Using ultrasound waves to blast bacteria back to the stone age

What was your stupid childhood injury? We’ve all had one — whether that’s a scrape to your knee falling off your bike, a bruise on the head falling off the swings, or a cut to your fingers when playing with scissors. We all have one. These memories are staples of our childhoods which most of us look back on very fondly but the unfortunate truth is, this may not be the case going forward. These light injuries may be the difference between life and death for many generations of young kids to come.

When you get an injury that breaks the skin, you are exposed to a great number of bacteria, most of which your natural immune system and modern medicine can fix. With the emergence of AMR though, these injuries may expose you to a bacteria that we won’t have a treatment for, putting your very life in peril.

AMR stands for antimicrobial resistance which is basically a microbe’s (in our case bacteria) ability to resist drugs/antibiotics, making it harder to treat. As time goes on, microbes become more and more resistant to current medicine making treatment near impossible. This could lead to simple cuts and bruises turning into tragic and fatal injuries…

The Problem With AMR

Our current approach to AMR bacteria is, well, antibiotics. If that doesn’t make sense to you, don’t worry, it really doesn’t. If we continue to tackle one of the most difficult issues of the 21st century with the same thing that’s fueling it, we’re doing nothing but screwing ourselves over. Antibiotics have done us well for a little under a century, since the advent of penicillin in 1928. So what changed between the bacteria in 2021 and the bacteria in 1928? To be frank, a lot.

A study conducted between 2000 and 2015 showed that our consumption of antibiotics increased by 40% in that span, and that’s mainly because of lack of care and supervision.

Our growing usage of antibiotics fuels AMR. The more antibiotics we use, the more mutations occur. This is simply due to natural selection; bacteria that have favorable traits that increase survivability live on to produce new offspring with their resistant traits. Once bacterial cells acquire resistance, exposure to antibiotics kills off non-resistance bacteria, while the antibiotic-resistant bacteria proliferate.

Basically, it’s survival of the fittest…

There are two main ways that bacterial cells can acquire antibiotic resistance. One is through random mutations that occur in the DNA of the cell during replication. The other way that bacteria acquire resistance is through horizontal gene transfer. Gene transfer is when genetic material is transferred from antibiotic-resistant bacteria to other bacterial cells, making them resistant to antibiotics as well.

Resistance mechanisms: mutations in the bacteria either support better protection of the cell or develop mechanisms to diminish the efficacy of the drug

• Protection by modification — mutations make the bacteria insensitive to antibiotic actions
• Protected by population — formation of a protective barrier secreted from the bacteria
• Drug modification— the contents of the antibiotics is altered to reduce the efficacy
• Drug destruction — the drug is cleaved or destroyed by the bacteria
• Drug expulsion — the drug is pumped out of the cell through structures on the membrane

Biofilms

Biofilm formation begins when free-floating microorganisms such as resistant bacteria come in contact with an appropriate surface and begin to put down roots, so to speak. This first step of attachment occurs when the microorganisms produce a gooey substance known as an extracellular polymeric substance (EPS). An EPS is a network of sugars, proteins, and nucleic acids. It enables the microorganisms in a biofilm to stick together.

Attachment is followed by a period of growth. Further layers of microorganisms and EPS build upon the first layers. Ultimately, they create a bulbous and complex 3D structure at the site of infection.

Biofilm formation increases the bacteria’s resistance against the defense mechanisms of the body as well as antimicrobial treatments, thereby promoting chronic infections. They might also work as an environment that accumulates different bacterial species as well as bacterial numbers in certain locations. This has a very negative effect on the body cells around the biofilm. If that wasn’t bad enough, the mere presence of persistent biofilms can influence the local immune system and trigger an inflammatory response or cause/aggravate tissue damage.

The 3 Phased-Process

Now, that’s enough of all that doomsday talk. The real question here is, what are we going to do about this crisis? Here at Blasteria, we’ve engineered a 3-step-repeat process that tracks-kills-cleans infectious biofilms. More important than anything, we don’t use antibiotics! In short, we simply use the power of ultrasound to blast bacteria to the point of destruction. Now although that’s stated simply, each step in the process involves an intricate application of the cutting-edge technology we have at our disposal.

Phase 1: Targeting

Before we can blast and destroy the bacteria, we have to first figure out where it is. In order to do this, we are going to be turning to the use of a contrast agent called microbubbles (MBs).

MBs are small — less than 5 μm! — gas-filled spherical voids that are generally stabilized by a material coating composed of a phospholipid or synthetic polymer. They basically act as our eyes to aid our ultrasound to image the site of infection (we’ll be going into this a little later). They can be manufactured with a variety of contents, structures, and appendages in order depending on the method of treatment. In our operation, we’ll be using tablets containing MBs that will be ingested. Once taken in and released from the tablet, the MBs will preferentially attach to desired cells or infection sites using targeting ligands.

