 Welcome. It's great to welcome you to today's biochemistry colloquium. There are many sponsors that made this possible, so thank you to all those sponsors for the donations. It's my pleasure to introduce today's speaker, Dr. Julian Davies, who's at the University of British Columbia in Vancouver. Julian was born in Wales and grew up between Wales and England, and this shows a picture of Julian when he was a boy. This is pre-UW. This is something that you can't really do these days. Which one? Well, of course he's the fine lad on the right, but this is not the sort of thing that's easy to do anymore is to have your picture taken with a primate. It may be you and Bernie, yes. Julian was educated as a chemist. In fact, as an organic chemist, he received his bachelor's in 53 and his PhD in 56 at Nottingham. And then when he was at Harvard Medical School, he made the shift to microbiology in the sense that he was still a chemist, once a chemist, always a chemist, but he started working on bacteria and he never looked back. He has a very interesting history. So he came to Madison in the 60s, I believe, as a faculty in this department and was here for many years. And then in 1980, he left to head research at Biogen, which was a biotech company right kind of in the early days of biotech in Geneva, Switzerland, which is the spy capital of the world, I'm told. So maybe other things went on there. And I believe he was affiliated with the University of Geneva at the time. Is that correct? Okay. And then he left Biogen and he went to Institute Pasteur, where he started a department with his friend Francois Jacop, who many of us are familiar with, won the Nobel Prize, among other things. After the Institute Pasteur, he was lured to the University of British Columbia in 1992 as the head of microbiology. And in 1996, he co-founded a company called, I believe it was 1996, called Terogen, which was focused on looking for new molecules in nature, but doing it in a new way, using kind of a more of a metagenomic approach. And in 1997, he became emeritus. Now, Julian has been emeritus since then, but he is super active. He is still in the lab. When I visited him last year, he was pulled me in the lab to show me his petri dishes and the things he was working on. And I have a short slideshow of pictures of him over time. So this is a pre-UW. And then this is his faculty picture at UW. We still take pictures like this these days. This is another picture of Julian at UW. And you'll notice that he's, that's Pap's blue ribbon. I'm told is a local beverage. And that's your, that's Julian's wife, Dottie. Is it right? No. Oh, it's not Julian's wife. Sorry. I just met Julian's wife for the first time this morning. It's Colleen. Hey, Colleen. That's Colleen and Julian. Enjoying a Pap's blue ribbon. We'll continue that tonight. This is post UW. You can see that once he broke away from the conservative Midwest, his personality really started to come out. And this is a picture I love, which is very recent, which shows Julian. And you'll notice in addition to this air punch he's doing, I notice he has these tie-dye colored socks. And in fact, he, that's a tradition he keeps to this day. You'll notice he is sporting said socks tonight. So today Julian's going to tell us about small talk, communication between back to your, my eyes are getting really bad. Julian, it's awesome to have you. Thank you so much. And so let's welcome Julian to the stage. Gosh, I'm glad my wife wasn't here. No offense, Colleen. I'm really delighted to be here to and to give a talk. I haven't been back to Madison for a little while. Well, I'm delighted to be here. I've had fun this morning talking to a lot of people and talking to students and this afternoon. And it's wonderful to be here and to meet a lot of old friends. When I mean old, I mean they're older than I am. And it's good to be to see them and to talk with them. And so it was, it was a very great honor for me to come here to the University of Wisconsin again. I mean, nothing looks the same. Everything's changed and things like this or modern stuff and things like this, but it's, it's still the University of Wisconsin and I'm glad to see that the biochemistry building is still standing. But in any event, I'm, this is just a great occasion for me. I wanted to, I wasn't quite sure what to talk about, but I decided I try to put lots of things together and give you some ideas of as to the thing, kinds of things that I've been doing and the reason why I've been doing them. And I wanted to start by telling you a story which when I came to Wisconsin in 19, let me see, 1960, no, 1970 something. Anyway, but when I came to Wisconsin, I had just come from the Pasteur Institute and I was asked what I should, what kind of research I wanted to do. And it was something which I hadn't really thought about. I've been doing lots of things and playing with them. And then I realized that I had a real prize in my possession. When I was in Paris, I met a Japanese scientist, Yukinori Hirota, who was one of the people who discovered our plasmids in Japan, our factors, you know. And he said, would you like to work on an our factor? And I said, yeah, I'd like to know what the mechanism of resistance is. He said, well, here you are. He gave me a strain. He gave me a strain in a tube and it was an NR1. And I did a little bit of work in Paris because I asked Jacob, could I test a few things? And all I could find out at that time was that it appeared to be some kind of an inactivation of the drugs. But there was no, I couldn't do any other experiments. I was supposed to be doing something else. So when I came to Wisconsin, I had to come up with a project. I had to write a grant. So I said I was going to study the mechanisms of resistance in our factors, in our factor carrying bacteria. And this has been a gold mine. I'm still working on it. And for different reasons. But it was great that when I came to Wisconsin, I was able to do some really unusual experiments because of the talented people around here. For example, Julius Adler helped me a great deal because he told me how to use ion exchange paper. I never knew such a thing existed. But ion exchange paper was a beautiful way of binding positively charged compounds