Background and Objectives:Transcranial Doppler (TCD) ultrasound is recommended for measuring cerebral blood flow velocity to guide therapeutic intervention in patients with cerebrovascular diseases. However, the widespread use of TCD is hampered by a shortage of trained practitioners. This study evaluated the efficacy of a simulator-based curriculum in teaching novices the foundational skill to perform TCD insonation. Methods:We developed a curriculum on a novel simulator that incorporates interactive practice with real-time feedback. We assessed change in the skill levels of medical and premedical students without previous exposure to TCD from pretraining and posttraining tests. Psychomotor skill was assessed based on accuracy in insonating the middle cerebral, anterior cerebral, ophthalmic, basilar, and vertebral arteries. Cognitive skill (knowledge) was assessed using multiple-choice questions (MCQs). Results:Training significantly improved skill: the correct insonation rate increased from 16 ± 24% (95% CI 9%-23%) on the pretest to 61 ± 34% (95% CI 52%-70%) on the posttest (N = 56, p < 0.0001, Cohen d = 1.5). First-attempt accuracy improved from 15 ± 23% (95% CI 9%-21%) to 49 ± 31% (95% CI 41%-57%) (N = 56, p < 0.0001, Cohen d = 0.9). Knowledge increased markedly, with MCQs answered correctly rising from 7 ± 26% (95% CI 0%-14%) to 64 ± 19% (95% CI 59%-69%) (p < 0.0001, Cohen d = 2.5). The course, including pre-test and posttests, was completed in 2.6 ± 1.0 hours. Discussion:Simulator-based training rapidly improved the ability of novices to locate intracranial arteries for measurement of blood flow velocity. The data support the employment of simulator-based training as a foundational step to help alleviate the shortage of trained providers. The simulator's self-directed practice allows for scalable, standardized training.

Methicillin-resistant Staphylococcus aureus (MRSA) is a gram-positive sphere-shaped bacterium and a common cause of abscesses. Focused ultrasound-induced cavitation can kill bacteria and potentially treat abscesses non-invasively by reducing bacterial populations. As an initial study to evaluate this approach, 10 mL MRSA suspensions were used as a simplified abscess model and treated in polycarbonate vials coupled to a focused ultrasound transducer (H-161, Sonic Concepts, Bothell, WA, F# 0.9). The pulse parameters were 1.2 MHz frequency with 20 μs pulse duration at 2% duty cycle, and 40-min treatments were conducted. The coupling water was heated to 37 °C, with input power adjusted to raise and maintain the sample temperature to 50 °C throughout the treatment. The cavitation dose was determined from PCD broadband noise using a co-axially aligned imaging probe (P4-2), while thermal dose (CEM50) was determined from thermocouple temperature measurements. Control experiments involving heating to 50 °C without ultrasound were also conducted. Results showed logarithmic relationship between MRSA inactivation and thermal dose, with cavitation contributing insignificantly to the effect. To understand this result in contrast to statistically significant inactivation of E. coli (gram-negative rod-shaped) by cavitation, we are investigating the effect of bacterial structural features on susceptibility to focused ultrasound-induced cavitation.

The Circle of Willis (CoW) is a ring-like network of blood vessels that perfuses the brain. Flow in the collateral pathways that connect major arterial inputs in the CoW change dynamically in response to vessel narrowing or occlusion. Vasospasm is an involuntary constriction of blood vessels following subarachnoid hemorrhage (SAH), which can lead to stroke. This study investigated interactions between localization of vasospasm in the CoW, vasospasm severity, anatomical variations, and changes in collateral flow directions. Patient-specific computational fluid dynamics (CFD) simulations were created for 25 vasospasm patients. Computed tomographic angiography scans were segmented capturing the anatomical variation and stenosis due to vasospasm. Transcranial Doppler ultrasound measurements of velocity were used to define boundary conditions. Digital subtraction angiography was analyzed to determine the directions and magnitudes of collateral flows as well as vasospasm severity in each vessel. Percent changes in resistance and viscous dissipation were analyzed to quantify vasospasm severity and localization of vasospasm in a specific region of the CoW. Angiographic severity correlated well with percent changes in resistance and viscous dissipation across all cerebral vessels. Changes in flow direction were observed in collateral pathways of some patients with localized vasospasm, while no significant changes in flow direction were observed in others. CFD simulations can be leveraged to quantify the localization and severity of vasospasm in SAH patients. These factors as well as anatomical variation may lead to changes in collateral flow directions. Future work could relate localization and vasospasm severity to clinical outcomes like the development of infarct.

