Metabolic Versatility of Shewanella oneidensis MR-1

M1 defined medium containing 0.02% (w/v) of vitamin-free Casamino Acids and 15 mM lactate was used in all physiological experiments [72 (link)]. Growth of the deletion strain under aerobic or anaerobic conditions was determined by recording growth curves in triplicate with a Bioscreen C microbiology reader (Labsystems Oy, Helsinki, Finland) with MR-1 as the control. For aerobic growth, exponential phase cultures were diluted to approximately ~1 × 10⁵ cells/ml in fresh medium, and 400 μl was transferred to the honeycomb plate wells of the Bioscreen C reader. The cultures were shaken at medium intensity continuously, and the turbidity was measured every 30 min at 600 nm and DO (dissolved oxygen) was recorded every hour with an Accumet XL40 meter (Fisher Scientific). For anaerobic growth, exponential phase cultures grown aerobically were centrifuged, purged in nitrogen and suspended in fresh medium to approximately ~1 × 10⁵ cells/ml in an anaerobic glove box. Electron acceptors tested in this study included fumarate (20 mM), nitrate (2 mM), nitrite (1 mM), thiosulfate (3 mM), TMAO (20 mM), and DMSO (20 mM). For electron acceptors containing metals including MnO₂ (5 mM), ferric citrate (10 mM), and cobalt(III)-EDTA (200 μM), growth was monitored by the color change of the cultures and cell counting under a microscope (Nikon Optiphot, Nikon, Japan).
Survival of MR-1 and the ΔarcA strain during the stationary phase was examined. Cultures were grown from a single colony under aerobic conditions with vigorous shaking. After the onset of stationary phase, the cultures were divided into two parts. One was kept in the incubator with vigorous shaking and the other was kept still. The cultures were serially diluted into LB and plated onto LB plates every 12 h. Plates from dilutions that gave 100 to 250 colony form units (CFU) per plate were used to minimize statistical variation due to small sample sizes. Experiments were done in triplicate.