and then counting the radioactivity of this being transferred to them during the reaction in activating the genes. I don't know if Julius remembers this, but it was, he was a great help. And a lot of other people were very helpful. And I settled down in Wisconsin very quickly. I worked on resistance a lot. I worked on mode of action of antibiotics and a variety of other things. So I want to tell you that in terms of resistance, everything started a long time ago. And I'm sure you know these people, Selman Waxman and Fleming. And they discovered the first two important antibiotics. Fleming discovered penicillin, which was sometime earlier. And Waxman in principle discovered streptomycin, but it was actually a graduate student of his who discovered streptomycin. And there has been a lot of bad news about that particular relationship subsequently. But in any event, I had this R-plasmid from Europe, from Japan, and I was able to go ahead and try out all kinds of experiments. And we didn't know anything about R-factor modification of antibiotics at that time. So there was a lot to try. This is really the story of Selman Waxman. He did not discover streptomycin. It was discovered by this young man here, Albert Schatz, who told Waxman that he wanted to be able to work on antibiotic isolation. And he chose a soil and he isolated some bacteria from the soil and one of the organisms produced streptomycin. He was looking specifically for drugs that were active against tuberculosis. And he found streptomycin. Unfortunately, as many of you may know, Waxman claimed all of the rights to this discovery and he got the Nobel Prize and Schatz got very little out of it. Anyway, it's a sad story, but that happens. You know, some supervisors are real mean and you have to be careful. So after this, I started working more and more on antibiotics. And I want to give you a brief history at the moment. This is a crowded slide and I don't want to go into any detail, but back 1940s was when we had penicillin. And the question of discovery of antibiotics was, well, it was covered. It's still going on. It's 70 years now in terms of work. And at the first, there was just streptomycin in penicillin. And then in about the 1940s, there was a discovery of many, many different kinds of antibiotics and many useful ones. And of course, the antibiotic industry really took off after that. So they were the golden years of discovery. And unfortunately, due to the excessive use of antibiotics, and you have no idea of the ways in which antibiotics were abused at that time. It wasn't just a question of feeding animals with antibiotics and feeding animals antibiotic residues and things like this. But there are many reports of physicians who used to spray their patients with penicillin when they came into the surgery so that it wouldn't cause any infections for the doctor. And this kind of thing was done completely at random. And it was really a disastrous situation. We know now that it's been a complete disaster. And without going on, we now have a situation where there's virtually no useful antibiotic that has been discovered for a number of years. This is very unfortunate because one needs new ideas. One needs new compounds. And it just has not really happened. But I believe, and as I will tell you shortly, I think there are many, many possibilities for reinvigorating the antibiotic industry that will hopefully provide a lot of new compounds. The only problem is obviously that we have to look out for resistance. And as you see here, this is another list. Any kind of resistance mechanism that you can imagine, bacteria have adopted and fungi have adopted in some cases to be able to protect themselves from being killed by the antibiotics. And these mechanisms have found time and time again. When I first started, I was looking at antibiotic inactivation. But I've since worked with a number of different mechanisms. And it is clear that the genetic dexterity of bacteria was something that was never, I don't think it's still appreciated what these organisms can do. I am a real bacteria lover. And I love them because of their wonderful characteristics, their ability to do many, many things in biochemical sense. And you know, you've got control of them all the time in principle. So the first thing that really happened was this reaction. Penicillin was hydrolyzed by a beta-lactamase which opens this beta-lactam ring here and inactivates the drug. This is the most expensive reaction in history. In the sense that when people sprayed penicillin over various places, bacteria with beta-lactamases were able to hydrolyze the antibiotic. And it was absolutely a disaster in terms of the way in which this was worked. And the interesting thing is that efforts to find penicillins and or beta-lactam antibiotics that have modified structures in such a way that they cannot, that they cannot be hydrolyzed have really failed. They're constantly coming because here we see this is an interesting comparison. This is more a chart which is very small but it shows the discovery, the lifetime of different antibiotics, different beta-lactam antibiotics. So you start here with penicillin but as you find more and more inactivation by penicillin, more and more types of penicillins were developed. And here you have cephalosporins in the way in which they were developed. So the industry has gone through hundreds of compounds in an effort to keep ahead of bacteria, in an effort to get, keep ahead of the beta-lactamases. The situation now as we speak is that there are, at the present time there are over a thousand beta-lactamases known in different bacteria and these are the MRSA everybody knows, they think that's the worst bug but it's a wimp really. And vancomycin resistant strains, the C.A. MRSA which was a highly drug- resistant MDM-1 and a whole series. There are all kinds of names for these particular organisms but they have all evolved. Novel beta-lactamases in order that they can hydrolyze even the newest of the beta-lactam antibiotics. So it's been a constant battle by the industry and the industry, if I can forgive them for one thing, one of the problems clearly is that the industry was not able to control the use of the antibiotics. So this shows the world transmission