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.

FIG. 1.

Schematic representation of the C. trachomatis genome. The omp1 gene, approximately 1.2 kb in length, was PCR amplified with various combinations of primers. Primers P1 and OMP2 were used in the primary PCR. DNA which failed to generate primary PCR products was subsequently amplified in a nested PCR with the following primer combinations: P1-CT6R, CT6F-OMP2, and NL-F-NL-R. Nucleotide sequence data from the C. trachomatis conjunctival samples was assembled into one contiguous sequence spanning the four variable domains (VDI to VDIV).

FIG. 2.

Distance neighbor-joining phylogenetic tree based on C. trachomatis nucleotide sequences spanning VDI to VDIV for the conjunctival samples. For clarity, only one representative from each genetic variant of the samples is illustrated (in bold type). Values in parentheses indicate the number of samples of that variant. Selected reference isolates stored in our laboratory are also included (underlined). The tree was rooted by using the equivalent MoPn omp1 nucleotide sequence region. Numbers at the nodes are bootstrap values (percentages) from 1,000 replicates. The bar corresponds to a graphic distance equivalent to 0.1 nucleotide substitution per site.

FIG. 3.

Distance neighbor-joining phylogenetic tree based on C. trachomatis amino acid sequences spanning VDI to VDIV for the conjunctival samples. For clarity, only one representative from each genetic variant of the samples is illustrated (in bold type). Values in parentheses indicate the number of samples of that variant. Selected reference isolates stored in our laboratory are also included (underlined). The tree was rooted by using the equivalent MoPn MOMP amino acid sequence region. Numbers at the nodes are bootstrap values (percentages) from 1,000 replicates. The bar corresponds to a graphic distance equivalent to 0.1 amino acid substitution per site.