BACKGROUND: For many years, syphilis treatment was considered straightforward due to the universal susceptibility of Treponema pallidum subsp. pallidum (TPA) to penicillin antibiotics. METHODS: Penicillin-binding protein genes from a ceftriaxone treatment failure T. pallidum isolate were assessed, and the introduction of identified mutations into two laboratory strains via natural competence was aimed for, followed by in vitro analysis of antibiotic susceptibility of the recombinants. RESULTS: TPA from the ceftriaxone treatment failure case contained A1873G and G2122A mutations in the TP0705 gene. Introduction of the A1873G mutation into laboratory strains DAL-1 and SS14 resulted in partial resistance to ceftriaxone and penicillin G in vitro. Furthermore, in silico analyses revealed that the majority of contemporary TPA SS14-like strains harbors this mutation and are thus partially resistant to ceftriaxone and penicillin G. CONCLUSIONS: This finding indicates that TPA strains accumulate mutations that increase their resistance to β-lactam antibiotics. Alternative approaches for controlling syphilis will be needed, including the development of the syphilis vaccine.

Penicillin antibiotics have been used to treat syphilis since the 1950s. Resistance to antibiotics is a growing concern. We investigated cases where antibiotics had failed to treat infection and found two mutations in a specific gene that could be responsible. Introduction of one of these mutations into two laboratory T. pallidum strains (the bacteria that cause syphilis) resulted in partial resistance to both ceftriaxone and penicillin antibiotics. Moreover, analysis of existing data revealed the presence of this mutation in numerous circulating T. pallidum strains, suggesting widespread partial resistance may already exist and increasing concerns about the future emergence of fully resistant syphilis strains.

• Publication type
• Journal Article MeSH

BACKGROUND: The increase in syphilis rates worldwide necessitates development of a vaccine with global efficacy. We aimed to explore Treponema pallidum subspecies pallidum (TPA) molecular epidemiology essential for vaccine research by analysing clinical data and specimens from early syphilis patients using whole-genome sequencing (WGS) and publicly available WGS data. METHODS: In this multicentre, cross-sectional, molecular epidemiology study, we enrolled patients with primary, secondary, or early latent syphilis from clinics in China, Colombia, Malawi, and the USA between Nov 28, 2019, and May 27, 2022. Participants aged 18 years or older with laboratory confirmation of syphilis by direct detection methods or serological testing, or both, were included. Patients were excluded from enrolment if they were unwilling or unable to give informed consent, did not understand the study purpose or nature of their participation, or received antibiotics active against syphilis in the past 30 days. TPA detection and WGS were conducted on lesion swabs, skin biopsies, skin scrapings, whole blood, or rabbit-passaged isolates. We compared our WGS data to publicly available genomes and analysed TPA populations to identify mutations associated with lineage and geography. FINDINGS: We screened 2802 patients and enrolled 233 participants, of whom 77 (33%) had primary syphilis, 154 (66%) had secondary syphilis, and two (1%) had early latent syphilis. The median age of participants was 28 years (IQR 22-35); 154 (66%) participants were cisgender men, 77 (33%) were cisgender women, and two (1%) were transgender women. Of the cisgender men, 66 (43%) identified as gay, bisexual, or other sexuality. Among all participants, 56 (24%) had HIV co-infection. WGS data from 113 participants showed a predominance of SS14-lineage strains with geographical clustering. Phylogenomic analyses confirmed that Nichols-lineage strains were more genetically diverse than SS14-lineage strains and clustered into more distinct subclades. Differences in single nucleotide variants (SNVs) were evident by TPA lineage and geography. Mapping of highly differentiated SNVs to three-dimensional protein models showed population-specific substitutions, some in outer membrane proteins (OMPs) of interest. INTERPRETATION: Our study substantiates the global diversity of TPA strains. Additional analyses to explore TPA OMP variability within strains is vital for vaccine development and understanding syphilis pathogenesis on a population level. FUNDING: US National Institutes of Health National Institute for Allergy and Infectious Disease, the Bill & Melinda Gates Foundation, Connecticut Children's, and the Czech Republic National Institute of Virology and Bacteriology.