of separate taxine in activating enzymes. You see all these places all over the world where you find different beta-lactamases. So if you go from South America, sorry, yes if you go from South America to Australia you're going to find a different beta-lactamase. And if you take it if you take a particular drug that worked in Australia, it won't work somewhere in another country. So there's a constant battle in terms of what is happening with respect to antibiotic resistance and the producing strains. And one good example here which summarizes everything is that bacterial resistance is a constant evolution. So that and it seems as most of the effect is due to mutation or inheritance due to stress. And one finds a constant cycle of organisms becoming resistant to the various antibiotics. So what one would like to do is to find new antibiotics. And I'm sure you know this and some of you may have tried, but in spite of that there are other difficulties. And the next slide gives you a remarkable example of a collection of genes which are found in one strain but are now found in many other strains. This particular cluster of genes as you see has 45 resistant genes to six different antibiotic clusters. That's in one package for an organism to pick up at any time. And one has different versions of these clusters in various parts of the world and even various parts of the country. You might find different ones in Boston compared to Madison. And these clusters are readily transferred by horizontal gene transfer. That's one of the major problems with resistance. We cannot stop these particular mechanisms happening. So when one looks at the classification of something like TB, TB is a very interesting case. TB has been a threat to humanity for thousands of years. And you see here that this is recent data from the TB Association that there are several classes of TB resistance. There's multi-drug resistance, there's extensively drug resistant and there's totally drug resistance. I don't know what happens next. How can you fuse totally and extensively? But the point is that these things are going on all the time. And the TB, the International TB Organization is very well aware of this and they've realized I'm not going to go into this in any detail but they realized that the only way to treat TB is with combinations of drugs and the combination of drugs that they use are more and more toxic. So the people who take these drugs because they have extensively or totally resistant TB very often have serious side effects. You can't avoid it. So TB is really the the worst situation and in the case of TB it's not transferable resistance. It's all by mutation. The multiple mutations in the bug give you constant rounds of resistance. So it's really difficult to know what to do and as I mentioned here just now, combinations are very very important and this is a slide from the TB Association. There are many targets mostly in metabolism and there are many ways of trying to hit protein synthesis. All of the drugs that are active against protein synthesis are essentially they're resisted by the TB drugs at the present time. It's a horrendous situation and we can't imagine that anything will ever really happen with TB unless somebody develops a vaccine. There is no vaccine at the present time but a vaccine, vaccines are being worked on and a vaccine is really the only solution. So here we go. The pharmaceutical industry is essentially out of it and so it's the time to do some science on natural products because the pharmaceutical industry spent years working on natural products and just took the products as they came and tried to use them without worrying about where they come from or what they might do but they were just new drugs and this is an interesting thing that I feel I have benefited from because in the 1970s when the pharmaceutical industry was going through the preparations and isolations of lots of different kinds of drugs, I wasn't a consultant for companies but there were a lot of companies since I was working on resistance. They would invite me to their factory or whatever or labs and give a talk and tell them about resistance and these companies, I can give you, they're all the great companies, they would always give me compounds. They'd give me a bunch of compounds, their newest compounds whatever and they even gave some compounds to Bernie Weisblum. Can you imagine what that means? But they gave, I still have a collection of about 300 compounds that I collected in those days. Many of them have never been used but the interesting, the most interesting thing for me about that time was that I didn't have to get transfer agreements. I didn't have to tell the university that I was taking these compounds and every, we trusted each other and I'm quite serious about that. The companies were reasonable, we were reasonable, we told them what the results were. If we published, we acknowledged the company. Nowadays it's just awful trying to work with a company. You have to go through all kinds of forms with some university industry liaison office who doesn't know what they're doing and we find that I think that the situation, the atmosphere in terms of universities and drug companies in terms of sharing things is just atrocious and they're making it worse all the time. You cannot do anything without signing your life away. Anyway, I was lucky at that time and I got lots of compounds and I don't have all of these but I have a lot of different compounds and the thing is that these compounds are really very important and they're very important to other living organisms and this is where the whole question of what are these compounds doing in the environment. When we use them as antibiotics, we're using them at concentrations that are high. They are not the low concentrations that you find in the environment. They're not sub-inhibitory. One uses them and within effort, in an effort to kill the micro-organism. But the fact in the manner is almost all of these compounds and others can be found in bacteria in the soil, in bacteria in various sources and they have other functions and here's a good example. We've been doing some work on lichen structures and within lichens, I can't show you this now, but lichens are