J. Anim. Sci. 2005. 83:1287-1302
© 2005 American Society of Animal Science
Feed physical form and formic acid addition to the feed affect the gastrointestinal ecology and growth performance of growing pigs
N. Canibe*,1,
O. Højberg*,
S. Højsgaard
and
B. B. Jensen*
* Department of Animal Health, Welfare, and Nutrition and
and
Department of Genetics and Biotechnology, Danish Institute of Agricultural Sciences, Research Center Foulum, Tjele, Denmark
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Abstract
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The effect of feeding a coarsely ground meal (COARSE) and a finely ground pelleted diet with 1.8% (as-fed basis) added formic acid (ACID) was compared with feeding a standard finely ground pelleted diet (STD) on the gastrointestinal ecology of growing pigs at different intervals after feeding. One hundred five castrated male growing-finishing pigs (initial BW 27 kg) were used. At a BW of 63 kg, 60 pigs were killed 0.5, 2.5, 4.5, 6.5, and 8.5 h after feeding, and samples from the gastrointestinal tract (GIT) were obtained. The remaining 45 pigs were kept on the experimental diets to a BW of 99 kg. Feeding the three diets resulted in a similar pattern of gastric pH with time, (i.e., highest pH values 0.5 h after feeding and decreasing values at the following sampling times, to reach a value of 2.12 at 8.5 h after feeding). The pH of the gastric digesta of pigs fed the ACID diet was below 4 at all sampling times, whereas the digesta from the other two dietary groups had values above pH 4 at the first sampling times. Feeding the ACID diet decreased the counts of total anaerobes in the proximal GIT (P 
0.007), and of lactic acid bacteria (P 0.001), enterobacteria (P 0.02), and yeasts (P 0.01) along the GIT compared with feeding the other two diets. Feeding the COARSE diet stimulated the growth of total anaerobes and lactic acid bacteria in the stomach and distal small intestine increased the microbial diversity mainly in the stomach (P = 0.001), compared with feeding the other two diets (P 0.09), and decreased the number of enterobacteria in the cecum compared with the STD diet (P = 0.03), with the same tendency in the mid-colon (P = 0.07). The concentration of lactic acid in the stomach was highest in the pigs fed the COARSE diet compared with the other two groups (P < 0.05). The concentration of formic acid was highest in the stomach and all segments of the small intestine of the pigs fed the ACID diet compared with those fed the STD and COARSE diets (P < 0.05). The results from this study suggest that feeding a coarsely ground diet and a finely ground diet with added formic acid affect the gastrointestinal ecology of pigs mainly by changing the environment in the proximal GIT. The presence of organic acids in the proximal GIT is a crucial factor contributing to the decrease in the number of enterobacteria along the GIT. The time after feeding at which samples are taken to measure characteristics describing the gastrointestinal ecology affects the results from the stomach and small intestine.
Key Words: Coarse Feed • Digestive Tract • Formic Acid • Growing Pigs • Microbial Flora
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Introduction
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Rearing healthy pigs with optimal growth performance and with the use of minimal medication is desirable from an economic and food safety point of view. Organic acids have been reported to decrease the number of potential pathogens, like enterobacteria, along the gastrointestinal tract (GIT; Kirchgessner et al., 1992
; Canibe et al., 2001) and to improve growth performance of growing pigs and piglets (Roth and Kirchgessner, 1998; Partanen et al., 2002). Feeding coarsely ground diets decreases the incidence of stomach lesions in pigs (Wondra et al., 1995; Nielsen and Ingvartsen, 2000) and the number of enterobacteria in the GIT of pigs (Mikkelsen et al., 2004). Growth performance can be compromised to some extent when pigs are fed coarsely ground diets (Wondra et al., 1995; Eisemann and Argenzio, 1999), which can make this alternative unattractive. An improvement of growth performance while maintaining health benefits would undoubtedly be preferable; however, rearing healthier pigs can have economic benefits for the producer besides the benefit obtained by decreasing the risk of transmission of disease to the consumer. In Denmark, for example, the presence of Salmonella in pigs is economically penalized, which can make it advantageous to the producer to avoid Salmonella in the animals, even if it is associated with somewhat lower growth performance.
The present study was designed with the main aim of elucidating the effects of feeding a finely ground and pelleted diet with formic acid, and a coarsely ground diet compared with a standard, finely ground, pelleted diet, on characteristics describing the gastrointestinal ecology of pigs and the mechanisms behind these effects. Elucidating the effect of time after feeding on these same characteristics was another primary goal of the study. A secondary aim of the study was to measure the effect of the experimental diets on growth performance through the grower-finisher stage.
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Materials and Methods
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This experiment was carried out at the Danish Institute of Agricultural Sciences. The Danish Ethical Commission approved the experimental protocol, and the animals were handled and slaughtered in a humane manner in accordance with the guidelines established by the Commission.
Animals and Housing
One hundred five (35 per treatment) castrated male growing-finishing pigs (Danish Landrace x Yorkshire x Duroc) from 34 litters were used. Pigs began the experiment at an initial BW of 27 kg (SD = 4.9). The animals were allotted on the basis of initial BW and litter to each of three dietary treatments. There were 21 pens (3 x 1.45 m, of which 0.7 x 1.45 m was slatted) corresponding to seven replicates of five pigs. No physical contact was allowed between pigs housed in different pens.
At a BW of 49 kg (SD = 7.7), 60 pigs (20 per treatment; four complete replicates) were moved to individual pens and later used to investigate the effect of the experimental diets on ecology of the GIT. These animals were killed, and samples from the GIT were obtained. The remaining 45 pigs were used to measure growth performance during the entire growing-finishing period (to a final BW of 99 kg, SD = 2.7).
Diets and Feeding
A grower diet was formulated (Table 1) and three dietary treatments were designed: 1) finely ground and pelleted (STD); 2) finely ground, pelleted, with 1.8% (as-fed basis) added formic acid (ACID); and 3) coarsely ground, nonheated, and nonpelleted (COARSE). The STD and ACID diets were ground to pass a 2-mm screen using a hammer mill, and the COARSE diet was obtained by grinding to pass a 5-mm screen in the same mill.
The pigs were allowed ad libitum access to the diets during the whole period, except for the animals to be killed. To obtain digesta samples from the GIT of the killed pigs at a defined time, the animals were offered the feed three times daily, at 0700, 1500, and 2200, during the last 10 d before being killed. During this 10-d period, the quantity of feed offered was adjusted every day so that the animals ate their ration approximately within 1 h after feeding.
