Gene expression profile of carbon starvation gene knockouts, ΔcstA and ΔyjiY, in Salmonella typhimurium
ABSTRACT: Two carbon starvation (cst) genes, cstA and yjiY, in Salmonella are predicted to mediate peptide utilization under nutrient stress. However, the mechanistic details of their function and their importance in the context of Salmonella pathogenesis is not addressed. In this study, the transcriptomes of cst gene knockouts were compared to that of wild type to check the global impact of cst genes on Salmonella metabolism as well as pathogenesis. Microarray was done for duplicate samples for each strain in late log phase of growth in LB, which is also marked by the expression of many virulence associated genes. Organism : Salmonella typhimurium , Agilent Custom Salmonella Typhimurium Gene Expression 8x15k Array (AMADID: 046639) designed by Genotypic Technology Private Limited.
Project description:Amino acid racemases are enzymes that catalyze the conversion between L and D- forms of amino acids. While the role of alanine and glutamate raemases in bacteria (specifically Salmonella) are well-studied, relatively less is known about the function of other non-canonical racemases. This study focuses on delineating the role of two such putative aspartate raceases viz. YgeA and AspR in Salmonella Typhimurium. For this purpose, Salmonella strains engineered to lack either one or both of these genes were compared to the wild type strain under planktonic and biofilm-inducing conditions to identify the global changes in gene expression orchestrated by these genes. Organism : Salmonella Typhimurium , Agilent Custom Salmonella Typhimurium str. 14028S Gene Expression 8x15k Array (AMADID: 046639) designed by Genotypic Technology Private Limited.
Project description:Bacterial pathogens adapt to multiple stress conditions; however, the consequences of these responses on infection are not well understood. Here, the adaptive responses of Salmonella enterica serovar Typhimurium (S. typhimurium) to the combination of nutritional deprivation and high temperature (NDHT) stress were investigated. Microarray analysis revealed extensive modulation of expression of genes involved in several metabolic pathways. Interestingly, the expression of a large number of genes involved in chemotaxis, invasion, motility etc are down regulated. Further studies involved S. typhimurium exposed to high temperature (HT) or grown in minimal media, i.e. nutritional downshift (ND), or the combination, i.e. NDHT. The amounts of secretory proteins reduce greatly during NDHT, but not ND or HT, resulting in lower number of colony forming units (CFU) in different organs post oral infection of mice. The expression of the global regulator, RpoS, is downregulated during NDHT. Deletion of rpoS greatly reduces the amounts of secretory proteins and intracellular replication of S. typhimurium during ND but not HT. Extension of these results to RAW macrophage cells demonstrated lower intracellular replication of S. typhimurium at HT. Overall, this study sheds insights on the adaptive responses of S. typhimurium to a combination of stress conditions, resulting in lower infection. Agilent one-color experiment,Organism: Salmonella Typhimurium ,Agilent-Custom GeneExpression 8x15k designed by Genotypic Technology Pvt. Ltd. (AMADID: 017809) , Labeling kit: Agilent Quick-Amp labeling Kit (p/n5190-0442)
Project description:Part of a study to characterise the two component regulatory system yehUT of Salmonella enterica serovar Salmonella Typhi and Typhimurium. 24 Samples examined, 12 of strain Salmonella Typhi BRD948 and 12 of strain Salmonella Typhimurium ST4/74.
Project description:Investigation of whole genome gene expression level changes in a Salmonella enterica serovar Typhimurium UK1 delta-iacP mutant, compared to the wild-type strain. IacP is resoponsible for the secretion of virulence effector proteins via the type III secretion system, thereby contributing the virulence of S. Typhimurium. The mutants analyzed in this study are further described in Kim et al. 2011. Role of Salmonella Pathogenicity Island 1 Protein IacP in Salmonella enterica Serovar Typhimurium Pathogenesis. Infection and Immunity 79(4):1440-1450 (PMID 21263021). A chip study using total RNA recovered from two separate wild-type cultures of Salmonella enterica serovar Typhimurium UK1 and two separate cultures of a mutant strain, Salmonella enterica serovar Typhimurium UK1 delta-iacP. Each chip measures the expression level of 4,302 genes from Salmonella enterica serovar Typhimurium.
