Dietary marker effects on fecal microbial ecology, fecal VFA, nutrient digestibility coefficients, and growth performance in finishing pigs.

Use of indigestible markers such as Cr2O3, Fe2O3, and TiO2 are commonly used in animal studies to evaluate digesta rate of passage and nutrient digestibility. Yet, the potential impact of indigestible markers on fecal microbial ecology and subsequent VFA generation is not known. Two experiments utilizing a total of 72 individually fed finishing pigs were conducted to describe the impact of dietary markers on fecal microbial ecology, fecal ammonia and VFA concentrations, nutrient digestibility, and pig performance. All pigs were fed a common diet with no marker or with 0.5% Cr2O3, Fe2O3, or TiO2. In Exp. 1, after 33 d of feeding, fresh fecal samples were collected for evaluation of microbial ecology, fecal ammonia and VFA concentrations, and nutrient digestibility, along with measures of animal performance. No differences were noted in total microbes or bacterial counts in pig feces obtained from pigs fed the different dietary markers while Archaea counts were decreased (P = 0.07) in feces obtained from pigs fed the diet containing Fe2O 3compared to pigs fed the control diet. Feeding Cr2O3, Fe2O3, or TiO2 increased fecal bacterial richness (P = 0.03, 0.01, and 0.10; respectively) when compared to pigs fed diets containing no marker, but no dietary marker effects were noted on fecal microbial evenness or the Shannon-Wiener index. Analysis of denaturing gradient gel electrophoresis gels did not reveal band pattern alterations due to inclusion of dietary markers in pig diets. There was no effect of dietary marker on fecal DM, ammonia, or VFA concentrations. Pigs fed diets containing Cr2O3 had greater Ca, Cu, Fe, and P (P ≤ 0.02), but lower Ti ( P= 0.08) digestibility compared to pigs fed the control diet. Pigs fed diets containing Fe2O3 had greater Ca (P = 0.08) but lower Ti (P = 0.01) digestibility compared to pigs fed the control diet. Pigs fed diets containing TiO2 had greater Fe and Zn (P ≤ 0.09), but lower Ti ( P= 0.01) digestibility compared to pigs fed the control diet. In Exp. 2, no effect of dietary marker on pig performance was noted. Overall, the data indicate that the inclusion of Cr2O3, Fe2O3, or TiO2 as digestibility markers have little to no impact on microbial ecology, fecal ammonia or VFA concentrations, nutrient digestibility, or pig growth performance indicating they are suitable for use in digestion studies.

Effects of feeding fiber-fermenting bacteria to pigs on nutrient digestion, fecal output, and plasma energy metabolites.

Inclusion of feedstuffs with higher plant cell wall (fiber) content in swine diets has increased in recent years due to greater availability and lower cost, especially coproduct feeds, such as corn distillers dried grains with soluble (DDGS). Limitations of feeding higher fiber diets include increased fecal output, which can exceed manure storage volumes, and decreased energy density, which can decrease growth performance; dietary treatments that ameliorate these limitations would benefit pork producers. Grower pigs (n = 48; 61.1 kg initial BW) were used to establish the effects of supplementation of fiber-fermenting bacteria in a 2 × 4 factorial, consisting of 2 diets (standard and high fiber) and 4 bacterial treatments (A, no bacteria; and B, C, and D bacterial supplements). Increased fiber came from inclusion of soybean hulls (10%) and corn DDGS (20%) in the diet. The 3 bacterial supplements (all Bacteroides strains) were isolated from fecal enrichment cultures and selected for their fiber-fermenting capacity. The high fiber diet increased fecal output, blood cholesterol, and triglyceride concentrations, and digestibility of NDF, ADF, and S; CP digestibility decreased (P ≤ 0.10). The improved fiber digestibility and altered energy status of pigs fed the high fiber diet was primarily due to fermentation of soybean hulls, resulting in increased short-chain fatty acid production and absorption, and decreased dietary starch content. Overall, pigs fed the bacterial treatments had only increased blood cholesterol concentrations (P = 0.10). When individual bacterial treatments were compared, pigs fed Bacteria B had decreased fecal output (P ≤ 0.10) and both blood glucose and cholesterol concentrations were increased (P ≤ 0.10) compared with the other 3 treatments, indicating an improved energy status. Pigs fed Bacteria B increased both CP and ADF (P ≤ 0.10), and tended (P = 0.16) to have increased NDF digestibilities compared with pigs fed no bacteria (Treatment A), whereas pigs fed the other 2 bacterial treatments did not differ from pigs fed Bacteria B for nutrient digestibility. Both had similar fecal outputs to pigs fed no bacteria. This is the first report of reduction in fecal output and increased fiber digestibility with pigs fed live bacteria. Successful application of this bacterial treatment could result in improved pig performance and decreased manure volumes, both of which would improve profitability of producers.

