[MUSIC] Hello, everyone welcome. This is the first video of a course on Antimicrobials, Antimicrobial resistance, and susceptibility testing. And today, I'm going to talk to you about antimicrobials and then on to Antimicrobial classification. My name is Lina Cavaco, I'm a senior researcher here at DTU at National Food Institute and I work in the research group for genetic epidemiology and this group is also the WHO collaboration. Centre for Antimicrobial Resistance and the European Union Reference Laboratory for Antimicrobial Resistance as well. This course has been built up with the support also of co-funding of an EU commission and we are going to have a basis module on antimicrobial susceptibility testing that starts with this video. So to start with, we are going to talk about Antimicrobials and the classification of these drugs. And first of all we would like to answer the first question, what is an antimicrobial? What is an antibiotic in general terms? Many of you have heard about them, many of you have taken some of these drugs when you are sick, and a lot of us actually still live because maybe at some point, we have been treated and cured by antimicrobials of an infection that could have a more serious ending. So, antimicrobials are drugs that we have since the 19th century and they have been very, very important for us. And their definition lies in the possibility of killing microorganism or of inhibiting, that's the general terms. But the most complete definition would say that, an antibiotic is actually a was produced by a microorganism because the first ones were like that, and that has the capacity to selectively inhibit or kill microorganisms. And that means that it doesn't really harm the host, and that's very special feature because it is a toxic substance and the host should not be affected by it. When we talk about antimicrobials, we broaden a bit the term because some of the drugs are not produced by microorganisms anymore. Humans have been able to make antimicrobials by synthesising them or by modifying the drugs that were produced by microorganisms. So, antimicrobial is a term that is used more broader. As I have mentioned before, since these are some of the very toxic substance that are available in the world. It is really an important feature that they affect microorganisms. That we want to get rid of, but not affect much the cells in our organisms, so the eukaryotic cells. So, how are they used? Well, we know that they are used in humans and they are also used in animals. They use in humans is quite widespread, so wherever there is an infections that can be treated by antimicrobials is not all infections that need them. But wherever there is, we can and may use antimicrobials to make this infection go away or to at least get it to a level where our immune system can take care about the rest. So, antimicrobials are used mainly to kill or inhibit bacteria that cause infection and in the end, disease of the humans. This is very widespread. These are some data from the US, but in EU is the same thing all over the world. Antimicrobials are used in patients in and outside of the hospitals and they really are necessary in the case of bacterial infections. In animals they are also used. Animals also get sick with infections. In here, there's the use for Therapeutic use, to treat really sick animals that have an infection. In some cases there is a more prophylactic or metaphylactic use, which means that if some animals are sick already in a group or we know that an infection could spread, then it's possible to prevent infections by treating already the group of animals that could get the infection spreading. In some terms and still in some countries this is possible. Antimicrobials have another effect that they not only have the antimicrobial effect, but they also have an effect over the growth rate of animals. And this has been used in the past, at antimicrobial were used to increase the growth of animals and increase the production in that way. This is forbidden in the EU for some years now, but some countries in the world still have a growth promoter use, which is called in this case. As we are going to talk in the course later on much more on antimicrobial resistance and also in susceptibility testing and detection of resistance. I introduce it briefly in this lecture, but it won't be the focus of the rest of the lecture. And here I just introduced that Antimicrobial will be. A relative term because it really depends on the situation. It depends on the bacteria. And it depends on the susceptibility levels, this bacteria has in terms of, which antimicrobials we can kill it with, or inhibit it with. And we are going a bit a head on some of the definitions in this area. In terms of resistance, we always think about the clinical definition because the end goal is to treat the patients, or to treat animals. And here the resistance of antimicrobials would be the possibility of an anti microbial acting, but not enough to inhibit or kill in that situation that the person, or the patient or the animal gets better. In that way resistance would be the ability to survive during treatment. But there's other definitions to resistance because we work in microbiological labs and we have also insides to genetics and therefore, the genetic definitions would be that there is a mechanism in these bacteria. That there is a mechanism that makes them more resistance as we can see then in the clinical case. So, we can find that background. And also in the laboratory when we don't have the genetic possibilities is to go into the whole resistance determinant, but we can also test phenotypically. We can do some in vitro tests and that's the main focus of this course is to do susceptibility testing, and see if resistance is there and in this case, resistance is the ability of one bacteria of a certain species to be able to survive in higher concentrations than other of the same species. So in that case, resistance is determined in that way. But we are not going to focus much more on resistance in this lecture As we are going back to antimicrobials again, to work a bit on the classification of this. So in terms of classification, we can have