the hearty quarter of the population who's held out for the last day of this meeting, it's really great to get to be here and talk about our stuff. And so I'm gonna, I also like how I think the organizers purposely, maybe it wasn't purposeful, but I think they kept a really nice percolated meeting between kind of theory talk, empirical talk, theory talk, empirical, so perhaps, you know, you can't get just like all of your excitement all at one moment, you have to kind of keep switching your brain, and I think it's actually really good to have this kind of back and forth. I'll have a little bit of math in mind, but not so much. And so I'm gonna talk, also I think about something that's a little bit at the borders of some of the other talks. We've had a fair number of talks about metabolism, and that metabolism is exciting, which I love. It's often been a rare thing to be the one in the crowd excited about metabolism. We've also heard about antibiotics and things that kill you. And now I'm gonna kind of bring those two things together and show kind of some commonalities between thinking about things like antibiotics and thinking about metabolism, because sometimes you even do things that hurt yourself. Maybe you did last night in downtown Trieste have a little too much metabolism in that sort of way. So first off, before I forget, I wanna just think, got some wonderful folks in the group. A lot of what I'm gonna talk about today was done by a postdoc, Jessica Lee, and some modeling by Siobha Shriyazi, a graduate student I share with Chris Rameen, and then little bits of things were from Tomaslav and Janelle, two other postdocs, as well as some former members of the lab. And all this, rather than talking about kind of old things, I've decided to also talk entirely about new stuff. And there are three parts to this. I'm gonna try to keep them really simple and just kind of convey a main message for a few of the parts and really get into the third one in a bit more detail. And so kind of, you know, the primo, segundo, you know, the go through a series of plates. So first, I'm gonna talk a little bit about communities, since this is a meeting about communities, and talk about toxins in the context of a community and how they can be made as some substrates are used and hurt the organism that's making them as well as go out there, could hurt other organisms, but some of them might actually be able to help the system from kind of a practical standpoint. I'm gonna zoom in and talk about a molecule that can actually sense these toxins and talks to the rest of cell physiology in a really unusual way. And then we're gonna really zoom in to the main thing I wanna try to talk about is heterogeneity and phenotypic heterogeneity and are kind of surprising, finding for at least surprising for us that that plays a critical role in terms of which of the cells are growing or not within this sort of scenario. And so, first of the organism in the compounds, so the organism is methylobacterium, and so this is a, it's in the bradyveraizobium group. It's basically a plant-associated microbe, but it happens to have kind of an unusual metabolism that can eat single-carbon compounds, and it's been a model system for about the last 50 years for this kind of lifestyle.  It's already a hint for what the bad compound decom will be. That can get oxidized through a pathway that depends upon a nice simple cofactor named tetrahydromethanopterine, try to say that. Oxidized through the formate, and that can either get burned off to CO2 or assimilated through kind of a modified amino acid cycle into biomass. And so all organisms have to do some of these reactions to make intermediates, to make amino acids, to make DNA, et cetera. This is their assimilatory main pathway, main dissimilatory pathways for growth if they're eating something like methanol, methane, et cetera. And there's lots of ways, both in terms of kind of applied aspects as well as environmental aspects, that C1 metabolism plays a critical role, both in terms of like micro-plan interactions. A lot of these are growth-promoting microbes in terms of things like oxidizing the methane that would otherwise be released into the atmosphere. And there's a whole bunch of interest in biotechnology based upon using methane or methanol as a feedstock. But I'm not gonna hit those topics, but I have to talk about it. And then of course the compound here is formaldehyde. Every single carbon when you're doing this goes through formaldehyde. This would be like if your diet was neat ethanol, right? Where then every single carbon that you ever took in went through acetaldehyde as an intermediate. This is a pretty hardcore way to go. And so this formaldehyde is produced at two millimolar a second. This would be the concentration that it would increase per second if you kept making formaldehyde but didn't use it. The briefest period of time you'd become a chip-chip experiment, right? Everything would be cross-linked to everything else. How does this not happen? And we're talking seconds, not minutes, not hours that it would take for this to occur. And so interestingly if you look at their growth and I'll come back to these aspects later, if you just throw in kind of outside formaldehyde they can take it up to about two millimolar no problem. Three millimolar begins to impart a little bit of a lag. But beyond that, they just will never grow. They don't grow. A little bit of interesting things that the board will talk about in the last part. And above about five, you get rapid death. And so this is a log scale here in terms of CFU per mill, just over a few hours in a concentration-dependent way, they die in a curve that look a lot like Bruce's work on antibiotics, for example, just they get killed. So it's definitely quite a challenge even for an organism that has this as its lifestyle. And so let me first mention the bit about communities. And so the community context, so this as well as some of the other later work was done by Jessica Lee. And a surprising place where you find C1 compounds is actually in plant biomass. There are a lot of methoxy groups in plant biomass because lignin is kind of a really complex polymer, you know, branched and non-repeating structure. And many of these different aromatic groups, they're all these OME, these ME are all methyl groups. And it's about 10% of the carbon in lignin and lignin is about say a third of a plant. And so it's actually a fairly sizable part of what's going on there. And critically for organisms that are degrading lignin and are trying to eat these different methoxylated aromatics, when they do so, take something like vanilla acid and convert it into proto-catechemic acid, this comes off as formaldehyde. And so they hurt themselves and it gets released into the media. And so it's a real challenge. The organisms universally will eat these compounds faster than those because they're making this nasty compound on the inside of themselves. And so on the inside, that's a private bad, right? But once it's outside, now it's a public bad. And any organisms in that community, whether it's a natural community or in these case kind of model practical synthetic ecologies would have to deal with that formaldehyde. And so in a project spearheaded by Daniel Segre and also involving Trent Northern, we have been looking at using a few different organisms. I won't get into all the details. We'd use organisms that degrade either the cellulose and hemicellulose or degrade the lignins, deal with this sort of aspect, feed organic acids to something like uroia to get nice and fat with a lot of lipids, make biodiesel, make DOE happy, we get funding. That's basically the cycle of things. And so the role for methylbacterium is here in terms of either being the one to take off these methoxy groups or to deal with this formaldehyde. And we've done some work in terms of this first bit but I'm not gonna talk about that today. But just I wanna briefly mention about this idea of being able to kind of purposely engineer into a synthetic ecology, a scavenger for this type of toxic molecule, the removal of a public bad. And so all I'm gonna do basically is show the kind of result that we can get from this, that if you look at growth, for example, of pseudomonaspeuta on vanilla acid alone versus in co-culture with methylbacterium, and this is formaldehyde buildup, having the methylbacterium around, not surprisingly, can dramatically prevent the formaldehyde from building up. It can speed up growth in the community. And when we go to then these higher order pairs and trios and quartets of interactions, having all the species present, including methylbacterium to do this detoxification, allows the entire community to grow much faster into higher yield. It turns out there's actually even some of the other organisms that I had on the last slide are even more sensitive to formaldehyde than either putida or exturcans are themselves. And so just wanted to make the simple points, when we think about metabolism, we often think about these excreted metabolites as being good things. pH as well as things like acetate were brought up as kind of these dual acetate can be a food, but it can also be a problem. Formaldehyde is really a problem, very strongly. And so it's a relevant challenge in this kind of system. And here's an example of having the removal of a public bad be rationale for inclusion in this kind of a system. So that's all I'm gonna talk about in terms of the community bits. So for a moment then to zoom in on how do these cells deal with formaldehyde. And so a really amazing former graduate student of mine, Dipty Nyak, she and I, we were puzzled by this fact that these guys grow at this amazing rate, two millimolar a second. And yet if you give them anything more than about two millimolar in the media, they really struggle and why is that? Could they get better? And so we decided to try to evolve them to grow and to grow on formaldehyde alone. And so just in a very simple kind of pilot experiment, just took a few populations evolved for a fairly short period of time on formaldehyde. We started on methanol and then put in some formaldehyde and kind of like Roy's morbidostat, we kept marching the formaldehyde up, up, up, up, making it harder on them every single batch. And now we were able to evolve these populations in a very short period of time to grow on 20 millimolar formaldehyde. That's the same concentration of formaldehyde that killed 10 to the seventh of the cells in two hours, now they're growing on it, right? So yes, this is possible. Amazing, what is it to, and they can actually grow on formaldehyde faster than methanol, these strains. So what did they do? We know so much about methylotrophic in this strain, had all these guesses. When we did resequencing, as is often the case, nothing that we knew about came up with any mutations and instead two loci in all three populations had different independent mutations. And of course there were genes of unknown function. Had no guess whatsoever what they were. I'm gonna ignore one of them for today. But the other, so we called these EFG, for Evolved Enhanced Formaldehyde Growth, this one really caught our eye because it is only found in methylotrophs, yet methylotrophs all over the phylogenetic tree, from alpha, beta, gamma, delta, et cetera, but even NC10, like one of these deep divisions, et cetera, only methylotrophs have this gene. And so that correlation between lifestyle and this made us feel like even though this formaldehyde selection was kind of a crazy ridiculous, just sadistic thing to do to our populations, clearly we found a part that has some relevance. We just have no idea what it is or why it would be relevant. And so in terms of very simple genetics, we could of course swap the alleles, the evolved allele in with the wild type one, or put the evolved allele into the original strain, and it causes you to either lose or gain from aldehyde growth. And so these alleles were necessary and sufficient for that to occur. And then we also made a deletion strain while we were at it, and that also allowed you to grow on formaldehyde. And so it means that these beneficial mutations were actually loss of function. And thus there's a gene, a protein there, EFGA, that actively is preventing you from growing. Why? Why would you have a protein that kind of does this sort of self restraint? What kind of scenario could that be? And one thing we saw is that if you don't have this protein, if we really hit them and with some very, very high level of formaldehyde, cells without EFGA have decreased survival. So there is some kind of ability to grow versus ability to survive sort of trade off, but I'll come back more to kind of what its purpose might be. So what does it do? In the past, this would be my entire 50 minute seminar, so I'm gonna put it here in four little panels. With a use of shimu down at rice, determine the crystal structure of this protein, because we had no idea what it was. And the structure made us realize it looks a lot like an oxidative stress sensor from some gram positives. But even though the amino acid similarity is very different, so maybe it's a sensor. Very remarkably, if you put just EFGA into E. coli, you can move the phenotypes with it. It will cause E. coli to stop growing on sublethal formaldehyde concentrations, but will increase survival of E. coli on lethal concentrations of formaldehyde all by itself, which also suggests it doesn't have kind of any particular methyl-by-caram specific partners, and must interact with something in a more general way. And from further evolution experiments, if it was fun the first time, why not do it again? We did a larger number of these. We found a whole bunch more EFGA alleles, but we also found mutations at other loci, including mutations in peptide deformalase.  