Footprint of positive selection in Treponema pallidum subsp. pallidum genome sequences suggests adaptive microevolution of the syphilis pathogen

Abstract

In the rabbit model of syphilis, infection phenotypes associated with the Nichols and Chicago strains of Treponema pallidum (T. pallidum), though similar, are not identical. Between these strains, significant differences are found in expression of, and antibody responses to some candidate virulence factors, suggesting the existence of functional genetic differences between isolates. The Chicago strain genome was therefore sequenced and compared to the Nichols genome, available since 1998. Initial comparative analysis suggested the presence of 44 single nucleotide polymorphisms (SNPs), 103 small (≤3 nucleotides) indels, and 1 large (1204 bp) insertion in the Chicago genome with respect to the Nichols genome. To confirm the above findings, Sanger sequencing was performed on most loci carrying differences using DNA from Chicago and the Nichols strain used in the original T. pallidum genome project. A majority of the previously identified differences were found to be due to errors in the published Nichols genome, while the accuracy of the Chicago genome was confirmed. However, 20 SNPs were confirmed between the two genomes, and 16 (80.0%) were found in coding regions, with all being of non-synonymous nature, strongly indicating action of positive selection. Sequencing of 16 genomic loci harboring SNPs in 12 additional T. pallidum strains, (SS14, Bal 3, Bal 7, Bal 9, Sea 81-3, Sea 81-8, Sea 86-1, Sea 87-1, Mexico A, UW231B, UW236B, and UW249C), was used to identify "Chicago-" or "Nichols -specific" differences. All but one of the 16 SNPs were "Nichols-specific", with Chicago having identical sequences at these positions to almost all of the additional strains examined. These mutations could reflect differential adaptation of the Nichols strain to the rabbit host or pathoadaptive mutations acquired during human infection. Our findings indicate that SNPs among T. pallidum strains emerge under positive selection and, therefore, are likely to be functional in nature.

Figure 1. TPChic0899 sequence alignment in T. pallidum isolates.

The single artifactual G deletion in the Nichols strain results in a frameshift and puts in frame the TGA triplet resulting in a premature stop codon and two ORFs (TP0899 and TP0900) was not confirmed in either the several Nichols lineages (Seattle, Dallas, Houston, UCLA, and Farmington) and in the eleven other non-Nichols T. pallidum isolates examined. Amino acid position is indicated according to the Chicago strain genome annotation. Nichols (Gen) and SS14 (Gen) refers to the genome sequences already available for these strains in GenBank (accession numbers are NC_000919 and CP000805.1, respectively).

Figure 2. tex gene sequence alignment in T. pallidum isolates.

The C→A transversion that generates a premature TAA (stop) codon in the TPChic0924 gene was found in eleven other non-Nichols T. pallidum isolates. No truncating mutation was found in the tex gene from the Nichols isolate currently propagated in our laboratory (Nichols Seattle), and in the Nichols strain used in the orginal T. pallidum genome project (Houston) , or in Nichols strains obtained from other laboratories (Dallas, UCLA, and Farmington). Amino acid position is assigned according to the published Nichols strain genome annotation . Nichols (Gen) refers to the genome sequences already available for this strain in GenBank (accession number is NC_000919).