Superoxide Generation and Its Involvement in the Growth of Mycobacterium smegmatis

Abstract

Superoxide generation is inevitable in aerobic organisms, most of which have developed mechanisms to detoxify superoxides. However, its significance has not been clearly understood in mycobacteria. This study demonstrates that NADH oxidase is the major source of superoxide in Mycobacterium smegmatis and elucidates the involvement of superoxide in M. smegmatis growth. The maximum inhibition of superoxide generation was observed in the presence of diphenyleneiodonium chloride (DPI), an NADH oxidase inhibitor, compared to other standard inhibitors. After incubation for 24 h, the number of colony forming units (CFUs) was reduced by 6.8 log10 compared to the untreated culture. The inhibitory effect of DPI on M. smegmatis was reversed when the same culture was supplemented with menadione and pyrogallol, which are superoxide generators. Thus, this study reports the source of superoxide generation and its involvement in the growth of M. smegmatis.

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

“…To determine comparative levels of superoxide production under different dormancy conditions, the dihydroethidium (DHE)-HPLC method was used [16]. Briefly, DHE (50 μM) was added to 17.5 mL of the following cultures: untreated aerobically growing culture (~0.35 OD 600 nm ), DPI treated culture (4 μg/mL, 24 h), Vitamin C treated culture (10 mM, 24 h) and Wayne hypoxic dormant culture (16 days) in 20 mm × 125 mm tubes and incubated for 90 minutes in the dark.…”

“…Ascorbic acid (10 mM), an antioxidant also reported to induce dormancy and VBNC phenotypes in Mtb [14–15], indicates the role of reactive oxygen species (ROS) scavenging during the development of a dormant form. Superoxide, a reactive oxygen species generated during electron transfer reactions in NADH oxidase, can be inhibited by diphenyleneiodonium (DPI) [16]. Yeware et al (2017) have described how DPI inhibits superoxide production as well as growth of M .…”

“…Yeware et al (2017) have described how DPI inhibits superoxide production as well as growth of M . smegmatis ; this inhibition is reversed by the addition of an external source of superoxide [16]. In contrast, nitric oxide (NO) and hydrogen peroxide have been shown to be either bacteriostatic or infective growth inhibitors at moderate concentrations [17].…”

The microbial contribution to reactive oxygen species dynamics in marine ecosystems

“…While traditionally considered a threat to cellular viability, there are cases where microbes exploit ROS for their own benefit. Intracellular superoxide can serve as a signal for development in eukaryotes (Bloomfield and Pears, ; Finkel, ; Mittra and Andrews, ), and as a growth factor for the bacterium Mycobacterium smegmatis (Yeware et al ., ) and the eukaryotic red tide phytoplankter Chattonella marina (Oda et al ., ). As we will see later, marine microbes may generate extracellular to enhance the uptake of iron.…”

Commentary: Superoxide Generation and Its Involvement in the Growth of Mycobacterium smegmatis

“…Yeware et al (2017), observed no effect of PEG-SOD on either growth or superoxide production in cells, which they interpret as non-permeability of PEG-SOD. Moreover, in crude membrane preparation PEG-SOD could barely inhibit 0.37 folds of 2-OH-E + .…”