I haven't talked about the ribosome, and certainly this was not at all in our ballet week, in terms of what we were thinking about. And as it turns out, EFGA binds to peptide deformalase in a formaldehyde dependent manner. The beneficial mutations all abrogate that interaction, and this causes translation to halt instantly, faster than our positive controls. We used some various antibiotics. They took a while to make the cells stop translating. With EFGA, that was active, you add formaldehyde, translation stops.  Whereas if you add something like canomycin, it takes a while for it to stop, and death happens rather quickly. So it was really fascinating. And so why would you have this, right? What's the advantage of stopping translation that doesn't seem like that's a good idea, right? It's like, please stop paying me my salary. Like, why would I ever want to say that, right? What could possibly be a rationale for this? And our first guess is it might have something to do of a growth survival trade-off. Maybe something like antibiotic persistence. And which is the idea that some cells phenotypically are actually tolerant to antibiotics, and they can switch back and forth between growing cells that are sensitive and non-growing cells that are resistant. And you see these sorts of kind of classical dynamics. Almost all of those mechanisms of persistence involve the ribosome. And so we thought perhaps EFGA is doing this sort of thing that maybe it creates some sort of persistent-like state. And so if you add formaldehyde and look at the kinetics of death, sorry, this is a little small, this is three hours, and this is a log scale again, going from like 10 to the eighth to non-detectable, you see killing that's kind of tri-phasic.  So it looks like persistence. There's this second tail. However, EFGA had nothing to do with it. You get this in either wild type or the strain lacking it. So it's neat, something else is causing this, but that's not what EFGA is doing. So unfortunately Roye's gone today, so I don't get to again give him credit for his really great paper that made us think about this direction. He showed that translational inhibition can actually speed up growth. Now you know, who of you amongst, you don't have to raise your hands, but thinks that that sounds like a crazy thought, right? How could you grow faster if I slowed down my own translation? So they studied antibiotic suppression. And antibiotic suppression is the idea that one antibiotic can actually help you grow on another. And it turns out this only really happens, and Bruce can correct me if I've gotten some of the details wrong. Only really happens between one class and another class, and it's only one direction.  And neat thing about it is it's generic. You can use essentially any replication inhibitor and any translational inhibitor, and you will see this sort of effect. And you can see this, like with growth rate in the Z-axis, you can go above the MIC on Trimethoprim, and as you march across levels of Spearomycin, you go from being not growing to faster, faster, faster, to the best you can do and back down. And this is, like I said, is stereotypical. Now, why is this? It's because the cells, even if they're not replicating, E. coli is actually pretty dumb when it comes to their ribosomes. They will continue to translate and they become longer and longer and longer. And so, with any replication inhibitor, you get these elongated cells, and here's a distribution of cell length, it doesn't matter which one you use, whereas if you use a translation inhibitor, of course, they're not translating, they don't get bigger, but they don't always keep dividing, right? The opposite is true. They're smart enough to know not to just divide every 30 minutes no matter what's happened, but they're not smart enough to stop translating if they're not replicating, and thus they elongate. And you can see this massive elevation in protein per cell. And the idea is that this unchecked translation itself is bad, right? Imagine you can translate a rate 10 and you can replicate a rate 10 and it's fine. You're in a steady state, you're happy. If your replication is now inhibited to five, but your translation won't slow down, they're both positive values, but you're not at steady state. You violated FBA assumptions and a whole lot of other things and you're out of whack. It's actually better for you to slow that translation down to match. So at least you can grow even though it's at a slower rate.  And that's exactly what they see with these sort of dynamics. And so the hypothesis was, even though E. coli never evolved to kind of, these at least particular drugs that we're using that we hit at them in this sort of way, that EFGA may be functioning like an internal antibiotic, so as to say, that does exactly this, that tunes the cell's translation to the level of damage being created by formaldehyde. If formaldehyde behaves relatively like one of these replication inhibiting drugs. And so I'm not gonna get into all of the various aspects about that. We've seen sensitivities to these various sorts of drugs, swap between the different genotypes. We've seen that EFGA, this protein, can suppress trimethyprim. So we get the exact same sort of chart as over here. So our protein can work just like something like canomycin. And then probably the neatest evidence that we have, this was worked with Andreas Vasdekis, a physicist at University of Idaho, looked at, did some image cytometry to look at what happened to the cells. And we wanted to see if you could stop this dysregulation of cell size. Does EFGA actually stop the cells from elongating if formaldehyde ever caused them to elongate the first case? And sure enough, it's a little bit hard to see, but formaldehyde does create these elongated cells that look just like we added trimethyprim or something. And you see this large increase in kind of the median cell size and a huge variation cell length, et cetera. If they have EFGA, this is even in E. coli, it stops. They can make themselves, they have restraint, they don't translate in the presence of this formaldehyde damage. And so these early experiments are consistent with the idea that EFGA functions kind of analogous to a translation inhibiting antibiotic. And so the thought then is, we know, this was the world that I thought was fun and this is in a textbook and I know it's going on in the cells, but that's housekeeping. And other people, of course, study translation and they know metabolism happens because it makes the amino acids that you use to translate, but you kind of ignore that. And so we knew that there was a connection from the point of view of damage, right?  But what we didn't realize is that there was this sort of linkage might be possible in terms of something that would actually directly sense this and directly modulate translation. And to our knowledge, this is the only case in a bacteria where a metabolite is sensed and it directly modulates translational activity. If anyone knows a counter example, please tell me, actually at the coffee break because I'm gonna be gone by the time Bruce is done talking. So by any questions or thoughts, I'd love to hear it, but it has to be over coffee, no, don't have lunch. You'll have to take off, I'm afraid. And so we're certainly pushing to understand much more in terms of exactly what's going on here, how exactly does this cause translation to stop? What's the broad cellular response, et cetera. One thing I wanna mention, there are EFGA homologs in almost every single bacterium. So if you study a microbe, which most of you do here, then chances are you might have a homologue of this. Some of these we've found to be encoded in operons that make aldehydes. And there's actually a lot of things that when they get used, make an aldehyde as an intermediate. So this actually may be a relatively more generic kind of interaction than previously appreciated. And the last thing I'll say too is we've seen because it behaves kind of like canamycin, you get metabolism drug interactions just like drug-drug interactions. So giving a little pre-dose of formaldehyde massively sensitizes the cells to some antibiotics and saves their butt against other antibiotics. So what metabolism is going on in our gut may greatly affect the sensitivity of antibiotics of the same bugs that have, if they have these sorts of systems going on. All right, so the third part is about the populations and finding some heterogeneity. And so I think it's also a lesson about, you only see what you look for and the kind of method that you use. And so we had been doing these growth experiments and we felt really good about ourselves because we were doing really nice fine kind of titration of formaldehyde and we got nice pretty data like these, et cetera. We saw these really long lags. You go to like four millimolar formaldehyde right at kind of the board of what they can take and not take, we thought, wow, look, there's this really long lag and it may be hard to see but this is like two days before they start growing. It's not a lag. It's not really a lag. This is CFU per mill, exponential death and then exponential increase. So they're not just all sitting there. There's massive death happening and then massive regrowth happening. And so we wanna understand what is causing these cells to change really rapidly from this death phase to this growth phase. And so the first easiest hypothesis, especially given EFGA mutations where a loss of function, these are just mutants, right? And it takes you about a day or two. Perhaps, you know, if you extrapolate this down, we're just getting a high number of EFGA knockout mutants or something and up they go. Short answer is no. There are no mutations and we can take these cells, grow them up in some other media, re-test them, you get exactly the same thing again. All right, so it's not genetics. Well, so then what? We thought maybe they're changing their environment. This has also been a theme of the meeting, right? Organisms change their environment. Maybe during this death phase formaldehyde's above some threshold that they can take and we've also heard about threshold tipping points, things like that a lot. If they can get below some threshold between net death and net growth, before everybody's dead, your population