Donna Paznar joins Absolute Gene-ius to discuss her research on Streptococcus pneumoniae, including how children can serve as hidden reservoirs and why serotype tracking matters for understanding bacterial spread. She also shares how molecular tools such as qPCR, nested PCR, and sequencing support surveillance research, along with the tick-borne disease story that first drew her into science.
Some science stories start with a textbook. Donna Paznar’s started with a tick.
In this episode of Absolute Gene-ius, Donna Paznar, head of Molecular Infection Diagnostics at the Medical University of Vienna, discusses her work studying Streptococcus pneumoniae and its impact across the Austrian population. She explains why this respiratory bacterium can be especially dangerous for infants and older adults, how children may act as asymptomatic reservoirs, and why tracking more than 100 bacterial serotypes is both important and technically challenging. Donna also walks through the molecular methods behind her work, including qPCR for identifying positive samples, nested PCR for targeting more specific DNA regions, and sequencing to help confirm serotypes. Along the way, she highlights the complexity of surveillance research and the goal of developing more efficient multiplex qPCR approaches.
The career corner takes a fun turn into ticks, workplace cliques, qPCR mistakes, and a proud sleuthing moment involving a rare tick-borne pathogen. Donna’s advice is simple and useful: try opportunities, expect failure, and stay open-minded.
Lisa Crawford 00:00
They have horse girls, and they also have tick girls.
Donna Paznar 00:02
They do; they really do well. Actually, they go hand in hand, Lisa, because usually you get your ticks from riding horses, actually, so
Lisa Crawford 00:10
Pathway.
Donna Paznar 00:11
Yeah, it is big time. Lifestyle.
Jordan Ruggieri 00:25
Welcome to Absolute Gene-ius, a podcast series from Thermo Fisher Scientific. I'm Jordan Ruggieri,
Lisa Crawford 00:31
and I'm Lisa Crawford. Today we're joined by Donna Paznar, whose research looks at how pneumonia-causing bacteria spread and affect communities.
Jordan Ruggieri 00:38
Donna shares how children can act as hidden carriers, why tracking infections matters and how her journey in science began with a fascination for ticks.
Lisa Crawford 00:47
Safe to say, she got bitten by the science bug. From early curiosity to tackling respiratory infections, today her story is full of insight and inspiration.
Jordan Ruggieri 00:57
We hope you enjoy the conversation.
Jordan Ruggieri 01:02
Donna. Thank you so much for joining us for today's episode of the Absolute Gen-ius. We are thrilled to have you, and really excited to dive into your science and your career, and I'm just really grateful for your time. Can you give a little bit of background for our listeners? What are you working on, and a little bit about what your research is on?
Donna Paznar 01:23
So, currently my research is on Streptococcus pneumonia and its impact throughout the current Austrian population. I've previously worked with sexually transmitted diseases, so I've switched from the STI world now into the respiratory world, in terms of bacteria. So that's currently what I'm, my, my study species at the moment.
Jordan Ruggieri 01:47
What is Streptococcus pneumonia? What is, what is that, and how does it impact people and the world?
Donna Paznar 01:55
Streptococcus pneumonia is, it's a bacteria that typically infects the respiratory tract of any age, but it's very, very dangerous, or can be life-threatening in the younger and the older population. So, in infancy, and then, of course, the geriatric population, it becomes quite the, quite a very severe disease in the end. Since we've had all of these kind of precautionary measures with masks and things like that, we definitely saw a decline in things in these respiratory illnesses, but since then, not only have immune responses been declining for certain bacteria like Streptococcus pneumonia, but it has also been a huge, huge issue for us specifically with the fact that now there's very, very colonized strains of this bacteria. So, the issue that we're facing right now is more so the fact that children, specifically, are these kind of really, really special reservoirs for the bacteria itself, and they don't actually show any signs or symptoms, and they're not seriously affected by the bacteria, but they're carriers, and then so that is then progressing further and further. And with myself right now, what I'm specifically working on is actually tracking the serotypes of the of the bacteria itself. So there's over 100, and so what we are doing is not only working with those that are showing, so anyone that comes with symptoms, signs of pneumonia, of course they're getting tested, but we're also testing children who are, who are thought to be reservoirs, just with regular, with regular testing, and getting these, these swabs from them, and then tracing and trying to track, okay, where are these serotypes kind of spreading throughout Austria.