several different classifications. The chemical one, depending on the chemical composition of the drug, and also in terms of the action and the spectrum of these drugs. So based on the chemical structure is one, and based on target organisms we can also have that a drug could be against bacteria only, against virus only, against fungi, and against parasites in some cases. In general terms we normally speak about antibiotics and antimicrobials in terms of antibacterial. The other drugs are normally designated as antivirals or antifungal or antiparasitic in that case. [COUGH] And in terms of activity, we are going to explain this a little a bit ahead. It has a division in terms of is the antimicrobial killing the bacteria, and in that case, it would be classified as bactericidal. Or is it stopping the growth to a level, and that is bacteriostatic. And in terms of spectrum of activity, we can have antimicrobials that have a very, very large spectrum that you can use for many species of bacteria that could be present in an infection, or a very narrow spectrum that are targeted to smaller groups. Just to note, some drugs are one thing in one case, and another thing in another. And some, for example, sulfonamides are actually so broad in spectrum that they also take some parasites, for example, and not only bacteria. In terms of origin, we all have heard probably the story of Alexander Fleming that found penicillin in 1928, and that was the first drug discovered, it was a natural drug produced by a fungus. And after that, other scientists try to find other organisms producing antimicrobials. And therefore, in the years after, several drugs were discovered. Many of them produced by microorganisms, either fungi or bacteria. And several groups of antimicrobial were found in this way. At a certain point, also the chemistry evolved, and some of these classes of antimicrobial were enlarged, not by finding more drugs in the natural world, but by modifying them. Just to show you in another page, this is a large table so you can probably, I'm not going to go through this, but I have another page where it will continue. It is chronologically and a little bit by groups, the classification that you will see that towards later on some drugs were not only the ones that are were natural or modified, but then some synthetic ones were coming up. Those were entirely created by man then. So, moving on into the activity, we have a definition that what is a bactericidal drug versus a bacteriostatic drug? And I tried to show this in a graph where here at the start of the graph there is an infection, there is a number of bacteria that are growing naturally in the organisms. And at this point you give a drug, and if it is a bactericidal effect drug, it will drop the number of bacteria in the site of infection. But if it is a bacteriostatic drug, it will actually not drop, it will just stop and inhibit the growth. And this means that the organism will have to take care of the rest, not letting it go further, so the immune system will have to take care. If we haven't given the drug, probably it would go up, and it could be getting much worse, the infection, or even the patient could die. So here there is in a schematic way, it's not so black and white always, but in a schematic way it's how bactericidal drugs and bacteriostatic drugs act. And as you can see here, it is quite a difference, and sometimes we really need these large drops of the bactericidal drugs, because the patient might be in risk or the patient might not have any immune system that can take it. So essentially in life threatening infection, immunocompromised patients and in situations where the infection has to be taken care of rapidly, bactericidal drugs are preferred. But, bacteriostatic drugs are not always like that. It might depends on how much of the drug is present, so the concentration, the dose. And it could also depend if we give other drugs together with that one, it could increase the effect and become bactericidal, and it might also depend on which species of bacteria, because bacteria don't have the same sensitivity to the drugs in that way. So, going a little bit ahead, I have here a table which I choose, some of the groups of antimicrobials in terms of if they are classified as bactericidal or bacteriostatic. And here you can see that, well many of them are bactericidal, but macrolides, chloramphenicols, and phenicols and tetracyclines and sulfonamides could be bacteriostatic in some occasions. And this has this effect that I just mentioned before. But as this course is focused on testing and on susceptibility testing as such, this could also have some implications in the reading of the tests and the interpretation of the tests, and that will be shown in other lectures ahead in this course. So just to show you with a little more space, some of the bactericidal drugs which are quite large groups that are normally used, and some of the bacteriostatic, as I just mentioned before as well. So in terms of treatment, while these are the side of the effect between the drug and the bacteria, but that's not the only thing that happens when we treat a patient or we treat an animal. The antimicrobial effect has also to be possible to see in the body of this patient or this animal. And therefore, we also have to take in account some pharmacokinetical and pharmacodynamical properties of these drugs as such. So these drugs have properties that should be taken, or parameters that should be taken, in account when calculating doses, when arranging how the frequency and how the treatment should go. And I will just give a brief definitions of pharmacodynamics and pharmacokinetics so you also have brushed up a little bit on that without going too much into detail. In terms of pharmacodynamics, this is basically what the drug does to the microorganism. So it is how it kills, the mechanism for killing. How it is in terms of time or in terms of concentration. This is more independent on time or in concentration. And then there are some measurements. You can measure the minimum inhibitory concentration which will be a major focus on this course and you will learn several ways of measuring this minimum inhibitory