Untargeted Metabolomics Investigation on Selenite Reduction to Elemental Selenium by Bacillus mycoides SeITE01

Bacillus mycoides SeITE01 is an environmental isolate that transforms the oxyanion selenite (SeO32−) into the less bioavailable elemental selenium (Se0) forming biogenic selenium nanoparticles (Bio-SeNPs). In the present study, the reduction of sodium selenite (Na2SeO3) by SeITE01 strain and the effect of SeO32− exposure on the bacterial cells was examined through untargeted metabolomics. A time-course approach was used to monitor both cell pellet and cell free spent medium (referred as intracellular and extracellular, respectively) metabolites in SeITE01 cells treated or not with SeO32−. The results show substantial biochemical changes in SeITE01 cells when exposed to SeO32−. The initial uptake of SeO32− by SeITE01 cells (3h after inoculation) shows both an increase in intracellular levels of 4-hydroxybenzoate and indole-3-acetic acid, and an extracellular accumulation of guanosine, which are metabolites involved in general stress response adapting strategies. Proactive and defensive mechanisms against SeO32− are observed between the end of lag (12h) and beginning of exponential (18h) phases. Glutathione and N-acetyl-L-cysteine are thiol compounds that would be mainly involved in Painter-type reaction for the reduction and detoxification of SeO32− to Se0. In these growth stages, thiol metabolites perform a dual role, both acting against the toxic and harmful presence of the oxyanion and as substrate or reducing sources to scavenge ROS production. Moreover, detection of the amino acids L-threonine and ornithine suggests changes in membrane lipids. Starting from stationary phase (24 and 48h), metabolites related to the formation and release of SeNPs in the extracellular environment begin to be observed. 5-hydroxyindole acetate, D-[+]-glucosamine, 4-methyl-2-oxo pentanoic acid, and ethanolamine phosphate may represent signaling strategies following SeNPs release from the cytoplasmic compartment, with consequent damage to SeITE01 cell membranes. This is also accompanied by intracellular accumulation of trans-4-hydroxyproline and L-proline, which likely represent osmoprotectant activity. The identification of these metabolites suggests the activation of signaling strategies that would protect the bacterial cells from SeO32− toxicity while it is converting into SeNPs.

Performance Evaluation of Selenite (SeO32−) Reduction by Enterococcus spp.

Lactic acid bacteria (LAB) such as Enterococcus spp. have an advantage over several bacteria because of their ability to easily adapt to extreme conditions which include high temperatures, highly acidic or alkaline conditions and toxic metals. Although many microorganisms have been shown to reduce selenite (SeO32−) to elemental selenium (Se0), not much work has been done on the combined effect of Enterococcus spp. In this study, aerobic batch reduction of different selenite concentrations (1, 3 and 5 mM) was conducted using Enterococcus hermanniensis sp. and Enterococcus gallinarum sp. (3.5 h, 35 ± 2 °C, starting pH > 8.5). Results from the experiments showed that the average reductions rates were 0.608, 1.921 and 3.238 mmol·(L·h)−1, for the 1, 3 and 5 mM SeO32− concentrations respectively. In addition, more selenite was reduced for the 5 mM concentration compared to the 1 and 3 mM concentrations albeit constant biomass being used for all experiments. Other parameters which were monitored were the glucose consumption rate, protein variation, pH and ORP (oxidation reduction potential). TEM analysis was also conducted and it showed the location of electron-dense selenium nanoparticles (SeNPs). From the results obtained in this study, the authors concluded that Enterococcus species’s high adaptability makes it suitable for rapid selenium reduction and biosynthesis of elemental selenium.

Multiple exposures to high concentration of selenate significantly improve selenate tolerability, red elemental selenium (Se0) and selenoprotein biosynthesis in Herbaspirillum camelliae WT00C

Herbaspirillum camelliae WT00C isolated from tea plant has an intact selenate metabolism pathway but its selenate tolerability is poor. In this study, microbiological properties between the strain WT00C and three strains CT00C, NCT00C and NT00C obtained respectively from 4, 6 and 8 rounds of 24-h exposures to 200 mM selenate were studied and compared. The selenate tolerability and the capability of generating red elemental selenium (Se0) and selenoproteins were significantly improved in H. camelliae WT00C via 4–6 rounds of multiple exposures to high concentration of selenate. The original strain WT00C grew in 200 mM selenate with the lag phase of 12 h and 400 mM selenate with the lag phase of 60 h, whereas the strains CT00C and NCT00C grew in 800 mM selenate and showed quite short lag phase when they grew in 50–400 mM selenate. Two stains also significantly improved the biosynthesis of red elemental selenium (Se0) and selenoproteins besides selenate tolerance. The stains CT00C and NCT00C exhibited more than 30% selenium conversion efficiency and 40% selenoprotein biosynthesis as compared to the original strain WT00C. These characteristics of the strains CT00C and NCT00C make them possible to be applied in pharmaceuticals and feed industries. The strain NT00C obtained from 8 rounds of 24-h exposures to 200 mM selenate was unable to grow in ≥ 400 mM selenate, and its selenium conversion efficiency and selenoprotein biosynthesis were similar to the strain WT00C. Too many exposures caused gene inactivation of some key enzymes involving in selenate metabolism and antioxidative stress. In addition, bacterial cells underwent obviously physiological and morphological changes including gene activity, cell enlargement and surface-roughness alterations during the process of multiple exposures to high concentration of selenate.