Hello everybody. Welcome to today's webinar hosted by the webinar Vet. My name is Doctor Stephanie Morley.
I'm the president of Vetlin, your sponsor for today's webinar. We are very pleased to bring to you Doctor Dustin Williams to speak to you about bacterial biofilm characteristics, related infection, and possible solutions. I'd like to thank all of you for joining and spending this hour with us, and I'd also like to thank Doctor Williams in advance for sharing his time and expertise with us.
Vetlin has recently launched a new product called the Vetlin pouch, and it happened to be invented by Dr. Williams. It's a local drug delivery device that allows for daily sustain antibiotic dosing directly at the site of infection.
It can be re-dosed daily with the correct antibiotic for up to 30 days and is removed similar to a drain. Dosing locally is the most effective method for treating biofilm-related infections while minimising systemic exposure. So please check us out at www.vetlin.com to learn more.
A few housekeeping items, or excuse me, a few housekeeping items to mention. If anyone has any questions for our speaker, please put them in the Q&A box. For comments or other queries, use the chat box.
And now allow me to introduce you to Doctor Dustin Williams. Doctor Williams is a professor of orthopaedics at the University of Utah, CSO to two companies, Purgo Scientific and Vetlin Advanced Veterinary Devices, and VP Microbiology and new technologies at Kurza Global Global. He received a BS of microbiology from Weber State University and a PhD of biomedical Engineering from the University of Utah.
Doctor Williams' research portfolio is broad and includes mitigating biofilm-related infections, such as those that affect bone fractures or soft tissue wounds, developing medical devices and technologies to address these infections, and investigating bone pathologies. Doctor Williams' most recently advanced technology, the Pergo pouch, received a breakthrough device designation from the FDA in November of 2023. The technology was launched into veterinary medicine in 2024 as the Vetlin pouch, where it currently is healing up dogs, cats, and horses around the world.
Over to you, Dustin. Well, thank you. You, I think you covered pretty much everything there.
Wonderful to be here today. I, I appreciate the opportunity. I always love the chance to talk a bit about biofilms, and it's fun, you know, hearing Stephanie give that overview.
Now we're getting to a point now where, we're, we're starting to, to get on, on top of this problem with biofilm-related infections. So, as she mentioned, I'll go through a little bit about biofilm's characteristics, some infections and kind of hallmarks of the infections of biofilms. And then solutions, what others are working on a little bit, and then I'm gonna give a kind of a, a, you know, a big dose of what we're working on as well with these, pouch technologies.
Brief disclosure, I, I am the original inventor of the technology. I do have financial interest in Vetlin and Purgo. I'm a co-founder of the company's, board member, CSO, and I also am a consultant with 3M.
So I thought I would just kind of lay out a little bit of my professional background. So yeah, as, as Stephanie mentioned, I have a Bachelor's of Science in Microbiology, great microbiology programme at Weaver State in Ogden, Utah, and I, it, it built a great foundation for me to, to really have a drive to try and provide solutions in this area. And then I went to the University of Utah, got my PhD, remarkable programme with biomedical engineering.
And, learned a lot about biomaterials. And so what you'll see is the fusion of these worlds. And I also did a postdoctoral fellowship in orthopaedics.
And again, that's kind of where you'll see the fusion of all of these, these areas and degrees. So, now I have a lot of fun, teaching, preschool kids, every, every once in a while. My alias is Dr.
Double O Darkwing. And I was just recently dubbed, given a new title last week by one of these kids, as the dermatologist. So, that, that was a lot of fun.
I thought I, I'm going to put that in my slides. But I am a professor now in orthopaedics here at the University of Utah. Wonderful department, great colleagues and surgeons that we work with, trying to tackle this problem on many levels.
I did edit this book a few years back on targeting biofilms and translational research and, you know, really trying to hone in on the biofilm problem to see what is it gonna take to solve the issues here. And I've also, of course, just wanted to share some fun. This stems from some technology that I developed, here at the University of Utah.
I'm going to talk about it a little bit later in my talk, but I, I had some fun writing a sci-fi action novel called Return to Duty based on osteointegrated implants, which are the, it's a technology that actually sparked my interest in biofilms and solving biofilm-related infection problems with medical technologies. So, my lab, I run a lab at the University of Utah called the, called the Bone and Biofilm Research Lab, and if we kind of take the core of our research is on biofilms. So we, you'll, you'll hear more later about how, you know, I took the concept of a contaminated wound, happening at the point of injury, being contaminated with biofilms, and, and so they're already in this established biofilm state and then have downstream problems.
And I'll again, I'll talk more about that in a bit. But we've, we've taken this kind of core concept and applied it to many different areas, including traumatic wounds, diabetic foot ulcers, for example, negative pressure wound therapies, which we've loaded with antibiofilm agents and seen some, significant promise there. Heterotopic ossification development is, catalysed a lot by, the presence of biofilms in traumatic injury sites after a, a, a blast wound is created.
We've also developed. Some antimicrobial blue light technologies for osteointegrated implants, found some cool, amazing discoveries wherein we've identified unique antibiotic classes that can kill bacteria in the core of a biofilm, which we'll talk about a little bit later. And then we do, antimicrobial development with pre-surgical approaches to reduce bio burden in skin prior to, surgeries, and then we, Of course, I've developed the, the Vtlin pouch, which is my kind of flagship programme right now, you know, in the market with the veterinary space in Betlin and, marching through the FDA process and the Pergo side.
So, I thought I'd start by, you know, asking, what is bacterial biofilm. But to answer that, we kind of need to go through the history of what, of biofilm discovery and research. So we, it's, you, you almost will never find any talk or presentation given about biofilms without mentioning Anthony van Leeuwenhoek.
So in the 1600s, he hand carved lenses and out of glass and made mini microscopes, right, that he held up to his eye. And observed the microbial world. He saw colour changes in water near his town, and he said, Yeah, there's got to be something in there changing the colour of those waters.
He took scrapings of his skin. He took scrapings of his teeth, and he observed anything he could get his hands on to look. Through them, to look through the microscopic world through these microscopes.
He made over 200 of them with 300x magnification or more and was the first who's kind of credited with documenting these discoveries of these microbes, these, you know, microscopic organisms. And not everybody believed him, of course. Because the germ theory was going on, but that's kind of a different story.
