Fecal Hyodeoxycholic Acid Is Correlated With Tylosin-Induced Microbiome Changes in Growing Pigs

Abstract

The changes in the gut microbiome play an important role in the promoting effects of antibiotics, such as tylosin, to the health, and productivity of farm animals. Microbial metabolites are expected to be key mediators between antibiotics-induced microbiome changes and growth-promoting effects. The objective of this study was to extend the identification of tylosin-responsive microbes to the identification of tylosin-responsive metabolites in growing pigs. The feeding trial was conducted on a commercial farm using two pens of pigs fed diets with and without tylosin (40 mg/kg of diet). Fecal samples were collected from 10 pigs per pen at weeks 10, 13, 16, 19, and 22 of age, and subsequently analyzed using liquid chromatography-mass spectrometry (LC-MS) analysis. The multivariate model of LC-MS data showed that time-dependent changes occurred in the fecal metabolome of both control and tylosin-treated pigs. More importantly, the metabolomic profiles were similar between the tylosin treatment and control groups in weeks 10 and 22, but diverged during weeks 13-19. Subsequent analyses of the fecal metabolites contributing to the separation of two groups of pigs showed that hyodeoxycholic acid (HDCA), together with tylosin and its metabolites in feces, was greatly increased during weeks 13-19 (P < 0.05) in the group of pigs fed tylosin. The integration of current metabolomics data and the microbiome data from a previous study revealed the consistency between HDCA and a specific genus of microbes in the Clostridia family. Further studies are required to determine the causative relations between tylosin-elicited changes in HDCA and the microbiome as well as the role of HDCA in the growth promoting effects of tylosin.

Keywords: antibiotics; bile acids; metabolomics; microbiome; pigs.

Figure 1

Identification of fecal metabolites induced by tylosin treatment through LC-MS-based metabolomics. (A) Scores plot of a PLS-DA model on fecal samples from the tylosin-treated and control pigs. The t(1) and t(2) values represent the scores of each data point in the principal component 1 and 2 of the model, respectively. These values are the averages of 10 pigs under the same treatment at weeks 10, 13, 16, 19, and 22. (B) S-plot of an OPLS model on week 13–19 control and tylosine treatment samples. The fecal metabolites contributing to the separation of two groups of pigs are labeled. The p(1) axis represents the magnitude of the fecal ions. The p(corr)(1) axis represents the correlation of the ions toward the predictive variation induced by tylosin treatment. (C) Extracted chromatograms of tylosin standard and a fecal sample. (D) Extracted chromatograms of HDCA standard and a fecal sample.

Figure 2

Concentrations of bile acids in fecal samples from control and tylosin-treated pigs from week 10 to week 22. (A) HDCA. (B) LCA. (C) deoxycholic acid (DCA). (D) CA. Values are mean ± S.D. (*P ≤ 0.05; **P ≤ 0.01; ***P ≤ 0.001).

Figure 3

Concentrations of SCFAs in fecal samples from control and tylosin-treated pigs from week 10 to week 22. (A) Acetic acid. (B) Propionic acid. (C) Butyric acid. (D) Valeric acid. Values are mean ± S.D. (*P ≤ 0.05).

Figure 4

Comparison of relative abundance for significantly correlated bacterial species over time for HDCA (A) and LCA (B,C).