Experimental Procedure
Body weight and feed intake were recorded weekly. The 60 pigs used in the slaughter experiment were moved to pens (2 x 1.8 m, of which 1.8 x 0.8 m was a resting place) where they were housed individually for 10 d. No physical contact between pigs from different treatments was allowed. After a 10-d period (average BW = 63 kg, SD = 7.2), the animals were killed by captive bolt 0.5, 2.5, 4.5, 6.5, and 8.5 h after the morning meal had been offered. The GIT was removed immediately and divided into eight segments: stomach, three equal segments of the small intestine, cecum, and three equal segments of the colon, including the rectum. The pars oesophagea region of the stomach was examined visually for possible alterations. The total content of each segment was weighed, and pH was measured within five minutes. Immediately after sample collection, approximately 5.0 g of digesta from each segment was extracted with 10.0 mL of perchloric acid/EDTA (2 M cold perchloric acid containing 10 mM EDTA) and stored at –80°C for ATP analyses. Samples from all segments of the GIT were taken and stored at –20°C to be analyzed for DM (5 to 10 g) and short-chain fatty acids (SCFA) and lactic acid concentration (approximately 10 g). Furthermore, digesta (10 g) from all segments were obtained for determination of in vitro production of SCFA and lactic acid. Microbiological determinations were performed immediately on digesta from the stomach, the distal segment of the small intestine, the cecum, and the middle segment of the colon. The remaining amount of sample from the stomach, the distal segment of the small intestine, the cecum, and mid-colon was stored at –20°C for terminal restriction fragment length polymorphism (T-RFLP) analysis.
Analytical Methods
Dry matter content of digesta was determined by freeze-drying the samples. To express the results of chemical analyses of the diets on a DM basis, DM was determined by drying at 103°C for 20 h. Ash and N were determined according to AOAC (1990) methods, energy by a Leco AC 300 automated calorimeter system 789-500 (Leco Corp., St. Joseph, MI). Amino acid analyses were carried out according to Mason et al. (1980) and EC Commission (2000). Fat was extracted with diethyl ether after acid hydrolysis and analyzed as described by Stoldt (1952). Low-molecular-weight sugars were measured following a modification of the method of Bach Knudsen and Li (1991) as described by Bach Knudsen (1997), starch by the method of Åman and Hesselman (1984), and soluble and insoluble fiber as described by Asp (1983). The concentration of SCFA and lactic acid was assayed by the method of Jensen et al. (1995). Particle size of the diets was determined by wet sieving through a series of sieves with the following mesh sizes: 1,400, 1,000, 500, 250, and 125 µm for the STD diet, and 3,150, 2,000, 1,000, 500, and 250 µm for the COARSE diet in a Retsch AS200 control "g" sieve shaker (Retsch GmbH and Co., Haan, Germany).
Microbiological Determinations.
Feed samples (10 g, as-fed basis) were transferred into flasks containing 90 mL of peptone water containing 1% peptone (Merck 1.07224, Darmstadt, Germany) and 0.1% (vol/vol) Tween 80. The suspension was then transferred to a plastic bag and homogenized in a stomacher blender (Interscience, St. Nom, France) for 2 min. Digesta samples (9 ± 2.8 g) were transferred rapidly after collection under a flow of CO2 into flasks containing 90 mL of a prereduced salt medium (Holdeman et al., 1977). The suspension was then transferred to a CO2-flushed plastic bag and homogenized as the feed samples. Then, 10-fold dilutions were prepared with the feed and digesta samples in prereduced salt medium by the technique of Miller and Wolin (1974). Samples (0.1 mL) were plated on or inoculated to both selective and nonselective media. Total anaerobic bacteria were enumerated by culturing the samples in roll tubes containing ruminal fluid-glucose-cellobiose agar (Holdeman et al., 1977) and incubating anaerobically at 38°C for 7 d. Total aerobes were enumerated on plate-count agar (Merck 1.05463) following aerobic incubation at 25°C for 1 d. Lactic acid bacteria were enumerated on de Man, Rogosa and Sharp agar (Merck 1.10660.0500) following anaerobic incubation at 38°C (digesta samples) or 25°C (feed samples) for 2 d. Enterobacteria were enumerated on McConkey agar (Merck 1.05465) following aerobic incubation at 37°C for 1 d. Yeasts were enumerated on malt chloramphenicol agar (10 g/L of glucose [Merck 1.08337.1000]; 3 g/L of malt extract (Merck 1.05347); 3 g/L of yeast extract (Merck 1.3753); 5 g/L of peptone (Merck 1.07224); 50 mg/L of chloramphenicol [Sigma-Aldrich Chemie GmbH C-0378, Steinheim, Germany]; 15 g/L of agar [Merck 1.01614]) following aerobic incubation at 37°C (digesta samples) or 25°C (feed samples) for 2 d.
The rate of SCFA and lactic acid production in the GIT of pigs fed the three diets was determined by incubating 10 g (as-is basis) of gastrointestinal contents from all eight segments with 40 mL of 100 mM Na-phosphate buffer (pH 6.5). The incubations were carried out essentially as described by Lærke et al. (2000), with an incubation time of 2 h in serum bottles. Samples were taken at 0, 1, and 2 h of incubation. The concentration of ATP was determined by the luciferin-luciferase method as described by Jensen and Jørgensen (1994). Terminal restriction fragment length polymorphism analysis was performed for fingerprinting microbial communities following the procedure described by Leser et al. (2000). The peaks of the T-RFLP profiles were subjected to tentative identification by comparing with the lengths of terminal restriction fragments (T-RF) obtained by in silico HhaI digestion of 16S rRNA gene sequences of a bacteria culture collection comprising 132 different commensal bacterial strains isolated from the GIT of pigs. According to this, some of the peaks seemed to represent unique strains, but often several strains rendered identical T-RF. The culture collection does not cover all species present in the GIT; therefore, some sample T-RF did not have a matching counterpart in the culture collection T-RF.
Calculations and Statistical Methods
When feed or digesta samples had counts below detectable levels, the minimum detectable level was applied. When the minimum detectable level was applied to one treatment, statistical differences between treatments only indicate minimal differences, and when it was applied to two or all three treatments for a given microbial group, the P-value is only approximate. Before carrying out statistical analysis of microbial counts, logarithmic conversion of the data was performed. The rate of organic acid production was linear during the 2 h of incubation, so the rate of production was calculated using the slope of the lines between 0 and 2 h.
