Serotyping of Chlamydia trachomatis by Polymerase Chain Reaction: Restriction Fragment Length Polymorphism Analysis

Seo, Yiel Hea; Park, Pil Whan; Kim, Wan; Kim, Duck An; Choi, Tae Yeal

Abstract

BACKGROUND: Chlamydia trachomatis is currently classified into 15 serovars (A, B, Ba, C, D, E, F, G, H, I, J, K, L1, L2, L3) and a large number of serovariants. Typing of C. trachomatis serovars has generally been performed by the MOMP (major outer membrane protein) serotyping method, which requires a large panel of polyclonal and monoclonal antibodies. Recently the PCR was successfully applied to the genotyping of different C. trachomatis serovars by means of restriction fragment length polymorphism (RFLP) analysis and direct sequencing of amplified omp1 DNA. The objective of this study was to evaluate the serotyping of C. trachomatis by PCR-FLP and to determine the serovars of C. trachomatis urogenital isolates.
METHODS
The 15 reference strains of C. trachomatis and 27 clinical isolates were analyzed by PCR-FLP. The C. trachomatis omp1 gene was amplified by PCR. For serotyping by RFLP analysis, the omp1 PCR products were digested with AluI and were electrophoresed through a 7% polyacrylamide gel with ethidium bromide staining. If necessary, the PCR products were analyzed with HinfI, a combination of EcoRI and DdeI, or CfoI.
RESULTS
In serotyping of 15 reference strains of C. trachomatis, serovars A, B, Ba, E, F, G, K, and L2 were clearly identified after AluI digestion. However serovars C, H, I, J, L3 and serovars D, L1 were respectively identical patterns after AluI digestion. Serovars C and J and serovars D and L1 were further discriminated by a second step of enzyme digestion with HinfI. Serovar H, I, and L3 were distinguished by enzyme digestion with EcoRI and DdeI. In serotyping of C. trachomatis from 27 urogenital isolates, 25 isolates were clearly typed. Nine were typed as serovar E, 8 were typed as serovar D, 6 were typed as F, 2 were typed as serovar H. Two isloates showed unidentifiable RFLP pattern which was not in accordance with any of the existing C. trachomatis prototypes.
CONCLUSIONS
PCR-FLP analysis is a rapid, simple, and powerful tool for differentiating serovars of C. trachomatis. Therefore, this approach is recommended for future epidemiological studies of C. trachomatis. And the serovar E, D, and F are the most prevalent types found in urogenital specimens, representing 92% of those investigated.