Dramatic reduction of culture time of Mycobacterium tuberculosis

Ghodbane, Ramzi; Raoult, Didier; Drancourt, Michel

doi:10.1038/srep04236

Cited by 64 publications

(58 citation statements)

References 19 publications

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“…We tried to optimize solid culture media and protocols in order to help speed up and simplify the routine isolation of mycobacteria, including MTC mycobacteria; these efforts yielded a new protocol and a new medium (MOD4) with good performance (6). Here, we modified the MOD4 medium formulation in order to improve its shelf life by eliminating the requirement for fresh blood.…”

Section: Discussionmentioning

confidence: 99%

“…The MOD9 culture medium was derived from the MOD4 culture medium (6) in which the 5% sheep blood and the 15% heat-inactivated bovine serum were eliminated and 15% heat-inactivated lamb serum (Life Technologies, Saint Aubin, France) was added. Also, 100 mg/liter ascorbic acid was added to mimic a microaerophilic atmosphere previously proven to be beneficial for M. tuberculosis growth (6). A mix of azorubine and Ponceau 4R normally used as food dye (Sainte Lucie, Gouvieux, France) was added to stain the medium red and increase the contrast with the white-beige colonies of mycobacteria.…”

Section: Methodsmentioning

confidence: 99%

“…Recently, we showed that the MOD4 culture medium incorporating blood and egg lecithin, combined with an improved laboratory workflow, significantly reduced the time to detection of cultured M. tuberculosis (6).…”

mentioning

confidence: 99%

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A Novel Solid Medium for Culturing Mycobacterium tuberculosis Isolates from Clinical Specimens

et al. 2015

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The laboratory diagnosis of tuberculosis usually relies on culture-based isolation of the causative Mycobacterium tuberculosis bacteria. We developed and evaluated the performance of MOD9, a new blood-free derivative of the MOD4 solid medium on which we previously reported for the isolation and culture of mycobacteria. First, inoculation of Lowenstein-Jensen medium with 21 M. tuberculosis isolates at 10 5 , 10 3 , and 10 CFU yielded colonies in 5.7 ؎ 1.5 days, 7.6 ؎ 0.8 days, and 10.8 ؎ 1.7 days versus 1.5 ؎ 0.4 days, 3.5 ؎ 0.6 days, and 4.9 ؎ 1 days in MOD9 (P < 0.05, Student's t test). Further, the time to detectable growth of M. tuberculosis was measured on MOD9 and Lowenstein-Jensen media after duplicate inoculation of 250 clinical specimens decontaminated with 0.7% chlorhexidine. The contamination rate was 1.6% (4/250) on MOD9 versus 4.4% (11/250) on Lowenstein-Jensen medium (P ‫؍‬ 0.11, Fisher's exact test). Chlorhexidine-MOD9 yielded 38/250 (15.2%) isolates versus 32/250 (12.8%) isolates for the chlorhexidine-LJ (P ‫؍‬ 0.5195, Fisher's exact test). All together, eight M. tuberculosis isolates were cultured solely from chlorhexidine-MOD9, and two M. tuberculosis isolates were cultured solely from chlorhexidine-LJ. The time to detection was 9.8 ؎ 3.9 (range, 5 to 18) days for chlorhexidine-MOD9 versus 17.4 ؎ 5.9 (range, 10 to 35) days for chlorhexidine-LJ (P < 0.05, Student's t test). These data indicate the superiority of the MOD9 medium over the standard LJ medium following chlorhexidine decontamination for the recovery of M. tuberculosis. P ulmonary tuberculosis caused by Mycobacterium tuberculosis complex (MTC) mycobacteria remains a global public health problem, causing on average 170 deaths every hour worldwide (1). Despite significant advances in the molecular diagnosis of tuberculosis over the past 2 decades (2), culture is still the universal gold standard for the laboratory diagnosis of tuberculosis, enabling complete postculture antimicrobial susceptibility testing and genotyping (3).MTC mycobacteria are fastidious organisms whose growth is routinely detected only after 7 to 12 days using advanced commercially available automated systems (4). These robots use various formulations of the Middlebrook liquid culture medium and indeed reduce the delay in growth detection compared to that with inoculation of standard solid culture media, such as Lowenstein-Jensen (LJ) medium (4). The World Health Organization (WHO) still recommends inoculating specimens in parallel into a liquid medium for accelerated diagnosis of high-titer specimens and onto a solid culture medium to increase the sensitivity of laboratory diagnosis of pulmonary tuberculosis (5).In an effort to conciliate speed and sensitivity of the culturebased diagnosis of tuberculosis, we developed new formulations of solid culture medium and new protocols for incubating and detecting colonies of M. tuberculosis. Recently, we showed that the MOD4 culture medium incorporating blood and egg lecithin, combined with an improved laboratory workflow, s...

