Are the Soft, Liquid-Like Structures Detected around Bacteria by Ambient Dynamic Atomic Force Microscopy Capsules?

Abstract

ABSTRACTHigh-resolution imaging of bacterial capsules by microscopy is of paramount importance in microbiology due to their role in pathogenesis. This is, however, quite a challenging task due to their delicate nature. In this context, recent reports have claimed successful exploitation of the capacity of atomic force microscopy (AFM) for imaging of extremely deformable (even liquid) surfaces under ambient conditions to detect bacterial capsules in the form of tiny amounts of liquid-like substances around bacteria. In order to further explore this supposed capacity of AFM, in this work, three staphylococcal strains have been scrutinized for the presence of capsules using such an AFM-based approach with a phosphate buffer and water as the suspending liquids. Similar results were obtained with the three strains. AFM showed the presence of liquid-like substances identical to those attributed to bacterial capsules in the previous literature. Extensive imaging and chemical analysis point out the central role of the suspending liquid (buffer) in the formation of these substances. The phenomenon has been reproduced even by using nonliving particles, a finding that refutes the biological origin of the liquid-like substances visualized around the cells. Deliquescence of major components of biological buffers, such as K2HPO4, CaCl2, or HEPES, is proposed as the fundamental mechanism of the formation of these ultrasmall liquid-like structures. Such an origin could explain the high similarity of our results obtained with three very different strains and also the high similarity of these results to others reported in the literature based on other bacteria and suspending liquids. Finally, possible biological/biomedical implications of the presence of these ultrasmall amounts of liquids wrapping microorganisms are discussed.