These ligands are basically DNA, RNA, or different types of proteins that bind to specific structures on the bacteria. Think of it as a lock and key. There are a couple of different common ligands that are used to target bacteria in modern medicine:

Surface proteins — these are attached to the membrane of the microbubbles and latch onto the bacterial cell and they include proteins and sugars such as lipopolysaccharides (LPS), peptidoglycan, teichoic acids, outer membrane proteins, oligosaccharides.

Linking biomolecules — this works by linking biomolecules between the microbubble and the surface of the target cell (for example, peptides, pH-responsive polymers) to increase targeting specificity.

Phase 2: Imaging

In our ultrasound examinations, a transducer (or probe) is placed directly on top of the targeted tissue or organ. The operators will scan around potential infection sites until they scan over the site with the MBs attached to it. The transducer will produce a frequency (usually 3.5 MHz) and receive an ultrasound signal at 7 MHz, which will specifically show MBs. It’s basically a flag for our imaging system to understand where to target with treatments.

In terms of picking a frequency for our ultrasound imaging, we have to take a few thinkings into account. The chosen diagnostic ultrasound frequency depends on the depth of the structures to be imaged and the spatial resolution required. Lower frequencies are able to penetrate deeper into tissue but produce images with poorer resolution. On the flip side, higher frequencies will produce images with higher spatial resolution but reduced tissue penetration. That said regardless of the frequency, the microbubbles are highly customizable and can accommodate for the imaging traits that we deem necessary.

Phase 3: HIFU Treatment

So that brings us to the last and most important step — blasting the bacteria to smithereens. But, before you can truly grasp the mechanics behind that, you need to be able to understand a process called inertial cavitation.

Have you ever seen a singer shatter glass with their voice? It’s an interesting scientific phenomenon that also plays a large role in our treatment method. When the frequency of the singer's voice matches that of the glass, it causes the glass to vibrate until it bursts into a bunch of pieces. Likewise, when the resonance frequency of our ultrasound matches that of the microbubbles, it causes the bubbles to expand until it reaches a maximum threshold after which, it implodes. This is inertial cavitation.

Our process uses high-intensity focused ultrasound (HIFU) to target the microbubbles that are attached to the site of infection causing them to rapidly implode one after the other. These implosions cause shockwaves and mechanical force on the bacteria and along with the biofilm as a whole, rupturing and destroying it. This treatment will be done in multiple waves during the same session since the first wave of HIFU blasts likely won’t be enough to kill all of the bacteria at the site of infection.

Our ultrasound is emitted via a machine called a phased array piezoelectric transducer, which enables fine control of focal depth and radius. The transducer looks a little like a bowl, concave in shape, with a bunch of piezoelectric components embedded in its surface area. By specifying which piezoelectric components to fire and at which timings, we can precisely pinpoint focal locations (the points at which the MBs are located). Focal radius ranges from 1mm to 10mm, and 10cm in depth. We can also control the specific frequency of the ultrasound by oscillating the current flow towards the piezoelectric transducers. An increase in voltage translates to an increase in ultrasound pressure and using these parameters, we can finetune our HIFU blasts.

Similar studies…

Various studies have been conducted using similar systems in vitro with promising results. Dr. Timothy Bigelow from Iowa State University has led multiple studies on cavitation and biofilm interactions in vitro, with a variety of different environmental conditions and bacteria types. His work with biofilms planar surfaces yielded very high efficacy after 1–2 minutes of HIFU, with almost 100% of the targeted bacteria destroyed each time.

His research provides a foundation for what our company is aiming to achieve, and with our targeting-based approach to HIFU, we aim to fill the gaps that Dr. Bigelow’s studies were met with.

The Scary Truth

After experiencing what we have with COVID-19 in the last year, it’s pretty clear to see that humanity might not be able to handle a pandemic. Whether that’s from a virus or rampant antibiotic-resistant superbug, we might not have the emotional or financial capabilities to go into a shutdown all over again. That’s why we here at Blasteria, are determined to use our innovative and unconventional ultrasound treatments to ensure that nightmare doesn’t become a reality.

KEY TAKEAWAYS

• Antibiotic resistance has the potential to cause pandemics or turn simple injuries into fatal ones
• The current treatment method of developing more antibiotics is just exacerbating the problem
• Our company, Blasteria, uses ultrasound waves to destroy bacteria with a mechanical force so we can avoid the process of mutation and resistant bacteria