beautiful organisms. But within lichens, it's now possible to thin section them and look for bacteria and look for compounds in the lichen structure, in the interstitial structure of the lichens. And there are many compounds being made there and many different compounds are in lichens and any structure you look at from an organism in the environment, you find these small molecules. And the question is, what are they all doing? When I say these small molecules, sometimes they're new, sometimes they're not, but they're all circulating within the structures of a whole series of organisms and most importantly, many of them are circulating in us and they are perfectly reasonable, they are involved in interactions. And so one really has to have a new think or new thought about what small molecules are doing. Here's some pictures of lichens and some various compounds that can be identified in different places. These bacteria are there and they're there for the health of the lichen. And the health of the lichen means that these organisms have to be stimulating or controlling some reactions. We just don't know what it is. And I like this statement by Goethe, of course, I am a little think myself, so I don't mind it. Okay, so the real question and we started asking this question a few years ago was, are antibiotics really antibiotics in nature? Are they antibiotics in nature? And Waxman was the person who first named an antibiotic. He called Streptomycin an antibiotic and that set off the whole tradition of calling a small molecule coming from a microbial antibiotic. And he then, not long before he died, he argued that antibiotics may play no part in modifying or influencing processes that normally occur in nature. So if that's the case, what are they doing? I mean clearly they have to be playing some role in nature and the question is, what are these roles? Now another incident, another situation where the effects of antibiotics or the production of antibiotics was that in pharmaceutical companies, the most important thing was to make as much of the product as possible, because the more you can make, the more you can sell. And there's some very interesting facts here. The antibiotic discovery and the antibiotic research was totally at random. People tried all kinds of things to get strained from the environment to make more products. And for example, in the environment and you look at the properties of the medium of a, to produce an antibiotic, in this environment you find that there are many properties and they're all very mild properties and you get very small amounts of materials. You only need small amounts of materials in nature. But then when you switch to a fermenter and you start doing production, you have to start changing these things a great deal. And so when originally the doubling time of a strain producing a small molecule might be days or weeks or months, in a fermenter it was a few hours. When you're also putting different substrates and aeration in soil is very low. Aeration is high and constant feeding with nutrients and various kinds of nutrients. Now the, and the whole situation was controlled in such a way that you make as much of the compound as possible. And this is the, the kinds of things they use. Now the most interesting thing of all of this slide is that sometime during the process of producing antibiotics in industry somebody came up with the use of hydrolyzed rabbit's fur to get a high concentration of a compound. And this turned out to be a very popular substrate for some strains and in order to produce antibiotics. Now whoever thought of this was a genius. But I think it's, it just gives you a sense of the, the illogic, the poor logic behind the use of bacteria to produce compounds and then to produce them in industrially, in industrial fermentations. So in nature they are simple molecules and they are present in nature in very low concentrations. And this is some experiments we did a few years ago. But the summary of this is here that at low concentrations the antibiotics have an important function. They, they stimulate reactions, they perform many functions and as you get up to the concentration which is close to the inhibiting concentration these things all decline and one has inhibition. So this is, this is nature, this is a fermentation tank and they are totally different. And what we also find is that with different compounds one has different interactions between the compounds and this is using a reporter assay to look for the macrolite antibiotics. You find there's many, there's a great deal of scattering depending on the concentration one gets different transcription assay activation and you can really easily separate these compounds based on their activity and their concentration and you can see that small molecules are really active and you don't need to use them at high concentrations unless you want to kill another bug. If you're using them in nature in small concentrations that's okay. And my hero is Paracelsus. I wish I had a hat like that actually but what he said is here, this is his statement and the fact of the matter is that the production of antibiotics is in the use of antibiotics and their effects is entirely due to concentration and this is now generally accepted that if you use low concentrations you get considerable metabolic effects between bacteria and you can show that a bacteria, a compound produced by one bacterium can activate functions in other bacteria. They're acting as signals. They're acting in ways that they can stimulate the organisms close to them. We don't find any conditions where microbes are making small molecules, inhibitory small molecules in nature where they are not high at low concentrations. So what Paracelsus says, the dose makes the poison and we all know that if you drink too much you get drunk and in these cases you don't want to get drunk. You don't want to die. You don't want to get blind drunk. You want to be able to use these compounds in such a way that they're carrying out activations in a way which is consistent with their presence in the environment. Now there's been a lot of work in recent years on signaling and particularly between bacteria and I give you here one of the probably the best known examples