Slide 1 Hi. My name is Neil Jaworski, and I'm a Ph. D. student in the Hans H. Stein Monogastric Nutrition Laboratory. And today, I'll be presenting the effect of Bacillus species direct-fed microbials on fecal volatile fatty acid concentrations, growth performance, and carcass characteristics of growing-finishing pigs, which I presented at the American Society of Animal Science Midwest Section meeting in Des Moines, Iowa, on March 18, 2014. Slide 2 The outline of my presentation will begin with a brief introduction to direct-fed microbials, and then I'll move into the materials and methods employed in the study, and then the results and discussion about fecal volatile fatty acid concentrations, growth performance, and carcass characteristics. And then I'll leave you with the conclusion and a take home message. Slide 3 Direct-fed microbials, which I'll abbreviate as DFM for the remainder of the presentation, may commonly be known as probiotics. But since 1989, the FDA has required feed manufacturers to use the term DFM. And the definition of a DFM is a source of live (or viable) naturally occurring microorganisms. Slide 4 Direct-fed microbials, as the name suggests, are fed microorganisms that modulate the intestinal microbial population in the pig. Previous research has shown that DFMs may control pathogens, enhance the immune response, increase nutrient digestibility, and increase growth performance. Slide 5 The suggested mode of action for Bacillus direct-fed microbials is that they begin as a metabolically inactive spore. And due to this, they are resistant to feed processing techniques. Slide 6 Next, the pig consumes the DFM with their feed, where it enters the stomach, and the Bacillus spores are resistant to the low pH and pepsin in the stomach. Slide 7 Then, the metabolically inactive spores enter the small intestine, where the pH ranges from 6 to 7, which is optimal for the spores to germinate and grow. The DFM continue to survive and grow by their unique ability to secrete fiber degrading enzymes. The nutrients and energy that may be trapped in the fiber complex, which are typically unavailable to the pig, are now released and available for absorption by the pig. Slide 8 Next, the DFM move with the digesta into the large intestine and promote greater fermentation and volatile fatty acid production, which the pig can utilize as a energy source. Also, the DFM promote a lower pH in the colon, which decreases the growth of harmful bacteria. Slide 9 The objective of this experiment was to determine the effects of graded levels of multiple Bacillus species direct-fed microbials included in complex, high fiber diets on fecal short chain and branched chain fatty acid concentrations, growth performance, and carcass characteristics. Slide 10 The test materials employed in this study were two different direct-fed microbials. The first was fed at 250 or 500 g/MT, which was DFM 1, and the second was fed at 500 g/MT, which was DFM 2. And both of these were a combination of three different bacilli species strains that were specifically selected for their high enzyme producing ability, and therefore, they may be more beneficial when supplemented to high fiber diets. Slide 11 The materials and methods for this experiment were the use of 256 pigs with an initial body weight of 35 ± 3.7 kg, allotted to a randomized complete block design using four dietary treatments, 16 replicate pens per treatment, and four pigs per pen. There were three experimental periods: the grower, which was from approximately 30 to 60 kg body weight, the early finisher, from 60 to 90 kg body weight, and the late finisher, from 90 to 110 kg body weight. Slide 12 Here are the experimental period diets. The ingredients that we used in the diets are listed in the first column, and then the three experimental period diets are shown: the grower, the early finisher, and the late finisher. These are the control diets for each period, with no DFMs included, but three additional diets per period were formulated by adding 250 or 500 g/MT DFM 1 or 500 g/MT DFM 2. These diets were formulated to contain 60% coproducts, which were DDGS, soybean hulls, wheat middlings, and corn germ meal to make up a complex, high fiber diet. Again, because the DFM were specifically selected for their ability to secrete fiber-degrading enzymes. Also, 500 units of microbial phytase per kg of complete diet was added to every diet. Slide 13 The measurements taken were fecal short chain fatty acid and branched chain fatty acid concentrations, and these were taken at the end of the growing and late finishing phase. At the end of each phase, the fresh feces were collected from one pig per pen, and they were analyzed for short chain branched chain fatty acid concentrations. Also, growth performance was measured at the end of each period. And finally, carcass characteristics were measured at the end of the late finishing period using 144 pigs within the final nine replicate pens per treatment. And we utilized ultrasound at the time of weighing. The pigs were ultrasonically scanned, and a transverse image was taken over the middle of the longissimus muscle at the 10th rib, and backfat depth and longissimus muscle area were measured on the image. Using these measurements, we were able to predict the fat-free lean percentage using an equation from Shull in 2013, which is listed at the bottom of this slide. Slide 14 The statistical analysis used for this study was the use of the Mixed procedure of SAS, and the experimental unit was equal to the pen. The model included the fixed effect of treatment, and the random effect of replicate, and four different orthogonal contrast statements were included to assess differences amongst means. Slide 15 Now we'll move into the results and discussion section of this presentation. Slide 16 Here, we're looking at the growing period fecal short chain fatty acid concentrations. On the y axis, we have micrograms per gram of feces, and this is on an as-is basis. And on the x axis, we have acetate, propionate, and butyrate concentrations. The orange bars correspond to the control diet, the blue bars correspond to the diet which included 250 