2.3. Analytical methods
Feed and fecal samples were analyzed for dry matter (DM), ash, crude protein (CP),and ether extract contents following AOAC (1995) procedures.
Samples of 400 mg of digesta preserved in ethanol were precipitated by centrifugation (13,000g for 5min).
The DNA in the precipitate was extracted and purified using a commercial kit (QIAamp DNA Stool Mini Kit, Qiagen, West Sussex, UK).
Lactobacillus and enterobacteria were quantified using real-time PCR following the methods described by Castillo et al.(2006).
Ileum samples were dehydrated and embedded in paraffin wax, sectionedat 4 mm, and stained with hematoxylin and eosin.