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Influence of the Microbial Community on Intestinal Function

Investigators
Applegate, Bruce; Patterson, John
Institutions
Purdue University
Start date
2007
End date
2012
Objective
The overall goal of this project is to improve animal health, performance and human food safety. There is an alliance between the intestinal microbiota, the epithelial lining of the intestinal tract and the mucosal immune system that influences animal health and shedding of pathogens, which influences human food safety. Various stressors (transportation, temperature, food restriction) may negatively impact one, or all of these systems, making the animal more susceptible to subclinical or clinical infections. There is increasing evidence of extensive cross talk between the three systems that influences the efficacy of the alliance against pathogens. The intestinal microbiota inhibits colonization of the intestinal tract by pathogens and the microbial community structure of the intestinal microbiota changes with diet, stress and antibiotic treatment. However, there is little information on the impact of specific components of the microbial community on animal health and performance, or on the mechanisms of action of specific components of the microbiota on the epithelial lining and the mucosal immune systems, or on pathogen colonization. Specific components of the microbiota may also negatively, or positively influence animal performance in the absence of pathogens.

The specific objectives of this research are:

  1. To determine the effect of stressors on the intestinal microbial community structure, epithelial function and immune status that affect animal health, performance, and human food safety.
  2. Determine components of the intestinal microbiota that are associated with either improving or decreasing animal health, performance and pathogen colonization.
  3. To determine the effect of dietary modifications (nutrient level, probiotics, prebiotics) on the microbiota, epithelial and immune alliance and subsequent effect on animal performance and human food safety.
More information
NON-TECHNICAL SUMMARY: The intestinal microbiota, lining and immune system work in concert to influence intestinal health, animal performance and human food safety. Common production practices provide a number of stressors that suppress one or all of these systems, thus increasing susceptibility to infection. This project addresses factors that influence interactions between the intestinal microbiota, lining and immune system that affect animal performance and shedding of pathogens. Dietary and management practices that enhance the interactions between these systems and improve animal health will be examined to determine effective approaches to improving animal health, productivity and human food safety.

APPROACH: To determine the effect of stressors on the intestinal microbial community structure, epithelial function and immune status that affect animal health, performance, and human food safety. In vitro approaches will determine conditions under which pathogens may be excluded by defined or undefined microbial populations. In vivo approaches will determine the impact of stressors on functioning of the intestinal microbiota and the effect of dietary supplements on the microbial community structure. Collaborators will address the influence of stressors and dietary supplements on functioning of the epithelial lining and the immune system.

PROGRESS: 2006/10 TO 2007/09
OUTPUTS: One experiment in swine and two in poultry were conducted and samples are being analyzed for changes in microbial community structure. The swine study addressed the transportation stress and the efficacy of a beta-glucan product on immune status, ileal morphology and microbial community structure. A broiler study was conducted to determine efficacy of replacing soybean meal with coconut meal on bird immune, intestinal morphology and intestinal microbiota response to challenge with either salmonella or IBDV. Broilers were subjected to a 24 h fast and the effect of diet on attachment of salmonella to ileal tissues was also conducted.
PARTICIPANTS: Patterson, J.A. Principal Investigator, in charge of project and microbial community structure analysis Applegate, T.J. Co-investigator, in charge of determining changes in tissue morphology Eicher, S.D. Co-investigator, in charge of determining changes in immune paramters Radcliffe, J.S. Co-investigator, in charge of determining changes in intestinal transport function Davidson, M.K. Collaborator, in charge of determining salmonella colonization and shedding Long, T. Collaborator, in charge of determining viral induced changes in tissues Wu, C.C. Collaborator, in charge of producing viral inoculum and determining viral colonization
TARGET AUDIENCES: Animal Nutritionists