Bacterial loads can be effectively reduced with cavitation-mediated focused ultrasound, or histotripsy. Our previous in vitro work with Escherichia coli (E. coli) established strong trends of bactericide with increasing peak negative pressure amplitude, pulse length, PRF, and treatment time. The current study correlates bactericide as a function of the histotripsy bubble cloud size produced for these treatment parameters at several frequencies. Histotripsy was applied to E. coli suspensions in 10-ml sample vials at 810 kHz, 1.2 MHz, or 3.25 MHz for 40 min. Separately, cavitation was recorded using a Photron Fastrax high-speed camera for acoustic parameters equivalent to those used in the E. coli studies. The images were used to quantify the maximum size of each bubble cloud, with the assumption that the cloud was axially symmetric around the propagation direction. A strong linear relationship exists between log kill versus cloud size (R2 = 0.96). Remarkably, across all variables studied, the log-reduction in viable bacteria exhibited a direct proportional relationship with the dimensions of the bubble cloud. This strong correlation between bacterial reduction and cavitation bubble cloud size could have significance for clinical applications where bacteria cannot easily be sampled. [Work supported by NIH R01AR080120 and R01EB023910.]

Blood velocities measured by Transcranial Doppler (TCD) are dependent on the angle between the incident ultrasound beam and the direction of blood flow (known as the Doppler angle). However, when TCD examinations are performed without imaging the Doppler angle for each vessel segment is not known. We have measured Doppler angles in the basal cerebral arteries examined with TCD using three-dimensional (3D) vessel models generated from computed tomography angiography (CTA) scans. This approach produces angle statistics that are not accessible during non-imaging TCD studies. We created 3D models of the basal cerebral arteries for 24 vasospasm patients. Standard acoustic windows were mapped to the specific anatomy of each patient. Virtual ultrasound transmit beams were generated that originated from the acoustic window and intersected the centerline of each arterial segment. Doppler angle measurements were calculated and compiled for each vessel segment. Doppler angles were smallest for the middle cerebral artery M1 segment (median 24.6°) and ophthalmic artery (median 25.0°), and largest for the anterior cerebral artery A2 segment (median 76.4°) and posterior cerebral artery P2 segment (median 75.8°). The ophthalmic artery had the highest proportion of Doppler angles that were less than 60° (99

Large abscesses are walled-off collections of pus and bacteria that often do not respond to antibiotic therapy. Standard of care involves percutaneous placement of indwelling catheter(s) for drainage, a long and uncomfortable process with high rehospitalization rates. The long-term goal of this work is to develop therapeutic ultrasound approaches to eradicate bacteria within abscesses as a noninvasive therapeutic alternative. Inertial cavitation induced by short pulses of focused ultrasound (histotripsy) is known to generate lethal mechanical damage in bacteria. Prior studies with Escherichia coli (E. coli) in suspension demonstrated that bactericidal effects increase with increasing peak negative amplitude, treatment time and duty cycle. The current study investigated correlates of bactericidal activity with histotripsy cavitation cloud size. Histotripsy was applied to E. coli suspensions in 10-mL sample vials at 810 kHz, 1.2 MHz, or 3.25 MHz for 40 minutes. The cavitation activity in the sample vials was separately observed with high-speed photography. The cavitation cloud area was quantified from those images. A linear relationship was observed between bacterial inactivation and cavitation cloud size (R-2 = 0.96), regardless of the acoustic parameters (specifically frequency, pulse duration and power) used to produce the cloud.

