ABSTRACT: Pseudomonas aeruginosa, Echerichia coli, Mycobacterium tuberculosis, and uncultivated soil bacterium Raw sequence reads and de novo assembled draft genomes
Project description:Transcriptional changes during early infection of macrophage-like THP-1 cell line with pathogenic bacterium Mycobacterium tuberculosis. RNAseq samples were taken at 0h (THP-1 cells growing in the RPMI medium), and after 4h, 24h and 48h post infection. Bacterial enrichment was performed to increase the amount of bacterial mRNA in the samples. Non-enriched samples were used to map THP-1 cells transcripts; enriched samples were used to map M. tuberculosis transcripts the corresponding genomes.
Project description:Mycobacteria can synthesize NAD+ using either the de novo biosynthesis pathway or the salvage pathway. The deletion of the three genes involved specifically in the NAD+ de novo biosynthesis pathway in the human pathogen Mycobacterium tuberculosis had no effect on the growth of the strain in vivo. In contrast, the same deletion in the bovine pathogen Mycobacterium bovis resulted in a strain that could not grow in vivo and could only grow in vitro with substantial nicotinamide supplmentation. This striking difference was attributed to the known defect in the nicotinamidase PncA of M. bovis, since introducing the M. tuberculosis pncA gene into the M. bovis strain defective for de novo NAD+ biosynthesis restored growth in vitro and in vivo. This study demonstrates that NAD+ starvation is a cidal event in mycobacteria and confirms that enzymes common to the de novo and salvage pathways may be good drug targets. We also propose that simultaneously targeting both the salvage and the de novo NAD+ biosynthesis pathways represents a potentially effective way to treat infection with tubercle bacilli. To characterize the lethality induced by nicotinamide starvation transcriptional profiling of the auxotrophs was performed. Triplicate 50 mL cultures of M. tuberculosis and M. bovis Delta nadABC mutants were grown in 7H9 OADC glycerol 0.05% tween broth in 500 mL roller bottles to an OD600nm= 0.1 in a roller incubator at 37°C. The cells were washed 1x in PBS and resuspended in 50 mL 7H9 OADC glycerol 0.05% tween broth with or without 20mg/L nicotinamide and returned to the incubator. After 7 days, cultures were harvested. Three biological replicates of each of two species with one dye-flip each
Project description:Mycobacteria can synthesize NAD+ using either the de novo biosynthesis pathway or the salvage pathway. The deletion of the three genes involved specifically in the NAD+ de novo biosynthesis pathway in the human pathogen Mycobacterium tuberculosis had no effect on the growth of the strain in vivo. In contrast, the same deletion in the bovine pathogen Mycobacterium bovis resulted in a strain that could not grow in vivo and could only grow in vitro with substantial nicotinamide supplmentation. This striking difference was attributed to the known defect in the nicotinamidase PncA of M. bovis, since introducing the M. tuberculosis pncA gene into the M. bovis strain defective for de novo NAD+ biosynthesis restored growth in vitro and in vivo. This study demonstrates that NAD+ starvation is a cidal event in mycobacteria and confirms that enzymes common to the de novo and salvage pathways may be good drug targets. We also propose that simultaneously targeting both the salvage and the de novo NAD+ biosynthesis pathways represents a potentially effective way to treat infection with tubercle bacilli. To characterize the lethality induced by nicotinamide starvation transcriptional profiling of the auxotrophs was performed. Triplicate 50 mL cultures of M. tuberculosis and M. bovis Delta nadABC mutants were grown in 7H9 OADC glycerol 0.05% tween broth in 500 mL roller bottles to an OD600nm= 0.1 in a roller incubator at 37°C. The cells were washed 1x in PBS and resuspended in 50 mL 7H9 OADC glycerol 0.05% tween broth with or without 20mg/L nicotinamide and returned to the incubator. After 7 days, cultures were harvested.
Project description:Earthworms show a wide spectrum of regenerative potential with certain species like Eisenia fetida, a terrestrial redworm, capable of regenerating more than two-thirds of their body while other closely related species, such as Paranais litoralis seem to have lost this ability. Earthworms belong to the phylum annelida, in which the genomes of the marine oligochaete Capitella telata, and the freshwater leech Helobdella robusta have been sequenced and studied. Herein, we report the de novo assembled transcriptome of Eisenia fetida (Indian isolate), along with an analysis of the transcriptomic changes during regeneration. We also used de novo assembled RNAseq data to identify genes that are differentially expressed during regeneration, both in the newly regenerating cells and in the adjacent tissue.
Project description:De novo assembled transcriptomics-assisted label-free quantitative proteomics analysis reveals sex-specific proteins in the intestinal tissue of Haemaphysalis qinghaiensis
Project description:The Zika outbreak, spread by the Aedes aegypti mosquito, highlights the need to create high-quality assemblies of large genomes in a rapid and cost-effective fashion. Here, we combine Hi-C data with existing draft assemblies to generate chromosome-length scaffolds. We validate this method by assembling a human genome, de novo, from short reads alone (67X coverage, Sample GSM1551550). We then combine our method with draft sequences to create genome assemblies of the mosquito disease vectors Aedes aegypti and Culex quinquefasciatus, each consisting of three scaffolds corresponding to the three chromosomes in each species. These assemblies indicate that virtually all genomic rearrangements among these species occur within, rather than between, chromosome arms. The genome assembly procedure we describe is fast, inexpensive, accurate, and can be applied to many species.
Project description:The Mycobacterium tuberculosis acyl-CoA carboxylases provide the building blocks for de novo fatty acid biosynthesis by fatty acid synthase (FAS) I and for the elongation of FAS I end-products by the FAS II complex to produce meromycolic acids. M. tuberculosis genome contains three biotin carboxylase subunits (AccA1-3) and 6 carboxyltransferase subunits (AccD1-6) of which AccD6 is located in a genetic locus that contains members of the FAS II complex. We found by microarray and quantitative real-time RT-PCR analysis that the transcripts of AccA3, AccD4, AccD5 and AccD6 are expressed at high levels during exponential growth phases of M. tuberculosis in vitro. Keywords: Time course, developmental stages