The Verification of Nucleic Acid Amplification Testing (Gen-Probe Aptima Assay) for Chlamydia trachomatis from Ocular Samples

• Published in: Ophthalmology (Rochester, Minn.) Vol. 122; no. 2; pp. 244 - 247
• Main Authors: Kowalski, Regis P., MS, M(ASCP), Karenchak, Lisa M., BS, M(ASCP), Raju, Leela V., MD, Ismail, Nahed, MD, PhD
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.02.2015
• Subjects: Chlamydia Infections - diagnosis; Chlamydia Infections - microbiology; Chlamydia trachomatis - genetics; Chlamydia trachomatis - isolation & purification; Conjunctiva - microbiology; Conjunctivitis, Inclusion - diagnosis; Conjunctivitis, Inclusion - microbiology; DNA, Bacterial - genetics; Eye Infections, Bacterial - diagnosis; Eye Infections, Bacterial - microbiology; False Positive Reactions; Humans; Nucleic Acid Amplification Techniques - methods; Ophthalmology; Polymerase Chain Reaction; Predictive Value of Tests; Retrospective Studies; RNA, Ribosomal - genetics; Sensitivity and Specificity; NAAT; ribosomal RNA; rRNA; Bartels Chlam Trans Transport Medium; RLU; CTM; nucleic acid amplification testing; PCR; polymerase chain reaction; relative luminescence
• ISSN: 0161-6420; 1549-4713
• DOI: 10.1016/j.ophtha.2014.08.038

Objective Chlamydia trachomatis conjunctivitis may present with extended symptoms, and it can have social ramifications as a sexually transmitted disease. For appropriate therapy, C. trachomatis conjunctivitis should be diagnosed definitively. This study presents the verification of nucleic acid amplification testing (NAAT; Gen-Probe Aptima Combo 2 assay) for detection of C. trachomatis ribosomal RNA (rRNA) from direct ocular samples. Design Retrospective laboratory verification study. Subjects Patients with infectious conjunctivitis. Methods A battery of 25 true-positive specimens (direct ocular specimens from patients with symptoms consistent with C. trachomatis conjunctivitis and with previously demonstrated positive polymerase chain reaction [PCR] results for C. trachomatis DNA by Roche Amplicor) and 25 true-negative specimens (direct ocular specimens with culture-positive results for herpes simplex virus [n = 5], adenovirus [n = 5], Haemophilus influenzae [n = 5], and Streptococcus pneumoniae [n = 5]), and transport medium (n = 5) were tested for C. trachomatis rRNA by NAAT. These true-negative specimens have differential etiologic agents of infectious conjunctivitis. The 25 C. trachomatis specimens with PCR-positive results (obtained May 1994–May 2012) and 20 true-negative infectious ocular specimens (obtained December 2008–August 2013) were collected with soft-tipped applicators and placed in transport medium. All excess specimens were stored at −80°C. All samples were centrifuged at 13,000 rpm for 1 hour at 6°C. For each sample, using the Aptima Unisex collection blue swab, a specimen was collected from the conical apex of the storage tube where a pellet was formed. The Aptima Unisex collection swab was placed in a tube of Aptima swab transport medium for testing. All samples were tested in duplicate. Main Outcome Measures Detection of C. trachomatis rRNA. Results Of 25 true-positive samples, 24 (96%) were positive by NAAT, whereas 25 of 25 true-negative samples (100%) showed negative results. The sensitivity, specificity, positive predictive value, negative predictive value, and efficiency were determined to be 96%, 100%, 100%, 96%, and 98%, respectively. Conclusions The detection of C. trachomatis in ocular specimens by NAAT was verified for laboratory diagnosis. The test will be evaluated prospectively to determine future test performance precisely.