Introduction We use a novel 4-dimensional (4D) volumetric M-mode (VMM) ultrasound (US) technique to assess intravascular volume by monitoring the inferior vena cava (IVC). The VMM method expands the spatial coverage of standard M-mode scanning (depth vs time) by including lateral image direction and adds transducer tilt to cover the region surrounding the IVC. Current ultrasound methods for volume assessment suffer from intra- and inter-operator variability. The VMM technique aims to address these limitations, aiding in early detection of hypovolemia/hemorrhage and guiding resuscitation. Methods/Technical Approach The 4D VMM technique was used on animals that underwent a swine hemorrhagic shock protocol with fluid resuscitation. 2D ultrasound images obtained were formatted in a 3D volume to capture changes over time in vessel size with respiration and volume status. Planes were then extracted from the 3D volume at multiple lateral locations to find and track the IVC. The vessel walls were manually traced on vertical planes (depth vs. time) to determine mean IVC diameter and IVC collapsibility at each measurement time point in the shock/resuscitation protocol. Planes at constant depth (lateral vs. time) were selected to extract respiratory and cardiac cycle data. Results Mean IVC diameter in the baseline phase was significantly greater than in the hemorrhage phase (p = 0.020). There was no significant different in mean IVC diameter between baseline and resuscitation (p = 0.064) or hemorrhage and resuscitation phases (p = 0.531). There was no statistically significant difference in mean collapsibility or ΔIVC diameter between protocol phases. The 4D VMM technique effectively measured heart and respiratory rates, consistent with monitored vitals. Conclusion 4D VMM identified IVC changes corresponding to blood loss and resuscitation during hemorrhagic shock as well as heart/respiratory rates. This innovative approach holds promise in reducing operator variability and providing actionable information during treatment of shock.

Infected abscesses are defined as localized collections of infected purulent material delineated by a capsule composed of granulation and connective tissue. They can affect any part of the body. Current treatment typically involves antibiotics with either long-term catheter drainage or incision and washout. Given the ubiquitous nature of bacteria and the serious threat of multi-drug resistant strains, abscesses have become a persistent global healthcare problem. Using histotripsy, a potential new noninvasive treatment modality, we have evaluated bacterial inactivation of Staphylococcus aureus (S. aureus) and Escherichia coli (E. coli) microbes. We have determined the peak negative pressure threshold for inactivation of E. coli in solution for cavitation histotripsy (low cycles, high pulse repetition rate) and boiling histotripsy (high cycles, high pulse repetition rate) for different treatment times. The effect of transducer frequency on bacterial inactivation has also been evaluated. The optimal regimen was used to evaluate the S. aureus inactivation. Bacterial inactivation has also been evaluated in an abscess phantom model. Custom transducers have been developed and evaluated in an animal model for abscesses. [Work supported in part by NIH NIBIB #R01EB019365 and NIAID #R01AR080120.]

The Circle of Willis (CoW) is a redundant network of blood vessels that perfuses the brain. The ringlike anatomy mitigates the negative effects of stroke by activating collateral pathways that help maintain physiological perfusion. Previous studies have investigated the activation of these pathways during embolic stroke and internal carotid artery occlusion. However, the role of collateral pathways during cerebral vasospasm-an involuntary constriction of blood vessels after subarachnoid hemorrhage-is not well-documented. This study presents a novel technique to create patient-specific computational fluid dynamics (CFD) simulations of the Circle of Willis before and during vasospasm. Computed tomographic angiography (CTA) scans are segmented to model the vasculature, and transcranial Doppler ultrasound (TCD) measurements of blood flow velocity are applied as boundary conditions. Bayesian analysis leverages information about the uncertainty in the measurements of vessel diameters and velocities to find an optimized parameter set that satisfies mass conservation and that is applied in the final simulation. With this optimized parameter set, the diameters, velocities, and flow rates fall within typical literature values. Virtual angiograms modeled using passive scalar transport agree closely with clinical angiography. A sensitivity analysis quantifies the changes in collateral flow rates with respect to changes in the inlet and outlet flow rates. This analysis can be applied in the future to a cohort of patients to investigate the relationship between the locations and severities of vasospasm, the patient-to-patient anatomical variability in the Circle of Willis, and the activation of collateral pathways.

