Heinzerling H F, Olivares M, Burne R A
Department of Dental Research, University of Rochester, New York 14642, USA.
Infect Immun. 1997 Jun;65(6):2041-51. doi: 10.1128/iai.65.6.2041-2051.1997.
Oral spirochetes possess many potential virulence factors, including the capacity for tissue invasion and persistence despite a vigorous host immune response. In an attempt to identify treponemal immunoreactive components, sera derived from individuals with advanced periodontal disease were used as a reagent to isolate recombinant bacteriophage lambda clones expressing antigens of the oral spirochete Treponema denticola ATCC 35405. Nucleotide sequence analysis of a clone expressing three immunoreactive products has revealed seven T. denticola genes which appear to encode homologs of flagellar basal body constituents, FlgB, FlgC, FliE, and FliF, a flagellar switch component, FliG, and the putative flagellar export proteins, FliH and FliI, initially characterized in Salmonella typhimurium. Also identified was a gene resembling fliJ. Primer extension analysis identified a transcriptional start site 5' to the treponemal flgB gene. Appropriately spaced with respect to this start site was a sigma28 binding motif. The absence of additional identifiable sigma factor binding motifs within the treponemal sequence and the proximity of adjacent genes suggested operonic arrangement, and reverse transcriptase PCR provided evidence of cotranscription. Supporting the identification of these genes as flagellar components, heterologous expression in enteric bacteria of the putative switch basal body genes from T. denticola interfered with motility. Specifically, the presence of a plasmid expressing treponemal fliG reduced swarming motility in S. typhimurium, while in Escherichia coli, this plasmid conferred a nonmotile phenotype and a reduction in flagellar number. Thus, while spirochetal flagella are subject to unique synthetic and functional constraints, the organization of flagellar genes and the presence of sigma28-like elements are reminiscent of the flagellar systems of other bacteria, and there appears to be sufficient conservation of constituent proteins to allow interaction between T. denticola switch-basal body proteins and the flagellar machinery of gram-negative bacteria.
口腔螺旋体具有许多潜在的毒力因子，包括组织侵袭能力以及即便在宿主免疫反应强烈时仍能持续存在的能力。为了鉴定密螺旋体的免疫反应成分，从患有晚期牙周病的个体中获取的血清被用作试剂，以分离表达口腔螺旋体齿垢密螺旋体ATCC 35405抗原的重组λ噬菌体克隆。对一个表达三种免疫反应产物的克隆进行核苷酸序列分析，揭示了七个齿垢密螺旋体基因，这些基因似乎编码鞭毛基体成分FlgB、FlgC、FliE和FliF的同源物、一个鞭毛开关成分FliG以及推定的鞭毛输出蛋白FliH和FliI，这些蛋白最初是在鼠伤寒沙门氏菌中鉴定出来的。还鉴定出一个类似于fliJ的基因。引物延伸分析确定了齿垢密螺旋体flgB基因5'端的一个转录起始位点。相对于该起始位点，有一个适当间隔的σ28结合基序。在齿垢密螺旋体序列中没有其他可识别的σ因子结合基序，且相邻基因距离较近，这表明存在操纵子排列，逆转录酶PCR提供了共转录的证据。将齿垢密螺旋体推定的开关基体基因在肠道细菌中进行异源表达会干扰运动性，这支持了将这些基因鉴定为鞭毛成分的结论。具体而言，表达齿垢密螺旋体fliG的质粒的存在会降低鼠伤寒沙门氏菌的群体运动性，而在大肠杆菌中，该质粒赋予了非运动表型并减少了鞭毛数量。因此，虽然螺旋体鞭毛受到独特的合成和功能限制，但鞭毛基因的组织以及σ28样元件的存在让人联想到其他细菌的鞭毛系统，并且组成蛋白似乎有足够的保守性，使得齿垢密螺旋体开关基体蛋白与革兰氏阴性菌的鞭毛机制之间能够相互作用。
