RNA AMPLIFICATION BY NUCLEIC-ACID SEQUENCE-BASED AMPLIFICATION WITH AN INTERNAL STANDARD ENABLES RELIABLE DETECTION OF CHLAMYDIA-TRACHOMATIS IN CERVICAL SCRAPINGS AND URINE SAMPLES

In the present study, the suitability of RNA amplification by nucleic acid sequence-based amplification (NASBA) for the detection of Chlamyd ia trachomatis infection was investigated, When comparing different pr imer sets for their sensitivities in NASBA use of both the plasmid and omp1 targets resulted in a detection limit of 1 inclusion-forming uni t (IFU), while the 16S rRNA appeared to be the most sensitive RNA targ et for amplification (10(-3) IFU), In contrast, for DNA amplification by PCR, the plasmid target was optimal (10(-2) IFU), which is 10 times less sensitive than rRNA NASBA. To exclude false negativity in NASBA detection because of inhibition of amplification and/or inefficient sa mple preparation, an internal standard was developed. The internal con trol was added prior to sample preparation, This 16S rRNA NASBA with a n internal control was compared with a plasmid DNA PCR by using a grou p of C. trachomatis-negative (n = 41) and -positive (n = 37) cervical scrapings, as determined by enzyme immunoassay (EIA). In addition, uri ne samples from the EIA-positive women were tested (n = 17). Both NASB A and PCR assays were able to detect C trachomatis in all EIA-positive cervical scrapings, the corresponding urine samples, and two samples from the EIA-negative group, The internal NASBA standard was found cle arly in all EIA-negative samples, In conclusion, these results indicat e that detection of C. trachomatis by RNA amplification by NASBA with an internal standard is a suitable and highly sensitive detection meth od, with potential use in the diagnosis of urogenital C. trachomatis i nfections with cervical scrapings as well as urine specimens.