Upper urinary tract stones are increasingly prevalent in pet cats and are difficult to manage. Surgical procedures to address obstructing ureteroliths have short- and long-term complications, and medical therapies (e.g., fluid diuresis and smooth muscle relaxants) are infrequently effective. Burst wave lithotripsy is a non-invasive, ultrasound-guided, handheld focused ultrasound technology to disintegrate urinary stones, which is now undergoing human clinical trials in awake unanesthetized subjects. In this study, we designed and performed in vitro testing of a modified burst wave lithotripsy system to noninvasively fragment stones in cats. The design accounted for differences in anatomic scale, acoustic window, skin-to-stone depth, and stone size. Prototypes were fabricated and tested in a benchtop model using 35 natural calcium oxalate monohydrate stones from cats. In an initial experiment, burst wave lithotripsy was performed using peak ultrasound pressures of 7.3 (n = 10), 8.0 (n = 5), or 8.9 MPa (n = 10) for up to 30 min. Fourteen of 25 stones fragmented to < 1 mm within the 30 min. In a second experiment, burst wave lithotripsy was performed using a second transducer and peak ultrasound pressure of 8.0 MPa (n = 10) for up to 50 min. In the second experiment, 9 of 10 stones fragmented to < 1 mm within the 50 min. Across both experiments, an average of 73–97

OBJECTIVE:Bacterial loads can be effectively reduced using cavitation-mediated focused ultrasound, or histotripsy. In this study, gram-negative bacteria (Escherichia coli) in suspension were used as model bacteria to evaluate the effectiveness of two regimens of histotripsy treatments: cavitation histotripsy (CH) and boiling histotripsy (BH).METHODS:Ten-milliliter volumes of Escherichia coli were treated at different negative focal pressure amplitudes and over time periods up to 40 min. Cavitation activity was characterized with coaxial passive cavitation detection (PCD) and synchronized plane wave B-mode imaging.RESULTS:CH treatments exhibited a threshold behavior that was consistent with PCD metrics of cavitation. Above the threshold, bacterial inactivation followed a monotonically increasing log-linear relationship that indicated an exponential inactivation rate. BH exhibited no threshold, but instead followed a different monotonically increasing inactivation rate. Inactivation rates were larger for BH at or below the CH threshold, and larger for CH substantially above the threshold. CH studies performed at different pulse lengths at the same duty cycle had similar inactivation rates, suggesting that at any given pressure amplitude, the "on time" was the most important variable for inactivating E. coli. The maximum inactivation was produced by CH at the highest pressure amplitudes used, leading to a log reduction >4.2 for a 40 min treatment.CONCLUSION:The results of this study suggest that both CH and BH can be used to inactivate E. coli in suspension, with the optimal regimen depending on the attainable peak negative focal pressure at the target.

Obesity is a common finding and a major pathogenetic factor in obstructive sleep apnea (OSA) in adults. To understand the mechanisms behind this, the present study investigated the tissue properties and respiratory kinematics of the tongue base and soft palate in the obese OSA minipig model. In 4 verified obese/OSA and 3 non-obese/non-OSA control minipigs, MRI fat-weighted images, ultrasound elastography (USE), and sleep video-fluoroscopy (SVF) were performed to quantify the fat composition, tissue stiffness, and respiratory kinematics of the tongue base and soft palate during sedated sleep. The results indicated that the fat composition gradually increased from the rostral to caudal tongue base, particularly in the posterior 1/3 of the tongue base, regardless of the presence of obesity and OSA. However, this trend was not seen in the soft palate and pharyngeal wall. The pharyngeal wall presented the highest fat composition as compared with the tongue base and soft palate. Overall, obese OSA minipigs showed stiffer tongue tissue than the controls, particularly in the rostral region of the tongue in obese Yucatan minipigs. The respiratory moving ranges of the soft palate were greater in both dorsal-ventral and rostral-caudal directions and during both respiratory and expiratory phases in OSA obese than control minipigs, and the largest moving ranges were seen in OSA obese Panepinto minipigs. The moving range of the tongue base was significantly smaller. These results suggest more fat infiltration in the caudal region of the tongue base regardless of the presence of obesity and/or OSA. The greater tissue stiffness of the tongue in obese OSA minipigs may result from altered neuromuscular drive.