I won't talk about it today, but fascinating story through microbiological discovery of microbes. In the context of biofilms though, in 1943, Claude Zobel, who worked at a marine biologist who worked at the Scripps Institute of Oceanography in San Diego, he's kind of considered the father of marine microbiology. He made a very interesting discovery where he took seawater samples and he used these glass vessels which he would dip all the way down 2 miles down into the ocean.
And collect samples, and he identified there were bacteria everywhere. But the thing he observed that was quite intriguing was that the bacteria in the seawater preferentially adhered to the sides of the, the glass vessels that he used for collection. And so, you know, these planktonic bacteria had the desire to stick to this surface and join a biofilm.
He didn't know it at the time. He didn't really know what he's observing formally yet, and that didn't come for, you know, a couple of decades later. Civil engineers who were working in the space of trickling philtres where they had rocks where they can, they now use polymeric philtres, but the bacteria, algae, protozoa, bacteria, they would adhere to the the philtre surfaces and create biofilms.
And what's interesting, I circled the word biofilm here because a lot of times you'll read through the literature that the word biofilm didn't come out till 1981 or so, but this publication from civil engineers is from 1975, so the term biofilm was out there and they called them these microbial films or biofilms, and, and they were beneficial, right? So they supported decomposition of wastewater material, and so they found that biofilms can be beneficial. But then in 1978, Bill Costerton made a unique discovery where they were looking at, he and the team were looking at surfaces of.
Rocks and streams and and the scanning electron microscope had just come out, which benefited the civil engineers as well as Bill Costerton team in transmission electron microscopes. They were coming into more popular use at this time. And so they used them to look deep into these bacterial films.
And, and, and Costerton and Group found that bacteria use a glycoccalyx or glycocalyx to stick to surfaces. So they had this sticky matrix that they used to adhere to surfaces, create a scaffold, and then build a three-dimensional structure. In his first paper, they kind of, talked about it as a 2D kind of film, if you will.
And it wasn't until sometime later, which we'll talk about here in just a moment, that it was kind of more three dimensional film. So, I just wanted to kind of share, it was wonderful. I had the opportunity in 2011, the year before Bill passed away, to, to host him here in Utah.
And it was, he, he was one of the reasons that I kept going in my PhD work. He was so kind, he came and did a review, spent two days here, reviewed my work, gave me tips, and, and he said, Dustin, your project is one of the top 10 projects I've ever seen. And that gave me motivation.
He probably said that to every student, but I don't know. He was very kind though, and, and it really motivated me to keep going to finish my project. And I, I published a paper with him using biofilms as initial inoculate in animal models of infection.
You'll see that model here in just a bit, and I'll talk more about it. But it also kind of began to help us understand. So over the years, clinicians were starting to see that, you know, materials that were being used in biomedicine, biomaterials, they were, they were being hampered by infection, difficult to treat infections.
And this was one of the first kind of, kind of, pieces of information that helped us see why phagocytes were having a hard time eating these bacteria. And it was because there was this frustrated phagocytosis. And phagocytes have a physical constraint.
They, in essence, cannot eat anything larger than their own body. And so, bacteria are developing into these biofilms. We started getting all these snippets of information showing.
That bacteria developing these biofilms is a survival mechanism in many, many ways. One of them is to resist phagocytosis. So by 1982, Costerton was partnering with many surgeons, clinicians across the world.
He had his fingers, as you watch the history of biofilm discovery, he had his fingers in almost everything, published over 600 papers on the topic. And in 1982, they published an interesting one, which was kind of a hallmark indicator of what of what biofilms can do in biomedicine and medicine in general, and some of the hallmark indicators of how they cause infection or how you can identify a biofilm related infection. So there was a patient, a 56-year-old patient, male, who was working in the garden or whatnot, and, and got an injury on his left elbow, and a few days later, he started complaining about pain in his upper right quadrant, and he had had a pacemaker put in a few years before.
And so he went into the doctor. They for 4 weeks, he was in the hospital. They gave him an antibiotic course for 4 weeks, and it, it resolved.
The infection resolved. Blood cultures for Staphylococcus aureus in this case. When he went into the hospital.
And then he went home. And then a few days later, he comes back to the hospital, and once again, this infection was raging, same infection recurring. And he, and then they, they treated him and he was fine, and then once again, it comes back, and then he goes back into the hospital.
And so, and it wasn't until they removed the pacemaker lead. And they imaged it here and they showed that there were biofilms on that lead that they found this kind of recurrence of infection, this problem, and and Costerton and group, they said, Hey, what's happening here is these biofilms on these medical devices are serving as a nitis or a reservoir of infection. So although systemic antibiotics are knocking them back because these biofilms can release planktonic bacterial cells or bio bioaggregates that come off their surface or their their community.
And, and the, the, the systemic antibiotics might be enough to kill those bacteria, but they're not quite enough to kill those biofilms that are on the surface. And just as another example, by 1984, this was seen in orthopaedic implants. Another patient for 7 years, had an intramedullary nail in the femur and suffered from infection that was treated and kept coming back.
It was a recurring infection for 7 more years. And it wasn't until they removed the intramedullary nail, gave a final course of antibiotics, that that infection finally resolved. And so these were some of the early indicators, hallmark indicators of biofilm-related infection, recurrence, and and it's also kind of a low lying quiescent type of infection.
Think of a cavity on your tooth. That really is a biofilm related infection. Low lying doesn't do a whole lot sometimes to you, doesn't cause a lot of damage, but over time it does.
So biofilms are are kind of these quiescent dormant low lying infective states. The same thing was kind of seen. Many, many clinicians were having problems with Foley catheters and the difficult, difficult to treat infections, and Nicolet al.
They found, this was kind of the seminal paper showing that there was a massive difference in the antibiotic tolerance or susceptibility of bacteria that live in planktonic state or the free floating single cell state versus those that live in this community of bacteria, this biofilm. And Nicolaidal showed that there that bacteria in the biofilm state were 1000 times more tolerant to Tobramycin, in this case, Pseudomonas aeruginosa biofilms. Than their planktonic counterparts.
And so that is where, this is where you hear, you know, quite often, and this has been documented and validated in hundreds and thousands of papers since, is, that, that bacteria in the biofilm are much more tolerant to antibiotics than planktonic bacteria. And then in the 80s, 90s, we started seeing the, the, invention slash formation and optimisation of confocal laser scanning microscopy. And in 1991, We Lawrence et al.
Again, Costerton was involved with this, but Lawrence et al. Published a paper showing that bacteria in these biofilms have 3 dimensional structures, and they actually have water channels that develop throughout these formations and structures within the biofilm community. Phil Stewart and group at the at the centre for Biofilm Engineering have amazing videos.