Determination and prediction of digestible and metabolizable energy from chemical analysis of corn coproducts fed to finishing pigs.

Twenty corn coproducts from various wet- and dry-grind ethanol plants were fed to finishing pigs to determine DE and ME and to generate equations predicting DE and ME based on chemical analysis. A basal diet comprised corn (97.05%), limestone, dicalcium phosphate, salt, vitamins, and trace minerals. Twenty test diets were formulated by mixing the basal diet with 30% of a coproduct, except for dried corn solubles and corn oil, which were included at 20 and 10%, respectively. There were 8 groups of 24 finishing gilts (n = 192; BW = 112.7 ± 7.9 kg). Within each group, gilts were randomly assigned to 1 of 5 test diets or the basal diet for a total of 4 replications per diet per group. Two groups of gilts were used for each set of coproducts, resulting in 8 replications per coproduct and 32 replications of the basal diet. The experiment was conducted as a completely randomized design. Gilts were placed in metabolism crates and offered 3 kg daily of their assigned test diet for 13 d, with total collection of feces and urine during the last 4 d. Ingredients were analyzed for DM, GE, CP, ether extract, crude fiber, NDF, ADF, total dietary fiber (TDF), ash, AA, and minerals, and in vitro OM digestibility was calculated for each ingredient. The GE was determined in the diets, feces, and urine to calculate DE and ME for each ingredient. The DE and ME of the basal diet were used as covariates among groups of pigs. The DE of the coproducts ranged from 2,517 kcal/kg of DM (corn gluten feed) to 8,988 kcal/kg of DM (corn oil), and ME ranged from 2,334 kcal/kg of DM (corn gluten feed) to 8,755 kcal/kg of DM (corn oil). By excluding corn oil and corn starch from the stepwise regression analysis, a series of DE and ME prediction equations were generated. The best fit equations were as follows: DE, kcal/kg of DM = -7,471 + (1.94 × GE) - (50.91 × ether extract) + (15.20 × total starch) + (18.04 × OM digestibility), with R(2) = 0.90, SE = 227, and P < 0.01; ME, kcal/kg of DM = (0.90 × GE) - (29.95 × TDF), with R(2) = 0.72, SE = 323, and P < 0.01. Additional equations for DE and ME included NDF in the instance that TDF data were not available. These results indicate that DE and ME varied substantially among corn coproducts and that various nutritional components can be used to accurately predict DE and ME in corn coproducts for finishing pigs.

Effect of dietary inorganic sulfur level on growth performance, fecal composition, and measures of inflammation and sulfate-reducing bacteria in the intestine of growing pigs.

