Trachomatis Assay Research Articles

The Finnish New Variant of Chlamydia trachomatis with a Single Nucleotide Polymorphism in the 23S rRNA Target Escapes Detection by the Aptima Combo 2 Test.

In 2019, more than 200 cases of Chlamydia trachomatis negative/equivocal by the Aptima Combo 2 assay (AC2, target: 23S rRNA) with slightly elevated relative light units (RLUs), but positive by the Aptima Chlamydia trachomatis assay (ACT, target: 16S rRNA) have been detected in Finland To identify the cause of the AC2 CT false-negative specimens, we sequenced parts of the CT 23S rRNA gene in 40 specimens that were AC2 negative/equivocal but ACT positive. A single nucleotide polymorphism (SNP; C1515T in the C. trachomatis 23S rRNA gene) was revealed in 39 AC2/ACT discordant specimens. No decrease in the number of mandatorily notified C. trachomatis cases was observed nationally in Finland in 2010–2019. When RLUs obtained for AC2 negative specimens were retrospectively evaluated in 2011–2019, a continuous increase in the proportion of samples with RLUs 10–19 was observed since 2014, and a slight increase in the proportion of samples with RLUs 20–84 in 2017–2019, indicating that the Finnish new variant of C. trachomatis might have been spreading nationally for several years. This emphasizes that careful surveillance of epidemiology, positivity rate and test performance are mandatory to detect any changes affecting detection of infections.

Finnish new variant of Chlamydia trachomatis escaping detection in the Aptima Combo 2 assay also present in Örebro County, Sweden, May 2019.

We identified the first two cases of the Finnish new variant of Chlamydia trachomatis (F-nvCT) beyond Finland in two clinical urogenital specimens in Örebro County, Sweden. These Aptima Combo 2 assay-negative specimens were Aptima Chlamydia trachomatis (CT) assay positive and had the characteristic C1515T mutation in the 23S rRNA gene. From 22 March to 31 May 2019, 1.3% (2/158) of the CT-positive cases in Örebro County were missed because of the F-nvCT. International awareness, investigations and actions are essential.

Contaminated fingers: a potential cause of Chlamydia trachomatis-positive urine specimens

ObjectivesThe detection of an STI agent in a urogenital tract (UGT) specimen from a young child is regarded as being indicative of sexual abuse. However, the probabilities of contamination events...

Field Evaluation of the Cepheid GeneXpert Chlamydia trachomatis Assay for Detection of Infection in a Trachoma Endemic Community in Tanzania

PurposeTo determine the sensitivity, specificity, and field utility of the Cepheid GeneXpert Chlamydia trachomatis (CT) Assay (GeneXpert) for ocular chlamydia infection compared to Roche Amplicor CT assay (Amplicor).MethodsIn a trachoma-endemic community in Kongwa Tanzania, 144 children ages 0 to 9 were surveyed to assess clinical trachoma and had two ocular swabs taken. One swab was processed at Johns Hopkins University, Baltimore MD, using Amplicor, (Roche Molecular Diagnostics) and the other swab was processed at a field station in Kongwa using the GeneXpert Chlamydia trachomatis/Neisseria gonorrhoeae assay (Cepheid). The sensitivity and specificity of GeneXpert was compared to the Amplicor assay.ResultsOf the 144 swabs taken the prevalence of follicular trachoma by clinical exam was 43.7%, and by evidence of infection according to Amplicor was 28.5%. A total of 17 specimens (11.8%) could not be processed by GeneXpert in the field due to lack of sample volume, other specimen issues or electricity failure. The sensitivity of GeneXpert when compared to Amplicor was 100% and the specificity was 95%. The GeneXpert test identified more positives in individuals with clinical trachoma than Amplicor, 55% versus 52%.ConclusionThe GeneXpert test for C. trachomatis performed with high sensitivity and specificity and demonstrated excellent promise as a field test for trachoma control.

Rapid homogeneous PCR assay for the detection of Chlamydia trachomatis in urine samples

Chlamydia trachomatis infection is the most common bacterial sexually transmitted disease and a major public health problem worldwide. Fast and sensitive point-of-care diagnostics including non-invasive sample collection would be of value for the prevention of C. trachomatis transmission. The aim of this study was to develop a fast, reliable, non-invasive and easy-to-use homogenous PCR assay for the detection of C. trachomatis. Bacteria were concentrated from urine by a simple and fast centrifugation-based urine pretreatment method. Novel automated GenomEra technology was utilized for the rapid closed-tube PCR including time-resolved fluorometric detection of the target using lanthanide chelate labeled probes. We have developed a rapid C. trachomatis assay which provides qualitative results in 1h with diagnostic sensitivity and specificity of 98.7% and 97.3%, respectively. The novel assay can be performed with minimal laboratory expertise and without sophisticated DNA-extraction devices and has performance comparable to current gold standard assays.

