Effects of Physical Properties of Feed on Microbial Ecology and Survival of Salmonella enterica Serovar Typhimurium in the Pig Gastrointestinal Tract

A two-by-two factorial experiment with pigs was conducted to study the effect of feed grinding (fine and coarse) and feed processing (pelleted and nonpelleted) on physicochemical properties, microbial populations, and survival of Salmonella enterica serovar Typhimurium DT12 in the gastrointestinal tracts of pigs. Results demonstrated a strong effect of diet on parameters measured in the stomachs of the pigs, whereas the effect was less in the other parts of the gastrointestinal tract. Pigs fed the coarse nonpelleted (C-NP) diet showed more solid gastric content with higher dry matter content than pigs fed the fine nonpelleted (F-NP), coarse pelleted (C-P), or fine pelleted (F-P) diet. Pigs fed the C-NP diet also showed significantly increased number of anaerobic bacteria (P < 0.05), increased concentrations of organic acids, and reduced pH in the stomach. In addition, pigs fed the C-NP diet showed increased in vitro death rate of S. enterica serovar Typhimurium DT12 in content from the stomach (P < 0.001). Pigs fed the C-NP diet had a significantly higher concentration of undissociated lactic acid in gastric content than pigs fed the other diets (P < 0.001). A strong correlation between the concentration of undissociated lactic acid and the death rate of S. enterica serovar Typhimurium DT12 was found. In the distal small intestine, cecum, and midcolon, significantly lower numbers of coliform bacteria were observed in pigs fed the coarse diets than in pigs fed the fine diets (P < 0.01). Pigs fed the C-NP diet showed the lowest number of coliform bacteria in these segments of the gastrointestinal tract. Pigs fed the coarse diets showed increased concentration of butyric acid in the cecum (P < 0.05) and colon (P < 0.10) compared with pigs fed the fine diets. It was concluded that feeding a coarsely ground meal feed to pigs changes the physicochemical and microbial properties of content in the stomach, which decreases the survival of Salmonella during passage through the stomach. In this way the stomach acts as a barrier preventing harmful bacteria from entering and proliferating in the lower part of the gastrointestinal tract.     

Roles of diet and the acid tolerance response in survival of common Salmonella serotypes in feces of finishing pigs

The persistence of Salmonella in the environment is an important factor influencing the transmission of infection in pig production. This study evaluated the effects of acid tolerance response (ATR), organic acid supplementation, and physical properties of feed on the survival of a five-strain Salmonella mixture in porcine feces held at 4 and 22°C for 88 days. Acid-adapted or non-acid-adapted nalidixic acid-resistant Salmonella strains were used to inoculate feces of pigs fed four different diets, which consisted of a nonpelleted, finely ground meal feed or a finely ground, pelleted feed that was left unsupplemented or was supplemented with K-diformate. Organic acid supplementation and physical properties of feed markedly influenced Salmonella survival, but the effects were highly dependent on storage temperature; survival was unaffected by ATR. The most pronounced effects were observed at 22°C, a temperature similar to that of finishing pig houses. The supplementation of meal diets with K-diformate significantly reduced the duration of survival (P < 0.1) and increased rates of decline (P < 0.0001) of salmonellae in feces compared to survival in feces of pigs fed unsupplemented meal. The pelleting of feed, compared to feeding meal, significantly reduced (P < 0.1) the duration of survival in feces held at 22°C. Only minor effects of feed form and acid supplementation on survivor numbers were observed at 4°C. Differences in the fecal survival of Salmonella could not be related to diet-induced changes in fecal physiochemical parameters. The predominant survival of S. enterica serovar Typhimurium DT193 and serotype 4,[5],12:i:- in porcine feces demonstrates the superior ability of these serotypes to survive in this environment. Fecal survival and transmission of Salmonella in pig herds may be reduced by dietary approaches, but effects are highly dependent on environmental temperature.     