survives. If you can't get there before everyone's dead, well then the whole population's dead. And so this seemed like a very reasonable scenario. And in fact, we went through the exercise of trying to really kind of model this out. Don't look at any of the equations, even if you want to, because I'm gonna tell you this is wrong, but I will mention that this was work of CIVOSH and like I said, co-advised by Chris Vermean and another faculty at Idaho. And so we made a model of what was going on and from this model, we could even felt really good about being able to make some quasi steady state assumptions, look at kind of phase planes and null clients and all sorts of funds and groovy things. And our model fit the data really nicely. Now mind you, what's the data? An exponential down and exponential up. It's not that hard to fit an exponential down and exponential up, but this all looked great. Until you do an experiment to ask about this sort of question, what is the basis for why cells would go from death to growth? Is that formaldehyde was going down in the media. And when you measure formaldehyde, it has not yet gone down. This whole transition happens long before when formaldehyde begins to go down. So they are not making their world better on a time scale that's causing this thing. So it looked great, the math was great, wrong reason entirely. So okay, the world's not gotten better and they haven't evolved. They have no genetic change. Then what about phenotypic differentiation? So back to this idea of something like persistence. So persistence like I said is this idea that some cells in the population randomly switch into growing essentially at no speed at all. And it's exactly because they're not growing that they are resistant to things like ampicillin. And then at some rate, they can switch back on to growth. And so that phenotypically, some number can survive a brief enough insult and come back and restart. Now the thing though about persistence is growth is reasonably by stable. Like typically you have the growers and a small proportion of non growers that rare and they're exactly resistant because they're not growing. This clearly can't be that because if it's involved, I need to explain the growth. Cause they're still growing while the antibiotics here. This switching back with persistence happens after you've taken the antibiotic away. And so the thought though is that maybe phenotypically, even though we see what looks like a lag, a lot of the cells may be just sitting there in terms of an optical density standpoint. And there's a rare set of phenotypes that they're able to grow they're different in some phenotypic way and they begin to outnumber and it's not until they reach a high enough number that you see the things take off again. If it was by stable, so let's imagine that you could have some like a persister but like a growing persister. And let's imagine it's one in a thousand and instead of only being able to take up to two millimolar you could take up to five. If we were to plot tolerance to formaldehyde phenotypic tolerance, you know, nearly all of our cells would be tolerant up to two then we'd have this kind of discrete class that could take it up to five and then that's it, right? This is what this kind of data would look like if this was discreet to, you know, bimodal distribution sort of scenario. What we see instead is a continuous distribution that fewer and fewer and fewer and fewer cells can make it to higher and higher and higher and higher and higher levels. So rather than just having kind of like some really discreet class and a lot of phenotypic heterogeneity that's been studied, I think because of ascertainment bias has been really discreet differences. Persisters are really different thing growers. Spoilation is really different thing growth.  And this more like kind of subtle quantitative variation phenotypically is often, I think, a little bit less followed out. And so we see instead, like I said, this kind of continuous distribution of fewer and fewer cells that can actually grow under the exact same conditions that are killing most of their neighbors. How is that possible? Yeah, Carol. Very easily. You have a series of plates with more and more and more formaldehyde and so you take them and you just plate.  How many do you get on one? How many do you get on? Yeah. Yes, you do. It seems to be totally, or massively fast genetic. So I mean, I will say, with a star, right? It could be some unknown, massively fast genetic thing that behaves on the same time scale as phenotypic. Does that sound good? Yeah. Yeah, I think, I would too for now. But frankly, they're gonna behave very similarly to each other. It would be easier if it was genetic because then we could sequence. We have something we can do to get right at it. All right, so back to the model. And so we wanted to ask, you know, if this was the case, what would kind of then the consequences be? Could we understand this from this perspective? And so we had a model that looks at kind of the use of the methanol and the formaldehyde that are present in the media across all the types of cells. And then the reason why this looks all nasty with these integrals and stuff like that is we're now using a PDE to explain the cells where we're following the cells through time and as well as in terms of X where X is the tolerance state. And so we have kind of this continuous rainbow of different tolerances that are around and that tolerance plays into three different parts. The growth rate seems to be relatively independent of the tolerance, so it doesn't show up there. Death, we're having, we can do different sorts of functions. This is kind of just a simple one. If your tolerance is above the formaldehyde in the world, then you don't die.  And then it also shows up in terms of the tolerance itself being able to change. And so this is basically a diffusion and an infection types of terms in terms of this tolerance. Whatever gene expression and such, we can get to kind of what might be going on here in these populations. So this sort of heterogeneity model, again, it can certainly reproduce fairly well the sorts of dynamics that we see including these rapid death followed by regrowth. But the critical thing is it gives us the insight what should we be looking for? And so at the beginning, we're saying that we've got this almost like exponential distribution of the phenotypic states. That I had plotted on a log scale before, so it was a straight line. This is not on a log scale. By the time the populations at Nadeer, then there should be very few, this is in total CFU, left of the sensitive types. But the rare tail of phenotypic types have been enriched as well as you got kind of continued diffusion off of that point. So we should see here kind of a mixture. And by the time this population is growing back up, they should all be in this kind of, this higher phenotypic state. But if you go a long period of time after where formaldehyde's not around, it should relax back to the original standing distribution. And indeed, this is what we see. So if we now do a dynamic experiment of exactly what Carol was asking, here we're just showing the zeroes, twos, and fours. So plate counts at zero, two, and four. Initially, you have this distribution, those that could take four were one in 50,000, something like that. And within a short period of time, kind of at the dip of viability, all the cells are of type four. You get the exact same counts on four, two, and zero. Every single cell there that could ever make a colony is able to make it even with four millimolar. And then that persists up. And then this goes away quickly. And so the sorts of questions that Bruce asked, not gonna show all the data for it right now, but if you take them immediately after recovery and you do the experiment again, they just grow right off. If you take them, you put them in succinate for a small period of time and grow them up, retest goes back to the original experiment. But, and if you do it at like short, soon after succinate, they're like intermediate. And so there's a time scale of a few generations that they have that keep some of this inheritance, but it will relax back to this initial state. This fully enriched population, because there's so many of them, you actually see some that can resist six and resist seven. So there is like an extended tail even further that would have just been too rare to have seen in the first population. And like I said, they rapidly lose this tolerance, but not instantly, if you take, put them in media now without formaldehyde around. And so, I'm making this case that it's phenotypic heterogeneity and it's rare cells that are actually doing this. There is an alternative hypothesis. And maybe some of you watch this show even all around the world. Anybody recognize this show? Right? So with zombies and the living dead and whatnot. So what if these cells are just partly dying? And then here they're undying, unpartly dying, right? And it may sound silly, like what do I mean by partly dead? Microbial ecologists have a word for this, right? Viable but not culturable, right? That they're there, they may be metabolically active, they just won't make a colony. Like asking them to make a colony is really, like that's a hard task. And we often forget that not every cell that's alive will necessarily make a colony, right? The plating efficiencies may change. And so if they basically have lost their plating efficiency, most of it, and then regain their plating efficiency, that could also explain this sort of dynamic. And thus this recovery phase is just the zombies going back to the regular humans. And so how do I really know that they're rare? And until yesterday I could only show one type of data for to be able to get at this. So there's an excellent assay that I learned about here three years ago. Matthias Heinemann gave an excellent talk about transitions and used exactly this assay. And thankfully I actually remembered that much later and we could give it a shot. So he added a membrane dye that interpolates into membranes but won't go cell to cell. Like you dye and the D-Y-E dye the cells and then you take away that and then you start growing them. Each time they grow, you have twice as many cells that are half as bright. And then you have four times as many cells that are as quarter as bright, et cetera. And so if it's just all the cells start growing at the same time, you'll just have this nice unimodal distribution fluorescence measured by the flow cytometry, nothing surprising. However, imagine the majority of the cells taking the dye and they never grow at all.  