2. Studying Evolution and Engineering Bug Guts w/ Jeffrey Barrick

In this episode, we interview Dr. Jeffrey Barrick, an Associate Professor at the University of Texas at Austin. We talk to Dr. Barrick about the famous long-term evolution experiment in the Lenski lab, evolutionary failure in synthetic biology, tools for engineering the bee gut microbiome, and more. Along the way, we talk about cricket farming, how a bad memory can be good for open science, and some career advice for young scientists.

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Hello and welcome to EBRC in Translation, a production of the Engineering Biology Research Consortium's Student and Postdoc Association. We are a group of graduate students and postdocs working to bring you engaging conversations with members of the engineering biology community. EBRC is a non-profit public-private partnership dedicated to bringing together an inclusive community committed to advancing engineering biology to address national and global needs. I'm your co-host, Katherine Brink, a graduate student in the Tabor Lab at Rice University. And I'm Fatima Nam, a postdoc in the Sonnenberg Lab at Stanford University. This episode, our guest is Professor Jeffrey Berrick, an associate professor in the Department of Molecular Biosciences at the University of Texas at Austin. Welcome to our show, Jeff. We're really excited to have you with us today. Thank you. I'm excited to speak with you. All right. So to get things started, could you tell us a little bit about yourself and what your lab has been working on? Sure. I guess my background is in a mixture of evolutionary biology, biochemistry, and microbiology. And then when I started my lab here at the University of Texas at Austin, I started getting more and more into synthetic biology and combining it with some of those interests. And so I would say that the kind of mission statement of my lab is to use these new tools for genome engineering to understand and also manipulate the evolutionary potential of microorganisms. And more recently, we've also started some work related to that, but new, on engineering bacterial endosymbionts of insects and for various applications, including in material science. That's really neat. I was wondering if you could talk a little about how you got interested in microbial evolution to start with. We know that you were involved in the long-term evolution experiment in the Lenski lab, and it'd be great to hear a little more about what that experiment is and what your involvement has been. Sure. So I've been interested in evolution ever since I was a freshman in college, where I heard a professor speak, Richard Roberts at Caltech, about using evolution for discovery of kind of novel molecules in that case that had new functions. And I'd actually been primed for this because I was raised by a micropaleontologist. My father is a professor of geology who works on microfossils. And so I had read a lot of evolutionary biology and everything kind of clicked that I could do the biochemistry, but maybe not design my own molecules rationally, but allow evolution to teach me a little bit about that process. And so I kind of got more and more involved in that over time. And by the time I wanted to choose my postdoc, I really wanted to up my background in evolutionary biology. And I also wanted to move from the level of molecules up to microorganisms becoming a little bit more complex. And so, as you said, I joined Richard Lenski's lab at Michigan State University, who has a kind of iconic experiment that's been going on for 30 years now, where he's just kept 12 independent lineages of bacteria evolving under laboratory conditions, and importantly, kept a frozen fossil record of these cells that has been able to take advantage of all kinds of new technology that develops after he started this experiment in 1988. Has COVID really affected the experiments? Are they at pause? Right, right. So, I mean, one thing that's great about this system for studying evolution is that you can pause it, like you said, or even like save your game, right? So the cultures are actually backed up across the world in many different labs to prevent against loss of the samples and interruptions. But yeah, they put it on hiatus for a few months here. It's only recently kind of started up doing the daily transfers again. But it's a really nice resource for lots of people to study these processes. Really cool. I know that when you started your lab, you started working on looking at the dynamics of microbial genome evolution in laboratory evolution experiments. And you showed how microbial evolutionary dynamics are often dominated by rapid adaptations. What were some of the key interesting findings in this area? Right. I think some of the long time stories that we investigated using this long term evolution experiment with E. coli that I was most interested in are some of the changes in what I would call the evolutionary potential of the microbes. So they get one mutation, then another mutation, then another mutation, and all of these mutants compete with one another, sweep through the population, drive extinct their ancestors. But as that happens, their ability to evolve changes in a few interesting ways. One of them is that a lot of them evolve high mutation rates early in the experiment because that allows them to essentially try to get the jackpot of good beneficial mutations more rapidly than other lineages. But it turns out one thing we learned that you could only see from this long experiment is that early on, that's great because there are many opportunities to improve. There's a large supply of beneficial mutations. But later, as they become more and more adapted to this environment, those opportunities become fewer and fewer. And it actually becomes worse to have mutations because most mutations are deleterious. And so we saw mutation rates evolve to be high and evolve to be low. There's interesting things that happened mechanistically thinking about how that can happen in a bacterial genome that occurred. And then so that's one thing, mutation rates. But also there's this contingency and the interactions between mutations. If you take one path early on, that can shut the door on other future paths. And so we have some really interesting results that we're still trying to figure out mechanistically what's going