You can go to their website and check them out, where they're showing antibiotics literally flowing through the system, so they may not even get in touch with the bacteria in the biofilm. And also this started prompting some thoughts that, hey, antibiotics might be getting gummed up in the matrix of the biofilm, and that is true for some, but not for all. So not all antibiotics have diffusion limitations through a biofilm, and they get to the core of the biofilm very quickly.
And the reason I mention that. Is because biofilms tolerate antibiotics, whether they get gummed up in the extracellular matrix or even if they get down into the core of the biofilm, they oftentimes do not have a significant effect. And this has to do with oxygen and nutrient gradient limitations.
Or oxygen and nutrient gradients develop in a biofilm and therefore limit the activity or effect of antibiotics within their community. Because bacteria that are deep down in the core of the biofilm, they kind of are going into a low metabolic state. I'll talk more about that in just a second.
So, well, right now, so we kind of see that deep down in the core of the biofilm, bacteria that are kind of in a low metabolic state. They tolerate antibiotic therapy more so than the bacteria that are on the exterior that are more metabolically active, metabolically rapidly dividing, and they can, the ones on the outside, they kind of swallow up the attack, if you will, of antibiotics, whereas the ones deep down, they hold. So if the ones that are killed off on the outside, you know, they might go away, but this biofilm matures over time toward this resistant fraction, if you will, is kind of seeding or helping the the biofilm to resist and tolerate and maintain its state.
And that kind of goes back to those medical devices that we saw, the pacemaker lead, the intramedullary nail, these biofilms that get onto there, the outside portions may be killed by systemic therapies, systemic antibiotics, because they're more metabolically active on the outside. The ones on the inside tolerate, they serve as a reservoir of infection. And then in 2005, you see the persistor cell theory where you have this, these bacteria and these resistant fractions that are persisting.
And resisting or tolerating antibiotics in this, in this state. So this makes biofilms very difficult to treat. So I kind of like this analogy just to kind of put a picture to this.
And one thing that we've observed in our lab is that different classes of antibiotics work differently against biofilms. And as you think about the biofilm phenotype, the biofilm state, the biofilm morphology, this begins to make sense. So if you have actively dividing cells on the outside of the biofilm and those that are not very actively dividing on the inside, think about a beta lactam.
How is it going to have an effect? If I throw, if I have a biker who's moving super fast down the road and I throw an umbrella into the spokes, I'm going to make that biker go flying off, and that's kind of like antibiotics working against highly metabolically active cells. They're going to have an effect.
They're going to kill them. But if I have a biker who's standing still and I throw an umbrella through those spokes, there might not be very much that happens, so that biker is going to be safe, no harm. So this is kind of like those cells deep in the core of the biofilm.
They're metabolically inactive and therefore antibiotics aren't going to have as much of an effect against them. And then start thinking about those classes of antibiotics. One thing we've observed.
Is that beta lactams, for example, that attack the cell wall of bacteria or attack the mechanisms that develop the cell wall of bacteria, they are going to have a higher effect against bacteria that are more metabolically active, that are dividing and need to create a cell wall. But if you have bacteria, like for example, down in that core, who don't really need to, who aren't dividing rapidly, who aren't really developing a cell wall, a beta lactam might not be very effective against them, but cells have to survive always. They're always producing protein.
So one thing in general that we see is that antibiotics that work more effectively against biofilms are those that target the inner workings of the cell, the inner cogs, the protein synthesis, for example, or DNA replication. So sometimes we say, hey, it might be more effective to try using antibiotics to treat a biofilm related infection with something that targets the inner core, and then even better so, can you combine those antibiotics to kill, you know, both, mechanisms, both molecular pathways or target both molecular pathways in the cell. So that's an observation we've seen.
And that kind of gives you an idea as as behind these, the biofilm history, biofilm discovery, why these things are so recalcitrant, why they're so tolerant, and why they're so difficult to treat in infection states. And, and that, that's what I hope that this has kind of painted the picture for you to begin to see clinically. I hope some of the clinicians, veterinarians are seeing, you know, that, oh yeah, I, I, I've seen this happen.
I've seen recurring infection. I've seen it's difficult to treat, and, and, and hopefully that helps us give some understanding to it. So just to kind of give some key points of a biofilm and some of the characteristics and morphology, biofilms thrive in, on and around us.
They're everywhere in nature, and I'll have a slide on that in just a second. But they preferentially adhere to solid surfaces. They love these avascular surfaces, but also Randy Walcott showed an interesting study that bacteria, they don't even care if it's, you know, avascular, they love to just stick to surfaces, even if there's blood flowing right past them.
They love to adhere to and and create their biofal surfaces for survival mechanisms. They love to stick to things including tissue and medical devices. And that can lower the infectious dose.
Now, I actually just read this week the Eleck and Conan experiments from 1958, and, you know, that's typically the paper that people that people point to saying, look, we can lower the infectious dose of bacteria 10,000fold if we, if a medical device is present. Well, it's quite interesting if you read that paper, Ehle and Conan, 1958, and they put sutures into medical students' arms actually that were contaminated with Staphylococcus aureus. And, and they, they actually only tested this in one person.
And so that one person's outcome is what has derived this whole thought process of a 10,000 fold reduction in infectious dose. So I, I, I take it with a little bit of a grain of salt because I'm not sure that in every single case, if, if an implant or a medical device is present, that it's going to have a lower infectious dose that's 10,000-fold difference, that needs to be fleshed out a little bit more. But the rule of thumb came out that paper where 105 CFU or 100,000 colony forming units per gramme of tissue is needed or used as a diagnosis for infection in tissues.
So also biofilms serve as reservoirs of infection, and they have this recurrence aspect or hallmark that we can see. They have 3D structures. They're very dynamic communities of bacteria, very much not a two dimensional film like was originally considered.
Water channels actively remove toxins or antibiotics away from their system if they want, and they're ever changing cells and aggregates. The system is turning over. They're releasing bioaggregates, planktonic cells, holding on to recalcitrant.
The, the refractive, core, and, they're highly tolerant to antibiotics in part because of the oxygen and nutrient gradients that exist. So developing, this gradient of, of me of metabolism throughout the biofilm. So, what is a bacterial biofilm?
I had some fun. I asked Grock and Gemini and perplexity and AI derivatives. I'm not going to read these definitions for you.
I do love the centre for Biofilm Engineering, and I give them the win on on defining what a biofilm is. And I had some fun with developing my own, you know, definition of a biofilm, including all of this information that we now know about biofilms. What does it tell us?
