[Music] thank you to add a sail for inviting me to speak to you today I'm going to jump right into it and if we're gonna talk about the effects of non AGP strategies on inflammation we really need to just think have a brief overview of the immune system and so if we look at the immune system we generally think of it as having two different arms they are in communication with each other they regulate each other in the first aspect is the adaptive immune system and these are the targeted responses involving antibodies and directed cytotoxicity mediated by B and T cells respectively and there is often memory associated with this part of the immune system now the important thing and one of the reasons why I'm not going to focus on this particular aspect in my presentation is that when the adaptive immune response is activated the adaptive immune response itself has a very low energy requirement from the body and so if we talk about the impact of challenges immune challenges on performance generally we're not talking about the effects of the adaptive immune response now because the adaptive immune response is so targeted it's so specific there is minimal collateral damage so the responses are against very particular antigens or cells presenting specific antigens and there's a very focused immune response now the problem with chickens in particularly broilers is that early in life the adaptive immune response is very poorly developed and so if we look at the ability of chicks from 0 to 4 days or from 4 to 8 days there's very little in the way of an antibody response in this case to the bovine serum albumin injection and it's not until about eight days after hatch that the chick is able to to mount much of an antibody response so for the first part of life the chick is really dependent on the innate immune response so the other side of the immune response then is the innate immune system and really this involves the nonspecific mechanisms the body has to prevent interaction with foreign challenges or - if the interaction occurs to respond to that either to eliminate the the challenge or to pass it on to the adaptive immune response and so what I'd like to stress is that when we're looking at innate immunity I think one of the most important things that we can do in poultry production is to maintain a barrier function so exclude the pathogens prevent their from being an interaction in the first place now of course we raise birds in an environment where there is bacteria where there are immunological challenges and so we can't completely eliminate these interactions but as we go through the presentation hopefully I'll show you that one of the ways that antibiotics and alternatives to antibiotics might work best is simply by reducing or preventing the interaction of immunogen's pro-inflammatory signals with the immune system and we can eliminate a lot of problems if we simply eliminate that those negative interactions so one of the aspects of innate immunity is also inflammation involving phagocytosis of invading foreign molecules cell killing and so although there's no memory we don't have memory in the innate immune system like we do in the adaptive immune system the innate immune response can be trained so it can be trained to respond more quickly that's beyond the scope of my talk today but there is an aspect of training the innate immune system now the problem if we're looking at a production environment the problem with in the inflammatory response which is necessary for the bird but the problem with the inflammatory response is not only are there low cool effects at the site of infection or the site of the challenge or encounter interaction with the foreign molecule but there can be systemic effects and really this is where the negative effects of inflammation have an impact a negative impact on performance and so when there's a systemic response we'll see a little bit what that looks like but when there's a systemic response that's when we start to see loss production so the innate immune system is incredibly important inflammation is incredibly important but we want to manage when that inflammation is expressed and also manage the effects that it has now because there can be systemic effects this can be an energetically costly system there can be collateral damage I'll speak more of that in a moment and as I pointed out early in life this is the main aspect of immune defense that the bird has so Kirk glazing did a series of experiments trying to quantify what is the mechanism or what is responsible for the reduction in performance when we see a strong systemic inflammatory challenge and he found that about 71 percent of the total loss in performance total loss of growth is associated with a voluntary reduction in feed intake so when there's a systemic inflammatory response the bird typically responds by reducing its feed intake the more severe the response the more severe the reduction in feed intake and so his question really was what is the other 29% what mechanisms if feed intake is responsible for 24 sorry for 71 percent of the loss of growth or loss of performance what contributes to the other 29% that can't be explained simply by the birds not wanting to eat as much so of that 29% through a series of experiments he found that digestive in efficiencies so reductions in nutrient absorption we saw some examples of that in previous talks accounts for about 9 percent of that 29 percent the energy expenditure related to increased body temperature the fever the systemic effects of increasing temperature and the energy that requires accounts for another three and a half percent metabolic inefficiencies mobilization so birds reduce their feed intake or stop eating but they still have a very high protein turnover the liver total body protein