Molecular Mechanisms Of Variation And Adaptation In Borr

伯氏疏螺旋体是莱姆病的病原体，在自然界中是通过野生哺乳动物和扁虱之间的感染循环维持的。像许多细菌病原体一样，伯氏杆菌必须应对一系列不断变化的环境条件，才能成功地持续、增殖并在宿主之间传播。我们的主要目标是用分子遗传学方法阐明伯氏杆菌适应和变异的分子机制及其在感染循环中的作用。布氏硬蜱在其自然生命周期的很大一部分时间里都栖息在硬蜱的中肠内。扁虱媒介经历了许多生理和代谢变化，驻留的细菌暴露在这些变化中。硬体硬壳层在取食和蜕皮准备过程中的膨胀需要合成新的角质层，N-乙酰氨基葡萄糖的聚合物甲壳素是角质层的组成部分。伯氏杆菌的基因组序列揭示了几个可能促进细菌利用甲壳素副产物的基因，包括甲壳二糖(甲壳素的二聚体亚单位)转运体的同源基因。我们已经开始了一项研究，以确定这些基因产物是否促进了壳二糖的利用，并有助于扁虱中的细菌生长。我们发现，在细菌的培养基中，壳二糖可以有效地替代N-乙酰氨基葡萄糖。我们灭活了编码转运蛋白跨膜组件的基因，并确定这些突变细菌无法利用壳二糖。我们还发现，转运蛋白基因受生长温度的调节，这与以前的结果一致，表明温度是调节扁虱和哺乳动物之间差异基因表达的重要环境变量。对伯氏假单胞菌利用壳二糖的遗传和调控进行了初步研究。我们最近在一个感染性克隆中灭活了壳二糖转运蛋白。我们已经开始了一系列的实验，以比较突变和互补形式与其同基因的野生型亲本在自然老鼠-壁虱感染循环中的差异。这种实验直到最近才成为可能，因为我们在致病伯氏杆菌的基因操作方面取得了进展。

Borrelia burgdorferi, the causative agent of Lyme disease, is maintained in nature through an infectious cycle between wild mammals and ticks. Like many bacterial pathogens, B. burgdorferi must cope with an array of changing environmental conditions to successfully persist, proliferate and be transmitted between hosts. Our broad objective is to use a molecular genetic approach to elucidate the molecular mechanisms of adaptation and variation in B. burgdorferi and their roles in the infectious cycle. B. burgdorferi resides in the midgut of an ixodid tick for a significant part of its natural life cycle. The tick vector undergoes a number of physiological and metabolic changes to which resident bacteria are exposed. Expansion of the ixodid tick integument during feeding and preparation for molting requires the synthesis of new cuticle, of which chitin, a polymer of N-acetyl glucosamine, is a component. The genome sequence of B. burgdorferi revealed several genes likely to facilitate chitin by-product utilization by the bacteria, including homologs of a transporter for chitobiose (the dimer subunit of chitin). We have begun a study to determine if these gene products facilitate chitobiose utilization and contribute to bacterial growth in ticks. We found that chitobiose efficiently substitutes for N-acetyl glucosamine during bacterial growth in culture medium. We inactivated the gene encoding the membrane - spanning component of the transporter and determined that these mutant bacteria were unable to utilize chitobiose. We also found that the transporter gene was regulated in response to growth temperature, consistent with previous results demonstrating temperature as an important environmental variable regulating differential gene expression between ticks and mammals. Initial studies on the genetics and regulation of chitobiose utilization in B. burgdorferi were conducted with an attenuated, nonpathogenic clone. We have recently inactivated the chitobiose transporter in an infectious clone. We have begun a series of experiments to compare the mutant and complemented form with its isogenic wild type parent in the natural mouse-tick infectious cycle. Such experiments have only recently become possible because of advances we have made in the genetic manipulation of pathogenic B. burgdorferi.