Metagenomic Characterization of Intestinal Regions in Pigs With Contrasting Feed Efficiency

Abstract

Greater feed efficiency (FE) is critical in increasing profitability while reducing the environmental impact of pig production. Previous studies that identified swine FE-associated bacterial taxa were limited in either sampling sites or sequencing methods. This study characterized the microbiomes within the intestine of FE contrasting Duroc × (Landrace × Yorkshire) (DLY) pigs with a comprehensive representation of diverse sampling sites (ileum, cecum, and colon) and a metagenomic sequencing approach. A total of 226 pigs were ranked according to their FE between weaning to 140 day old, and six with extreme phenotypes were selected, three for each of the high and low groups. The results revealed that the cecum and colon had similar microbial taxonomic composition and function, and had higher capacity in polysaccharide metabolism than the ileum. We found in cecum that the high FE pigs had slightly higher richness and evenness in their micriobiota than the low FE pigs. We identified 12 phyla, 17 genera, and 39 species (e.g., Treponema porcinum, Treponema bryantii, and Firmicutes bacterium CAG:110) that were potentially associated with swine FE variation in cecum microbiota through LEfSe analysis. Species enriched in the cecum of the high FE pigs had a greater ability to utilize dietary polysaccharides and dietary protein according to the KEGG annotation. Analysis of antibiotic resistance based on the CARD database annotation indicated that the macB resistant gene might play an important role in shaping the microbial community in the cecum of pigs with contrasting FE. The bacteria from the genus Prevotella was highly enriched in the cecum of low FE pigs, which may impair the establishment of a more effective nutrient harvesting microbiota because of the interaction between Prevotella and other benefical microbes. These findings improved our understanding of the microbial compositions in the different gut locations of DLY pigs and identified many biomarkers associated with FE variation wich may be used to develop strategies to improve FE in pigs.

Keywords: DLY pigs; feed efficiency; intestinal microbiome; macB gene; metagenome.

FIGURE 1

Overview of the experimental design. Feed efficiency (FE) was defined as the weight gained between weaning and 140 days of age, divided by total feed intake.

FIGURE 2

Distribution of abundance of genes in the gene catalog. The gene catalog was constructed by merging genes identified in the present study and a previous reference set (Xiao et al., 2016). (A) Venn diagram of intersection between the genes in the reference set and identified in the present study. (B) Boxplot showing the distribution of gene abundance in each category based on the Venn diagram.

FIGURE 3

Alpha diversity of microbial community among three intestinal locations and between high and low FE groups. (A) The Chao1 index. (B) The Shannon index. Each point represents one sample as indicated by the axis label and color of the point.

FIGURE 4

Comparison of microbial composition based on species abundance among samples. (A) Non-metric multidimensional scaling (NMDS) plot according to FE performance and intestinal location based on the abundance of species. The plot is based on the Bray–Curtis distances between pairs of samples. Each point represents one sample; ellipses represent the 95% confidence for all points within each cluster. Stress, the value used to estimate the NMDS ordination fitness. R, the statistic from the analysis of similarities (ANOSIM) that compares the mean of ranked dissimilarities between groups to the mean of ranked dissimilarities within groups. p, the p-value from ANOSIM analysis between groups. (B) The abundance ratio of different species between the ileum microbiota and hindgut microbiota. The species are ordered along the X-axis according to their rank of abundance ratio between the ileum and non-ileum.

FIGURE 5

Linear discriminant analysis (LDA) effect size (LEfSe) analysis. The same analysis was performed at the (A) phylum, (B) genus, and (C) species levels to compare the cecum microbiota profiles between the high and low FE groups.

FIGURE 6

Functional annotation of the microbiomes. (A) Heatmap of the 10 most abundant (based on TPM value) carbohydrate active enzyme (CAZy) families in any of the groups. Color scale shows the abundance of CAZy enzyme family within each group. Z-score, calculated with the formula z = (x − μ)/σ, where x is the log10 of abundance of enzymic families in each group, μ is the mean value of the log10 of abundance in all groups, and σ is the standard deviation of the log10 of abundance. (B) NMDS plot of high and low FE of cecum samples based on the abundance of CAZy families. (C) LEfSE analysis for KEGG orthology (KO) to compare the cecum microbiota functional profiles between the high and low FE group. (D) Heatmap of the 10 most abundant (based on TPM value) antibiotic resistance genes (ARGs) in any of the groups. (E) Ternary plot showing the abundance comparison of the 10 most abundant ARGs of each group in the ileum (green), cecum (brown), and colon (blue). The sum of the abundance for one specific ARG in these three types of gut was set as 100%. The percentage (%) of one specific ARG in each gut location is equal to its corresponding abundance divided by the abundance sum of this ARG in the three intestinal locations. The symbol size indicate the total abundance of the ARGs. The symbol color indicated the types of the ARGs.