Whole-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

Zhao

et al. 2015

Appl Environ Microbiol

dAlthough fructooligosaccharides (FOS) can selectively stimulate the growth and activity of probiotics and beneficially modulate the balance of intestinal microbiota, knowledge of the molecular mechanism for FOS metabolism by probiotics is still limited. Here a combined transcriptomic and physiological approach was used to survey the global alterations that occurred during the logarithmic growth of Lactobacillus plantarum ST-III using FOS or glucose as the sole carbon source. A total of 363 genes were differentially transcribed; in particular, two gene clusters were induced by FOS. Gene inactivation revealed that both of the clusters participated in the metabolism of FOS, which were transported across the membrane by two phosphotransferase systems (PTSs) and were subsequently hydrolyzed by a ␤-fructofuranosidase (SacA) in the cytoplasm. Combining the measurements of the transcriptome-and membrane-related features, we discovered that the genes involved in the biosynthesis of fatty acids (FAs) were repressed in cells grown on FOS; as a result, the FA profiles were altered by shortening of the carbon chains, after which membrane fluidity increased in response to FOS transport and utilization. Furthermore, incremental production of acetate was observed in both the transcriptomic and the metabolic experiments. Our results provided new insights into gene transcription, the production of metabolites, and membrane alterations that could explain FOS metabolism in L. plantarum. Prebiotics are defined as nondigestible food ingredients that selectively stimulate the growth and activity of beneficial microbial strains residing in the host gastrointestinal tract (GIT) (1). Among the sugars that are qualified as prebiotics, fructooligosaccharides (FOS) are fructose polymers of diverse lengths that can be either derivatives of simple fructose polymers or fructose moieties attached to a sucrose molecule (2). Because of the linkage configuration, FOS are not digested in the upper GIT and have been shown in vivo to beneficially modulate the composition of the intestinal microbiota by preferentially increasing the numbers of bifidobacteria and lactobacilli (3, 4).Despite considerable commercial and research interest in the beneficial effects of FOS, the molecular basis of FOS metabolism by specific members of the intestinal microbiota has only recently been examined. In order to understand the influence of environmental conditions on genome-wide gene expression levels, wholegenome DNA microarrays have often been used to survey the gene expression patterns of strains in the presence and absence of oligosaccharides (2, 5-8). On the basis of in silico analysis of the Lactobacillus acidophilus NCFM genome sequence, Barrangou et al. (2) identified a multiple-sugar metabolism (msm) operon that was involved in the metabolism of FOS. The msm operon encodes an ATP-dependent binding cassette-type transport system and a cytoplasmic ␤-fructosidase, which mediates FOS uptake and intracellular hydrolysis. Moreover, expression of the operon was i...

“…As stated before, a number of microarray or RNA-seq experi- (46). Although the results are different, some commonalities could be found.…”

Section: Discussionmentioning

confidence: 75%

Section: Discussionmentioning

confidence: 99%

Metabolism of Fructooligosaccharides in Lactobacillus plantarum ST-III via Differential Gene Transcription and Alteration of Cell Membrane Fluidity

Zhao

et al. 2015

Appl Environ Microbiol

“…T (11) and L. ruminis L5 (31). The organization and genes of the L. curvatus NRIC 0822 and L. acidipiscis KCTC 13900 motility operons are very similar (98.8% identity between the concatenated proteins), suggesting that the L. curvatus NRIC 0822 motility genes were horizontally acquired recently.…”

Section: Discussionmentioning

confidence: 95%

Detection and Genomic Characterization of Motility in Lactobacillus curvatus: Confirmation of Motility in a Species outside the Lactobacillus salivarius Clade