What does it teach us? And I've come up with this massive definition, right? So I'm trying to include everything in here, and I asked my friend and colleague, who's oftentimes a much simpler than I am, how would you define a biofilm?
And Nick Ashton said they're microbes stuck to a surface and to each other. So that's the simple takeaway of what biofilms are. So just to kind of show you, for, so for about 20 years I've been working with biofilms, published various papers on this.
We have confirmed, right, just validated what's been found over the years, and then also made some very important advancements, I would say in this space. But we've observed the biofilm matrix, this extracellular polysaccharide substance that helps create a 3D, that helps create a scaffold to which the cells can attach and adhere and create these amazing three dimensional structures that come off the surface. We have confirmed that biofilms increase in density the longer you give them to grow, in particular under ideal conditions, but the longer you give them to grow and develop, the more mature they'll become, the denser they'll become, and that density can correlate to the gradients, the oxygen and nutrient gradients that then increase their tolerance to antibiotic therapy.
So, Ideally, you would have a situation where you can start targeting or attacking these biofilms early in a clinical case, if you started seeing something develop rather than late, it might be more difficult to treat them. And of course mechanical removal can be super beneficial, just like every day we're brushing our teeth, we're removing biofilms by brushing our teeth. And so the mechanical removal is is of utmost importance.
And we've developed many, many animal models of infection using biofilms as initial inocula, which I'll talk a little bit more about in just a bit. So back to that kind of continuum of care from a biofilm perspective, you can see we've taken this biofilm theory, the biofilm concept, and we've applied it to many different areas, particularly my lab is fully funded by the Department of defence, and so I've been very interested in developing technologies for the DOD. And this, the Vetlin pouch and the Purgo pouch on the human side, is one of the flagship programmes.
We've received a lot of DOD funding to develop it. It's a major fracture-related infection is a major unmet need in, in the, in the military. So where are bacterial biofilms?
I remember sitting in a class with Dr. Craig Oberg at Weber State University talking about how he helped at the Utah State University creamery. This was years ago.
They kept on having contamination. In one of their food processing plants, and they said, we can't figure out where this is. And my professor's like, oh, I know about biofilms.
I know where they like to stick. They love to stick to O-rings of gaskets in between pipelines. And so he took apart one of their systems and sure enough there was this massive biofilm.
They cleaned that out, contamination was gone. They took care of it for the rest of the time. Biofilms are everywhere.
They're in the food industry, milk production, dairy production. They're in pipelines, oil pipelines all over the world. And what's kind of fascinating about that, just as a fun thought, is oil pipelines are typically treated with glutaraldehyde.
Glutaraldehyde is a fixative, so the same product that they're using to kill the biofilm is actually also locking it in place, which is one of the reasons over years and years that you see the pipeline decreasing with its inner diameter because it keeps on coalescing dead bacterial biofilms on the inside. And they also mineralize these biofilms. And so it becomes a very difficult thing to treat and manage.
Like I mentioned, teeth are teeth along that gum line. Bacterial biofilms love three things. They love a triple interface, air, water, solid interface.
That's where they thrive. It's where they grow. Bill Costerton made fun of bachelors in his book, The Biofilmer, because that's where you find the most biofilms is on the toilet ring around the bowl.
In, in essentially every area. Wherever there is a medical device, you have risk of a biofilm-related infection being present. And that's true for, osteointegrated implants, which I wanted to just share a brief bit about.
So this is What got me going in biofilm related infection research. So I helped, I was brought onto the team as a microbiologist to help develop this technology called osteointegrated implants, which is a titanium post that can be used to, to, surgically implant into the medullary canal of a residual femur, for example. And then the patient will have that post protrude through the skin, and they can dock, a prosthetic or bionic leg onto it, and it helps maintain functionality.
And so just as to give you a little example of how this works, how it goes in, just so you kind of get a, get a an appreciation for how this relates to biofilms. So surgically, what they do is remount that medullary canal, they're going to place that post into there. And then that will protrude out of the skin to which they can dock that abutment or bionic leg.
And a lot of them have a breakaway technology that's quite awesome, so that they break away the leg instead of the post out of the leg first. And then this just kind of gives you an idea. It's directly loading the skeleton with this system.
And this is all, I borrowed this from the Austin Irrigation International. I'm not involved with them, no, no financial interest with this company or anything, but it's been awesome seeing a technology that I was able to help develop. Led to improved quality of life for many, many patients all across the world, and it's still being used now.
So that was really neat. But as you can imagine, wherever I mentioned, I mentioned, wherever there are medical devices, there's going to be the potential for this biofilm problem to exist. And you can imagine having a skin implant interface where this percutaneous exit site of a post can have problems with infection development in that area.
And that was one of the number one problems. Still is one of the risks with this technology. They've dialled it in to where it's, it's a lower risk now, but for many, many decades this was one of the greatest problems that they had to deal with and figure out how do we mitigate infection at that skin implant interface, and that's what I helped with initially.
And then I went into graduate school. I learned about biofilms, learned about a lot of different technologies that are being used to try and manage biofilms. And then my research shifted a bit to fracture-related infection, which is amazing.
If you look at the 1970s and 1980s when open fractures were being defined and classified more formally by Gustilo and Anderson, you find that in their 1984 publication, Rates of fracture of type 3B, open fracture-related infection, were 52%. And if you look at fracture rates, fracture-related infection rates today, they can still be as high as 50%. And in fact, in military situations, we've seen 67% rates of open fracture-related infection, and I just got back a month ago from the Military Health System Research symposium, MHSRS meeting.
Where in, in the Ukraine, they're reporting infection rates and multifocal injuries of 98%. I mean, these soldiers have essentially a 100% chance of getting an infection because of those battle wounds they are, they are suffering. So this is a major unmet need, and the FDA recognises that.
And that's why we've already heard the Pergo pouch got FDA breakthrough device designation. It is meeting a major unmet need. Fracture-related infection.
We need to make progress in this space. Surgeons are doing all they can, all that they have their hands on. They're putting local beads in there, powders in there to try and manage these infections, but what we're currently doing just isn't working well or good enough.
And, so as you can see from the characteristics of the biofilm, just how difficult they are to manage, we can see why. And I'll talk about what we're doing to, to tackle the problem. So, something happens when you have a traumatic injury, or let's just say it's an open fracture.
