inherited from the in vitro model . In vitro , animal and clinical studies support M. tuberculosis subpopulation(s) that are culture‐negative on solid media but can grow in liquid media …”

Section: Discussionmentioning

confidence: 97%

Application of the Multistate Tuberculosis Pharmacometric Model in Patients With Rifampicin‐Treated Pulmonary Tuberculosis

Svensson

Simonsson

2016

CPT Pharmacom & Syst Pharma

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This is the first clinical implementation of the Multistate Tuberculosis Pharmacometric (MTP) model describing fast‐, slow‐, and nonmultiplying bacterial states of Mycobacterium tuberculosis. Colony forming unit data from 19 patients treated with rifampicin were analyzed. A previously developed rifampicin population pharmacokinetic (PK) model was linked to the MTP model previously developed using in vitro data. Drug effect was implemented as exposure‐response relationships tested at several effect sites, both alone and in combination. All MTP model parameters were fixed to in vitro estimates except Bmax. Drug effect was described by an on/off effect inhibiting growth of fast‐multiplying bacteria in addition to linear increase of the stimulation of the death rate of slow‐ and nonmultiplying bacteria with increasing drug exposure. Clinical trial simulations predicted well three retrospective clinical trials using the final model that confirmed the potential utility of the MTP model in antitubercular drug development.

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Persister populations of Mycobacterium tuberculosis in sputum that grow in liquid but not on solid culture media

Abstract: About 90% of the bacilli in sputum are persisters that can grow in liquid media but not on solid plates.

Cited by 58 publications

References 11 publications

Simultaneous staining of sputum smears for acid-fast and lipid-containing Myobacterium tuberculosis can enhance the clinical evaluation of antituberculosis treatments

Simultaneous staining of sputum smears for acid-fast and lipid-containing Myobacterium tuberculosis can enhance the clinical evaluation of antituberculosis treatments

High-dose rifampicin, moxifloxacin, and SQ109 for treating tuberculosis: a multi-arm, multi-stage randomised controlled trial

Application of the Multistate Tuberculosis Pharmacometric Model in Patients With Rifampicin‐Treated Pulmonary Tuberculosis

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