Cousin

Lynch

Harris

et al. 2015

Appl Environ Microbiol

Lactobacillus is the largest genus within the lactic acid bacteria (LAB), with almost 180 species currently identified. Motility has been reported for at least 13 Lactobacillus species, all belonging to the Lactobacillus salivarius clade. Motility in lactobacilli is poorly characterized. It probably confers competitive advantages, such as superior nutrient acquisition and niche colonization, but it could also play an important role in innate immune system activation through flagellin-Toll-like receptor 5 (TLR5) interaction. We now report strong evidence of motility in a species outside the L. salivarius clade, Lactobacillus curvatus (strain NRIC 0822). The motility of L. curvatus NRIC 0822 was revealed by phase-contrast microscopy and soft-agar motility assays. Strain NRIC 0822 was motile at temperatures between 15°C and 37°C, with a range of different carbohydrates, and under varying atmospheric conditions. We sequenced the L. curvatus NRIC 0822 genome, which revealed that the motility genes are organized in a single operon and that the products are very similar (>98.5% amino acid similarity over >11,000 amino acids) to those encoded by the motility operon of Lactobacillus acidipiscis KCTC 13900 (shown for the first time to be motile also). Moreover, the presence of a large number of mobile genetic elements within and flanking the motility operon of L. curvatus suggests recent horizontal transfer between members of two distinct Lactobacillus clades: L. acidipiscis in the L. salivarius clade and L. curvatus in the L. sakei clade. This study provides novel phenotypic, genetic, and phylogenetic insights into flagellum-mediated motility in lactobacilli.M otility in bacterial species is often mediated by a sophisticated molecular structure called the flagellum. This chief organelle of bacterial motility is self-assembled using dozens of different proteins and rotates to propel the cell forward (1). The filament of the bacterial flagellum is composed of one or more flagellin proteins, a microbe-associated molecular pattern (MAMP) which is recognized by the host Toll-like receptor 5 (TLR5) (2) and which, via activation of the nuclear factor kappa B (NF-B) signaling pathway, engages defense responses both systemically and at epithelial surfaces (3). Several flagellate bacterial pathogens (alphaproteobacteria and epsilonproteobacteria) have evolved flagellin proteins with sequence changes that avoid TLR5 recognition while maintaining motility (4). In an ecosystem, flagellum-mediated motility may confer a competitive advantage on motile species over nonmotile species with respect to niche colonization, biofilm formation, and the secretion of virulence proteins by pathogenic bacteria (5).Lactobacillus spp. constitute a very diverse group and the largest genus within the lactic acid bacteria (LAB). Lactobacilli are associated mainly with food production and probiotics (6). Some Lactobacillus species colonize the gastrointestinal, oral, and genital tracts of humans, making them important members of the human microbiota (7)....

“…Bacterial species in which flagellar synthesis is known to be suppressed by glucose (Adler and Templeton, ; Dobrogosz and Hamilton, ; Kang et al ., ; Lawley et al ., ) use the phosphoenolpyruvate:sugar PTS to transport glucose. Because members of the family Vibrionaceae also take up glucose by group translocation via the PTS, we assumed that a glucose‐specific PTS component may participate in regulation of flagellar motility in the human pathogen V. vulnificus .…”

Section: Resultsmentioning

confidence: 99%

Glucose induces delocalization of a flagellar biosynthesis protein from the flagellated pole

Park

Lee

et al. 2016

Molecular Microbiology

To survive in a continuously changing environment, bacteria sense concentration gradients of attractants or repellents, and purposefully migrate until a more favourable habitat is encountered. While glucose is known as the most effective attractant, the flagellar biosynthesis and hence chemotactic motility has been known to be repressed by glucose in some bacteria. To date, the only known regulatory mechanism of the repression of flagellar synthesis by glucose is via downregulation of the cAMP level, as shown in a few members of the family Enterobacteriaceae. Here we show that, in Vibrio vulnificus, the glucose-mediated inhibition of flagellar motility operates by a completely different mechanism. In the presence of glucose, EIIA(Glc) is dephosphorylated and inhibits the polar localization of FapA (flagellar assembly protein A) by sequestering it from the flagellated pole. A loss or delocalization of FapA results in a complete failure of the flagellar biosynthesis and motility. However, when glucose is depleted, EIIA(Glc) is phosphorylated and releases FapA such that free FapA can be localized back to the pole and trigger flagellation. Together, these data provide new insight into a bacterial strategy to reach and stay in the glucose-rich area.