Analysis of dye binding by and membrane potential in spores of Bacillus species.

摘要

This study sought to determine roles of coats in staining of Bacillus subtilis spores, and whether spores have membrane potential. Staining by 4 dyes and autofluorescence of B. subtilis spores that lack some (cotE, gerE) or most (cotE gerE) coat protein was measured. Wild-type, cotE and gerE spores autofluoresced and bound dyes, but cotE gerE spores did not autofluoresce and were stained only by 2 dyes. A membrane potential-sensitive dye DiOC6(3) bound to dormant B. megaterium and B. subtilis spores. This binding was abolished by the protonophore FCCP, DiOC6(3) bound to heat-killed spores, but not to dormant B. subtilis cotE gerE spores. However, DiOC6(3) bound well to all germinated spores. It is concluded that autofluorescence of dormant B. subtilis spores and binding of some dyes are due to the coat. There is no membrane potential in dormant Bacillus spores, although membrane potential is generated when spores germinate. The elimination of the autofluorescence of B. subtilis spores may allow assessment of the location of low abundance spore proteins using fluorescent reporter technology. The dormant spore's lack of membrane potential may allow tests of spore viability by assessing membrane potential in germinating spores.