And instead there's a minority of cells that are somehow phenotypically different. They're gonna start to lose their membrane dye long before you can ever see them in the population if they were initially rare enough. So by the time you see them, they're gonna be a little bump in the distribution far out to the side and the original distribution will just get outnumbered and those new cells will rise up in number and also I continue to kind of finish diluting out the membrane dye that they have. So it's a really lovely assay for being able to, in a simple way, look at phenotypic heterogeneity. And so this is what it looks like for just a plain experiment. Just grow cells on methanol from time zero to 53 hours, they were all bright. They became less, less, less, less, less. Nothing very exciting at all, right? But for this experiment, we see exactly what I was mentioning. The original time point, this fluorescence stays basically exactly the same and just gets lost and instead you see this little bump that rises, like you can see this blue trace, right? And rises and rises and as it's rising, it's actually continuing to dilute the dye out but you can see it popped up all on its own. It was a rare small number of cells that were actually able to make this sort of transition. And we're saying, you know, our numbers seem to be something on the one in 10,000. If it's harder than that, even one in a million cells that actually allow the population to survive and yet it's just a one in a million phenotypic. And then just yesterday, I finally got images from Andreas Fasteckis. We've been doing this with using automated microscopy and this just shows one particular field. And so you put cells down just on methanol and essentially every cell will grow up and make a nice little colony and have a wonderful little life for themselves. However, if you put this two and a half million molar formaldehyde, only a very rare number and it's exactly the same numbers. It was about 2% of the cells here. And we also had an estimate from the plating of 2%. And so direct observation in this. And the other nice thing about direct observation that you never get with plating is what happened to the cells that didn't make a colony? Did they like have so much formaldehyde damage, they managed like a couple of doublings and then just kind of ground to a halt? Did they at least try to extend their cells and maybe get all messed up in their morphology? Turns out they do absolutely nothing. Those that do not make colonies because their translation is probably halted, they never change the way they look at all. And so we get that kind of information from this sort of assay that we never would have gotten from the dye assay, of course. And then our last approach for quantifying is we're gonna use a barcoding technique, put in a population of barcodes and see how many barcodes survive various sorts of bottlenecks. And I think that'll be even the most quantitative for being able to hit really high stress levels. And so what I think is really fun about this is I think it's really shows that kind of a minority of cells can be incredibly important for what a population is doing. And in this case, it was really just these very, very, very few, these ones that can make a colony that end up making this sort of bump that allow these cells to be able to navigate these sort of transitions. Pankaj alerted me too, there was a paper about cancer cells and anti-cancer drugs where this sort of phenomenon has even been worked out mechanistically, where there's a minority of cancer cells that have an expression pattern that allows them to resist a higher level of drug than the rest. That's the kind of direction that we wanna go in terms of, of course, understanding what the hell's going on here, what are presumably either the regulatory circuits or DNA methylation patterns or something, like what is the basis, of course, of being able to create this heterogeneity. Yeah, Wenying. Oh, that's a high, this is a four. The experiment we did was at only two and a half percent. It's a challenge, unfortunately, to directly observe them. You want it to be rare, but not too rare. So we can't hit them as hard as we want to because we're mean. And so like, I'd love to see them at four, but I can't ask Andreas to look at 100,000 cells to find the five that are growing. So we had to back down for that. EFGA, yeah, we're making a construct for that. So like, yeah, the ideal is to, so the nice thing is we can enrich phenotypically for a population that we can even just use bulk transcriptomics or proteomics and look, what's different? How are they different, how are they similar or different to being genetically resistant, the Delta EFGA strain, for example? And we can try to understand, like, what's our marker for this? Once we have a marker, then we make a fluorescent reporter go and look in single cells. Because I want to know, two minutes before we hit them with formaldehyde, what made you, what can I predict if you're going to survive or not, right? That's where we want to go with this. But right now we don't, so EFGA is guilty just, all right, like just by association with these sorts of things. And I think it's possible, or it may be more complicated, it may be the tails of the dystopic cells that have abnormally low EFGA and abnormally high peptide deformalase.  Like they have to, EFGA has to stop all the peptide deformalases from working, probably. And so like that ratio might be, who knows? Yeah, great question. And I think last kind of soapbox point is, you know, in terms of thinking about how common this heterogeneity is, so this first paper from Matthias Heinemann's group was using the extraordinarily exotic organism, Escherichia coli, and the transition from glucose to acetate, right? And saw in fact, there's a huge long lag and like 100,000 cells will make the transition from glucose to acetate. This doesn't sound like the hardest thing in the world to do. And yet it's very, very rare. Showed up as a lag, but it's not really a lag. It was some cells were doing it and some cells never will and they have excellent work in terms of understanding why that was true. And the thing that's really kind of exciting or demoralizing, however you want to think about it, is myself and others have done a lot of work studying this transition, for example, from succinate to methanol and looking at fluxes and transcripts and proteins and metabolites and blah, blah, blah, blah. It turns out only about a quarter of the cells go through that transition. So during those, the eight hours of transition, what have we really been studying? We're studying three quarters of the cells that are going nowhere and one quarter of the cells, they're not in lag, they're growing. And so it's really, it's the linear interpolation then in terms of one quarter growing cells and three quarters of whatever. And we've been, we see various things spike early on and like, oh, these are the genes important for the transition. No, that's actually what like, you're never gonna grow again, looks like. And so I think the way we interpret and think about things is massively different, even in relatively more mundane sorts of transitions. And so, again, just a number of excellent collaborators both at Idaho, Yusuf Shremu in particular on this project, Daniel on other projects, et cetera, thankfully some people who give us money and happy to take any questions.


Gain nice smoke Salutations Salutations for our loved ones Salutations to those who can't be with us Salutations for those who might be on their way From our existence from our reality Salutations to you Salutations to freedom


Hi everyone, Joe for Jaspy's casebreaks.com coming at you with 2019, 2020, Panini, Daunress, Optic, Basketball, Mega Box Edition, four box, pick your team number 11. Two ones right there. Number 11, big thanks to all of these folks for getting into the action. I think Ryan got double last spot in Mojo with the Clippers and Timberwolves right there. We're giving away the Pelicans in this break. Now a lot of times you see two entries, two teams, one entry, four teams, two entries, so on and so forth, right? This one, if you read carefully, one spot is one entry, two spots, two entries, so on and so forth. So let's grab all of your names right here. Let's take out the little placeholder for the Pelicans. Let's roll it, let's randomize it. One and three, four times, name on top. After four, it gets the Pelicans. One, two, three, and fourth and final time, four. Thanks everyone for giving this a shot. Name on top, Kang, there you go. Nice, you have the Pelicans. What's your name here? There it is, you bought the Kings. You still have the Kings, ended up with the Pelicans. That's not too shabby at all. All right, there it is, 2019, 2020, Donner's Optic Basketball Mega Box Edition. We've got four of these. Is that a 75 orange, or out of origins? I think you could probably get away with a 130 top loader, Ken. Are there usually, I don't remember these mega boxes, are there usually two? All right, good luck everybody. We got a hollow rated rookie, Terrence Mann. That'll be, oh, can I put this out? That'll be for the Clippers. That'll be for Ryan. There's the official printout right there. Whoa, they're taking Clevenger out. I kind of feel bad for the guy. I think, what do you have, like an elbow or shoulder thing already? A little Dodger's Playoff game on the background. There you go, that's for the Knicks, Brenton. All right, and all of these parallels, of course, these pink hypers will go. There's another Terrence Mann right there and a Darius Basely for OKC. You had a pretty good time in the bubble, John. The Thunder, nice. And Bam out of bio, I think they're in half time right now. My Lakers are playing as well. A lot of sports happening, folks, a lot of sports. I guess there are two bonus packs in each of these mega boxes. Oh, they're back already. We need to play off Rondo, Lakers down a couple of points. Miami up 54-52. Nice hollow Yanis right there for the Bucks. That's gonna go to Blaine. Even the second year Luca Donchich cards. I know we're doing all right in the secondary market, Danny. Especially up when the season kicks off again and Shabazz Napier in the back. Ooh, nice steal. Jared Culver could be on a team with, spend all season with Carl Anthony Towns, the Andrew Russell, the number one overall pick. Could be good for his stock. There's Darius Basely again. Culver going to the Timberwolves. That's gonna be for Ryan. All right, a couple more here. Ricky, what's up? Your hands hurt after opening so many packs. No, no, I got strong hands. Ricky, I got strong hands. Also, I play like musical instruments, like guitar and stuff like that. So I've always used hands to do things. You know what I mean? I was built for this. Case-breaking machine. And I do a lot of hand stretches. Try to keep the hands loose. Gotta keep the hands loose. You work in IT, so you're soft is what I, I didn't say you were soft. I'm just saying anyone can improve their hand technique. I just happen to get lucky and have a lot of background that will help this job. I couldn't do what you do. All right, there's a hollow Brandon Clark and a Kobe White and a Tyler Hero, nice. Miami Heat, Kevin Mendoza with that one. How's, what are, what's Tyler's stat lines right now? Tyler Hero's got nine points, plus minus of minus 11, 0 for three from three. Come on, Tyler Hero. Lakers defense is too good, but I want Tyler Hero to play well and lose as a Lakers fan, but also as someone in the hobby. Kobe White goes to Chris Gerard, Tyler Hero going to Kevin, another Yanis right there. Bam out of Iowa, coming off that injury, 13 points already, Jimmy Butler has 14 points. LeBron, Danny Green, KCP, all in double digit points. All right, maybe some pink hyper LeBron, or LeBron would be nice, but John Moran's and Zion's, that'd be nice. Ricky, you're waiting on Tyler Hero, RPA Redemptions? Well, he's been busy. I'll cut him some slack on that. If he wasn't in the bubble or the playoffs, then I'd be like, come on, guys got to sign his cards. All right, here's the fourth and final mega box. We have some more, so we might be able to run this back tonight if you want to. Good luck, last box, 2019, 2020, Panini Donner's Optic Basketball, four box, mega box, pick your team number 11. These packs are a lot of strong glue on them. All right, there is a My House, John Moran. That'll be for the Gris, that's going to be for Chris Gerard. Why not take another three, LeBron, heat check? He's going to do it. Take the three, he's going to drive. Hey, there's LeBron James right there. That might be a parallel too. There you go, that goes to Christopher P. and the Lakers. Kevin Porter Jr., I think he's certainly got a lot of upsides. If I'm, Cabs, Kevin Mendoza, I would hold on to that guy and see what he can turn into. Townhorn Tucker, are you going to get some minutes tonight? Has not entered the game yet? Man, you never, what if he turned out to be the reason why the Lakers win this game? Game four. There's Deandre Hunter and Danilo Gallinari. All right, so no Zion, only a Ja Moran, insert. So nothing too crazy there, but I think we got some more that we're going to post in the store. So check it out on jaskycasebrakes.com and let me tell you, Townhorn looks surprised in that photo like he wasn't expecting it. He's like, there you go, it happens folks. Four box, pick your team number 11 in the books. I'm Joe, I'll see you next time for the next break.


