Prokaryotes

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    Protocols in Current Issue
    Optimized Protocol for the Incorporation of FDAA (HADA Labeling) for in situ Labeling of Peptidoglycan
    Authors:  Katharina Peters, Manuel Pazos, Michael S. VanNieuwenhze and Waldemar Vollmer, date: 08/05/2019, view: 517, Q&A: 0
    [Abstract] The essential peptidoglycan (PG) layer surrounds the cytoplasmic membrane in nearly all bacteria. It is needed to maintain the shape of the cell and protect it from lysis due to high turgor. Growth of the PG layer is a complex process that involves the activities of PG synthases and hydrolases during elongation and cell division. PG growth sites ...
    Microfluidics-Based Analysis of Contact-dependent Bacterial Interactions
    Authors:  Robert Cooper, Lev Tsimring and Jeff Hasty, date: 08/20/2018, view: 2180, Q&A: 0
    [Abstract] Bacteria in nature live in complex communities with multiple cell types and spatially-dependent interactions. Studying cells in well-mixed environments such as shaking culture tubes or flasks cannot capture these spatial dynamics, but cells growing in full-fledged biofilms are difficult to observe in real time. We present here a protocol for ...
    Adhesion of Enteroaggregative E. coli Strains to HEK293 Cells
    Authors:  Jorge Luis Ayala-Lujan and Fernando Ruiz-Perez, date: 04/20/2018, view: 2608, Q&A: 0
    [Abstract] Enteroaggregative Escherichia coli (EAEC) is a recognized cause of acute diarrhea among both children and adults worldwide. EAEC strains are characterized by the presence of aggregative adherence fimbriae (AAF), which play a key role in pathogenesis by mediating attachment to the intestinal mucosa and by triggering host inflammatory ...
    In vivo Analysis of Cyclic di-GMP Cyclase and Phosphodiesterase Activity in Escherichia coli Using a Vc2 Riboswitch-based Assay
    Authors:  Ying Liu, Hyunhee Kim and Ute Römling, date: 03/05/2018, view: 2932, Q&A: 0
    [Abstract] Cyclic di-guanosine monophosphate (c-di-GMP) is a ubiquitous second messenger that regulates distinct aspects of bacterial physiology. It is synthesized by diguanylate cyclases (DGCs) and hydrolyzed by phosphodiesterases (PDEs). To date, the activities of DGC and PDE are commonly assessed by phenotypic assays, mass spectrometry analysis of ...
    Bacterial Survival in Dictyostelium
    [Abstract] We performed an assay to test the ability of different E. coli strains to survive inside amoebal cells after ingestion. In the assay we incubated bacteria together with cells of Dictyostelium discoideum for six hours. After co-incubation most of the uningested bacteria were removed by centrifugation and the remaining uningested ...
    Isolation and Infection of Drosophila Primary Hemocytes
    Authors:  Charles Tracy and Helmut Krämer, date: 06/05/2017, view: 5175, Q&A: 0
    [Abstract] Phagocytosis of invading pathogens and their subsequent clearance in lysosomes is important for organismal fitness. We have devised the following protocol to extract phagocytic hemocytes from wild-type and mutant Drosophila larvae and infect the isolated hemocytes with GFP-labeled E. coli to measure the rate of phagocytosis and ...
    Identification and Characterization of Bacterial Chemoreceptors Using Quantitative Capillary and Gradient Plate Chemotaxis Assays
    [Abstract] Bacterial chemotaxis is a motility-based response that biases cell movement toward beneficial molecules, called attractants, and away from harmful molecules, also known as repellents. Since the species of the genus Pseudomonas are characterized by a metabolic versatility, these bacteria have developed chemotactic behaviors towards a wide range of ...
    Fitness Measurements of Evolved Esherichia coli
    Authors:  Migla Miskinyte and Isabel Gordo, date: 09/05/2014, view: 7960, Q&A: 0
    [Abstract] Bacteria can adapt very rapidly to novel selective pressures. In the transition from commensalism to pathogenicity bacteria have to face and adapt to the host immune system. Specifically, the antagonistic interaction imposed by one of the first line of defense of innate immunity cells, macrophages, on commensal bacteria, such as Escherichia coli ...
    Iodine Staining of Escherichia coli Expressing Genes Involved in the Synthesis of Bacterial Glycogen
    [Abstract] The presence of intracellular glycogen can be detected by the following iodine staining technique. Cells with glycogen stain dark brown, whereas in its absence they remain with a pale yellowish color. It is hypothesized that iodine atoms fit into helical coils formed by the α-polyglucan to form a coloured glycogen-iodine complex. Here, we have ...
    Preparation of Outer Membrane Vesicle from Escherichia coli
    [Abstract] Outer membrane vesicles (OMVs) are spherical bilayered phospholipids of 20-200 nm in size produced from all Gram-negative bacteria and Gram-positive bacteria investigated to date. Previous biochemical and proteomic studies have revealed that the Gram-negative bacteria-derived OMVs are composed of various components like outer membrane proteins, ...



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