Improved costs and turnaround time for Treponema pallidum detection utilising a real-time PCR assay developed for the...

The aims of this study were to evaluate the performance and workflow characteristics of a laboratory-developed test to detect Treponema pallidum on the Hologic Panther Fusion system compared with an existing commercial assay. A Hologic Panther Fusion-based real-time polymerase chain reaction assay was optimised for T. pallidum (TP RT-PCR) using previously published primer and probe sequences and validated with a simplified preprocessing protocol. A total of 124 clinical and external quality assurance (EQA) samples were tested in parallel by the new method and the Viasure T. pallidum RT-PCR (Certest Biotech). Cross reactivity, limit of detection, PCR efficiency and coefficient of variation (CV) were measured. Simplified preprocessing methods involving either concentration or vortexing were compared with the existing overnight chemical/enzymatic preprocessing method for a subset of positive samples. Workflow impacts were assessed before and after implementation. The Panther Fusion T. pallidum assay correctly detected 65 of 65 (100%) positive clinical and EQA samples and 33 of 33 (100%) negative samples. No cross reactivity was observed for 45 of 45 (100%) samples. The limit of detection was 15 copies/reaction, and intrarun CV was <1.66%. The simplest preprocessing protocol detected 34 of 34 (100%) positive samples with an average Δ cycle threshold (Ct) <0.82 compared with the Viasure workflow. The new workflow reduced median turnaround time from 3.83 to 1.73 days (p<0.001) and estimated costs from US$35,004.96 to US$19,390.15 in the 3 months post-implementation. The laboratory-developed Panther Fusion T. pallidum assay is a sensitive and specific method for detection of syphilis and a cost-effective option to help improve workflow and turnaround time in a diagnostic laboratory.