PROGRESS: 2005/10/01 TO 2006/09/30
Recent basic research has expanded our knowledge of the importance of interactions between intestinal microorganisms; and interactions between the intestinal microbiota, intestinal epithelium and mucosal immune system, which are involved in maintaining animal health and wellbeing. We have a collaborative group of researchers interested in these interactions, the effect of stressors on these interactions, and dietary and management approaches to manipulate these systems to enhance animal health and wellbeing. Probiotics and prebiotics, short term and long term heat stress and food withdrawal increase variability in microbial community structure in vivo and attachment of salmonella to ileal tissues in an in vitro ileal challenge model, suggesting diet additions and environmental stressors reduce the ability of intestinal epithelial tissues to resist colonization. Transportation stress alters immune function in pigs and lairage may mitigate some of these responses. Transportation also alters intestinal microbial community structure and these alterations are influenced by lairage. Lairage increases variation in microbial community structure, initially in jejunal contents and subsequently in cecal contents. This increase in variability is consistent with the concept that disruption of the intestinal microbiota decreases intestinal resistance to infection. Replacing soybean meal with coconut meal had no influence on carriage of salmonella in broilers orally challenged with salmonella. However, salmonella did not significantly colonize pasture-raised broilers over 3 days, but did colonize the ceca of intensively raised broilers. These data indicate that dietary, environmental and stress factors influence the ability of livestock to respond to pathogen exposure and to resist pathogen colonization. Approaches that minimize management and environmental stressors may improve livestock health and wellbeing and diet manipulation may be useful for enhancing health of livestock.

IMPACT: 2005/10/01 TO 2006/09/30
Clinical and sub-clinical enteric infections in livestock significantly reduce animal performance and negatively impact human food safety. Interactions between the intestinal microbiota, the intestinal lining and the intestinal immune system are important in maintaining health of livestock. Mechanisms by which these systems interact are being examined. We have shown that diet and environmental/management stressors (feed withdrawal, heat stress, transportation) negatively influence the immune system and disrupt the intestinal microbiota. Fasting and heat stress significantly increase attachment of salmonella to the intestinal lining in in vitro studies. This data demonstrates the importance of intestinal interactions in maintaining intestinal health and hold promise for developing dietary and management approaches to minimize enteric infections in livestock and enhance human food safety.

PROGRESS: 2004/10/01 TO 2005/09/30
Past dogma on host animal defenses against infection suggested that the intestinal microbiota, epithelial lining and the mucosal immune system combat pathogens that colonize the intestine. Function of all of these systems is detrimentally affected by stressors. Recently, is has become clear that there is extensive crosstalk between the three systems and that normally they act as an integrated system and each is important for the development and function of the other systems. Stress tends to suppress the function of all of the systems. However, there is sparse information on the effect of stress on these systems in livestock species. Enhancing beneficial functions of each system and their interactions that influence animal health and production would be beneficial to producers and may lead to reduction of the use of growth promotant antibiotics. We have conducted several studies to address these issues in poultry and swine. Copper sulfate is used to suppress pathogenic bacterial populations. In vitro, high concentrations (250 ppm) had little effect on lactobacilli, whereas 250 ppm of copper sulfate significantly decreased growth of E. coli. Beta glucan has been shown in vitro to modulate phagocytosis and TLR expression and ascorbic acid affects a number of immune functions. We fed different forms of beta glucan and ascorbic to young calves prior to challenge with Salmonella Dublin. Different types of beta glucan tended to affect parameters of immune function differently. Feeding a combination of beta glucan and ascorbic did not affect TLR 2 and TLR 4 expression. However, fewer calves had temperatures above 40C for the beta glucan-ascorbic acid treatments than did the control calves and feed conversion at 42 days after challenge was improved over the control. In a current study we are examining the effect of lairage on behavior, immune function and intestinal microbial populations. Sampling is completed and samples are being analyzed.

IMPACT: 2004/10/01 TO 2005/09/30
Stress alters functioning of the mucosal immune system, the epithelium and the intestinal microbiota and their interactions. We have examined the effect of diet additions on functioning of these systems in the presence of bacterial challenge and transportation stress. The data suggests that dietary treatments may be beneficial to animal health and performance, however, there is much that is not understood.

PROGRESS: 2003/10/01 TO 2004/09/29
We have conducted trials with pigs and poultry addressing dietary and stressor effects on animal performance, intestinal function and intestinal microbial community structure. Pigs fed a US based diet with growth promotant antibiotics performed better than pigs fed a European vegetable-based diet. Pigs fed an EU based diet with added lactitol had performance values between the two. Differences in microbial community structure were observed. Performance of broilers raised at 30C and fed a prebiotic was between the negative control and the positive (growth promotant antibiotic fed) control. Samples are being processed for microbial community structure and intestinal function.

IMPACT: 2003/10/01 TO 2004/09/29
We have shown that diet and stressors influence animal performance. We are addressing the underlying mechanisms by which this occurs, with the long-range goal of identifying dietary and management practices that can improve animal health and performance by increasing the synergistic protective actions of the intestinal microbiota, epithelial tissues and mucosal immune system.

