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Chlamydia trachomatis (C. trachomatis), an obligate intracellular parasite, is a type of prokaryotic microbes with a special life cycle. C. trachomatis infection in human would cause a series of diseases, among which the urogenital infection by serotypes D-K is the most common sexually transmitted disease. Though producing no obvious clinical manifestations, the infection in women specifically damages the columnar epithelial cells in reproductive tract, and is correlated with vaginitis, cervicitis, endometritis, salpingitis, pelvic inflammatory disease, infertility, and abortion. The incidence of reproductive tract infection (RTI) by C. trachomatis in the world is about 93 million per year, and the incidence in China is rising year by year, making it a serious public health concern.Studies on animal models of C. trachomatis caused RTI could contribute to the understanding of its pathological mechanisms. Given the highest detection rate of serotype E in clinical practices, this study established a mice model of serotype E caused RTI. In vitro amplification of C. trachomatis was first conducted with HeLa229 cells to determine the time-effect relationship. After infecting BALB/C mice through reproductive tract perfusion with low, medium, and high doses, we recorded a number of indexes at 5 time points (3,7,14,21,28 days after the infection), including body weight and other clinical indicators, pathological test findings, isolation of chlamydia from vaginal and cervical swab samples, quantitative analysis of nucleic acids of cervical tissue samples, and measurement of serum antibody and cytokines such as IFN-y, IL-2, and IL-10, in order to establish a standardized model evaluation system.Class 1 outer membrane protein gene (OMP1 gene) is a chlamydia specific gene, which was targeted in the present study for the measurement of chlamydia in infected tissues, and for revealing the correlation between pathogen replication and other indexes. DNA of tissue samples was extracted with protein cleavage and salt precipitation. A real time PCR system was established with the interference of mouse genome excluded to quantify the exact amount of chlamydia in the tissue samples. This method improves the sensitivity and specificity of chlamydia detection.In this study, we first determined the in vitro replication pattern of chlamydia, prepared chlamydia with high purity for infection. After infection with three different doses, chlamydia was isolated from cervical swabs of all the infected mice. The results of nucleic acid quantification identified the replication and proliferation of chlamydia in infected tissues. Clinical indicators and pathological analysis confirmed the clinical and pathological changes of the infection. The analysis of the peripheral blood of the infected mice revealed significantly increased anti-chlamydia antibodies in the first 2 weeks after infection, after that the antibody level remained stable. The levels of IFN-y, IL-2, and IL-10 showed that immune response against pathogens and changes of related factors occurred in mice infected by chlamydia.This study established a stable mouse model of lower reproductive tract infection by human C. trachomatis. The medium and high doses produce much similar etiological and pathological indexes in mice as in human, with cervicitis being the major manifestation, accompanied with mild vaginitis. However, the immunological indexes are different from those in infected human, since only short-term lower reproductive tract infection is induced in mice because of the quicker spontaneous clearance.In the present study, we established the methods for in vitro amplification of chlamydia, nucleic acid quantification, infection vaccination, sample collection after infection, and detection of etiological, pathological, and immunological indicators after infection. We also analyzed the impact of different doses on those indicators, and primarily developed a standardized operating procedure, which may provide some helpful information for other types of chlamydia caused RTI models in the future.
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