Isolation, passage and culture of one clinical strain of Treponema pallidum in Guangdong province and the detection of its nonsynonymous single nucleotide polymorphisms

Objective To isolate and culture a clinical strain (GD1) of Treponema pallidum (Tp) in Guangdong province, and to investigate the difference in nonsynonymous single nucleotide polymor-phisms (nsSNPs) between the GD1 strain and Tp Nichols strain. Methods The GD1 strain was isolated from the hard chancre in a patient with primary syphilis in Guangdong province, and continuously subcultured in the testes of New Zealand white rabbit. The serial subcultivation of GD1 was multi-verified by dark-field microscopy, polymerase chain reaction (PCR) for TP0548 gene, DNA sequencing and genotyping. Meglumine diatrizoate density gradient centrifugation was performed to isolate rabbit tissues and concentrate GD1, and DNA sequencing was used to verify the nsSNPs in the TP0443 and TP0584 genes. Results The GD1 strain was successfully isolated from the lesions of the patient with syphilis, and classified as a subtype f of TP0548. Compared with the American Tp (Nichols strain) , there were nsSNP mutations in the GD1 strain. One mutation was located in the TP0443 gene, leading to the the substitution of threonine by alanine at amino acid position 120, and another one was located in the TP0584 gene, which caused a change from alanine to threonine at amino acid position 314. Conclusion The GD1 strain was successfully isolated firstly from the lesions in a patient with syphilis in Guangdong province, and nsSNP mutations were confirmed in the GD1 strain on the etiology. Key words: Syphilis; Treponema pallidum; Polymorphism, single nucleotide; TP0443 gene; TP0584 gene