Growth phase-associated changes in Lactobacillus rhamnosus GG
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ABSTRACT: Transcriptional profiling of probiotic Lactobacillus rhamnosus GG during growth in industrial-type whey medium in pH-controlled bioreactor cultures at two different growth pH: 4.8 and 5.8. Keywords: growth phase, growth pH Cell samples from three biological replicates were harvested at mid-exponential (4 h), late exponential (12 h), stationary transition point (16 h), early stationary (20 h) and late stationary (31 h) phases, all these samples both from pH 4.8 and 5.8 cultivations. A total of 66 hybridizations were performed according to ANOVA design where all possible direct pair-wise comparisons within biological replicates were conducted. Growth series between treatments (pH 4.8 or pH 5.8) were combined together by hybridizing RNA samples from the treatments at time points 12 h and 31 h to microarrays using dye-swap replicates. Treatments were paired twice and each biological replicate was used only once to form the pairs. Other comparisons between the two pH treatments were estimated computationally.
Project description:Transcriptional profiling of probiotic Lactobacillus rhamnosus GG during growth in industrial-type whey medium in pH-controlled bioreactor cultures at two different growth pH: 4.8 and 5.8. Keywords: growth phase, growth pH
Project description:For a pathogen such as Streptococcus suis serotype 2, ecological success is determined by its ability to sense the environment and mount an appropriate adaptive transcriptional response. Thus, determining conditions for analysis of gene expression in vitro that are representative of the in vivo environment is critical for understanding the contributions of transcriptional response pathways to pathogenesis. In this study, we used analysis of the global expression profile in response to acidic pH in vitro and identified a set of regulated genes involved in diverse cellular processes. 196 (11%) genes were differentially regulated by the acid stress: 92 (47%) were down-regulated at low acid (pH 5.8) relative to the neutral condition (pH 7.2), whereas 104 (53%) were up-regulated at pH 5.8 versus pH 7.2. To confirm the microarray data, 16 genes were measured by quantitative RT-PCR. There was a strong positive correlation (r = 0.96) between the results obtained by microarray and quantitative RT-PCR. The data showed that S. suis S2 is equally capable of inducing an acid tolerance response with maximal protection provided after adaptation at pH 5.8 for survival. A cDNA microarray imprinted with 2156 genes representing about 98% of the Streptococcus suis serotype 2 genome was used for transcriptome analysis. For two-sample (reference vs. test) microarray hybridization, four independent bacterial cultures from each condition (pH 5.8, pH 7.2) were prepared as biological replicates for RNA isolation. Accordingly, four dual-fluorescence-labeled cDNA probes were prepared to hybridize with four slides. Pairwise comparisons were made using dye-swaps to avoid labeling bias. A ratio of mRNA levels (test/reference) was calculated for each gene. Significant changes of gene expression were identified with the SAM software. After the SAM analysis, only genes with at least 2-fold changes in expression were collected for further analysis.
Project description:Lactobacillus plantarum WCFS1 was differentially fermented in 29 different fermentations according to a factorial, combinatorial scheme that included variations in growth temperature (28 or 37C), NaCl concentration (0 or 0.3M), pH (5.2, 5.8, 6.4), as well as oxygen (N2 or air) and amino acid availability (1.1 or 2x concentration). Samples were hybridized in a star design using five samples in the center all connected to each other. The remaining samples were linked (evenly distributed) to the samples in the inner circle in a loop design with additional links (shortcuts) to the center sample. All samples were hybridized in duplicate (at minimum) using each of the two dyes (Cy3/Cy5) at least once per sample.
Project description:Lactobacillus plantarum WCFS1 was differentially fermented in 29 different fermentations according to a factorial, combinatorial scheme that included variations in growth temperature (28 or 37C), NaCl concentration (0 or 0.3M), pH (5.2, 5.8, 6.4), as well as oxygen (N2 or air) and amino acid availability (1.1 or 2x concentration).