synthesis increases dramatically because the liver is synthesizing acute-phase proteins and so the body actually mobilizes skeletal muscle in order to feed the requirement for amino acids from aunt acute phase protein synthesis in the liver and that accounts for about another eight percent and then the systemic immunity the cost of the immune descent defenses accounts for about another five percent so overall that accounts for most of the loss performance and so I'm not going to go into a lot of detail but just keep in mind as we go through this presentation when we activate inflammation when we cause a systemic inflammatory response most of the effect is due to reduced feed intake but there are also things going on physiologically in response inside the bird metabolically that are causing decreased performance and so when we look at antibiotic growth motors and their replacements and how they might function really what we're trying to do is reduce the cost first of all of bird voluntarily decreasing feed intake and second of all these mechanisms that influence the metabolism of the bird when it's faced with an inflammatory challenge so I'm not going to go through this entire list there are a number of proposed mechanisms by which growth moding antibiotics work but what I really want to focus on is the reduction or prevention of the inflammatory interaction between the host and the immunogen and so that interaction that causes the host to respond now there are pathogenic bacteria present in the digestive tract of chickens there are pathogenic bacteria present in the digestive tract of each one of us it's when there's a an imbalance or an over reaction and I don't mean that in a s in a protective sense when when there's a strong reaction to something that's come out of balance this is when we typically see a reduction in performance and typically where we see a beneficial effect of antibiotic growth promoters so if we can do that if we can reduce that interaction or reduce the severity of the interaction we can reduce the maintenance costs associated with the turnover the intestinal epithelium we saw again some examples of that earlier in this symposium and if we can do that we can reduce the inflammation induce growth depression which again of course is associated with reduced feed and water consumption and also a reduced efficiency of the nutrients that are utilized and absorbed so this was some classic work out of Kirk's lab again and the basic design was birds were raised in either a clean or a dirty environment and fed a diet containing either no antibiotics or with antibiotics and you can see that in a clean environment antibiotics had no effect on performance they had no effect on feed efficiency but when they looked at inflammatory mediators in this case interleukin 1 they found that in the clean environment antibiotics had no effect on systemic inflammation but in the dirty environment the antibiotics reduced the systemic inflammation and of course over the years there's a lot of indication that at least part of the mechanism of antibiotics growth promoting antibiotics is reduction in systemic inflammation and whether that's a direct or an indirect effect we can talk about so when we remove antibiotic growth motors there we have an opportunity or a series of opportunities for a wide range of different pathogens to take hold to interact more strongly with the immune system and particularly the innate immune system of the bird and so when we talk about removing antibiotic growth motors really what we want to do protect birds from poultry pathogens that could cause disease or reduce performance probably a little bit outside of the scope of my talk I'll touch on a briefly later we want to protect humans from foodborne pathogens which are not always recognized or at least not always responded to by the birds immune system very often foodborne pathogens are tolerated by the immune system of the bird and at a certain point every analogy falls apart but really what I want to illustrate with this is we want to we want to minimize the chance that a pathogen is going to access the bird okay now I'm aware of course of the importance of communication between the microbiome and the bird I'm not trying to imply that we want to completely eliminate the microbes from the gut but what I want to do is say we want to have a separation from bacteria that will cause a decrease in performance and separate that from from the immune system the reaction that would cause systemic inflammation and so ultimately we want to reduce negative interactions between the host and the gut microbes of course there are lots of interactions but we want to focus on the negative interactions that are going to decrease performance okay so if we use that analogy then and we think of that innate immune system as a fence if we think about the goal whether it's with antibiotics or antibiotic replacers to decrease that interaction that negative consequences of the interaction between the host and the the microbiome we can think of the antibiotics as a fence and if we're going to replace them we would need to replace them with effective alternatives and so as I go through these various options for replacements we need to be aware of the fact that antibiotics have been very effective those of you that heard Greg Mathis is talk this morning he believes and I agree with them that nothing is as good as antibiotics and certainly nothing individually is as good as antibiotics and so if we're going to replace antibiotics we can't just think about one thing that's going to cover a small part of the potential problems now we might have a good system or a good series of products but if there's a hole in the system we're still not offering protection sometimes the problem might be bigger