Two experiments investigated the impact of dietary inorganic S on growth performance, intestinal inflammation, fecal composition, and the presence of sulfate-reducing bacteria (SRB). In Exp. 1, individually housed pigs (n = 42; 13.8 kg) were fed diets containing 2,300 or 2,100 mg/kg of S for 24 d. Decreasing dietary S had no effect on ADG, ADFI, or G:F. In Exp. 2, pigs (n = 64; 13.3 kg) were fed diets containing 0, 0.625, 1.25, 2.5, or 5.0% CaSO(4), thereby increasing dietary S from 2,900 to 12,100 mg/kg. Two additional diets were fed to confirm the lack of an impact due to feeding low dietary S on pig performance and to determine if the increased Ca and P content in the diets containing CaSO(4) had an impact on growth performance. Pigs were fed for 35 d. Ileal tissue, ileal mucosa, and colon tissue were harvested from pigs fed the 0 and 5% CaSO(4) diets (low-S and high-S, respectively) to determine the impact of dietary S on inflammation-related mRNA, activity of mucosal alkaline phosphatase and sucrase, and pathways of inflammatory activation. Real-time PCR was used to quantify SRB in ileal and colon digesta samples and feces. Fecal pH, sulfide, and ammonia concentrations were also determined. There was no impact on growth performance in pigs fed the diet reduced in dietary S or by the increase of dietary Ca and P. Increasing dietary S from 2,900 to 12,100 mg/kg had a linear (P < 0.01) effect on ADG and a cubic effect (P < 0.05) on ADFI and G:F. Real-time reverse-transcription PCR analysis revealed that pigs fed high-S increased (P < 0.05) the relative abundance of intracellular adhesion molecule-1, tumor necrosis factor-α, and suppressor of cytokine signaling-3 mRNA, and tended (P = 0.09) to increase the relative abundance of IL-6 mRNA in ileal tissue. Likewise, pigs fed high-S had reduced (P < 0.05) abundance of nuclear factor of κ light polypeptide gene enhancer in B-cells inhibitor-α and increased (P < 0.05) phospho-p44/p42 mitogen-activated protein kinase in ileal tissue, but there was no effect of dietary S on mucosal alkaline phosphatase or sucrase activity. Pigs fed the high-S diet had decreased (P < 0.05) total bacteria in ileal digesta, but increased (P < 0.05) prevalence of SRB in colon contents. Fecal sulfide was increased (P < 0.05) and fecal pH was deceased (P < 0.05) in pigs fed high-S. The data indicate that growing pigs can tolerate relatively high amounts of dietary inorganic S, but high dietary S content alters inflammatory mediators and intestinal bacteria.

Changes in bacterial communities from swine feces during continuous culture with starch.

Bacteria from swine feces were grown in continuous culture with starch as the sole carbohydrate in order to monitor changes during fermentation and to determine how similar fermenter communities were to each other. DNA extracted from fermenter samples was analyzed by denaturing gradient gel electrophoresis (DGGE). A significant decrease in diversity was observed, the Shannon-Weaver index dropped from 1.92 to 1.13 after 14 days of fermentation. Likewise, similarity of fermenter communities to those in the fecal inoculum also decreased over time. Both diversity and similarity to the inoculum decreased most rapidly in the first few days of fermentation, reflecting a period of adaptation. Sequencing of DGGE bands indicated that the same species were present in replicate fermenters. Most of these bacteria were placed in the Clostridium coccoides/Eubacterium rectale group (likely saccharolytic butyrate producers), a dominant bacterial group in the intestinal tract of pigs. DGGE proved useful to monitor swine fecal communities in vitro and indicated the selection and maintenance of native swine intestinal bacteria during continuous culture.

Fate and transport of zoonotic, bacterial, viral, and parasitic pathogens during swine manure treatment, storage, and land application.

Members of the public are always somewhat aware of foodborne and other zoonotic pathogens; however, recent illnesses traced to produce and the emergence of pandemic H1N1 influenza virus have increased the scrutiny on all areas of food production. The Council for Agricultural Science and Technology has recently published a comprehensive review of the fate and transport of zoonotic pathogens that can be associated with swine manure. The majority of microbes in swine manure are not zoonotic, but several bacterial, viral, and parasitic pathogens have been detected. Awareness of the potential zoonotic pathogens in swine manure and how treatment, storage, and handling affect their survival and their potential to persist in the environment is critical to ensure that producers and consumers are not at risk. This review discusses the primary zoonotic pathogens associated with swine manure, including bacteria, viruses, and parasites, as well as their fate and transport. Because the ecology of microbes in swine waste is still poorly described, several recommendations for future research are made to better understand and reduce human health risks. These recommendations include examination of environmental and ecological conditions that contribute to off-farm transport and development of quantitative risk assessments.