Jordan Ruggieri 03:47
Sounds like there's a couple aspects when it comes to your research. We're not necessarily talking the diagnostic, the clinic side of it, where a subject comes in and is looking for some sort of treatment, but post that, looking at research that can help inform some of the decisions, or inform some of the future directions for, for this particular, you know, research, and this particular bacteria. Is that a correct statement? You're it's you're looking at that research post that the actual positive or negative test?
Donna Paznar 04:24
Exactly, so in my case, with my research, I get samples post whatever situation they may, the subject may have presented in a clinic with. So aside from that, aside from any sort of diagnostic or clinical setting, my research specifically focuses on everything post that. And so I do, I have the samples, then, and whatever clinically, diagnostically the patient has before that is of no interest to me, really. So I'm just looking at everything after it's already been in that setting, so to say.
Jordan Ruggieri 05:07
Do all the different serotypes cause pneumonia? And I'm assuming because of the name of the of the bacteria, it's pneumonia we're talking about here? Is that, is that correct?
Donna Paznar 05:17
It's one of several pneumonias, so
Jordan Ruggieri 05:19
Okay.
Donna Paznar 05:19
That's another thing. Streptococcus pneumonia is one that's more associated with septic pneumonia. So, what it's commonly seen, we do see it a lot in, in patients that are presenting with more so these very severe cases. The walking pneumonia, which is also something that I'm a lot of people are aware of, and it's also going around everywhere at the moment, not only just here but worldwide, that's the Mycoplasma pneumonia. So that's a different species that does also present with the respiratory infection, the bacteria that resides typically in the cilia and in the lungs, and then Streptococcus pneumonia can actually become systemic, where it eventually affects all organs, and then that's when we have these septic patients, and it gets really life-threatening actually.
Jordan Ruggieri 06:14
How does this tracking work, or the research work?
Donna Paznar 06:18
I can answer that in two parts. Basically, when somebody comes in presenting with some sort of lung infection, whether we don't know yet, and then they come back and test positive for Streptococcus pneumonia. Of course, first things first, you're treating the subject. It's very important as well which antibiotics you use, so that's another thing that's like you want to make sure and test right away, test quickly and ensure that the subject is receiving the correct treatment. Now, there's two sides to that coin, because, of course, one, we want to make sure, as well, you know, if there's any elderly or young children in the household, because that's another huge risk at that point as well, and then that sample from that subject will then come to me, and so that's where I then perform the testing and the research to figure out what stereotype it is, and that's a really long process. It's like one sample would take you like almost three and a half days, so in terms of the sereotyping that I'm doing as well right now, in terms of the tracking and the surveillance, the thing is Austria doesn't have a surveillance system at the moment for these bacteria. And so my research focus is mainly, first of all, we want to get as much information as we can. So we've partnered with these pediatric clinics on a voluntary basis to gather that information, and then we have these different surveillance sites, sort of set up throughout Austria, and those are just routine tests, so anytime a child comes in for just their routine checkup, or maybe it's a vaccine appointment, or things like that, they are, of course, then asked if they would like to, you know, test. But the problem with this, and it's something that we were really in the early stages of figuring this out as well, is the fact that what do we do with the information if the patient is positive.
Jordan Ruggieri 08:13
That's really interesting and makes a lot of sense. I mean, that was, that was actually going to be my, my next question is talking a little bit about colonization, and could you maybe elaborate on that. And, like, you know, help me understand, right, hey, if I'm, if I am positive for, for this bacteria, but am a, you know, a colonized individual, not a necessarily somebody that has the pneumonia aspect of it or the sepsis aspect of it, what does that mean? How do I have the bacteria but not have the disease it causes?
Lisa Crawford 08:51
I mean, it's like being a carrier, right. Like, some people just have these things and don't even realize it.
