title: Development, validation and testing costs of an in-house real-time PCR assay for the detection of Chlamydia trachomatis.
abstract: OBJECTIVE
  To improve the screening of Chlamydia trachomatis(C. trachomatis) in Brazil, an accurate and affordable method is needed. The objective of this study was to develop and assess the performance and costs of a new in-house real-time PCR (qPCR) assay for the diagnosis of C. trachomatis infection.
  
  
  METHODS
  Asymptomatic women aged 14-25 years who attended primary health services in Manaus, Brazil, were screened for C. trachomatis using the Digene Hybrid Capture II CT-ID (HCII CT-ID) DNA test. A subset of cervical specimens were tested using an in-house qPCR and a commercial qPCR, ArtusC. trachomatis Plus RG PCR 96 CE (Artus qPCR) kit, as a reference test. A primer/probe based on the sequence of cryptic plasmid (CP) was designed. An economic evaluation was conducted from the provider's perspective.
  
  
  RESULTS
  The primers were considered specific for C. trachomatis because they did not amplify any product from non-sexually transmitted bacterial species tested. Overall, 292 specimens were tested by both the commercial kit (Artus qPCR) and the in-house qPCR. Of those, one resulted in no amplification and was excluded from the analysis. The sensitivity, specificity, and positive and negative predictive values of the in-house qPCR were 99.5 % [95 % confidence interval (CI): 97.1-100], 95.1 % (95 % CI: 89-98.4), 97.4 % (95 % CI: 94-99.1) and 99.0 % (95 % CI: 94.5-100), respectively. The cost per case of C. trachomatis was £0.44 ($0.55) for HCII CT-ID, £1.16 ($1.45) for Artus qPCR and £1.06 ($1.33) for in-house qPCR.
  
  
  CONCLUSIONS
  We have standardized an in-house qPCR to detect cervical C. trachomatis targeting CP. The in-house qPCR showed excellent accuracy and was more affordable than the commercial qPCR kit.
authors.affiliation: Programa de Pós-graduação em Medicina Tropical, Universidade do Estado do Amazonas, Manaus, Amazonas, Brazil.|Fundação Alfredo da Matta, Manaus, Amazonas, Brazil.
authors.forename: Camila Gurgel Dos
authors.initials: CGD
authors.lastname: Santos
authors.affiliation: Fundação de Medicina Tropical Doutor Heitor Vieira Dourado (FMT-HVD), Manaus, Amazonas, Brazil.|TransLab, Departament of Medical Sciences, Universitat de Girona, Catalunya, Spain.|CIBER de Epidemiologia y Salud Pública, CIBERESP, Spain.
authors.forename: Meritxell
authors.initials: M
authors.lastname: Sabidó
authors.affiliation: Fundação Alfredo da Matta, Manaus, Amazonas, Brazil.
authors.forename: André Luiz
authors.initials: AL
authors.lastname: Leturiondo
authors.affiliation: Fundação Alfredo da Matta, Manaus, Amazonas, Brazil.
authors.forename: Cynthia de Oliveira
authors.initials: CO
authors.lastname: Ferreira
authors.affiliation: Fundação Alfredo da Matta, Manaus, Amazonas, Brazil.
authors.forename: Thielle Pereira
authors.initials: TP
authors.lastname: da Cruz
authors.affiliation: Fundação Alfredo da Matta, Manaus, Amazonas, Brazil.|Fundação de Medicina Tropical Doutor Heitor Vieira Dourado (FMT-HVD), Manaus, Amazonas, Brazil.|Departamento de IST, Aids e Hepatites Virais, Secretaria de Vigilância em Saúde, Ministério da Saúde, Brasília, Brazil.
authors.forename: Adele Schwartz
authors.initials: AS
authors.lastname: Benzaken
mesh_terms: D000293:Adolescent; D000328:Adult; D058345:Asymptomatic Infections; D001938:Brazil; D002584:Cervix Uteri / Q000382:microbiology; D002690:Chlamydia Infections / Q000175:diagnosis* / Q000382:microbiology; D002692:Chlamydia trachomatis / Q000235:genetics* / Q000302:isolation & purification; D017931:DNA Primers; D004269:DNA, Bacterial / Q000235:genetics; D005260:Female; D006801:Humans; D010957:Plasmids; D011933:Reagent Kits, Diagnostic; D060888:Real-Time Polymerase Chain Reaction / Q000191:economics* / Q000379:methods* / Q000592:standards; D012680:Sensitivity and Specificity; D055815:Young Adult
chemical_list: D017931:DNA Primers; D004269:DNA, Bacterial; D011933:Reagent Kits, Diagnostic