Reliability of Nucleic Acid Amplification Methods for Detection of Chlamydia trachomatis in Urine: Results of the First International Collaborative Quality Control Study among 96 Laboratories

Abstract

The first European Quality Control Concerted Action study was organized to assess the ability of laboratories to detect Chlamydia trachomatis in a panel of urine samples by nucleic acid amplification tests (NATs). The panel consisted of lyophilized urine samples, including three negative, two strongly positive, and five weakly positive samples. Ninety-six laboratories in 22 countries participated with a total of 102 data sets. Of 204 strongly positive samples 199 (97.5%) were correctly reported, and of 506 weakly positive samples 466 (92.1%) were correctly reported. In 74 (72.5%) data sets correct results were reported on all samples, and 17 data sets (16.7%) showed either one false-negative or one false-positive result. In another 11 data sets, two or more incorrect results were reported, and two data sets reported a false-positive result on one negative sample. The Roche COBAS Amplicor test was performed in 44 (43%) data sets, the Abbott LCx assay was performed in 31 (30%) data sets, the Roche Amplicor manual assay was performed in 9 (9%) data sets, an in-house PCR was performed in 9 (9%) data sets, the Becton Dickinson ProbeTec ET assay was performed in 5 (4.9%) data sets, and the GenProbe TMA assay was performed in 4 (3.9%) data sets. The results of the Roche Amplicor manual (95.6% correct), COBAS Amplicor (97.0%), and Abbott LCx (94.8%) tests were comparable (P ‫؍‬ 0.48). The results with the in-house PCR, BD ProbeTec ET, and GenProbe TMA tests were reported correctly in 88.6, 98, and 92.5% of the tests, respectively. Freeze-drying of clinical urine specimens proved to be a successful method for generating standardized, stable, and easy-to-transport samples for the detection of C. trachomatis by using NATs. Although the results, especially the specificity, for this proficiency panel were better than most quality control studies, sensitivity problems occurred frequently, underlining the need for good laboratory practice and reference reagents to monitor the performance of these assays.

External Quality Assessment for Detection of Chlamydia trachomatis

“…A further EQA study of detection of C. trachomatis in urine samples, based in Australia, found varying performance depending on the concentration (0 to 100%) and reported that low-level positives could not be detected consistently by a single test (18). In contrast to the results reported by Verkooyen et al (26), Land et al (18) found that at lower concentrations of C. trachomatis, the Roche AMPLICOR CT/NG assay was more sensitive than the Abbott LCx. However, Goessens et al (14) examined the detection of C. trachomatis in female specimens (urine and swabs) and found no difference among three commercial tests (Abbott LCx, GenProbe AMP-CT, and Roche COBAS AMPLICOR CT/NG).…”

“…Nonetheless, EQA provides valuable insight into the standard of clinical diagnostic testing by laboratories. In addition to monitoring standards, the process of EQA collates information that can be utilized to provide insight into the performance of specific laboratory methods (18,26). In this study, EQA enabled the comparison of C. trachomatis detection by differing assay types (EIA and NAAT) and also the comparison of the three main NAAT used by participants in the study (Abbott LCx, BDProbeTecET, and the Roche AMPLICOR/COBAS CT/NG method group), with two distinct sources of C. trachomatis.…”

“…Verkooyen et al (26) described the results obtained from an international EQA scheme consisting of freeze-dried urine specimens. The authors reported difficulty in the detection of C. trachomatis at lower concentrations and found no difference between the NAAT methods Roche AMPLICOR CT/NG, Roche COBAS AMPLICOR CT/NG, and Abbott LCx.…”

Yeast populations associated with Ghanaian cocoa fermentations analysed using denaturing gradient gel electrophoresis (DGGE)

“…Freeze-drying has previously been shown to be a suitable method for preserving DNA in various matrixes (Huckenbeck and Bonte, 1992;Verkooyen et al, 2003) and was thus chosen as the preservation method. Efficient lysis of the cells were ensured by treating the freeze-dried cocoa pulp enzymatically with protinase K and lysozyme, chemically with SDS, and mechanically by freeze-thaw cycles and bead beating.…”

Armored RNA as Virus Surrogate in a Real-Time Reverse Transcriptase PCR Assay Proficiency Panel

“…Key to the success of these laboratory networks is the use of standardized procedures and assays in all of the associated laboratories, which in turn is reliant on specific training programs as well as a demonstrated proficiency of laboratory workers to perform the assays in question. For molecular biology-based assays, evaluations of proficiency test practices have identified analytic errors associated with all stages of the testing process as well as errors specific to the physical setup of individual laboratories, emphasizing the need for on-site proficiency testing (2,3,11,15,16). There are, however, biosecurity risks associated with the distribution of live agents for training or proficiency test purposes, as documented by the inadvertent global distribution of a pandemic strain of influenza A/H2N2 virus in a public health laboratory proficiency panel during early 2005.…”