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Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

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A Novel Solid Medium for Culturing Mycobacterium tuberculosis Isolates from Clinical Specimens

et al. 2015

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“…One major drawback of tuberculosis culture is the slow growth of Mtb (Ghodbane et al, 2014). While recent advances have made the culture of Mtb in less than 48 h a possibility (Ghodbane et al, 2015), currently these techniques are technologically sophisticated and limited to reference laboratory/research settings.…”

Section: Mycobacterial Culturementioning

confidence: 99%

Resuscitation-promoting factors are important determinants of the pathophysiology inMycobacterium tuberculosisinfection

Rosser

Stover

Pareek

et al. 2017

Critical Reviews in Microbiology

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Resuscitation promoting factors (Rpf) are peptidoglycan-hydrolyzing enzymes that are pivotal in the resuscitation of quiescent actinobacteria including Mycobacterium tuberculosis. From the published data, it is clear that Rpf are required for the resuscitation of non-replicating bacilli and pathogenesis in murine infection model of tuberculosis, although their direct influence on human Mycobacterium tuberculosis infection is ill-defined. In this review, we describe the progress in the understanding of the roles that Rpf play in human tuberculosis pathogenesis and importance of bacilli dependent upon Rpf for growth for the outcome of human tuberculosis. We outline how this research is opening up important opportunities for the diagnosis, treatment and prevention of human disease, progress in which is essential to attain the ultimate goal of tuberculosis eradication.ARTICLE HISTORY

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“…It is known that, the growth of lactobacilli can be strongly affected by Tween 80 which is often included in the culture media as a growth factor for fastidious bacteria as well as for lactic acid bacteria and Mycobacterium tuberculosis (Li et al, 2011;Ghodbane et al, 2014). The oleic acid present in Tween 80 is incorporated into the lipid membranes of bacteria altering the fatty acid composition, fluidity and its permeability ( Nikkila et al, 1995;Kankaanpää et al, 2004).…”

Section: Growth Of the Lactobacillus Speciesmentioning

confidence: 99%

Activity of metabolites produced by new strains of Lactobacillus in modified de Man, Rogosa and Sharpe (MRS) medium against multidrug-resistant bacteria

Mesquita¹,

Costa²,

Frutuoso³

et al. 2017

Afr. J. Microbiol. Res.

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The emergence of nearly untreatable infections caused by multidrug-resistant bacteria has led to a new public health concern in which a need for development of alternative non-antibiotic strategies has become urgent. The activity of metabolites produced by new strains of Lactobacillus against multidrugresistant bacteria was investigated. The objective of this work was to isolate and identify lactobacilli from artisanal kefir by 16S rRNA gene sequencing as well as to evaluate the effect of the growth of Lactobacillus species in the Man Rogosa and Sharpe broth, supplemented with Tween 80 at concentrations of 0.6, 0.4, and 0.2 %. Cell Free Supernatants (CFSs) were obtained from these lactobacilli cultures and with them, organic acids (lactic acid and acetic acid) and ethanol were quantified by HPLC followed by the determination of their antimicrobial activities against eight strains of Staphylococcus aureus-MRSA and Klebsiella pneumonia KPC strains. The GenBank BLAST analysis revealed that, the isolated lactobacilli belonged to Lactobacillus paracasei (n=4) and Lactobacillus plantarum (n=5) strains. Increasing concentrations of Tween 80 did not affect the growth of Lactobacillus species significantly when compared to their controls (MRS broth). HPLC analysis of CFSs showed concentrations greater than 18.0, 4.0 and 1.0 g/L for lactic acid, acetic acid and ethanol, respectively. All CFSs were able to inhibit all pathogenic microorganisms evaluated. The percentage of inhibition was on average greater than 88% for MRSA and KPC strains. The antimicrobial activity was dependent on the CFSs tested. Based on these experimental conditions, organic acids and ethanol are likely to be responsible for this antimicrobial activity.

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Dramatic reduction of culture time of Mycobacterium tuberculosis

Cited by 64 publications

References 19 publications

A Novel Solid Medium for Culturing Mycobacterium tuberculosis Isolates from Clinical Specimens

A Novel Solid Medium for Culturing Mycobacterium tuberculosis Isolates from Clinical Specimens

Resuscitation-promoting factors are important determinants of the pathophysiology inMycobacterium tuberculosisinfection

Activity of metabolites produced by new strains of Lactobacillus in modified de Man, Rogosa and Sharpe (MRS) medium against multidrug-resistant bacteria

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