The model used to estimate the effect of diet and time after feeding on the various response variables describing the gastrointestinal ecology was a mixed model that included diet, time after feeding, segment, block, diet x time after feeding, diet x segment, and time after feeding x segment as fixed effects. To capture the positive correlation among pigs from the same litter and from pigs previously kept in the same pen, random litter and pen effects were included in the model. To capture the correlation between measurements in different segments of the GIT on each pig, the random errors were allowed to be correlated. The correlation structure was a heterogeneous compound structure (csh in the terminology of the SAS Mixed procedure; SAS Inst., Inc., Cary, NC), under which the variances in different segments are different, but where the correlation between different segments is constant. For some variables, SAS (v. 8.2) had numerical problems in fitting this model due to the heterogeneous variance structure. To overcome this difficulty, the variance in each segment was estimated first (assuming segments to be independent), and these estimated variances were then used as weights when fitting a model with homogeneous compound correlation structure. The resulting model was one with a heterogeneous compound correlation structure. Because the design was balanced, the SEM for treatments should be identical; however, due to the estimation procedure adopted to overcome the computational difficulties, the SEM were not in practice completely identical. Thus, the largest SEM is presented to protect for overinterpretation of the results.
The effect of diet on ADG and ADFI was estimated using the GLM procedure of SAS, with diet and block as fixed effects and with initial BW included as a covariate. The effect of diet on G:F was estimated using a GLM with diet and block as fixed effects.
When there was an overall effect of diet or time after feeding at an alpha of P = 0.05, differences between means were compared pairwise using an F-test.
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Results
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Diets
The chemical composition of the diets, partly shown in Table 2, was very similar among the three diets. In addition, the DM, ash, fat, AA, and carbohydrate contents were very similar in all three diets (data not shown). The counts of some selected microbial population revealed a higher density of enterobacteria, yeasts, and total aerobes in the COARSE diet compared with the STD and ACID diets (Table 2).
All diets contained similar proportions of particles with a size smaller than 250 µm as measured by wet sieving: 61% (SD = 5.2) in the STD and ACID diet and 60% (SD = 13.0) in the COARSE diet. Nonetheless, the median particle size of the material larger than 250 µm was 358 µm (SD = 6.4) for the STD and ACID diets, and 747 µm (SD = 211.5) for the COARSE diet.
Digesta from the Gastrointestinal Tract
The quantity of digesta collected from the various segments of the GIT was similar for all diets, and more importantly, high amounts of digesta were obtained from all segments (data not shown). The stomach of the animals from all treatments contained an average of 1,025 to 1,174 g (as-is basis) of digesta.
The percentage of DM was greatest in the digesta from the stomach of pigs fed the COARSE diet and least in the digesta of those fed the ACID diet (Table 3). Although the differences were much smaller, a similar pattern was observed in the proximal small intestine. In the mid-colon, DM percent was less in the COARSE treatment compared with the other two treatments. Percentage of DM in the stomach was highest in the COARSE-fed animals at all times after feeding (data not shown). Although fluidity of gastric contents was not measured, the visual examination of the material showed a clear lower fluidity of the contents of pigs fed the COARSE diet compared with those from the other two dietary groups.
The pH in the gastric digesta was not different between pigs fed the various diets when testing the diet effect, that is, when considering the mean of all sampling times (Table 4). Only a tendency (P = 0.11) for a lower value for the ACID diet compared with the COARSE diet was observed; however, a considerable sampling time effect on gastric pH was observed, as shown in Figure 1. Feeding the three diets resulted in a similar pattern of gastric pH with time (Figure 1a; i.e., highest pH values 0.5 h after feeding followed by decreasing values at the following sampling times, reaching a value of 2.12 at 8.5 h after feeding). An important observation was made when looking at each diet separately: the pH of the gastric digesta of pigs fed the ACID was below 4 at all sampling times, whereas the digesta from the other two dietary groups had values above pH 4 at the first sampling times (Figure 1b). The effect of time after feeding on pH was most evident in the stomach. However, an effect of time also was detected along the small intestine, with pH decreasing during the first hours after feeding, and increasing again at 6.5 and 8.5 h after feeding (Figure 1a). The pH values were relatively constant with time in the cecum and colon.
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Figure 1. The pH a) along the gastrointestinal tract (mean of diets; n = 12) and b) in the stomach (by diet; n = 4) of pigs fed the experimental diets at various times after feeding. STD = finely ground and pelleted; ACID = finely ground, pelleted, and 1.8% added formic acid; COARSE = coarsely ground, nonheated, and nonpelleted. Sto = stomach; SI1 = proximal small intestine; SI2 = mid small intestine; SI3 = distal small intestine; Ce = cecum; Co1 = proximal colon; Co2 = mid colon; Co3 = distal colon. Values are least squares means and standard errors. x,y,zValues within gastrointestinal segment or diet without a common superscript differ, P < 0.05.
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Feeding the ACID diet decreased the counts of total anaerobes in the proximal GIT (P 0.007), and of lactic acid bacteria (P 0.001), enterobacteria (P 0.02), and yeasts (P 0.01) along the GIT compared with feeding the other two diets (Table 5). Feeding the COARSE diet resulted in a strong stimulation of the growth of total anaerobes and lactic acid bacteria in the stomach compared with the STD and ACID diet (P < 0.05). The COARSE diet also resulted in a higher density of lactic acid bacteria in the distal small intestine compared with the other two diets (P 0.04) and a higher density of total anaerobes compared with the ACID diet (P < 0.05); the same tendency was observed compared with the STD diet (P = 0.09). The number of enterobacteria in the cecum of pigs fed the COARSE diet was lower than of those fed the STD diet (P = 0.03), and the same pattern was detected in the distal small intestine (P = 0.13) and mid-colon (P = 0.07).
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Table 5. Counts of selected microbial populations (cfu/g digesta, as-is basis) in digesta from the gastrointestinal tract of pigs fed the experimental dietsa
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There was no effect of time after feeding on total anaerobes, lactic acid bacteria, and yeasts along the GIT (Figure 2). The counts of enterobacteria were affected by time after feeding in the stomach and distal small intestine (P 0.02), whereas no effect of time was observed in the cecum and mid-colon. In the gastric digesta, enterobacteria were present at higher levels during the first 2.5 h after feeding (4.0 to 4.4 log cfu/g of digesta) compared with the period from 4.5 to 8.5 h after feeding (2.9 to 3.2 log cfu/g of digesta). In the distal small intestine, enterobacteria counts were lowest 0.5 h after feeding (P 0.02), and no differences amongst the remaining sampling times were observed.
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Figure 2. Counts of selected microbial populations (log cfu/g of digesta, as-is basis) in a) the stomach, b) distal small intestine, c) cecum, and d) mid-colon of pigs fed the experimental diets at various times after feeding. Values are least squares means and standard errors (n = 12). y,zValues within microbial group and gastrointestinal segment without a common superscript differ, P < 0.05.