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This is covering the spread part of the Fandall podcast network We have got some fantastic games on tap across week 14 in the NFL We have got the Cowboys and the Eagles chiefs and bills with big playoff implications Both in terms of seating and he will make in the playoffs as well We're gonna break down those games plus others to get you ready for week 14 by talking to Dr. Ed Fang and picking his brain and what his number say about week 14's games This is covering the spread right here on the Fandall podcast network and Fandall research My name is Jim Sonnis I am a managing editor of digital media for Fandall research joined here as I am every Thursday by Dr. Ed Fang you can find his work at the power bank comm check him out on Twitter at the power Icon Ed didn't get to talk to you about college football this week because we were into the postseason there now But obviously a very interesting weekend across the conference championships in the playoffs election with Michigan Getting the one seeds. How are you doing today? I'm doing great. It was nice to see Michigan get a win. It was kind of a weird game against Iowa I actually thought they I thought it was worse looking at watching the game I thought I was defense played really well really bottled up a bunch of things that Michigan wanted to do but then I looked at the metrics and like I Mean, I didn't really change Michigan at all based on what happened And then you consider the pretty sure-handed Colson Loveland dropped a couple third down passes that could have really changed You know the narrative in terms of you know when you were watching the game, so yeah, obviously all the fireworks happen Sunday Yeah, all the fireworks happening Sunday with FSU being omitted despite the fact they won their conference championship They were 13 and 0 and and you understand why for sure and You understand why you understand why FSU fans be very frustrated You get the argument from their behalf and I guess Ed It's situation where I get every argument here and I do feel bad for FSU people I understand people saying they should have been let in because of what they did but also like as a Very selfish more casual college football fan. I'm a lot more intrigued by Michigan, Alabama than I am by Michigan FSU personally, so I see it from all sides But like just like a football watching perspective selfishly I kind of did want the committee to do what they did Exactly Michigan, Alabama's I mean, it's essentially a coin flip. Yeah, I think that's what the markets are saying I think that's what my numbers are saying and it's gonna be exciting Florida State look the committee has always talked about putting the four best teams in there And they've never really put the four best teams in there but the fact that they've been talking about putting the four best teams in there kind of gave them in and out with You know, let me Florida States not one of the best four teams right now In fact, they weren't one of the four best teams with Jordan Travis. I would make the argument But that that's a discussion for another day You know, they did put the better team in there. So, you know, I guess hats off to them I understand the frustration with Florida State I would not be happy if I were someone like but Elliot whose income kind of depends on You know a Florida State podcast and you know interest in that and then Yeah Anyway, I like I think one thing that is pretty interesting I think we have a lot of people complaining about like 12 team playoff and like oh, there's gonna be a three-loss team in there Yada yada yada, but I actually don't hear anyone saying man, this four team playoff sucks. Let's go back to the bcs Let's just have Michigan Washington in a game. Like I don't hear that. Maybe I'm missing it Maybe I'm not scrolling my my Twitter screen enough, but we're gonna get to the playoff next year It's gonna be great. It's gonna be I mean just the bonanza in terms of money. It's great for my business It's great for your business gym, right? Everything's gonna be good. No one's gonna be like, hey, let's go back to four teams Um, so, you know, we just gotta wait out one more year and it's it's gonna be fun You know, this is kind of like my glory week, uh, out of like my entire career doing college football because I've decided not to put too much effort in the bowls really just not my Strong house, you know numbers are just not going to be the best when there's so many opt outs and late breaking news It's essentially become like betting NFL preseason Which is something I don't do Next year when the playoff comes going to be really busy this past week Uh, getting ready, you know getting ready for the playoff, but this is the one year It's kind of like, yeah, this this has kind of been a nice week Of course, uh a whole bunch of NFL stuff like dropped in my lap this week So it's not like I was doing nothing But it's an interesting year out of You know the decade Past and hopefully decades future that that I do college football. It's an interesting year Well, like the 12 team playoff like I get, you know, again I understand what people might not want it and stuff like that But like from my perspective like there were a lot of fun teams that were ranked 5 through 12 this year like George is a good football team and they were the six ranked team Like there are a really good football team Do I want to watch Georgia play more meaningful games? Heck, yeah, I do like absolutely that'd be a lot of fun Ohio State a good football team I'm assuming if they had been the playoff Kyle McCord would not have entered the transfer portal at that time Bo Nix in Oregon would have been ranked eighth like That's fun setup. Like that's some fun football teams and like I I get it from a lot of different angles like it does D value regular season a tiny bit compared to what it is right now But like those games will still matter a lot for seeding perspective like those those buys for the top four seeds Are going to matter a lot. So I I'm just excited for it. Honestly. I think it's going to be great getting to watch more Brock Bowers and meaningful games like I think that that's all that's all super positive. So I I'm just pumped for next year it's bummer for teams like fsu, but like You know, we're getting more fun games this year. We're going to get a lot more fun games next year, too So I'm all in favor of all of it for sure Absolutely I mean, I do I do get that it devalues some of the regular season games and Michigan Ohio State this year was truly special because everything was on the line for for both these teams But it doesn't mean that people aren't going to watch it next year. It's still going to be a good game between two great programs So I think I think I think people like to complain about a lot of things. Um I think next year is going to be great It could still mean a lot too Like if one of those teams the one loss team and a second loss knocks them out Like knocks them out of the big 10 championship game when there are no divisions Like there are a lot of ways those games can mean a lot even if it's not, you know, single loss elimination type things So that'll be a whole lot of fun for sure next year and the more immediate future We've got some nfl to break down in week 14 And like I said really fun slate will break down here and just one second But first a reminder to make sure you're subscribed to covering the spread wherever you get your podcasts are Preview of the banger game between these Steelers and Patriots is already up with Tom Vecchio Breaking down his thoughts on a that game for Thursday night football That is in the covering the spread podcast feed and the fan dual tv plus app as well Tomorrow we're talking to tom as well for some nfl props austin castle swing by to break down epl uh match week 15 as well getting more soccer on the show Once again all right here in the covering the spread podcast feed the fando youtube page and fan dual tv plus Score early this nfl season with fan dual america's number one sports book right now new customers Get 150 dollars in bonus beds with any winning five dollar money line bet That's 150 bucks if your team wins if you've been thinking about joining fan dual There is no better time to get in on the action the app is so easy to use There is a wide range of betting options including spreads player props totals and more So visit fan dual and kick off the nfl season fan dual official partner of the nfl Must be 21 plus in president select states fan dual is offering online sports wagering in kansas Under an agreement with kansas stark as you know llc first online real money wager only Five dollar pregame money line wager required $10 first deposit required Bonus issued as non-modrable bonus bets that expire seven days after a seat See terms at sportsbook dot fan dual dot com gambling problem call 1 800 gambler or visit fan dual dot com slash rg In colorado iowa michigan new jersey ohio pennsylvania illinois kentucky tennessee and virginia call 1 800 next step Or text next step to five three three four two in arizona one eight eight seven eight nine seven seven seven Over the ccpg dot org slash chat in connecticut one eight hundred nine with it in indiana One eight hundred five two two forty seven hundred visit ks gambling all common kansas One eight seven seven seven seven seven zero stop in louisiana visit md gambling health and organ maryland 1 800 gambler dot net in west virginia 1 800 five two two forty seven hundred in wyoming hope is here visit gambling helpline m a dot org Or call 803 to 750 50 for 24 seven support in massachusetts or call 187 70 hope and why or text hope and why in new york Now i mentioned before thursday night football at the topic i want to touch on at least partially here ed because looking at week 14 There are a lot of super super low totals It's not just that game but also the browns jags game total is 31 and a half for that game right now and For most this year my model has been able to keep pace with the market in terms of having very low totals I've had a lot of the lowest totals i've had i've been this year but This week i'm showing value a lot of overs and i don't know if it's like Unders just got a lot of steam or whatever it may be But i want to ask you to kind of get a sanity check here to make sure i'm not losing my mind because i like a lot of He's overs but do you have any interest in betting toward the mean with these outlier spots or Is it hard to grasp how bad some offenses truly are right now? Yeah, i think it is hard to grasp how bad offenses are and core back injuries are Definitely the first culprit everybody came on my podcast the football analytics show and i asked them what his biggest takeaway from the season was and and he talked about how offensives sucked and He talked about a lot of different things but one thing that really stuck out is that teams are playing light boxes more on defense and Offenses are not being as efficient running against those light boxes. So maybe that's noise