Dataset Information

Characterization of Treponema pallidum Dissemination in C57BL/6 Mice.

ABSTRACT: The spirochetal pathogen Treponema pallidum causes 5 million new cases of venereal syphilis worldwide each year. One major obstacle to syphilis prevention and treatment is the lack of suitable experimental animal models to study its pathogenesis. Accordingly, in this study, we further evaluated the responses of mice to Treponema pallidum. Quantitative polymerase chain reaction showed that Treponema pallidum could colonize the heart, liver, spleen, kidneys, and testicles of C57BL/6 mice, and the organism may be able to rapidly penetrate the blood-brain barrier in mice by 24 h after infection. In subsequent rabbit infectivity tests, we observed evident signs of the microorganism in the mouse lymph node suspension. After infection, bacterial loads were higher in the tissues than in the blood of C57BL/6 mice. Moreover, a significant Th1 immune response was recorded by cytokine assays. Flow cytometric analysis suggested an obvious increase in the proportion of CD3⁺ T and CD4⁺ T cells in the spleen cells in the infected mice. Thus, improving our understanding of the response of C57BL/6 mice for Treponema pallidum will help to comprehensive elucidate the pathogenic mechanisms of this bacterium and lay the foundation for the development of a new research model of Treponema pallidum.

SUBMITTER: Lu S

PROVIDER: S-EPMC7819853 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature

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Characterization of Treponema pallidum Dissemination in C57BL/6 Mice.

Lu Simin S Zheng Kang K Wang Jianye J Xu Man M Xie Yafeng Y Yuan Shuai S Wang Chuan C Wu Yimou Y

Frontiers in immunology 20210108

The spirochetal pathogen Treponema pallidum causes 5 million new cases of venereal syphilis worldwide each year. One major obstacle to syphilis prevention and treatment is the lack of suitable experimental animal models to study its pathogenesis. Accordingly, in this study, we further evaluated the responses of mice to Treponema pallidum. Quantitative polymerase chain reaction showed that Treponema pallidum could colonize the heart, liver, spleen, kidneys, and testicles of C ...[more]

PMID: 33488577

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Project description:Despite more than a century of research, genetic manipulation of Treponema pallidum subsp. pallidum (T. pallidum), the causative agent of syphilis, has not been successful. The lack of genetic engineering tools has severely limited understanding of the mechanisms behind T. pallidum success as a pathogen. A recently described method for in vitro cultivation of T. pallidum, however, has made it possible to experiment with transformation and selection protocols in this pathogen. Here, we describe an approach that successfully replaced the tprA (tp0009) pseudogene in the SS14 T. pallidum strain with a kanamycin resistance (kanR) cassette. A suicide vector was constructed using the pUC57 plasmid backbone. In the vector, the kanR gene was cloned downstream of the tp0574 gene promoter. The tp0574prom-kanR cassette was then placed between two 1-kbp homology arms identical to the sequences upstream and downstream of the tprA pseudogene. To induce homologous recombination and integration of the kanR cassette into the T. pallidum chromosome, in vitro-cultured SS14 strain spirochetes were exposed to the engineered vector in a CaCl2-based transformation buffer and let recover for 24 hours before adding kanamycin-containing selective media. Integration of the kanR cassette was demonstrated by qualitative PCR, droplet digital PCR (ddPCR), and whole-genome sequencing (WGS) of transformed treponemes propagated in vitro and/or in vivo. ddPCR analysis of RNA and mass spectrometry confirmed expression of the kanR message and protein in treponemes propagated in vitro. Moreover, tprA knockout (tprAko-SS14) treponemes grew in kanamycin concentrations that were 64 times higher than the MIC for the wild-type SS14 (wt-SS14) strain and in infected rabbits treated with kanamycin. We demonstrated that genetic manipulation of T. pallidum is attainable. This discovery will allow the application of functional genetics techniques to study syphilis pathogenesis and improve syphilis vaccine development.

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Characterization of Treponema pallidum Dissemination in C57BL/6 Mice.

Publications

Characterization of <i>Treponema pallidum</i> Dissemination in C57BL/6 Mice.

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