Analysis Of Molecular Properties Of Leptospira

Leptospirosis is one of the most popular global zoonotic diseases caused by pathogenic species of the Leptospira genus. The clinical symptoms of Leptospirosis in humans consist of fever, sore and ache all over, weakness, conjunctival congestion, and lymph nodes swelling. Although most patients can be recovered, some severe cases with multiple organ failure have been reported which can cause death fianlly. At present, leptospira is mainly classified by traditional serological method and molecular genetic approaches. Notwithstanding the serological method has been widely applied in the past decades, it is found that certain cross reaction exists between different Leptospira serogroups or serovars because those sera used are mostly polyclonal antibodies,. With the development of biotechnology, various molecular typing methods have been used for epidemiology of Leptospira in many countries.In this study,16S rRNA, multilocus sequence analysis (MLST), and pulsed field gel electrophoresis (PFGE) methods were used to analyze the 73 Chinese Leptospira strains and 63 international strains from National Institutes for Food and Drug Control (NIFDC). 16S rRNA gene sequence analysis was performed to identify the geno-species for various Leptospria serogroup strains. Total 85 strains were pathogenic Leptospira species, which was comprised of 8 distinct species. Then 7 housekeeping gene required by MLST were amplified using polymerase chain reaction (PCR) and sequenced. The sequence types (STs) of pathogenic species were identified and total 47 ST were found, of which 32 STs were from China, suggesting molecular epidemic trend in China was different from other countries. Furthermore, the chromosomal DNA of Leptospira strains were also performed by using restriction enzymes Not I digestion and PFGE analysis. The results showed that distinct DNA fragments from 20 KB to 700 KB size were separated, and the number, size and distribution characteristic of DNA fragments were obviously different among Leptospira strains studied. PFGE patterns of Leptospira strains were clustered using the unweighted pair group with arithmetic averages (UPGMA) of Bionumerics software, the similarity coefficients ranged from 28.50% to 90.91% among Leptospira strains studied.In general, we obtained the first-hand data on molecular characteristics information of Leptospira species circulating in China, and analyzed the difference between Leptospira strains from different countries at molecular level by using MLST and PFGE for typing Leptospira species. Furthermore, such molecular tools used in the study were proved to be reliable and reproducible. These data will provide the basis on tracking source, determining media and timely controlling epidemic situation, and can also offer some references for clinical diagnosis, treatment, as well as the development of the vaccine.