Evaluation of elevated dietary corn fiber from corn germ meal in growing female pigs.

To evaluate the effects of dietary hemicellulose from corn on growth and metabolic measures, female pigs (n = 48; initial BW 30.8 kg) were fed diets containing 0 to 38.6% solvent-extracted corn germ meal for 28 d. Increasing the hemicellulose level had no impact on ADG or ADFI, but resulted in a quadratic response (P < 0.03) on G:F. To investigate physiological changes that occur with increased dietary hemicellulose, blood, colon contents, and tissue samples from the liver and intestine were obtained from a subset (n = 16; 8 pigs/treatment) of pigs fed the least and greatest hemicellulose levels. The abundance of phospho-adenosine monophosphate-activated protein kinase (AMPK) and the mitochondrial respiratory protein, cytochrome C oxidase II (COXII) were determined in liver, jejunum, ileum, and colon by Western blotting. The mRNA expression levels of AMPKalpha1, AMPKalpha2, PPAR coactivator 1alpha (PGC1-alpha), PPARgamma2, and sirtuin 1 (Sirt1) were determined in liver and intestinal tissues. When compared with pigs fed the control diet, pigs fed the high hemicellulose diet had increased (P < 0.02) plasma triglycerides, but there was no difference in plasma cholesterol, glucose, or insulin. Absolute and relative liver weights were decreased (P < 0.03) in pigs consuming the high hemicellulose diet. The high-fiber diet led to a tendency (P < 0.12) for decreased liver triglyceride content. In pigs fed the high hemicellulose diet, ileal mucosal alkaline phosphatase activity was increased (P < 0.08) and sucrase activity tended (P < 0.12) to be increased. The high hemicellulose diet had no effect on phospho-AMPK, AMPK mRNA, or colonic VFA, but in pigs consuming the high fiber diet there was a greater (P < 0.05) abundance of COXII in colon tissue. The expression of PGC1-alpha, PPARgamma, or Sirt1 mRNA was not altered by dietary fiber in liver, jejunum, or ileum tissue. In colon tissue from pigs fed the high fiber diet there was an increase (P < 0.09) in Sirt1 mRNA and a trend (P < 0.12) toward increased of PGC1-alpha mRNA. These data suggest that alterations in metabolism involved in adaptation to a diet high in hemicellulose are associated with increased colonic Sirt1 mRNA and COXII expression, indicating an increased propensity for oxidative metabolism by the intestine.

Comparative sulfur analysis using thermal combustion or inductively coupled plasma methodology and mineral composition of common livestock feedstuffs.

The objective of this study was to compare the use of thermal combustion (CNS) and inductively coupled plasma (ICP) to measure the total S content in plant-, animal-, and mineral-based feedstuffs, and to provide concentrations of other macro- and micro-minerals contained in these feedstuffs. Forty-five feedstuffs (464 total samples) were obtained from suppliers as well as swine feed and pet food manufacturers throughout the United States. Mineral data from IPC analysis were summarized on a DM basis using sample mean and SD, whereas the comparison of total S content between CNS and ICP was examined by bivariate plot and correspondence correlation. Analyses of a wide range of feedstuffs by CNS and ICP for total S were comparable for all but a few feedstuffs. For potassium iodide and tribasic copper chloride, ICP estimated total S to be lower than when analyzed by CNS (bias = 2.51 +/- 0.15 SE, P < 0.01). In contrast, for defluorinated phosphate and limestone, ICP estimated total S to be greater than when analyzed by CNS (bias = -1.46 +/- 0.51 SE, P < 0.01). All other samples had similar estimates of total S, whether analyzed by CNS or ICP. As expected, S composition varied greatly among feedstuffs. For total S, plant-based feedstuffs generally had lower total S compared with animal-based feedstuffs, whereas minerals supplied in sulfate form had the greatest concentration of total S. In addition to total S, mineral composition data are provided for all feedstuffs as obtained by ICP analysis. Within specific feedstuffs, mineral composition was quite variable, potentially due to low concentrations in the feed-stuff causing high mathematical variation or due to the source of feedstock obtained. In general, analyzed values of P were similar to previous tabular values. These data provide feed formulators a database from which modifications in dietary minerals can be accomplished and from which mineral requirements can be met more precisely to reduce losses of minerals into the environment.