on. But we know that certain mutations were very successful early in the experiment, but they always went extinct in the long term. So for some reason, they made them less evolvable in terms of these interactions between future paths. And then one of the most interesting things that happened in that experiment lately was that one of the 12 populations evolved the ability to use a new nutrient, citric acid, that had actually been sitting there in the media unused all along for 15 years. And in 11 of the 12 flasks, it is still sitting there unused. And so trying to understand why this is very rare and why that one particular population was able to evolve to access it is a really long story of contingency that we're trying to unravel. But it's challenging. It's really challenging. We can count all the mutations. We can do RNA-seq. We can do all these things to characterize the cells. But connecting that to fitness is still really a challenging thing that is related to a lot of things in engineering biology. We have metabolic flux analysis and things like that, but they still have an incomplete picture of the fitness of a cell. So thinking a little bit about that fitness aspect, it seems like you're mostly studying microbes kind of in isolation or like one microbe at a time. Do the results that you've found also apply to more complex and established ecosystems? So thinking about human or maybe even insect gut microbiomes. Right. That is a great question. And I mean, there is a large and vibrant field called experimental evolution, where people are adding back some of that complexity on top of this kind of like simplest hydrogen atom model of evolution that we have with the Linsky Lab experiment and making it more realistic. Many of these studies are starting to look at evolution of microbes associated with animals. So there's some really nice studies looking at just taking E. coli in the gut of a mouse and seeing how its evolution occurs over time. It turns out things like bacteriophage are very important and the dynamics are quite different and the types of mutations and genetic exchange that can happen really influence the dynamics. Right. And then there's some people starting to try to look at communities of microbes in various ways. And I think the ecological aspect is really interesting. It constrains the evolution a lot of the time. If there are a lot of microbes there that are already very good at all the different things that you can do to become fit in an environment, there are fewer opportunities for you to evolve and kind of colonize those niches. Right. So we're learning a lot about that. And there's also people studying design consortia, which is another really active and interesting area in this, in the lab, you know, for where there are lots of applications that people want for consortia, but we need to understand their stability. Right. And then the real question is when you engineer a microbe that you want to add as a living therapeutic to the microbiome, will it continue to function over time and how should you add it and how can you keep it to genes from flowing into the native microorganisms? So there's a lot of unknowns and we certainly need more studies in this area. That's actually a great segue into our next question, which is thinking about evolutionary failure in synthetic biology in particular. And I think since our audience is mostly engineering biology folks, it would be great to hear a little bit about your evolutionary failure mode calculator and kind of what you think are the biggest places where synthetic biologists make mistakes when it comes to instability and what they can do to improve their circuit design. Right. So this is kind of flipping on its head. My other studies of evolution where I like, I love studying microbes. I love studying how they evolve, but it becomes inconvenient, confusing, frustrating when you try to engineer a cell and then it does not behave the way you expect. And sometimes you don't know why. Right. So I see this as one of the things that's in that black box of failure where often we take the approach where we make a lot of constructs. We try them all. Some of them work as we expect. Some of them never work. Some of them, we never know why they didn't work. And one potential reason for that is evolution. Right. Because these are DNA sequences we are putting in a cell. So obviously mutations can arise in them and they can affect the cell population. Now, the real question is when does this happen? When does this crop up and become important? And so in talking to people who work on different systems, bacteria especially are kind of prone to this because they can move a lot of their resources towards the one thing that you engineered. Right. You can put a huge burden on the cell. And so I think learning more and more about that and just being aware of this is great. Now we can sequence genomes. We can sequence whole plasmids. You know, we can really characterize what's going on. So the kind of one of the viewpoints of my lab is saying, okay, we can design cells. We can make a DNA sequence. It encodes a protein. You know, it includes a circuit. It encodes something very complicated, a metabolic pathway. But you know, there's a lot of choices for those DNA sequences we could use to make that thing. So we should actually remove some of them from consideration. We should cut out the ones that potentially have problems. One of the problems being that there's certain sites in DNA that are hyper mutable. So that's what the evolutionary failure mode calculator is kind of designed to do is try to alert you of those things so you can not use those particular sequences and remove them. Now I think there's a lot of interesting work here too, where people are learning to more rigorously measure the burden that genetically engineered constructs has on a cell. And you know, if it only burdens the cell and slows its growth rate to about like 90% or 80% of what's normal, it turns out that's probably not going to affect your experiment in the lab. There's not enough time for the mutant that eliminates your function and starts growing more rapidly to take over a population of cells. When you start getting to the level of 30 or 40% growth rate reduction, that's when you become unclonable, where even by the time you grow a colony, your plasmids have already mutated, right? So something