San Diego State University View Institution's Website 15 articles published in JoVE Immunology and Infection T4 Bacteriophage and E. coli Interaction in the Murine Intestine: A Prototypical Model for Studying Host-Bacteriophage Dynamics In Vivo Nicola Pett1, Michael Hunter1, Natalia A. Carranza García1, Jung Hee Seo1, Samuel R. Collins1, Forest Rohwer2, Lisa C. Osborne1, Carolina Tropini1,3,4 1Department of Microbiology and Immunology, Life Sciences Institute, University of British Columbia, 2Department of Biology, San Diego State University, 3School of Biomedical Engineering, University of British Columbia, 4Humans and the Microbiome Program, Canadian Institute for Advanced Research Bacteriophages (phages), viruses that infect bacteria, are an integral component of the gut microbiome. Though these symbiotic inhabitants drive bacterial fitness and population dynamics, little is understood about how they impact gut homeostasis and disease. This protocol studies isolated T4 phages within a mouse model, adaptable to other phage-bacterial pairs. Environment Coral Reef Arks: An In Situ Mesocosm and Toolkit for Assembling Reef Communities Jason L. Baer1, Jessica Carilli2, Bart Chadwick3, Mark Hatay1, Anneke van der Geer1, Yun Scholten4, William Barnes4, Jenna Aquino1, Ashton Ballard1, Mark Little1, Jared Brzenski5, Xiaofeng Liu6, Gunther Rosen2, Pei-Fang Wang2, Jose Castillo5, Andreas F. Haas4, Aaron C. Hartmann7, Forest Rohwer1 1Department of Biology, San Diego State University, 2Energy and Environmental Sciences Branch, Naval Information Warfare Center (NIWC) Pacific, 3Coastal Monitoring Associates, 4Department of Marine Microbiology and Biogeochemistry, NIOZ Royal Netherlands Institute for Sea Research, 5Computational Science Research Center, San Diego State University, 6Department of Aerospace Engineering, San Diego State University, 7Department of Organismic and Evolutionary Biology, Harvard University Moored midwater geodesic structures called Coral Arks provide a modular, scalable, and vertically adjustable research platform that can be used to build, monitor, and perturb coral reef communities in previously inoperative areas, including offshore. Immunology and Infection RNA Fluorescence in situ Hybridization (FISH) to Visualize Microbial Colonization and Infection in Caenorhabditis elegans Intestines Dalaena E. Rivera1, Vladimir Lažetić2, Emily R. Troemel2, Robert J. Luallen1 1Department of Biology, San Diego State University, 2School of Biological Sciences, University of California, San Diego Intestinal microbes, including extracellular bacteria and intracellular pathogens like the Orsay virus and microsporidia (fungi), are often associated with wild Caenorhabditis nematodes. This article presents a protocol for detecting and quantifying microbes that colonize and/or infect C. elegans nematodes, and for measuring pathogen load after controlled infections in the lab. Immunology and Infection Selective Cleaning of Wild Caenorhabditis Nematodes to Enrich for Intestinal Microbiome Bacteria Emily Morgan1, Jonah Faye Longares1, Marie-Anne Félix2, Robert J. Luallen1 1Department of Biology, San Diego State University, 2 Wild Caenorhabditis nematodes are associated with many microbes, often in the gut lumen or infecting the intestine. This protocol details a method to enrich unculturable microbes colonizing the intestine, taking advantage of the resistance of the dauer cuticle. Biology Swabbing the Urban Environment - A Pipeline for Sampling and Detection of SARS-CoV-2 From Environmental Reservoirs Maria I. Rojas1,2, Steven S. Giles3, Mark Little1,2, Rafael Baron1,2, Isabella Livingston1,2, Taylor R. T. Dagenais3, Jason Baer1,2, Ana G. Cobián-Güemes1,2, Brandie White1,2, Forest Rohwer1,2 1Department of Biology, San Diego State University, 2Viral Information Institute, San Diego State University, 3Big Rose Web Design, LLC A citizen science project was designed to recruit San Diego residents to collect