Effects of adding fibrous feedstuffs to the diet of young pigs on growth performance, intestinal cytokines, and circulating acute-phase proteins.

The effects of feeding different types of fiber to weanling pigs on growth performance, intestinal and liver cytokine expression, circulating acute-phase proteins, and IGF-I were evaluated. Intestinal tissue abundance of DNA, protein, and phosphorylated S6 kinase were also determined. Pigs (n = 120; initially 5.2 kg and 24 d of age) were randomly assigned to diets containing 1 of 4 fiber sources: 1) control diets containing no added fiber source, 2) diets containing 7.5% distillers dried grains with solubles (DDGS), 3) diets containing 7.5% soybean hulls, or 4) diets containing 7.5% citrus pulp. The experimental diets were fed for 4 wk in 2 phases (phase 1, wk 1 and 2; phase 2, wk 3 and 4). Intestinal tissue samples, liver samples, and blood samples were collected from a subset (n = 24; 6 pigs/treatment) of the pigs on d 7, and blood samples were collected from another subset (n = 24; 6 pigs/ treatment) of pigs on d 28 of the experiment. Dietary treatment had no effect on ADG, ADFI, or G:F throughout the experiment. Likewise, pig BW variability (CV), plasma IGF-I, or the plasma concentration of the acute-phase proteins, alpha(1)-acid glycoprotein, C-reactive protein, and haptoglobin, were not affected by dietary treatment. Real-time RT-PCR analysis revealed that on d 7, pigs fed DDGS had a greater (P < 0.05) relative abundance of the mRNA encoding IL-6, IL-1beta, and IL-10 in ileum tissue than pigs fed all other diets. Diets containing DDGS had no effect on the relative abundance of tumor necrosis factor alpha or interferon-gamma mRNA in ileum tissue on d 7. The d-7 mRNA expression of cytokines was not altered in jejunum, colon, or liver tissue by dietary treatment. Intestinal tissue protein content or jejunum and ileum DNA concentrations were not affected by diet. Western blot analysis found no effect of dietary treatment on the activation of S6 kinase in jejunum, ileum, or colon tissue on d 7. These results indicate that feeding 7.5% of a fiber source as DDGS, soybean hulls, or citrus pulp does not affect growth performance or circulating markers of inflammation in weanling pigs and that feeding DDGS increases the expression of both proinflammatory and antiinflammatory cytokines in intestinal tissue.

Manure composition of swine as affected by dietary protein and cellulose concentrations.

