Project description:Listeria monocytogenes (Lm) cells can attach to both cantaloupe surface and food contact surfaces and promote biofilm growth. This study was to understand the impact of cantaloupe juice on the physiology and transcriptome of Lm planktonic cells and biofilm cells grown on stainless steel coupons using confocal laser scanning microscopy (CLSM), Cryo-Scanning Electron Microscopy (Cryo-SEM) and RNA Seq technology. Lm showed a strong autoaggregation phenotype when grown in cantaloupe juice at room temperature. It is interesting to note that Lm formed significantly more biofilms on stainless steel (SS) coupons when grown in cantaloupe juice than in TSB. SEM images revealed a different attachment profile of Lm on SS coupons. In TSB, Lm cells were mainly found in scratches/groves of the metal surface, whereas, in cantaloupe juice they attached to the smooth surface as well. Interestingly, Lm planktonic and biofilm cells in cantaloupe juice showed an elongated cell shape which might be a stress-induced phenotype in cantaloupe juice. Cantaloupe juice induced a distinct transcriptional profile of biofilm and planktonic cells of Lm from TSB. Functional annotation indicated that the significantly differentially expressed genes (DEGs, Padj < 0.05, log2foldchange ≥ 1) from the comparison mainly participated in metabolism, signaling and stress response. Notably, certain pathways downregulated for planktonic cells were significantly upregulated for biofilm cells in cantaloupe juice compared to TSB, including ABC transporters, two-component system, quorum sensing, chemotaxis, and flagellar assembly. These data highlighted the interaction of Lm with food matrix (i.e., cantaloupe) and the role of food matrix on Lm survival and adaptation. These results provided the basis for future functional characterization of genes with potential roles in biofilm formation and persistence of Lm in cantaloupe juice, as well as for development of mitigation practices for Lm biofilms on produce and food contact surfaces.
Project description:Transcriptional profiles of Salmonella Typhimurium str. ST4/74 air-dried onto stainless steel for 4 h was compared to an early stationary phase (ESP) culture control. Cells that had been air-dried for 4 h were then subsequently rehydrated with water for a 30 min period, after which the transcriptional profile was compared to an ESP control.
Project description:Transcriptional profiles of Salmonella Typhimurium str. ST4/74 air-dried onto stainless steel for 4 h was compared to an early stationary phase (ESP) culture control. Cells that had been air-dried for 4 h were then subsequently rehydrated with water for a 30 min period, after which the transcriptional profile was compared to an ESP control. Carried out using 2 biological replicates for each sample; hybridised in a two-channel hybridization against Salmonella genomic DNA as the comparator/reference.
Project description:Antimicrobials have been shown to select for changes in biofilm formation and multidrug susceptibility in common human pathogens. We investigated whether common food preservatives selected for these changes in the food pathogen Salmonella enterica serovar Typhimurium. Bacteria were exposed to four food preservatives in either planktonic cultures or biofilms grown on stainless steel beads. Cultures were passaged into fresh media supplemented with the food preservative every 72 hours. Following approximately 1000 generations of continuous preservative exposure, populations were sequenced to determine the single nucleotide polymorphisms that were selected for over evolutionary time.
Project description:Previously, we performed DNA array-based transcriptomic analysis of Clostridium acetobutylicum biofilm adsorbed onto fibrous matrix in batch fermentation. Here, to further shed light on the transcriptomic modulation of maturing Clostridium acetobutylicum biofilm, we performed the DNA array-based transcriptomic analysis in repeated-batch fermentation. Significant time course changes in expression levels were observed for the genes involved in amino acid metabolism, oligopeptide ABC transporter, nitrogen fixation, and various other processes. Repeated-batch fermentation was carried out in 2-L stainless steel columns packed with 40 g of cotton towel ?cut into pieces?approximately 3 cm × 5 cm) containing 1.5 L of P2 medium. Medium circulation rate was maintained at 35 mL/min via a peristaltic pump and the temperature was controlled at 37°C. Fermentation broth was replaced with fresh P2 medium every 12 h. Samples were withdrawn at 6 h after the medium replacement at predetermined interval, except for the last 3 samples. The last 3 samples were withdrawn at 12 h, 15 h, and 17 h after the medium replacement, respectively, to study the transcriptomic response to the adverse condition at the end of fermentation. A total of 8 samples were withdrawn over a period of 7 days, and time course gene expression profiles were studied.