PROGRESS: 2002/10/01 TO 2003/09/30
Stress disrupts the intestinal microbiota, epithelial tissue function and immune system, resulting in increased incidence of clinical and subclinical infection. We have determined the effect of feed withdrawal, acute and long term heat stress on broiler performance, epithelial tissue function, immune status, microbial community structure and ability of salmonella to attach to ileal tissues. There was a 1.5 log increase in attachment of salmonella to ileal tissues after a 24 h fast, with less of an increase with short term heat stress and little effect of long term heat stress. Birds fed an arabinoxylan diet as a prebiotic alternative to antibiotics tended to gain more than control and antibiotic fed birds under heat stress, but not when raised at normal temperatures. Salmonella attachment was higher for antibiotic fed birds at control temperatures, but did not differ for other treatments at either temperature.

IMPACT: 2002/10/01 TO 2003/09/30
Stress reduces the intestinal barrier against infection. We have shown that fasting and acute heat stress increase attachment of salmonella to the intestine, whereas long term heat stress does not increase attachment. Intervention stratagies may be developed to reduce the impact of stress on intestinal infections.

PROGRESS: 2001/10/01 TO 2002/09/30
Both animal and human health are affected by the actions and interactions of the intestinal microbiota, intestinal epithelial lining and immune system. Stress decreases the efficacy of these systems in preventing or reducing infections. We are developing an interdiciplinary collaboration to determine the effects of stressors on the intestinal immune, epithelial and microbial systems.

IMPACT: 2001/10/01 TO 2002/09/30
We expect that stressors will reduce the ability of the animal defense system to resist infection. With this understanding, dietary and management approaches for enhancing resistance to infection may be developed. This research should impact animal health, performance, well-being and human food safety.

PROGRESS: 2000/10/01 TO 2001/09/30
Several studies were conducted to identify alternatives to sub-therapeutic antibiotic use on pig farms. Weanling pigs were fed control diets or diets containing sub-therapeutic antibiotics or potential alternatives to sub-therapeutic antibiotics. Animal performance was measured throughout the nursery period and rectal microbial populations were obtained prior to, during and at the end of the nursery period. Several probiotic organisms and prebiotic compounds have been tested. In this system, the pigs on the sub-therapeutic antibiotic treatment had numerically improved performance parameters and several of the probiotic and probiotic-prebiotic combinations had performance parameters between the control and sub-therapeutic antibiotic treatments. Model systems need to be developed where significant differences can be detected. The large intestine was removed from six pigs and the ileum was attached to the rectum to simulate human colonectomy patients. Pigs were fed different diets to determine the effect of feeds on intestinal function. Physical parameters (DM, pH, VFA), in vitro gas production and microbial community structure (using denaturing gradient gel electrophoresis) were determined. Diet influenced a number of these parameters. This technique is currently being used in The Netherlands in human patients.

IMPACT: 2000/10/01 TO 2001/09/30
Although not as effective as sub-therapeutic antibiotics, several probiotics and prebiotics have shown promise. Additional research to determine conditions under which they are effective is needed. Pigs are useful as models for a number of human diseases.

PROGRESS: 1999/10/01 TO 2000/09/30
Several studies were conducted to identify alternatives to sub-therapeutic antibiotic use on pig farms. Weanling pigs were fed control diets or diets containing sub-therapeutic antibiotics or potential alternatives to sub-therapeutic antibiotics. Animal performance was measured throughout the nursery period and rectal microbial populations were obtained prior to, during and at the end of the nursery period. Several probiotic organisms and prebiotic compounds have been tested. In this system, the pigs on the sub-therapeutic antibiotic treatment had numerically improved performance parameters and several of the probiotic and probiotic-prebiotic combinations had performance parameters between the control and sub-therapeutic antibiotic treatments. Model systems need to be developed where significant differences can be detected.

IMPACT: 1999/10/01 TO 2000/09/30
Although not as effective as sub-therapeutic antibiotics, several probiotics and prebiotics have shown promise. Additional research to determine conditions under which they are effective is needed.