Project description:Investigation of whole genome gene expression level changes in a Salmonella enterica serovar Typhimurium 14028 delta GidA mutant The mutant described in this study is further analyzed in Shippy, D. C., N. M. Eakley, P. N. Bochsler, and A. A. Fadl. 2011. Biological and virulence characteristics of Salmonella enterica serovar Typhimurium following deletion of glucose-inhibited division (gidA) gene. Microb Pathog. A single chip study using three separate cultures of wild-type Salmonella enterica serovar Typhimurium 14028 and three separate cultures of a single mutant, delta GidA Salmonella enterica serovar Typhimurium 14028.
Project description:BACKGROUND: This study was conducted to analyse M. leprae gene expression profiles in biopsy of leprosy patient and in environmental soil samples. Investigation was carried out to find out the expression profile for the minimal gene set necessary for growth of environmental M. leprae and its genes potentially require for infection and pathogenesis of leprosy. METHOD: RNA was extracted from human biopsy and environmental soil samples and total RNA obtained by subtractive hybridization of rRNA, was reverse transcribed and amplified using random primers. The whole genome was tiled at 10bp to obtain probes having 60 mer oligonucleotides in sense orientation. 179963 probes were designed in both sense and antisense orientation. Blast was performed against the mRNA sequence databases to check the specificity of the probes. Finally, 359926 probes were designed and 56579 specific probes were replicated to fill the remaining spots. RESULTS: Up regulation of several functional genes of M. leprae genome were observed from soil samples responsible for metabolism of Carbon compounds , Amino acids and amines  , Fatty acids synthesis , Lipid Biosynthesis , Phosphorous compounds . Up regulation of genes were observed from soil samples in energy metabolism function such as Glycolysis , Pyruvate dehydrogenase , TCA cycle , Glyoxylate bypass . In up regulation of respiration function  genes were observed and some of genes responsible for miscellaneous oxidoreductases and oxygenases  function, ATP-proton motive force  were also observed. Up regulation of genes for cell envelope function , conserved membrane proteins , cell processes , central intermediary metabolism , polyamine synthesis, biosynthesis of cofactors, prosthetic groups and carriers , broad regulatory functions , macromolecule metabolism, degradation of macromolecules , miscellaneous transferases , conserved hypothetical  and unknowns  were observed. Significant up regulation of Virulence genes , PE and PPE families  were also observed from soil samples. CONCLUSION: Transcriptome analysis indicated that up regulation of essential genes of M. leprae transcripts in environmental samples as compared to M. leprae from human biopsy samples which strongly suggested that several M. leprae genes are expressed more in environment for its survival and may act as source of infection. RNA was extracted from human biopsy and environmental soil samples and total RNA obtained by subtractive hybridization of rRNA, was reverse transcribed and amplified using random primers. The whole genome was tiled at 10bp to obtain probes having 60 mer oligonucleotides in sense orientation. 179963 probes were designed in both sense and antisense orientation. Blast was performed against the mRNA sequence databases to check the specificity of the probes. Finally, 359926 probes were designed and 56579 specific probes were replicated to fill the remaining spots.