environmental samples for SARS-CoV-2. A multilingual web-based platform was created for data submission using a user-friendly mobile device interface. A laboratory information management system facilitated the collection of thousands of geographically diverse samples with real-time outcome tracking. Biology Simultaneous Isolation and Culture of Atrial Myocytes, Ventricular Myocytes, and Non-Myocytes from an Adult Mouse Heart Erik A. Blackwood1, Alina S. Bilal1, Khalid Azizi1, Anup Sarakki1, Christopher C. Glembotski1 1San Diego State University Heart Institute and the Department of Biology, San Diego State University A method is described for the simultaneous isolation of myocytes and non-myocytes from both the atria and ventricles of a single adult mouse heart. This protocol results in consistent yields of highly viable cardiac myocytes and non-myocytes and details optimal cell-specific culture conditions for phenotyping and in vitro analysis. Developmental Biology A Semi-high-throughput Imaging Method and Data Visualization Toolkit to Analyze C. elegans Embryonic Development Renat N. Khaliullin1,2,3, Jeffrey M. Hendel1,2, Adina Gerson-Gurwitz1,2, Shaohe Wang1,4,5, Stacy D. Ochoa1,6, Zhiling Zhao1,7, Arshad Desai1,2, Karen Oegema1,2, Rebecca A. Green1,2 1Ludwig Institute for Cancer Research, San Diego, 2Department of Cellular and Molecular Medicine, University of California, San Diego, 3Recursion Pharmaceuticals, 4Biomedical Sciences Graduate Program, University of California, San Diego, 5Cell Biology Section, National Institute of Dental and Craniofacial Research, National Institutes of Health, 6Department of Biology, San Diego State University, 7Developmental and Stem Cell Biology Graduate Program, University of California, San Francisco This work describes a semi-high-throughput protocol that allows simultaneous 3D time-lapse imaging of embryogenesis in 80–100 C. elegans embryos in a single overnight run. Additionally, image processing and visualization tools are included to streamline data analysis. The combination of these methods with custom reporter strains enables detailed monitoring of embryogenesis. Neuroscience Disruption of Frontal Lobe Neural Synchrony During Cognitive Control by Alcohol Intoxication Ksenija Marinkovic1,2, Lauren E. Beaton1, Burke Q. Rosen1,3, Joseph P. Happer4, Laura C. Wagner1 1Department of Psychology, San Diego State University, 2Department of Radiology, University of California San Diego, 3Department of Neurosciences, University of California San Diego, 4San Diego State University/UC San Diego Joint Doctoral Program in Clinical Psychology This experiment uses an anatomically-constrained magnetoencephalography (aMEG) method to examine brain oscillatory dynamics and long-range functional synchrony during engagement of cognitive control as a function of acute alcohol intoxication. Immunology and Infection A Murine Model of Group B Streptococcus Vaginal Colonization Kathryn A. Patras1, Kelly S. Doran1,2 1Department of Pediatrics, Division of Host-Microbe Systems & Therapeutics, University of California San Diego School of Medicine, 2Department of Biology and Center for Microbial Sciences, San Diego State University The purpose of this protocol is to imitate human group B Streptococcus (GBS) vaginal colonization in a murine model. This method may be used to investigate host immune responses and bacterial factors contributing to GBS vaginal persistence, as well as to test therapeutic strategies. Immunology and Infection Phage Phenomics: Physiological Approaches to Characterize Novel Viral Proteins Savannah E. Sanchez1, Daniel A. Cuevas2, Jason E. Rostron1, Tiffany Y. Liang3, Cullen G. Pivaroff1, Matthew R. Haynes1, Jim Nulton4, Ben Felts4, Barbara A. Bailey4, Peter Salamon4, Robert A. Edwards1,5,6, Alex B. Burgin7, Anca M. Segall1, Forest Rohwer1 1Department of Biology, San Diego State University, 2Computational Science Research Center, San Diego State University, 