Early syphilis in Guangzhou, China: presentation, molecular detection of Treponema pallidum, and genomic sequences in clinical specimens and isolates obtained by rabbit infectivity testing

Background: The global resurgence of syphilis requires novel prevention strategies. Whole genome sequencing (WGS) of Treponema pallidum (TPA) using different specimen types is essential for vaccine development. Methods: Patients with primary (PS) and secondary (SS) syphilis were recruited in Guangzhou, China. We collected ulcer exudates and blood from PS participants, and skin biopsies and blood from SS participants for TPA polA polymerase chain reaction (PCR); ulcer exudates and blood were also used to isolate TPA strains by rabbit infectivity testing (RIT). TPA WGS was performed on 52 ulcer exudates and biopsy specimens and 25 matched rabbit isolates. Results: We enrolled 18 PS and 51 SS participants from December 2019 to March 2022. Among PS participants, TPA DNA was detected in 16 (89%) ulcer exudates and three (17%) blood specimens. Among SS participants, TPA DNA was detected in 50 (98%) skin biopsies and 27 (53%) blood specimens. TPA was isolated from 48 rabbits, with a 71% (12/17) success rate from ulcer exudates and 69% (36/52) from SS bloods. Twenty-three matched SS14 clade genomes were virtually identical, while two Nichols clade pairs had discordant tprK sequences. Forty-two of 52 unique TPA genomes clustered in an SS14 East Asia subgroup, while ten fell into two East Asian Nichols subgroups. Conclusions: Our TPA detection rate was high from PS ulcer exudates and SS skin biopsies and over 50% from SS whole blood, with RIT isolation in over two-thirds of samples. Our results support the use of WGS from rabbit isolates to inform vaccine development. ### Competing Interest Statement The authors have declared no competing interest. ### Clinical Protocols ### Funding Statement This study was funded by the US National Institutes of Health National Institute for Allergy and Infectious Disease (NIAID) (U19AI144177 to JDR and MAM), the Bill and Melinda Gates Foundation (INV-036560 to ACS), strategic research dollars from Connecticut Childrens, NIAID (T32 T32AI007151 to FFA), the National Nature Science Foundation of China (82220108006), the National Institute of Virology and Bacteriology (Program EXCELES, ID Project No. LX22NPO5103, Funded by the European Union - Next Generation EU to DS). ### Author Declarations I confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained. Yes The details of the IRB/oversight body that provided approval or exemption for the research described are given below: This research was approved by the Medical Ethics Committee Dermatology Hospital of Southern Medical University (GDDHLS-20181202). Rabbit experimentation was approved by the Ethics Committee of Dermatology Hospital of Southern Medical University (approval no. GDDHLS-20181202,12/12/2018) and South China Agricultural University Experimental Animal Ethics Committee (approval no. 2020C004, 07/05/2020). All rabbits were housed under approved biosafety conditions at South China Agricultural University. Their diet, care, and maintenance conformed to institutional regulations. I confirm that all necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived, and that any patient/participant/sample identifiers included were not known to anyone (e.g., hospital staff, patients or participants themselves) outside the research group so cannot be used to identify individuals. Yes I understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance). Yes I have followed all appropriate research reporting guidelines, such as any relevant EQUATOR Network research reporting checklist(s) and other pertinent material, if applicable. Yes All data produced in the present work are contained in the manuscript