like this. So I think just being aware that you have a certain threshold and if you start seeing certain things, you should become more and more worried about what's going on, you know, super small colonies, just characterizing what's going on. So I want people to be aware of that. And one thing I'm really excited about is that our 2020 iGEM team that I kind of co-mentor has started putting this burden value on iGEM biobrick web pages as part of their project. So actually 300 web pages now have this plasma that you could get from the iGEM registry, slows the growth rate of E. coli by this percentage. And you know, it's only a small percentage of these plasmids that on their own burden the cell enough to be worried about and are probably unstable actually in the iGEM collection itself. But when you start adding things together and engineering a cell more and more, you're more likely to cross this threshold, which as we get better and better tools, you know, we are pushing those limits. Or when you're trying to eke out the last bit of production of a biofuel or something like this, you really need to worry about the stability as one aspect of your design. That's fascinating. Yeah, it has been super helpful to see tools like this come out for synthetic biologists. And people like me who have worked with model organisms like E. coli, it's great. But one of the more exciting projects that your lab has recently been working on is this work on the insect gut microbiota, which is super fascinating. And we hear a lot about the efforts of engineering plants directly or engineering different microbial symbiotes in plants. But engineering these insects and biotes is something new, and especially looking at the potential and using it on the academic side as well as commercially. Like there are companies that spun out from Ginkgo Bioworks. All of these, what do you think has really led you to decide on insects and biotes specifically? Right. So that's actually kind of a simple answer. And I would give this as career advice. So you are in a certain scientific environment of colleagues. You learn which of those colleagues you like to talk to, learn from, work with. And so I was very lucky to end up in a situation where I have many great colleagues in synthetic biology here at UT Austin. Also evolutionary biology, which is one of the reasons that I was excited to work here, start my lab here. But also more recently, we've had some great colleagues who are experts in symbiotes of insects, in particular, Nancy Moran, who has studied all kinds of different systems like this. And for us, you know, as engineers, we often want to apply our tools to interesting problems, right? And so having an expert who knows about a certain study system is invaluable because then you can learn from them and use your tools and their system to kind of move things forward. And also in this case, I would say that insects are, you know, they're still micro. They're only a slight step up. There are a lot of advantages to studying insects compared to, say, mice, right? They're invertebrates. So it's a lot easier to do the research and get up and running, house them, work with them and things like this. So I think in a lot of ways, they're a nice test tube for studying some of the problems we've talked about. But also, I mean, insects are the most diverse, most numerous in terms of species, animals on the planet. And so there's a huge variety there. And what I was really interested to learn was that a lot of insects have bacteria growing within them. In some cases, it is a lot like the human microbiome. So honey bees is kind of what we're talking about here, where they have characteristic microbes that colonize their gut. They have a kind of strange diet, right? So they're feeding on vector and pollen. So it's a very specific types of microbes there. But there's a core set of them that you find in honey bees and also in other bees. So they have a defined microbiota. It is important for their health and protecting them from pathogens and other things. So it's a cool study system for that. But also, a lot of insects have microbes that live inside of their cells and have essentially started evolving into organelles of various kinds. So Nancy has also studied aphids for a long time in this capacity. But a lot of other sap sucking insects have the same problem where they feed on a diet that is super rich in sugar, but it does not have many amino acids in it. And so animals can't make certain amino acids. So they essentially co-opted a bacterial cell to become an amino acid factory for them. And it's turned out that now they cannot live without the bacterial cell that is inside of their cells. And also that cell can no longer live without the insect. So really stable, important types of interactions between a microbe and its host. And then there's all kinds of things in between. And there's more stories about Wolbachia and other reproductive parasites of insects. There's all kinds of bacteria associated with insects in really intimate and crucial ways for the biology. So I think that's an opportunity for engineering these systems. And then insects can be beneficial, like honeybees, where these are important pollinators, also bumblebees, where we are worried about their health because agriculture depends on it very strongly for many, many, many crops. But also many of them are pests. Many more insects are pests. And so understanding it from both perspectives, there's potential for applications in engineering the bacteria, which is an easier lever for us to work on genetically than engineering the insect host in these cases. So I think it's really interesting for all of those reasons. So could you actually delve into some of the more specific aspects of your honeybee and aphid engineering? It'd be great to hear. Sure. The honeybee work is kind of published and out there and the furthest along. So maybe I'll concentrate on that first at least. So in that case, again, due to this great collaboration with the Moran Lab and other labs here at UT Austin, we took one of the native gut symbionts of honeybees and we engineered it. And one thing that's cool about insects too is that their kind of internal workings are a little bit more open than than other animals, mammals, for example. And so we were able to overexpress double-stranded RNA from this microbe and it is able