g/MT DFM 1, green bars correspond to the diet that included 500 g/MT DFM 1, and the red bars correspond to the diet that included 500 g/MT DFM 2. The graphs are set up the same throughout the presentation. So here, we see that pigs supplemented with direct-fed microbial 1 and direct-fed microbial 2 had an increase concentration of acetate compared to pigs fed the control diet. There was no effect of DFM supplementation on the concentration of propionate in the feces; however, there was a tendency for pigs fed the diet containing 500 g/MT DFM 1 to have an increased concentration of butyrate compared with pigs fed the control diet. What this indicates is that the direct-fed microbials, in fact, increase fermentation of carbohydrates that are reaching the large intestine, therefore promoting fatty acid production. Slide 17 Here, we're looking at the growing period fecal branched chain fatty acid concentrations. And on the x axis, we have isobutyrate, isovalarate, and valarate. And here we can see that isobutyrate and isovalarate were increased in the feces of pigs fed DFM 1 and DFM 2 compared with pigs fed the control diet. And there was no effect of DFM supplementation on the concentrations of valarate in the feces of pigs. And what this may indicate is that the direct-fed microbials are not particular in fermenting certain substrates. And so undigested branched chain amino acids will also be fermented to produced branched chain fatty acids in the large intestine. Slide 18 Here, we're looking at the total fecal short chain and branched chain fatty acid concentrations during the growing period and at the end of the late finishing period, which are presented on the x axis. And we see that during the growing periods, pigs supplemented with direct-fed microbial 1 or direct-fed microbial 2 had increased concentrations of fecal short chain and branched chain fatty acids compared with pigs fed no DFMs. However, when we look at the end of the late finishing period, there was no effect of direct-fed microbial supplementation to the diets on fecal short chain fatty acid or branched chain fatty acid production. And what this may indicate is that the microbial population in the hindgut of growing pigs may not as capable of handling a high fiber diet as a finishing pig. So if you add direct-fed microbials to the growing diet, it may enhance fermentation, supplying more energy in the form of volatile fatty acids to the pig. Slide 19 Now, moving on to growing period performance. And on the x axis, we have average daily gain, average daily feed intake, and gain:feed. And we saw that growing pigs fed diets supplemented with 250 g/MT DFM 1 had and increased average daily gain. But we saw no effects on average daily feed intake or gain:feed. And if you remember, we saw an increase in total fecal volatile fatty acid concentrations at the end of the growing period. And that may correspond to the slight increase in growth performance shown here. But if we move to the early finishing period… Slide 20 We see the average daily gain and gain:feed were increased due to DFM 1 or DFM 2 supplementation. Therefore, it may have been that the increased VFA production seen on the last day of the growing period may have just begun occurring due to an adaptation of the DFMs in the gut. But once adapted, VFA production increased, increasing the energy available to the pig, which increased average daily gain and gain:feed during the early finishing period. Slide 21 But when we look at cumulative performance, we see no effect due to DFM supplementation. This is associated with the lack of response of DFM supplementation on VFA production during the late finishing period. And therefore, it may be concluded that by the late finishing stage, the pigs have become capable of handling the increased fiber diet, and the DFM no longer increase fermentation; or, the dose of DFM must be increased in order to increase fermentation and VFA production. Slide 22 Finally, moving on to carcass characteristics, with backfat depth, longissimus muscle area, and fat-free lean percentage on the x axis, we see that DFM supplementation had no effect on backfat depth. However, longissimus muscle area was increased in pigs fed diets supplemented with DFM 1 or DFM 2 compared with pigs fed the control diet. Also, there is a tendency for pigs fed diets supplemented with DFM 1 or DFM 2 to have an increased fat-free lean percentage compared with pigs fed the control diet. And this may indicate that the metabolizable energy in the diets was potentially overestimated. And in my opinion, it was the metabolizable energy of the fermentation of the fibrous coproducts that was overestimated. And therefore, when the direct-fed microbials were added to diets, fermentation was increased, and subsequently, VFA production increased, which allowed the DFM-fed pigs to have more energy available for protein synthesis, thus increasing the longissimus muscle area, and fat-free lean percentage. Slide 23 In conclusion, when supplementing diets with multiple Bacillus species direct-fed microbials, we saw that there was an increase in growing period fecal acetate, butyrate, isobutyrate, and isovalarate concentrations as well as an increase in the total volatile fatty acids concentrations in the feces. Also, early finishing average daily gain increased 3.45%, and gain:feed increased 4.44%. Also, a 5.4% larger longissimus muscle area and 4.2% greater fat-free lean percentage was seen in pigs supplemented with direct-fed microbials. Slide 24 The take home message is that using multiple Bacillus species direct-fed microbials, specifically selected for their high enzymatic properties allowed growing and early finishing pigs to increase fermentation, which increased average daily gain and gain:feed when fed complex, high fiber based diets. Slide 25 I would like to acknowledge DuPont and Danisco Animal Nutrition for the project funding. Slide 26 Thank you for your attention. and if you have any other questions pertaining to this podcast or any other information in terms of monogastric nutrition, you can visit the Hans H. Stein Monogastric Nutrition Group at the website listed on this slide.