Effects of particle size and hydro-thermal treatment of feed on performance and stomach health in fattening pigs

Effects of grinding and hydro-thermal treatment of feed on growth performance, slaughter traits, nutrient digestibility, stomach content and stomach health were examined by using 96 crossbred fattening pigs. Pigs were fed a grain-soybean meal-based diet processed by various technical treatments. Feeding groups differed in particle size after grinding (finely vs. coarsely ground feed) and hydro-thermal treatment (without hydro-thermal treatment, pelleting, expanding, expanding and pelleting). Fine grinding and hydro-thermal treatment showed significant improvements on the digestibility of crude nutrients and content of metabolisable energy. Hydro-thermal treatment influenced average daily gain (ADG) and average daily feed intake (DFI) significantly. Finely ground pelleted feed without expanding enhanced performances by increasing ADG and decreasing feed-to-gain ratio (FGR) of fattening pigs. Coarsely ground feed without hydro-thermal treatment resulted in the highest ADG and DFI, however also in the highest FGR. Expanded feed decreased DFI and ADG. Slaughter traits were not affected by treatments. Coarsely ground feed without hydro-thermal treatment had protective effects on the health of gastric pars nonglandularis, however, pelleting increased gastric lesions. Hydro-thermal treatment, especially expanding, resulted in clumping of stomach content which possibly induced satiety by slower ingesta passage rate and thus decreased feed intake. Pigs fed pelleted feed showed less pronounced development of clumps in stomach content compared with expanded feed.     

Effect of fermented feed on the microbial population of the gastrointestinal tracts of pigs.

An in vivo experiment was performed with pigs to study the inhibitory effect of fermented feed on the bacterial population of the gastrointestinal tract. Results demonstrated a significant positive correlation between pH and lactobacilli in the stomach contents of pigs in dry feed as well as in the stomach contents of pigs fed fermented feed. Furthermore, a significant positive correlation between the pH and the numbers of bacteria in the family Enterobacteriaceae in the contents of the stomach of pigs fed dry feed was found. In the stomach contents of pigs fed fermented feed, a significant negative correlation was found between the concentration of the undissociated form of lactic acid and the numbers of Enterobacteriaceae. The numbers of Enterobacteriaceae in the contents of the stomach, ileum, cecum, colon, and rectum of pigs fed fermented feed were significantly lower compared with the contents of the stomach, ileum, caecum, colon, and rectum of pigs fed dry feed. The numbers of total lactobacilli were significantly higher in the stomach contents of pigs fed fermented feed and in the ileum contents of one pig group fed fermented feed compared with the contents of pigs fed dry feed. However, the influence of lactobacilli on numbers of Enterobacteriaceae could not be demonstrated. It was concluded that fermented feed influences the bacterial ecology of the gastrointestinal tract and reduces the levels of Enterobacteriaceae in the different parts of the gastrointestinal tract.     

Effect of low-protein amino acid-supplemented diets on the growth performance, gut morphology, organ weights and digesta characteristics of weaned pigs

A 21-day study was conducted to determine whether isoleucine might limit the performance of piglets fed low-crude protein (CP), amino acid (AA)-supplemented diets and to investigate the potential benefits of low-CP diets on gastrointestinal health in weaned pigs. Ninety-six piglets (initial BW = 6.44 ± 0.14 kg), housed four per pen, were randomly assigned to one of four diets, resulting in six replicate pens per diet. Dietary treatments were as follows: (1) 210 g/kg CP diet, (2) 190 g/kg CP diet deficient in isoleucine, (3) 190 g/kg CP diet supplemented with crystalline isoleucine up to the level in the 210 g/kg CP diet and (4) 170 g/kg CP diet supplemented with isoleucine and valine on the ideal protein ratio basis (60% and 70% relative to lysine, respectively). Pigs were allowed to adapt to the new environment for 4 days before the experiment commenced. Overall, pigs fed the 210 g/kg CP diet had higher (P < 0.05) average daily gain and lower (P < 0.05) feed : gain ratio compared with those fed the other diets. The faecal consistency score of pigs fed the 210 g/kg CP diet was higher (P < 0.05) than those fed the other diets. Pigs fed the 170 g/kg diet had lower (P=0.02) small intestine weight than those fed the 210 g/kg CP diet. Pigs fed the 210 g/kg CP diet had deeper (P < 0.05) crypt in the duodenum and ileum and higher (P < 0.05) ammonia N concentration in caecal digesta than those fed the other diets. There were no effects of diet on microbial population and volatile fatty acid concentration in the caecal digesta except for propionic acid whose concentration was higher (P < 0.05) for pigs fed the 170 g/kg diet than those fed the 190+isoleucine and the 210 g/kg CP diets. The results indicate that the low-CP, AA-supplemented diet reduced crypt hypertrophy, ammonia N concentration in the caecal digesta, small intestine weight and the performance of piglets. Also, the results of the current study were inconclusive with respect to whether isoleucine may limit the performance of pigs fed a low-CP, AA-supplemented diet.     