An experiment was conducted to investigate the effects of reducing dietary CP and increasing dietary cellulose concentrations on manure DM, C, N, S, VFA, indole, and phenol concentrations. Twenty-two pigs (105 kg initial BW) were fed diets containing either 14.5 or 12.0% CP, in combination with either 2.5 or 8.7% cellulose. Pigs were fed twice daily over the 56-d study, with feed intake averaging 2.74 kg/d. Feces and urine were collected after each feeding and added to the manure storage containers. Manure storage containers were designed to provide a similar unit area per animal as found in industry (7,393 cm2). Before sampling on d 56, the manure was gently stirred to obtain a representative sample for subsequent analyses. An interaction of dietary CP and cellulose was observed for manure acetic acid concentration, in that decreasing CP lowered acetic acid in pigs fed standard levels of cellulose but increased acetic acid in pigs fed greater levels of cellulose (P = 0.03). No other interactions were noted. Decreasing dietary CP reduced manure pH (P = 0.01), NH4 (P = 0.01), isovaleric acid (P = 0.06), phenol (P = 0.05), and 4-ethyl phenol (P = 0.02) concentrations. Increasing dietary cellulose decreased pH (P = 0.01) and NH4 (P = 0.07) concentration but increased manure C (P = 0.03), propionic acid (P = 0.01), butyric acid (P = 0.03), and cresol (P = 0.09) concentrations in the manure. Increasing dietary cellulose also increased manure DM (P = 0.11), N (P = 0.11), and C (P = 0.02) contents as a percentage of nutrient intake. Neither cellulose nor CP level of the diet affected manure S composition or output as a percentage of S intake. Headspace N2O concentration was increased by decreasing dietary CP (P = 0.03) or by increasing dietary cellulose (P = 0.05). Neither dietary CP nor cellulose affected headspace concentration of CH4. This study demonstrates that diets differing in CP and cellulose content can significantly impact manure composition and concentrations of VFA, phenol, and indole, and headspace concentrations of N(2)O, which may thereby affect the environmental impact of livestock production on soil, air, and water.

Application of group specific amplified rDNA restriction analysis to characterize swine fecal and manure storage pit samples.

Group specific amplified ribosomal-DNA restriction analysis was evaluated as a method to rapidly assess microbial community structure in swine fecal and manure storage pit samples. PCR primer sequences were evaluated for their specificity to ribosomal DNA from selected bacterial groups by optimizing annealing temperatures and determining specificity using a set of primer target and non-target organisms. A number of primer sets were identified targeting the following groups: Bacteroides-Prevotella, clostridial clusters I and II, clostridial clusters IX and XI, clostridial clusters XIVa and XIVb, Lactobacillus, Desulfovibrionaceae and Streptococcus-Lactococcus, as well as an universal primer set to represent total populations. Each bacterial group was digested with at least three restriction enzymes. We applied the group specific amplified ribosomal-DNA restriction analysis to swine fecal and manure storage pit samples obtained on two separate occasions. Fecal and manure storage pit samples obtained on the same day were more similar to each other than to any other samples. Results were consistent with 16S ribosomal DNA sequencing data from bacterial isolates and clones obtained from swine feces and manure storage pit. The group specific amplified ribosomal-DNA restriction analysis technique was able to rapid detect gross bacterial community differences among swine fecal and manure storage pit samples and determine groups of interest for more detailed examination.

Evaluation of the specificity of Salmonella PCR primers using various intestinal bacterial species.

AIMS: Nine sets of PCR primers targeting Salmonella were evaluated for their specificity with pure cultures of intestinal-associated bacteria prior to their application to Salmonella detection in faecal samples. METHODS AND RESULTS: Gene targets of PCR primers included: 16S rDNA, a Salmonella pathogenicity island I virulence gene, Salmonella enterotoxin gene (stn), invA gene, Fur-regulated gene, histidine transport operon, junction between SipB and SipC virulence genes, Salmonella-specific repetitive DNA fragment, and multiplex targeting invA gene and spvC gene of the virulence plasmid. Fifty-two Salmonella strains were used to determine sensitivity; five strains from related genera and 45 intestinal bacteria were used to evaluate specificity. All primers amplified DNA from Salmonella strains, although two primer sets failed to amplify Salmonella DNA from either Salmonella bongori (hilA) or subgroups VI or VII (16S rDNA). There was no detected amplification of DNA from related bacterial genera with any of nine PCR assays. Six of the PCR assays amplified DNA for some intestinal bacteria. CONCLUSIONS: Only three primer pairs were determined to be suitable for application of PCR amplification of Salmonella in faecal samples - 16S rDNA, stn and histidine transport operon. We are currently evaluating their sensitivity of detection of Salmonella in faecal samples. SIGNIFICANCE AND IMPACT OF THE STUDY: This study demonstrated the importance of internal lab validation of PCR primers prior to application to the type of samples of interest. Information from this evaluation can be applied in other labs to facilitate choosing Salmonella PCR primers.