but that It's it's an interesting little fact and stuff that i don't think you can look up publicly I think edward has access to some nfl next gen data So again, you know, we're kind of playing the signal versus noise kind of thing and I went back and looked at some of my numbers and it's not just a decrease this year it's kind of a Decrease since 2020 you look at how points per game have dropped from 24.7 to 21.7 this season Success rate has dropped every single season We're looking at 42.1 percent passing success rate compared to about almost 46 yards for past attempt has dropped to So it's overall Decrease in efficiency overall and the passing metrics that I use I don't know bad ball rate is up to it's 12.2 percent compared about 11.6 percent over the last couple of seasons So there probably is some systematic Thing going on with offenses being less efficient And I don't think it's necessarily just this year with the quarterback injuries I think it's an overall trend over the last four years. It'll be interesting to see the league Changes rules or whatever to to fix it. I do think there probably is some signal in the decrease That certainly doesn't mean you shouldn't bend over if your model is saying that But I think the overall trends are kind of pointing down and and not just the season Yeah, definitely and I think that for me that's part of why I've been okay with having Having my model say it should have valued unders previously is because like okay I can project for a back of quarterback and stuff like that and to this point the year was never an issue It's just like this this week specifically like looking at totals for this week Like it's a lot of overs and it's not all overs and low totals like I have the over on C ox 49ers or 46 and a half. So like that's a that's a higher total too, but It is like The the jags browns game. It's gone up a point.  So I mean 30 is just so low, right? It's the lowest in like decades. Yeah And like defensive touchdowns can happen so like Yeah, you know, I just I'm having a hard time getting there despite the fact like my model knows how bad these quarterbacks have been I can project Bailey zappy to be terrible Um, and it's still showing value in the over but I don't know It's just it's tough to get there for a lot of these. So We'll see how it goes. Luckily Ed. We're not talking about those games We break down games for today And so we're talking about some fun games beginning with the seahawks in the 49ers right now The 49ers are 10 and a half point favorites in the total in this game is 46 and a half We just saw these two teams square off back on Thanksgiving in that game 49ers took care of business covered the spread They scored 31 points. Can they do so again? Given this lofty spread here Yeah, I don't know. I mean the Niners have been really on a heater ever since the Cincinnati game over the last four games They won every game by 13 or more points and um So it's not surprising that this spread would be 10 and a half And it's not surprising that my model cannot catch up with it because my model tends to assume more Regression to the mean I've Niners by about eight Um, you know Seattle is the definition of NFL average I had them within a point of NFL average this pre season and Both my market model based on closing point spreads adjusted for who you played And then all the data from this current season they all say that Seattle's with within a point Of of of NFL average. So let's just assume, you know, they have a rating of zero This is a division game. So I'm giving about 1.3 points to The Niners for being at home. So you have to believe that the Niners are Nine or more points better than NFL average That's a tall order. I mean, that's that's like some of the best teams that we've seen Over the past decade. Could they be one of these teams? Sure, maybe but um, you know This is a big spread. I like Seattle plus 10 and a half. Um I I can't I can't quite get there. Seattle is not a terrible team Uh, the Niners, I I just don't think are quite that good. I do think there are some out here And I think there's also a bit of like recency bias Because we did just see this game and the Niners covered there They I think it was seven and a half was the spread for that game. They won by 18 in that one Um, and then they beat the Eagles in that high profile game pretty handily On the road there now they come back home Seattle's on extra rest. That's a benefit in their favor as well So a lot of factors pointing to the seahawks in this game I guess like It is tough mentally to bet the seahawks given how good the formula is Look, that's kind of the point, right? Like is your betting against the market and like we have those biases We're betting and people are reluctant to bet the seahawks here Then that may imply there's value in the 49 or there's value in the seahawks because people are reluctant to bet them No matter what the number may be Yeah, absolutely.  I think they're I think they're uh, good They have a lot of talent young talent on defense. You know, Smith's not terrible They have a pretty good set of wide receivers Not many injuries on either side either. Uh, so It's a game that you can really trust the numbers Also, Gino and that Thanksgiving game had that banged up elbow that he hurt on the sunday before it and then Still played thursday, but then looked really good against Dallas. That was indoors So that does matter, but this game I think it's like three mile per hour wins at santa clara for sunday So not a lot of win to discuss there part of why I like the over in this game is that so I think it should be a fun one out in California Let's talk now about the bills and the cheats. We're right now fangirl sportsbook has the cheats as very slim favorites point A half in their favor right now Total of this game is 48 and a half and that's spread as tight from where it was earlier on this week It was minus two and a half. It's now minus one and a half Do you agree with that movement towards the bills in this game? I think I can make the case for why it's moving that way Look buffalo is probably one of the most talented teams. It really hasn't worked out from there for them this year Uh, I kind of feel a little bit like a broken record But we've talked about how bad their defense has performed this year They're 31st when I look at passing success rate adjusted for opponent But I really don't think even with other injuries that um, that they're that bad I think they have some decent players in the secondary Um, you know, there's some signs. They're 13th when you look at adjusted yards for pass attempt Success rate is stickier in terms of predicting the future, but you know when you have that wide gap you can kind of I mean, I think they're better better than that and I think buffalo is a pretty good team Josh Allen the offense has been great. They're fourth in my adjusted passing success rate. Um, you know, they have they have all those weapons But on the other hand, you know, my model has kansas city by four point two points So I do like not having to bet. I mean two and a half is fine A fine number, but if you're gonna if you're gonna bet kansas city one and a half is is also good Uh with kansas city, um, there are some injuries. Isaiah pacheco is uh, not practicing although I feel like he should play um, you know in patrick mchomes hasn't been great, uh So, I mean not up to as usual lofty standards, right?  Uh on monday night against green bay Could have really won that game. I guess they got to convert a two point conversion as well. Um I mean if I lean any direction it would be kansas city minus one and a half here Yeah, I did take the bill's money line at plus 118 on tuesday or I took that on monday, I think but then talked about on the show on tuesday and The reason why I was there is because I have this is basically a toss-up I have 49.8 for the bills, uh, which implied there is value there at plus 102 That means there's no value left in their money line right now I do agree with you that the pacheco injury is Significant for the cheese if he doesn't want to play now the situation for him was He had a run I was watching pretty closely because I had him as uh in the mvp slot and uh My single game lineups and I had a couple of You know, they give you like the bonus bet same game parlays. I'm not a huge fan in general But like hey, it's if it's a bonus bet sure, whatever. I'll I'll play my part Um, and so I had like some of that stuff tied to pacheco There was a play where he came off and like he was like It was I think it was the last play in the third quarter He like kind of motioned toward the sideline, but then like stayed in the game I could be totally making that up But I thought I saw a motion towards the sideline, but then he stayed in And I'm not sure if that's where it happened. They're calling the shoulder confusion And I always assume that means he'll play But not practicing again thursday is concerning the reason that that matters is Like they they just been a very good rushing team recently Which they hadn't been earlier on this year and like it keeps the bills honest So I I think that like pacheco does matter. I still think there's a good shot that he plays like a Probably 50 chance that he plays in this game despite missing practice again Uh, but like it would be a downgrade dot to have him, uh for sure. So I don't know. I I like the bills of plus 118 plus one or two. I don't see any value anymore So I would stay away from it personally, uh, but I do think that I get what you're saying with the minus one and a half Just as a as a by-law spot on the chiefs Um, right given that there's kind of some bad sentiments in the betting market around them right now right, I mean in general like I mean my Numbers probably I think they're probably spot on with kc and it may be a little bit low on buffalo but I mean there's a little bit of wiggle room where I have it by You know over four points Right, exactly. So ed leaning towards the chiefs minus one and a half in this one Let's finish up though with the headliner from week number 14 That is the eagles at the cowboys are right now total in this game Is 51 and a half and the spread is a cowboys minus three and a half and we finally saw the eagles Kind of get punched in the mouth last week and now they get another tough task on the road So ed can the eagles bounce back and cover here or the cowboy is moving to a tie for the lead in the nfc east Yeah, I mean look my numbers have dialed by about a point. So kind of showing value but Uh, unlike last week when I had value on philly and I was able to I don't know. I just thought san francisco being Favorite at philly was just too much didn't work out. Um I think I mentioned that I think philly is overrated. I do think philly is overrated I I wouldn't bet this even though my numbers have it. I do think dallas is probably going to get this win at home Look philly seven and one and one score games, which is which is kind of crazy I actually looked up some of uh jaylon herds' pff numbers So he he had a pff