PROGRESS: 1998/10/01 TO 1999/09/30
Oligosaccharides have been shown to improve human and animal health and to enhance animal performance. As with subtherapeutic antibiotic usage, greater response would be expected under conditions where animals are producing well below their genetic potential due to health or environmental conditions. Proposed mechanism of action of oligosaccharides is thought to be by stimulation of beneficial bacterial populations. These bacterial populations then inhibit pathogens by: competitive exclusion of pathogenic bacteria from the intestinal lining, competition for limiting nutrients, production of toxic compounds and stimulation of the immune system. We have examined the efficacy of several oligosaccharides on animal performance and on intestinal microbial populations. Although not significant, nursery pigs fed Virginiamycin or FOS gained more weight than did control pigs in a clean nursery and feed efficiency was improved in pigs fed the FOS diet that were housed in a dirty nursery. E. coli counts were significantly reduced and Bifidobacterial counts were significantly increased in FOS fed pigs housed in the clean nursery and similar trends occurred in the dirty nursery. Feeding arabinogalactan to broilers had little effect on performance, but decreased E. coli counts. Continuous culture studies indicated that competition for limiting carbohydrates is not a selective mechanism against E. coli. We are currently examining non-culture methods to follow changes in bacterial populations in response to feeding prebiotic compounds such as oligosaccharides.

IMPACT: 1998/10/01 TO 1999/09/30
We have shown that oligosaccharides tend to impact animal performance to a similar extent as subtherapeutic antibiotics under conditions where performance is below optimal. We are examining mechanisms by which oligosaccharides may work to better define how and when the may have efficacy. Feeding oligosaccharides reduces ammonia concentrations in the intestinal tract and we are looking at the effect on concentrations of other odor compounds.

PROGRESS: 1997/10/01 TO 1998/09/30
Oligosaccharides have been shown to improve human and animal health and to enhance animal performance. Sucrose thermal oligosaccharide caramel (STOC) is a thermally produced oligosaccharide. Broilers fed diets containing either STOC or a growth promotant had significantly higher average daily gains and improved feed conversion. Cecal bifidobacterial populations were generally increased by feeding STOC. Animal performance was improved to a similar extent by doubling NRC recommended levels of trace minerals and vitamins, or by feeding STOC. This data is consistent with other data on improved absorption of major minerals and suggests that by feeding oligosaccharides one may be able to maintain animal performance and reduce mineral excretion by the animal. A salmonella challenge model is being developed for finishing pigs to examine the effect of oligosaccharides and other intervention measures on shedding of salmonella prior to shipping and during lairage. Several management studies have indicated that SEW raised finishing pigs may be more susceptible to colonization than their conventionally raised littermates. This may indicate that additional preventative methods need to be used with SEW raised pigs. We are in the initial stages of determining the effect of oligosaccharides and other dietary compounds on fecal and pit odors.

PROGRESS: 1996/10/01 TO 1997/09/30
Methanogenesis from H2:CO2 in the rumen results in increased energy for certain ruminal microorganisms, yet the methane eructated represents a loss of energy to the animal. Acetogenic bacteria predominate over methanogens in some animals, thus it was of interest to determine if there is potential for acetogens to replace the function of methanogenic bacteria in the rumen, with the endproduct being utilizable by the animal. Ruminal acetogens were isolated from rumen contents of dairy cattle fed a 55:45 concentrate:roughage diet. Acetogens were isolated in H2 limited continuous cultures either at pH 6.7 or pH 6.0. Acetogenic bacteria were characterized as to their H2 threshold, growth on H2 or a variety of sugars and acetate production. Based upon this information, most of the research was conducted using isolates A10, 3H and G3.2a. Known cultures of Acetobacterium woodii, Acetitomaculum ruminis, and Sporomusa termitida were included in some experiments for comparison. It has been shown that some acetogens utilize H2 and sugars mixotrophically and it has been proposed that this capability may allow acetogens to outcompete methanogens in some species. Using 13C labeling A10 was shown to utilize glucose first and produced formate during growth on glucose. Hydrogen utilization did not start until the glucose was consumed. From growth data it appears that the other ruminal acetogen isolates are not mixotrophs. Isolate A10 was shown to have a PTS transport system for sugars, which may be involved in regulation of H2 utilization. Concentrations of glucose, maltose and cellobiose necessary for initiation of growth of 3H and A10 were 111, 14, 14 and 56,28, 139 uM, respectively. Soluble glucose concentrations found in the rumen of cattle fed high and low concentrate diets was around 14 uM. Cellobiose concentration was around 122 uM and no soluble maltose was detected. The role of soluble sugars in regulation of H2 utilization is not clear at present. A series of batch culture studies were conducted to determine the efficacy of acetogens for utilizing H2 produced by ruminal contents when methanogenesis was inhibited by BES or lumazine. Addition of acetogens at 107-9 cells/ml culture reduced H2 concentrations significantly. Hydrogen concentrations were always lower than the control + BES and were lower than the ruminal fluid control cultures at 12h and 24h, but H2 concentrations were always lower in the ruminal fluid control after 24h of incubation. Continuous cultures, containing BES and inoculated with ruminal contents, were run at D=0.42 and 0.84. H2 concentration was not affected by addition of isolate A10 or G3.2a to give 108 cells/ml at each turnover. Although acetogens can effectively utilize H2 in batch culture when methanogenesis is inhibited, the isolates tested were not able to utilize H2 in continuous culture. Further work will be needed to determine conditions where acetogens can effectively replace methanogenesis in the rumen.