than we anticipated we may have to deal with that and then sometimes the problem might be something that we just really never ever thought would happen so antibiotics have been very effective they've been used very effectively for many years they've covered a lot of bases but as we're looking for antibiotics we need to make sure that if we're looking for a replacement of that function that we're covering as broad a series of possible outcomes that we could anticipate again this has been or aspects of this have been talked about earlier so I'm not going to spend a lot of time about but when the innate immune system responds with inflammation we tend to see reduction in barrier function we tend to see a reduction in absorptive function of the intestine and so when we have birds that are clinically ill we see those negative effects of systemic inflammation so the point I'm trying to make here is that we can have a negative impact at the site of the infection the level of the gut but we can also have systemic effects at the level of the burden so as we look for replacements for antibiotics we may need to look at both both levels of effect I'm going to spend very little time talking about nutritional immunomodulation there are many examples of nutrients that are truly immunomodulatory vitamin E and carotenoids selenium 25 hydroxy vitamin d3 I'm just going to spend very briefly a couple of minutes talking about omega-3 fatty acids as an example of a way that even with a pathogenic interaction even with a local inflammatory response we may be able to change the metabolism of the bird and reduce the negative effects so this is some work that we did many years ago we fed a diet high in omega-3 fatty acids corn oil based diet we fed a diet that was high in omega sorry corn oil based diet high in omega-6 fatty acids a fish oil based diet high in omega-3 fatty acids we fed an experimental anti-inflammatory called low foreign so within each diet we had a an anti-inflammatory and no anti-inflammatory and then within each of those treatments we had infected birds infected with I'm area and non infected and then we looked at performance post injection so when we fed the omega-6 enriched diet without the anti-inflammatory drug we saw a negative effect on growth rate with the I'm area infection but when we blocked inflammation we saw no effect on growth rate following that induction of coccidiosis in the fish oil based diet so high levels of omega-3 fatty acids omega-3 fatty acids are typically anti-inflammatory we saw no impact of performance on performance whether or not we had the the anti-inflammatory drug so what we are seeing is the blocking of inflammation reduced the systemic effects of the I'm area infection now when we looked at the histopathology we did see a local response so what we what we did was we saw a separating and unlinking of the local inflammatory response the response to the pathogen being present the parasite being present from the systemic effects on growth rate so this is happening within the body of the bird this is happening after the infection and of course if we look at the effects of fish oil on inflammation as we increase the level of fish oil in the diet we see a decrease in in the interleukin 1 activity which is a pro-inflammatory cytokine so we have a couple of lines of evidence here that an anti inflammatory environment decreases the systemic effects of inflammation but I think when we want to look at replacements we're probably talking about dietary ingredients so things that we're going to add to the diet not necessarily that are going to affect metabolism like nutrients but ingredients as opposed to too nutrients so these products that are going to potentially replace antibiotics are generally going to function at the level of the gut lumen rather than being absorbed and having an effect physiologically they may or may not have anti-inflammatory effects I think if they're effective at least part of it will be a direct or an indirect effect on inflammation and I'd like to make clear that I'm not endorsing any one particular class of products if I don't talk about your favorite antibiotic replacement product that doesn't mean it's not relevant just in the limited time that we have you know I think there's a lot of potential products out there and in some cases there's lots of data in other cases there's limited data so I've just picked a few examples of some possible antibiotic replacements and then I'm also going to talk about the idea that as I said if we only have a fence that six inches wide we're probably going to miss a lot of the potential challenges and so I think we need to have combinations of products with different mechanisms so now the other caveat I'd like to give is that I would encourage my students when we talk about things like this look for a trail of evidence so it's interesting when there's a study that shows a certain product has an anti-inflammatory effect in cell culture or at the level of gene expression in a particular tissue or even at the expression of protein and levels in various tissues but I would remind you that really what we want to do if we're looking for effective replacements for antibiotics we're looking for products that show an effect on performance and in order to do that to show that smoking gun we need to be able to tie those anti-inflammatory measures into the performance of the birds and I'll just mention of course the the proper the importance of properly designed and controlled experiments so often when we read the literature we see antibiotics were fed product a was fed they were both fed to birds in clean university environments there was no difference in performance so product a obviously as a replacement for antibiotics so be a little bit critical when you read