Whole genome sequence of the Treponema pallidum subsp. endemicum strain Bosnia A: the genome is related to yaws treponemes but contains few loci similar to syphilis treponemes

Abstract

Background: T. pallidum subsp. endemicum (TEN) is the causative agent of bejel (also known as endemic syphilis). Clinical symptoms of syphilis and bejel are overlapping and the epidemiological context is important for correct diagnosis of both diseases. In contrast to syphilis, caused by T. pallidum subsp. pallidum (TPA), TEN infections are usually spread by direct contact or contaminated utensils rather than by sexual contact. Bejel is most often seen in western Africa and in the Middle East. The strain Bosnia A was isolated in 1950 in Bosnia, southern Europe.

Methodology/principal findings: The complete genome of the Bosnia A strain was amplified and sequenced using the pooled segment genome sequencing (PSGS) method and a combination of three next-generation sequencing techniques (SOLiD, Roche 454, and Illumina). Using this approach, a total combined average genome coverage of 513× was achieved. The size of the Bosnia A genome was found to be 1,137,653 bp, i.e. 1.6-2.8 kbp shorter than any previously published genomes of uncultivable pathogenic treponemes. Conserved gene synteny was found in the Bosnia A genome compared to other sequenced syphilis and yaws treponemes. The TEN Bosnia A genome was distinct but very similar to the genome of yaws-causing T. pallidum subsp. pertenue (TPE) strains. Interestingly, the TEN Bosnia A genome was found to contain several sequences, which so far, have been uniquely identified only in syphilis treponemes.

Conclusions/significance: The genome of TEN Bosnia A contains several sequences thought to be unique to TPA strains; these sequences very likely represent remnants of recombination events during the evolution of TEN treponemes. This finding emphasizes a possible role of repeated horizontal gene transfer between treponemal subspecies in shaping the Bosnia A genome.

Figure 1. Unrooted tree based on the alignment of the Bosnia A genome with additional treponemal genomes.

An unrooted tree was constructed from the complete genome sequences of TPA strains (Nichols, Chicago, DAL-1, SS14, and Mexico A), TPE strains (CDC-2, Gauthier, Samoa D, and Fribourg-Blanc), and the TEN strain (Bosnia A) using the Maximum Parsimony method and MEGA5 software . The bar scale corresponds to a difference of 200 nucleotides. Bootstrap values based on 1,000 replications are shown next to the branches. All positions containing indels in at least one genome sequence were omitted from the analysis. There were a total of 1,128,391 nucleotide positions aligned in the final dataset.

Figure 2. Representation of the Bosnia A chromosome with location of Bosnia A-, TPE-, and TPA-specific sequences.

Bosnia A-specific sequences are shown in green while TPE-specific sequences (TPA and Bosnia A sequences are identical in these loci) are shown in blue. TPA-specific sequences (TPE and Bosnia A sequences are identical in these loci) are shown in red. Bosnia A-specific sequences comprised 406 loci (encompassing a total of 2772 bp) while TPE- and TPA-specific sequences were found in 197 (635 bp) and 1422 (2335 bp) loci, respectively.

Figure 3. Sequence alignments of TENDBA_0133, TENDBA_0968, TENDBA_0577, TENDBA_0326 and TENDBA_0856 loci with the orthologous sequences of TPA and TPE genomes.

Sequences of five TPA strains (Nichols, DAL-1, Chicago, Mexico A and SS14) and four TPE strains (CDC-2, Gauthier, Samoa D and Fribourg-Blanc) are shown. Numbers above the alignment represent gene coordinates in the re-sequenced TPA Nichols strain (CP004010.2 [14]). While the alignment of TENDBA_0133 showed locus completely identical to TPE strains, TENDBA_0968, TENDBA_0577 and TENDBA_0326 loci showed the presence of TPA sequences in the genome of Bosnia A. The TENDBA_0856 locus represent Bosnia A specific region with sporadic nucleotide positions identical to TPA sequences. The TPE-like sequence was found in most of the Bosnia A loci while the pattern found in TENDBA_0968 was also found in TENDBA_0858 (not shown) and the pattern identified in the TENDBA_0577 was found also in the TENDBA_1031 (not shown). The alignment pattern in TENDBA_0326 was previously found in TENDBA_0488 and the pattern in TENDBA_0856 in TENDBA_0865.