Adaptive responses of Salmonella enterica serovar Typhimurium DT104 and other S. Typhimurium strains and Escherichia coli O157 to low pH environments

AIMS: Cattle are a known main reservoir for acid-resistant Escherichia coli O157 and Salmonella enterica serovar Typhimurium DT104. We studied the response of S. Typhimurium DT104 to extreme low pH environments and compared their response to that of acid-resistant E. coli O157 and other S. Typhimurium phage types. METHODS AND RESULTS: Bacteria were grown in nutrient-rich medium and subsequently acid challenged at pH 2.5. We found that stationary phase cultures of various S. Typhimurium strains were able to survive a challenge for 2 h at pH 2.5. As in E. coli, the ability of S. Typhimurium to survive at pH 2.5 was shown to be dependent on the presence of amino acids, specifically arginine. The amount of proton pumping H+/ATPase, both in E. coli O157 and S. Typhimurium strains, was lower when grown at pH values <6 than after growth at pH 7.5. Cyclo fatty acid content of membranes of bacteria grown at pH values <6 was higher than that of membranes of bacteria grown at pH 7.5. CONCLUSIONS: Various S. Typhimurium strains, both DT104 and non-DT104, are able to survive for a prolonged period of time at pH 2.5. Their response to such low pH environment is seemingly similar to that of E. coli O157. SIGNIFICANCE AND IMPACT OF THE STUDY: Food-borne pathogens like S. Typhimurium DT104 and E. coli O157 form a serious threat to public health since such strains are able to survive under extreme low pH conditions as present in the human stomach. The emergence these acid-resistant strains suggests the presence of a selection barrier. The intestinal tract of ruminants fed a carbohydrate-rich diet might be such a barrier.     

Apparent absorption of methionine and 2-hydroxy-4-methylthiobutanoic acid from gastrointestinal tract of conventional and gnotobiotic pigs

The effect of commensal microbiota and feeding corn or wheat/barley-based diets on the apparent gastrointestinal absorption of dl-methionine (MET) and 2-hydroxy-4-methylthiobutanoic acid (MHA-FA) was studied in conventional (n = 32) and gnotobiotic pigs (n = 24). Conventional pigs (CON) were vaginally delivered and sow-reared until weaning at 14 days of age. Gnotobiotic pigs were derived by caesarian section and reared in HEPA (high efficiency particulate air)-filtered isolator units with ad libitum access to a milk-based formula. Corn or wheat/barley-based diets were fed to all pigs from 14 to 24 days of age. At 24 days of age, after an overnight fast, pigs were fed 20 g/kg BW of experimental diet supplemented with 107 Bq of either 3H-l-MET or 3H-l-MHA-FA per kg of feed and chromic oxide (0.5% wt/wt). Pigs were killed for sample collection 3 h after consuming the meal. Residual 3H-MET and 3H-MHA-FA were estimated in gastrointestinal contents as the ratio of 3H : chromic oxide in digesta samples to the ratio of 3H : chromic oxide in feed. In CON pigs, feeding a wheat/barley-based diet increased (P < 0.05) total aerobes, whereas supplementation with MHA-FA increased (P < 0.05) total aerobes and lactobacilli populations in proximal small intestine (SI). Among the gnotobiotic pigs, bacterial contamination occurred such that eight pigs (two isolators) were monoassociated with a Gram-negative bacteria closely related to Providencia spp. and 16 pigs (four isolators) were monoassociated with Gram positive Enterococcus faecium. Species of monoassociated bacterial contaminant and diet composition did not affect residual methionine or MHA-FA in digesta. In both CON and monoassociated (MA) pigs, methionine and MHA-FA were retained in stomach (92%) but disappeared rapidly from proximal SI. Residual methionine and MHA-FA in digesta was not different in MA pigs; however, in CON pigs, less (P < 0.01) apparent residual methionine was found in digesta recovered at 25% (from cranial to caudal) and 75% of SI length compared with MHA-FA. Apparent residual methionine was 16% and 8% compared with 34% and 15% for MHA-FA, at the 25% and 75% locations, respectively. In proximal SI tissue, significantly (P < 0.05) higher radioactivity (cpm/mg wet tissue) was associated with MET pigs (8.56 ± 0.47) as compared to MHA-FA ones (5.45 ± 0.50). This study suggests that microbial metabolism of MHA-FA increases retention in small intestinal digesta relative to methionine and contributes, in part, to the lower bioefficacy of MHA-FA compared to methionine.     

