The Damage In A Murine Model And Phylogeography Of Orientia Tsutsugamushi

【Objective】To understand the molecular characteristics of the 56 k Da outer membrane protein of Orientia tsutsugamushi(Ot)isolated from rodents and humans.Pathological changes caused by Ot mouse infection model.The Chinese and global phylogeographic reconstruction of Ot provides scientific guidance for the control of scrub typhus.【Methods】From August 2020 to August 2021,samples of rodents and suspected scrub typhus patients were collected in Dali City,Yunnan Province,China.The Ot conserved gene was amplified by nested PCR.Positive samples from rodents and different genotypes of Ot were screened out through clinical data and molecular experiments.Ot was isolated by intraperitoneal inoculation of mice.The isolated Ot was identified by Giemsa staining and indirect immunoassay.Primers were designed to amplify the full-length 56 k Da type-specific antigen(TSA)gene of Ot.Using MEGA X,DNAStar 7.1,Seq Man,Tree Time and other bioinformatics software,the homology,genetic evolution,differentiation time and mutation of the Ot sequence obtained in this study and the national and global sequences were systematically analyzed.【Results】A total of 76 rodents of 5 genera and 6 species were captured,and the Ot sequence was amplified from the tissue grinding liquid of one Rattus tanezumi,with a positive rate of2.38%(1/42).The strain Ot was isolated by intraperitoneal inoculation of mice,and was identified as TA763 type by molecular identification.The full-length nucleotide sequence of the 56 k Da TSA gene was 1566 bp.Among the 108 suspected scrub typhus patients,the positive rate was 70.37%(76/108).The three genotypes of Karp,TA763,Kawasaki isolated from the patient were evolutionarily similar with 95.60% to 99.94% to the global isotype Ot.Mice were intraperitoneally inoculated with an infectious dose of Ot after 15 days,it was observed that the liver and spleen of the experimental group were significantly larger than those of the control group.There was no difference in aspartate aminotransferase and alanine aminotransferase among the three genotypes of Ot,and they were significantly higher than those in the control group.Hepatocyte edema can be seen in the liver tissue,typical hepatocyte ballooning.Around the capillaries and bile ducts,there are a large number of lymphocytes.Immunohistochemistry showed hypersplenism and accumulation of Ot.In the heart,lungs,kidneys,and brain,the predominant manifestation is interstitial inflammation.Phylogeographic reconstruction results showed that Taiwan Province was identified as a major transmission hub for scrub typhus within China.Globally,China was identified as the transmission center of scrub typhus and maintained the most genotypes.【Conclusion】(1)No differences in molecular 56 k Da of Ot were found between hosts.(2)Inflammatory cell infiltration was found in several organs in a mouse model of Ot infection.(3)The consistency of the 56 k Da conserved region is one of the reasons why the same pathological changes occur in scrub typhus infections providing a theoretical basis for our future development of a scrub typhus vaccine.(4)China is an important hub for the transmission of scrub typhus and Yunnan is the second hub for the transmission of scrub typhus in China.