Evaluation of a High Resolution Genotyping Method for Chlamydia trachomatis Using Routine Clinical Samples

Abstract

BackgroundGenital chlamydia infection is the most commonly diagnosed sexually transmitted infection in the UK. C. trachomatis genital infections are usually caused by strains which fall into two pathovars: lymphogranuloma venereum (LGV) and the genitourinary genotypes D–K. Although these genotypes can be discriminated by outer membrane protein gene (ompA) sequencing or multi-locus sequence typing (MLST), neither protocol affords the high-resolution genotyping required for local epidemiology and accurate contact-tracing.Principal FindingsWe evaluated variable number tandem repeat (VNTR) and ompA sequencing (now called multi-locus VNTR analysis and ompA or “MLVA-ompA”) to study local epidemiology in Southampton over a period of six months. One hundred and fifty seven endocervical swabs that tested positive for C. trachomatis from both the Southampton genitourinary medicine (GUM) clinic and local GP surgeries were tested by COBAS Taqman 48 (Roche) PCR for the presence of C. trachomatis. Samples tested as positive by the commercial NAATs test were genotyped, where possible, by a MLVA-ompA sequencing technique. Attempts were made to isolate C. trachomatis from all 157 samples in cell culture, and 68 (43%) were successfully recovered by repeatable passage in culture. Of the 157 samples, 93 (i.e. 59%) were fully genotyped by MLVA-ompA. Only one mixed infection (E & D) in a single sample was confirmed. There were two distinct D genotypes for the ompA gene. Most frequent ompA genotypes were D, E and F, comprising 20%, 41% and 16% of the type-able samples respectively. Within all genotypes we detected numerous MLVA sub-types.ConclusionsAmongst the common genotypes, there are a significant number of defined MLVA sub-types, which may reflect particular background demographics including age group, geography, high-risk sexual behavior, and sexual networks.

Multilocus Variable-Number Tandem-Repeat Analysis Scheme for Chlamydia felis Genotyping: Comparison with Multilocus Sequence Typing

“…Interestingly, in samples from two cats in two different catteries, where both eyes were sampled, two different genotypes were identified, which in one case differed by only one tandem repeat. These findings provide the first evidence that mixed infections with strains of different C. felis genotypes occur in cats, similar to those described for C. trachomatis in humans (39). The possibility that the different genotypes identified in these two cases may represent divergent populations resulting from instability in the MLVA loci cannot be excluded; however, analysis of two C. felis isolates passaged 10 times through chicken eggs suggests that the MLVA genotype is highly stable.…”

Chlamydia trachomatis infection prevalence and serovar distribution in a high-density urban area in the north of Italy

“…Although high-resolution genotyping methods, such as multilocus variable number of tandem repeat analysis (MLVA), multilocus sequence typing (MLST) or microarray assay (Christerson et al, 2011;Ruettger et al, 2011;Bom et al, 2011;Wang et al 2011;Satoh et al, 2014;Hermann et al, 2015), have been recently described, their use in routine diagnostic practice is largely restricted by their complexity and high costs. So far these assays have basically played a role in epidemiological and phylogenetic analysis or in the evolutionary surveillance of specific clones, but what is really needed in the near future is the advent of rapid and cost-saving methods for faster and reliable management of patients (Pedersen et al, 2009;Xia & Xiong, 2014).…”

Highly diverse MLVA-ompA genotypes of rectalChlamydia trachomatisamong men who have sex with men in Brighton, UK and evidence for an HIV-related sexual network

“…There are now several very highly discriminating multilocus molecular typing systems, including multi locus sequence typing (MLST), for C. trachomatis that allow differentiation and subgrouping of isolates within a genotype 13 16 17 . W4–6 The multilocus variable number tandem repeat (VNTR) analysis plus analysis of the omp A gene (MLVA- omp A) high-resolution genotyping system of Pedersen et al ,13 has been successfully evaluated,18 and recent work has shown that the VNTR marker sequences are stable and therefore ideal for genotyping 19. The system consists of short, easy to amplify markers (the longest being part of the omp A gene), which are simple to manipulate and yield a high efficiency return of results as demonstrated in the quoted surveys.…”

“…The system consists of short, easy to amplify markers (the longest being part of the omp A gene), which are simple to manipulate and yield a high efficiency return of results as demonstrated in the quoted surveys. It also has an optimal ability to differentiate between genotypes,20 as measured by the Simpson's discriminatory index 13 18…”