to be taken up by the insect cells. We don't completely understand that mechanism, I'll say, but taken up by insect cells and induce their RNA interference response, which means that essentially it can knock down a targeted gene. Now we can target that against a bee gene. And that's great because this becomes a tool for studying honeybees, which are actually not easy to study. You cannot do genetics because you need that quinging, right? So you need a whole hive as your unit if you were trying to do breeding or things like this. So that's really hampered kind of study of honeybees for a long time. And so potentially we could colonize them with a microbe that knocks down a gene to study that gene's function. But in this study, we've applied it to try to protect bee health in a few ways. And so one way is to target that response against a bee pathogen, a virus that could infect the bee. So now you've essentially immunized them kind of up to their immune system response before they were exposed to a virus. And that can prevent the virus from being able to replicate when it gets into their cells. And I should say that also this spreads throughout the bee's body. There are systems in the RNA response that make it systemic in this animal. So even though it's coming from the gut, it affects the cells in the brain. For example, we've been able to alter their feeding behavior and other things like this. But kind of the coolest thing is that bacteria have these parasitic mites. They're kind of like ticks a little bit. They kind of feed on the hemolymph, the blood and the fat bodies that live within the mite. So they're parasites that you just see on them and they cause problems and they become too many of them in a hive. And it's one of the problems with colony collapse. They're thought to be implicated in that. But they're also an arthropod. So it turns out you can induce their RNA response by making this double-stranded RNA in the microbe, in the bee gut. It gets into the bee. It is replicated in this process. Then when the mite feeds on the bee, it gets into the mite and it also induces a response. Now you target it against a gene that the mite needs to survive, an essential gene, and we were able to show that it decreases mite survival when they feed on the bees that we've colonized with this microplenum. So I just think that's an incredible kind of ability to use this system for potential applications. I should say all of this was, of course, in a lab. These are genetically engineered microbes and there's all kinds of interesting questions about whether and when and how you would be able to use these technologies out there in the real world. Do you have plans to move it to the field? We do have a patent application on this technology and have been talking to various potential partners. Things obviously kind of slow for developing new projects right now due to COVID and everything like that. Has there been any work towards biocontainment and safeguard systems? I know you talked about any sort of horizontal gene transfer or issues that could happen, especially in field settings. Right. We've tried to think about that carefully. I think the biggest answer is that tests are needed, but one of the things is what should you be testing for? What should you be more worried about? What should you be less worried about? One thing about these bacterial symbionts of insects is that for the honeybee example, these are very specific to bees. It's unlikely that this microbe is going to survive in the environment. It's going to be passed to a different kind of insect. Some of the strains are even specific to honeybees and not a different one is found in bumblebees. There's something specific on that level. We think that there's a little bit of physical containment built into the system, at least within bees. Now, do we know if a worker bee goes into the wrong hive if it will spread it to that hive? No, we don't. Under some circumstances, you might want that if you were trying to inoculate it and protect the hive. You might want a certain amount of transfer, but most of the time you want no transfer. We have some ideas. If we can get some new funding, we have ideas to try to make these non-transmissible so that you could put that bee back in a hive and maybe study a gene in certain bees in a hive, see how it makes them behave differently in around the same environment of the hive, but make sure that the one we put in cannot be transferred to the other ones. Normally, the bee got microbiota is transferred via the same fecal-oral route that it is for mammals and things like this. It is spread amongst the workers in a hive. When they emerge, they are colonized by the new microbiota. That's the normal way it's transmitted in this case. For other insects, they actually inherit it from parent to offspring. In that case, you have a different type of biocontainment going on. That's the case for some of the symbionts with aphids, for example. I know another project that you have going on that also relates to insects is looking at these brocosomes in leaf hoppers, which I found out are native to Texas. I was wondering, if I go into my yard and find a leaf hopper, how can I use that to contribute to your brocosome studying efforts? If you could explain a little what a brocosome is and what the potential applications of that might be. Sure. Let's start with the insect here. Leaf hoppers are a type of sap-sucking insect. They are related to aphids. For example, they're in the same group, but they have this different behavior where they are quite rapid moving and jump. That's why they are hoppers. They have evolved this very interesting nanostructure that they produce. Within their gut, within their malpiggy and tubules specifically, they have certain cells that have specialized to secrete and they, in fact, excrete it the same way they would waste. Little droplets that contain these nanoparticles. If you want to picture these, if you're listening to this, they look a bit like bucky balls. They have pentagonal and hexagonal faces or kind of like a soccer ball. They are cage-like and hollow when you look at them under an electron microscope. The insect makes a milky white kind of droplet and then it uses its legs to anoint it to rub it all over its body. They essentially coat themselves in this. Insects molt, so they do this kind of every time after