Interrelations between the microbiotas in the litter and in the intestines of commercial broiler chickens

Abstract

The intestinal microbiota of broiler chickens and the microbiota in the litter have been well studied, but the interactions between these two microbiotas remain to be determined. Therefore, we examined their reciprocal effects by analyzing the intestinal microbiotas of broilers reared on fresh pine shavings versus reused litter, as well as the litter microbiota over a 6-week cycle. Composite ileal mucosal and cecal luminal samples from birds (n = 10) reared with both litter conditions (fresh versus reused) were collected at 7, 14, 21, and 42 days of age. Litter samples were also collected at days 7, 14, 21, and 42. The microbiotas were profiled and compared within sample types based on litter condition using PCR and denaturing gradient gel electrophoresis (PCR-DGGE). The microbiotas were further analyzed using 16S rRNA gene clone libraries constructed from microbiota DNA extracted from both chick intestinal and litter samples collected at day 7. Results showed significant reciprocal effects between the microbiotas present in the litter and those in the intestines of broilers. Fresh litter had more environmental bacteria, while reused litter contained more bacteria of intestinal origin. Lactobacillus spp. dominated the ileal mucosal microbiota of fresh-litter chicks, while a group of bacteria yet to be classified within Clostridiales dominated in the ileal mucosal microbiota in the reused-litter chicks. The Litter condition (fresh versus reused) seemed to have a more profound impact on the ileal microbiota than on the cecal microbiota. The data suggest that the influence of fresh litter on ileal microbiota decreased as broilers grew, compared with temporal changes observed under reused-litter rearing conditions.

FIG. 1.

DGGE banding profiles (A) and corresponding PCA analysis (B) of the litter microbiotas collected at days 7, 14, 21, and 42. In figure A, “M” represents the electrophoresis marker. In figure B, symbols are as follows: triangle, day 7; diamond, day 14; circle, day 21; square, day 42; open symbols, fresh litter samples; closed symbols, reused litter samples.

FIG. 2.

DGGE banding profiles (A) and corresponding PCA analysis (B) of the ileal mucosal microbiotas collected at days 7, 14, 21, and 42. In figure A, “M” represents the electrophoresis marker. Bands 1 to 3 and 4 to 7 changed intensity and disappeared over time, respectively. In figure B, symbols are as follows: triangle, day 7; diamond, day 14; circle, day 21; square, day 42; open symbols, fresh-litter birds; closed symbols, reused-litter birds.

FIG. 3.

DGGE banding profiles (A) and corresponding PCA analysis (B) of the cecal luminal microbiotas collected at days 7, 14, 21, and 42. In figure A, “M” represents the electrophoresis marker. In figure B, symbols are as follows: triangle, day 7; diamond, day 14; circle, day 21; square, day 42; open symbols, fresh-litter birds; closed symbols, reused-litter birds.

FIG. 4.

Rarefaction curves of clones sequenced from reused litter (A), fresh litter (B), ileal mucosa of reused-litter chicks (C), or fresh-litter chicks (D) or cecal content of reused-litter chicks (E) or fresh-litter chicks (F). Sequence similarity level: asterisks, 97%; squares, 95%; triangles, 90%; circles, 85%. There is no 95 or 90% similarity curve for the ileal clones of reused-litter chicks (see Fig. 3C) because the few OTUs identified for this sample differed only by a count of 1 between 97 and 85% similarity.

FIG. 5.

Phylogenetic tree of OTUs defined at 97% similarity. LR, reused litter; LF, fresh litter; IR, ileal mucosa of reused-litter chicks; IF, ileal mucosa of fresh-litter chicks; CR, cecal content of reused-litter chicks; CF, cecal content of fresh-litter chicks.

FIG. 5.

Phylogenetic tree of OTUs defined at 97% similarity. LR, reused litter; LF, fresh litter; IR, ileal mucosa of reused-litter chicks; IF, ileal mucosa of fresh-litter chicks; CR, cecal content of reused-litter chicks; CF, cecal content of fresh-litter chicks.