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The concentration of lactic acid in the stomach was greater in pigs fed the COARSE diet compared with the other two dietary groups (P < 0.05; Table 6). In the proximal and mid-small intestine, the concentrations of lactic acid were greater for the COARSE group compared with the ACID group (P = 0.02), with the same tendency observed compared with the STD group in the proximal small intestine (P = 0.06). The lowest numerical concentration of lactic acid in the stomach and small intestine was observed in pigs fed the ACID diet. The effect of time after feeding on gastric lactic acid concentration for each diet (Figure 3a) showed a clearly different picture for the COARSE compared with the STD and ACID diets. The levels in the pigs fed the latter diets were low at all sampling times, whereas in the pigs fed the COARSE diet, lactic acid concentration reached maximal levels of approximately 49 mmol/kg of digesta at 4.5 and 6.5 h after feeding. The concentration of formic acid was greatest in the stomach and all segments of the small intestine of the pigs fed the ACID diet compared with those fed the STD and COARSE diets (P < 0.05; Table 6). The effect of time on the concentration of formic acid in the stomach is shown in Figure 3b. The STD and COARSE groups had very low levels at all sampling times (0.2 to 1.1 mmol/kg of digesta), whereas the ACID group had levels of 44 to 47 mmol/kg digesta during the first 2.5 h after feeding. The gastric concentration of total SCFA was highest in pigs fed the COARSE diet, mainly due to acetic acid and propionic acid (Table 6). The concentration in the distal small intestine was greatest for the animals fed the ACID (21.9 mmol/kg digesta) diet compared with the other two dietary groups (11.6 to 13.5 mmol/kg digesta) (P < 0.05), whereas in the colon, the levels were numerically greater for the ACID and COARSE groups compared with the STD group, the differences being significant only in the mid-colon (P 0.04). The effect of time on the concentration of total SCFA was most clearly observed in the stomach (data not shown). The concentration followed the same pattern as that of the counts of total anaerobes and lactic acid bacteria shown in Figure 2
, with increasing concentration during the first sampling times (from 6.4 mmol/kg digesta at 0.5 h to 13.9 mmol/kg digesta at 6.5 h) and decreasing concentrations at the last sampling time (8.0 mmol/kg digesta at 8.5 h). The concentrations of acetic acid in the distal small intestine, cecum, and colon were greatest when feeding the ACID diet compared with the other two diets (P 0.05). In the mid- (P = 0.17) and distal colon (P = 0.10), the differences between the ACID diet and the COARSE diet were not significant. The concentration of propionic acid was similar for all diets along the GIT, except in the stomach, where it was greatest for the COARSE group. In addition, the concentration of butyric acid was greatest in the stomach of pigs fed the COARSE diet, and least in the mid- and distal colon of those fed the STD diet. The levels of the branched-chain fatty acids, isobutyric and isovaleric acid, were similar for the three dietary groups.
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Table 6. Concentration of organic acids (mmol/kg of digesta, as-is basis) in digesta from the gastrointestinal tract of pigs fed the experimental dietsa
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Figure 3. Concentration (mmol/kg digesta, as-is basis) of a) lactic acid and b) formic acid in the stomach of pigs fed the experimental diets at various times after feeding. STD = finely ground and pelleted; ACID = finely ground, pelleted, and 1.8% added formic acid; COARSE = coarsely ground, nonheated, and nonpelleted. Values are least squares means and standard error (n = 4). y,zValues within diet without a common superscript differ, P < 0.05.
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The in vitro production of lactic acid in the stomach was greatest in the digesta obtained from pigs fed the COARSE diet (10.9 mmol·kg of digesta–1·h–1]), and lowest in that from pigs fed the ACID diet (1.2 mmol·kg of digesta–1·h–1; Table 7). In the small intestine, the values were least for the pigs fed the ACID diet compared with the other two dietary groups (P 0.03). The in vitro production of formic acid was very low and did not differ among diets along the GIT. The production of total SCFA of gastric digesta was greatest (P < 0.05) in animals fed the COARSE diet compared with animals in the other two groups. The almost exclusive contribution to production of total SCFA in the stomach was by acetic and propionic acid. No differences between diets were observed in the remaining GIT.
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Table 7. In vitro production of organic acids (mmol·kg of digesta–1·h–1, as-is basis) of digesta from the gastrointestinal tract of pigs fed the experimental dietsa
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The microbial activity along the GIT of the pigs, measured as ATP, is presented in Table 8. The highest gastric ATP concentration was measured in the pigs fed the COARSE diet (P < 0.05). In the cecum and proximal colon, the highest ATP concentration was detected in those animals fed the ACID diet (P 0.02), with the same tendency in mid-colon (P 0.10).
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Table 8. Adenosine 5'-triphosphate (µg/g of digesta, as-is basis) in digesta from the gastrointestinal tract of pigs fed the experimental dietsa
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The T-RFLP profiles were determined in samples from the stomach, distal small intestine, cecum, and mid-colon. The data showed, in accordance with the other characteristics measured, that the main effect of diet on microbial diversity was observed in the stomach (stomach and mid-colon in Table 9; distal small intestine and cecum, not shown). The COARSE diet resulted in the highest number of T-RF, that is, in the highest microbial diversity in the stomach of pigs (P = 0.001). The effect of diet on microbial diversity in the remaining segments of the GIT followed the same pattern as in the stomach, but it was less evident. The number of T-RF in digesta from the ACID group was least along the GIT.
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Table 9. Fragment size and frequency (percentage of pigs out of the total pigs per diet) of the most frequent terminal restriction fragments (T-RF) in digesta from the gastrointestinal tract of pigs fed the experimental dietsa
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The concentration of starch in the distal small intestine, cecum, and distal colon was measured in 15 samples per treatment. These data showed that the animals fed the COARSE diet had higher concentration of starch in all segments of the GIT measured (4.1 to 6.0%) compared with those fed the other diets (0.0 to 2.7%; P < 0.001; data not shown).
Gastric Epithelium
The region of the pars oesophagea of the pigs fed the STD and ACID diets showed alterations, which were similar for both dietary groups; however, all of the pigs fed the COARSE diet had white, smooth, and healthy-looking tissue.
Growth Performance
Average daily gain and ADFI did not differ among diets in the interval of 27 to 50 kg BW, whereas G:F tended to be greatest for pigs fed the STD diet (475 g/kg) and least for those fed the COARSE diet (438 g/kg; P = 0.08; Table 10). During the interval of 27 to 99 kg BW, the ADG for the animals fed the ACID diet (952 g) tended to be highest, whereas ADG was the lowest for those fed the COARSE diet (855 g; P = 0.11). During this interval, G:F was higher for the pigs fed the ACID (392 g/kg) diet than for those fed the COARSE diet (351 g/kg; P = 0.02).
