Extensive microbial and functional diversity within the chicken cecal microbiome

Abstract

Chickens are major source of food and protein worldwide. Feed conversion and the health of chickens relies on the largely unexplored complex microbial community that inhabits the chicken gut, including the ceca. We have carried out deep microbial community profiling of the microbiota in twenty cecal samples via 16S rRNA gene sequences and an in-depth metagenomics analysis of a single cecal microbiota. We recovered 699 phylotypes, over half of which appear to represent previously unknown species. We obtained 648,251 environmental gene tags (EGTs), the majority of which represent new species. These were binned into over two-dozen draft genomes, which included Campylobacter jejuni and Helicobacter pullorum. We found numerous polysaccharide- and oligosaccharide-degrading enzymes encoding within the metagenome, some of which appeared to be part of polysaccharide utilization systems with genetic evidence for the co-ordination of polysaccharide degradation with sugar transport and utilization. The cecal metagenome encodes several fermentation pathways leading to the production of short-chain fatty acids, including some with novel features. We found a dozen uptake hydrogenases encoded in the metagenome and speculate that these provide major hydrogen sinks within this microbial community and might explain the high abundance of several genera within this microbiome, including Campylobacter, Helicobacter and Megamonas.

Figure 1. Summary of 16S analysis.

(A) Rarefaction curves of the OTUs clustered at 97% sequence identity for each chicken, based on the averages of the four replicates per bird. (B) Hierarchical clustering visualizing similarities among cecal samples. The number in each label indicates each individual chicken and the letter each individual ceca (either A or B). Technical replicates of each ceca were carried out. (C) Bar chart showing the proportion of reads assigned to the top 5 most abundant OTUs in each chicken.

Figure 2. Polysaccharide-degrading enzymes in the chicken cecal metagenome.

The figure shows each class of enzyme as judged by SEED/KEGG annotation and GH (see methods) for three types of NSP. The size of pie chart reflects abundance of the enzyme class; numbers indicate quantity of genes assigned to each bacterial Taxon at the class level.

Figure 3. Gene clusters associated with polysaccharide degradation and utilization.

NSP degrading genes were identified by SEED/KEGG annotation and GH domain (see methods). (A) Gene clusters encoding putative PUL systems from various Bacteroidetes. (B) Gene cluster encoding putative integrated polysaccharide degradation and utilization systems from various Firmicutes. (C) NSP degrading enzymes associated with sporulation genes * indicates predicted signal peptide.

Figure 4. Pathways and gene clusters associated with propionoate production in the chicken cecal metagenome.

(A) Pathways involved in propionoate production showing the putative genes identified coding for the enzymes involved. Size of pie chart reflects abundance of the gene class; numbers indicate quantity of genes assigned to each taxon. (B) Operon structure of two Methylmalonyl decarboxylase loci.

Figure 5. Pathways and gene clusters associated with butyrate production in the chicken cecal metagenome.

(A) Pathways involved in butyrate production showing the putative genes identified coding for the enzymes involved. Size of pie chart reflects abundance of the gene class; numbers indicate quantity of genes assigned to each taxon. (B) Operon structure of two butyryl-CoA:acetate-CoA transferase (BCD) loci.

Figure 6. Potential Hydrogen sinks in the Chicken cecal metagenome.

The key genes involved in each pathway that could potentially use hydrogen are shown. The size of pie chart reflects abundance of the gene class; numbers indicate quantity of genes assigned to each taxon.