• MeSH
• Bacterial Vaccines immunology   administration & dosage   MeSH
• Adult MeSH
• Phylogeny MeSH
• Genetic Variation genetics   MeSH
• Genome, Bacterial MeSH
• Genomics MeSH
• Middle Aged MeSH
• Humans MeSH
• Young Adult MeSH
• Molecular Epidemiology * MeSH
• Cross-Sectional Studies MeSH
• Whole Genome Sequencing * MeSH
• Syphilis * epidemiology   microbiology   MeSH
• Treponema pallidum * genetics   immunology   MeSH
• Treponema MeSH
• Check Tag
• Adult MeSH
• Middle Aged MeSH
• Humans MeSH
• Young Adult MeSH
• Male MeSH
• Female MeSH
• Publication type
• Journal Article MeSH
• Multicenter Study MeSH
• Geographicals
• United States epidemiology   MeSH
• Names of Substances
• Bacterial Vaccines MeSH

The treponemes infecting lagomorphs include Treponema paraluisleporidarum ecovar Cuniculus (TPeC) and ecovar Lepus (TPeL), infecting rabbits and hares, respectively. In this study, we described the first complete genome sequence of TPeL, isolate V3603-13, from an infected mountain hare (Lepus timidus) in Sweden. In addition, we determined 99.0% of the genome sequence of isolate V246-08 (also from an infected mountain hare, Sweden) and 31.7% of the genome sequence of isolate Z27 A77/78 (from a European hare, Lepus europeaus, The Netherlands). The TPeL V3603-13 genome had considerable gene synteny with the TPeC Cuniculi A genome and with the human pathogen T. pallidum, which causes syphilis (ssp. pallidum, TPA), yaws (ssp. pertenue, TPE) and endemic syphilis (ssp. endemicum, TEN). Compared to the TPeC Cuniculi A genome, TPeL V3603-13 contained four insertions and 11 deletions longer than three nucleotides (ranging between 6 and2,932 nts). In addition, there were 25 additional indels, from one to three nucleotides long, altogether spanning 36 nts. The number of single nucleotide variants (SNVs) between TPeC Cuniculi A and TPeL V3603-13 were represented by 309 nucleotide differences. Major proteome coding differences between TPeL and TPeC were found in the tpr gene family, and (predicted) genes coding for outer membrane proteins, suggesting that these components are essential for host adaptation in lagomorph syphilis. The phylogeny revealed that the TPeL sample from the European brown hare was more distantly related to TPeC Cuniculi A than V3603-13 and V246-08.

• MeSH
• Phylogeny * MeSH
• Genome, Bacterial MeSH
• Rabbits MeSH
• Syphilis * microbiology   MeSH
• Treponema * genetics   isolation & purification   MeSH
• Hares * microbiology   MeSH
• Animals MeSH
• Check Tag
• Rabbits MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH

Syphilis is a multistage sexually transmitted disease caused by Treponema pallidum ssp. pallidum. In the Czech Republic, there are around 700-800 new syphilis cases annually, continuously increasing since 2012. This study analyzed a total of 1228 samples from 2004 to 2022. Of the PCR-positive typeable samples (n = 415), 68.7% were fully-typed (FT), and 31.3% were partially-typed. Most of the identified isolates belonged to the SS14-clade and only 6.3% were the Nichols-like cluster. While in the beginning of sample collection isolates have been macrolide-susceptible, recent isolates are completely resistant to macrolides. Among the FT samples, 34 different allelic profiles (APs) were found. Most of the profiles (n = 27) appeared just once in the Czech population, while seven profiles were detected more than twice. The most frequent APs belonged to two separate groups of SS14-like isolates, including group of 1.3.1 (ST 1) and 1.26.1 (ST 25) profiles, and the second group containing 1.1.8 (ST 3), 1.1.1 (ST 2), and 1.1.3 (ST 11) (representing 57.5%, and 25.3% of all detected APs, respectively). Both groups consistently differed in 6 nucleotide positions in five genes (TP0150, TP0324, TP0515, TP0548, and TP0691) coding amino-acid replacements suggesting that one or more of these differences could be involved in the higher success of the first group.