The effect of chlortetracycline treatment and its subsequent withdrawal on multi-resistant Salmonella enterica serovar Typhimurium DT104 and commensal Escherichia coli in the pig

AIMS: To investigate the effect of a therapeutic and sub-therapeutic chlortetracycline treatment on tetracycline-resistant Salmonella enterica serovar Typhimurium DT104 and on the commensal Escherichia coli in pig. METHODS AND RESULTS: Salmonella Typhimurium DT104 was orally administered in all pigs prior to antibiotic treatment, and monitored with the native E. coli. Higher numbers of S. Typhimurium DT104 were shed from treated pigs than untreated pigs. This lasted up to 6 weeks post-treatment in the high-dose group. In this group, there was a 30% increase in E. coli with a chlortetracycline minimal inhibitory concentration (MIC) > 16 mg l-1 and a 10% increase in E. coli with an MIC > 50 mg l-1 during and 2 weeks post-treatment. This effect was less-pronounced in the low-dose group. PCR identified the predominant tetracycline resistance genes in the E. coli as tetA, tetB and tetC. The concentration of chlortetracycline in the pig faeces was measured by HPLC and levels reached 80 microg g-1 faeces during treatment. CONCLUSION: Chlortetracycline treatment increases the proportion of resistant enteric bacteria beyond the current withdrawal time. SIGNIFICANCE AND IMPACT OF THE STUDY: Treated pigs are more likely to enter abattoirs with higher levels of resistant bacteria than untreated pigs promoting the risk of these moving up the food chain and infecting man.     

Effects of phytase and 25-hydroxyvitamin D3 inclusions on the performance, mineral balance and bone parameters of grower-finisher pigs fed low-phosphorus diets

Two experiments, a performance experiment and a mineral balance study, were conducted on grower-finisher pigs (42 to 101 kg live weight) to investigate the effects of Peniophora lycii phytase enzyme and 25-hydroxyvitamin D3 (25-OHD3) on growth performance, carcass characteristics, nutrient retention and excretion, and bone and blood parameters. The two experiments were designed as a 2 × 2 factorial (two levels of phytase and two levels of 25-OHD3). The four diets were T1, low-phosphorous diet; T2, T1 + phytase; T3, T1 + 25-OHD3 and T4, T1 + phytase + 25-OHD3 diet. In all, 25 μg of 25-OHD3 was used to replace 1000 IU of vitamin D3 in diets T3 and T4. Diets were pelleted (70°C) and formulated to contain similar concentrations of energy (13.8 MJ DE/kg), lysine (9.5 g/kg) and digestible phosphorus (P; 1.8 g/kg). Neither the inclusion of phytase nor 25-OHD3 in the diet had any effect on pig performance. There was an interaction between phytase and 25-OHD3 on calcium (Ca) and P retention (P < 0.01) and on the apparent digestibility of ash (P < 0.01), P (P < 0.001) and Ca (P < 0.001). Pigs offered phytase diets only, had a higher retention of Ca and P and digestibility of ash (P < 0.01), P (P < 0.001) and Ca (P < 0.01) compared with pigs offered unsupplemented diets. However, when the combination of phytase and 25-OHD3 were offered, no effects were detected compared with 25-OHD3 diets only. Pigs fed phytase diets had higher bone ash (P < 0.01), bone P (P < 0.01) and bone Ca (P < 0.05) concentrations compared with pigs offered non-phytase diets. In conclusion, pigs offered phytase diets had a significantly increased bone ash, Ca and P than pigs offered unsupplemented phytase diets. However, there was no advantage to offering a combination of phytase and 25-OHD3 on either bone strength or mineral status compared to offering these feed additives separately.     