•Automated analysis of transcranial ultrasound images identifies brain tissue.•A K-means clustering algorithm and anatomy-based processing create an ultrasound image mask for brain tissue.•The clustering algorithm incorporates both ultrasound echo strength and tissue displacement.•The ultrasound brain masks are compared to a computed tomography brain mask.

Abscesses are infected walled-off collections of pus and bacteria. They can affect any part of the body. Current treatment is typically limited to antibiotics, catheter drainage and hospitalization, or surgical wash-out. Although bacteria can develop drug resistance, they remain susceptible to thermal and mechanical damage. Histotripsy generates localized cavitation without heating and represents a potential new noninvasive treatment modality. Successful histotripsy treatments were demonstrated with gram-negative E-coli in both in vitro and in vivo studies [Brayman et al., UMB 44, 1996–2008 (2018); Matula et al., UMB 47, 603–619 (2020)]. Interestingly, preliminary data showed that boiling histotripsy was less effective than cavitation histotripsy in killing bacteria. In this talk, we will describe more in-depth studies that compare cavitation histotripsy (low number of cycles, high-pulse repetition rate) with boiling histotripsy (high number of cycles, low-pulse repetition rate) on inactivation rates of microbes. [Work funded in part by NIH NIBIB No. R01EB019365.]

Phase aberration induced by soft tissue inhomogeneities often complicates high-intensity focused ultrasound (HIFU) therapies by distorting the field and, previously, we designed and fabricated a bilayer gel phantom to reproducibly mimic that effect. A surface pattern containing size scales relevant to inhomogeneities of a porcine body wall was introduced between gel materials with fat- and muscle-like acoustic properties-ballistic and polyvinyl alcohol gels. Here, the phantom design was refined to achieve relevant values of ultrasound absorption and scattering and make it more robust, facilitating frequent handling and use in various experimental arrangements. The fidelity of the interfacial surface of the fabricated phantom to the design was confirmed by three-dimensional ultrasound imaging. The HIFU field distortions-displacement of the focus, enlargement of the focal region, and reduction of focal pressure-produced by the phantom were characterized using hydrophone measurements with a 1.5 MHz 256-element HIFU array and found to be similar to those induced by an ex vivo porcine body wall. A phase correction approach was used to mitigate the aberration effect on nonlinear focal waveforms and enable boiling histotripsy treatments through the phantom or body wall. The refined phantom represents a practical tool to explore HIFU therapy systems capabilities.

Infected abscesses are walled-off collections of pus and bacteria. They are a common sequela of complications in the setting of surgery, trauma, systemic infections, and other disease states. Current treatment is typically limited to antibiotics with long-term catheter drainage, or surgical wash-out when inaccessible to percutaneous drainage or unresponsive to initial care efforts. Antibiotic resistance is also a growing concern. Although bacteria can develop drug resistance, they remain susceptible to thermal and mechanical damage. In particular, short pulses of focused ultrasound ( i . e ., histotripsy) generate mechanical damage through localized cavitation, representing a potential new paradigm for treating abscesses non-invasively, without the need for long-term catheterization and antibiotics. In this pilot study, boiling and cavitation histotripsy treatments were applied to subcutaneous and intramuscular abscesses developed in a novel porcine model. Ultrasound imaging was used to evaluate abscess maturity, for treatment monitoring and assessment of post-treatment outcomes. Disinfection was quantified by counting bacteria colonies from samples aspirated before and after treatment. Histopathological evaluation of the abscesses was performed to identify changes resulting from histotripsy treatment and potential collateral damage. Cavitation histotripsy was more successful in reducing the bacterial load while having a smaller treatment volume compared with boiling histotripsy. The results of this pilot study suggest focused ultrasound may lead to a technology for in situ treatment of acoustically accessible abscesses.

