Assessment of Chlamydia trachomatis infection by Cobas Amplicor PCR and in-house LightCycler assays using PreservCyt and 2-SP media in voluntary legal abortions

Abstract

Chlamydial infection of the upper genital tract after abortion is well recognized, but routine screening for infection before termination is rare, and few centres are aware of the prevalence of post-abortion complications in their patient population. Knowledge of the patient population is the best guide for developing screening strategies. The aim of this study was to determine the prevalence of chlamydial infection in patients presenting for legal termination of pregnancy, and to assess the presence of Chlamydia trachomatis by PCR on specimens collected in either PreservCyt (ThinPrep) or 2-sucrose phosphate (2-SP) transport medium. Two hundred and eleven single, sexually active women, aged 15–26 years, attending the Gynaecology and Obstetric Hospital, Amiens, France, for surgical termination of pregnancy were enrolled in this study from June 2002 to June 2003. C. trachomatis detection using a Cobas Amplicor PCR test (Roche Diagnostics) targeting a 207 bp segment of the common cryptic plasmid and a quantitative LightCycler real-time PCR (LC-PCR) (Roche Diagnostics) targeting a 123 bp fragment within the highly conserved constant domain 3 of the single-chromosome-copy ompA gene were performed on endocervical swabs in 2-SP, and on specimens collected using a cytobrush and placed in PreservCyt medium. The in-house LC-PCR was used as a chromosomal diagnosis method and to determine the load of C. trachomatis. This method was able to detect the mutant Swedish variant with a deletion of 377 bp in the target area in the cryptic plasmid, which is the region targeted by the Cobas Amplicor PCR test. C. trachomatis was detected in 19/211 patients (9 %) by both PCR methods. Among the 19 infected women, C. trachomatis was detected by the Cobas Amplicor PCR in 16 specimens in PreservCyt (7.6 %) and in 12 endocervical swabs in 2-SP (5.7 %). Specimens from only nine women were PCR-positive in both PreservCyt and 2-SP media by this method. Cobas Amplicor PCR revealed that 10.9 and 2.3 % of the PreservCyt and 2-SP samples, respectively, contained inhibitors. The same 19 infected women were LC-PCR positive in both PreservCyt and 2-SP samples. No additional infected women were found by this last method; thus, it was concluded that none of the samples contained the new variant of C. trachomatis. The load in each sample varied from 102 to 107 copies ml−1.

Detection of chlamydia trachomatis by immunological methods in adult and adolescent female population in Cuiabá, Mato Grosso

“…Among the ethnic group, there was a higher number of positive cases in the mulatto group, perhaps owing to the prevalence of this phenotype in the population of Mato Grosso. However, there were no significant differences between white or black skin groups, although some authors consider the existence of differences in the prevalence of Chlamydia infection among ethnic groups (30,31) . Various methods are employed for the diagnosis of Chlamydia.…”

“…However, the diagnosis by PCR has replaced other techniques for its speed, more reliable reproducibility and, currently, low cost. The methods based on the amplification of nucleic acids have demonstrated a high positive predictive value, presenting the advantage of being usable with urethral, cervical, vaginal, and urine specimens (30,31) . Several factors can interfere with the determination of the prevalence of this STD, such as the laboratory resources available, ecology of the bacteria, the sexual behavior of population groups, therapeutic interference, among others.…”

Diagnosis and follow-up of genital chlamydial infection by direct methods and by detection of serum IgG, IgA and secretory IgA

“…[13] The NAATs, which have become the method of choice have high sensitivity and specificity and are considered to be a more suitable method either for screening or diagnosing chlamydial infections. [12,14] Produced by submucosal plasma cells, C. trachomatis sIgA antibodies were reported to be a better indicator of a manifest immunoresponse caused by an antigenic stimulus than circulating antibodies. [8,15] Studies have shown that infection of the genital tract by C. trachomatis is associated with a rise in levels of cervical sIgA and in the number of IgA-producing plasma cells.…”

“…These tubes were then centrifuged at 13000 g for 5 minutes and, incubated at 95°C for 10 minute, which reduces the effects of inhibitors. [11,12] Fifty microliters of the processed specimens were immediately transferred to A-rings containing 50 µl of PCR Master Mix for amplification. An internal control (IC) was included in the master mix to monitor inhibition and was amplified with the C. trachomatis target DNA.…”

Diagnosis of Chlamydia trachomatis Infection

“…Commercially available techniques utilized have largely focused on PCR [94,99,100], ligase chain reaction [101,102] and strand displacement assays (SDA) [103], amongst others [104,105]. Further development into real-time [106,107] and multiplexed based platforms [108], as well as nesting steps [109] has also improved diagnostic utility.…”