Pilot study of the utility and acceptability of tampon sampling for the diagnosis of Neisseria gonorrhoeae and Chlamydia trachomatis infections by duplex realtime polymerase chain reaction in United Kingdom sex workers

We aimed to evaluate the acceptability of self-collected tampon samples for the screening of female sex workers for sexually transmitted infections. We recruited 65 sex workers, and 63 agreed to provide tampon samples. The tampon samples were processed by realtime polymerase chain reaction (PCR) targeting Neisseria gonorrhoeae and Chlamydia trachomatis. Urethral and endocervical swabs were also obtained from 61 of 63 participants and tested using culture (N. gonorrhoeae) and the BD ProbeTec strand displacement amplification (SDA) (C. trachomatis) assay. Tampon sampling was preferred by 95% of the women and all favoured being tested away from genitourinary medicine clinics; the most common reasons cited were avoidance of embarrassment (40%) and convenience (30%). Besides near-universal acceptability of tampon sampling, the tampon sampling-PCR approach described in this study appeared to have enhanced sensitivity compared with conventional testing, suggesting the possibility of a residual hidden burden of N. gonorrhoeae and/or C. trachomatis genital infections in UK female sex workers.

Rapid Hybridization Probe Assay and PCR for Detection of Chlamydia trachomatis in Urinary Tract Infections: A Prospective Study

Chlamydia trachomatis is a widespread bacterium that causes trachoma and genital tract infections in humans. The fact that the growth of this pathogen does not normally occur outside living cells poses a challenge in its diagnosis. The present study aimed to compare the efficacies of different molecular and cultural methods in the detection of C. trachomatis in urine samples collected from patients with urinary tract infections. Examined detection methods involved the Gen-Probe C. trachomatis (GP-CT) assay, direct antigen detection by enzyme-linked immunosorbent assay (ELISA), and polymerase chain reaction (PCR) method. The efficacies of these methods were compared to that of the cell culture technique depending on sensitivity, specificity, and accuracy. C. trachomatis was detected in 25 out of 50 (50%) of examined urine samples using the cell culture method. Compared with this standard technique, the GP-CT assay was the most sensitive procedure, being able to detect the pathogen in all positive samples, followed by PCR and ELISA, which showed 60% and 40% sensitivities, respectively. PCR and ELISA displayed the highest level of specificity (100%) compared to the cell culture method with the GP-CT assay showing 40% specificity. The rate of accuracy was comparable between the GP-CT, PCR, and ELISA methods ranging from 70-80% of the accuracy of the cell culture method. The above results suggest that C. trachomatis is a frequent pathogen associated with upper and lower urinary tract infections. Both the GP-CT assay and PCR method can be recommended as reliable detection methods for C. trachomatis, and the GP-CT can be used as a screening tool.

Screening of Chlamydia trachomatis urogenital infections among the male and female population of the Republic of Macedonia

Noninvasive urine screening for Chlamydia trachomatis infections offers a valuable public health tool, that could be of vast importance in Chlamydia control programs. The goal was to determine the prevalence of C. trachomatis infections among a sexually active population, to define the epidemiological factors associated with it, and to develop potential selective screening strategies among asymptomatic individuals in the Republic of Macedonia, using a highly sensitive and specific DNA amplification method for C. trachomatis. A total of 1435 urine samples, divided into two main groups: asymptomatic individuals (n = 1210) and symptomatic patients (n = 225), were tested. Samples from the asymptomatic group were collected during routine screening programs, while the symptomatic group consisted of patients with symptoms of urogenital tract infection, attending sexually transmitted diseases (STD) clinics. The presence of C. trachomatis was determined using commercial AMPLICOR C. trachomatis Assay (Roche Diagnostic Systems, Inc., Branchburg, NJ, USA). The prevalence of C. trachomatis infections among different groups was: recruits 0%, soldiers 0.4%, policemen 3.5%, clerks 4.6%, pregnant women 4%, and students 4.4%. The average prevalence for both groups (asymptomatic and symptomatic) was 2.3%[95% confidence interval (CI): 1.5-3.1%]. The average prevalence for the asymptomatic group was 1.6% (95% CI: 0.8-2.4%), while the average prevalence for the symptomatic group was 6.2% (95% CI: 3.1-9.3%) which were significantly different (P = 0.00003). Testing first void urine specimens by AMPLICOR C. trachomatis assay is a highly sensitive and specific method for diagnosing C. trachomatis infections in men and women. This method provides health care workers and public health officials with a new molecular amplification assay that uses noninvasive urine specimens for population-based screening purposes. The prevalence of C. trachomatis was relatively low among asymptomatic individuals. However, selective screening strategies are highly recommended for testing the student population in the Republic of Macedonia.

In vitro assessment of the APTIMA Combo 2 assay for the detection of Chlamydia trachomatis using highly purified elementary bodies

The Gen-Probe APTIMA Combo 2 assay has previously been reported to have a high sensitivity. The end point of this assay was evaluated using highly purified chlamydial elementary bodies (EBs). The performance of the APTIMA Combo 2 assay was compared with a commercially available PCR kit, AMPLICOR Chlamydia trachomatis. The number of inclusions of C. trachomatis at the end point of the APTIMA Combo 2 assay was 0.005 inclusion-forming units (i.f.u.) ml(-1), which was equivalent to 0.008 EBs per assay. The end point of the AMPLICOR kit was 5 i.f.u. ml(-1) (equivalent to 0.5 EB per assay). The efficacy of the AMPLICOR C. trachomatis assay was inhibited by phosphate or Fe2+ ion, while these had no effect on the APTIMA Combo 2 assay. In conclusion, the APTIMA Combo 2 assay appears to have a greater sensitivity than the AMPLICOR C. trachomatis assay for detection of C. trachomatis, while demonstrating few problems with inhibitory substances such as phosphate and Fe2+ ion.