concentration. Which is the minimum concentration that is able to inhibit or kill the bacteria. And then, for example, also this minimum bactericidal concentration which would not be just inhibiting or killing, it's really only that nothing is left alive so minimum bactericidal would be just killing in this case. In terms of pharmacokinetics, I also have here some of the parameters, but here's it's different. Here's what the body of the patient or the animal does to the drug. So this all about how the drug enters the organism, how it's administrated, how the drug is formulated, how is it absorbed, how is it distributed, and how does it get to the place where it should act, basically. So the parameters here are a lot about, well the concentration, the maximum concentration that will be achieved in the plasma or in another organ for example. The time at which this concentration reaches the maximum, the half life of the drug where the drug is reduced to a half of the concentration. And the area under the curve, which represents the overall presence of the drug in the targets. So these parameters, the pharmacodynamic and the pharmacokinetics, are studied by the pharmacologist before even the drugs are launched in the market. And when they do some of the studies, before the drugs are launched, so that doses and frequency and how it is administered and so on can be decided. So, here we go towards the basic pharmical-dynamic parameters that we actually test in our labs, so the others normally are studies done by the pharmacologists, but in our microbiology labs. We need sometimes to do this inventory concentration. This also mean in bactericidal concentrations sometimes and that is a rather simple essay. We have several lectures on this and we are going to tell you about the methods and how these are standardized and so on, but here just for a basic slides for how a MIC is determined. A MIC is determined with a row of, well it could be in tubes, it could be in agar plates. We are going to focus on that later, but it is determined with two-fold concentrations of a drug. Where each tube or each plate has double concentration of the next one and of course the control that doesn't have any drug. Then the bacteria are inoculated and inserted in an amount which is very standardized, and then we are going to observe growth after the incubation. That's normally 24 hours, depending on the organism. And we are looking for growth in the smallest concentration, because there's not enough concentration to kill or inhibit the bacteria and then at certain point, the growth disappears. And the minimum concentration where the growth has disappeared is our minimum inhibitory concentration and that's the term visually or maybe by some equipment that can make this reading automatically as well. And in this case, where we have this growth of concentration and the growth is here and the growth can be observed there, our minimum individual concentration can be one and can individual concentration is one and normally it is expressed through milligrams per liter or micro-grams per milliliter, which is the same. But this doesn't tell us about the bactericidal effect. For that we would need to take up from these tubes or these space and take this up into a solid media and then try to grow them again. And in some of these tubes where we don't see any growth here, actually something might be surviving and goes up. So the minimum bacterial concentration is not necessarily the bacteria side 01. In this case, for example, there's still growth for a few concentrations and then here there is no growth, so actually the minimum bactericidal concentration would be 8 which is a little bit higher than for the MICs, than the MIC. And this tells us also something about the effect of the drug. Normally for bactericidal drugs, the MIC is very close or really just one step or something like that from the MBC. In bacteria static drugs, there could be a little bit more distance between them. So, moving ahead. We also I have doctors and veterinarians, when they do a treatment, they have assess the cases and not always it's possible to use one drug, one antimicrobial. It sometimes is necessary to associate antimicrobials and use two at the same time or even three, depending on the cases. Some of these associations are very classical like amino penicillins with clavulanic acids and sulfonamides with trimethoprim which come already in the same package and everything. And sometimes we will have to administer two drugs at the same time to the same patient. This is done normally for several reasons and among them to increase the efficacy, to increase the spectrum. If you give one drug that is more for gram negatives and another that is more for gram positives, you increase the range of bacteria that you can kill or inhibit. And sometimes for synergy, and this means that both drugs alone have an effect, but when they are added together, they have more effect than the two effects added up only. So it has a synergy effect, but this is not always true that when you give two drugs or three drugs together its better. Sometimes it could be that actually worse and one of the classical mm, cases for this is of course chemical interference or side effects issues but also the bactericidal, bacteriostatic effect. Which, in that case, if you give a bactericidal drug together with a bacteriostatic, it could be that the bacteriostatic drug inhibits the growth but the bactericidal drug actually needs bacteria that are actively growing to be able to kill them. So, in the end it doesn't have such a good effect because the bacteria are not actively growing. So, in that way there could be some antagonist in these drugs. So it's not always good and, of course, the doctors have an idea which combinations are beneficial and which are not. So this was all for today. I hope you have learned a lot and I'm happy to show you other videos and the other videos will focus much more on susceptibility testing and on the methods and the standardization of the methods. If you have other issues you would like to look up on in our laboratory, you also welcome to go to our website and to search for them. Thank you very much. [MUSIC]
