Tracking Inhibitory Alterations during Interstrain <i>Clostridium difficile</i> Interactions by Monitoring Cell Envelope Capacitance

Global threats arising from the increasing use of antibiotics coupled with the high recurrence rates of <i>Clostridium difficil</i>e (<i>C. difficile</i>) infections (CDI) after standard antibiotic treatments highlight the role of commensal probiotic microorganisms, including nontoxigenic <i>C. difficile</i> (NTCD) strains in preventing CDI due to highly toxigenic <i>C. difficile</i> (HTCD) strains. However, optimization of the inhibitory permutations due to commensal interactions in the microbiota requires probes capable of monitoring phenotypic alterations to <i>C. difficile</i> cells. Herein, by monitoring the field screening behavior of the <i>C. difficile</i> cell envelope with respect to cytoplasmic polarization, we demonstrate that inhibition of the host-cell colonization ability of HTCD due to the S-layer alterations occurring after its co-culture with NTCD can be quantitatively tracked on the basis of the capacitance of the cell envelope of co-cultured HTCD. Furthermore, it is shown that effective inhibition requires the dynamic contact of HTCD cells with freshly secreted extracellular factors from NTCD because contact with the cell-free supernatant causes only mild inhibition. We envision a rapid method for screening the inhibitory permutations to arrest <i>C. difficile</i> colonization by routinely probing alterations in the HTCD dielectrophoretic frequency response due to variations in the capacitance of its cell envelope

Tracking Inhibitory Alterations during Interstrain <i>Clostridium difficile</i> Interactions by Monitoring Cell Envelope Capacitance

Global threats arising from the increasing use of antibiotics coupled with the high recurrence rates of <i>Clostridium difficil</i>e (<i>C. difficile</i>) infections (CDI) after standard antibiotic treatments highlight the role of commensal probiotic microorganisms, including nontoxigenic <i>C. difficile</i> (NTCD) strains in preventing CDI due to highly toxigenic <i>C. difficile</i> (HTCD) strains. However, optimization of the inhibitory permutations due to commensal interactions in the microbiota requires probes capable of monitoring phenotypic alterations to <i>C. difficile</i> cells. Herein, by monitoring the field screening behavior of the <i>C. difficile</i> cell envelope with respect to cytoplasmic polarization, we demonstrate that inhibition of the host-cell colonization ability of HTCD due to the S-layer alterations occurring after its co-culture with NTCD can be quantitatively tracked on the basis of the capacitance of the cell envelope of co-cultured HTCD. Furthermore, it is shown that effective inhibition requires the dynamic contact of HTCD cells with freshly secreted extracellular factors from NTCD because contact with the cell-free supernatant causes only mild inhibition. We envision a rapid method for screening the inhibitory permutations to arrest <i>C. difficile</i> colonization by routinely probing alterations in the HTCD dielectrophoretic frequency response due to variations in the capacitance of its cell envelope

Tracking Inhibitory Alterations during Interstrain <i>Clostridium difficile</i> Interactions by Monitoring Cell Envelope Capacitance

Global threats arising from the increasing use of antibiotics coupled with the high recurrence rates of <i>Clostridium difficil</i>e (<i>C. difficile</i>) infections (CDI) after standard antibiotic treatments highlight the role of commensal probiotic microorganisms, including nontoxigenic <i>C. difficile</i> (NTCD) strains in preventing CDI due to highly toxigenic <i>C. difficile</i> (HTCD) strains. However, optimization of the inhibitory permutations due to commensal interactions in the microbiota requires probes capable of monitoring phenotypic alterations to <i>C. difficile</i> cells. Herein, by monitoring the field screening behavior of the <i>C. difficile</i> cell envelope with respect to cytoplasmic polarization, we demonstrate that inhibition of the host-cell colonization ability of HTCD due to the S-layer alterations occurring after its co-culture with NTCD can be quantitatively tracked on the basis of the capacitance of the cell envelope of co-cultured HTCD. Furthermore, it is shown that effective inhibition requires the dynamic contact of HTCD cells with freshly secreted extracellular factors from NTCD because contact with the cell-free supernatant causes only mild inhibition. We envision a rapid method for screening the inhibitory permutations to arrest <i>C. difficile</i> colonization by routinely probing alterations in the HTCD dielectrophoretic frequency response due to variations in the capacitance of its cell envelope

Label-Free Quantification of Intracellular Mitochondrial Dynamics Using Dielectrophoresis

Mitochondrial dynamics play an important role within several pathological conditions, including cancer and neurological diseases. For the purpose of identifying therapies that target aberrant regulation of the mitochondrial dynamics machinery and characterizing the regulating signaling pathways, there is a need for label-free means to detect the dynamic alterations in mitochondrial morphology. We present the use of dielectrophoresis for label-free quantification of intracellular mitochondrial modifications that alter cytoplasmic conductivity, and these changes are benchmarked against label-based image analysis of the mitochondrial network. This is validated by quantifying the mitochondrial alterations that are carried out by entirely independent means on two different cell lines: human embryonic kidney cells and mouse embryonic fibroblasts. In both cell lines, the inhibition of mitochondrial fission that leads to a mitochondrial structure of higher connectivity is shown to substantially enhance conductivity of the cell interior, as apparent from the significantly higher positive dielectrophoresis levels in the 0.5–15 MHz range. Using single-cell velocity tracking, we show ∼10-fold higher positive dielectrophoresis levels at 0.5 MHz for cells with a highly connected versus those with a highly fragmented mitochondrial structure, suggesting the feasibility for frequency-selective dielectrophoretic isolation of cells to aid the discovery process for development of therapeutics targeting the mitochondrial machinery