they molt. They also do it at some other times, so that it's some kind of coating for the insect. We don't exactly know what this does for the insect. I should say the biological function of these is really interesting. There's a lot of hypotheses, but I'll tell you about some of the applications we think they might have, which are definitely related to what we also suspect are the functions in nature. It is known that these give a kind of superhydrophobicity to the insect's surface, meaning water droplets bead up and easily do not get stuck. That's important for sap-sucking insects because they have to feed a lot of phloem or xylem liquid through their system to get nutrition. They're essentially continually pumping liquid through their system and secreting it. This is this honeydew stuff that sometimes gets under your car and makes it sticky, for example. That's caused by these or related insects. And so you can imagine that if you're a small insect, it might be very bad to get stuck in that sticky fluid. It might die, right? So not getting stuck in their own secretions or other sources of water is important for a very small insect. So that's probably what the superhydrophobicity helps with. Although I should say most insects don't make interesting nanostructures to deal with this. They just make wax or something simple like that. So it's kind of a mystery why evolution would have evolved something so complex that there's not other functions going on in this case. One other thing that they do is they have interesting interactions with light. So they have some optical properties. They have what's called omnidirectional anti-reflectivity, meaning light from different directions does not reflect back. So this is related to things like stealth technology. It might somehow make it so certain predators can't sense them or maybe more specifically their eggs. Some of them put the brocosomes on their eggs. Or it also might turn out that they coat predators. People have also thought that these might have antimicrobial properties. So they're superhydrophobic and their coating surface might protect them against bacterial or fungal pathogens, which could also be important, and also grow in an environment that is rich in sticky, sugary substances. So we have a really interesting multidisciplinary multi-investigator grant going on right now with collaborations with material scientists, some of whom are studying the optical properties of these structures and also structures that are made of metals that are inspired by the morphology of these. So there's bio-inspired and biological materials research going on. Some who study membranes that you would use for water purification, where fouling of the membrane is an important problem and potentially embedding these or coating the membranes with these could prevent certain types of fouling. And then also people who are more interested in the nano properties of these things and being able to take what on the insect is put on in a random pattern and making it an ordered pattern or a pattern that you have put other functional groups on the surface of these in order to produce new types of coatings or substances. So we're still at the very beginning of this project. It is very exciting to have a lot of funding and a lot of collaborators to work on what is essentially biological mystery. And this is just one amongst many things like this that insects make. There's all kinds of weird nanostructures, nanopillars, photonic crystals that beetles, butterflies, all kinds of insects make. But this is a particularly interesting one. So when you're talking about working with these collaborators with the brocosomes, I'm assuming you mean purified brocosomes and I'm curious how you purify them. Do you have to mash up leaf hoppers to be able to access these brocosomes? Is there some alternative way of producing them? Right. So I would say that in the short term, we're using the insects to produce them and they coat the insect pretty well and they stick pretty well, but you can actually sonicate them off. And the thing that I should also say that's surprising about brocosomes is that you imagine these are made of proteins or biological, they should be kind of unstable, right? So you would imagine that if you like dry them out, the insects out, or you throw them in the freezer or something, you might damage these. No, they are incredibly robust. So I would think of them more like something like silk, keratin, collagen, these other property materials that are made mostly of proteins. These are made majority proteins, it's kind of uncharacterized families of proteins that are only in these types of insects. But they also have some lipids in them and we don't completely understand their biochemistry yet or their structure. Those are things we're working on. So you can sonicate it, you can purify them in acetone, they have these hydrophobic properties, and you can get nice brocosomes. It turns out these are also floating around in the air all of the time and we're probably breathing them. When people filter pollen and they start looking at it under an electron microscope, sometimes they see brocosomes. So they're fairly stable in the environment as well. One cool application that's stuck in my head is you have to build invisibility cloaks. Unless you're protecting us from, say, ladybird beetles will do much good for us, but yeah. Okay, so we have talked about multiple different projects, but I think I'd like to go back to talking a little bit about the environmental release. So this has been a concern for any type of genetic engineering or synthetic biology, and we see a lot of public controversy over different field tests with engineered insects, especially looking at gene drives that has been done in mosquitoes. And at the same time, evolutionary stability is a big hurdle for many synthetic biology applications, whether it's in the lab or an industry setting. Do you think there is a place for really engineering instability into genetic circuits, like designing to help with containment of genetically engineered organisms? Could this be a feature and not a bug? Right. So I think that is a really interesting question. And I think that there definitely is a place. The question is, how effective can you be and what are the most effective ways to essentially make a disappearing message? It's kind of like you