Identification of novel Salmonella enterica serovar Typhimurium DT104-specific prophage and nonprophage chromosomal sequences among serovar Typhimurium isolates by genomic subtractive hybridization

Genomic subtractive hybridization was performed between Salmonella enterica serovar Typhimurium LT2 and DT104 to search for novel Salmonella serovar Typhimurium DT104-specific sequences. The subtraction resulted mainly in the isolation of DNA fragments with sequence similarity to phages. Two fragments identified were associated with possible virulence factors. One fragment was identical to irsA of Salmonella serovar Typhimurium ATCC 14028, which is suggested to be involved in macrophage survival. The other fragment was homologous to HldD, an Escherichia coli O157:H7 lipopolysaccharide assembly-related protein. Five selected DNA fragments-irsA, the HldD homologue, and three fragments with sequence similarity to prophages-were tested for their presence in 17 Salmonella serovar Typhimurium DT104 isolates and 27 non-DT104 isolates by PCR. All five selected DNA fragments were Salmonella serovar Typhimurium DT104 specific among the serovar Typhimurium isolates tested. These DNA fragments can be useful for better detection and typing of Salmonella serovar Typhimurium DT104.     

Potential rates of fermentation in digesta from the gastrointestinal tract of pigs: effect of feeding fermented liquid feed

Microbial catabolic capacity in digesta from the gastrointestinal tract of pigs fed either dry feed or fermented liquid feed (FLF) was determined with the PhenePlate multisubstrate system. The in vitro technique was modified to analyze the kinetics of substrate catabolism mediated by the standing stock of enzymes (potential rates of fermentation), allowing a quantitative evaluation of the dietary effect on the catabolic capacity of the microbiota. In total, the potential rates of fermentation were significantly reduced in digesta from the large intestine (cecum, P < 0.1; colon, P < 0.01; and rectum, P < 0.0001) of pigs fed FLF compared to pigs fed dry feed. No effect of diet was observed in the stomach (P = 0.71) or the distal part of the small intestine (P = 0.97). The highest rates of fermentation and the most significant effect of diet were observed for readily fermentable carbohydrates like maltose, sucrose, and lactose. Feeding FLF to pigs also led to a reduction in the large intestine of the total counts of anaerobic bacteria in general and lactic acid bacteria specifically, as well as of microbial activity, as determined by the concentration of ATP and short-chain fatty acids. The low-molecular-weight carbohydrates were fermented mainly to lactic acid in the FLF before being fed to the animals. This may have limited microbial nutrient availability in the digesta reaching the large intestine of pigs fed FLF and may have caused the observed reduction in activity and density of the cecal and colonic microbial population. On the other hand, feeding FLF to pigs reduced the viable counts of coliform bacteria (indicator of Escherichia coli and Salmonella spp.) most profoundly in the stomach and the distal part of the small intestine, probably due to the bactericidal effect of lactic acid and low pH. The results presented clearly demonstrate that feeding FLF to pigs had a great impact on the indigenous microbiota, as reflected in bacterial numbers, short-chain fatty acid concentration, and substrate utilization. However, completely different mechanisms may be involved in the proximal and the distal parts of the gastrointestinal tract. The present study illustrates the utility of the PhenePlate system for quantifying the catabolic capacity of the indigenous gastrointestinal tract microbiota.     

