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TitleWhole-transcriptome shotgun sequencing (RNA-seq) screen reveals upregulation of cellobiose and motility operons of lactobacillus ruminis l5 during growth on tetrasaccharides derived from barley β-Glucan
Publication TypeJournal Article
Year of Publication2013
AuthorsLawley, B., Sims I.M., and Tannock G.W.
JournalApplied and Environmental Microbiology
Pagination5661 - 5669
Date Published2013
ISSN00992240 (ISSN)
Keywordsarticle, bacterium, Batch cell culture, Batch culture, beta glucan, beta-Glucans, bioassay, cellobiose, chemistry, Cross-feeding, Early growth, enzyme activity, gene expression, genetics, glucose, Glucose-grown cells, Growth phase, growth rate, growth, development and aging, Hordeum, Hydrolysis, Lactobacillus, Locomotion, Mammals, Metabolic Networks and Pathways, metabolism, motility, oligosaccharide, Oligosaccharides, operon, phenotype, Phenotypic changes, Polymerase Chain Reaction, real time polymerase chain reaction, Real-Time Polymerase Chain Reaction, RNA, ruminant, Strain, Tetrasaccharides, Transcription, transcriptome, Up-regulation, upregulation
AbstractLactobacillus ruminis is an inhabitant of human bowels and bovine rumens. None of 10 isolates (three from bovine rumen, seven from human feces) of L. ruminis that were tested could utilize barley β-glucan for growth. Seven of the strains of L. ruminis were, however, able to utilize tetrasaccharides (3-O-β-cellotriosyl-D-glucose [LDP4] or 4-O-β-laminaribiosyl-D-cellobiose [CDP4]) present in β-glucan hydrolysates for growth. The tetrasaccharides were generated by the use of lichenase or cellulase, respectively. To learn more about the utilization of tetrasaccharides by L. ruminis, whole-transcriptome shotgun sequencing (RNA-seq) was tested as a transcriptional screen to detect altered gene expression when an autochthonous human strain (L5) was grown in medium containing CDP4. RNA-seq results were confirmed and extended by reverse transcription-quantitative PCR assays of selected genes in two upregulated operons when cells were grown as batch cultures in medium containing either CDP4 or LDP4. The cellobiose utilization operon had increased transcription, particularly in early growth phase, whereas the chemotaxis/motility operon was upregulated in late growth phase. Phenotypic changes were seen in relation to upregulation of chemotaxis/flagellar operons: flagella were rarely seen by electron microscopy on glucose-grown cells but cells cultured in tetrasaccharide medium were commonly flagellated. Chemotactic movement toward tetrasaccharides was demonstrated in capillary cultures. L. ruminis utilized 3-O-β-cellotriosyl-D-glucose released by β-glucan hydrolysis due to bowel commensal Coprococcus sp., indicating that cross feeding of tetrasaccharide between bacteria could occur. Therefore, the RNA-seq screen and subsequent experiments had utility in revealing foraging attributes of gut commensal Lactobacillus ruminis. © 2013, American Society for Microbiology.

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