which is the it's a canonical system the Lux-R system where you have one function, one small molecule which activates a particular pathway and that particular pathway then can affect the production of target genes and there are many cases now where you can see that the Lux-R system is operating in a very controlled way to regulate metabolism, to regulate pigment production, to regulate gene transfer any of these things they are done at low concentrations and there is not there are other things besides Lux-R and this is why I feel that antibiotics as we know them are not don't fit this way this pattern but antibiotics at very low concentrations really do fit this pattern but these are unknown activations that we have at the moment that if we have a small molecule and binds to receptor then we have very simple and defined effects electro, gaseous, light and nanotube functions are also present and they affect the interactions between bacteria. Bacteria have lives, they have signals, they have a language that is between them and so we must think about the signaling responses and the specific transcription functions. Now when you think about activity of small molecules and their effects on cells one of the best examples is the ribosome. This is the small subunit, the large subunit and it is known now largely because of binding and then also by electron microscopy studies and cryo electron microscopy studies that you can find the binding sites for these small molecules on the ribosome and they are there in low concentrations and they clearly affect the ribosome in different ways. We are not talking about inhibition, I am confident that a lot of these molecules are binding to the ribosome as one of the central core structures in the cell which responds to small molecules. The ribosome is a target, a receptor for small molecule activation. But let me now go into this, this is a question of signaling. This is something which I am sure you all know Margaret MacFall and the guy, she came out with this great chart a few years ago and about signaling and what is talking about here is signaling between different organs in the body and also the external environment and these are signals which are important. These signals are important for life. They are responsible for interactions being stimulated in one cell or being turned down in another cell. So we must think more about the use of small molecules in terms of regulatory effects, up or down regulation and it is very, very easy to detect these things. This is a few examples, for example here. This strain makes, does not make an inhibitor but as you see along here, as you move, as it moves, this is the same strain here and right here you see the induction of the formation of an inhibitor. You find the same kind of situation here. Here in Roberto Coulter's work he shows that in this particular strain you can get a biofilm formed by the addition of a small molecule. These are cases where the lowest concentrations of the compounds stimulate an enormous swarming effects of the bacteria and they are made pretty by using a reporter but the swarming is very concentration dependent. So you are making bacteria, you are inducing bacteria to swarm to different places at low concentrations and these are other cases of swarming, low concentration, the rest of the isin doesn't make them swarm at all but the compound named Kosuga isin, which Jim Darberg knows well, which is a very active in producing swarming of bacteria and it will produce swarming of various different kinds of bacteria. Here's another few cases where swarming is detected by, oh these are interesting because a norepinephrine and epinephrine which I assume to be human hormones are also bacterial hormones, microbial hormones because these compounds will stimulate swarming of different bacteria and this one will inhibit but this one will swarm, this is a case of swarming which is induced by the compound that is placed here, things like that you can get absolutely marvelous patterns of bacteria swarming and swimming based on the presence of small molecules. Let's see and this is a weird one, so you put spots of bacteria here, there's one here and if you're lucky you can get a real picture out of it. Right, so what I want to summarize at the moment is that there are certain bacteria that are very, very adept at swarming, swarming and interacting and the ones that we find that are perhaps the most useful are the bacilli and they've been called the smartest bacteria by Ben Jacob but they have, they produce many small molecules, the bacilli produce a whole series of cyclic dipeptides, they can produce as many as 10 cyclic dipeptides of different types and those cyclic dipeptides have effect on the swarming of other bacteria. So when bacteria are in your gut living, they're having fun, they're swarming, they're moving around, they're moving, they're swarming, they're swimming and that's being run by small molecules. This is a case where a presence of a small molecule, a bioactive small molecule leads to an enormous increase in the yield of conjugants. This is a strain radiobacter capsulata, which is a water organism and it undergoes gene transfer by means of a particle which carries the DNA and if you increase, if you put it in the presence of very low concentration of a small molecule, nova biasing, you can see you get increase of gene transfer of 100 fold or more so that these antibiotics, the so-called antibiotics and other molecules are actually doing a lot of things. They are acting as sex hormones in some cases. Okay, so what you must realize and what has become very common in microbiology at the moment is that all microbes in the environment exist as connections between their microbial communities, their distributed metabolic networks. What that means is that one organism will produce a compound which will stimulate another organism which will stimulate another organism and et cetera, et cetera. And these distributed metabolic networks are believed to be the basis of microbial communities in the environment. And they're all based on signaling molecules. The signaling molecules have not been identified in many of these cases, but you can see interactions between two organisms, easily in three organisms, but