Two protocols for the detection of oleaginous bacteria using Oil Red O

Abstract

The selection of oleaginous bacteria, potentially applicable to biotechnological approaches, is usually carried out by differentexpensive and time-consuming techniques. In this study, we used Oil Red O (ORO) as an useful dye for staining of neutrallipids (triacylglycerols and wax esters) on thin-layer chromatography plates. ORO could detect minimal quantities of bothcompounds (detection limit, 0.0025 mg of tripalmitin or 0.005 mg of cetylpalmitate). In addition, we developed a specific,rapid, and inexpensive screening methodology to detect triacylglycerol-accumulating microorganisms grown on the agarplate. This staining methodology detected 9/13 strains with a triacylglycerol content higher than 20% by cellular dry weight.ORO did not stain polyhydroxyalkanoates-producing bacteria. The four oleaginous strains not detected by this screeningmethodology exhibited a mucoid morphology of their colonies. Apparently, an extracellular polymeric substance producedby these strains hampered the entry of the lipophilic dye into cells. The utilization of the developed screening methodologywould allow selecting of oleaginous bacteria in a simpler and faster way than techniques usually used nowadays, based onunspecific staining protocols and spectrophotometric or chromatographic methods. Furthermore, the use of ORO as a stainingreagent would easily characterize the neutral lipids accumulated by microorganisms as reserve compounds.