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Discussion
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The lower percentage of DM and greater fluidity of gastric contents of pigs fed a finely ground and pelleted diet compared with those fed a coarsely ground meal diet, as measured in our study, have been observed by various authors (Maxwell et al., 1970; Regina et al., 1999; Nielsen and Ingvartsen, 2000). Maxwell et al. (1970) and Ange et al. (2000) measured a higher water to feed disappearance ratio, and Maxwell et al. (1972) reported a greater water intake in pigs fed a finely ground and pelleted diet compared with pigs fed a coarsely mash diet, which was suggested to be the reason for the differences in DM percent and fluidity of stomach contents between diets. The actual intake of water was not measured in our experiment, so differences in water consumption may account for some of the differences in percentage of DM noted between the COARSE, STD, and ACID diets. The lower DM percent of gastric contents of the ACID fed pigs compared with those fed the STD diet is difficult to explain. Whether the relatively high concentration of formic acid in the ACID diet stimulates water consumption has, to our knowledge, not been reported. However, Roth et al. (1992) did not observe any differences in gastric DM percent in piglets fed diets with 0 to 2.4% added formic acid. Differences in DM percent and fluidity of gastric contents are thought to be related to the higher incidence of lesions and keratinization of the nonglandular esophageal region of the stomach, the pars oesophagea, of pigs fed finely ground and pelleted diets (Maxwell et al., 1970, 1972; Regina et al., 1999), which was clearly observed in the present study. Pepsin and hydrochloric acid are not secreted in the proximal but in the more distal region of the stomach. According to Maxwell et al. (1970) and Regina et al. (1999), greater mixing of the gastric content of pigs fed a fine diet, due to greater fluidity, results in higher levels of pepsin and hydrogen ions in the esophageal region of the stomach of these animals compared with those fed a coarse meal. This would contribute to tissue damage. Studies by Lang et al. (1998) suggested that a combination of low pH and high concentration of bile acids, again due to an easier reflux of distal gastric and duodenal contents to the proximal region of the stomach, contribute to ulceration in the animals fed finely ground and pelleted diets. Furthermore, results by Argenzio and Eisemann (1996) suggested that microbial production of SCFA may be important in the pathogenesis of porcine gastric ulcers. The present results, and those by Regina et al. (1999) and Mikkelsen et al. (2004), in which a higher concentration of SCFA was noted in the stomach of pigs fed a coarse meal compared with those fed a finely ground diet, do not support the latter hypothesis.
Feeding coarsely ground diets and the addition of organic acids to the diets are two feeding strategies under investigation as ways of producing healthy pigs without the use of antibiotic growth promoters (Gabert and Sauer, 1994; Jensen, 1998; Hansen et al., 2001; Partanen et al., 2002). To our knowledge, characteristics of a healthy GIT have not been properly defined. But rearing pigs with low levels of enterobacteria (i.e., Salmonella and coliform bacteria) in the GIT is a common aim. Low pH, high concentration of organic acid, and high numbers of lactic acid bacteria contribute to obtaining low numbers of enterobacteria along the GIT of pigs (van Winsen et al., 2001; Canibe and Jensen, 2003). Feeding the ACID diet decreased the levels of all microorganisms measured, including enterobacteria. Moreover, the T-RFLP profiles suggested a lower microbial diversity in the stomach of these animals. The pH values in the stomach were not significantly different between diets when all times after feeding were pooled. Nonetheless, when evaluating pH at various times after feeding for each diet separately, an important observation was made: the gastric pH in animals fed the ACID diet did not exceed a value of 4 at any time, whereas the other two groups had values higher than 4 during the first hours after feeding. Additionally, a higher concentration of formic acid was measured in the stomach and along the small intestine of the pigs fed the ACID diet compared with the other two dietary groups. The present results agree with previous data obtained by Canibe et al. (2001) with potassium diformate fed-piglets, which showed a higher concentration of formic acid in the stomach and small intestine, and a reducing effect of potassium diformate on total anaerobes, lactic acid bacteria, yeasts, and the same tendency for enterobacteria. In vitro studies with gastric contents obtained from pigs showed that at pH 5, the population of coliform bacteria remained constant, whereas at pH 4, the growth of these bacteria was inhibited (Knarreborg et al., 2002). Furthermore, the addition of potassium diformate to the digesta increased the magnitude of the response. The impairment of bacterial metabolism by a combination of low pH and high concentration of organic acids has been well described (Russell and Diez-Gonzalez, 1998). The constant low pH in the stomach in combination with the high concentration of formic acid in the stomach and small intestine were the most probable contributors to the reduction of microbes along the GIT of pigs fed the diet added formic acid observed in the present experiment. Microorganisms, including enterobacteria, would be able to survive or proliferate in the stomach of the pigs from the other dietary groups during the first hours, where the pH was high and the level of organic acids was still low.
The effect of time on pH (and to a lower extent on counts of microorganisms) along the GIT was evident in the proximal segments, especially in the stomach. The observed effect of time on pH indicates that the results from the proximal GIT in studies carried out with the slaughter method, in which pigs are killed at a given time after feeding, will be affected by the time after feeding at which the animals are killed. The lack of a significant effect of addition of formic acid or its salts on gastric pH observed by several authors (Roth et al., 1992; Canibe et al., 2001) in animals slaughtered 3 to 4 h after feeding could be explained by the effect of time after feeding on pH observed in the present study. Results from the distal segments of the GIT will not or will only slightly be affected by time of killing.