Preslaughter holding environment in pork plants is highly contaminated with Salmonella enterica.

The objective of this study was to determine whether abattoir pens can provide a Salmonella enterica infection source during the 2 to 4 h of preharvest holding. Previous work has suggested that pigs may be getting infected, but little has been reported on the environmental contamination of abattoir holding pens. For 24 groups of pigs studied ( approximately 150 animals/group) at two high-capacity abattoirs, six pooled fecal samples (n, 10 per pool) were collected from each transport trailer immediately after pigs were unloaded. Holding pens were sampled (one drinking water sample and six pooled floor samples consisting of swabs, residual liquid, and feces) prior to entry of study pigs for the routine holding period ( approximately 2.5 h). After slaughter, cecal contents and ileocecal lymph nodes were collected, on the processing line, from 30 pigs in each studied group. All samples were cultured for the isolation and identification of S. enterica by primary enrichment in GN-Hajna and tetrathionate broths, secondary enrichment in Rappaport-Vassiliadis broth, and plating on brilliant green sulfa and xylose-lysine-tergitol-4 agars, followed by biochemical and serological identification. The study pens were highly contaminated with S. enterica; all holding pens sampled had at least one positive sample. Additionally, 33% (8 of 24) of drinking water samples were positive for S. enterica. All 24 groups of pigs had S. enterica-positive cecal contents and ileocecal lymph nodes, including those groups from transport trailers with no positive samples. From pigs, trailers, and pens, 586 isolates representing 36 different Salmonella serovars were isolated. Of the 353 isolates from pigs (109 from ileocecal lymph nodes plus 244 from cecal contents), 19% were identified as belonging to the same serovars as those isolated from the respective pens; 27% were identified as belonging to the same serovars as those isolated from the trailers. Sixteen percent of the unique serovars were isolated from both pigs and pens, suggesting that pens served as the infection source. This study demonstrates highly contaminated abattoir holding pens and watering sources. It also demonstrates that holding pens can serve as an infection source. This study identifies the abattoir holding pens as a significant hazard and a potential control point for Salmonella contamination in the preharvest pork production chain.

Persistence and functional impact of a microbial inoculant on native microbial community structure, nutrient digestion and fermentation characteristics in a rumen model.

Small sub-unit (SSU) rRNA-targeted oligonucleotide probes were used to monitor the persistence of a genetically engineered bacterium inoculated in model rumens. Eight dual flow continuous culture fermenters were operated with either standard artificial saliva buffer or buffer with chondroitin sulfate (0.5 g/l) added. After 168 h of operation, fermenters were inoculated with Bacteroides thetaiotaomicron BTX (BTX), at approximately 1% of total bacteria. B. thetaiotaomicron was quantified using a species-specific probe and shown to persist in fermenters 144 h after inoculation (relative abundance 0.48% and 1.42% of total SSU rRNA with standard and chondroitin sulfate buffers, respectively). No B. thetaiotaomicron SSU rRNA was detected in fermenter samples prior to inoculation with strain BTX. Relative abundances of Bacteria, Eucarya and Archaea were not affected by either inoculation or buffer type. Fiber digestion, in particular the hemicellulose fraction, increased after strain BTX addition. Chondroitin sulfate addition to the buffer increased bacterial nitrogen flow in fermenters, but did not alter fiber digestion. Neither inoculum nor buffer type altered total short chain fatty acid (VFA) concentrations but proportions of individual VFA differed. In model rumens, B. thetaiotaomicron BTX increased fiber digestion when added to mixed ruminal microbes, independent of chondroitin sulfate addition; but further study is needed to determine effects on other fiber-digesting bacteria.