Project description:Endothelial cell (EC)-enriched protein coding genes, such as endothelial nitric oxide synthase (eNOS), define quintessential EC-specific physiologic functions. It is not clear whether long noncoding RNAs (lncRNAs) also define cardiovascular cell-type specific phenotypes, especially in the vascular endothelium. Here, we report the existence of a set of EC-enriched lncRNAs and define a role for STEEL (spliced transcript – endothelial enriched lncRNA) in angiogenic potential, macrovascular/microvascular identity and shear stress responsiveness. STEEL is expressed from the terminus of the HOXD locus and is transcribed antisense to HOXD transcription factors. STEEL RNA increases the number and integrity of de novo perfused microvessels in an in vivo model and augments angiogenesis in vitro. The STEEL RNA is polyadenylated, nuclear-enriched and has microvascular predominance. Functionally, STEEL regulates a number of genes in diverse endothelial cells. Of interest, STEEL upregulates both eNOS and the transcription factor Kruppel-like factor 2 (KLF2), and is subject to feedback inhibition by both eNOS and shear-augmented KLF2. Mechanistically, STEEL upregulation of eNOS and KLF2 is transcriptionally mediated, in part, via interaction of chromatin-associated STEEL with the poly-ADP ribosylase, PARP1. For instance, STEEL recruits PARP1 to the KLF2 promoter. This work identifies a role for EC-enriched lncRNAs in the phenotypic adaptation of ECs to both body position and hemodynamic forces, and establishes a newer role for lncRNAs in the transcriptional regulation of EC identity.
Project description:Pseudomonas aeruginosa is a pathogenic micro-organism responsible for many hospital-acquired infections. It is able to adhere to solid surfaces and develop an immobilised community or so-called biofilm. Many studies have been focusing on the use of specific materials to prevent the formation of these biofilms, but the reactivity of the bacteria in contact to surfaces remains unknown. In order to evaluate the impact of different materials on the physiology of Pseudomonas aeruginosa during the first stage of biofilm formation, i.e. adhesion, we investigated the total proteome of cells adhering to three materials: stainless steel, glass and polystyrene. Using tandem mass spectrometry performed at the PAPPSO proteomic platform, 930 proteins were identified, 70 of which were differentially expressed between the materials. Dysregulated proteins belonged to 19 PseudoCAP (Pseudomonas Community Annotation Project) functional classes, with a particular abundance of proteins involved in small molecule transport and membrane proteins. Notably, ten porins or porin precursors were under-produced in bacteria adhering to stainless steel when compared to those adhering to polystyrene and glass. Although adhesion to solid surfaces is an extracellular phenomenon, it involves not only extracellular proteins but also intracellular reactions, as observed with the dysregulation of 11 proteins involved in various metabolisms and five in protein translation. Overall, this work showed that during bacterial adhesion, P. aeruginosa senses the materials concerned and is able to modulate its physiology accordingly.
Project description:Cronobacter sakazakii is well-known for its desiccation tolerance in the powdered infant formula (PIF) food production environment and the bacterium has been linked with high fatality rates in neonates who consume contaminated product. In this study, using deep-level RNA-sequencing, differentially expressed genes were studied in C. sakazakii ATCCTM29544 grown in simulated low-moisture environment designed to mimic the PIF production environment. Desiccation of bacteria was carried out on stainless steel coupons from which total RNA was subsequently recovered and sequenced. During 4 h of desiccation from the early stationary phase (ESP) grown culture, an approximately 3 log10 reduction was recorded for C. sakazakii viable cell count, with the largest change in viable cells occurring between desiccation hour 1 and 2 during which the culture medium was completely dried. Transcriptomic data obtained after 4 h of desiccation highlighted several highly-up regulated osmotolerance-related genes which were associated with the secondary response mechanism. These actively expressed genes mainly modulate pathways that synthetize glycine betaine and trehalose as well as the transport of these two and other compatible solutes. Understanding the activities of these genes and pathways will assist the development of technologies that mitigate the survival of C. sakazakii in the PIF production process.