title: Rapid detection and simultaneous antibiotic susceptibility analysis of Yersinia pestis directly from clinical specimens by use of reporter phage.
abstract: Yersinia pestis is a tier 1 agent due to its contagious pneumopathogenicity, extremely rapid progression, and high mortality rate. As the disease is usually fatal without appropriate therapy, rapid detection from clinical matrices is critical to patient outcomes. We previously engineered the diagnostic phage ΦA1122 with luxAB to create a "light-tagged" reporter phage. ΦA1122::luxAB rapidly detects Y. pestis in pure culture and human serum by transducing a bioluminescent signal response. In this report, we assessed the analytical specificity of the reporter phage and investigated diagnostic utility (detection and antibiotic susceptibility analysis) directly from spiked whole blood. The bioreporter displayed 100% (n = 59) inclusivity for Y. pestis and consistent intraspecific signal transduction levels. False positives were not obtained from species typically associated with bacteremia or those relevant to plague diagnosis. However, some non-pestis Yersinia strains and Enterobacteriaceae did elicit signals, albeit at highly attenuated transduction levels. Diagnostic performance was assayed in simple broth-enriched blood samples and standard aerobic culture bottles. In blood, <10(2) CFU/ml was detected within 5 h. In addition, Y. pestis was identified directly from positive blood cultures within 20 to 45 min without further processing. Importantly, coincubation of blood samples with antibiotics facilitated simultaneous antimicrobial susceptibility profiling. Consequently, the reporter phage demonstrated rapid detection and antibiotic susceptibility profiling directly from clinical samples, features that may improve patient prognosis during plague outbreaks.
authors.affiliation: Guild BioSciences, Charleston, South Carolina, USA.
authors.forename: J P
authors.initials: JP
authors.lastname: Vandamm
authors.affiliation: Department of Molecular Genetics and Microbiology, University of Florida, Gainesville, Florida, USA.
authors.forename: C
authors.initials: C
authors.lastname: Rajanna
authors.affiliation: Guild BioSciences, Charleston, South Carolina, USA.
authors.forename: N J
authors.initials: NJ
authors.lastname: Sharp
authors.affiliation: Molecular Biosciences, Institute for Cellular and Molecular Biology, University of Texas at Austin, Austin, Texas, USA.
authors.forename: I J
authors.initials: IJ
authors.lastname: Molineux
authors.affiliation: Guild BioSciences, Charleston, South Carolina, USA dschofield@guildbiosciences.com.
authors.forename: D A
authors.initials: DA
authors.lastname: Schofield
mesh_terms: D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D001431:Bacteriological Techniques / Q000379:methods*; D001435:Bacteriophages / Q000254:growth & development* / Q000302:isolation & purification*; D005189:False Positive Reactions; D017930:Genes, Reporter; D006801:Humans; D008156:Luciferases / Q000032:analysis / Q000235:genetics; D008163:Luminescent Measurements; D012680:Sensitivity and Specificity; D013997:Time Factors; D015010:Yersinia pestis / Q000187:drug effects* / Q000302:isolation & purification* / Q000821:virology
chemical_list: D000900:Anti-Bacterial Agents; D008156:Luciferases