Project description:Salmonella enterica Serovar Typhimurium (S. Typhimurium) causes enterocolitis in humans and calves characterized by diarrhea and polymorphonuclear cell (PMN) influx to the intestinal mucosa. The Salmonella Type III Secretion System encoded at Pathogenicity Island I (SPI-1) translocates the Salmonella effector proteins SipA, SopA, SopB, SopD, and SopE2 into the host epithelial cell cytoplasm. These five effector proteins act in concert to induce fluid secretion and transcription of C-X-C chemokines, which serve to recruit PMNs to the intestine. While the individual molecular interactions of these Salmonella proteins with cultured host cells have been extensively characterized, their combined role in the generation of fluid secretion and inflammation is less well understood. A bovine ligated ileal loop model was used in conjunction with a custom bovine microarray to determine intestinal response to acute S. Typhimurium infection in the calf. Gene expression responses to both wild type S. Typhimurium a delta sipA, sopABDE2 mutant were measured at seven times during the initial 12 hours of infection. Microarray analysis confirmed increased expression of genes encoding proteins previously associated with immune response to Salmonella spp. infection. Gene expression changes were mapped to molecular interaction pathways and changes in expression of mechanistic genes, which are defined as perturbed genes identified by Bayesian genetic network modeling, were strongly involved in the mechanisms of the host immune response. In addition to correctly identifying known effects of wild type S. Typhimurium on host (bovine) gene expression, Bayesian genetic network modeling identified novel effects of S. Typhimurium on several molecular interaction pathways. Novel effects impacted gene regulation in the following pathways: adipocytokine signaling, insulin signaling, complement and coagulation cascades, axon guidance, gap junction, neuroactive ligand-receptor interaction, long-term depression, long-term potentiation, melanogenesis, and natural killer cell mediated cytotoxicity. Known effects were observed in the following pathways: regulation of actin cytoskeleton, apoptosis, cytokine-cytokine receptor interaction, cell adhesion molecules (CAMs), MAPK signaling, calcium signaling, Jak-STAT signaling, leukocyte transendothelial migration, adherens junction, tight junction, and ECM-receptor interactions, phosphatidylinositol signaling system, and antigen processing and presentation. Quantitative real-time PCR was used to verify the expression of some of these mechanistic genes. Microarrays were used to examine the transcriptional profiles of bovine intestinal epithelia infected with wild type Salmonella enterica serotype Typhimurium (control and wild type or delta sipA sopABDE2 mutant infected) across seven time points (15 min, 30 min, 1, 2, 4, 8, and 12 hours). Experiments were performed in quadruplicate (bovine ligated ileal loops surgeries were performed with four calves), generateing a total of 84 arrays.
Project description:Transcriptomic analysis in a Salmonella enterica Serovar Typhimurium SL 1344 that constitutively expresses stdE and stdF compared with a strain carrying an stdEF deletion A four chip study using total RNA recovered from two separate cultures of Salmonella enterica Serovar Typhimurium SL 1344 constitutively expressing stdE and stdF and two separate cultures of Salmonella enterica Serovar Typhimurium SL 1344 lacking stdE and stdF.
Project description:Part of a study to determine the impact of genome evolution on host adaptation of a group of Salmonella Typhimurium. The experiment is designed to determine the impact of culture at elevated temperature on gene experssion. strains SL1344 and 94-213 at 37 and 42C
Project description:The alternative subunit of RNA polymerase RpoS/σS controls a global adaptive response allowing many Gram-negative bacteria to survive nutrient deprivation and various stresses and contributes to biofilm formation and virulence of Salmonella enterica serovar Typhimurium. We have addressed an important but poorly understood aspect of σS-dependent control; that of a repressor. The general assumption is that negative regulation of gene expression by σS is mainly a passive phenomenon, due to competition between σS and other sigma factors for binding to a limited amount of core RNA polymerase (RNAP). Genome expression profiling and physiological characterization of mutants of Salmonella ser. Typhimurium deleted for rpoS or producing a σS protein efficient for binding to RNAP, but not to the -10 promoter element, revealed that gene repression by σS requires its binding to DNA. Therefore, gene repression by σS is an active phenomenon rather than the sole cause of σS competing with other sigma factors for RNAP binding. RNA profiles of the wild type strain and two rpoS mutants of Salmonella enterica serotype Typhimurium were generated by RNA sequencing, using three biological replicates. One rpoS mutant is deleted for rpoS (delta-rpoS), while the other (rpoSdb) contains two point mutations in rpoS (C421A and G469A) and produces an σS protein inefficient for binding to the -10 promoter element.