Yigong Ge

Date of Graduation


Document Type



Motility and chemotaxis have been considered as important candidates for the study of virulence in many pathogenic spirochetes, including B. burgdorferi. Little is known about the underlying mechanisms of motility and chemotaxis. In this study, we have carried out an extensive search for the motility and chemotaxis genes in B. burgdorferi. The unique technique of "semi-random PCR chromosome walking" was developed and used to clone several large motility and chemotaxis gene clusters. Together with the previously reported flaB and flgE genes, 38 motility and chemotaxis related genes (more than 36 kb) were identified. These genes demonstrated extensive homology with their bacterial counterparts, especially with those from Treponema and Bacillus subtilis. Motility and chemotaxis genes constitute four major clusters on the chromosome of B. burgdorferi. Transcript analyses using RT-PCR and primer extension revealed that at least five operons were present in these motility and chemotaxis clusters; they were flgB, flgK, flaB, flaA/che, and fliD operons. The conserved {dollar}\\sigma\\sp{lcub}70{rcub}{dollar}-like promoters were involved in all of these operons, which is distinct to those of other bacteria. These results indicate that although the motility and chemotaxis genes in B. burgdorferi are well conserved, the transcriptional regulation of flagellar gene expression in B. burgdorferi is different from those of other bacteria. To further analyze these motility and chemotaxis genes, we overexpressed several motility and chemotaxis proteins in E. coli. Serological analyses using recombinant motility and chemotaxis proteins (FlgE, FliG, FliI, CheY and FlaA) indicate that except for FlgB and FlgE, these proteins were not immunodominant antigens during infection. Thus, they are not good candidates for diagnostic purposes. In order to further study the functions of these genes, we made an extensive effort to develop a gene inactivation system. Several different antibiotic constructs (gyrB-coumermycin resistant gene, cat and kanamycin resistant genes) were inserted into several motility genes. Unfortunately, so far none of these constructs worked successfully in B. burgdorferi. This study is the first important step toward the achievement of a better understanding of the genetics of spirochetal motility and chemotaxis.