Department of Biology, Mt. Allison University.
Department of Molecular Biology, Umeå University.
J Vis Exp. 2022 Nov 25(189). doi: 10.3791/64431.
The Borrelia consists of three groups of species, those of the Lyme borreliosis (LB) group, also known as B. burgdorferi sensu lato (s.l.) and recently reclassified into Borreliella, the relapsing fever (RF) group Borrelia, and a third reptile-associated group of spirochetes. Culture-based methods remain the gold standard for the laboratory detection of bacterial infections for both research and clinical work, as the culture of pathogens from bodily fluids or tissues directly detects replicating pathogens and provides source material for research. Borrelia and Borreliella spirochetes are fastidious and slow growing, and thus are not commonly cultured for clinical purposes; however, culture is necessary for research. This protocol demonstrates the methodology and recipes required to successfully culture LB and RF spirochetes, including all recognized species from B. burgdorferi s.l. complex including B. afzelii, B. americana, B. andersonii, B. bavariensis, B. bissettii/bissettiae, B. burgdorferi sensu stricto (s.s.), B. californiensis, B. carolinensis, B. chilensis, B. finlandensis, B. garinii, B. japonica, B. kurtenbachii, B. lanei, B. lusitaniae, B. maritima, B. mayonii, B. spielmanii, B. tanukii, B. turdi, B. sinica, B. valaisiana, B. yangtzensis, and RFspirochetes, B. anserina, B. coriaceae, B. crocidurae, B. duttonii, B. hermsii, B. hispanica, B. persica, B. recurrentis, and B. miyamotoi. The basic medium for growing LB and RF spirochetes is the Barbour-Stoenner-Kelly (BSK-II or BSK-H) medium, which reliably supports the growth of spirochetes in established cultures. To be able to grow newly isolated Borrelia isolates from tick- or host-derived samples where the initial spirochete number is low in the inoculum, modified Kelly-Pettenkofer (MKP) medium is preferred. This medium also supports the growth of B. miyamotoi. The success of the cultivation of RF spirochetes also depends critically on the quality of ingredients.
伯氏疏螺旋体由三个物种群组成，即莱姆病（LB）群，也称为伯氏疏螺旋体 sensu lato（s.l.），最近重新分类为 Borreliella；回归热（RF）群疏螺旋体和第三个与爬行动物相关的螺旋体群。基于培养的方法仍然是用于研究和临床工作的细菌感染实验室检测的金标准，因为从体液或组织中培养病原体可以直接检测到复制的病原体，并为研究提供源材料。伯氏疏螺旋体和 Borreliella 螺旋体是苛刻和缓慢生长的，因此通常不用于临床培养；然而，培养对于研究是必要的。本方案展示了成功培养 LB 和 RF 螺旋体所需的方法和配方，包括伯氏疏螺旋体 s.l. 复合体中所有已识别的物种，包括阿氏疏螺旋体、美洲疏螺旋体、安德森氏疏螺旋体、巴伐利亚疏螺旋体、Bissettii/bissettiae、伯氏疏螺旋体严格意义上（s.s.）、加利福尼亚疏螺旋体、卡罗莱纳疏螺旋体、智利疏螺旋体、芬兰疏螺旋体、加氏疏螺旋体、日本疏螺旋体、库滕巴赫氏疏螺旋体、Lane 氏疏螺旋体、卢氏疏螺旋体、葡萄牙疏螺旋体、海洋疏螺旋体、Mayonii、spielmanii、tanukii、Turdi、中华疏螺旋体、瓦莱州疏螺旋体、扬氏疏螺旋体和 RF 螺旋体、anserina、coriaceae、crocidurae、duttonii、hermsii、hispanica、persica、recurrentis 和 miyamotoi。生长 LB 和 RF 螺旋体的基本培养基是 Barbour-Stoenner-Kelly（BSK-II 或 BSK-H）培养基，该培养基可可靠地支持已建立的培养物中螺旋体的生长。为了能够从蜱或宿主来源的样本中生长新分离的伯氏疏螺旋体分离株，其中接种物中的初始螺旋体数量较低，首选改良 Kelly-Pettenkofer（MKP）培养基。该培养基也支持 miyamotoi 的生长。RF 螺旋体的培养成功也严重依赖于成分的质量。
