Distribution of calcium and other elements in cryosectioned Bacillus cereus T spores, determined by high-resolution scanning electron probe x-ray microanalysis

Abstract

The distribution of a number of key elements in B. cereus T spores was determined by high-resolution scanning electron probe X-ray microanalysis. To circumvent the redistribution of soluble or weakly bound elements, freeze-dried cryosections of spores, which had been rapidly frozen in 50% aqueous polyvinyl pyrrolidone, were employed. The sections were examined by using a modified Philips EM400 electron microscope fitted with a field emission gun, scanning transmission electron microscopy attachment and a computer-linked energy-dispersive X-ray microanalysis system. X-ray maps for selected elements and the corresponding electron image were produced simultaneously by scanning the cryosections with a fine electron beam in a raster pattern, using the scanning transmission electron microscopy attachment. Almost all of the Ca, Mg and Mn, together with most of the P, was located in the core region. An unexpectedly high concentration of silicon was found in the cortex/coat layer. Granules containing high concentrations of Ca, Mn and P were demonstrated in spores containing reduced levels of dipicolinic acid. Spot mode analyses, in which a stationary beam was located over the region of interest in the spore cryosection, confirmed the results obtained with the scanning mode and also provided a more accurate quantitation of the elemental concentrations on a dry weight basis.