Establishment Of Pulse-Field Gel Electrophoresis Procedure In Bartonella And Application In Bartonella Epidemiology

Pulsed-field gel electrophoresis has been invented by Schwartz and Canter in 1984 and eventually became a key molecular biological technique after a series of development.Pulsed-field gel electrophoresis has been widely used in the study of bacteria and has shown its strong value in controlling diseases outbreak.It not only can be used to track the source of infection alone,but also can support the relationship of bacteria different species.Now it becomes an essential tool for tracking the source of disease.Recently,we found some species in Bartonella can caused different disease and its have strong correlation.In other words,different kinds of Bartonella can lead to different disease.In addition,whether different genotypes strains can lead to different diseases need to do further studies.In such requirment,we need one technique which is a simple,reliable and has strong ability to distinguish species of Bartonella.This study has established PFGE procedure of Bartonella by bacteria concentration, selected the appropriate digestion enzyme,optimized the electrophoretic parameters and characterized reproducibly two Bartonella species strains.Its parameters are as following:bacterium concentration≧ 4.5(turbidity instrument readings),restriction endonuclease preferred SmaI,electrophoresis parameters parameters(EP-C).In addition, we found that Bartonella henselae has 20%probability of occurrence of the loss of bands by two passages and B.elizabethae found no variation.This study has analyzed the 52 strains of Bartonella from Shandong,Beijing,Henan province and get a cluster of these strains by Bionumerics database software.The result indicates that PFGE can be used to distinguish species.In addition,epidemiological analysis of Bartonella from cats and dogs discovered that Bartonella genotypes are different and presumably are the different species of Bartonella for less than 60%homology,.Through GltA gene amplification and sequencing,the study discovered that isolates from dogs and cats are clusted to two clusters and do not cross each other.In addition,Bartonella henselae has high homology with the strains from cats.All of these results are consistent with the results of PFGE.To sum up,we have established the PFGE of Bartonella and used to distinguish Bartonella species through its cluster analysis.We discovered Bartonella from cats and dogs may be different species.In addition,the majority of Bartonella may be Bartonella henselae in cats and distributions of genotypes in three region are different by statistic analysis.