title: Optical Imaging of Paramagnetic Bead-DNA Aggregation Inhibition Allows for Low Copy Number Detection of Infectious Pathogens.
abstract: DNA-paramagnetic silica bead aggregation in a rotating magnetic field facilitates the quantification of DNA with femtogram sensitivity, but yields no sequence-specific information. Here we provide an original description of aggregation inhibition for the detection of DNA and RNA in a sequence-specific manner following loop-mediated isothermal amplification (LAMP). The fragments generated via LAMP fail to induce chaotrope-mediated bead aggregation; however, due to their ability to passivate the bead surface, they effectively inhibit bead aggregation by longer 'trigger' DNA. We demonstrate the utility of aggregation inhibition as a method for the detection of bacterial and viral pathogens with sensitivity that approaches single copies of the target. We successfully use this methodology for the detection of notable food-borne pathogens Escherichia coli O157:H7 and Salmonella enterica, as well as Rift Valley fever virus, a weaponizable virus of national security concern. We also show the concentration dependence of aggregation inhibition, suggesting the potential for quantification of target nucleic acid in clinical and environmental samples. Lastly, we demonstrate the ability to rapidly detect infectious pathogens by utilizing a cell phone and custom-written application (App), making this novel detection modality fully portable for point-of-care use.
authors.affiliation: Department of Chemistry, University of Virginia, Charlottesville, VA, United States of America.
authors.forename: Jacquelyn A
authors.initials: JA
authors.lastname: DuVall
authors.affiliation: Department of Chemistry, University of Virginia, Charlottesville, VA, United States of America.
authors.forename: Juliane C
authors.initials: JC
authors.lastname: Borba
authors.affiliation: National Center for Biodefense and Infectious Diseases, George Mason University, Manassas, VA, United States of America.
authors.forename: Nazly
authors.initials: N
authors.lastname: Shafagati
authors.affiliation: Department of Microbiology, Immunology, and Cancer Biology, University of Virginia, Charlottesville, VA, United States of America.
authors.forename: Deborah
authors.initials: D
authors.lastname: Luzader
authors.affiliation: Department of Computer Science, University of Virginia, Charlottesville, VA, United States of America.
authors.forename: Nishant
authors.initials: N
authors.lastname: Shukla
authors.affiliation: Department of Chemistry, University of Virginia, Charlottesville, VA, United States of America.
authors.forename: Jingyi
authors.initials: J
authors.lastname: Li
authors.affiliation: National Center for Biodefense and Infectious Diseases, George Mason University, Manassas, VA, United States of America.
authors.forename: Kylene
authors.initials: K
authors.lastname: Kehn-Hall
authors.affiliation: Department of Microbiology, Immunology, and Cancer Biology, University of Virginia, Charlottesville, VA, United States of America.
authors.forename: Melissa M
authors.initials: MM
authors.lastname: Kendall
authors.affiliation: Center for Comparative Medicine, University of Virginia, Charlottesville, VA, United States of America.
authors.forename: Sanford H
authors.initials: SH
authors.lastname: Feldman
authors.affiliation: Department of Chemistry, University of Virginia, Charlottesville, VA, United States of America; Department of Mechanical Engineering, University of Virginia, Charlottesville, VA, United States of America; Department of Pathology, University of Virginia, Charlottesville, VA, United States of America.
authors.forename: James P
authors.initials: JP
authors.lastname: Landers
mesh_terms: D000818:Animals; D040421:Cell Phone; D004247:DNA / Q000097:blood* / Q000737:chemistry*; D017931:DNA Primers / Q000737:chemistry; D004927:Escherichia coli Infections / Q000175:diagnosis* / Q000382:microbiology; D019453:Escherichia coli O157 / Q000235:genetics / Q000302:isolation & purification; D006801:Humans; D008280:Magnetics; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D061848:Optical Imaging / Q000379:methods*; D019095:Point-of-Care Systems; D016133:Polymerase Chain Reaction; D012295:Rift Valley Fever / Q000175:diagnosis* / Q000821:virology; D012296:Rift Valley fever virus / Q000235:genetics / Q000302:isolation & purification; D019779:Salmonella enterica / Q000235:genetics* / Q000302:isolation & purification; D012822:Silicon Dioxide / Q000737:chemistry
chemical_list: D017931:DNA Primers; D012822:Silicon Dioxide; D004247:DNA