passing grade of 84 last year And it's only down a little bit to like 81. Um It seems like it should be down a little bit more But when I was looking at that at that, uh, it was interesting to see we talked about how scoring is down But you know Last year, let's see. Sorry this year. There are 13 quarterbacks that have a passing grade over 80 And last year there were six So there are actually more quarterbacks that are doing well this year Maybe it's balanced by the just awfulness of of some of the lower lower graded quarterbacks Um Anyways jaylon herds is outside the top 10 this year. That's how I view him as a passer I just don't think he is in a top 10 when we're talking about passing. Obviously he brings other things to this game I just think there's a ton of questions with it with the zealots team With dallas, we kind of wrote him off after that san francisco game a long time ago But before that like we were really pumped about I think I have a bet on dallas winning the division I mean, I like dallas coming into the season uh Kind of sold on him a little bit there in the middle, but they've looked pretty good recently I think we're back to where we think dallas is uh, you know one of the one of the primary contenders to make the super bowl out of the nfc um I don't know. I think the market's fine. I wouldn't bet this Yeah, I'm pretty much in the same spot. I mentioned before that I don't trust my one model when it comes to the eagles because Because the way of use passing efficiency and they don't grade out as well there The other one actually does have dallas minus four and again, that's the one That's a bit higher on philly. So even that one's showing a bit of value But it's not a ton not enough where I'd be willing to bet it but Um, the minus three and a half is actually even money right now fando sportsbook. So potentially A bit more enticing there, but again, it's not a big enough Um big enough gap to for me to get super intrigued. I think the most interesting thing here is actually the total So I talked before about how like, you know, I'm seeing a lot of value in overs on low totals I actually have this one is pretty much spot on now It's very hard typically for me to show value in a high total with a decently tight spread because high totals correlate Towards larger spreads So you wouldn't expect to see a high total be a fair like number when the spread is tight like this But I've actually got this total at 51.1 so What that says to me yet is this game is going to rock because it's it's a good offensive environment It is indoors. There is no wind here So like that does nothing to you for for you from a betting perspective because it's saying the market's efficient But I think the fact that market is efficient given how high the total is for a game with a tight spread I'm pretty jazzed about that. So from a a football fan perspective that gets me excited Yeah, and it's got the nice sunday night slot. It should be a pretty good football game I'm hoping the Eagles can kind of do their best to stay in there and maybe potentially win it but um But yeah, I think Dallas is right. I feel like favored. I did have one question for you Ed when you were looking at Jalen Hertz's pff grade Uh, this is overall grade not just passing grade. Did you happen to notice the player right in front of him? Uh, I did not It is Brock Purdy who I've been told is not deserving of the the mvp award because he he benefits too much from his environment Although Jalen Hertz is third in mvp odds right now Brock purdy is above him in pff grade and like I'm not saying like you should Like Brock purdy should be mvp. I just think it's very funny the way the discourse has dismissed him of like, oh His stats are propped up by his his environment by his coaching staff, which they are that's true But like, you know, it's not like he's terrible And I think the people kind of take that a bit too far sometimes and like dismissing Brock purdy Just because he has good circumstances Yeah, yeah for sure. Um Yeah, I don't know. I don't I don't know what I mean, it's interesting to think what we'll be saying about Brock purdy in two years Right. Well, let's just be a blip Yeah, um, you know, I mean Daniel Jones came out of the the gate roaring. Yeah in the NFL Uh, he was he was pretty good when they when they first threw him out there as a rookie That hasn't worked out quite so well. So we'll see what we're saying about purdy in the future You know, whether he's with the Niners whether he's with another team Um That's the fun part about being an analyst Absolutely. Alrighty Ed beyond those games any other spots where you see value across week 14 Yeah, so I want to talk a little bit about this Thursday night game because I think you can make the case for, uh some value in Pittsburgh So what actually has been keeping me busy this week is that I actually reworked my model over the summer um, I was able to figure out I was able to Uh adopt my algorithm for making schedule adjustments to account for individual quarterbacks, right? You could say yeah, you should have done this since beginning Ed, but look, I got it done this off season and uh Finally with all the injuries and with the lack of college football this week I got around to to implementing it and it's pretty interesting because uh Bailey zappy is is kind of historically bad when you when you look at it When you get look, it's only 71 attempts, but you know, he's predicted to have a passing success rate of 27 percent Against an average NFL defense the NFL average is about 15 percent So, you know the second worst on this list is Tommy DeVito at 32 percent. So there's there's a huge gap and um When when you make a number, okay So before what I would do when accounting for quarterback injuries would just Would be to rely on the market model um You get a couple of games and and you get some sense of What the market thinks of that team compared to NFL average and now I actually have a way of doing a database Uh way of doing it both through success rate and yards for past attempts But when you did it was zappy, I mean you you got something like Pittsburgh by like 10 or something and So you and you can argue like if zappy's going to continue to be that bad Like Pittsburgh, which should be a 10 point favorite, but then you sit back and think oh Pittsburgh is going to be a 10 point favorite with mr. Biskey at the quarterback position It didn't make a ton of sense, right and so um You I actually get about Pittsburgh minus six when you assume that new england is about three I I use the market value for for new england saying, uh, okay We don't have a ton of don't have a ton of stats on on zappy. Let's just go with that If you assume that they are about three points worse Maybe four maybe more. I don't remember the number but That would give you uh what the market has for Pittsburgh So I do think you can make the case for Pittsburgh side here But it's just when you sit back and think about it. It doesn't make a ton of sense. So Uh, it's an interesting game. I I mean I I haven't I haven't bet on Pittsburgh here But I thought it was an interesting thing to talk about for this week I'm curious what the number said about trabisky in that sense because it's a larger sample on him But it's also been a pretty long time since we've seen him actually get reps at quarterback and like for me when I'm I don't know if this is wrong But like I view there as being almost no difference between picket and trabisky So that's part of why I should value the over here is because like I don't even make a difference I think picket kind of sucks. So like Like that's part of it why I should value the over there part of why bet steward's minus six Uh, so I'm curious what the number said regarding those two. Yeah, I mean for trabisky It's it's 50 past temps right and I have about 3% better than nfl average when you make the adjustments But again small sample size. I don't expect him to be better than nfl average Especially with the weapons that they has on that on that pittsburgh offense um, so I think overall When you looked at everything I think pittsburgh was Like exactly nfl average when you looked at just trabisky's numbers and the market model with him Okay, well Putting my money on uh, mitch trabisky via the over and via steward's minus six What could possibly go wrong who is ever regretted having financial? Implications tied to mitchell trabisky in their entire life That is all that we have here for today here on covering the spread But add a lot of stuff cooking for you over at the power bank. What's going on over there and on the football analytic show I had a great conversation with professor edward egros on the football analytic show We talked about the nfl and why offenses sucked talked about the college football semifinal games So get that wherever you get your podcasts and then check out my uh free sports betting email newsletter If you're looking for action on any given weekend, this is the free service for you You can check that out at the power rank dot com. It's my curated list. Uh, sometimes it's my bet. Sometimes it's other people's bets Um, but check that out at the power rank dot com All right and find ed on twitter at the power rank. You can find me on twitter at jim sonnis I am on threads at jim dot sonnis. You can find fan dual research on twitter at fan dual research I want to thank you all for tuning in for today. Good luck to you with your bets across week 14 Back again tomorrow to break down some props and some epl bets. We'll talk to you all then This has been covering the spread right here on the fan dual podcast network