Project description:For a pathogen such as Streptococcus suis serotype 2, ecological success is determined by its ability to sense the environment and mount an appropriate adaptive transcriptional response. Thus, determining conditions for analysis of gene expression in vitro that are representative of the in vivo environment is critical for understanding the contributions of transcriptional response pathways to pathogenesis. In this study, we used analysis of the global expression profile in response to acidic pH in vitro and identified a set of regulated genes involved in diverse cellular processes. 196 (11%) genes were differentially regulated by the acid stress: 92 (47%) were down-regulated at low acid (pH 5.8) relative to the neutral condition (pH 7.2), whereas 104 (53%) were up-regulated at pH 5.8 versus pH 7.2. To confirm the microarray data, 16 genes were measured by quantitative RT-PCR. There was a strong positive correlation (r = 0.96) between the results obtained by microarray and quantitative RT-PCR. The data showed that S. suis S2 is equally capable of inducing an acid tolerance response with maximal protection provided after adaptation at pH 5.8 for survival.
Project description:Limited data are available on the impacts of copper (Cu)-pH-interaction-responsive genes in roots. Citrus sinensis seedlings were treated with 300 (Cu-toxicity) or 0.5 uM (control) CuCl2 x pH 3.0 or 4.8 for 17 weeks. Thereafter, we investigated the impacts of Cu-pH interactions on transcriptomics in roots.
Project description:Survival of the foodborne pathogen Listeria monocytogenes in acidic environments (e.g., stomach and low pH foods) is vital to its transmission. L. monocytogenes grows at temperatures as low as 2M-BM-0C, and refrigerated, ready-to-eat foods have been sources of L. monocytogenes outbreaks. The purpose of this study was to determine whether growth at a low temperature (i.e., 7M-BM-0C) affects the response of L. monocytogenes to sudden acid shock. A full genome microarray was used to determine changes in L. monocytogenes 10403S gene expression after exposure to acidified brain-heart infusion (BHI; pH 3.5) for 5 or 15 min. To determine changes in gene transcription after acid treatment, separate competitive hybridizations were performed between cDNA from untreated cells (grown at 7M-BM-0C or 37M-BM-0C to log or stationary phase) and (i) cells acid treated for 5 min or (ii) cells acid treated for 15 min. For L. monocytogenes grown to log or stationary phase, competitive hybridizations were performed between total cDNA from non-acid-treated cells grown to 7M-BM-0C and non-acid-treated cells grown to 37M-BM-0C to determine baseline differences in gene transcription between growth temperatures prior to acid treatment. For each experiment, four biological replications were completed. Hybridizations were carried out with dye swapping (i.e., for each comparison, each cDNA from each condition was labeled with each dye exactly twice) to help minimize dye incorporation bias.
Project description:Transcriptional profiling of probiotic Lactobacillus rhamnosus strain GG mid-exponential pH-controlled bioreactor cultures before and after exposure to bovine bile (0.2% ox gall). Keywords: bile, stress response Cell samples from four biological replicates were harvested right before (time point 0 min) and 10, 30 and 120 min after bile treatment. Each sample was compared to a common reference sample (time point 0 min, mid-exponential growth phase Lactobacillus rhamnosus GG cultures). A total of 12 hybridizations were performed using balanced dye-swap design. Dyes were balanced between compared sample pairs and between biological replicates.
Project description:We revealed that an enhancement of rice growth by 2'-deoxymugineic acid (DMA) application was observed not only under high pH conditions where iron availability for plant uptake was reduced but also under normal pH conditions. This result indicates that DMA application improves not only Fe availability for plants but also plant productivity. To explain a mechanism caused by the DMA application, molecular regulation in rice treated with or without DMA was analyzed using microarray analysis and qRT-PCR. Results provide insight into advantages of DMA application in rice seedlings. Gene expression patterns induced by DMA and EDTA in root and shoot were analyzed with control experiment (no chelator). One independent experiment was performed at each pH (pH 5.8 or pH 8.0).