XII - 12th International Symposium - Digestive Physiology of Pigs

Digestive
Physiology
of Pigs
Key words: in vitro, digestion, viscosity
3033 Discrepancies in microbiota composition along
the pig gastro-intestinal tract between in vivo observations
and an in vitro batch fermentation model. C.
Boudry* 1 , C. Poelaert 1 , D. Portetelle 2 , A. Thewis 1 , and J.
Bindelle 1 , 1 Animal Science Unit, Gembloux Agro-Bio Tech,
University of Liege, Gembloux, Belgium, 2 Animal and
Microbial Biology Unit, Gembloux Agro-Bio Tech, University
of Liege, Gembloux, Belgium.
In vitro fermentation models are increasingly used to assess
prebiotic potential of novel indigestible carbohydrates
(ICHO). A trial was performed to assess the validity of such
approaches by comparing the influence of fermentation of
inulin (INU) and cellulose (CEL) on microbiota in vivo and
in vitro. Three INU and CEL based semi-purified diets (5%
INU, 5% CEL and 2.5% of both) were fed to 3 groups of
4 pigs (≈25 kg). After 3 weeks, the pigs were slaughtered
and digesta was sampled from jejunum, ileum, cecum and
3 parts of the colon to measure pH, SCFA and microbiota
population. One week before slaughter, an in vitro gas
fermentation test was performed on INU and CEL with fresh
feces of the experimental pigs as bacterial inoculum. The
gas production kinetics were modeled and fermentation
broth samples were taken after 5, 8, 12, 24 and 72 h.
Total bacterial DNA was extracted from the samples and
qPCR was performed to quantify total bacteria, Lactobacilli,
Bifidobacteria, Bacteroides, Clostridium Cl. I and E. coli.
Total bacteria quantification showed similarities between
both systems. In vivo, total bacteria increased along the gut
until the second part of the colon (from 10.5 7 to 10 10 cfu mg −1 ,
P < 0.001) and then decreased to 10 9 cfu (P < 0.05), while in
vitro, it increased until 12 to 24 h of fermentation (+5 10 8 cfu
ml −1 ) and then decreased to initial level. This evolution was
in relation with fermentation kinetics. In both models, INU
increased Bifidobacteria and E. coli populations compared
with CEL (P < 0.05). However, in vivo this was observed
only in the first parts of the gut while in vitro, the effect
lasted during the 72 h fermentation. Bacteroides genus was
not influenced by the ICHO source in the 2 systems (P >
0.05). Finally, evolution of Lactobacilli and Clostridium Cl. I
populations in both systems were not consistent. This can
be ascribed to specific bacterial properties as e.g., adhesive
properties or sensitivity to sulfur reducing agent used in the
in vitro model. Further developments of the in vitro method
are required to properly assess prebiotic potential of ICHO
Key words: in vitro fermentation, microbiota, gut
3034 Comparison of three internal markers in flow
and recovery of feed enzymes in pigs. H. Jørgensen* 1 ,
L. Salmon 2 , H. N. Lærke 1 , and K. E. B. Knudsen 1 , 1 Aarhus
University, Department of Animal Science, Foulum,
Denmark, 2 Danisco Animal Nutrition, Marlborough, United
Kingdom.
To quantify the flow and survivability of feed enzymes
through the intestine a reliable marker is a prerequisite.
The study investigated 3 internal markers use to test in
vivo survivability of feed enzymes in cannulated pigs. The
markers were: titanium dioxide (TiO2), chromium III oxide
XII INTERNATIONAL SYMPOSIUM ON
DIGESTIVE PHYSIOLOGY OF PIGS
131
Session VI
(Cr2O3) and Celite analyzed as acid insoluble ash (AiA).
Three male castrate crossbred growing pigs (initial weight
30 kg) were surgically fitted with 2 T-cannula (16 mm
diameter) one at the duodenum and one anterior to the ileocecal
junction. Pigs were kept in individual pens with half the
floor area covered with raised plastic gratings throughout
the whole experiment and the pens were equipped with
toys and drinking nipples. The diet was based on barleywheat-soybean
meal fortified with vitamins and minerals
and the diet was heat treated and pelleted to reduce the
dietary phytase level. Pigs were fed according to scale 3
times daily (0800, 1600 and 2300h). In the morning meal
(0800h) on Monday, Wednesday and Friday the marker
and the enzyme preparation were added according to a 3
× 3 Latin Square design. Collection of digesta took place
at 0800h and 0900h from the duodenum cannulas (300 g)
and at 0900h and 1030h from the ileal cannulas (200 g).
The digesta was immediately frozen (−20°C) until further
analysis. The concentration of the markers in the bolus
meal was as expected. The variation of the markers in both
duodenum and ileal digesta was as expected higher than
in the diet. The relative variation (CV, Std dev/Mean*100)
was in the duodenum digesta 13, 41 and 18% for Cr2O3,
TiO2 and AiA, respectively. For ileum collected at 1030h the
variation was in the order of 25, 50 and 21%. The lowest
concentration of markers and highest relative variations
was found at ileum collected at 0900h, indicating that only
a small portion of the bolus meal had arrived at this time.
Furthermore in 2 cases the concentration of TiO2 in ileal
digesta collected at 0900 was close to the detection limit
causing very high variation. For this kind of study chromic
oxide and Celite was equal suitable.
Key words: duodenum, bolus meal, ileum
3035 ex vivo model for investigating the bacterial
association to the gut epithelium of pigs. S. Sugiharto,
B. Jensen, and C. Lauridsen,* Aarhus University, Foulum,
Denmark.
Diarrhea-like conditions are difficult to reproduce in pigs. To
study Enterotoxigenic E. coli (ETEC) association, in vitro
cellular models have been used, but these are less suitable
for ETEC pathogenesis study in pigs. This experiment
aimed to establish a model to study the association of
ETEC to the gut epithelium of pig. Intestine cultures were
prepared from 4 weaned pigs susceptible to both E. coli
O149:F4 (homo- and heterozygotic, 2 pigs each) and
O138:F18 (all homozygotic). Five segments (15 cm each)
were taken from 50% of the intestinal length measured
from duodenum (mid-small intestine [SI]), and a further 5
segments were taken from 90% (distal-SI), immersed in
Dulbecco’s Modified Eagle Medium (DMEM) and kept on
ice. Polyethylene tubing was inserted into either end of the
segment and tied. The tissue was washed with 50 mL of
PBS. The other end of segment was tied, 10 mL of DMEM
alone or DMEM containing either E. coli F4 or F18 was
inoculated and the segment was sealed with Teflon plug.
The culture was immersed in DMEM in a 300-mL infusion
bottle in a shaking water bath (150 rpm) at 37°C. After 1 h
the culture was removed, tissue was washed with 50 mL of
PBS, weighed and homogenized in PBS. Final dilution of