The effects of cereal particle size and pelleting on the nutritive value of oat-based diets for the growing pig

Oat grains from a single source were ground in a hammer mill, either with no screen present (coarse) or through 1.56 or 4.68-mm screens, to produce three samples with different particle size distributions. At inclusion levels of 850 g kg^?1 the three samples were used as the bases of three otherwise identical diets. Half of each diet was steam pelleted to give pellets of 7-mm diameter and the other half retained as a meal form. Factorially designed metabolism experiments using 30 pigs of 20 kg initial live weight and a growth experiment using 72 individually fed pigs of 25 kg initial live weight were made.Pelleting or particle size of the diet had no significant effect on (a) apparent digestibility of dry matter (DM), modified acid detergent fibre and gross energy (b) digestible (DE) and metabolisable energy (ME) contents (the DE contents of the meal and pelleted diets were 12.81 and 13.03 MJ kg^?1 DM, respectively) and (c) nitrogen retention. Apparent digestibility of the ether-extractable fraction was significantly poorer in meals (cf. pellets) and in the diets based on coarse grains compared with those based on the two finer-ground samples. Also, apparent digestibility of nitrogen was significantly poorer in the diets based on the coarse grains (0.803) compared with that based on the 1.56-mm ground grain (0.832). Growth and efficiency of feed conversion were significantly improved by pelleting the coarse grain diet, and the meal diet based on the coarse grain gave significantly inferior results compared with those from the two other meal diets.     

Capsular polysaccharide surrounds smooth and rugose types of Salmonella enterica serovar Typhimurium DT104

The biofilms and rugose colony morphology of Salmonella enterica serovar Typhimurium strains are usually associated with at least two different exopolymeric substances (EPS), curli and cellulose. In this study, another EPS, a capsular polysaccharide (CP) synthesized constitutively in S. enterica serovar Typhimurium strain DT104 at 25 and 37 degrees C, has been recognized as a biofilm matrix component as well. Fluorophore-assisted carbohydrate electrophoresis (FACE) analysis indicated that the CP is comprised principally of glucose and mannose, with galactose as a minor constituent. The composition differs from that of known colanic acid-containing CP that is isolated from cells of Escherichia coli and other enteric bacteria grown at 37 degrees C. The reactivity of carbohydrate-specific lectins conjugated to fluorescein isothiocyanate or gold particles with cellular carbohydrates demonstrated the cell surface localization of CP. Further, lectin binding also correlated with the FACE analysis of CP. Immunoelectron microscopy, using specific antibodies against CP, confirmed that CP surrounds the cells. Confocal microscopy of antibody-labeled cells showed greater biofilm formation at 25 degrees C than at 37 degrees C. Since the CP was shown to be produced at both 37 degrees C and 25 degrees C, it does not appear to be significantly involved in attachment during the early formation of the biofilm matrix. Although the attachment of S. enterica serovar Typhimurium DT104 does not appear to be mediated by its CP, the capsule does contribute to the biofilm matrix and may have a role in other features of this organism, such as virulence, as has been shown previously for the capsules of other gram-negative and gram-positive bacteria.     

Induction and resuscitation of viable but nonculturable Salmonella enterica serovar typhimurium DT104

Salmonella enterica serovar Typhimurium DT104 11601 was tested for its ability to maintain viability in minimal, chemically defined solutions. Periodic monitoring of growth and survival in microcosms of different ion concentrations, maintained at various temperatures, showed a gradual decline in culturable organisms ( approximately 235 days) at 5 degrees C. Organisms maintained at a higher temperature (21 degrees C) showed continuous, equivalent CFU per milliliter ( approximately 10(6)) up to 400 days after inoculation. Fluorescence microscopy with Baclight revealed that nonculturable cells were actually viable, while observations with scanning electron microscopy showed that the cells had retained their structural integrity. Temperature upshift (56 degrees C +/- 0.5, 15 s) of the nonculturable organisms (5 degrees C) in Trypticase soy broth followed by immediate inoculation onto Trypticase soy agar (TSA) gave evidence of resuscitation. Interestingly, S. enterica serovar Typhimurium DT104 from the microcosms at either 5 degrees C (1 to 200 days) or 21 degrees C (1 to 250 days) did not show enhanced growth after intermittent inoculation onto catalase-supplemented TSA. Furthermore, cells from 21 degrees C microcosms exposed to oxidative and osmotic stress showed greater resistance to stresses over increasing times of exposure than did recently grown cells. It is possible that the exceptional survivability and resilience of this particular strain may in part reflect the growing importance of this multidrug-resistant organism, in general, as a cause of intestinal disease in humans. The fact that S. enterica serovar Typhimurium DT104 11601 is capable of modifying its physiological characteristics, including entry into and recovery from the viable but nonculturable state, suggests the overall possibility that S. enterica serovar Typhimurium DT104 may be able to respond uniquely to various adverse environmental conditions.     