The substantial stimulating effect of the COARSE diet on the growth of lactic acid bacteria, mainly in the stomach but also in the small intestine, coincided with a high concentration of lactic acid in these segments. Furthermore, the gastric concentration of lactic acid with time after feeding followed a similar pattern to that of the number of lactic acid bacteria (differences with time not significant for lactic acid bacteria). That is, increasing levels of both characteristics during the first hours after feeding and decreasing levels at the last sampling times. So, whereas the concentration of formic acid was greatest during the first hours after feeding because it was given in the diet, the high levels of lactic acid were observed after the lactic acid bacteria had proliferated and produced the lactic acid. This time gap with low levels of lactic acid during the first hours after feeding theoretically would be detrimental because, as seen here, during the first hours after feeding the gastric pH was highest and therefore, the need for high concentrations of organic acids to avoid the proliferation of pathogens would be greater. The reason for the proliferation of lactic acid bacteria in the stomach of pigs fed coarsely ground meal compared with those fed finely ground pellets observed in this and other studies (Jørgensen et al., 1999) has not been elucidated. The consistency of gastric material obtained from the two types of diets was very different (i.e., a much more firm material after feeding the COARSE diet), which resulted in a lack of separation between the solid and liquid phase of the digesta, which was opposite to what was observed after feeding the other diets (i.e., a more fluid content that separated into a solid and a liquid phase very rapidly after sampling). This firm material could result in a more beneficial environment for the lactic acid bacteria than for other microbial groups; however, this postulate needs further investigation. On the other hand, several studies have reported a longer retention time of gastric digesta after feeding a coarsely ground meal compared with feeding a finely ground pellet (Maxwell et al., 1970; Regina et al., 1999). The longer time available to the gastric microbial population, in which lactic acid bacteria predominate (Jensen, 1999), could result in a dominance of this bacterial group with respect to the other groups.
Besides the observed higher number of lactic acid bacteria and total anaerobes in the stomach of the COARSE fed-animals, the T-RFLP profile of the same material showed a higher microbial diversity. Most of the T-RF found with higher frequency in the stomach of the COARSE group could be tentatively identified as microorganisms of the lactic acid bacteria group, whereas a few were not of this group or could not be identified. The higher concentration of not only lactic acid but also of SCFA in the stomach of pigs fed the COARSE diet suggests that both lactic acid bacteria and bacteria from other groups were stimulated by feeding the COARSE diet. Moreover, most of the T-RF found in the colon of pigs fed the COARSE diet also were found in the stomach of the same pigs, which could indicate that these pigs practiced coprophagy. One of the studies found in the scarce literature on T-RFLP profiles of digesta from the GIT of pigs is that of Leser et al. (2000). Leser et al. (2000) found more T-RF in the colon of pigs fed various diets than we did in the present study. These authors found T-RF with length of 34 bp, whereas our shortest T-RF had 88 bp, and the longest T-RF found by Leser et al. (2000) had 612 bp, whereas we found T-RF with lengths up to 717 bp. Several of the most frequent T-RF found in our study also were detected in the work of Leser et al. (2000), whereas others were not. More work is needed to evaluate the usefulness of T-RFLP analysis in gut ecology studies; however, it seems to be a method that supplements the other methods already used to describe the ecology of the GIT.
The lower number of enterobacteria in the small intestine, cecum, and colon of the pigs fed the COARSE diet compared with those fed the STD diet probably is due to the high concentration of lactic acid in the stomach and small intestine of the animals fed the COARSE diet, which is a result of the proliferation of lactic acid bacteria. This hypothesis is supported by data from the literature (Ratcliffe et al., 1986; van Winsen et al., 2000; Jørgensen et al., 2001; Knarreborg et al., 2002), in which a bactericidal effect of lactic acid on enterobacteria (Salmonella and coliform bacteria) has been observed. The fact that the inhibiting effect was not immediate could be due to the gastric concentration of lactic acid with time discussed above. The highest concentrations of lactic acid were obtained after 4.5 to 6.5 h after feeding, which presumably gave enterobacteria the possibility to proliferate during the first hours after feeding. The latter is supported by the data on the effect of time on enterobacteria counts in the stomach. These data showed a higher number of enterobacteria during the first hours after feeding than during the following hours after feeding. Although the interaction of the effect of diet x time was not significant, the level of enterobacteria in the ACID group was more constant with time, that is, the numbers were only slightly higher during the first hours. This is in accordance with the high levels of formic acid already measured during the first hours after feeding, and it supports the hypothesis on the relation between the concentration of organic acids and the numbers of enterobacteria proposed above.
The somewhat higher concentrations of SCFA in the colon of pigs fed the ACID diet are supported by the higher microbial activity measured as ATP at this site in the same pigs. These results could be partially a consequence of the observations made on microbial counts (i.e., reducing effect of formic acid addition on total anaerobes in the proximal segments of the GIT and no effect in the colon). The previous result could suggest that more fermentable substrates arrived to the colon of the ACID fed animals because, due to a lower number of microorganisms in the more proximal segments, there was a decreased extent of fermentation in the stomach and small intestine. Similar observations have been reported with piglets fed potassium diformate and slaughtered 7 d after weaning, although at 29 d after weaning, the concentration of SCFA was similar in the control and potassium diformate group (Canibe et al., 2001). Furthermore, the T-RFLP profiles showed a somewhat lower diversity in the colon of pigs fed the ACID diet compared with the other two dietary groups, which again indicates that although the density of total anaerobes was not significantly affected by formic acid addition, qualitative changes occurred in the colon.
Thus, these data suggest that addition of formic acid decreased the growth of microorganisms in the proximal segments of the GIT, thereby decreasing somewhat the extent of fermentation, but allowing for high fermentation in the large intestine, which contributes with energy to the host. This idea is supported by the somewhat higher concentration of starch in the small intestine and cecum of the ACID group compared with that found in the STD diet-fed pigs. That is, microorganisms contribute to the fermentation of starch or improve the accessibility of endogenous enzymes to the starch, which is decreased when the microbial numbers are decreased. In the colon, where the number of total anaerobes was not decreased by the formic acid, no starch was detected.
The lower in vitro production of lactic acid in the stomach of the ACID-fed animals and the higher production of those fed the COARSE diet confirmed the data on lactic acid bacteria counts and lactic acid concentration in this segment of the GIT. The results emphasize the reducing effect of formic acid and the stimulating effect of a coarse diet on the lactic acid bacteria. Furthermore, these data clearly show, as expected, that the high levels of formic acid measured in the stomach of the ACID fed animals were a result of addition of the acid to the diet and not of production in the stomach.
The presence of high concentration of starch in the distal small intestine, cecum, and mid-colon of pigs fed the COARSE diet compared with the other dietary groups shows that feeding diets with greater particle size can impede access of endogenous and microbial enzymes to the nutrients inside these feed particles. The data on starch concentration agree with the growth performance results, which showed a lower G:F for the COARSE-fed animals compared with the others, and a tendency to lower ADG. The tendency for greater ADG and greater G:F measured in pigs fed the diet added formic acid when the whole growing period was considered is in line with previous studies, in which improved growth and/or G:F were noted when grower diets were supplemented with formic acid or formates (Øverland et al., 2000; Partanen et al., 2002). The trend for a lower G:F in the period from 26 to 50 kg of pigs fed the ACID diet was unexpected and is difficult to explain when the positive data from the literature on addition of formic acid or formates on growth performance is considered. Whether the low microbial density in the proximal segments of the GIT of these pigs had a negative influence on growth performance during this period, as indicated by the starch data, cannot be ruled out; however, the results are not in accordance with the literature.
