Molecular typing of Treponema pallidum in the Czech Republic during 2011 to 2013: increased prevalence of identified genotypes and of isolates with macrolide resistance

Abstract

From January 2011 to December 2013, a total of 262 samples, from 188 patients suspected of having syphilis were tested for the presence of treponemal DNA by PCR amplification of five chromosomal loci, including the polA (TP0105), tmpC (TP0319), TP0136, TP0548, and 23S rRNA genes. Altogether, 146 samples from 103 patients were PCR positive for treponemal DNA. A set of 81 samples from 62 PCR-positive patients were typeable, and among them, nine different genotypes were identified. Compared to a previous study in the Czech Republic during 2004 to 2010, the number of genotypes detected among syphilis patients in a particular year increased to six in both 2012 and 2013, although they were not the same six. The proportion of macrolide-resistant clinical isolates in this 3-year study was 66.7%.

FIG 1

Flow chart of the patients investigated in this study. Seronegative samples that were PCR negative were excluded from this study. We analyzed 262 samples from 188 patients. Altogether, we analyzed 153 whole-blood samples, 103 swabs, 3 CSF samples, and 3 tissue samples. Among this set of samples, we detected 146 PCR-positive samples from 103 patients.

FIG 2

Alignment of unique sequences of the TP0136 and TP0548 loci of clinical isolates collected in the Czech Republic between 2004 and 2013 (; this study). Only positions showing nucleotide differences (indels, substitutions) are shown. Coordinates correspond to TP0136 and TP0548 in the T. pallidum subsp. pallidum (TPA) SS14 genome (accession no. CP004011) (37). Nucleotide differences shown on a gray background indicate differences between the SS14-like and Nichols-like groups of strains. Nucleotide changes shown on black and white backgrounds represent unique differences found in clinical isolates.

FIG 3

Distribution of T. pallidum subsp. pallidum genotypes in the Czech Republic from 2004 to 2013. Data for 2004 to 2010 are from a previous study performed in the Czech Republic (5). Eleven completely typed genotypes and three partially typed genotypes (XU3R8, XU4R9, XU8R9) with unique TP0548 sequences are shown. Allele numbering reflects not the year of isolation but the date of sequence analysis; therefore, genotype XU3R8 appeared earlier than genotype SU2R8. Similarly, genotype SU5R8 appeared later than genotypes U3U6R8, SU7S, and XU8R9. Note that 14 different genotypes of T. pallidum subsp. pallidum isolates were identified in the Czech Republic from 2004 to 2013. The number of genotypes detected in a particular year increased over time, while some genotypes disappeared over time. The relative abundance of genotypes in a particular year is color coded. Three of the 91 genotyped T. pallidum subsp. pallidum isolates represented were from patients in the early latent stage of syphilis. All of these genotypes are shown 1 year before the diagnosis of the disease to reflect the prevalence of the genotypes in a particular year.

FIG 4

(A) Prevalence of the A2058G and A2059G mutations in treponemal 23S rRNA genes isolated from clinical samples from syphilis patients in the Czech Republic from 2007 to 2013. Data from 2007 to 2010 are from a previous study in the Czech Republic (5). While the prevalence of A2058G appears to be increasing, the prevalence of A2059G is relatively low and appears to be decreasing. The relative prevalence of mutations in the 23S rRNA gene locus since 2007 is shown because no isolates harboring these mutations were found in 2004 or 2005. In 2006, only one isolate harboring A2058G was identified. (B) Macrolide antibiotic use in the Czech Republic from 2007 to 2013. Data were provided by the State Institute for Drug Control in the Czech Republic. Antibiotic use is shown as the number of medical product packages representing the dose most frequently used for the treatment of a single patient.