title: Performance of the GeneXpert Ebola Assay for Diagnosis of Ebola Virus Disease in Sierra Leone: A Field Evaluation Study.
abstract: BACKGROUND
  Throughout the Ebola virus disease (EVD) epidemic in West Africa, field laboratory testing for EVD has relied on complex, multi-step real-time reverse transcription PCR (RT-PCR) assays; an accurate sample-to-answer RT-PCR test would reduce time to results and potentially increase access to testing. We evaluated the performance of the Cepheid GeneXpert Ebola assay on clinical venipuncture whole blood (WB) and buccal swab (BS) specimens submitted to a field biocontainment laboratory in Sierra Leone for routine EVD testing by RT-PCR ("Trombley assay").
  
  
  RESULTS
  This study was conducted in the Public Health England EVD diagnostic laboratory in Port Loko, Sierra Leone, using residual diagnostic specimens remaining after clinical testing. EDTA-WB specimens (n = 218) were collected from suspected or confirmed EVD patients between April 1 and July 20, 2015. BS specimens (n = 71) were collected as part of a national postmortem screening program between March 7 and July 20, 2015. EDTA-WB and BS specimens were tested with Xpert (targets: glycoprotein [GP] and nucleoprotein [NP] genes) and Trombley (target: NP gene) assays in parallel. All WB specimens were fresh; 84/218 were tested in duplicate on Xpert to compare WB sampling methods (pipette versus swab); 43/71 BS specimens had been previously frozen. In all, 7/218 (3.2%) WB and 7/71 (9.9%) BS samples had Xpert results that were reported as "invalid" or "error" and were excluded, leaving 211 WB and 64 BS samples with valid Trombley and Xpert results. For WB, 22/22 Trombley-positive samples were Xpert-positive (sensitivity 100%, 95% CI 84.6%-100%), and 181/189 Trombley-negative samples were Xpert-negative (specificity 95.8%, 95% confidence interval (CI) 91.8%-98.2%). Seven of the eight Trombley-negative, Xpert-positive (Xpert cycle threshold [Ct] range 37.7-43.4) WB samples were confirmed to be follow-up submissions from previously Trombley-positive EVD patients, suggesting a revised Xpert specificity of 99.5% (95% CI 97.0%-100%). For Xpert-positive WB samples (n = 22), Xpert NP Ct values were consistently lower than GP Ct values (mean difference -4.06, 95% limits of agreement -6.09, -2.03); Trombley (NP) Ct values closely matched Xpert NP Ct values (mean difference -0.04, 95% limits of agreement -2.93, 2.84). Xpert results (positive/negative) for WB sampled by pipette versus swab were concordant for 78/79 (98.7%) WB samples, with comparable Ct values for positive results. For BS specimens, 20/20 Trombley-positive samples were Xpert-positive (sensitivity 100%, 95% CI 83.2%-100%), and 44/44 Trombley-negative samples were Xpert-negative (specificity 100%, 95% CI 92.0%-100%). This study was limited to testing residual diagnostic samples, some of which had been frozen before use; it was not possible to test the performance of the Xpert Ebola assay at point of care.
  
  
  CONCLUSIONS
  The Xpert Ebola assay had excellent performance compared to an established RT-PCR benchmark on WB and BS samples in a field laboratory setting. Future studies should evaluate feasibility and performance outside of a biocontainment laboratory setting to facilitate expanded access to testing.
authors.affiliation: Public Health England, Porton Down, United Kingdom.
authors.forename: Amanda E
authors.initials: AE
authors.lastname: Semper
authors.affiliation: Partners In Health, Boston, Massachusetts, United States of America.
authors.forename: M Jana
authors.initials: MJ
authors.lastname: Broadhurst
authors.affiliation: Public Health England Laboratory, Port Loko, Sierra Leone.|Mid Essex Hospital Services NHS Trust, Chelmsford, United Kingdom.
authors.forename: Jade
authors.initials: J
authors.lastname: Richards
authors.affiliation: Public Health England Laboratory, Port Loko, Sierra Leone.|Liverpool School of Tropical Medicine, Liverpool, United Kingdom.
authors.forename: Geraldine M
authors.initials: GM
authors.lastname: Foster
authors.affiliation: Public Health England, Porton Down, United Kingdom.|Public Health England Laboratory, Port Loko, Sierra Leone.
authors.forename: Andrew J H
authors.initials: AJ
authors.lastname: Simpson
authors.affiliation: Public Health England, Porton Down, United Kingdom.
authors.forename: Christopher H
authors.initials: CH
authors.lastname: Logue
authors.affiliation: Wellbody Alliance, Freetown, Sierra Leone.
authors.forename: J Daniel
authors.initials: JD
authors.lastname: Kelly
authors.affiliation: Partners In Health, Boston, Massachusetts, United States of America.|Harvard Medical School, Boston, Massachusetts, United States of America.
authors.forename: Ann
authors.initials: A
authors.lastname: Miller
authors.affiliation: Public Health England, Porton Down, United Kingdom.|Public Health England Laboratory, Port Loko, Sierra Leone.
authors.forename: Tim J G
authors.initials: TJ
authors.lastname: Brooks
authors.affiliation: Partners In Health, Boston, Massachusetts, United States of America.|Harvard Medical School, Boston, Massachusetts, United States of America.
authors.forename: Megan
authors.initials: M
authors.lastname: Murray
authors.affiliation: Partners In Health, Boston, Massachusetts, United States of America.|Department of Laboratory Medicine, Boston Children's Hospital, Boston, Massachusetts, United States of America.
authors.forename: Nira R
authors.initials: NR
authors.lastname: Pollock
mesh_terms: D000293:Adolescent; D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D002648:Child; D002675:Child, Preschool; D029043:Ebolavirus / Q000235:genetics*; D005260:Female; D006023:Glycoproteins / Q000235:genetics*; D019142:Hemorrhagic Fever, Ebola / Q000097:blood / Q000175:diagnosis*; D006801:Humans; D007223:Infant; D008297:Male; D008875:Middle Aged; D009698:Nucleoproteins / Q000235:genetics*; D019095:Point-of-Care Systems; D012367:RNA, Viral / Q000097:blood*; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D012807:Sierra Leone; D055815:Young Adult
chemical_list: D006023:Glycoproteins; D009698:Nucleoproteins; D012367:RNA, Viral