Rumen Microbiota Distribution Analyzed by High-Throughput Sequencing After Oral Doxycycline Administration in Beef Cattle

Abstract

The beef cattle rumen is a heterogenous microbial ecosystem that is necessary for the host to digest food and support growth. The importance of the rumen microbiota (RM) is also widely recognized for its critical roles in metabolism and immunity. The level of health is indicated by a dynamic RM distribution. We performed high-throughput sequencing of the bacterial 16S rRNA gene to compare microbial populations between rumens in beef cattle with or without doxycycline treatment to assess dynamic microbiotic shifts following antibiotic administration. The results of the operational taxonomic unit analysis and alpha and beta diversity calculations showed that doxycycline-treated beef cattle had lower species richness and bacterial diversity than those without doxycycline. Bacteroidetes was the predominant phylum in rumen samples without doxycycline, while Proteobacteria was the governing phylum in the presence of doxycycline. On the family level, the top three predominant populations in group qlqlwy (not treated with doxycycline) were Prevotellaceae, Lachnospiraceae, and Ruminococcaceae, compared to Xanthomonadaceae, Prevotellaceae, and Rikenellaceae in group qlhlwy (treated with doxycycline). At the genus level, the top predominant population in group qlqlwy was unidentified_Prevotellaceae. However, in group qlhlwy, the top predominant population was Stenotrophomonas. The results revealed significant RM differences in beef cattle with or without doxycycline. Oral doxycycline may induce RM composition differences, and bacterial richness may also influence corresponding changes that could guide antibiotic use in adult ruminants. This study is the first to assess microbiota distribution in beef cattle rumen after doxycycline administration.

Keywords: MiSeq sequencing; bacterial richness; beef cattle; doxycycline; dysbacteriosis; oral antibiotics; rumen microbiota.

Figure 1

Collation results of DNA sequence data and microbial diversity index analyses. (A) ACE index, (B) Good's coverage index, (C) observed species index, (D) PD whole tree, (E) Chao1 index, (F) Shannon index, (G) Venn diagram. The numbers represent the unique or common OTUs of each group. (H) ANOSIM analysis. “Between” represents the difference between groups; an R-value closer to 1 indicates greater difference. “qlqlwy” and “qlhlwy” represent the two groups.

Figure 2

Differences in bacterial community structures and relationships among all samples. (A,B) PCoA of bacterial community structures of the RM in the three sample groups. Each blue point represents a sample. The distance between two points represents the difference of RM. PCoA shows distinct bacterial communities between different samples. (C) The UPGMA tree analysis of samples in evolution in weighted unifrac distance. (D) The UPGMA tree analysis of samples in evolution in unweighted unifrac distance.

Figure 3

Sample feasibility analysis. Each curve represents a sample. (A) Rarefaction curves depicting the effect of sequences on the number of identified OTUs in the rank abundance curve. (B) The abscissa indicates the OTU (species) abundance order, and the ordinate corresponds to the relative abundance ratio of OTU (species).

Figure 4

Microbial composition of different samples. Each bar represents the average relative abundance of each bacterial taxon within a group. The top 11 abundant taxa are shown. (A) Taxa assignments at Phylum level. (B) Taxa assignments at Family level. (C) Taxa assignments at Genus level.

Figure 5

Bacterial taxa significantly differentiated rumens samples of cows with or without doxycycline identified by LEfSe using the default parameters. (A) Histogram of LDA scores computed for bacterial taxa differentially abundant between groups. (B) Bacterial taxa that were differentially abundant visualized using a cladogram.

Figure 6

Differences in bacterial abundance between the groups qlqlwy and qlhlwy. Left, abundance ratios of different strains in two samples. Middle, difference in bacterial abundance within the 95% confidence interval. Right, P-values of significance testing.