You have compromised vasculature. The minute that injury occurs, you've opened that leg, you have these, these wounds that are created. You compromise the tubes that take the drugs to where they need to go.
You also have, at the point of injury, biofilm contamination. If I go outside just like the original paper in 1978 from Bill Costerton showed how bacteria stick, they looked at bacterial biofilms in natural ecosystems, and they found that the way that bacteria prefer to dwell, their survival mechanism, their dwelling state, is as a biofilm. So if I go outside and I grab a handful of dirt, for every 1 gramme of nutrient-filled soil and moist soil as well, I'm going to have about 10 billion bacteria per gramme of soil.
And 99% of those are going to be dwelling in a biofilm. And just think about the same thing for a dog bite. Dogs have biofilms all over on their teeth.
They bite another dog, they are inoculating a dog wound with biofilms. Open fractures are contaminated at the point of injury with already established biofilms. So even if I get down to the doctor, I get a shot in my arm or we give a shot to our our pet, those antibiotics are going to distribute systemically.
But they might not be at high enough concentrations to kill the biofilm that inoculated the space, and if I have compromised vasculature, the antibiotic might not even get to where it needs to be because the tubes are busted. So these are two major problems in meeting traumatic and treating traumatic injury wounds, and how, and one of the underlying reasons why bile film related infections, these difficult to treat infections, can take hold. So this is kind of a as a visual of what I've just mentioned.
So if we have intact vasculature, the systemic antibiotics are going to get to where they need to be. We're gonna have these systemic antibiotics, but if the vasculature is compromised, the antibiotics may not get to where they need to be. The biofilms can flourish, these bacteria can flourish and form into these biofilms, or like I said, the biofilm could already be there.
So a lot of the traditional thinking with biofilms is that you have to have planktonic bacteria that seed a space and then become a biofilm. What I propose and what my work is kind of hinged on is the fact that biofilms are already there, contaminating a traumatic injury at the point of injury. So there are several technologies being developed or being used, and then others of course being developed that are attempting to give local high dose antibiotic therapy because that's the way to go.
If we, if our systemic antibiotics are limited, we need to give the antibiotic locally where it needs to be. And so use many clinicians, veterinarians may be on this call who do this routinely. And even if we use lavages and then we develop cements and beads and we put tobramycin powder or vancomycin powder directly into a wound site.
This does give an initial bolus of antibiotic, but it might not be high enough, even so, to kill these biopics. So this Venn diagram shows that we have this clinical need. Here's what we're doing right now.
Local applications. Each, each local application that's being used right now, one of the limitations is that they, they taper so quickly in their payload. If you put antibiotic powders into a wound site, within about 12 to 24 hours, it's gone.
So you've only had a 12 to 24 hour effect. Is that enough to kill a biofilm? Might not be.
Beads can begin to dissolve if those, if they're dissolvable, or polymethylmethacrylate bone cement beads, they only ever release 3 to 7% of their payload, the the polymethylmethacrylate bone bone cements. They only ever release 3 to 7% of their payload, which may not be enough to kill those biofilms, and reservable beads, they're going to dump their payload within about 72 hours, their effective payload. Now, it's not just about releasing and maintaining something above the minimum inhibitory concentration.
The MIC has essentially no relevance to biofilms. They could care less about an MIC. You have to have dozens to hundreds to thousands of times more antibiotic to kill biofilms than just above an MIC level.
So what I, what I wanted to do with my kind of career and life's work was develop something, a technology that could sustain. That could give the high dose therapy, local therapy, and sustain that local high dose therapy. And I will, I, I, I'll talk about that more in just a moment.
I did want to share there are, there are other technologies under development right now trying to address this with local therapies and non-antibiotic therapies. For example, Garwood Medical and many other companies are developing kind of like electrocution, if you will, for a simple term. Where if there's metal in the body, they can stick a needle through the skin, touch the metal, and zap it to sensitise the bacteria, essentially, this is like electroporation, which is very common in microbiology, and sensitise the bacteria to antibiotics, getting in and killing them more effectively.
Selennik Medical is taking the approach where they have a really cool copper coil that they put around a leg, for example, let's say you have a total knee, a total knee replacement that's becoming infected or whatnot. They have this coil that they can put around it. And it creates an alternating magnetic field to warm up the metal, to heat the metal, and then sensitise the bacteria, to kill some of the bacteria and sensitise them also to antibiotics.
And then luminos is creating an antimicrobial blue light technology that's going into systems as well. So 11 technology I worked on for over a decade was active release antimicrobial coatings. The concept is you have a polymer coating that's releasing a drug into the local space and it gets down into kill these biofilms that are on the surface of the bone, and that's kind of the concept behind this.
But once again, these are limited in that whatever you put, whatever antibiotic you put in the coating, you're locked into that antibiotic, and you can't put a whole tonne of antibiotic into a thin film coating, maybe 1 gramme in a large implant. And also, if, if the antibiotic, if the bacteria are not susceptible to the antibiotic, you know, you can't swap it out. It's not, it's a non-refillable system.
So that's kind of some of the limitations that I wanted to address when, when I was developing, thinking about how do I develop a technology to address these limitations. So, I like to give a visual, and I think of this kind of like in different weight classes of boxing, you know, if we think of, of, of lightweight boxing, lightweight class, that might be the lavages, you know, they're very temporary. They do have high antimicrobial concentration, but they might be pretty limited in how much bacteria they can kill, given the amount of time that they're, they're there.
It's not that they're useless, they have their place, but they may not be as effective. And I put kind of powders, beads, and coating into a middleweight class, if you will. And so they, they might give a little bit more extended release, a little bit higher dose over a little bit longer period of time, but they're still just not cutting it when it comes to really moving the needle in this space of bile from implant-related infection.
So I wanted to develop a heavyweight class product. And I think of this where, you know, we need, we need to meet these three criteria. It's, it's all about antibiotics, antibiotics contacting what they need to contact.
Having the concentration needed to kill the biofilm, and also they have to have time. One thing that my lab has found is that you really have to just continuously wallop the biofilm for time to kill it sufficiently. You gotta wear him down, like Rocky did in Rocky 4.
He just kept on letting Drago hit him, hit him, hit him. I think they're in the 15th round, until Rocky finally says, just keep hitting me. He wore him down to a point where he could finally take him out.
I also love Iron Man. I love how he built the Hulkbuster to manage an unmanageable Hulk. We're in the heavyweight class now, and that's, I love to kind of unofficially call the Ventlin pouch the biofilm buster.
