Nanosecond electric pulses rapidly enhance the inactivation of Gram-negative bacteria using Gram-positive antibiotics.

Physically disrupting microorganism membranes to enable antibiotics to overcome resistance mechanisms that inhibit or excrete antibiotics has great potential for reducing antibiotic doses and rendering resistance mechanisms inert. We demonstrate the synergistic inactivation of a Gram-positive (Staphylococcus aureus) and two Gram-negative (Escherichia coli and Pseudomonas aeruginosa) bacteria by combining 222 30 kV/cm electric pulses (EPs) or 500 20 kV/cm EPs with 300-ns EP duration with various antibiotics with different mechanisms of action is demonstrated. Doses of antibiotics that produced no inactivation in 10 min of exposure in solution with bacteria induced several log reductions under the influence of nanosecond EPs. Combining 2 µg/L or 20 µg/mL of rifampicin with the 30 kV/cm EPs enhanced Staphylococcus aureus inactivation compared with EPs alone, while only a few of the other combinations demonstrated improvement. Combining 2 µg/L or 20 µg/mL of mupirocin or rifampicin with either EP train enhanced E. coli inactivation compared with EPs alone. Combining 2 µg/L or 20 µg/mL of erythromycin or vancomycin with the 30 kV/cm EPs enhanced E. coli inactivation compared with EPs alone. These results indicate that EPs can make Gram-positive antibiotics efficient for inactivating Gram-negative bacteria with future studies required to optimize EP parameters for other antibiotics and Gram-negative bacteria.

Nanosecond pulsed electric field induced proliferation and differentiation of osteoblasts and myoblasts.

Low-intensity electric fields can induce changes in cell differentiation and cytoskeletal stresses that facilitate manipulation of osteoblasts and mesenchymal stem cells; however, the application times (tens of minutes) are of the order of physiological mechanisms, which can complicate treatment consistency. Intense nanosecond pulsed electric fields (nsPEFs) can overcome these challenges by inducing similar stresses on shorter timescales while additionally inducing plasma membrane nanoporation, ion transport and intracellular structure manipulation. This paper shows that treating myoblasts and osteoblasts with five 300 ns PEFs with intensities from 1.5 to 25 kV cm-1 increased proliferation and differentiation. While nsPEFs above 5 kV cm-1 decreased myoblast population growth, 10 and 20 kV cm-1 trains increased myoblast population by approximately fivefold 48 h after exposure when all cell densities were set to the same level after exposure. Three trials of the PEF-treated osteoblasts showed that PEF trains between 2.5 and 10 kV cm-1 induced the greatest population growth compared to the control 48 h after treatment. Trains of nsPEFs between 1.5 and 5 kV cm-1 induced the most nodule formation in osteoblasts, indicating bone formation. These results demonstrate the potential utility for nsPEFs to rapidly modulate stem cells for proliferation and differentiation and motivate future experiments to optimize PEF parameters for in vivo applications.

Synergistic bacterial inactivation by combining antibiotics with nanosecond electric pulses.

Antibiotic resistance mechanisms render current antibiotics ineffective, requiring higher concentrations of existing drugs or the development of more powerful drugs for infection treatment. This study demonstrates the synergistic inactivation of a gram-positive (Staphylococcus aureus) and a gram-negative (Escherichia coli) bacteria by combining either tobramycin or rifampicin with 300-ns electric pulses (EPs). For EPs depositing the same total energy density into the sample with no drug, higher electric fields induced greater inactivation, indicating a threshold for irreversible electroporation at these fields and membrane recovery in between lower intensity EPs. Synergistic inactivation generally increased with increasing drug concentration up to 20 µg/mL compared to strictly EP treatment. Combining even 1/20 of the clinical dose of tobramycin with a train of EPs induced between 2.5 and 3.5 log inactivation after only 10 min of exposure compared to hours to induce inactivation with a clinical dose with no EPs. Similarly, combining a train of EPs with a clinically relevant dose of rifampicin induced 7 to 9 log inactivation over the same time of exposure. These results indicate the promise of combining EPs with antibiotics to rapidly inactivate antibiotic-resistant bacteria in localized treatment areas.