Donna Paznar 08:56
Exactly. Thankfully, it almost always, if, if you are colonized for the bacteria, you almost always will test positive. And so the reservoir is very similar to what we have with the COVID-19 virus, where it's in that nasopharyngeal tract. So it basically, the bacteria kind of just stays there, and it event, it doesn't. So again with these latent phases and these active phases, it doesn't necessarily mean that the bacteria is active within its infection period, so to say, which basically means that when it's in these latent phases, it's not replicating, it's not doing, it's not affecting, it's not causing any, it's not causing any interference, so to say, with the area that it is in. But it just means that you essentially are a carrier, and a lot of the time, as well, depending on the bacteria. So, what can also happen is, let's say somebody comes in for, I don't know, any sort of, maybe bacterial infection, they're given some sort of general antibiotic that can also clear a colonization of something else, which is also an interesting facet to this whole project is again, when are we catching these colonized stages. Because you can't exactly tell when, that's the other, that's the problem, you can't tell when this with the research, you can't say, “Okay, they acquired it on this day, at this hour, at this time, and they've had it colonized for this long.” But it essentially just means that you have the bacteria within your body, but your immune system is strong enough to not allow it to spread, and usually it's in some sort of latent phase of its replication cycle, where it's not active at the moment, so it just, it's just hanging around, basically. And then what can happen is the second that it leaves that latent phase, for whatever reason, you also likely won't show any signs or symptoms, but that's when you can pass it on, and so that's where there's this again, this tracking is very, we really see so far from like the early research that we've done, typically a child is a colonized, has a colonized case, and then other family members within the household, so usually younger siblings, so let's say there's a baby or an elderly grandparent that may live in the household, those are the ones getting affected. So, the parents are usually in the clear. Sometimes the parents are also colonized, but it's very rare from what we've seen so far. But the children have this colonized strain, and again, we might, we also can, the further, there's further testing you can do to actually tell the phase that the actual, that the bacteria is in, but likely when we catch it, it's almost always in this replicative phase, where then it's again being able to be passed on.
Jordan Ruggieri 11:52
I'd love to talk about some of the technologies that you use in your research. Can you? We love digital PCR; we love qPCR and sequencing here at Absolute Gene-ius. Can you talk a little bit about how maybe qPCR, dPCR, sequencing play a role in what you do? How do you use these technologies to get the answers that you're looking for?
Donna Paznar 12:13
Everything that I do is reliant first on we use qPCR. With Streptococcus pneumonia, we don't actually have a threshold level, so to say. So we don't have, “Okay, if the Ct value is below this, you know, we might have to think about whether it's really positive or negative kind of situation.” Anything that's under the Ct of 40 we consider as positive, and then we always, if it's very, very, very low Ct, we always do a retesting of it. But it's huge for me, because you know these lower Ct values, they are very, very impactful for me to know, okay, what stage of the disease, or which stage, you know, might be very, very highly infectious at that stage. And then those later on, you know, again, also for testing subjects throughout, we will also do, you know, testing of the day that they maybe had presented with these symptoms to us, then three days later, then six days later, a week, you know, and then again, seeing that difference in the, in the values of their of the qPCR data that I have. But the biggest thing that comes from that is again positivity or negativity. So, as soon as I have that information, which is huge, that's when I then go to further down the line of the sequencing. So, from there I'll usually run a nested PCR, so I'll run a nested PCR with the extracted DNA from the subject sample. We will basically then send that off the product off to sequencing, or we'll sequence it ourselves. And then that's when the fun begins, because then it's basically, you know, aligning sequences and figuring out all of exactly what serotype it is, and things like this, but that also is very difficult with Streptococcus pneumonia, because there's over 100. So you have to kind of have these very specific primers, probes, things like that to actually figure out what you've got and what you're dealing with. And my PhD focus is actually going to be on designing these kind of multiplex qPCR assays, where I can detect what will be, we will eventually actually group the serotypes. So without those molecular methods, there would just be no, no chance of being able to actually carry out this research.
Jordan Ruggieri 14:48
Can you elaborate, what is nested PCR? Can you, can you maybe give just a description on that for, for our listeners?