We're managing unmanageable infections, really moving the needle in this space. Some of the benefits, I've mentioned limitations, some of the benefits of this technology are it can deliver one or more drugs. You can also swap out the therapy, you could swap out the drug if you need to.
You get a culture that says, I need this, you can swap it out and give it to him. You can also, if you had to, if there were adverse reaction, you could remove this pouch, as we'll show here in just a minute, in, in a matter of seconds if you needed to. It also gives high dose delivery, maximum daily dose is the is how much we ever load into the pouch.
So even if the device failed, it would be safe to the patient. It's also a standalone technology. I wanted something that was standalone, that I could put essentially anywhere in the body and not have it locked into a single implant or medical device.
I could put it wherever I need it. And then because this device is refillable, it sustains local high dose antibiotic therapy. And I have a video I want to share because the video tells 10,000 words of how this technology works.
So I have Progo Scientific, the parent company there, but the rate controlling membrane is kind of the brains behind the device. And what we do to implant this surgically is, let's say we have an infected open fracture wound here, or it could be any kind of infected wound. We use a trocar to externalise the tube, and if the surgeon wants to, they can use sutures to lock that device into place.
And then cut away the trochar, bring in a barbed connector and a needleless connector to create a refilling port. Make sure that there aren't any leaks in the device. If the surgeon wants to, they could do a patency check with some saline, remove that, suture, they'll suture the site closed here, and then bring in a concentrated antibiotic solution.
Fill up that pouch underneath the skin in the wound site, in the treatment site, and then over a 24 hour period, The antibiotic will passively diffuse across the membrane, giving local high dose antibiotic therapy into that space. And then after 24 hours that solution is depleted of antibiotic. We remove it, come in with another concentrated antibiotic solution, and again fill it up.
And over the 24 hours again it's going to give that local high dose therapy, so we're just walloping, walloping, walloping the biofilm in that space. And we can do this for up to 30 days. It's refillable for up to 30 days.
After that 30 day, remove the depleted solution, and when you remove it, it kind of creates a vacuum and it collapses the pouch down to about the same diameter as the as the as the catheter. Pop those suture anchors, and then, without having to do a second surgery, we remove the device out of that exit site. So pretty slick and easy.
We designed it to go in and come out just like a surgical drain. So just, I wanted to just share some of the data we collected and how we collected it to develop this technology. So first of all, I want to share that I wanted to develop something.
That was going to go up against a kill test. I had spent 13 years of my life developing osteointegrated implant technology. And, you know, it's a long time.
It's a long time to dedicate your life to something in your career. And I wanted to know from the get-go, is this technology, this idea that I have even worth working on for 10 or however many years, right? 20 years, I don't know.
But, so I wanted, I, I used my animal model that I developed in graduate school, which is a super challenging animal model, to test the efficacy of, of the, of the Purgo and Wetland pouch. So, to begin, I take, we took sheep and we did some testing in the proximal medial aspect of the sheep tibia. The first thing we do, you'll see this air cannon punch the leg.
The first thing we do is we compromise the soft tissue with a 300 pound punch of air, and you can see that it immediately does compromise the the tissue. We create simulated fractures in that flat part of the medial, proximal medial aspect of the tibia. And then we take simulated fracture fixation plates on which we've grown biofilms.
So once again, we use biofilms as initial inoculant in our animal models of infection. To make them immediately recalcitrant to standard antibiotic therapies, the kill test. And so we grow monomic we've grown monomicrobial, polymicrobial, various different bacteria, MRSA, Staphylococcus aureus, pseudomonas aeruginosa.
And we've, we've tested, clinical standards and the pouch in this, in this model. So we can put beads in there, we can put powders in there. We have, we have, done all of this work.
We've put the pouch in there and we put them right over those simulated fracture fixation plates. And you'll see here as we fill up the pouch, many on this call may have used the pouch, and you'll see it, it fills up right underneath the skin, and it holds that antibiotic solution in it. And then it could begin to deliver its payload passively diffusing across the membrane.
No pumps or batteries required. It's all based on a concentration gradient-driven process. And as a just a brief snapshot, we've tested the Pergo pouch, Betlin pouch, in over 200 sheep to date with a wide variety of antibiotics combinations and compared to another 150 sheep as clinical standards of care comparators.
One of the reasons we got such a large data set was to help kind of, you know, get through that FDA process and really convince ourselves that this technology works. So you'll see right away some of the outcomes. Positive controls of infection, the wounds never even healed.
We use beads in many of these, these technologies. Wounds did not close. Most of the beads fell out because the wound was pushing them out and it was not healing.
IV vancomycin, what was interesting on the IV vancomycin, just like the clinical case I shared earlier, when we were during the period where we were giving IV vancomycin, the infections were kind of healing. But after we stopped the IV therapy, the wound opened up and started becoming more infected. The biofilms serving as a reservoir of infection.
Whereas sheep treated with the Pergo pouch, the suture lines heal, and they're, they're healing up great. And these are some of the subdermal pictures you can see without the pouch. Positive controls obviously massively infected, but whenever we've had the pouch in there, tissues have been very healthy, healing up well.
I can confirm, right, we've done a lot of, of, release kinetics work. That for the entire 30 day period, the Wetland pouch, Pergo pouch delivers its payload without having biofouling problems involved, and we have tremendous amounts of data to support this. It's delivering about 99% of its payload each day.
Microbiologically, we quantify everything we can in that space. We take soft tissue, we take bone, we take all the hardware, we quantify it. The gist of the outcome is every sheep group we've tested that has received a Pergo pouch or Wetlin pouch has has reduced more bacteria than any clinical standard, and it's typically between 10 and 1000 times more bacteria we reduce or kill to a larger degree than clinical standards of care.
And in particular, when we layer. The pouch technology with systemic antibiotics, we see an even greater outcome. We've reduced all bacteria, all detectable amounts of bacteria in our animal models when we layer these therapies.
So as just a quick brief histological overview, we've assessed bones, histologically, those that are treated with the patch, those that are not, and the promising outcome is. That we're seeing that the bone is healing in, for example, the stimulated fracture line by this dark peak regions, osteoids coming in, osteoblasts are healing. And I'm gonna just jump forward to this graph.
So, whenever we've used the Pergo pouch, or Etlin pouch, we retain more bone and we have lower osteoclast counts, osteoclasts being a direct correlator to the severity of infection. So very promising. We're testing this in the spine space as well, tremendous outcomes in that area, but I wanted to close with just two quick case studies, two of, two of my favourite, where this has been used in a dog and a horse with that Vtland pouch.