The SopEPhi phage integrates into the ssrA gene of Salmonella enterica serovar Typhimurium A36 and is closely related to the Fels-2 prophage

Salmonella spp. are enteropathogenic gram-negative bacteria that use a large array of virulence factors to colonize the host, manipulate host cells, and resist the host's defense mechanisms. Even closely related Salmonella strains have different repertoires of virulence factors. Bacteriophages contribute substantially to this diversity. There is increasing evidence that the reassortment of virulence factor repertoires by converting phages like the GIFSY phages and SopEPhi may represent an important mechanism in the adaptation of Salmonella spp. to specific hosts and to the emergence of new epidemic strains. Here, we have analyzed in more detail SopEPhi, a P2-like phage from Salmonella enterica serovar Typhimurium DT204 that encodes the virulence factor SopE. We have cloned and characterized the attachment site (att) of SopEPhi and found that its 47-bp core sequence overlaps the 3' terminus of the ssrA gene of serovar Typhimurium. Furthermore, we have demonstrated integration of SopEPhi into the cloned attB site of serovar Typhimurium A36. Sequence analysis of the plasmid-borne prophage revealed that SopEPhi is closely related to (60 to 100% identity over 80% of the genome) but clearly distinct from the Fels-2 prophage of serovar Typhimurium LT2 and from P2-like phages in the serovar Typhi CT18 genome. Our results demonstrate that there is considerable variation among the P2-like phages present in closely related Salmonella spp.     

Effect of Fermented Feed on the Microbial Population of the Gastrointestinal Tracts of Pigs

An in vivo experiment was performed with pigs to study the inhibitory effect of fermented feed on the bacterial population of the gastrointestinal tract. Results demonstrated a significant positive correlation between pH and lactobacilli in the stomach contents of pigs in dry feed as well as in the stomach contents of pigs fed fermented feed. Furthermore, a significant positive correlation between the pH and the numbers of bacteria in the family Enterobacteriaceae in the contents of the stomach of pigs fed dry feed was found. In the stomach contents of pigs fed fermented feed, a significant negative correlation was found between the concentration of the undissociated form of lactic acid and the numbers of Enterobacteriaceae. The numbers of Enterobacteriaceae in the contents of the stomach, ileum, cecum, colon, and rectum of pigs fed fermented feed were significantly lower compared with the contents of the stomach, ileum, caecum, colon, and rectum of pigs fed dry feed. The numbers of total lactobacilli were significantly higher in the stomach contents of pigs fed fermented feed and in the ileum contents of one pig group fed fermented feed compared with the contents of pigs fed dry feed. However, the influence of lactobacilli on numbers of Enterobacteriaceae could not be demonstrated. It was concluded that fermented feed influences the bacterial ecology of the gastrointestinal tract and reduces the levels of Enterobacteriaceae in the different parts of the gastrointestinal tract.     

Identification of specific gene sequences conserved in contemporary epidemic strains of Salmonella enterica

Genetic elements specific to recent and contemporary epidemic strains of Salmonella enterica were identified using comparative genomic analysis. Two epidemic multidrug-resistant (MDR) strains, MDR Salmonella enterica serovar Typhimurium definitive phage type 104 (DT104) and cephalosporin-resistant MDR Salmonella enterica serovar Newport, and an epidemic pansusceptible strain, Salmonella serovar Typhimurium DT160, were subjected to Salmonella gene microarray and suppression subtractive hybridization analyses. Their genome contents were compared with those of coexisting sporadic strains matched by serotype, geographic and temporal distribution, and host species origin. These paired comparisons revealed that epidemic strains of S. enterica had specific genes and gene regions that were shared by isolates of the same subtype. Most of these gene sequences are related to mobile genetic elements, including phages, plasmids, and plasmid-like and transposable elements, and some genes may encode proteins conferring growth or survival advantages. The emergence of epidemic MDR strains may therefore be associated with the presence of fitness-associated genetic factors in addition to their antimicrobial resistance genes.     