want to engineer something into the microbe, and then it's like mission impossible. You know, this message will self-destruct, or burn after reading. And the most difficult thing, though, is that you have to make sure it's not just 99% effective, that it's 99.999999. You get a lot of decimals of assurance that the message will disappear on a certain time scale. Now, certainly you can design genomes, you can design plasmids, you can design all kinds of things such that there is this planned obsolescence built in, I would say, and that's really interesting. I think there needs to be some testing and seeing which mechanisms are best at getting rid of a function. Because on the flip side, if that function is beneficial at all to an organism, it's going to keep it around, even if it mutates most of the time, right? Even if a lot of the offspring lose it, some of them will keep it around. So you have to make sure that there's no potential benefit in the organism you evolved, and probably no potential benefit in other organisms that might take up the DNA. Because we are learning that there's a lot more gene exchange in a lot of these complex microbiomes than I think people would have thought through some of the experimental evolution I've described in other studies. Do you have any thoughts for how to do those types of measurements to be able to determine whether a genetic sequence that's been engineered would pose any benefit to an organism potentially? Oh, right. So this is one of those questions where biology, ecology, evolution just become this exploding problem of complexity. The weather, the butterfly flapping its wings and having all kinds of unintended consequences and forming hurricanes. It's very difficult to do risk assessment in this type of field. I think that one thing that would be very useful is being able to put some innocuous sequences out there in the environment and seeing where they spread to and how long they last. So doing a controlled release of nothingness, just barcodes or something like this, and being able to track that in the environment. And we're getting better with sequencing technology, ways of enriching certain sequences and complex mixtures, where I think you could do some really interesting studies in the lab. And I think these would be allowed in natural ecosystems as well, as long as it's completely non-coding the sequence you're adding. Of course, once it becomes coding, it's another question and something else you would want to look at. But I really hope that people will take the time to figure out the regulations and be able to do some of these studies. And that will inform what's going on. One thing that's interesting is that probably there have also been unintended experiments in a lot of these areas. So we have very good protocols that everyone follows to autoclave their organisms to destroy the DNA when we work with recombinant organisms in the lab. But those are probably not 100% effective. And now that people are doing things like sewer surveillance, where they just look at sequences and microbes and things going that are making into the waste stream, which is one way people are starting to monitor for COVID, or there's all kinds of other interesting things you can do. You could look for sequences that are highly used in engineering in those environments and see if some of this is already occurring at some low level. If it is, that would provide at least a little bit of peace of mind that things are not becoming super widespread because of it. I mean, we should be worried about the antibiotic resistance genes in some cases, although the ones we use in the lab are usually not clinically relevant in most cases, which is a good design on our parts. But there's a lot of unintended experiments like that, I think. We've been studying where people take their plasmids and they use them, but they almost never sequence the backbone of the plasmid. So they don't know that maybe the copy number isn't changing or the antibiotic resistance marker isn't changing. And as we're getting more data, we want to kind of look at those as well. So there's a lot of natural experiments that might have happened, which are another potential way people could study those processes. This is a great segue for something I really wanted to ask you, something about open science, about sharing protocols. So historically, the culture of science has been really solitary. And I think your lab has made huge strides in making science more open and accessible. I love your lab wiki. I have been using it since day one of my PhD. And even last week I was doing an RNA-seq experiment and I had to go to the very lab wiki. You also had a fun paper about a DIY agro replacement, which was really cool for culturing microbes at home. Do you think open science is being pushed in the right direction? Right. So I would say that I did not do these things on purpose. You know, it wasn't like I sat down and I said, how should I design the way I do science to check some boxes? It was very natural for me to share things because I think all of us know that over time you kind of learn stuff. And for me, I have a very poor memory. So essentially, our lab wiki was my notes to myself, right? So why not share those notes with other people so that I don't have to try to find the information again somewhere else on the web? And I think that I would encourage this sort of viewpoint in people in general, being open with what you're doing. Sometimes it seems like not a great idea to teach other people to do what you are doing in science, right? Because you're worried that they might compete with you. But I would say that that has essentially never happened to us. By sharing our science and sharing reagents and sharing, you know, we also develop open source computational tools. I have made so many friends that I didn't know I had that when I go to a conference, I get to hear great things like Fatima just told me where people are using these resources or using our computational tool or emailed me about some bug and then I fixed it for them and they were grateful, right? So that's led to new opportunities to do science and just be involved in the scientific community, which is really rewarding, right? In the long term. So I think those trends are very good. I know that the bar is very high for you young scientists when you publish a paper now about