the literature so there there should be a negative control there should be a challenge and then evidence that the product replaces or restores the level of performance to that of the of the positive control okay so going through some examples probiotics or competitive exclusion culture cultures these have typically been marketed as competitive exclusion cultures and certainly that is possibly part of the effect but there is also likely other effects that that the effective probiotics are showing I mentioned the importance of properly controlled experiments and here's an example so if you look on the left hand of each panel is the control the positive control with oxytetracycline and on the right is the negative control with nothing and then going from right to left is increasing levels of the of the probiotic so in the first panel we can see a response in terms of body weight and on the bottom left hand panel a response in terms of feet converters and racial and that is in fact linked with a decreasing level of inflammation in terms of - expression in the jejunum we heard the difference between prebiotics and probiotics earlier today prebiotics are compounds that are indigestible by the bird but they are digestible by specific classes of commensal microbes and these may provide a competitive advantage advantage for those microbes against bacteria that can't digest the prebiotics and so typically we would hope that those are pathogens and so the idea the simplified idea is that we're feeding the good bacteria and so it's not only that we're feeding the bacteria them a competitive advantage it's that many times the metabolites that are produced by the beneficial microbes when they consume these prebiotics are metabolites that may have a positive impact on the health of the gut or gut function gut barrier function and again we saw some examples of that earlier today this is some work that we did in collaboration with Steve Rickey at the University of Arkansas we fed laying hens a number of different products so we had our control feed we had a it was an in vitro incubation of cecal contents from laying hens with Salmonella typhimurium enteritidis so when we incubated the Salmonella with the feed only or with the cycle contents from birds fed no prebiotics we saw almost no reduction in Salmonella but as we added different yeast products we saw a significant reduction over time during incubation associated with most of these yeast products so again these different East products different fractions and so without getting into what product is which here's an example where when we provide what likely contained some prebiotic components the microflora present in the Sica were able to out-compete the Salmonella Salmonella numbers went down over time another category would be the all of the saccharides man and all of us a carides derived from yeast so there were there was very likely a man in all of the saccharide effect in the previous slide fructooligosaccharides derived from various plants containing fructose chains and of course Xylo oligosaccharides derived from non-structural plant carbohydrates Xylo oligosaccharides I'll talk about again in a minute the idea here is that the the all of those saccharides will minimize or reduce the interaction between potential pathogenic bacteria in the gut and the host by binding to the pathogenic bacteria the pathogenic bacteria are bound to the all of the saccharides they cannot bind to the intestinal wall and they need to bind to the intestinal wall in order to colonize and cause problems and they're essentially flushed out through with the died with the excreta enzymes I think enzymes show a lot of promise in terms of being part of the solution again I well I don't hesitate I just outright don't recommend thinking about replacements for antibiotics and saying we're taking antibiotics out we're putting one product back in but I think enzymes can play an important part in this particularly if we're feeding wheat based diets but we've got some evidence that I'll show later on that even in corn based diets these soluble non-starch polysaccharide asses may have a positive influence so typically we think of them as reducing the viscosity making a less hospitable environment for for the the pathogenic bacteria we also think of them as breaking down or making nutrients more accessible for the bird and therefore less successful for the pathogens but there's a lot of evidence accumulating that the all of us saccharides that are released as part of the enzymatic action may also have a prebiotic effect and and feed the the beneficial bacteria there's some work going on in various labs looking at using enzymes to degrade stuff that's in the in the digestive tract in the diet that can trigger the immune response or can can degrade molecules that are essential for microbial survival or growth and again in particular we're focusing on the pathogenic bacteria so enzymes that break down beta Manas or proteoglycans basically reduce the viability of bacteria by cleaving flagellum and others and I think this is a rich area of potential potential antibiotic replacement work organic acids organic acids are those that or organic acids don't dissociate completely in water when they're fed in the diet and in a Qui environment they're taken up by the bacterial cells and inside the cells they do dissociate and essentially what happens then is the bacteria spend so much energy pumping out the excess hydrogen ions that they have very little energy left to think about reproducing so they there's again a lot of evidence that they have a positive effect in terms of reducing numbers of potentially pro-inflammatory bacteria we heard a lot about butyrate earlier on so i'm not going to say too much about that other than just to remind