title: Monitoring biothreat agents (Francisella tularensis, Bacillus anthracis and Yersinia pestis) with a portable real-time PCR instrument.
abstract: In the event of suspected releases or natural outbreaks of contagious pathogens, rapid identification of the infectious agent is essential for appropriate medical intervention and disease containment. The purpose of this study was to compare the performance of a novel portable real-time PCR thermocycler, PikoReal™, to the standard real-time PCR thermocycler, Applied Biosystems® 7300 (ABI 7300), for the detection of three high-risk biothreat bacterial pathogens: Francisella tularensis, Bacillus anthracis and Yersinia pestis. In addition, a novel confirmatory real-time PCR assay for the detection of F. tularensis is presented and validated. The results show that sensitivity of the assays, based on a dilution series, for the three infectious agents ranged from 1 to 100 fg of target DNA with both instruments. No cross-reactivity was revealed in specificity testing. Duration of the assays with the PikoReal and ABI 7300 systems were 50 and 100 min, respectively. In field testing for F. tularensis, results were obtained with the PikoReal system in 95 min, as the pre-PCR preparation, including DNA extraction, required an additional 45 min. We conclude that the PikoReal system enables highly sensitive and rapid on-site detection of biothreat agents under field conditions, and may be a more efficient alternative to conventional diagnostic methods.
authors.affiliation: Centres for Military Medicine and for Biological Threat Preparedness, Helsinki, Finland. Electronic address: markos.molsa@mil.fi.
authors.forename: Markos
authors.initials: M
authors.lastname: Mölsä
authors.affiliation: Centres for Military Medicine and for Biological Threat Preparedness, Helsinki, Finland. Electronic address: heidi.hemmila@mil.fi.
authors.forename: Heidi
authors.initials: H
authors.lastname: Hemmilä
authors.affiliation: Centres for Military Medicine and for Biological Threat Preparedness, Helsinki, Finland. Electronic address: anna.katz@mil.fi.
authors.forename: Anna
authors.initials: A
authors.lastname: Katz
authors.affiliation: Natural Resources Institute Finland, Vantaa, Finland. Electronic address: jukka.niemimaa@luke.fi.
authors.forename: Jukka
authors.initials: J
authors.lastname: Niemimaa
authors.affiliation: Natural Resources Institute Finland, Suonenjoki, Finland. Electronic address: kristian.forbes@helsinki.fi.
authors.forename: Kristian M
authors.initials: KM
authors.lastname: Forbes
authors.affiliation: Natural Resources Institute Finland, Suonenjoki, Finland. Electronic address: otso.huitu@luke.fi.
authors.forename: Otso
authors.initials: O
authors.lastname: Huitu
authors.affiliation: Natural Resources Institute Finland, Suonenjoki, Finland. Electronic address: peterdstuart@hotmail.com.
authors.forename: Peter
authors.initials: P
authors.lastname: Stuart
authors.affiliation: Natural Resources Institute Finland, Vantaa, Finland. Electronic address: heikki.henttonen@luke.fi.
authors.forename: Heikki
authors.initials: H
authors.lastname: Henttonen
authors.affiliation: Centres for Military Medicine and for Biological Threat Preparedness, Helsinki, Finland. Electronic address: simo.nikkari@mil.fi.
authors.forename: Simo
authors.initials: S
authors.lastname: Nikkari
mesh_terms: D001408:Bacillus anthracis / Q000235:genetics / Q000302:isolation & purification*; D054045:Biological Warfare Agents; D004269:DNA, Bacterial / Q000235:genetics; D005604:Francisella tularensis / Q000235:genetics / Q000302:isolation & purification*; D063731:Mobile Applications; D060888:Real-Time Polymerase Chain Reaction / Q000295:instrumentation / Q000379:methods*; D015010:Yersinia pestis / Q000235:genetics / Q000302:isolation & purification*
chemical_list: D054045:Biological Warfare Agents; D004269:DNA, Bacterial
keywords: Biothreat; Francisella tularensis; PikoReal; Portable; Rapid identification; Real-time PCR