Aberrations induced by soft tissue inhomogeneities distort acoustic fields and often complicate HIFU therapies. An aberration-generating phantom was designed and fabricated to mimic the acoustic effects of a porcine body wall. A rippled surface pattern replicating inhomogeneities of size scales relevant to the porcine body wall was transferred to a large phantom made of fat-like and muscle-like gel materials. The acoustic distortions produced by the phantom and an ex vivo porcine body wall were characterized by hydrophone measurements with a 256-element 1.5 MHz focused ultrasound array (diameter = 144 mm, F# = 0.83). Insertion of the phantom generated large distortions for both linear and nonlinear beam focusing including displacement of the focus (up to 0.3 mm transversely), enlargement of the focal region, and reduction in focal pressures due to aberration (up to 2.5-fold peak positive pressure reduction). The phantom and porcine body wall produced similar levels of aberration and attenuation effects. An aberration correction approach was leveraged to isolate the quantitative effect of aberration and to restore shocks in nonlinear waveforms in the focus. Boiling histotripsy treatments through the phantom or body wall were subsequently demonstrated in a hematoma sample. [This work was supported by NIH R01EB7643 and R01EB25187, and RSF 20-12-00145.]

Abscesses are walled-off collections of infected fluids that often develop as complications in the setting of surgery and trauma. Treatment is usually limited to percutaneous catheterization with a course of antibiotics. As an alternative to current treatment strategies, a histotripsy approach was developed and tested in a novel porcine animal model. The goal of this article is to use advanced ultrasound imaging modes to extract sonographic features associated with the progression of abscess development in a porcine model. Intramuscular or subcutaneous injections of a bi-microbial bacteria mixture plus dextran particles as an irritant led to identifiable abscesses over a 2 to 3 wk period. Selected abscesses were imaged at least weekly with B-mode, 3-D B-mode, shear-wave elastography and plane-wave Doppler imaging. Mature abscesses were characterized by a well-defined core of varying echogenicity surrounded by a hypoechoic capsule that was highly vascularized on Doppler imaging. 3-D imaging demonstrated the natural history of abscess morphology, with the abscess becoming less complex in shape and increasing in volume. Furthermore, shear-wave elastography demonstrated variations in stiffness as phlegmon becomes abscess and then liquefies, over time. These ultrasound features potentially provide biomarkers to aid in selection of treatment strategies for abscesses.

Burst wave lithotripsy (BWL) is a new non-invasive treatment for comminuting kidney stones. Although exposure parameters used in current clinical trials have produced negligible renal injury in preclinical pig studies, it remains of interest to understand how quickly injury accumulates, should it occur. BWL treatments lasting ≤30 min were delivered to 36 sites in 20 kidneys of 11 pigs. Treatments at 350 kHz utilized pulses with supra-clinical 8–9 MPa peak negative pressure, 20 cycles/pulse, and a pulse repetition frequency (PRF) of 10 or 40 Hz. Real-time monitoring was performed with B-mode and Doppler ultrasound to identify the presence and duration of cavitation activity, which has been correlated with the onset of injury. After treatment, kidneys were excised for magnetic resonance imaging (MRI) within 3 h of euthanasia. MRI was used to quantify the volume of any hemorrhagic injury. For sites with injury at 40 Hz PRF, the average (n = 5) injury volume increased with the number of BWL pulses (72, 720, 7200, or 72 000 pulses) delivered with cavitation. Notably, the rate of lesion growth slowed above 7200 pulses while all lesion volumes were smaller for equivalent cavitation exposures at 10 Hz PRF. [Funding support by NIH P01-DK043881, K01-DK104854.]