So there was a dog, this is in the DVM 360 report. You may have seen a tremendous case where the dog was, you know, unfortunately hit by a car, had degloving injury, and the surgeon had tucked up. The, the, the hind limb into the stomach to hope that a skin graft would take, it became infected.
So they put a Vetlin pouch in, treated, and that infection resolved, and that skin graft was able to start being, start grafting and that's just a wonderful outcome. Another one of my favourite, my, my dad passed away 18 years ago. We loved watching the Kentucky Derby.
And so this is one of my kind of favourite stories as well. About 1 year ago, there was a horse. My understanding is that it was a, a, a training horse for the Kentucky Derby, and it, it, bucked the jockey, split open its fetlock ankle joint by trying to jump the barrier.
They went in, put the Vetlin pouch in, cast it for a couple of weeks, 2 weeks, 2500 milligrammes of amicain a day, and that horse, healed up like a champ and even was back on the track a year later. So that's our goal is to get animals, to get humans back on the track, back with their quality of life. I give tremendous thanks to a whole lot of people who's helped get us where we are today.
It does, I mean, you can't do this work without tremendous manpower and help. And also a huge thank you to our funders. The Department of defence has funded us to date to, you know, create the Pergo pouch.
Vetlin, of course, has been kind of the company effort, but it's all been kind of a fusion of getting us to where we can heal animals and humans up. So thank you and I'm happy to answer any questions. Thank you, Dustin.
I appreciate it so much. We do actually have a couple of questions. One was curious if you have any experience with adjunctive hyperbaric oxygen therapy for wound care.
We find that it decreases the healing time for most wounds by an average of 35 to 40%. Oftentimes it is tissue sparing, in extreme cases, limb sparing. Great question.
So, I do not, I personally do not have experience with that. But I, I do not foresee that there would be any problem using hyperbaric treatment with the pouch. So that, that's kind of my take on it.
OK, second question, what are your thoughts on utilising nitric oxide wound pads and or gels? So that's another space I don't have a tonne of experience in. But what do I think about it?
I, I think, go for it. Local therapies, right? I think any kind of local therapy might have a better shot than just systemic therapies alone if you, you think or you are dealing with a bile from related infection.
So, I mean, I've actually had several people approach me about putting, kind of the nitric oxide in, in the pouch, and we haven't tested it yet, but we're excited to see if it might work. I can speak from the Vetlin case study perspective. Those cases we have received back.
We have had several clinicians that have used that kind of multi-modalities of local opportunities for infection reduction, and they've worked well together. So, oh good. Well, that's great.
OK. Next question. What analgesia was used in the sheep studies, the tibial fracture infected implant model?
Yeah, great question. So just as far as overall, treatment throughout the course of the study, so we begin with the fentanyl patch and also buprenorphine. So we do buprenorph or the fentanyl patch, the day before buprenorphine, I believe the day of our veterinarian would be a much better person to answer specifically.
And then we use Arimidil as needed. So we keep that fentanyl on board if they're holding their limb or they're limping, we just keep it on board for, you know, a minimum of 72 hours, and then we extend as needed. And then, Rimmel is typically on board for I believe the 1st 5 days and the model that works pretty well.
We really haven't had to have anything additional, than that, we do, I mean we give. At the end, when we're going to remove the pouch, we remove, the antibiotic solution, and then we'll fill up the pouch with lidocaine solution. I believe it's a 2.5% lidocaine solution.
And then we leave it in there for 15 to 60 minutes, kind of depending. And then, we also do Emla cream around the exit site. And that just helps to kind of numb up the region, reduce any kind of pain, and, and then we'll, pop the sutures in and pull it out.
So, and we, we haven't seen any problems, with that. The pouches slides out very well. Animals do not seem disturbed at all.
They're, and they're walking again, no problem. Great. We're, we're getting some questions in the chat as well as the Q&A, but I'm trying to attach the ones that are related, in order just to keep us on track.
So someone asked a question, I assume this is relating to the sheep studies, but potentially not. It says, are these patients also receiving systemic antibiotics or just Belin pouch high-dose antibiotics? So it's a mix and match.
We've done a lot of studies where we only use, the pouch, and then where we only use systemic antibiotics, so we have the comparison, and then we layer them with systemic and the pouch. In essence, the outcomes that we're seeing is whenever we have systemic antibiotics, and this kind of depends on On the antibiotics. So sometimes you'll have an apples to oranges comparison, or, you know, but apples to apples, but in general, what we're seeing is that sheep treated with systemic antibiotics, let's just say levofloxacin and and rifampin, for example.
They have between 10 and 100 times more bacteria in the site than those that are treated with the Pergo pouch or the Ventolin pouch only. But when you layer those, we reduce the bacteria by 1000 times more. So it's a very synergistic outcome and effect with, you know, the local and systemic therapy, but in general, we're seeing that the pouch alone, either, depending on the systemic antibiotic, either kills as many bacteria or about 100 times more than those with systemic antibiotics.
All right. This one says maybe a silly question, but I'll say there's no silly questions. But isn't treating for 30 days a promoter of antibiotic resistance?
That's a great question. So one of the things I love about the pouch is you can remove it whenever you need to. It doesn't have to be 30 days.
The other consideration with antibiotic resistance, and this is what I was kind of getting at with the MIC level, right? So whenever you're flirting with the minimum inhibitory concentration, I've done, we've done passage studies in this lab, you know, up to wazoo to generate antibiotic resistant organisms. And really it's when you flirt with that line or that concentration that you are more a little bit more concerned about the antibiotic resistance.
So the cool thing about the pouch is we are delivering. Hundreds to thousands of times more antibiotic than the MIC. So our local tissue concentrations are on the milligramme levels immediately around that pouch.
So the risk of resistance development is lowered just because you have such a higher concentration, way above that MIC. And that's, and the nice thing too that I love is that you pull it so quickly, right? After you've given this massive dose, you pull it, whereas if you're using an active release antimicrobial coating, I spent 10 years of my life working on, you're always looking at how long it's releasing along that MIC, and it can be for literally 10 to 20 days, and you would be concerned about antibiotic resistance in that case, because you're flirting with the MIC line for so long, you're given the chance for bacteria.
With the, with the pouch. You're way above the entire time and then you pull it, so you're not tapering and asymptoting to that line of MIC. Yeah, and I, I would add, we actually did a, a webinar with the webinar vet earlier this year with Doctor Grannock, talking about antimicrobial resistance and the judicious use of antibiotics and being an antibiotic steward.