PROGRESS: 1995/10 TO 1996/09
Methanogenesis from H2:CO2 in the rumen results in increased energy for certain ruminal microorganisms, yet the methane eructated represents a loss of energy to the animal. Acetogenic bacteria have potential to replace the function of methanogenic bacteria in the rumen, with the endproduct being utilizable by the animal. Previously we have shown that a ruminal acetogenic isolate A10 is incapable of simultaneously utilizing carbohydrates and H2 for growth and that soluble carbohydrate concentrations in the rumen are very low. Efficacy of bromo ethane sulfonic acid and lumazine in inhibiting methanogenesis and potential toxicity to acetogens has been determined. When added to rumen contents lumazine only slightly inhibited methanogenesis. In the presence of added acetogens, methane production was significantly reduced in the presence of lumazine. Methanogenesis may not be inhibited by Lumazine, however when there is competition for hydrogen, methanogenesis may be inhibited enough for acetogenesis to compete favorably for hydrogen. Research with a thermally produced fructooligosaccharide (sucrose thermal oligosaccharide caramel, STOC) has shown an interaction between level of vitamin/trace minerals on weight gain and levels of Bifidobacteria in Peking ducks. Weight gains and Bifidobacterial numbers were higher for STOC fed birds irrespective of vitamin or mineral supplementation level. Highest gains were with the highest level of vitamin supplementation.

PROGRESS: 1994/10 TO 1995/09
Methanogenesis from H2:CO2 in the rumen results in increased energy for certain ruminal microorganisms, yet the methane eructated represents a loss of energy to the animal. Acetogenic bacteria have potential to replace the function of methanogenic bacteria in the rumen, with the end product being utilizable by the animal. Previously we have shown that a ruminal acetogenic isolate A10 is incapable of simultaneously utilizing carbohydrates and H2 for growth. Current studies show that the predominant soluble carbohydrates in ruminal fluid from animals fed production diets are glucose, cellobiose and an unidentified pentose at 13, 135, and 139 uM, respectively. Maltose was not found and the unidentified pentose was not free arabinose or xylose. Minimum concentrations of glucose, cellobiose and maltose required for growth were below the concentrations of these substrates in ruminal fluid, suggesting that these compounds may not regulate H2 utilization in the rumen. In vitro studies where methanogenesis has been inhibited have shown that acetogens can replace the function of methanogens. Sucrose thermal oligosaccharide caramel (STOC) has been shown to selectively enrich for bifidobacteria in the intestinal tract of poultry and increase growth performance. The effect of STOC has been more dramatic when animals were fed a vitamin/mineral premix at NRC recommended levels than at twice NRC levels. Current research is designed to determine if this response is related to vitamins or to minerals.

PROGRESS: 1993/10 TO 1994/09
The ruminal isolate A10 rapidly utilized glucose and produced formate and hydrogen during growth on glucose. Isolate A10 utilized glucose, formate and then hydrogen. Use of NaH(superscript 13) CO(subscript 3) confirmed that A10 grew diauxically on glucose and hydrogen, that A10 utilized the Wood pathway and thus A10 is a true acetogen. However, when either isolate A10 or another isolate, H3HH, were added to batch cultures where methanogenesis was inhibited, both isolates rapidly utilized hydrogen. Isolate H3HH reduced hydrogen concentrations to similar levels as cultures containing uninhibited methanogens. Thus, under appropriate conditions acetogens may be able to replace the function of methanogens in the rumen. Oligosaccharides have been shown to selectively enrich for bifidobacteria in a number of species. We are able to produce a mixture of oligosaccharides from sucrose by thermolysis (Sucrose Thermal Oligosaccharide Caramel, STOC) and have conducted a number of poultry studies and are currently conducting swine studies. Poultry studies have generally shown 10 to 30% increases in weight gain. There have been 5 to 15 fold increases in bifidobacterial numbers and occasionally increases in numbers of lactobacilli. Similar results were obtained in ducks. In the first swine study with individually fed weanling pigs, six pigs per treatment, pigs fed a diet containing 2% STOC had gains and feed efficiencies numerically greater than control animals and similar to antibiotic fed pigs.