you that butyrate is a preferred preferred fuel for intestinal cells and there are direct and indirect effects on systemic and local inflammation and then essential oils essential oils there's a lot of work been done with essential oils these are typically typically fairly good smelling oils from various herbs and spices and they have a direct effect on the microbial cell permeability again decreasing the viability of of pathogens or potential pathogens by decreasing microbial membrane activity integrity this is an example from laying hens so again here's an example where if we look at a performance so over a six-week period the inclusion of increasing levels of a combination of essential oils derived from from trees we see an increase in egg production with increasing levels of the the essential oils we see an increase in egg mass and a decrease in feed conversion ratio so there's our effect on performance that is associated with changes in the gut microflora so a decrease in potential pathogens staph aureus and e-coli an increase in what would commonly be considered a beneficial microbial species lactobacillus in a related experiment they looked at response to LPS injection LPS is a bacterial cell wall component the immune system responds to it to it as if it has an infection and so the inflammatory response is very strong and so when the birds are fed increasing levels of these essential oils we typically see an expression of cytokines that would be associated with a more oh sorry a less inflammatory situation so again here's an example of a physiological effect at the level of the bird a performance response and an indication that the effect is mediated by a reduction in inflammation okay so coming back again if we're looking at antibiotic free production antibiotics were very effective they are very effective where they're being used but the world is moving towards a reduction or an elimination of antibiotics and so the microbiologists are probably going to throw things at me when I use broad-spectrum in this way please beware I understand but antibiotics covered a lot of sins it allowed allowed a lot of mistakes to be made without bad consequences when we take the antibiotics away there are a lot of things that can directly or indirectly predispose the birds to a pro-inflammatory situation the stress associated with vaccination poor nutrition or changes in nutrition poor water quality a poor level of maternal antibodies that might predispose the birds to viral diseases that could cause secondary or could allow secondary bacterial infections brooding temperatures cleaning and disinfection all these things could potentially increase the risk of a bacterial infection that would negatively impact performance so again you individual products maybe may be good they may have an effect but I think if we're going to look at a commercial environment differences from farm to farm from barn to barn over time we need to look at having a lot more bases covered with our strategy to replace antibiotics so we we probably need an approach that involves multiple products with multiple mechanisms because different pathogens require different strategies things change over time the challenges change with the source of your chicks with the source of your feed from season to season and so on so if we have complementary strategies if we have multiple products that complement each other we may be more likely to cover that broad range of potential challenges that antibiotics were able to do this is some work we did in Ecuador this is in a commercial integrator we had we had believe five million birds in this experiment all treatments were replicated within the same farm we had our standard diet that contained our contained the antibiotic remote growth motor we had a treatment that removed the antibiotic growth motor and we added some a probiotic we added a high level of phytase and then we had a third treatment where we removed a few more of the antibiotic potential antibiotic replacements so again I acknowledge this is a field trial we didn't have a negative control treatment because nobody is willing to take antibiotics out and not replace them with something so we had no negative control what we did find under commercial conditions with five million broilers distributed across nine different farms there was no effect on body weight of taking the antibiotics out when we had these replacements in and no effect on European performance efficiency factors so again we're lacking a negative control that limits our interpretation or what we can say about the trial but under commercial conditions we were to do that so to summarize we need to be aware of what antibiotics did from a functional standpoint but we need to be aware of what antibiotics replacements can't do they cannot cure bad management we can't simply say now that we've got a reliable antibiotic replacement we can let things slip antibiotic replacements can't overcome every situation we will still have outbreaks we will still need to treat with therapeutic antibiotics but we need to be proactive we need to prevent the establishment of disease we need to prevent as much as possible the negative interaction between the microbiome and the inflammatory system of the bird and they can't provide complete protection from infectious disease so when there's an Ebola outbreak the people working in those conditions take a lot of precautions they're not given an aspirin and a thing yogurt and said here you go so we need to be aware just like antibiotics we can't rely too heavily on them we have to have good management we have to recognize when we need to go in and treat therapeutically and so really the antibiotic replacements are the cherry on top to good management overall thank you very much [Applause] [Music]