when you're looking at a million organisms in the soil, it's clearly a rather difficult proposition. Kendra Ronda, Rumbaugh a few years ago, showed that a whole series of molecules will lead to an interaction from between bacteria and other organisms. These are the compounds. They will interact with fungi. They will interact with plants and animals so that the bacteria and us are communicating with us all the time and they're doing it for a good purpose. They're doing it for our health. It is now possible to detect some of these bacteria and the compounds by high resolution mass spectroscopy. And so you can find different bacteria in an environment. An environment that you have to establish producing a variety of different molecules and these molecules will stimulate or enhance and in some way interact with other compounds with other organisms. This type of system is not readily available but it's becoming more available and we'll be able to detect what these compounds are by direct mass spectroscopy. Don't do that. Sorry, going back there. So, in terms of antibiotics and the production of antibiotics, we're in a situation where even though we know that these compounds are not really antibiotics, we still need them. We still need to use them as antibiotics. But they're harder and harder to find. Every pharmaceutical company in the world and many laboratories are looking for compounds that are going to be active for some disease or another and the failure rate for example here, these two compounds here are actually used in agriculture. They're not in, they're not human, they're not useful in humans. So, one has a tremendous source of compounds that is available in nature and we have to try to find them. This is what I believe is the source in nature. Actually, Stuart Treiber who first came up with it and he said that life cannot exist without with macromolecules alone. And this chart just simply shows you that the central dogma should now include not only the genome, transcriptome and proteome, but what we call the parvoam which are small molecules, natural products distributed in nature. Here's an example of penicillin. Penicillin has many different functions and so we have in the environment a marvelous collection and set of interactions between molecular weight compounds from very small to very large. It's really very important. So, question is without antibiotics what can we do? What are going to be able to use to treat infections? And my last few slides may think that I've become a quack. I assume you know what a quack is. But I probably am already a quack. But anyway, I want to tell you about some which concerns natural medicines. I never thought that I would be interested in natural medicines. Well, I used to take it a lot I had to cod live royal when I was a boy but nothing much in terms of natural medicines. And these are the escape pathogens. These are the seven most serious pathogens that one finds in hospitals. These strains have been identified as the most serious human pathogens that occur in hospitals and they almost always are found in hospital situations and in mostly in surgical wards. And the death rate from infections by these organisms is very high. And these organisms are all multi-drug resistant. There are no drugs that are effective against all of the escape organisms. Escape means here we go. Enterococcus, Staphylococcus, Clepsiola, Acinobacter, you see escape. But they're very, they're in a serious problem and they're considered to be a a major project for discovery in the industry. Now, I want to tell you a little bit about antimicrobial activities of a natural mineral clay. And you'll say, I've gone mad. I have not gone mad and I'll show you why. This, this natural, there are many minerals found which are natural clays and you find that they're very complex structures, very complex structures and can have a variety of different minerals. The reasons for these structures are really quite difficult to anticipate. But the natural clay minerals have, some of them have quite different structures from others and the clay that we've started to work with is Kisimid clay which is found in northern British Columbia. And this, this shows you what antimicrobial clays actually can do. They are known already to have a variety of effects on bacteria and probably also on people. So you're not necessarily going to be using clays on, shall we say, precious parts of your body because you might do the wrong thing. However, this is northern and the coast of the northwest coast of British Columbia and this, here there's an Indian band that lives here and this here is a huge mound of clay. It's, it's unique clay in terms of its structure and in terms of its environment. This clay has not been found anywhere else but it's been used in the past by native populations and also by a number of physicians and many people with bad burns have been treated with clay. But anyway, what's this clay? What is it going to do? Well, we harvest, we've taken lots of samples of this clay and we've cut cores and analyzed the cores and we know exactly, we know a great deal about the different types of clay that are in different places. But here, Hauser did a lot of work on the activity of the clay. Now Hauser is dead so we can't get much help from him but the point is we feel that the clay has really interesting activities and here we look at different mineral clays the chisimi clay. It's very unique. It has virtually no pyrite which is iron sulfide and it has a number of other different characteristics which are different from other clays. Now, it's easy to do these kinds of experiments because you can just separate them by centrifugation. The clay, you might say, well, so what are you going to be able to do with this clay? And how can you use it? And this slide shows you experiments where we take water aqueous suspensions of the clay, centrifuging to remove all the solid matter and we have solvent treatment of methanol-resistant staph aureus, E. coli, Klebsiella, A. snedobacter and you see that at relatively low concentrations you get complete inhibition and complete killing of the bacteria. That's nice. That's nice. And we've also repeated this with a variety of other suspensions, so the leachate