PROGRESS: 1992/10 TO 1993/09
Work is continuing on characterization of several new acetogenic ruminal isolates. These isolates have been shown to be different from known bacterial species and we are in the process of naming the novel isolates. Currently we are assessing conditions under which these acetogens can replace the role of methanogens in ruminal fermentations. Manipulation of the lower intestinal tract of nonruminant species to increase numbers of bifidobacteria has been shown to improve animal health and efficiency of production. We have shown that a sucrose thermal oligosaccharide (STO, Techno-Taffy) product consistently increases bifidobacterial numbers in cecal contents of broiler chickens and has increased four week weight gain by 32%. STO is a series of low molecular weight complex carbohydrates produced by controlled thermolysis of sucrose. We have shown that bifidobacterial species can grow on STO and that many potentially pathogenic bacterial species (E. coli, Clostridium perfringes, etc.) do not grow on STO. It is suggested that STO acts as a selective nutrient for bifidobacteria and thus allows for selective enrichment of particular bacterial species in the intestinal tract.

PROGRESS: 1991/10 TO 1992/09
Several ruminal acetogens have been isolated during the preliminary stages of this project using a hydrogen limited continuous culture system. These isolates are gram positive rods and coccii and grow on hydrogen and carbon dioxide as well as on a variety of carbohydrates. These isolates have hydrogen threshold values lower than other ruminal acetogens and are only twice as high as those for methanogens. Thus, these acetogenic isolates may be more able to compete with methanogens than other previously isolated ruminal acetogens. Two of nine isolates contain plasmids and isolate H3HH had five plasmids. The smallest plasmid from H3HH has been ligated into pBR322 and inserted into both the host acetogenic isolate and into E. coli by electroporation. A temperate bacteriophage has been isolated from H3HH by induction by mitomycin C. Either plasmids or bacteriophage may be useful in developing vectors for genetic manipulation. Several isolates have both glucokinase and PTS activity. Preliminary data suggests that isolate H3HH grows mixotropically on glucose and hydrogen. We are developing a fundamental understanding of how acetogens utilize hydrogen and other energy sources. This fundamental knowledge may help lead to reduction of methanogenesis from ruminants.

PROGRESS: 1989/10 TO 1990/09
Research has just been completed and data is being analyzed on a project to determine the effect of ensiling dairy cattle waste with wheat straw. Another project has just been initiated to isolate and characterize chitin degrading microorganisms.

PROGRESS: 1988/10 TO 1989/09
Use of 2-ethoxyethanol in the neutral detergent fiber determination of feed cellwall content was re-evaluated because justification for its use has diminished and it poses a potential health hazard. Timing of addition of alpha-amylase, during or after boiling, was also evaluated. A total of 17 grasses, six legumes and five concentrate or processed feeds were analyzed. Delection of 2-ethoxyethanol resulted in slightly lower NDF values. Addition of alpha-amylase 30 min after onset of boiling resulted in slightly lower NDF values than with no alpha-amylase addition in solutions containing 2-ethoxyethanol, but not in solutions without 2-ethoxyethanol. Observed treatment differences were small and probably of no practical significance. No differences in NDF values were observed due to timing of alpha-amylase addition. Thus, there is no justification for continued use of 2-ethoxyethanol, and alpha-amylase can be added to the neutral detergent solution after boiling instead of half-way through. Research is being conducted on methane emissions from ruminants and from ruminant waste material. No results are available at present.

PROGRESS: 1987/10 TO 1988/09
Microbial interactions are thought to play an important role in fiber digestion.Effects of bacterial interactions, pH and low cellulase production on rate and extent of NDF digestion by Ruminococcus albus were determined using defined cultures. Digestion of NDF by R. albus was increased (P interactions was still present. Bacterial interactions and tolerance to low pH affected NDF digestion by R. albus and may affect its ability to compete with other cellulolytic species in the rumen.