And so that was a webinar vet, webinar, and then we also have one on our website we did with, on the large animal side, speaking to equine practitioners about, antibiotic stewardship. So you might wanna check those out. OK, another question.
What would be the benefit of the Vetlin pouch over a refillable pump delivering the antibiotic to the site? So it depends on what pump we're talking about. So one of the approaches that often is taken is, at least in human clinical care is a Hickman catheter approach, where you put like an, an infusion pump in, into, the site.
So there's a couple of potential issues with it. And a lot of clinicians that I work with here at the University of Utah have experienced this. So whenever you're dumping a very, very high amount of liquid into a space or a site, You're doing a couple of things.
One is you're diluting the immune system. So you're diluting the patient's immune system. If you're dumping, you know, 500 millilitres or 1 litre into that space over a 24 hour period, they're going to have a little bit harder time, healing because you're diluting the healing factors.
The second thing is by putting so much fluid or liquid into that space, you increase the risk of fungal infection as well. So if you're only using antibiotics, you're going to increase the risk of fungal infection. That's one of the, it's called the white sides method.
It's one of the limitations, that does accompany that, that type of approach. And also, whenever you, you're, you're, you're dumping water into that space. Water is going to travel the path of least resistance.
You don't know 100% for sure if it stays in the site that you need it. With the pouch, what we're creating is a diffusive field of antibiotic around it. And so we're giving that local high dose concentration as this diffusive field to potentially increase the opportunity of, of antibiotic to to touch the bacteria that are in the local space as opposed to kind of tracking away down either following the lymphatic clearance or following some channel that it might find.
Thank you. Another alternative therapy, it says, hi, nice lecture. Do you have experience with blue light and chromophor gel like and fovea veterinary lamp?
So I, I have quite a bit of experience with antimicrobial blue light, but not the second component. And I will say the immacobi blue lights intriguing. It's interesting.
It has pretty, actually, you can see my publications on nimicrobi blue light that are out there. It has, it has some effect, in particular, if you, if you're able to get contact with the bacteria. The limitation of immacobiia blue light is its diffusive capability.
It only can penetrate skin or tissue, maybe 1 millimetre. If you're lucky, 2 millimetres, depending on, you know, the state of the skin. But what if the bacteria are 3 millimetres away?
So that's what you, you, you, in essence, can't reach it with it. So it's a very interesting, approach of a non-antibiotic or a non-chemical antimicrobial, but it does have its diffusive capabilities and it's not overly fantastic at reducing biofilm burden because it it's just, it just doesn't have quite the, the punch. OK.
Another, Alternative solution, question, . Sorry, I think I just lost it, but I was talking about the use of manuka honey. Can honey be used in the pouch?
Good question. I mean, it would release potentially, this is just kind of off the, off the cuff here. Potentially release sugar.
And if you can get enough sugar to create an osmotic tension, because basically the only way that honey really kills bacteria is twofold. It's osmotic tension, and it's, honey itself doesn't actually have antimicrobial components in it. It's the antimicrobial peptides from the bees that come into the honey that give it some antimicrobial properties from antimicrobial peptides.
And one of the things that's challenging with honey is you have no idea. How many antimicrobial peptides the bees put in there, what bees produce the best amount of antimicrobial peptides, right? So it's a, it's kind of the wild west with, with honey.
And so I think in theory, you could, I do not know how effective it would be. Because sugar is handled really well by the body. So it might be degraded enzymatically very quickly.
Another question. Thank you for excellent talk. How do you choose antibiotics to use first?
So kind of back to that, what I was mentioning is, you know, what is the problem you're tackling? Do you think you have this biofilm related infection? We typically go with an antibiotic that's going to attack those inner cogs.
So maybe it's, it's a ribosome inhibitor. Maybe it's a DNA replicator inhibitor, because no matter the state, no matter the phenotypic or the morphologic or metabolic state of the bacteria. They're always going to have to be producing something internally, but they might not be turning over their cell wall.
So if I were just kind of going off the cuff, I would say let's try and choose an antibiotic first of all, that's susceptible. If the, if the bacteria is not susceptible to the antibiotic, it's resistant, right? It's not even going to work if it is in the biofilm or not.
So you've got to get a susceptibility profile. But then it would be, OK, what can I use that has the best chance of, of killing the inner cogs, of disrupting the inner cogs, but then. I might go with, if I, if safety is a factor, you know, I might have to go with a beta lactam that could be dosed super high.
In addition, it's, it's more beneficial to use antibiotics that are highly soluble. Because then you can load them up higher. And remember, it's a concentration gradient driven process to create that diffusive field.
So the higher the concentration you can get in the pouch, the better. So, an antibiotic that's more soluble is better. It's one of the reasons we say amicain.
The immunoglycosides are, you know, 100 milligrammes per mL. They're very, very soluble. I'm gonna ask you one more question.
A couple of these, I could probably answer and I'm gonna encourage people for the sake of time to maybe follow up with me via email. But what's the risk of seroma formation with the pouch after its removal? Yeah, so we have seen, you're, you're creating, you know, a void in there.
So there's this space. The risk has been relatively low in the models that we've used. We have not seen complications with it.
But when you remove it, what we do is we use kind of a rolled gauze to compress it. So the typical approach you would use to manage a sero a possible seroma, even after removing a drain, right, you're gonna have that risk because you've created a void in the body. So I think with standard management, they resolve or do not form, you know, quite readily.
So it's been, we've been good. OK. The other questions about, can pet owners do this at home?
Yes, we have plenty of pet owners who have been dosing at home. And we've actually had a lot of great feedback, particularly from cat owners because it is so much easier to dose. So yes, that it can happen.
And also, how do you keep patients from pulling the pouch? Bandaging is the, the best thing, keeping the animal from it. All of us know as veterinarians, it's not always easy to do.
But, standard bandaging practises are gonna be the best thing you can do. Outside of that, if there's any other questions, these have all been amazing. I appreciate it.
You can reach me actually at [email protected] or [email protected], either one.
And if there are any questions I can't answer, I will get them to Dustin and make sure, we get you your answers, but I truly appreciate everyone's, interest and questions. I wanna thank you, Dustin, for your time and of course, thanks to Vetlam for sponsoring. But most importantly, I'd like to thank all of you for taking the time and spending time with us.
We really appreciate it. And the recording of the webinar and the CE certificates will be available in 24 hours' time. So thank you again, everybody, and have a wonderful evening.