Donna Paznar 14:57
Yeah, yeah, of course. So, um, basically, so it's again a polymerase chain reaction. So, what's different about a nested PCR compared to a qPCR is usually there's two sets of primers that are used, so, and you do two successive PCR reactions. So you do your, you run your first round, so you have round one, so to say, with your first set of primers. And specifically with my research, that first round is actually meant to amplify a broader range of the DNA region of Streptococcus pneumonia. So we know, all right, we've got Streptococcus pneumonia from our qPCR, we found out, okay, we're positive. From there, that's when it goes into the first round of the nested PCR, where we kind of amplify this broad region of the Streptococcus DNA. And then the second primer, this sometimes is like referred to as the inner primer, and it uses a small sample of the first round, so you actually use the product from your first round of PCR, and then you use that in your second master mix with your second set of inner primers, and this usually, or is meant to, which is like the fun part of designing your own primers and assays, is targeting a smaller sequence of the bacteria that you're looking at. Another reason why we use that, is that nested PCR usually can target DNA in really small amounts as well. So you don't need as much bacteria in a, in a patient sample, so to say, to carry out that reaction. And so that's basically why I've used it, and then again it does really, really help with the further sequencing of it. Because I have, I have more of a targeted region of that of that area, and then I can then use that data that I get from sequencing. So the second round of the PCR is actually the product that we send off for sequencing, so we run it's called an E-Gel, and it's a, it's an E-Gel gel electrophoresis system. It's the best thing I think has ever been invented. Truly, this thing I've never run a gel. I can run like 30 gels in a day, and they're all done. And the nice thing, too, is that you can have multiple - I mean, you could have multiple samples in the old gels, as well. But you need nothing, like you just need a plug, and that's it. I mean, I don't need to, I don't need to wear my UV protection glasses, go into the dark room, then take the scalpel out, and cut my products. It's like the fastest thing ever. So that's another side piece that we, I couldn't do this research without.
Jordan Ruggieri 17:41
I remember back in, back in when I was in the lab, trying to cut out gels, and you're sitting there, and you're, you know, we didn't have any of this high-tech stuff either, it was a, you know, UV lamp and a cardboard box kind of thing, right. And you're trying to, like, looking in, where is it? Okay, I'm going to cut there. Oh, I cut my band, I got to redo it. Agh!
Donna Paznar 18:02
Agh! It's the worst.
Jordan Ruggieri 18:04
So for the nested PCR, the reason for let's say you amplify a broader range and then you go a little bit more narrow, is it simply because you're trying to look at a narrower range for serotyping? So you're looking at just having a larger amount of that very particular segment of DNA, and so you do nested to make sure it's a little bit more focused on the exact area you're looking at to have sequence for serotyping?
Donna Paznar 18:34
Yeah, absolutely. And like another way you could like also say that is the fact that it's so highly specific. It's so, so, so highly specific compared to other methods, or just a normal conventional PCR, for example, where you're just using one primer set. So because there's over 100 serotypes, you can imagine there is very many genetic differences between each of the strains, but they're very small, like they're not jumping out at you, so to say. So you also, the genes that you're, that you're targeting, you are working with very, very small pieces of very, or small variations, so to say. And so nested PCR also, like the second part, is very, very specific to the serotype itself. So then you know that the product that you're getting, not only are you reducing your chance of getting a false positive, but you're also, you know, that your, your product that you're getting is going to be very, very highly specific for that serotype, specifically.
Jordan Ruggieri 19:36
And then do you sequencing, you know, as a, as a kind of confirmation that you've hit that serotype? So are those primers, those inner primers kind of set for the specific serotype, and then you use the sequencing as, as a method to make sure you're looking at the right one? Kind of confirm that that that is the serotype that you, that you think you have? Is that again a good assumption there?
Donna Paznar 20:00
Yeah, absolutely. And that's one of the things that, for us, it's, now that's the tricky part is again designing these primers, because you're the first. So, before we get to the nested PCR, the goal is actually to design a qPCR assay that targets the groups. So then you maybe have 10 or 15 serotypes in the group, and then from there you're going to have to run multiple nested PCRs with the same sample to then know which serotype you're going to get. But it starts to get narrower and narrower and narrower until you get, again what you said, the product, and then the sequencing is a confirmation for us.
Jordan Ruggieri 20:41
That was going to be my next question, is you know, can you, can you just set up primers and probes for each of the serotypes? I mean, I imagine it's different, it's difficult when you're trying to narrow down from 100 serotypes, right, or 100 plus plus. But if you, you can get it into 10 groups, it makes it a little bit more feasible to maybe use that nested PCR technique to identify, you know, okay, it's within x number of serotypes based on this group, and then we can, we can identify from there in a way that's pretty rapid, where you're not necessarily waiting for results to come back or spending a lot even to try and get that answer? You'll confirm, but, but seems like it's a little bit faster, helps narrow down a little bit more, and a little bit less expensive as well? Again, I'm assuming. Is that, are these good, good assumptions?
