Effects Of Dietary Protein Levels On Bacterial Diversity In Ileum And Colon Of Weaned Piglets

Protein in diet is one of the sources of animal nutrients.Undigested and unabsorbed dietary protein can provide nutrition for intestinal bacteria and promote bacterial reproduction,thus affecting the composition and quantity of intestinal flora.The effects of dietary protein levels on intestinal flora and diarrhea in weaned piglets in a short period of time have been less reported.For this reason,niety-six weaned three-way hybrid piglets[weaning at 25-day-old] were randomly divided into 4 groups.After a week of pre-feeding,18%,20%,22% and 24% protein levels were fed respectively.Four piglets in each group were randomly selected to collect ileum and colon chyme at 6,24 and 72 hours after feeding.Total bacterial DNA was extracted and the changes of bacterial community structure and quantity were analyzed by high throughput sequencing and real-time fluorescence quantitative PCR,the following results are obtained:(1)Increased dietary protein level could increase the intestinal bacterial alpha-diversity of weaned piglets,but the difference was not significant(P>0.05).The intestinal diversity of piglets tended to increase with the increase of feeding time.(2)In all intestinal content samples(except H1: colon samples of 24% protein group at 6h and B2: ileum samples of 20% protein group at 24h),the highest relative content of Firmicutes(43.40%-82.74%)was found,followed by Bacteroides(0.87%-35.43%)and Proteobacteria(1.51%-27.18%).The relative content of Teneribacteria in sample B2 was12.17%,higher than that of Bacteroides(6.39%).The relative content of Actinomycetes in sample H1 was 4.51%,higher than that of Proteobacteria(2.17%).Lactobacillus was the dominant genus in all samples(4.03 %-64.84 %),except samples B3,D3,G3 and H3:ileum samples with 20%,24% protein group and colon samples with 22% and 24% protein group at 72 hours of feeding.The relative contents of Megacoccus in samples B3,D3,G3 and H3 were 15.08%,17.86%,7.02% and 8.12%,respectively,higher than those of Lactobacillus14.71%,13.84%,4.03% and 4.48%.(3)At phylum level,increasing dietary protein level could increase the relative content of Proteobacteria and Bacteroides in intestine.After 72 hours of feeding,the relative content of colonic Proteobacteria in 22% CP group was significantly higher than that in the other three groups(P<0.01).Increased dietary protein level could reduce the relative content of Firmicutes,and the change was most obvious after 72 hours of feeding(P<0.01).At genus level,increasing dietary protein level could increase the relative contents of Roseburia and Streptococcus,Megasphaera and Streptococcus(P<0.05 orP<0.01),and decrease the relative contents of Lactobacillus and Bifidobacterium(P<0.05),especially after 72 hours of feeding.(4)Increased dietary protein level could increase the number of Clostridium difficile and Escherichia coli in ileum and colon of weaned piglets,decrease the number of Lactobacillus and Bifidobacterium,and increase the number of Roseburia in ileum and decrease the number of colonic segments.At 72 hours,there were significant differences between 22% and 24% protein groups and 18% and 20% protein groups(P<0.05 or P<0.01).Conclusion:Firmicutes was the most abundant phyla in ileum and colon of weaned piglets,Lactobacillus was the most abundant genus.The increase of dietary protein level on bacterial alpha-diversity in ileum and colon of weaned piglets was not significant,but it can increase the bacterial alpha-diversity in intestinal tract of weaned piglets,increase the relative content of Proteobacteria and Bacteroides,reduce the relative content of Firmicutes;increase the relative content of Streptococcus in ileum and colon,ileal Roseburia and colonic Megasphaera.With the increase of dietary protein level,the number of Clostridium difficile and Escherichia coli in ileum and colon of weaned piglets increased,while the number of Lactobacillus,Bifidobacterium decreased,and the number of Roseburia in ileum incresased and the number of colonic segments decreased,increased risk of diarrhea in piglets.