title: Universal extraction method for gastrointestinal pathogens.
abstract: A universal stool extraction method for recovery of nucleic acids (NAs) from gastrointestinal pathogens was developed to support rapid diagnostics for the London 2012 Olympics. The method involved mechanical disruption (bead beating) of the stools, followed by automated extraction and detection using real-time PCR. This method had been used extensively in the Second Infectious Intestinal Disease Study (IID2) for the isolation of NA from bacteria and parasites (and was effective for the robust recovery of Cryptosporidium spp.) but had not been used for enteric viruses. To ensure this method was universally suitable, panels of samples known to contain target bacteria, viruses or parasites were processed in triplicate using the pre-treatment method routinely used for each target and the new extraction method (bead beating). The extracts were tested using real-time PCR and the cycle threshold values were compared. The results from this study showed that bead beating improved yields for the bacterial and parasitic targets and was suitable for the viral targets. The implementation of this universal method should confer cost- and time-saving benefits and streamline the processes required for the characterization of an array of pathogens from faecal samples.
authors.affiliation: PHE Microbiology Services, Colindale, London, UK.|Royal Free London NHS Trust, London, UK.
authors.forename: Fenella D
authors.initials: FD
authors.lastname: Halstead
authors.affiliation: Burnet Institute, Melbourne, Australia.|PHE Public Health Laboratory London, Barts Health NHS Trust, London, UK.
authors.forename: Adele V
authors.initials: AV
authors.lastname: Lee
authors.affiliation: PHE Public Health Laboratory London, Barts Health NHS Trust, London, UK.
authors.forename: Xose
authors.initials: X
authors.lastname: Couto-Parada
authors.affiliation: University College London Hospitals NHS Trust, London, UK.
authors.forename: Spencer D
authors.initials: SD
authors.lastname: Polley
authors.affiliation: PHE Public Health Laboratory London, Barts Health NHS Trust, London, UK.
authors.forename: Clare
authors.initials: C
authors.lastname: Ling
authors.affiliation: PHE Microbiology Services, Colindale, London, UK.
authors.forename: Claire
authors.initials: C
authors.lastname: Jenkins
authors.affiliation: Cryptosporidium Reference Laboratory, Public Health Wales Microbiology, Swansea, UK.
authors.forename: Rachel M
authors.initials: RM
authors.lastname: Chalmers
authors.affiliation: Cryptosporidium Reference Laboratory, Public Health Wales Microbiology, Swansea, UK.
authors.forename: Kristin
authors.initials: K
authors.lastname: Elwin
authors.affiliation: Norfolk and Norwich University Hospitals NHS Trust, UK.|PHE Microbiology Services, Colindale, London, UK.
authors.forename: Jim J
authors.initials: JJ
authors.lastname: Gray
authors.affiliation: PHE Microbiology Services, Colindale, London, UK.
authors.forename: Miren
authors.initials: M
authors.lastname: Iturriza-Gómara
authors.affiliation: PHE Microbiology Services, Colindale, London, UK.
authors.forename: John
authors.initials: J
authors.lastname: Wain
authors.affiliation: PHE Public Health Laboratory London, Barts Health NHS Trust, London, UK.
authors.forename: Duncan A
authors.initials: DA
authors.lastname: Clark
authors.affiliation: PHE Public Health Laboratory, Manchester, UK.
authors.forename: Frederick J
authors.initials: FJ
authors.lastname: Bolton
authors.affiliation: PHE Public Health Laboratory London, Barts Health NHS Trust, London, UK.
authors.forename: Rohini J
authors.initials: RJ
authors.lastname: Manuel
authors.lastname: The Olympics Gi Group
mesh_terms: D001424:Bacterial Infections / Q000175:diagnosis; D004247:DNA / Q000235:genetics / Q000302:isolation & purification*; D005243:Feces / Q000382:microbiology* / Q000469:parasitology*; D005759:Gastroenteritis / Q000209:etiology*; D006801:Humans; D007411:Intestinal Diseases, Parasitic / Q000175:diagnosis; D008131:London; D025202:Molecular Diagnostic Techniques / Q000191:economics / Q000379:methods*; D060888:Real-Time Polymerase Chain Reaction / Q000191:economics / Q000379:methods*; D013048:Specimen Handling / Q000191:economics / Q000379:methods*; D013997:Time Factors
chemical_list: D004247:DNA