Using diphenyleneiodonium to induce a viable but non-culturable phenotype in Mycobacterium tuberculosis and its metabolomics analysis

Abstract

Depletion of oxygen levels is a well-accepted model for induction of non-replicating, persistent states in mycobacteria. Increasing the stress levels in mycobacterium bacilli facilitates their entry into a non-cultivable, dormant state. In this study, it was shown that diphenyleneiodonium, an inhibitor of NADH oxidase, induced a viable, but non-culturable state in mycobacteria, having similar features to dormant bacilli, like loss of acid-fastness, upregulation of stress-regulated genes and decreased superoxide levels as compared to actively growing bacilli. Comprehensive, untargeted metabolic profiling also confirmed a decrease in biogenesis of amino acids, NAD, unsaturated fatty acids and nucleotides. Additionally, an increase in the level of lactate, fumarate, succinate and pentose phosphate pathways along with increased mycothiol and sulfate metabolites, similar to dormant bacilli, was observed in the granuloma. These non-cultivable bacilli were resuscitated by supplementation of fetal bovine serum, regaining their culturability in liquid as well as on agar medium. This study focused on the effect of diphenyleneiodonium treatment in causing mycobacteria to rapidly transition from an active state into a viable, but non-cultivable state, and comparing their characteristics with dormant phenotypes.

Bovine serum albumin promotes reactivation of viable but non-culturableMycobacterium tuberculosisvia the activation of protein kinase-dependent cell division processes

“…Figure 2 shows the DPI treatment results for the induction of VBNC state in H37Rv and H37Ra (18). As shown in Fig.…”

“…Previously, NADH oxidase inhibitor diphenyleneiodonium chloride (DPI) was reported to simply and rapidly induce a VBNC state in the Mtb H37Ra strain, and incubation with fetal bovine serum, which is a common supplement for mammalian cell cultures, could facilitate the reactivation of VBNC cells, while incubation with OADC (oleate-albumin-dextrose-catalase) supplementation could not (18). This report gave us an important clue for constructing a simple and rapid assay system to induce and reactivate VBNC Mtb cells.…”

Adherence of Trichomonas vaginalis to SiHa Cells is Inhibited by Diphenyleneiodonium

“…As such, DPI treatment may induce a viable but nonculturable (VBNC) state in T. vaginalis . DPI has been similarly reported to induce the rapid transition from an active to a VBNC state in Mycobacterium tuberculosis [ 40 ]. Collectively, these data strongly suggest that DPI contribute to parasite detachment from host cells during T. vaginalis infection.…”

Mass spectrometry-based identification of new serum biomarkers in patients with latent infection pulmonary tuberculosis

“…Lau SKP et al [10] reported that the content of 24 metabolites in the culture supernatant of M. tuberculosis was significantly higher than that of non- M. tuberculosis group, of which 4 metabolites were identified as 1-tuberculin adenosine derivatives, which may be used as a new marker of metabolism of M. tuberculosis . Yeware A et al [11] focused on the mechanism of rapid transition of M. tuberculosis from the active state to the viable but non-culturable state caused by ammonium diphenylene alkylate treatment through metabolomics and compared it with the dormant phenotype.…”