Donna Paznar 21:35
Yes. The only thing I would say that's a little bit maybe not so accurate. It's expensive. It's still really expensive. But I think in the future what the goal will be post research. So, the research right now, of course, the sequencing is expensive. It's, you know, you've got a lot of time invested as well. But I think right now that's only because that's part of our confirmatory process, is again, you know, like getting the sequence and then confirming that we have what we thought we had from the primers. I think in the future the goal would be to not have to do the sequencing and be able to fully rely on the primer sets themselves. So that, and then again, I think part of my research will also encompass the fact that I'm going to group the serotypes based on those sereotypes that are also covered within and with, like, those that are covered with the vaccines and those without. And then again further on it'll be, so it will be. The goal is to be much more cost effective for the healthcare system, but right now, of course, we're spending the money to figure all these little pieces out. But yes, it is, it is definitely much more cost effective than running qPCRs or conventional PCRs with every single individual serotype.
Jordan Ruggieri 23:06
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Lisa Crawford 23:14
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Lisa Crawford 24:01
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Jordan Ruggieri 24:05
And now let's get back to our guest.
Lisa Crawford 24:11
All right, yeah, so I want to back it out a little bit and just talk more high level about your, how you got into science, your experience in science, and just, you know, your journey to end up where you are now. So, if you think back to your childhood, little Donna, what was it that you remember as being the thing that got you interested in science to begin with?
Donna Paznar 24:33
I can truly remember the day. I don't know, in the U.S., if you guys have this as well, but in Canada, when you were in your high school years, you are, you're mandated to do a certain number of volunteer hours. So I decided, because we lived, I lived, I grew up in a really small town north of Toronto, it's called Collingwood, and we had a vet that we were like, my parents were friends with, and so she was like, "You know what, you can come, you can volunteer at the vet. And you know, get your hours." And so I was like, "Perfect, this is great." And when I was there, there was one dog that was there for a long time, and they couldn't really figure out what was wrong with him. And I was like; my job was basically just cleaning the cages and like just keeping things organized. I didn't do anything other than that, basically. And I was cleaning his cage one day, and I took him out, and I was like, there's something really weird, like on his nose, so I went and I got the vet, and then she came back, and she's like, "Oh my God, it's a tick". And so actually, what was ailing this dog at the time was some sort, I don't remember what tick-borne disease it was, but from that day I was like, I went home, and I was like, "Mom, I need to know everything there is to know about ticks," like I'm like, "I want to study ticks, I want to study tick-borne disease." And then I remember, like, there was all this hype and talk about Lyme disease back then, and that's like where my, I was so hooked on that. And so basically my whole, since then, my whole life, or so to say, my career has revolved around bacteria and vector-borne disease, or you know, human to human disease, like we see with the STIs, and I just got so into it. I went and I did my bachelor's degree at Queen's University in Canada. And I did it in science. I just got really, really into figuring out, okay, what do I want to, what really do I want to do, and then I decided that studying molecular biology, and then figuring out how these bacteria or viruses actually are physiologically inside of you. So then I ended up coming to Austria just randomly to visit some friends that were living here and ended up saying, "Okay, you know what, I like it here a lot. I think I'm going to do my master's here." So then I applied for the molecular biology master's program here and did it in Innsbruck, and then my tick love came back, you know, it came back again. So it wasn't just a phase, Mom. I'm sorry.
Lisa Crawford 27:01
When you go to the cafeteria at lunch, do the tick people all sit together? Like, “Oh, those are the tick people, we don't talk to them.”
Donna Paznar 27:10
Yeah, it's like, those are the tick people. It's like the STIs. We have our water hygiene. You know, we've got it all in the cafeteria there. But I will say that the tick group does throw the best parties, that is one thing I will say.
Jordan Ruggieri 27:23
You really latched on to tick-borne diseases.