• MeSH
• Alleles * MeSH
• Adult MeSH
• Genotype MeSH
• Middle Aged MeSH
• Humans MeSH
• Macrolides pharmacology   MeSH
• Multilocus Sequence Typing * MeSH
• Syphilis * microbiology   epidemiology   genetics   MeSH
• Treponema pallidum * genetics   isolation & purification   MeSH
• Check Tag
• Adult MeSH
• Middle Aged MeSH
• Humans MeSH
• Male MeSH
• Female MeSH
• Publication type
• Journal Article MeSH
• Geographicals
• Czech Republic MeSH
• Names of Substances
• Macrolides MeSH

Yaws is an endemic disease caused by Treponema pallidum subsp. pertenue (TPE) that primarily affects children in rural regions of the tropics. The endemic character of yaws infections and the expected exclusive reservoir of TPE in humans opened a new opportunity to start a yaws eradication campaign. We have developed a multi-locus sequence typing (MLST) scheme for TPE isolates combining the previously published (TP0548, TP0488) and new (TP0858) chromosomal loci, and we compared this typing scheme to the two previously published MLST schemes. We applied this scheme to TPE-containing clinical isolates obtained during a mass drug administration study performed in the Namatanai District of Papua New Guinea between June 2018 and December 2019. Of 1081 samples collected, 302 (28.5%) tested positive for TPE DNA, from which 255 (84.4%) were fully typed. The TPE PCR-positivity in swab samples was higher in younger patients, patients with single ulcers, first ulcer episodes, and with ulcer duration less than six months. Non-treponemal serological test positivity correlated better with PCR positivity compared to treponema-specific serological tests. The MLST revealed a low level of genetic diversity among infecting TPE isolates, represented by just three distinct genotypes (JE11, SE22, and TE13). Two previously used typing schemes revealed similar typing resolutions. Two new alleles (one in TP0858 and one in TP0136) were shown to arise by intragenomic recombination/deletion events. Compared to samples genotyped as JE11, the minor genotypes (TE13 and SE22) were more frequently detected in samples from patients with two or more ulcers and patients with higher values of specific TP serological tests. Moreover, the A2058G mutation in the 23S rRNA genes of three JE11 isolates was found, resulting in azithromycin resistance.

• MeSH
• Child MeSH
• Yaws * epidemiology   MeSH
• Genotype MeSH
• Humans MeSH
• Multilocus Sequence Typing MeSH
• Mutation MeSH
• Treponema pallidum * genetics   MeSH
• Treponema genetics   MeSH
• Ulcer MeSH
• Check Tag
• Child MeSH
• Humans MeSH
• Publication type
• Journal Article MeSH
• Geographicals
• Papua New Guinea epidemiology   MeSH

Globally, 94% of Treponema pallidum subsp. pallidum (TPA) clinical strains belong to the SS14-like group and 6% to the Nichols-like group, with a prevalence of macrolide resistance of 90%. Our goal was to determine whether local TPA strain distribution and macrolide resistance frequency have changed significantly since our last report, which revealed that Buenos Aires had a high frequency of Nichols-like strains (27%) and low levels of macrolide resistance (14%). Swab samples from patients with suspected syphilis were collected during 2015-2019 and loci TP0136, TP0548, TP0705 were sequenced in order to perform multilocus sequence typing. Strains were classified as Nichols-like or SS14-like. The presence of macrolide resistance-associated mutations was determined by examination of the 23S rDNA gene sequence. Of 46 typeable samples, 37% were classified as Nichols-like and 63% as SS14-like. Macrolide resistance prevalence was 45.7%. Seven allelic profiles were found, five were SS14-like and two were Nichols-like. The frequency of Nichols-like strains increased between studies (26.8% vs. 37%, p = 0.36). A dramatic increase was found in the frequency of macrolide resistant strains between studies (14.3% vs. 45.7%, p = 0.005). Our results are in agreement with international trends and underscore the need to pursue further TPA molecular typing studies in South America.

• MeSH
• Anti-Bacterial Agents pharmacology   MeSH
• Drug Resistance, Bacterial genetics   MeSH
• Treponemal Infections * MeSH
• Humans MeSH
• Macrolides pharmacology   MeSH
• Multilocus Sequence Typing MeSH
• DNA, Ribosomal MeSH
• Treponema pallidum * genetics   MeSH
• Treponema MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Geographicals
• Argentina epidemiology   MeSH
• Names of Substances
• Anti-Bacterial Agents MeSH
• Macrolides MeSH
• DNA, Ribosomal MeSH

