Inactivation of Planktonic Escherichia coli by Focused 1-MHz Ultrasound Pulses with Shocks: Efficacy and Kinetics Upon Volume Scale-Up

Abstract

This study addresses inactivation of E. coli in either 5- or 10-mL volumes, which were 50- to 100-fold greater than used in an earlier study (Brayman et al. 2017). Cells were treated with 1-MHz pulsed high-intensity focused ultrasound (10 cycles, 2-kHz repetition frequency, +65/−12.8 MPa focal pressures). The surviving fraction was assessed by coliform assay, and inactivation demonstrated curvilinear kinetics. The reduction of surviving fraction to 50% required 2.5 or 6 min in 5- or 10-mL samples, respectively. Exposure of 5 mL for 20 min reduced the surviving fraction to ~1%; a similar exposure of 10-mL samples reduced the surviving fraction to ~10%. Surviving cells from 5-min exposures appeared normal under light microscopy, with minimal debris; after 20 min, debris dominated. Transmission electron microscopy images of insonated samples showed some undamaged cells, a few damaged but largely intact cells and comminuted debris. Cellular damage associated with substantive but incomplete levels of inactivation can be variable, ranging from membrane holes tens of nanometers in diameter to nearly complete comminution.

Treating Porcine Abscesses With Histotripsy: A Pilot Study

“…Bacterial inactivation by intense (non-thermal) ultrasound -termed histotripsy -results from inertial cavitation via direct mechanical effects related to the shear forces associated with bubble collapse (Gao, et al 2014) and via sonochemical effects (Joyce, et al 2003). Earlier results obtained with E. coli in suspension are consistent with this general conclusion (Brayman, et al 2017, Brayman, et al 2018. Generally, inactivation rates increased above a pressure threshold (P -~ 7 MPa) and with exposure time.…”

“…Three sample pairs (before and after treatment) were fixed in half-strength Karnovsky's fixative for ultrastructural analysis. Methods similar to those used in Brayman et al were used to process the samples (Brayman, et al 2018). Briefly, samples were pelleted by centrifugation (8,000 g x 20 minutes).…”

“…13b). Both of these observations are likely due to the macroscopic streaming which circulates cells and other particles in the liquid pus through the focus (Brayman, et al 2018). It is also possible that post-treatment aspiration and post euthanasia tissue processing could cause some mixing of the treated and non-treated pus.…”

“…Unlike the procedure for tumors, histotripsy treatment for abscesses should not require that the focus be moved near the abscess capsule. This was demonstrated in an in vitro study which used a fixed-focus system to treat a 10-cc sample by drawing bacteria into the comparatively small focal volume (Brayman, et al 2018).…”

A porcine bimicrobial abscess model for testing interventional treatments

“…Given the morbidity and financial burden of percutaneous drainage, as well as the increasing problem of antibiotic resistance, the risk for secondary infection, wound management and reduced patient quality of life, new methods to treat abscesses are needed [8,9]. As such, appropriate animal models are required in order to perform preclinical studies.…”