Donna Paznar 27:27
I really latched. Honestly, the moment that I found out that the Hadid’s and Justin Bieber had some sort of tick-borne disease, Lyme disease, I was like, "I'm in, I want to treat them, I want to meet them." But that was, that was, yeah, that was 13-year-old Donna, like, “Oh my God, we found our, we found it.”
Lisa Crawford 27:47
Couple more questions. Something we like to ask everybody is what, you know, having spent all this time in science and doing all this lab work, what comes to mind if I ask you A, your proudest lab moment that you can remember, and B your most embarrassing lab moment that you've had?
Donna Paznar 28:05
Proudest. Well, okay, let's go most embarrassing first. So the amount of times, specifically in my master's, that I started a qPCR without selecting a target, I should be arrested for. Because then you've just like not only wasted the money, the time, the time that that takes to then see how did I not select a target on that, and then you just have to redo everything all over again, truly, should be a crime. I would say my proudest moment was we had a tissue biopsy sample from a child who was presenting with crazy symptoms. This was like one of my first, first things I ever did in my new job. So this was like I was like one and a half months in, I remember we had this subject, so to say, or this child, and it was like a "House" episode. We were like all sitting around this table, and we were like, what? And we only had like a very, very, very small amount of cerebral spinal fluid from this, and we were like, “What can we do here? Like, what is possible with this?” And I had suggested, actually, a tick-borne disease that we rarely see, it's so rare, it's called Borrelia miyamotoi. It's also part of the Borrelia Lyme family. I remember my supervisor was like, like, “Do we really want to waste this, you know, we really want to waste this on such a seldom seen bacterial pathogen?” And we ended up running three tests, one of those, which I was able to convince to say, "Okay, let's do this," and they came back positive, and we were able to treat and find it out, and it was like, yes, like, yeah, we were really, that was probably the proudest moment.
Lisa Crawford 29:57
I'm the smartest person in the world.
Donna Paznar 29:59
Yeah. You're like, call me Dr. House from now on.
Lisa Crawford 30:03
One last quick question. I know we're getting to the end here, but I do like to ask this. What advice would you give to someone who maybe get interested in getting into your field, getting into science in general? What is something maybe you wish you had known before you started?
Donna Paznar 30:18
I think one huge piece of advice is take any opportunity you can. Like I think that that is also, there's so many areas of science that you will be overwhelmed with how or where to start, which I think I was as well. And I think from trying so many different things, so from going from tick to STI to respiratory to tick again, and then back to respiratory. It's one of these things where the most experience that you can give yourself is only going to benefit you in the end. Because it's only going to point you in the direction that you want to stay in. And I think for me it's something that I wish that I knew also back then was like, you are going to fail. I know that that is something that is so often, you know, you just think, “Okay, I'm doing this now, it has to work.” You know, you're on a project and your supervisor, or whoever has given you an idea of what should be the outcome, but it doesn't work, and you just think, “I'm failing, or I'm doing something wrong,” or but things just sometimes don't work. And you have to also, that's also part of the research as well, is to know that you are going to fail, and sometimes it's just, you know, that's an answer in itself. So no answer is also an answer. And really to remember just to keep an open mind. More like I think that's a huge thing as well, that just experience everything you possibly can, anything, any opportunity, whether it's you know sitting in on a different lecture that day that maybe you wouldn't normally have access to, but been invited, or somebody says come on. Just as much as you can do is the best, I think, advice I can give, because it really just helps you figure out where you want to be and what you want to be focusing on.
Jordan Ruggieri 32:11
Donna, thank you so much for joining us for today's episode. Really, really enjoyed it. Really, thank you, thank you so much for your time. I learned a ton. I hope our listeners did as well.
Donna Paznar 32:21
Yeah, it was so much fun to talk to you guys. It was like, like hanging out with friends, you know, talking science. It's truly the best.
Jordan Ruggieri 32:30
That was Donna Paznar, head of Molecular Infection Diagnostics at the Medical University of Vienna. Until our next episode, we encourage you to stay curious and to remember to check for ticks after hiking or walking through tall grasses. This episode of Absolute Gene-ius was produced by Sarah Briganti, Matt Ferris, and Matthew Stock.
Jordan Ruggieri 32:50
Products mentioned in this episode are for research use only, not for use in diagnostic procedures.