PROGRESS: 1986/10 TO 1987/09
Examination of fiber degradation by pure or defined mixed cultures of rumen bacteria requires that the fiber material be sterilized prior to use. Autoclaving removed a significant amount of soluble material from forages, caused a significant increase in lag time before digestion was initiated and decreased extent of digestion. Lag, rate and extent of digestion were not significantly affected by ethylene oxide sterilization. Sterilization of particulate material by ethylene oxide does not alter kinetics of fiber digestion and is the sterilization method of choice when using pure or defined cultures of rumen bacteria. The effect of interspecies H(2) interactions on microbial fermentation patterns has been well documented; however, all theoretical calculations on the thermodynamics of the reactions have assumed standard temperature, pressure and a pH of 7. We have examined the effect of pH on interspecies H(2) interaction between Selenomonas ruminantium and Methanobrevibacter ruminantium. Hydrogen production by Selenomonas rumination was increased as pH decreased. Hydrogen concentration was decreased in the presence of Methanobrevibacter ruminantium, but total H(2) production (H(2) + CH(4)) was increased. Total H(2) production was increased by decreasing pH. Propionate production increased as pH decreased and was lower in the presenceof Methanobrevibacter ruminantium.

PROGRESS: 1985/10 TO 1986/09
Examination of fiber degradation by pure or defined mixed cultures of rumen bacteria requires that the fiber material be sterilized prior to use. Autoclaving removed a significant amount of soluble material from forages, caused a significant increase in lag time before digestion was initiated and decreased extent of digestion. Lag, rate and extent of digestion were not significantly affected by ethylene oxide sterilization. Sterilization of particulate material by ethylene oxide does not alter kinetics of fiber digestion and is the sterilization method of choice when using pure or defined cultures of rumen bacteria. The effect of interspecies H(2) interactions on microbial fermentation patterns has been well documented, however, all theoretical calculations on the thermodynamics of the reactions have assumed standard temperature, pressure and a pH of 7. We have examined the effect of pH on interspecies H(2) interactions between Selenomonas ruminantium and Methanobrevibacter ruminantium. Hydrogen production by Selenomonas ruminantium was increased as pH decreased. Hydrogen concentration was decreased in the presence of Methanobrevibacter ruminantium, but total H(2) production (H(2) + CH(4)) was increased. Total H(2) production was increased by decreasing pH. Propionate production increased as pH decreased and was lower in the presence of Methanobrevibacter ruminantium.

PROGRESS: 1985/01 TO 1985/12
Enzymatic hydrolysis of cellulosic materials is increased by pretreatment with NaOH and a further increase in cellulose hydrolysis is obtained when the material is pretreated with FeTNa (iron tartrate in NaOH). The potential for using the FeTNa pretreatment with ruminant feedstuffs was investigated in this study. The in vitro neutral detergent fiber (NDF) digestabilities of untreated corn stover, NaOH-treated corn stover and FeTNa-treated corn stover were: 25.3%, 58.2% and 47.2%, respectively. In repeated batch fed cultures there was a significant decrease in NDF digestion of FeTNa-treated corn stover with time. Further studies indicated that the iron complexed with NH(3), thus making it unavailable to the microorganisms for growth. Surface area has been proposed as a major factor in determining the extent of cellulose hydrolysis. A solute exclusion technique was used to examine the effects of fermentation and NaOH or FeTNa pretreatments on micropore size distribution. The data showed that: 1) removal of hemicellulose and lignin by pretreatments increased dry matter disappearance of the remaining material upon fermentation; 2) relative to the size of bacterial cellulases (40 to 160 angstrom), the pretreatments and fermentation had little effect on increasing accessibility of the surface area internal to the cellulose particles, and 3) the micropore changes caused by pretreatments do not explain the enhanced digestion of corn stover by rumen microorganisms.