BACKGROUND: Treponema pallidum subspecies pallidum (TPA) and subsp. endemicum (TEN) are the causative agents of syphilis and bejel, respectively. TEN shows similar clinical manifestations and is morphologically and serologically indistinguishable from TPA. Recently, bejel was found outside of its assumed endemic areas. Using molecular typing we aimed to discover bejel and characterize circulating TPA types among syphilis cases with Surinamese, Antillean and Dutch ethnicity in Amsterdam. METHODS: DNA was extracted from 137 ulcer swabs, which tested positive in the in-house diagnostic PCR targeting the polA gene. Samples were collected between 2006 and 2018 from Surinamese, Antillean and Dutch patients attending the Amsterdam STI clinic. Multilocus sequence typing was performed by partial sequence analysis of the tp0136, tp0548 and tp0705 genes. In addition, the 23S rRNA loci were analyzed for A2058G and A2059G macrolide resistance mutations. RESULTS: We found 17 distinct allelic profiles in 103/137 (75%) fully typed samples, which were all TPA and none TEN. Of the strains, 82.5% were SS14-like and 17.5% Nichols-like. The prevalence of Nichols-like strains found in this study is relatively high compared to nearby countries. The most prevalent types were 1.3.1 (42%) and 1.1.1 (19%), in concordance with similar TPA typing studies. The majority of the TPA types found were unique per country. New allelic types (7) and profiles (10) were found. The successfully sequenced 23S rRNA loci from 123/137 (90%) samples showed the presence of 79% A2058G and 2% A2059G mutations. CONCLUSIONS: No TEN was found in the samples from different ethnicities residing in Amsterdam, the Netherlands, so no misdiagnoses occurred. Bejel has thus not (yet) spread as a sexually transmitted disease in the Netherlands. The strain diversity found in this study reflects the local male STI clinic population which is a diverse, mixed group.

• MeSH
• Alleles MeSH
• Genes, Bacterial * MeSH
• Adult MeSH
• Ethnicity statistics & numerical data   MeSH
• Humans MeSH
• Syphilis epidemiology   ethnology   microbiology   MeSH
• DNA Barcoding, Taxonomic MeSH
• Treponema pallidum classification   genetics   pathogenicity   MeSH
• Check Tag
• Adult MeSH
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Geographicals
• Netherlands Antilles MeSH
• Netherlands MeSH
• Suriname MeSH

A recently introduced Multilocus Sequence Typing scheme for Treponema pallidum subsp. pallidum was applied to clinical samples collected from 2004 to 2017 from the two largest cities (Prague and Brno) in the Czech Republic. Altogether, a total of 675 samples were tested in this study and 281 of them were found PCR-positive for treponemal DNA and typeable. Most of the typed samples (n = 281) were swabs from primary or secondary syphilis lesions (n = 231), and only a minority were whole blood or tissue samples (n = 50). Swab samples from patients with rapid plasma regain (RPR) values of 1-1024 were more frequently PCR-positive (84.6%) compared to samples from patients with non-reactive RPR test (46.5%; p-value = 0.0001). Out of 281 typeable samples, 136 were fully-typed at all TP0136, TP0548, and TP0705 loci. Among the fully and partially typed samples, 25 different allelic profiles were identified. Altogether, eight novel allelic variants were found among fully (n = 5) and partially (n = 3) typed samples. The distribution of TPA allelic profiles identified in the Czech Republic from 2004 to 2017 revealed a dynamic character with allelic profiles disappearing and emerging over time. While the number of samples with the A2058G mutation was seen to increase (86.7% in 2016/2017), the number of samples harboring the A2059G mutation was found to have decreased over time (3.3% in 2016/2017). In addition, we found several allelic profile associations with macrolide resistance or susceptibility, the gender of patients, as well as patient residence.

• MeSH
• Alleles MeSH
• Anti-Bacterial Agents pharmacology   MeSH
• Drug Resistance, Bacterial genetics   MeSH
• DNA, Bacterial genetics   MeSH
• Adult MeSH
• Genotype MeSH
• Humans MeSH
• Young Adult MeSH
• Multilocus Sequence Typing * MeSH
• RNA, Ribosomal, 23S genetics   MeSH
• Syphilis genetics   microbiology   pathology   MeSH
• Treponema pallidum genetics   pathogenicity   MeSH
• Check Tag
• Adult MeSH
• Humans MeSH
• Young Adult MeSH
• Male MeSH
• Female MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Geographicals
• Czech Republic epidemiology   MeSH
• Names of Substances
• Anti-Bacterial Agents MeSH
• DNA, Bacterial MeSH
• RNA, Ribosomal, 23S MeSH