Distribution and abundance of Gram-positive bacteria in the environment: development of a group-specific probe.

We developed a 16S rRNA-targeted oligonucleotide probe (S-P-GPos-1200-a-A-13) for the Gram-positive bacteria, confirmed its specificity by database searches and hybridization studies, and investigated the effects of humic acids on membrane hybridizations with this probe. S-P-GPos-1200-a-A-13 was used to estimate the abundance of Gram-positive populations in the bovine rumen and Lake Michigan sediments. This probe should be useful for studies of the environmental distribution of Gram-positive bacteria and the detection of uncultured, phylogenetically Gram-positive bacteria with variable or negative Gram staining reactions, and could serve for Gram staining in some diagnostic settings.

Comparison of microbial populations in model and natural rumens using 16S ribosomal RNA-targeted probes.

A model rumen system, dual-flow continuous culture fermenters, was evaluated by two comparative criteria in two experiments using ribosomal (r)RNA-targeted DNA probes to compare key microbial groups in samples. The initial experiment measured temporal changes in population structure during adaptation of ruminal microbial populations in fermenters over 240 h. The fermenter inoculum contained 34.9% Bacteria, 60.1% Eukarya and 6.8% Archaea measured as a fraction of total small subunit (SSU) rRNA quantified using a universal probe. The cellulolytic bacterial genus Fibrobacter comprised 9.5% of total SSU rRNA in the inoculum. After 240 h of fermenter operation, the average abundance was 80.9% Bacteria, 6.1% Eukarya, 5.1% Archaea and Fibrobacter genus accounted for 6.6% of the total SSU rRNA. Divergence between ruminal and fermenter population structure was evaluated in the second experiment and samples were classified as ruminal, inoculum or fermenter (96, 120, 144 and 168 h of fermenter operation). Fermenter samples had higher relative abundances of Bacteria (84.5%) and Archaea (2.1%) and lower relative abundances of Eukarya (1.8%) than ruminal samples (average 48.0% Bacteria, 1.3% Archaea and 61.5% Eukarya). The relative abundance of Fibrobacter was similar in all samples, averaging 2.5%. The ruminal and fermenter samples had similar proportions of F. succinogenes and F. succinogenes subgroup 3 (as a percentage of Fibrobacter SSU rRNA). Fibrobacter succinogenes subgroup 1 and F. intestinalis proportions of Fibrobacter were lower in fermenter samples (8.2% and 0.7% respectively) than in ruminal samples (28.4% and 2.2% respectively). Fermenters were able to maintain a core prokaryotic community structure similar to the native microbial community in the rumen. Although protozoa populations were lost, maintenance of Fibrobacter and archaeal populations indicated that the model system supported a functional community structure similar to the rumen. This model rumen system may serve as a suitable tool for studying aspects of ruminal microbial ecology and may resolve some of the relationships between microbial community structure and function by providing control of experimental conditions.

Chemical drying agents for alfalfa hay: effect on nutrient digestibility and lactational performance.

Effects of chemical drying agents for hay on plasma profile, lactational performance, and nutrient digestibility by cows were studied. First-cutting alfalfa (late bud) was harvested as hay and treated when cut by applying untreated control, a commercial drying agent (7.64 kg/ha), or a mixture of active ingredients, potassium carbonate and sodium carbonate (7.47 kg/ha). Six multiparous Holstein cows (120 to 150 d postpartum) were fed diets (two cows per treatment) containing 55% hay and 45% concentrate (DM basis) in a balanced two period changeover design. Plasma profiles were similar for all cows, regardless of hay treatment. There were no differences in milk yield, milk fat, or milk protein percentages for control, drying agent, or mixture. Intake of DM was not different among treatments. Apparent digestibilities of DM, OM, CP, NDF, and ADF were not different among treatments. Treatment of alfalfa hay with chemical drying agents did not alter plasma profile, milk production, or nutrient digestibility in midlactation cows.