Human Treponema pallidum 11q/j isolate belongs to subsp. endemicum but contains two loci with a sequence in TP0548 and TP0488 similar to subsp. pertenue and subsp. pallidum, respectively

Abstract

Background: Treponema pallidum subsp. endemicum (TEN) is the causative agent of endemic syphilis (bejel). An unusual human TEN 11q/j isolate was obtained from a syphilis-like primary genital lesion from a patient that returned to France from Pakistan.

Methodology/principal findings: The TEN 11q/j isolate was characterized using nested PCR followed by Sanger sequencing and/or direct Illumina sequencing. Altogether, 44 chromosomal regions were analyzed. Overall, the 11q/j isolate clustered with TEN strains Bosnia A and Iraq B as expected from previous TEN classification of the 11q/j isolate. However, the 11q/j sequence in a 505 bp-long region at the TP0488 locus was similar to Treponema pallidum subsp. pallidum (TPA) strains, but not to TEN Bosnia A and Iraq B sequences, suggesting a recombination event at this locus. Similarly, the 11q/j sequence in a 613 bp-long region at the TP0548 locus was similar to Treponema pallidum subsp. pertenue (TPE) strains, but not to TEN sequences.

Conclusions/significance: A detailed analysis of two recombinant loci found in the 11q/j clinical isolate revealed that the recombination event occurred just once, in the TP0488, with the donor sequence originating from a TPA strain. Since TEN Bosnia A and Iraq B were found to contain TPA-like sequences at the TP0548 locus, the recombination at TP0548 took place in a treponeme that was an ancestor to both TEN Bosnia A and Iraq B. The sequence of 11q/j isolate in TP0548 represents an ancestral TEN sequence that is similar to yaws-causing treponemes. In addition to the importance of the 11q/j isolate for reconstruction of the TEN phylogeny, this case emphasizes the possible role of TEN strains in development of syphilis-like lesions.

Fig 1. A schematic representation of the sequenced chromosomal loci of the TEN 11q/j isolate.

The TEN 11q/j chromosome is shown as a circle and the pink regions represent sequenced chromosomal loci similar to corresponding chromosomal loci of TEN Bosnia A and Iraq B. The sequenced regions are not shown to scale, with respect to the whole genome length. The locus names of the sequenced loci are shown outside the circle together with the length of each sequenced region. Blue region within the TP0488 locus represents a sequence related to TPA treponemes, while the green region within the TP0548 locus represents a sequence related to TPE treponemes. Sanger sequenced regions are denoted as black circles and chromosomal loci sequenced using the Illumina method are shown as grey circles. The chromosomal regions amplified within Pool 1 of the PSGS sequencing approach are indicated by the black line.

Fig 2

Sequence alignments at the TEND11qj_0488 (A.) and TEND11qj_0548 (B.) loci with the orthologous sequences of the selected TEN, TPA, and TPE genomes. Sequences of two TEN strains (Bosnia A and Iraq B), two TPE strains (Gauthier and Samoa D) and two TPA strains (Nichols and SS14) are shown. Numbers above the alignment represent gene coordinates in the corresponding genes of TEN Bosnia A (CP007548.1; [16]). The alignment of TEND11qj_0488 showed a sequence very similar to TPA strains, and TEND11qj_0548 was similar to TPE strains.

Fig 3. Unrooted tree based on the alignment of the TEND11qj_0488, TEND11qj_0548 and the remaining sequenced loci of the TEN 11q/j isolate.

A. An unrooted tree was constructed from a 505 bp-long sequence of the locus TP0488, shown in Fig 2A between coordinates 715–1221 corresponding to TEN Bosnia A numbering (CP007548; [16]). B. An unrooted tree constructed from a 613 bp-long sequence of the locus TP0548, shown in Fig 2B between coordinates 69 of TENDBA_0547a and coordinate 623 of TENDBA_0548 (CP007548; [16]). C. An unrooted tree was constructed from concatenated sequences of 42 genomic regions of the 11q/j isolate, except for the TP0488 and TP0548 genes, and respective sequences of TPA strains (Nichols, SS14), TPE strains (CDC-2, Gauthier, Samoa D, and Fribourg-Blanc), and the TEN strains (Bosnia A, Iraq B) using the Maximum Likelihood method and MEGA7 software [31]. 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 was a total of 29,447 nucleotide positions aligned in the final dataset. The bar scale corresponds to a difference of 0.005 (A.), 0.005 (B.) and 0.001 (C.) nucleotides. While the 11q/j isolate cluster in the TP0488 locus with TPA strains and in the TP0548 locus with TPE strains, a tree constructed from concatenated sequences of 42 genomic regions of the 11q/j isolate cluster with TEN strains. A position within the tree where the putative recombination events, involving TP0488 and TP0548, likely occurred is shown, respectively.