Analysis of As- and Hg-species in metal-resistant oral bacteria, by imaging ToF-SIMS.

Bacterial strains were isolated from the oral cavity of healthy volunteers and grown in the presence of Hg-ions (1-10 ppm) or arsenate ions at concentrations of 0.1-1.0%. To elucidate how bacteria take up and transform toxic metals inside the cells, we performed ion imaging and depth profiling with time-of-flight secondary ion mass spectrometry (ToF-SIMS). This analysis relies on the use of a pulsed ion beam to ionize surface molecules that can be extracted into a time-of-flight mass spectrometer. By combining the pulsed ion beam with another ion beam in direct current (DC) mode, depth profiles are obtained as a result of consecutive removal of surface layers. The spatial resolution of the depth profile along the y-axis is in the range of a few nanometres, and the spatial resolution along the x- and z-axes is in the μm range. The ToF-SIMS analysis was performed on crude biofilms of bacteria air-dried at aluminium foil surfaces, allowing subcellular resolution along the y-axis. The mercury ions were found transformed to methylmercury preferably in the periplasmic space, and the arsenate ions were found reduced to arsenite inside the cells, close to the cell membrane. The data are discussed in relation to current concepts in bacterial resistance to metals and antibiotics.