Treponema pallidum subsp. pallidum, the causative agent of sexually transmitted syphilis, detected in clinical samples from France, was subjected to molecular typing using the recently developed Multilocus Sequence Typing system. The samples (n = 133) used in this study were collected from 2010-2016 from patients with diagnosed primary or secondary syphilis attending outpatient centers or hospitals in several locations in France. Altogether, 18 different allelic profiles were found among the fully typed samples (n = 112). There were five allelic variants identified for TP0136, 12 for TP0548, and eight for TP0705. Out of the identified alleles, one, seven, and three novel alleles were identified in TP0136, TP0548, and TP0705, respectively. Partial allelic profiles were obtained from 6 samples. The majority of samples (n = 110) belonged to the SS14-like cluster of TPA isolates while 7 clustered with Nichols-like isolates. Patients infected with Nichols-like samples were more often older (p = 0.041) and more often diagnosed with secondary syphilis (p = 0.033) compared to patients infected with SS14-like samples. In addition, macrolide resistance caused by the A2058G mutation was found to be associated with allelic profile 1.3.1 or with strains belonging to the 1.3.1 lineage (p<0.001). The genetic diversity among TPA strains infecting the European population was surprisingly high, which suggests that additional studies are needed to reveal the full genetic diversity of TPA pathogens infecting humans.

• MeSH
• Alleles MeSH
• Biodiversity MeSH
• Child MeSH
• Adult MeSH
• Infant MeSH
• Middle Aged MeSH
• Humans MeSH
• Adolescent MeSH
• Young Adult MeSH
• Multilocus Sequence Typing MeSH
• Infant, Newborn MeSH
• Child, Preschool MeSH
• Aged MeSH
• Syphilis epidemiology   microbiology   MeSH
• Bacterial Typing Techniques MeSH
• Treponema pallidum genetics   isolation & purification   MeSH
• Check Tag
• Child MeSH
• Adult MeSH
• Infant MeSH
• Middle Aged MeSH
• Humans MeSH
• Adolescent MeSH
• Young Adult MeSH
• Male MeSH
• Infant, Newborn MeSH
• Child, Preschool MeSH
• Aged MeSH
• Female MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Geographicals
• France epidemiology   MeSH

BACKGROUND: Treponema pallidum subsp. pertenue (TPE) is the causative agent of yaws, a multi-stage disease, endemic in tropical regions of Africa, Asia, Oceania, and South America. To date, four TPE strains have been completely sequenced including three TPE strains of human origin (Samoa D, CDC-2, and Gauthier) and one TPE strain (Fribourg-Blanc) isolated from a baboon. All TPE strains are highly similar to T. pallidum subsp. pallidum (TPA) strains. The mutation rate in syphilis and related treponemes has not been experimentally determined yet. METHODOLOGY/PRINCIPAL FINDINGS: Complete genomes of two TPE strains, CDC 2575 and Ghana-051, that infected patients in Ghana and were isolated in 1980 and 1988, respectively, were sequenced and analyzed. Both strains had identical consensus genome nucleotide sequences raising the question whether TPE CDC 2575 and Ghana-051 represent two different strains. Several lines of evidence support the fact that both strains represent independent samples including regions showing intrastrain heterogeneity (13 and 5 intrastrain heterogeneous sites in TPE Ghana-051 and TPE CDC 2575, respectively). Four of these heterogeneous sites were found in both genomes but the frequency of alternative alleles differed. The identical consensus genome sequences were used to estimate the upper limit of the yaws treponeme evolution rate, which was 4.1 x 10-10 nucleotide changes per site per generation. CONCLUSIONS/SIGNIFICANCE: The estimated upper limit for the mutation rate of TPE was slightly lower than the mutation rate of E. coli, which was determined during a long-term experiment. Given the known diversity between TPA and TPE genomes and the assumption that both TPA and TPE have a similar mutation rate, the most recent common ancestor of syphilis and yaws treponemes appears to be more than ten thousand years old and likely even older.

• MeSH
• Time Factors MeSH
• Escherichia coli genetics   MeSH
• Yaws epidemiology   microbiology   MeSH
• Genome, Bacterial * MeSH
• Humans MeSH
• Chromosome Mapping MeSH
• Mutation MeSH
• Papio microbiology   MeSH
• Sequence Analysis, DNA MeSH
• Treponema pallidum classification   genetics   isolation & purification   MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Geographicals
• Asia epidemiology   MeSH
• Ghana epidemiology   MeSH
• South America epidemiology   MeSH