Feeding a coarsely ground diet to pigs resulted in a healthier GIT by affecting mainly characteristics of the stomach. Addition of formic acid to the feed also improved gastrointestinal health by resulting in digesta less suitable for growth of potential pathogens. Coarsely ground meal stimulates proliferation of lactic acid bacteria and increases microbial diversity in the stomach, and addition of formic acid to the diet, inhibits the growth of lactic acid bacteria and other microorganisms. Both feeding strategies result in lower counts of enterobacteria along the GIT. Thus, pigs with decreased numbers of enterobacteria can be produced through different mechanisms. The results from this study suggest that the presence of organic acids in the proximal segments of the GIT of pigs is a crucial factor contributing to a reduction of the number of enterobacteria along the GIT. The time after feeding at which samples are taken to measure characteristics describing the ecology of the GIT has effects on results from the stomach and small intestine, but time after feeding had almost no effect on those from the large intestine. Therefore, it is important to consider an optimal and standarized time after feeding to obtain digesta samples.
1 Correspondence: P.O. Box 50, 8830 Tjele (phone: +45 89 99 11 48; fax: +45 89 99 13 78; e-mail: nuria.canibe{at}agrsci.dk).
Received for publication January 23, 2004.
Accepted for publication March 16, 2005.
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Literature Cited
|
|---|
Åman, P., and K. Hesselman. 1984. Analysis of starch and other main constituents of cereal grains. Swedish J. Agric. Res. 14:135–139.
Ange, K. D., J. H. Eisemann, R. A. Argenzio, G. W. Almond, and A. T. Blikslager. 2000. Effects of feed physical form and buffering solutes on water disappearance and proximal stomach pH in swine. J. Anim. Sci. 78:2344–2352.[Abstract/Free Full Text]
Argenzio, R. A., and J. Eisemann. 1996. Mechanisms of acid injury in porcine gastroesophageal mucosa. Am. J. Vet. Res. 57:564–573.[Medline]
Asp, N. G. 1983. Rapid enzymatic assay of insoluble and soluble dietary fiber. J. Agric. Food Chem. 31:476–482.[Medline]
AOAC. 1990. Official Methods of Analysis. Assoc. Off. Anal. Chem., Washington, DC.
Bach Knudsen, K. E. 1997. Carbohydrate and lignin contents of plant materials used in animal feeding. Anim. Feed Sci. Technol. 67:319–338.
Bach Knudsen, K. E., and B. Li. 1991. Determination of oligosaccharides in protein-rich feedstuffs by gas-liquid chromatography and high-performance-liquid chromatography. J. Agric. Food Chem. 39:689–694.
Canibe, N., and B. B. Jensen. 2003. Fermented and non-fermented liquid feed to growing pigs—Effect on aspects of gastrointestinal ecology and on growth performance. J. Anim. Sci. 81:2019–2031.[Abstract/Free Full Text]
Canibe, N., S. H. Steien, M. Øverland, and B. B. Jensen. 2001. Effect of K-diformate in starter diets on acidity, microbiota, and the amount of organic acids in the digestive tract of piglets, and on stomach alterations. J. Anim. Sci. 79:2123–2133.[Abstract/Free Full Text]
EC Commission. 2000. Directive 2000/45/EC: Establishing EC methods of analysis for the determination of vitamin A, vitamin E and tryptophan in feedingstuffs. Off. J. L 174:32–50.
Eisemann, J., and R. A. Argenzio. 1999. Effects of diet and housing density on growth and stomach morphology in pigs. J. Anim. Sci. 77:2709–2714.[Abstract/Free Full Text]
Gabert, V., and W. C. Sauer. 1994. The effects of supplementing diets for weanling pigs with organic acids. A review. J. Anim. Feed Sci. 3:73–87.
Hansen, C. F., K. E. Bach Knudsen, B. B. Jensen, and H .D. Kjærsgaard. 2001. Effect of meal feed and coarser grinding of pelleted feed on productivity, microbiology, and physico-chemical properties in the gastrointestinal tract of finishers. Pages 106–108 in Proc. 4th Int. Symp. Epidemiol. Control of Salmonella and Other Foodborne Pathogens in Pork, Leipzig, Germany.
Holdeman, L. V., E. P. Cato, and E. C. Moore. 1977. Anaerobe Laboratory Manual. Virginia Polytechnic Inst. State Univ., Blacksburg.
Jensen, B. B. 1998. The impact of feed additives on the microbiological ecology of the gut in young pigs. J. Anim. Feed Sci. 7:45–64.
Jensen, B. B. 1999. Impact of feed composition and feed processing on the gastrointestinal ecosystem in pigs. Pages 43–56 in Nutrition and Gastrointestinal Physiology—Today and Tomorrow. Presented at the Symposium Held on the Occasion of the Retirement of J. Huisman. Wageningen, The Netherlands.
Jensen, B. B., and R. M. Engberg. 2004. Antibiotic growth promoters. Pages 1030–1034 in Encyclopedia of Meat Sciences. W. K. Jensen, C. Devine, and M. Dikeman, ed. Academic Press, New York, NY.
Jensen, B. B., and M. T. Jensen. 1993. In vitro measurements of microbial production of skatole in the digestive tract of pigs. Pages 99–105 in Measurements and Prevention of Boar Taint in Entire Male Pigs. Les Colloques No. 60. M. Bonneau, ed. Inst. Natl. Rech. Agron., Roskilde, Denmark.
Jensen, B. B., and H. Jørgensen. 1994. Effect of dietary fiber on microbial activity and microbial gas production in various regions of the gastrointestinal tract of pigs. Appl. Environ. Microbiol. 60:1897–1904.[Abstract/Free Full Text]
Jensen, M. T., R. P. Cox, and B. B. Jensen. 1995. Microbial production of skatole in the hind gut of pigs given different diets and its relation to skatole deposition in backfat. Anim. Sci. 61:293–304.
Jørgensen, L., J. Dahl, B. B. Jensen, and H. D. Poulsen. 1999 Effect of expanding, pelleting and grinding on Salmonella, growth performance and gastrointestinal health in growing pigs and on phytase activity and vitamin stability in feed. Danish Bacon and Meat Council, No. 426, Copenhagen, Denmark. [In Danish]
Jørgensen, L., H. D. Kjærsgaard, H. Wachmann, B. B. Jensen, and K. E. Bach Knudsen. 2001. Effect of pelleting and use of lactic acid in feed on Salmonella prevalence and productivity in weaners. Pages 109–111 in Proc. 4th Int. Symp. Epidemiol. Control Salmonella Other Foodborne Pathogens in Pork, Leipzig, Germany.
