Viability-PCR Shows That NAAT Detects a High Proportion of DNA from Non-Viable Chlamydia trachomatis

被引:43
作者
Janssen, Kevin J. H. [1 ]
Hoebe, Christian J. P. A. [1 ,2 ]
Dukers-Muijrers, Nicole H. T. M. [1 ,2 ]
Eppings, Lisanne [1 ,2 ]
Lucchesi, Mayk [1 ]
Wolffs, Petra F. G. [1 ]
机构
[1] Maastricht Univ, Med Ctr MUMC, Sch Publ Hlth & Primary Care CAPHRI, Dept Med Microbiol, Maastricht, Netherlands
[2] South Limburg Publ Hlth Serv GGD South Limburg, Dept Sexual Hlth Infect Dis & Environm Hlth, Geleen, Netherlands
关键词
POLYMERASE-CHAIN-REACTION; PROPIDIUM MONOAZIDE; GENITAL-INFECTION; PERFORMANCE; TIME; DIAGNOSIS; ASSAYS;
D O I
10.1371/journal.pone.0165920
中图分类号
O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];
学科分类号
07 ; 0710 ; 09 ;
摘要
Objectives According to the current guidelines for laboratory diagnosis of sexually transmitted infections (STIs), nucleic acid amplification tests (NAATs) are the preferred diagnostic method for Chlamydia trachomatis (CT) infections. However, NAATs amplify the available target DNA without discriminating between DNA originating from viable or non-viable CT. Assessing CT viability will provide more insights in the clinical and public health relevance of a CT positive test result. The aim of this study was to technically validate and implement viability-PCR (V-PCR) to asses CT viability. Methods Technical validation of V-PCR was performed by the assessment of predefined viability ratios of CT. Samples were subjected to V-PCR which consisted of propidium monoazide (PMA) treatment prior to DNA extraction followed by quantitative PCR (qPCR) targeting the ompA gene for the detection of CT DNA. Finally, V-PCR was applied to vaginal swabs of 50 CT positive patients, as indicated by routine NAAT, collected at our outpatient STD clinics before antimicrobial treatment. Results Technical validation of V-PCR showed that PMA treatment of heat-inactivated CT culture resulted in an almost complete loss of qPCR signal. PMA treated samples of the fresh viable CT culture showed no marked reduction of PCR signal, indicating that all DNA from viable CT could be detected. Applying V-PCR to clinical samples showed that in 36% of samples (18/50) less than 1% of CT DNA originated from viable bacteria. Conclusions V-PCR showed to be a fast and easy method to assess CT viability in clinical samples, without the need of traditional challenging cell culture methods. Furthermore, V-PCR results of clinical samples have indicated that a substantial amount of the amplified CT DNA originated from non-viable cells. Although results might be influenced by cell death during transport, this study suggests that there is a potential overestimation of quantitative CT positivity by currently used NAATs.
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共 30 条
[21]   2015 UK national guideline for the management of infection with Chlamydia trachomatis [J].
Nwokolo, Nneka C. ;
Dragovic, Bojana ;
Patel, Sheel ;
Tong, C. Y. William ;
Barker, Gary ;
Radcliffe, Keith .
INTERNATIONAL JOURNAL OF STD & AIDS, 2016, 27 (04) :251-267
[22]   Time to clearance of Chlamydia trachomatis ribosomal RNA in women treated for chlamydial infection [J].
Renault, Cybele A. ;
Israelski, Dennis M. ;
Levy, Vivian ;
Fujikawa, Bruce K. ;
Kellogg, Timothy A. ;
Klausner, Jeffrey D. .
SEXUAL HEALTH, 2011, 8 (01) :69-73
[23]   Opportunities and pitfalls of molecular testing for detecting sexually transmitted pathogens [J].
Trembizki, Ella ;
Costa, Anna-Maria G. ;
Tabrizi, Sepehr N. ;
Whiley, David M. ;
Twin, Jimmy .
PATHOLOGY, 2015, 47 (03) :219-226
[24]   Performance of the cobas CT/NG Test Compared to the Aptima AC2 and Viper CTQ/GCQ Assays for Detection of Chlamydia trachomatis and Neisseria gonorrhoeae [J].
Van Der Pol, Barbara ;
Liesenfeld, Oliver ;
Williams, James A. ;
Taylor, Stephanie N. ;
Lillis, Rebecca A. ;
Body, Barbara A. ;
Nye, Mindy ;
Eisenhut, Carol ;
Hook, Edward W., III .
JOURNAL OF CLINICAL MICROBIOLOGY, 2012, 50 (07) :2244-2249
[25]   Alarmingly poor performance in Chlamydia trachomatis point-of-care testing [J].
van Dommelen, Laura ;
van Tiel, Frank H. ;
Ouburg, Sander ;
Brouwers, Elfi E. H. G. ;
Terporten, Peter H. W. ;
Savelkoul, Paul H. M. ;
Morre, Servaas A. ;
Bruggeman, Cathrien A. ;
Hoebe, Christian J. P. A. .
SEXUALLY TRANSMITTED INFECTIONS, 2010, 86 (05) :355-359
[26]   Quantitative real-time PCR analysis of total and propidium monoazide-resistant fecal indicator bacteria in wastewater [J].
Varma, M. ;
Field, R. ;
Stinson, M. ;
Rukovets, B. ;
Wymer, L. ;
Hauglanda, R. .
WATER RESEARCH, 2009, 43 (19) :4790-4801
[27]   The Epidemiology of Chlamydia trachomatis Organism Load During Genital Infection: A Systematic Review [J].
Vodstrcil, Lenka A. ;
McIver, Ruthy ;
Huston, Wilhelmina M. ;
Tabrizi, Sepehr N. ;
Timms, Peter ;
Hocking, Jane S. .
JOURNAL OF INFECTIOUS DISEASES, 2015, 211 (10) :1628-1645
[28]   CHLAMYDIA-TRACHOMATIS INFECTION IN A HIGH-RISK POPULATION - COMPARISON OF POLYMERASE CHAIN-REACTION AND CELL-CULTURE FOR DIAGNOSIS AND FOLLOW-UP [J].
VOGELS, WHM ;
VADER, PCV ;
SCHRODER, FP .
JOURNAL OF CLINICAL MICROBIOLOGY, 1993, 31 (05) :1103-1107
[29]   Duration of Polymerase Chain Reaction-Detectable DNA After Treatment of Chlamydia trachomatis, Neisseria gonorrhoeae, and Trichomonas vaginalis Infections in Women [J].
Williams, James A. ;
Ofner, Susan ;
Batteiger, Byron E. ;
Fortenberry, J. Dennis ;
Van Der Pol, Barbara .
SEXUALLY TRANSMITTED DISEASES, 2014, 41 (03) :215-219
[30]   LONG-TERM ERADICATION OF CHLAMYDIA-TRACHOMATIS GENITAL-INFECTION AFTER ANTIMICROBIAL THERAPY - EVIDENCE AGAINST PERSISTENT INFECTION [J].
WORKOWSKI, KA ;
LAMPE, MF ;
WONG, KG ;
WATTS, MB ;
STAMM, WE .
JAMA-JOURNAL OF THE AMERICAN MEDICAL ASSOCIATION, 1993, 270 (17) :2071-2075