the amount of extra documentation, about methods, about the number of supplemental figures, data, tables, protocols, githubs, you know, everything that has to go into those right now. So I really, I feel for you in terms of the amount of work that it can be, but I would say that it is really worth it, you know, getting that data set up there that is properly annotated that other people use will bring more citations to your paper, more attention to your science. It takes a little bit longer for those things to pay off, so you might not see it for a few years down the road, but it's really important. And I will say, I want to give a quick shout out to an experience I had during graduate school that I think really speaks to this and maybe was formative in the way I think about these things looking back, which is that I was in Ron Breaker's lab at Yale. We were looking at new riboswitches, and I was doing a lot of bioinformatics, finding these sequences that folded into secondary structures that had functions binding the metabolites and regulating gene expression of bacteria. So I was doing bioinformatics and aligning these sequences to try to figure out what the structures were. I was working a lot with the sequences, and I was using some tools and a database called RFAM, the RNA families database, and some of the tools related to that. And I just started submitting my stuff to the database. And then a few months later, I got this basically email from them, which was like, you submitted like 90% of the new things we've seen over this period of time. We would like you to come and visit us for a month so we can just watch what you're doing with your bioinformatics and stuff like that. So I got to go to the Sanger Center near Cambridge, England, spend a month there, stayed in a bed and breakfast. I'd had a full English breakfast every morning. I also got to meet a collaborator who I'd only known by email who was doing some of the development bioinformatics tools they brought us both in. And I got to experience that environment where they are really doing a service for all of the community by keeping these databases running, right? So RFAM and PFAM are heavily used, right? So I got to experience that culture for a little while. And it was a really great experience. And I definitely appreciate what they do and try to make sure my stuff makes it out there into these types of databases, partially because of that. But as I said, it was kind of an accident. I was doing, I just wanted my stuff to get out there and was appreciative of what was already going on. Kind of going off of that point, do you have any advice for young scientists who would like to pursue an academic route? Right. So it's a big question. I don't know, we could talk about that for hours, probably. But I would say that you have to recognize a few things. And again, I can speak to my own experiences and the way things developed for me a little bit. After undergraduate, where I'd done research for four years in a lab, so I knew quite a bit about what I was doing. I had a really specific idea about what I wanted to work on in graduate school. So I went to a lab, I wanted to do in vitro evolution of proteins, kinds of things. But while I was in that lab, other people discovered really cool things going on in biology. And in fact, I worked on my experimental project for five years there, never got it to work. But I took on these back up projects and I actually used kind of my, I guess, latent skills in programming, applied them to biology, where I'd programmed computer games for fun for years. So I kind of had these skills, but it never applied them to science. But the problems came up in the lab, I was there, and I was able to help a lot of different people in that lab study their particular RNA sequences. And it became a really great experience. And so, you know, I had a thesis on this background project by the end that worked out quite well. I could not have been luckier in terms of where I was at a certain time. So I think the lesson there is, you want to have kind of your true north, the things that you care about that inspire you. And, you know, I have followed that, you know, I still study applied evolution, I still study evolution. It's not in the systems I started out on at all. And now I'm looking at insects and actually their evolution of these brocosomes, for example. But that's there's a through line there that really inspires me. But also I've let I've taken advantage of things going on around me and tried to get involved in those projects where you're just trying to catch the wave, you know, that's coming by and benefit from the expertise of the people around you in a complementary fashion. So I think you have to mix those two things. You have to figure out when to switch between those two things, what's going on with those things. And then, of course, you have to also just keep high standards. I think that's another thing is that keeping high standards, contributing to open science, doing these things pays off in the long run, far more than the tendency to try to be a little bit too opportunistic to go after, you know, this potential funding source, write a grant for this thing, when maybe it's not really your kind of underlying interests, it will it will catch up to you eventually, and that you won't be inspired to do it, you'll be stuck doing something you don't want to do. So to the extent that it's possible, I would try to try to mix together those things. Now, there's obviously a lot of luck, a lot of contingency in this process, we want to bring it back to some of the evolution words, right. But having that true north and having a really good environment are important to success. So one question that has come to our mind, when thinking about people who have careers in academia, but also work with microbes, is how did you choose to go into academia as opposed to, for example, start a brewery? And if you have thought about starting a brewery, why hasn't that been a path that you've chosen or any other sort of career that might be a little more eccentric and off the beaten path? Right, right. So I do have several backup careers, I'll tell you about them at the end of this response. Have I brewed beer? I've only brewed mead, actually, which is fermented honey. I did this during my postdoc before I ever knew I would work on honeybees. But