Effect of Water Washing Purification on the Surface Morphology of Bacillus cereus Spores

The architecture of spore surfaces plays an important role in the pathogenesis and natural ecology of many Bacillus organisms. However, changes to the laboratory preparation method for Bacillus spores may be a source of phenotypic variability in cell surfaces that can affect our understanding of in situ morphology and make cross-study comparisons more difficult. In this work, we examined how variation in the number of water washing steps during spore purification influenced the three-dimensional morphology of Bacillus cereus spore surfaces. Two strains of B. cereus, str. 14579 and T-strain, were cultured in liquid medium and purified using 0, 1, 3, or 5 water washing steps. Results showed that the nanoscale morphology of the spore surface is affected by the number of water washes such that early wash step samples had higher levels of roughness, quantified with a Root Mean Square (RMS) algorithm, which decreased with successive washes. The presence of large (>200 nm) surficial features, representing cell fragments and/or debris from the culture, was also inversely correlated with the number of washes. The presence of cellular debris in unwashed spore samples also changed the nanomechanical signatures of the outermost surface but did not affect the properties of the underlying cell. This suggests that variation in purification method can affect the nanoscale morphology and mechanics and should be considered when analyzing microto nano-scale surface properties of Bacillus cereus spores. Journal of Nature and Science, 1(6):e116, 2015