Nile red fluorescence demonstrates lipid in the envelope of vesicles from N2-fixing cultures of Frankia

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Abstract
Using Nile red as a fluorescent stain for lipid, we investigated the composition of the envelope of vesicles of Frankia strain HFPCcI3 from cultures induced in N-free medium at 1 and 20 kPa O2. Vesicles and nitrogenase activity appeared in the cultures at both pO2; on average, vesicles viewed by dark-field microscopy were larger and had thicker envelopes at 20 kPa O2 than at 1 kPa O2. Envelopes of Nile red-stained vesicles fluoresced red under incident green light. When samples of CcI3 were extracted through a lipid-solvent series and then stained, vesicles still fluoresced red but lacked a distinct peripheral fluorescent ring. These results are consistent with the view that the envelope of Frankia vesicles consists largely of lipid and serves as a barrier to diffusion of O2.