Hey guys, it's Monica. I am going to share with you some of the things that I have been doing with my new face device. Do you think at 66 I can get a new face? With new face? Anyways guys, if you are interested in finding out exactly how I'm using this device, what I'm doing and potentially something that could save you some time and money if you're thinking about buying a new face, stay tuned. So oh my gosh, first of all, let me just tell you that I've only been using it at the time of this filming about two and a half weeks. So I still have a long way to go, but I have really discovered some things that helped me use it better and also really just like anything, an exercise program, a diet program, anything you do. If you don't commit to it, if you are not consistent, it isn't going to help you. It really isn't going to help you and the new face device is amazing, but you need to commit. So I had gotten an email. First of all, this is not a sponsored video. I want to make that perfectly clear. I am not an affiliate member. I won't make a penny off of anything that you purchase from the website I'm going to talk about, but I was contacted by a company called Skinstore and they asked me to take a look at their website and take a look at the products they have and pick something I might be interested in reviewing for them. And of course, it was all about facial toners and all of that, the page that I landed on. And for those of you that have been following me for a long time, you know, I have been wanting the new face for a long, long time. Now, I never bit the bullet because it is pricey. And again, it's going to go back to you needing to make sure that you are committed to using this the right way. So when the company reached out to me, I checked their website, I was so totally impressed. I'm going to show you in a superimpose. I was so totally impressed by their product line, what they were talking about. They have been around for 20 years. I had never heard of them. And they carry a number of products, as you can see from many, many different brands.  And they have, I think, an amazing service. They offer a free, fast shipping for any orders over $49. And because of the huge product line that they have, to me, I thought, wow, this would be interesting to all of you. So they have gifted me with this device in exchange for review. And they have also given me a 25% off coupon. There are some restrictions that may apply on some of the products. They don't have to check it out. But a 25% off coupon, which I think is absolutely amazing, especially for the new face device. So I'm super excited to try this. I'm going to, I have taken photos that this picture that I'm going to include here was actually the day I started using the device. And I had no makeup on. I just had a little bit of mascara on, but no other makeup. So the way I have done this device, the way I have used it, because I have the L head, which I really like because that has been, I can focus in on some of my really fine, fine wrinkles. But I'm going to take it off and just show you.  And then you can insert the regular one. So this is basically how most of you will get it. This is the new face trinity. And the thing for me, if you have like, say you're hot of hearing this, the, the beep is kind of a low tone. Some of you may hear it perfectly fine. And some of us will not hear it. So I can hear it fine on my good ear side, but not good on, or at all on the other side. In fact, if I sleep on my good ear side, I don't hear any sounds because I'm pretty well one ear, totally deaf just about.  They do have a gel, a conductor gel, a primer gel. I'm going to call it a conductor gel. Now I know that you can look and you can probably buy other gels and all that, but just be super careful on the ingredients that are in there. But anyways, this is the gel primer that comes with new face. And I basically wash my face. I only put on, I have a little rubber, a little rubber, which I didn't bring in here, spatula type thing that I use as a brush. I only put on enough gel to cover the section of the face that I'm doing because I think it dries fast. I put it on with the damp skin first and then I will put the gel on. I start usually up my neck. And the trick is three go-arounds, three times each in three different sections on my neck. So I usually start low and you also have to make sure these are microcurrents. You get a positive and negative. They both have to be flush against your skin when you are gliding for it to work correctly. If you lift up one or one is kind of like at a different angle, that's not good. So you want to make sure that you're definitely flush and then you start. And I don't, I avoid the middle here, but I start and I just go all the way up. Now on this side, I can hear it. I go slow enough that it's a five seconds roughly before the beep. If I don't hear the beep, I don't move. So then I go back and I go to the middle and I do it in three sections and I do the same thing right under my neck and I always go up. I wait till I hear the beep. The beep is roughly, like I said, five seconds. So then I'm going to go and repeat that whole pass and then I go to my face and I'll wet my skin again if I have to.  I wet my skin if I have to and then I will apply the new batch of the primer, the gel. Then I'll turn it back on. It kind of remembers that I was on a high. I didn't even mention you can have a lower intensity, intensity and higher. I keep mine on the highest and then I do the same thing. I very slowly make sure that my skin is connected to this device.  I hear the beep. I move to this. I go up.   I just do. And then I wet my neck and I start on the other side. This side I can't hear a darn thing. So I make sure that when I have it on here, I make sure that I've got my connection and all my hair out of the way, my necklace out of the way and I do the very same thing. I glide up but I'm counting and I usually go, you know, one, one thousand, two, one thousand, three, one thousand, four, one thousand, five, one thousand and lift up. And that's how I monitor it because I literally can't hear the tone is low. I can't hear it at all on this side and I will do that again. The repeat the same thing three times and then I move up here. I put the gel on here, wet my skin, damp my skin. Make sure I put the gel on here and I do the exact same thing and I'm focusing on lifting up, lifting up. When I'm all done that, if I want, I will do my forehead. I have a hard time with my forehead. So I'm tending to, you know, I just do one pass three times instead of going all over.  It just feels weird. So I just go up and I'm counting if I can't hear it. Again, it's one, one thousand, two, one thousand, three, one thousand, four, one thousand, five, one thousand. That's how I do it. And that's pretty well. The beep comes at five seconds. When I'm all done that I pop this off and then I put on the attachment, snap it in. This is the attachment I do for my eyes. Also, I go back into my nasal lumbia folds or whatever they're called and here. And when I do this, and if you guys want a more, a better, more detailed tutorial on this afterwards, let me know and I'll share it with you. But I basically put the gel on and I start, I don't glide here because I have a lot of loose skin. So I just kind of hold it and count and hold it and count, hold it and three times. Again, everything is three. So you do three sections and everything.  I am paying a lot of attention to my eyes because of my hooded eyelids and I'm holding it up there and doing the same thing. Ah, you know, I post it on Instagram, I have the new face and people go, oh, you're going to love it. You're going to love it. And I know people that have used it, that have loved it. I have over the course of time watched videos with women that use it and love it. I was super psyched when the skin store contacted me and this was one of their products that they were willing to send me. I was like, oh yeah, baby. Now, for those of you that are going to ask, you all know that I do the red light therapy. I have the trophy skin device, I use the red light therapy. I do that every morning when I'm home. So one of the things I do want to buy, they have a light therapy attachment. I don't have it, but I do want, if I, when I continue to use this long term, I do want to get that because I think that red light therapy would be perfect, especially when I'm traveling. Because that's the one thing about my big trophy skin. I lie there under it in the morning. I love it.  And then when I'm all done this, I kind of wrench the gel off and then I do my regular skincare. So I'm loving it. But you guys should check out the website. I'm going to link it all below skinstore.com. The amount of products they have on there is amazing. Makeup, skincare tools, hair, I mean hair tools, all sorts of stuff. So check out their website and yeah, and I'm going to continue to update you on the progress of my attempt at 66 to get a new face. But you know what? Even in the short period of time, I think I've seen a difference. I was talking to Marlene, Fab and Glam and I said, oh my gosh, I think I see a difference. And she goes, you're going to notice a difference. So here's my big tip. If you can't commit to doing this consistently, I've been doing it every day. I think for the next three months, I'm going to do it every day as much as possible. If I skip a day, that's okay. They say at least five times a week. But if you can't continue to do this for whatever reason, it's just like when you stop going to the gym, when you stop lifting the weights, your muscles start to sag. So this is an awesome product as long as you use it because your muscles have a memory. And when you stop, and I'm talking longer than a week or two, you know, I mean, we all have issues that come up when we can't go to the gym. We can't do this or that. But when you stop using this, your face will go back to the way it used to be, I think, although I don't want to test it because I'm loving it. So anyways, that's my new face for my new face at 66 sent to me complimentary from a company called Skin Store. And I'm so thrilled they found me and sent me this device. So thanks so much guys for watching. I'll chat with you later and I'll keep you updated on my progress.


The new Special Advisor to the President on Media and Publicity during Gilali has reiterated the commitment of President Bola Tenobu to serve Nigeria and open up the presidency to all in order to ensure that public office holders deliver renewed hope to Nigerians both home and abroad. Gilali, who resumed this morning, was having his first meeting with the State House Correspondents when the assurance of disseminating prompt information on the activities of the presidency to all Nigerians in terms of the execution of the policies and programs. President Bola Tenobu is a man of empathy. He is committed to the cause of opening the presidency to Nigerians for the benefit of not just Nigerians but to the benefit of public office holders because it is in the interest of public office holders to know that they are being watched so that they can be held accountable to deliver on the promises of the president in terms of the execution of government policy. And we are going to, in very short order, announce a series of very tangible measures that have to do with the opening up of this institution on a daily basis in a deeper and more comprehensive way than has ever been seen before. The new media aide noted that the era of international or institutional arrogance where Nigerians are kept in the dark over happenings in the State House has ended as every Nigerian has the right to have access to accurate factual and reliable information on all that happens in the presidential villa. This institution belongs to Nigerians. It does not belong to any political party. It does not belong to any public official. It belongs to over 200 million citizens of this country, both at home and abroad. And it is with that understanding that we will be engaging the world. We believe it is the right, not a privilege, but the right of every Nigerian to have full access into the affairs of the presidential villa, into the affairs of the office of the president and the office of the vice president. We will be more open in terms of providing access to information to radio networks, television networks, newspapers and online news media than ever before. So you get notified about fresh news updates.