3.3. Correlated superoxide radical levels and silver nanoparticle levels of individual bacteria
Having evidenced the differing SR levels of bacteria using fluorescence microscopy and varying silver nanoparticle concentration inside a bacterium using back reflection imaging, we finally investigate the possible correlation between the levels of two biologically redox active products at the single bacterium level. Experimentally, the DHE fluorescence probes were added to a bacterial mixture after the incubation with Ag+ to create DHE-labelled Ag particle-containing bacteria. In turn, bright field microscopy, back reflection imaging and fluorescence microscopy were applied to the immoblised bacteria in a same optical field. Consequently, the optical intensity of respective images are compared for each single entity.
Surprisingly, as the bright field image in Figure 4a presents all the adhered bacteria in the optical field, a small fraction (<20%) of the bacteria exhibiting an exceptional back reflection intensity (Figure 4b) are shown to be exactly complementary to those exhibiting a high fluorescent intensity (Figure 4c). Ultimately, the fluorescent intensity and the back reflection intensity of a total number of 1626 bacteria were individually measured and plotted in the correlation chart (Figure 4d). Overall, a distinct negative correlation between the two kinds of optical signals is evident. This strongly suggests that a bacterium which has a potent ability to reduce silver ions tends to possess a low superoxide radical level. This conclusion further implies that the two different kinds of biological activities – superoxide radical reduction and silver ion reduction – may share to some extent the same reducing system withinD.r. bacteria: in this case, the pool of antioxidant molecules in the bacterial cytosol as discussed in Introduction. On the ground that most single entity research focuses on structure-activity relationship, our work suggests an interesting perspective of looking into the relationship between different activities of a same microbe, which may gain fundamental implications for chemical aspects of biology.