suspensions and we find also that we get very rapid killing by this clay. Now, I didn't believe all this stuff for a long time. I must say. And I haven't had a hallucination or anything like this. But this clay is most unusual in the sense that we've looked at the microbiome of this clay and it has 3,000 taxa in the clay in each sample and we're still working on it but it's enormously complicated material and there are no other clays that have this complexity of being significantly microbes. And we're hoping that we might be able to get some use out of this. So this is the constitution of the different microbiomes. They vary depending on where you take the sample from. Galeonera. Galeonasia are a marine organism and we find here Galeonera again and they are very, they're very accurate, spatially distributed within the clay and we can take out clays which we feel that we really want to be able to use. These are some more figures showing you the activity of the clay against the escape pathogens and they're very effective. Now there's one very interesting thing about this clay and that is that the native population uses them for geophagy. They eat it. They eat the clay and they use it for treating stomach infections. And as far as we know there have not been any fatalities of using this clay. So the question now is is this clay going to be a useful alternative for the treatment of escape pathogens? Perhaps under the various special conditions or is it going to be just another what can I say, a dream of a group of people who think that they can find something from the environment. But this is an environmental sample. There are a number of people that have used it on themselves which is of course completely illegal but we've done it and it appears to have no side effects so far and kills all of the escape pathogens. Now whether it's going to be useful against the escape pathogens, I don't know. We know very little about the mode of action but that's something that is worked upon. So I would end by saying last few slides with a microbiology. What is needed at the moment is more compounds. We need many more small molecules if we're to continue with standard antimicrobial treatment. If without more compounds from whatever source we can it's not going to be possible to carry out the kinds of treatments that we've done in the past because of the problems of resistance. Without recycling drugs you cannot overcome resistance. And I believe firmly that the possibilities of getting more compounds pure from the microbiome is really good. We can look at microbiomes from different sources and find whether or not there are novel microbes. But on the other hand I think there are things like clay and possibly other similar complexes that might turn out to be very useful. I mean it takes us back about 10,000 years of course but on the other hand if it works then one would be happy about it. So with that I want to thank you for coming to hear me. Well thank you for inviting me first and thank you for coming and it's been a great pleasure for me. Thank you. Sure, Julian would be interested in having some questions. Maybe we can have a question first from a student. Sorry Colleen. Any student questions? Darrell? No? Colleen, please. Small compounds and they are active in a petri dish against a discape pathogen. So if you look at a core sample and you try to extract different levels of the core can you tell where these are coming from? I assume you have a sterile extract and it's not. We know that there are some of the core samples that are cut have no activity but we know that on other sites nearby the core samples have very good activity but we don't know why. Have you done any of the classic biochemical tests to determine what it is? I don't do biochemistry anymore. Water... No, it's not. Peptide or... What do you mean by classical tests? Well, is it heat stable? Is it a small molecule? A large molecule? We don't know. We don't find any small molecules. We don't find... We've run mass spec on the aqueous leachate. And we don't find anything. We don't find any compounds. We have... Let me see. We've centrifuged as hard as we can and we can't get anything out of it but the material is suspended in pure sterile distilled water and we don't know anything more about it. I'm not kidding. Very interesting talk. I want to see if I get one of the punchlines correctly and that's that without more compounds we cannot treat microbial infections as we have in the past. I maybe try to be provocative and say maybe you have to do a different way to treat and what you've told us is that we are microbiomes. We are ecosystems. You've also showed us interesting data that the microbiome is a function of where it is, so at least in your soil samples. So I suggest that maybe what you want to do is not try to kill off the bacteria the way we have traditionally but manage the ecosystem. So if you have ways of understanding how components of the signaling affect the ecosystem distribution you might shift the distributions towards lesser pathogenic strains and favor those that are more healthful for us. I wonder if you could comment on that. Well, I agree with you completely. I think that the question of using microbial populations in order to treat different kinds of infections or even things which are sort of more fundamental in terms of life. They, I think they, let me put it this way. When it used to be thought that microbiology was a life science, now microbiology is the science of life and we have to use microbiology that way. And so I have no question that compounds, that mixtures, that various components of microbial preparations are going to be very, very useful in future, in the future treatment of disease. And I would say, I have no question about this. We just have to do it. Well, then there's one problem. That's the FDA. How do we do these things and get around the FDA? I hope so, yes. Who will get the money to do it in the first place? What? Who will get the money? The FDA's got lots of money. But no, but the food and drug rate, yes. But they must be able to support the kind of analyses and work that one needs to be able to use microbial populations. Really interesting talk. A lot of what you touched on is something I've been