2.9. Survival of the four probiotic strains in the piglet feed stored at room temperature
The survival of the probiotic bacteria in piglet feedwas studied using the freezing cultures
of each LAB obtained as described above. The commercial piglet feed was mixed with the
corresponding defrosted culture (20 mL/kg of feed) and stored at room temperature. Daily,
duplicate samples (10 g) of probiotic supplemented piglet feed were mixed 1:10 with sterile
PBS and vortexed for 2 min. Both samples were serially diluted using sterile PBS and each
dilution was plated in triplicate in MRS or Rothe agar. The plates were incubated at 30 ◦C
for 2 days. Incubated plates were observed for the optimum number of CFU, between 30
and 300 colonies per plate. The results were expressed as the number of colonies counted
per gram (wet weight) of feed.
2.10. Treatment of piglets, preparation of experimental diets and analysis
On the 21st day of age, a total of 80 piglets were distributed into 4 groups of 20: the
non-treated control group, 2 probiotic supplemented fed groups and the antibiotic (colistin
sulfate) supplemented fed group. Each group was housed separately in individual cages.
The composition of the basal diet used is shown in Table 3. Each experimental group was
fed ad libitum with its own diet for 42 days. The temperature of the room with continuous
lighting was maintained at 28 ◦C initially, and reduced 1 ◦C/week until reached 24 ◦C, at
which the room temperature was maintained for the end of the experiment.
This experimentwas divided in two trials. In the first, the two potentially probiotic strains
tested were Ped. acidilactici and Ent. faecium. In the second trial, which was developed
in the same conditions, the two probiotic strains assayed were L. lactis and Lact. casei.