Reliability of the Amplicor ™ internal control to disclose false-negative Chlamydia trachomatis PCR results

Four possibly false-negative samples were detected when 514 male urine specimens were tested in the Amplicor™ Chlamydia trachomatis assay. In three of the four samples, the inhibition could be reduced by removal of urine supernatant. Under partially inhibitory conditions, after spiking with 50 C. trachomatis elementary bodies/ml specimen, a selective inhibition of the C. trachomatis target amplification and a preferential internal control amplification was observed. We conclude that a positive internal control signal might be misleading in inhibitory specimens with low amount of C. trachomatis.

Semiautomation of Nucleic Acid-based Assays for Chlamydia trachomatis and Neisseria gonorrhoeae

The US prevalence rates of sexually transmitted diseases associated with Chlamydia trachomatis (CT) and Neisseria gonorrhoeae (GC) are as high as 27% (1) and 56% (2), respectively. Routine screening for both conditions is recommended for several risk groups, and infections with CT and GC frequently coexist (3). Because both infections are often asymptomatic, screening is important in efforts to slow the spread of infection. Recent studies compared amplified with nonamplified testing (4)(5)(6)(7)(8)(9)(10) and swab with urine specimens (4)(8)(11)(12)(13)(14)(15)(16)(17)(18)(19)(20). Because recommendations vary (21)(22)(23)(24)(25), each laboratory must decide which test(s) to use by considering the prevalence of infection, assay sensitivity and specificity, cost-effectiveness, laboratory work flow, and types of specimens received. Several tests may be required to best serve customers with different needs. We developed a menu of semiautomated tests, including amplified CT, nonamplified CT, and nonamplified GC assays. We describe two novel menu choices here: (a) a combination of the nonamplified PACE® 2 GC assay (Gen-Probe, Inc.) with the AMPLIFIEDTM Chlamydia Trachomatis Assay (AMP CT; Gen-Probe, Inc.) for swab specimens; and (b) AMP CT for urine specimens. We used the TECAN GENESIS RSP instrument (1.426 m × 0.780 m × 1.200 m; TECAN US, Inc.). It has an eight-tip arm and uses 1-mL, extended length, disposable, filter pipette tips with level-sensing capability. We designed and machined custom deck plates to support the necessary specimen racks, assay racks, and reagent reservoirs. With approval from the Institutional Review Board, four clinical sites collected urogenital swab specimens using the standard PACE 2 specimen collection method, and two clinical sites collected …

An exploratory study of information-giving used to promote chlamydial test-seeking by students at a college family planning clinic

Partnership working, involving workers in various aspects of sexual health and a large UK further education college, took place to give information about genital tract chlamydial infection in order to...

Detection of PCR inhibitors in cervical specimens by using the AMPLICOR Chlamydia trachomatis assay.

To determine that susceptibility of AMPLICOR Chlamydia trachomatis PCR to inhibitory factors possibly present in cervical specimens, we obtained cervical specimens from 200 gynecology patients attending our outpatient clinic. The prevalence of C. trachomatis infection was 4.1%, as determined by cell culture. All AMPLICOR specimens were tested in one procedure as described by the manufacturer, and after the specimen was spiked with C. trachomatis, several other pretreatment protocols were used. Complete inhibition of the PCR was observed in 38 (19%) cervical specimens. Heat treatment at 95 degrees C, freeze-thawing, or 10-fold dilution of the samples reduced the initial inhibition to 9, 16, or 9%, respectively. A combination of heat treatment and 10-fold dilution reduced the inhibition to 4% of the samples. A second specimen type (swabs inoculated in 0.2 M sucrose phosphate buffer [2SP]) was also evaluated. A 10-fold dilution of the spiked 2SP specimen resulted in an inhibition rate of 6%, which was comparable to that obtained by centrifugation of the 2SP specimen prior to processing. Furthermore, it was shown that the inhibition was not correlated with blood contamination. Processing the specimens on the day of collection or the day after resulted in a higher inhibition rate than did delayed processing (27.6 versus 15.5%, respectively). An inverse correlation was found between the concentration of C. trachomatis added to the sample and the rate of inhibition observed. The inhibition was partly correlated with the pH of the cervical mucosa. Decreased inhibition was found at pH values of > or = 7.5. The effects of blood, pH, and delay in processing were all evaluated by using the AMPLICOR specimen. We conclude that the susceptibility of AMPLICOR C. trachomatis PCR to inhibiting factors in cervical specimens can be significantly reduced if the pretreatment procedure includes heat treatment or the use of 2SP transport medium. Also, a 10-fold dilution of the clinical specimen followed by heat treatment will largely prevent the inhibition of this PCR.