PROGRESS: 1984/01 TO 1984/12
Pyrophosphates have been used as a phosphorous source and as a buffer in ruminant rations. Recently it has been shown that certain anaerobic bacteria can use pyrophosphates as an energy source. Experiments using rumen contents indicate that the major rumen bacteria can not use pyrophosphates as energy source. Water activity, or osmotic pressure, affects the growth and metabolic activities of microorganisms. In vitro studies were conducted to determine the effect of osmotic pressure between 200 to 450 mOsm/kg, which brackets the normal rumen osmotic pressure, on rumen microorganisms. As osmotic pressure was increased by additions of polyethylene glycol (3,350 MW) growth of rumen microorganisms was inhibited, but the concentrations and ratios of fermentation products were not affected. Zeolites are cation exchange compounds and may have potential as ruminal buffers. In vitro studies showed that addition of .4, .8, or 1.6 g zeolite/100 ml unbuffered medium resulted in an immediate increase in pH over the control medium. The final pH, after incubation with rumen microorganisms was also increased by each addition of zeolite. In vivo trials were conducted where either 0, 100, 200, or 300 g zeolite was added to the rumen prior to feeding and rumen pH and VFA concentrations were determined for 6 hr after feeding. Rumen pH tended to be higher in animals given zeolite. Molar % VFA was not changed by zeolite additions, but total VFA was increased by the addition of 300 g zeolite.

PROGRESS: 1983/01 TO 1983/12
Low pH has a significant negative effect on the degradation of plant material inthe rumen, thus buffers have been added to ruminant diets to maintain a higher pH. The effect of a cation exchange material (zeolite A) on pH was studied invitro using media containing no buffer or low levels (15 mM) of the major buffer in the rumen (NaHCO3) with the following levels of zeolite A (g/100 ml): 0.0, 0.4, 0,8, or 1.6. Tubes were inoculated with strained rumen contents and pH values were recorded with time. Initial pH values for the nonbuffered media were 5.5, 5.7, 5.9 and 6.1 and final (24 hr) pH values were 4.2, 4.6, 5.2 and 6.0 for zeolite treatments of 0.0, 0.4, 0.8, and 1.6 g/100 ml), respectively. Initial pH of the weakly buffered medium was higher than the non buffered medium with inital pH values of 6.2, 6.3, 6.4 and 6.5 for the respective zeolite additions. Final pH values for the weakly buffered media were 4.8, 5.2, 6.0 and 6.3, respectively. Invivo trials were conducted using 4 mature fistulated steers. Aniamls were fed a corn silage/grain diet once daily and were dosed with either 0, 100, 200 or 300 g of zeolite A. Samples were taken before and at 1/2, 1, 1 1/2, 2, 3, 4 and 6 hr after the zeolite addition. The 2 hr samples have been analyzed and rumen pH was increased from 6.1 to 6.2 by all of the zeolite additions. Volatile fatty acid concentrations were not altered by zeolite additions but there was a trend for lactate and NH3 concentrations to decrease as zeolite levels increased.

PROGRESS: 1983/01 TO 1983/12
Low pH has a significant negative effect on the degradation of plant material inthe rumen, thus buffers have been added to ruminant diets to maintain a higher pH. The effect of a cation exchange material (zeolite A) on pH was studied invitro using media containing no buffer or low levels (15 mM) of the major buffer in the rumen (NaHCO3) with the following levels of zeolite A (g/100 ml): 0.0, 0.4, 0,8, or 1.6. Tubes were inoculated with strained rumen contents and pH values were recorded with time. Initial pH values for the nonbuffered media were 5.5, 5.7, 5.9 and 6.1 and final (24 hr) pH values were 4.2, 4.6, 5.2 and 6.0 for zeolite treatments of 0.0, 0.4, 0.8, and 1.6 g/100 ml), respectively. Initial pH of the weakly buffered medium was higher than the non buffered medium with inital pH values of 6.2, 6.3, 6.4 and 6.5 for the respective zeolite additions. Final pH values for the weakly buffered media were 4.8, 5.2, 6.0 and 6.3, respectively. Invivo trials were conducted using 4 mature fistulated steers. Aniamls were fed a corn silage/grain diet once daily and were dosed with either 0, 100, 200 or 300 g of zeolite A. Samples were taken before and at 1/2, 1, 1 1/2, 2, 3, 4 and 6 hr after the zeolite addition. The 2 hr samples have been analyzed and rumen pH was increased from 6.1 to 6.2 by all of the zeolite additions. Volatile fatty acid concentrations were not altered by zeolite additions but there was a trend for lactate and NH3 concentrations to decrease as zeolite levels increased.

Funding Source
Nat'l. Inst. of Food and Agriculture
Project source
View this project
Project number
IND010984
Accession number
89786
Categories
Salmonella
Parasites
Natural Toxins
Viruses and Prions
Bacterial Pathogens
Chemical Contaminants
Escherichia coli
Commodities
Meat, Poultry, Game