Fikrig E, Chen M, Barthold S W, Anguita J, Feng W, Telford S R, Flavell R A
Department of Internal Medicine, Yale University School of Medicine, New Haven, CT 06520, USA.
Mol Microbiol. 1999 Jan;31(1):281-90. doi: 10.1046/j.1365-2958.1999.01171.x.
The expression of a Borrelia burgdorferi gene, erpT, was investigated throughout the spirochaete life cycle in the arthropod vector and the murine host. Three phage clones from a B. burgdorferi DNA expression library synthesized a 30 kDa antigen that was recognized by antibodies in the sera of B. burgdorferi-infected mice but not mice hyperimmunized with B. burgdorferi lysates. Differential antibody binding suggested that this protein was preferentially expressed in vivo. This antigen was designated ErpT, based upon 99.6% homology with the BBF01 sequence in the B. burgdorferi genome. ErpT was not detected on spirochaetes cultured in BSK II medium by indirect immunofluorescence or in B. burgdorferi lysates by immunoblotting, implying that ErpT is not readily produced in vitro. erpT mRNA was not discernible by Northern blot but was identified by RNA polymerase chain reaction in vitro, indicating that erpT is expressed at low levels by cultured spirochaetes. erpT expression was then investigated in the vector and mice because B. burgdorferi do not normally reside in culture medium. RNA polymerase chain reaction and immunofluorescence studies demonstrated that erpT was expressed by a small minority of B. burgdorferi (11/500, 2.2%) within unfed ticks and then repressed during engorgement. erpT mRNA or ErpT antibodies were first detected in B. burgdorferi-infected mice at 4 weeks, suggesting that erpT was not expressed in the early stages of murine infection. Then, during persistent infection, RNA polymerase chain reaction showed that erpT was expressed by B. burgdorferi within the joints, heart and spleen, but not by spirochaetes in the skin. Immunization of mice with ErpT was antigenic but was not protective. These studies demonstrate that B. burgdorferi erpT is differentially expressed throughout the B. burgdorferi life cycle, in both the vector and the mammalian host, and is primarily expressed in extracutaneous sites during murine infection.
对伯氏疏螺旋体基因erpT在节肢动物媒介和小鼠宿主的螺旋体生命周期中的表达进行了研究。来自伯氏疏螺旋体DNA表达文库的三个噬菌体克隆合成了一种30 kDa的抗原,该抗原能被伯氏疏螺旋体感染小鼠的血清中的抗体识别,但不能被用伯氏疏螺旋体裂解物进行超免疫的小鼠的血清中的抗体识别。抗体结合差异表明该蛋白在体内优先表达。基于与伯氏疏螺旋体基因组中BBF01序列99.6%的同源性,该抗原被命名为ErpT。通过间接免疫荧光在BSK II培养基中培养的螺旋体上未检测到ErpT,通过免疫印迹在伯氏疏螺旋体裂解物中也未检测到,这意味着ErpT在体外不易产生。通过Northern印迹法未检测到erpT mRNA,但通过体外RNA聚合酶链反应鉴定到了,表明培养的螺旋体中erpT表达水平较低。由于伯氏疏螺旋体通常不存在于培养基中,因此随后对其在媒介和小鼠中的表达进行了研究。RNA聚合酶链反应和免疫荧光研究表明,在未进食的蜱中,一小部分伯氏疏螺旋体(11/500,2.2%)表达erpT,但在饱食过程中受到抑制。在感染伯氏疏螺旋体的小鼠中,在4周时首次检测到erpT mRNA或ErpT抗体,这表明erpT在小鼠感染的早期阶段不表达。然后,在持续感染期间,RNA聚合酶链反应表明,关节、心脏和脾脏中的伯氏疏螺旋体表达erpT,但皮肤中的螺旋体不表达。用ErpT免疫小鼠具有抗原性,但无保护作用。这些研究表明,伯氏疏螺旋体erpT在伯氏疏螺旋体的整个生命周期中,在媒介和哺乳动物宿主中均有差异表达,并且在小鼠感染期间主要在皮肤外部位表达。