again, connections develop over time. So it turned out that was a little too sweet and too much of it to drink. So I kind of didn't follow that up. I have a lot of great friends in the experimental evolution fields who do work on yeast. And increasingly, they are working on how yeast evolve when they are reused and repitched several times in microbreweries. And so really interesting stories about how yeast evolves and has become domesticated and changes over time, which can change the taste and the properties of beer, for example, right. So yeah, my interests don't quite go to the brewing. Actually, the career that I always joked I would have if I was not in science was that I would be a mushroom farmer, trying to figure out ways to culture difficult types of interesting culinary fungus. More recently, you know, semi-seriously, I've learned about what they call mini agriculture, which goes back to insects. Farming crickets, for example, is a thing. Cricket meal is a thing. I could imagine being interested kind of in that as a problem to solve and working on that sort of thing. Sorry, if I can interrupt for a second, is that intended for human consumption or for animal consumption? I guarantee you that you could go into a somewhat fancy supermarket and find a energy bar of some kind that has cricket meal in it. So yeah, it absolutely is. And these things are high in protein. So the idea is that they can replace less sustainable types of agriculture potentially, right. So, you know, it takes a lot more resources for a cow versus crickets. It turns out crickets actually also have lots of problems when you try to culture them. Most of the people who've cultured these started having them to feed reptiles, right. So often you have crickets as a food for reptiles. But there've been a few epidemics where a virus got in and it completely wiped out these companies, destroying all their crickets, right. So, you know, there's also that kind of weird interesting angle where if you could protect the crickets from the viruses, you could have better cricket agriculture. Yeah, I don't know. This is clearly a very strange rabbit hole we're going down. But I think I would choose interesting things like that maybe to work on if I was not in academia. But being a professor can be a difficult job with lots of demands. But that's also the upside is that it's so diverse, you know. You spend some of your time writing, some of your time being forward thinking, a lot of your time mentoring, teaching. You get to mix together a lot of different things. And if I've learned one thing from being an assistant professor and kind of maturing a little bit in my career, it's that over time you get a chance to kind of shape the career a little bit to which of those parts you like more or less. And so I think that's really unique and that's invaluable. In a lot of ways, a lab is a small business. You know, you're trying to bring in funding, keep your people funded, work on it, bring out, you know, papers and things like this. So you get a little bit of that entrepreneurship or, I guess, freedom, danger, everything mixed in and then academic career as well. We are glad you took the academic route. I think I prefer my cliff bar over any cricket. No, thank you for all of that advice. And it's like to me, if I had to use a word to describe your research, your life experiences, all of the advice you had for us is symbiosis. So it all fits perfectly together. So at the end of this conversation, we wanted to ask you if you had anything to plug. Right. So I was going to use this to put out a postdoc ad of source. So one of the things that we've started to also work on is engineering bacterial viruses, bacteriophage. And I lucked into a nice collaboration with a computational biologist here at UT Austin, Klaus Wilke. And he has worked for years with a group of people here working on bacteriophage T7 as a model system. And that includes Jim Bull, who worked on the evolution of this. So very similar to Linsky experiments using this bacteriophage to understand how it can evolve. And also Ian Molinou, who's a longtime expert, one of the people who's really built our understanding from the ground up over decades on this thing. So we have a great group of people who are experts on this. This particular grant ties back to the stopping evolution thing. The idea is to take a genome, try to engineer it in ways that decrease its virulence, but in such a way that it cannot re-evolve high virulence. The potential application is thinking about this, not this particular bacteriophage, but viruses in general, where you might want to make an attenuated live vaccine such that you can put it in someone. It can elicit a very good immune response and potentially even replicate a little bit as part of that process. The polio vaccine is the main example of this type of vaccine. But the polio vaccine has a problem where sometimes it re-evolves into polio and all of the polio in the world is actually reverting from the vaccine strain at this point. So the idea is to use some of those tools of synthetic biology where we can change a lot more about the sequence than people did in making that vaccine where they just passage it for a while and saw what naturally arose to weaken it. In that sort of context, I think there's also applications in phage therapy and thinking about how you might want to engineer phages that are used in that direction. We're really excited about this project. If anybody wants to build bacterial virus genomes, see how they evolve, try to control that evolution, this would be a really great position for them. Well, thanks again so much for joining us today. This has been a really fascinating conversation. Great. It was a lot of fun. Great questions. Thanks a lot. This has been another episode of EBRC In Translation, a production of the Engineering Biology Research Consortium's Student and Postdoc Association. For more information about EBRC, visit our website at ebrc.org. If you're interested in becoming a member of the EBRC Student and Postdoc Association, you can find our membership application on our website. A big thank you to the entire EBRC SPA podcast team, Catherine Brink, Fatima Anam, Andrew Hunt, Adam Silverman, and Kevin Reed. Thanks also to EBRC for their support and to you, our listeners, for tuning in. We look forward to sharing our next episode with you soon.