Knarreborg, A., N. Miquel, T. Granli and B. B. Jensen. 2002. Establishment and application of an in vitro methodology to study the effects of organic acids on coliform and lactic acid bacteria in the proximal part of the gastrointestinal tract of piglets. Anim. Feed Sci. Technol. 99:131–140.
Kirchgessner, M., B. Gedek, S. Wiehler, A. Bott, U. Eidelsburger, and F. X. Roth. 1992. Influence of formic acid, calcium formate and sodium hydrogen carbonate on the microflora in different segments of the gastrointestinal tract. 10. Communication. Investigation about the nutritive efficacy of organic acids in the rearing of piglets. J. Anim. Physiol. Anim. Nutr. 68:73–81.
Lang, J., A. Blikslager, D. Regina, J. Eisemann, and R. Argenzio. 1998. Synergistic effect of hydrochloric acid and bile acids on the pars esophageal mucosa of the porcine stomach. Am. J. Vet. Res. 59:1170–1176.[Medline]
Leser, T. D., R. H. Lindecrona, T. K. Jensen, B. B. Jensen, and K. Møller. 2000. Changes in bacterial community structure in the colon of pigs fed different experimental diets and after infection with Brachyspira hyodysenteriae. Appl. Environ. Microbiol. 66:3290–3296.[Abstract/Free Full Text]
Lærke, H. N., B. B. Jensen, and S. Højsgaard. 2000. In vitro fermentation pattern of D-tagatose is affected by adaptation of the microbiota from the gastrointestinal tract of pigs. J. Nutr. 130:1772–1779.[Abstract/Free Full Text]
Maribo, H., B. B. Jensen, and M. S. Hedemann. 2000. Various doses of organic acids to piglets. Danish Bacon and Meat Council, No. 469, Copenhagen, Denmark. [In Danish]
Mason, V. C., S. Bech-Andersen, and M. Rudemo. 1980. Hydrolysate preparation for amino acid determination in feed constituents. J. Anim. Physiol. Anim. Nutr. 43:146–164.
Maxwell, C. V., E. M. Reimann, W. G. Hoekstra, T. Kowalczyk, N. J. Benevenga, and R. H. Grummer. 1970. Effect of dietary particle size on lesion development and on the contents of various regions of the swine stomach. J. Anim. Sci. 30:911–922.
Maxwell, C. V., E. M. Reimann, W. G. Hoekstra, T. Kowalczyk, N. J. Benevenga, and R. H. Grummer. 1972. Use of tritiated water to assess, in vivo, the effect of dietary particle size on the mixing of stomach contents of swine. J. Anim. Sci. 34:212–216.
Mikkelsen, L. L., P. J. Naughton, M. S. Hedemann, and B. B. Jensen. 2004. Effects of physical properties of feed on microbial ecology and survival of Salmonella enterica serovar Typhimurium in the pig gastrointestinal tract. J. Appl. Microbiol. 70:3485–3492.
Miller, T. L., and M. J. Wolin. 1974. A serum bottle modification of the Hungate technique for cultivating obligate anaerobes. Appl. Microbiol. 27:985–987.[Medline]
Nielsen, E. K., and K. L. Ingvartsen. 2000. Effect of cereal type, disintegration method and pelleting on stomach content, weight and ulcers and performance in growing pigs. Livest. Prod. Sci. 66:271–282.
Øverland, M., T. Granli, N. P. Kjos, O. Fjetland, S. H. Steien, and M. Stokstad. 2000. Effect of dietary formates on growth performance, carcass traits, sensory quality, intestinal microflora, and stomach alterations in growing-finishing pigs. J. Anim Sci. 78:1875–1884.[Abstract/Free Full Text]
Partanen, K., H. Siljander-Rasi, T. Alaviuhkola, K. Suomi, and M. Fossi. 2002. Performance of growing-finishing pigs fed medium-or high-fibre diets supplemented with avilamycin, formic acid or formic acid-sorbate blend. Livest. Prod. Sci. 73:139–152.
Ratcliffe, B., C. B. Cole, R. Fuller, and M. J. Newport. 1986. The effect of yoghurt and milk fermented with a porcine intestinal strain of Lactobacillus reuteri on the performance and gastrointestinal flora of pigs weaned at two days of age. Food Microbiol. 3:203–211.
Regina, D. C., J. H. Eisemann, J. A. Lang, and R. A. Argenzio. 1999. Changes in gastric contents in pigs fed a finely ground and pelleted or coarsely ground meal diet. J. Anim. Sci. 77:2721–2729.[Abstract/Free Full Text]
Roth, F. X., B. Eckel, M. Kirchgessner, and U. Eidelsburger. 1992. Influence of formic acid on pH-value, dry matter content, concentration of volatile fatty acids and lactic acid in the gastrointestinal tract. 3. Communication. Investigations about the nutritive efficacy of organic acids in the rearing of piglets. J. Anim. Physiol. Anim. Nutr. 67:148–156.
Roth, F. X., and M. Kirchgessner. 1998. Organic acids as feed additives for young pigs: Nutritional and gastrointestinal effects. J. Anim. Feed Sci. 7:25–33.
Russell, J. B., and F. Diez-Gonzalez. 1998. The effects of fermentation acids on bacterial growth. Adv. Microb. Physiol. 39:205–234.[Medline]
Stoldt, W. 1952. Vorschlag zur vereinheitlichung der fettbestimmung in lebensmitteln. [Suggestions to standardize the determination of fat in foodstuffs.] F. Seif. Anst. 54:206–207.
van Winsen, R. L., L. J. A. Lipman, S. Biesterveld, B. A. P. Urlings, J. M. A. Snijders, and F. van Knapen. 2000. Mechanism of Salmonella reduction in fermented pig feed. J. Sci. Food Agric. 81:342–346.
van Winsen, R. L., B. A. P. Urlings, L. J. A. Lipman, J. M. A. Snijders, D. Keuzenkamp, J. H. M. Verhejden, and F. van Knapen. 2001. Effect of fermented feed on the microbial population of the gastrointestinal tracts of pigs. Appl. Environ. Microbiol. 67:3071–3076.[Abstract/Free Full Text]
Wondra, K. J., J. D. Hancock, K. C. Behnke, and C. R. Stark. 1995. Effects of mill type and particle size uniformity on growth performance, nutrient digestibility, and stomach morphology in finishing pigs. J. Anim. Sci. 73:2564–2573.[Abstract]
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Abstract
Introduction