Changing Dietary Calcium-Phosphorus Level and Cereal Source Selectively Alters Abundance of Bacteria and Metabolites in the Upper Gastrointestinal Tracts of Weaned Pigs

Several dietary ingredients may affect the bacterial community structure and metabolism in the porcine gut and may therefore influence animals'' health and performance. This study investigated the effects of cereal source and calcium-phosphorus (CaP) level in the diet on bacterial microbiota and metabolites, nutrient intake, and gut environment in weaned pigs. Pigs (n = 8/treatment) were fed wheat-barley- or corn-based diets with an adequate or high CaP level for 14 days. Effects on microbiota in the stomach, ileum, and midcolon were assessed using quantitative PCR. Data showed that Enterobacteriaceae, Campylobacter spp., and Helicobacter spp., which all contain highly immune reactive lipopolysaccharide (LPS), were abundant at all gut sites. Diet effects on bacteria and metabolites were moderate and occurred mainly in the upper gut, whereas no effects on bacteria, fermentation products, and LPS could be observed in the colon. Differences in carbohydrate intake with corn versus wheat-barley diets selectively stimulated Bifidobacterium in the stomach and ileum. There was a growth advantage for a few bacterial groups in the stomach and ileum of pigs fed the high versus adequate CaP level (i.e., gastric Enterobacteriaceae and ileal Enterococcus, Bacteroides-Prevotella-Porphyromonas, and Campylobacter). Interestingly, gastrointestinal pH was not affected by dietary CaP level. The present findings demonstrate the stability of the bacterial community and gut environment toward dietary changes even in young pigs. The results on stimulation of gastric and ileal Bifidobacterium by corn diets may be employed in nutritional strategies to support gut health after weaning.     

Dose determination for acute Salmonella infection in pigs

Pigs were exposed to various levels of Salmonella enterica subsp. enterica serovar Typhimurium by either intranasal inoculation or by subjecting them to a contaminated environment. More than 10(3) salmonellae were required to induce acute Salmonella infection. These results indicate that intervention against acute Salmonella infection in lairage may be more readily achieved than previously thought.     

Effects of diets differing in protein source and technical treatment on digestibility,performance and visceral and biochemical parameters of fattening pigs

The aim of the experiment on 100 cross-bred barrows was to compare commercial diets for fattening pigs based on either soya bean meal (SBM) imported from non-European countries with diets based on a mixture of locally produced rape seed meal, distillers’ dried grains with solubles and soya beans as main protein sources. In addition, these both types of diets were processed by two different technical feed treatments, i.e. coarse grinding without hydrothermal treatment or fine grinding and pelleting. With only few exceptions, nutrients of the diet without SBM were more digestible (p < 0.05) resulting in a higher metabolisable energy (ME) content. Fine grinding and pelleting increased also the ME content and the nutrient digestibility with the exception of crude fibre. Higher feed intake of animals that fed diets without SBM (p < 0.01) resulted in higher average daily gain (p < 0.01). However feeding this diet, the higher digestibility was not reflected in a decreased feed-to-gain ratio (FGR), but fine grinding and pelleting reduced FGR (p < 0.001). A higher pH value and a lower DM content of caecal chymus were detected in animals that received coarsely ground feed (p < 0.05). Animals that fed finely ground and pelleted feed had higher slaughter and relative liver weights and higher blood cholesterol concentrations (p = 0.040). The urea concentrations of blood were lower (p = 0.019) after feeding diets without SBM. In conclusion, SBM imported from non-European countries can be replaced by alternative local protein sources without compromising digestibility or performances of animals. Although fine grinding and thermal treatment particularly seemed to be advantageous for digestibility and performance, the possible risk of development of stomach lesions should be considered.