Characterization of Chlamydia trachomatis omp1 Genotypes Detected in Eye Swab Samples from Remote Australian Communities

Abstract

Chlamydia trachomatis conjunctival samples collected over a 6-month period from individuals with clinical signs of trachoma and located in remote communities in the Australian Northern Territory were differentially characterized according to serovar and variants. The rationale was to gain an understanding of the epidemiology of an apparent increased prevalence of acute trachoma in areas thought to be less conducive to this disease. Characterization was performed through sequencing of a region of the omp1 gene spanning the four variable domains and encoding the major outer membrane protein. Nucleotide and deduced amino acid sequences were genotyped by using a BLAST similarity search and were examined by phylogenetic analyses to illustrate evolutionary relationships between the clinical and GenBank reference strains. The predominant genotype identified corresponded to that of serovar C (87.1%), followed by the genotype corresponding to serovar Ba (12.9%). All nucleotide and amino acid sequences exhibited minor levels of variation with respect to GenBank reference sequences. The omp1 nucleotide sequences of the clinical samples best aligned with those of the conjunctival C. trachomatis reference strains C/TW-3/OT and Ba/Apache-2. All clinical samples (of serovar C) exhibited four or five nucleotide changes compared with C/TW-3/OT, while all serovar Ba samples had one or two nucleotide differences from Ba/Apache-2. Phylogenetic analyses revealed close relationships between these Northern Territory chlamydial samples and the respective reference strains, although the high proportion of sequence variants suggests an evolutionarily distinct C. trachomatis population causing eye infections in Australia. Given that such genotypic information has gone unreported, these findings provide knowledge and a foundation for trachoma-associated C. trachomatis variants circulating in the Northern Territory.

Evaluation of a Novel PCR-Based Assay for Detection and Identification of Chlamydia trachomatis Serovars in Cervical Specimens

“…If the isolated DNA contains only a very low concentration of bacterial DNA, it is possible that only cryptic-plasmid DNA but no genomic DNA would be included in the PCR mixture and that only the plasmid PCR yields positive results. Similar findings of a positive plasmid-directed PCR and a negative omp1 PCR have been reported for genital and conjunctival samples (22,28).…”

Development and Evaluation of anompAQuantitative Real-Time PCR Assay forChlamydia trachomatisSerovar Determination

“…In cases where amplification did not result in sufficient amplicons for sequ encing, smaller overlapping PCR products were targeted using the following nested primer pairs: MS-1F/CT6-R (655 to 679 bp), 518-bp fragment spanning VDI to -II; CT6-F (655 to 679 bp)/MS-1R, 443-bp fragment spanning VDIII to -IV; and NL-F (394 to 413 bp) and NL-R (856 to 875 bp), 482-bp fragment spanning VDII to -III. The nucleotide sequences of these primers have been described previously (25). The PCR cycling conditions used were as described above with the exception of a lowered annealing temperature of 50°C and use of 5 l of purified PCR template.…”

“…DNA extracts obtained from the clinical specimens were sequenced across the four variable domains (VDs) of the ompA gene. Sequencing of the eye swab DNA extracts has been previously described (25). For the purpose of orientation, the following primer positions are based on the ompA gene of serovar A strains.…”

“…These specimens comprised first-pass urine (n ϭ 210) (42.3%) and anal swabs (n ϭ 284) (57.7%). In addition, six DNA extracts obtained from eye swab specimens collected during the 2002 outbreak of trachoma in the Northern Territory (25) were selected from a collection of 31 specimens to evaluate detection of the A to C serovars. All clinical specimens (anal swabs, urine, and eye swabs) were stored at Ϫ80°C until processing.…”

Chlamydia trachomatis Serovars among Strains Isolated from Members of Rural Indigenous Communities and Urban Populations in Australia

“…While numerous studies have addressed the epidemiology of C. trachomatis in Australia, very little is known about the distribution of C. trachomatis serovars. One of these studies characterized the C. trachomatis serovars in eye swab samples from people living in remote Australian communities (7). Among the C. trachomatis serovars A, B, and C, which are usually associated with trachoma, only serovar C (87%) and serovariant Ba (13%) were found in conjunctival samples from 31 patients.…”