Fig 1.
Antibiotics alters diversity and composition of intestinal microbiota.
A. Antibiotics statistically reduced Ace, Chao, Shannon and Sobs, which are the indices of intestinal microbiota. *p<0.05, ***p<0.001 compared with control. B. Percent of community abundance on phylum level. Antibiotics significantly increased the percentage of Proteobacteria.
Fig 2.
NMDS and LEfSe analysis of intestinal microbiota composition after antibiotics treatment.
A. The green dots, representing the distributions of samples in control group, mainly distribute at the lower right area. The red triangles, standing for samples of antibiotics group, locate at the left area. B. Green symbolizes the differential bacteria in control group, and red stands for bacteria with significant difference in antibiotics group. Species with in-apparent variation are represented by yellow dot. The circle is segmented into five layers, respectively representing phylum, class, order, family, and genus levels from the inside out.
Fig 3.
Potential biomarkers and correlation analysis.
A. Wilcoxon rank sum test analysis to figure out the potential biomarkers. X axis is percentage value of specific species abundance, and Y axis represents species name at various classification levels. *p<0.05, **p<0.01, ***p<0.001 when compared with control group. B. X axis is environmental factors including intestinal permeability, acetate, propionate, and butyrate. Y axis represents specific species on phylum level. Spearman correlation coefficients are represented by color ranging from blue, negative correlation (-0.4), to red, positive correlation (0.6). *p<0.05, **p<0.01, ***p<0.001 when compared with control group.
Fig 4.
Functional prediction of intestinal microbiota community.
A. COG function classification of antibiotics group. X axis represents function class, and Y axis stands for abundance. B. Comparison of COG function between control and antibiotics group. X axis represents samples and Y axis stands for relative abundance. C. Comparison of abundance related to short chain fatty acids biosynthesis and metabolism. X axis represents different functions and Y axis stands for abundance value. D. Antibiotics decreased SCFAs concentrations. Concentrations of SCFAs (acetate, propionate and butyrate) in ileocecal feces were assessed after antibiotics treatment. Antibiotics remarkably decreased the concentrations of all SCFAs. Data represent the mean±SEM. *p<0.05, **p<0.01, ***p<0.001 compared with control.
Fig 5.
Antibiotics impairs intestinal barrier function.
A. Intestinal permeability to 4.4 kDa FITC-dextran was significantly increased after antibiotics treatment. Data represent the mean±SEM (n = 6). *p<0.05, **p<0.01, ***p<0.001 compared with control. B. Antibiotics significantly decreased proteins expressions of ZO-1, occludin and claudin-1. Data represent the mean±SEM (n = 6). *p<0.05, **p<0.01, ***p<0.001 compared with control. OCC: occludin, CL-1: claudin-1. C. ZO-1 (red) was mainly localized to the epithelial tight junctions in the ileum of control mice. In contrast, antibiotics disrupted the normal arrangement of ZO-1 at the apical junctions. Data are representative of three independent experiments. Scar bar = 10.0μm.
Fig 6.
Antibiotics activates NLRP3 inflammasome.
A. After antibiotics treatment, protein expressions of NLRP3, ASC, cleaved caspase-1, cleaved IL-1β and IL-18 increased significantly. But expressions of caspase-1 and IL-1β decreased gradually. Data represent the mean±SEM (n = 6). *p<0.05, **p<0.01, ***p<0.001 compared with control. B. Antibiotics remarkably augmented the amount of ASC protein (red punctate). Scale bar = 25.0μm.
Fig 7.
Antibiotics activates autophagy.
A. Antibiotics increased the expressions of Atg5 and ratio of LC3-II/LC3-I, but decreased P62 expression. Data represent the mean±SEM (n = 6). **p<0.01, ***p<0.001 compared with control. B. Antibiotics increased the amount of LC3B (red punctate). Scale bar = 25.0μm.