3Bioinformatics and Medical Informatics Research Center, San Diego State University, 4Department of Mathematics and Statistics, San Diego State University, 5Department of Computer Science, San Diego State University, 6Mathematics and Computer Science Division, Argonne National Laboratory, 7SPARC Committee, Broad Institute Here, we present phenomic approaches for the functional characterization of putative phage genes. Techniques include a developed assay capable of monitoring host anabolic metabolism, the Multi-phenotype Assay Plates (MAPs), in addition to the established method of metabolomics, capable of measuring effects to catabolic metabolism. Biology Genetic Barcoding with Fluorescent Proteins for Multiplexed Applications Cameron A. Smurthwaite1, Wesley Williams1, Alexandra Fetsko1, Darin Abbadessa1, Zachary D. Stolp1, Connor W. Reed1, Andre Dharmawan1, Roland Wolkowicz1 1Department of Biology, San Diego State University Since the discovery of the green fluorescent protein gene, fluorescent proteins have impacted molecular cell biology. This protocol describes how expression of distinct fluorescent proteins through genetic engineering is used for barcoding individual cells. The procedure enables tracking distinct populations in a cell mixture, which is ideal for multiplexed applications. Biology Purifying the Impure: Sequencing Metagenomes and Metatranscriptomes from Complex Animal-associated Samples Yan Wei Lim1, Matthew Haynes2, Mike Furlan1, Charles E. Robertson3, J. Kirk Harris4, Forest Rohwer1 1Department of Biology, San Diego State University, 2DOE Joint Genome Institute, 3Department of Molecular, Cellular and Developmental Biology, University of Colorado, 4Department of Pediatrics, School of Medicine, University of Colorado Using the cystic fibrosis airway as an example, the manuscript presents a comprehensive workflow comprising a combination of metagenomic and metatranscriptomic approaches to characterize the microbial and viral communities in animal-associated samples. Environment Unraveling the Unseen Players in the Ocean - A Field Guide to Water Chemistry and Marine Microbiology Andreas Florian Haas1, Ben Knowles1, Yan Wei Lim1, Tracey McDole Somera2, Linda Wegley Kelly1, Mark Hatay1, Forest Rohwer1 1Department of Biology, San Diego State University, 2Scripps Institution of Oceanography, University of California San Diego Here, we present a comprehensive protocol to assess the organic and inorganic nutrient availability and the abundance and structure of microbial and viral communities in remote marine environments. Biology Fluorescent Labeling of Drosophila Heart Structures Nakissa N. Alayari1,2, Georg Vogler2, Ouarda Taghli-Lamallem2, Karen Ocorr2, Rolf Bodmer2, Anthony Cammarato1,2 1Biology Department, San Diego State University, 2Development and Aging Program, NASCR Center, The Sanford Burnham Institute for Medical Research Here we describe a basic protocol for fluorescent labeling of different elements of heart tubes from larva and adult Drosophila melanogaster. These specimens are well-suited for imaging via fluorescent or confocal microscopy. This technique permits detailed structural analysis of the features of the hearts from a powerful model organism. Biology Semi-automated Optical Heartbeat Analysis of Small Hearts Karen Ocorr1, Martin Fink2, Anthony Cammarato1,3, Sanford I. Bernstein3, Rolf Bodmer1 1Development and Aging Program, The Sanford Burnham Institute for Medical Research, 2Cardiac Electrophysiology Group, Dept. of Physiology, Anatomy and Genetics, The Sanford Burnham Institute for Medical Research, 3Biology Department and Heart Institute, San Diego State University We have developed a Semi-automated Optical Heartbeat Analysis method (SOHA) for analyzing high speed optical recordings from Drosophila, zebrafish and embryonic mouse hearts. We demonstrate the application of our methodology to the analysis of heart function in fruit fly and embryonic mouse hearts.