Fig. 3: Effect of reuterin on rumen microbial composition and isolation of the bacterial host.
From: Plasmid-encoded toxin defence mediates mutualistic microbial interactions
a, Distribution plot representing the concentration of reuterin (in mM) detected in rumen samples from 20 cows. b, Phylogenetic analysis of the cultured microbial rumen community after serial passaging with or without reuterin addition at the genus level. The bar graph represents the normalized abundances of the genera. Genera with low abundances in control communities, including Adlercreutzia, Bacteroides, Bifidobacterium, Blautia, Butyricicoccus, Butyrivibrio, Dialister, Peptococcus, Prevotella, Roseburia and Sediminibacterium, are not represented. c, Top gel: PCR of part of the 1,3-PD gene (expected product length 560 bp), amplified from the plasmid-purified fraction of five isolates. Bottom gel: PCR of the whole plasmid encoding the 1,3-PD gene, using reverse primers (expected product length ~5,300 bp), amplified from the plasmid-purified fraction of five isolates. In both gels, plasmid pBR322:1,3-PD was used as a positive control template. Both gels were reproducible in two distinct experiments. Illustrations on the right represent the plasmid in blue, and the 1,3-PD gene in orange and arrows indicate the primers used for the PCR. d, STORM and wide-field (inset) imaging of E. faecalis MM1 cells labelled by CARD-FISH with specific probes for the 1,3-PD defence plasmidic gene (top) or GroEL chromosomal gene (bottom). White dashed lines delimit the bacterial cell walls. Similar images were obtained from two distinct experiments. e, Comparative resistance of E. faecalis MM1 plasmid-carrying strain and E. faecalis DSM 8630 pMM1 transformed with the rumen plasmid with the 1,3-PD defence gene against reuterin and E. faecalis DSM 8630 ∅ (with an empty plasmid backbone lacking the 1,3-PD defence gene). The strains were grown at increasing concentrations of reuterin (0 to 0.9 mM) in 1,000 μl basal defined medium and incubated for 24 h at 37 °C (volumes of cultures were homogenized across samples using sterile water). Turbidity measurements at 600 nm were used to determine the protection of the 1,3-PD defence gene against reuterin. The y axis represents the % growth of a specific strain as compared to growth of the same strain without reuterin. The measurements were carried out in biological triplicates, for which actual data, averages and standard deviations are presented.