also kind of trying to think a lot about is like a five o'clock project, maybe sort of looking at with a lab mate of mine, possibly exploring chemotaxis and swarming. So one of the questions that I've kind of been butted up again, maybe similar to your situation, is how these microbials, not microbials, but these small molecules actually persist in real space in the soil and how to test that. So I'm curious if you and your lab have looked at whether or not there's adsorption of specific compounds to the clay, that maybe it's not the aqueous, that maybe it's somehow being in physical contact with the weird biophysics of clay and how that structures around bacteria about if that's having some kind of an effect on these molecules. Okay, that's a very interesting question and you find that people are talking about signaling in soil and things of this type by small molecules. There are no publications that have claimed that they can isolate bioreactive small molecules from soil. They're not there in sufficient quantity or they're bound in some way, but nobody, to my knowledge, has ever isolated what we might call an antibiotic from soil. And that's all there is to it. Very interesting talk. So you said that there are microorganisms in this clay. So are they resistant to the clay? Well, I don't think the clay is necessarily... Wait a minute. I think the clay contains the microbes which will produce the compounds, but I think this happens when you harvest the clay. I can't give you any better explanation than that. We cannot find compounds there and we cannot find compounds in the clay when we've isolated and washed it. It's a mystic... I don't know what you would call it. So if you sterilize the clay, is it still active? If you sterilize the clay, it is still active and we don't have any riddle solutions as to what are the active components. It must be multifunctional. I'll send you some if you like. Just in case I didn't get it when you were talking about it, have you looked at whether the material from the clay can kill escape pathogens with existing antibiotic-resistant mechanisms? It will kill that sort of. It will kill... With existing mutations that have dealt with various kinds of antibiotics. So I guess my question is whether these material in the clay have a new mechanism of action comparing to existing antibiotics because those existing antibiotics even though they're not effective against escape pathogens, they work somewhat. So if you have an escape pathogen that has a very effective antibiotic-resistant mechanism, can this clay still be effective against... Okay, we don't know. We have not tried to isolate strange-resistant clay. Fun? The bacteria in the clay, for example, certainly the acinetobacter is sensitive to the clay. If we isolate clay bacteria, the ones we've tested are sensitive to clay. Colleen. Is this a dinner talk? Yeah, you know, I would... There are just things we can't explain about this. But it's true. Fun? Oh yes, it's probably mineral as much as anything. It must be the right combination of minerals. Yes. So this wouldn't tell you exactly what's in the clay, but I don't know if you can do these experiments on animals, but it'd be interesting to see how the microbiome changes in animals fed with the clay because that might give you some clues as to how it's acting. We're planning to do that with mice. Are you able to release these active ingredients into just the aqueous extraction? Just water extraction? You know, you're releasing these active ingredients from the clay, from the bacteria. Just water? We just sterile distilled water and we respring, we take the clay, we stir it up in sterile distilled water and we centrifuge it. Because we have to use very strong detergent to release bacteria. That's possible. We have not tested detergent. We have not been able to use just aqueous water. We have only tried water. We have not tried detergent. So these are unique bacteria that have very labile membrane? I don't know. Maybe I'll end with the last question, John, and if you can catch Julian after. Julian, when we look out over this crowd, there are lots of young and very bright scientists, kind of, you know, just about to launch their career, their biochemical careers. And you chose an area that was very fertile, right? Which is resistance genes and understanding the biochemistry of it and then beyond that the impact of that, of those genes and those proteins. And so I guess the question I would have for you in the context of this audience is, you know, with your experience, were you to choose again? Do you have any advice to these students and postdocs on how do you find, you know, what's a good kind of step towards really finding that fertile area to launch your career? Well, I wouldn't want them to come and work on clay. That's the one thing. But I would say that the most important thing in terms of developing a career, when you're going ahead to get a PhD and things like this, is to do a good postdoc. That to me is most important. I had some pictures I was going to show you, but I mean, I was very fortunate. I was an organic chemist in Nottingham. I don't think I was a very good one, but I managed to work for Gilbert's talk for two years and it was incredible. I was trained in the UK and I went to Columbia and Gilbert had a bunch of good students and there was a Scott there with me and for about a month we really couldn't understand what the American students were talking about because all they were doing on blackboards was drawing arrows and we couldn't understand what all these arrows were and then finally we realized that they were electrons and we then suddenly realized that you have to think about things in a very different way. I was never taught to think in terms of looking at chemical reactions in this way and that's the way Gilbert did it and obviously a lot of Woodward did the same way. But to do something like that and you have to go somewhere and you have to learn from a postdoc. You're not going to a postdoc to do something cool. You have to learn and learn new approaches. This is critical and I was lucky in doing that, but that's what I would tell anybody. Great, well thanks for ending on the high note and thank you so much for your visit. Thank you.