Kingry Luke C, Batra Dhwani, Replogle Adam, Rowe Lori A, Pritt Bobbi S, Petersen Jeannine M
Division of Vector-Borne Diseases, Bacterial Diseases Branch, Centers for Disease Control and Prevention, Fort Collins, Colorado, United States of America.
Division of Scientific Resources, Biotechnology Core Facility Branch, Centers for Disease Control and Prevention, Atlanta, Georgia, United States of America.
PLoS One. 2016 Dec 28;11(12):e0168994. doi: 10.1371/journal.pone.0168994. eCollection 2016.
Borrelia mayonii, a Borrelia burgdorferi sensu lato (Bbsl) genospecies, was recently identified as a cause of Lyme borreliosis (LB) among patients from the upper midwestern United States. By microscopy and PCR, spirochete/genome loads in infected patients were estimated at 105 to 106 per milliliter of blood. Here, we present the full chromosome and plasmid sequences of two B. mayonii isolates, MN14-1420 and MN14-1539, cultured from blood of two of these patients. Whole genome sequencing and assembly was conducted using PacBio long read sequencing (Pacific Biosciences RSII instrument) followed by hierarchical genome-assembly process (HGAP). The B. mayonii genome is ~1.31 Mbp in size (26.9% average GC content) and is comprised of a linear chromosome, 8 linear and 7 circular plasmids. Consistent with its taxonomic designation as a new Bbsl genospecies, the B. mayonii linear chromosome shares only 93.83% average nucleotide identity with other genospecies. Both B. mayonii genomes contain plasmids similar to B. burgdorferi sensu stricto lp54, lp36, lp28-3, lp28-4, lp25, lp17, lp5, 5 cp32s, cp26, and cp9. The vls locus present on lp28-10 of B. mayonii MN14-1420 is remarkably long, being comprised of 24 silent vls cassettes. Genetic differences between the two B. mayonii genomes are limited and include 15 single nucleotide variations as well as 7 fewer silent vls cassettes and a lack of the lp5 plasmid in MN14-1539. Notably, 68 homologs to proteins present in B. burgdorferi sensu stricto appear to be lacking from the B. mayonii genomes. These include the complement inhibitor, CspZ (BB_H06), the fibronectin binding protein, BB_K32, as well as multiple lipoproteins and proteins of unknown function. This study shows the utility of long read sequencing for full genome assembly of Bbsl genomes, identifies putative genome regions of B. mayonii that may be linked to clinical manifestation or tissue tropism, and provides a valuable resource for pathogenicity, diagnostic and vaccine studies.
伯氏疏螺旋体狭义种（Borrelia burgdorferi sensu lato，Bbsl）中的马尤尼伯氏疏螺旋体（Borrelia mayonii）最近被确定为美国中西部上游地区莱姆病（Lyme borreliosis，LB）患者的致病原因。通过显微镜检查和聚合酶链反应（PCR），估计感染患者每毫升血液中的螺旋体/基因组载量为10⁵至10⁶。在此，我们展示了从两名此类患者血液中培养出的两株马尤尼伯氏疏螺旋体分离株MN14 - 1420和MN14 - 1539的完整染色体和质粒序列。使用PacBio长读长测序（Pacific Biosciences RSII仪器），随后通过分层基因组组装流程（HGAP）进行全基因组测序和组装。马尤尼伯氏疏螺旋体基因组大小约为1.31 Mbp（平均GC含量为26.9%），由一条线性染色体、8条线性质粒和7条环状质粒组成。与其作为新的Bbsl基因种的分类命名一致，马尤尼伯氏疏螺旋体的线性染色体与其他基因种的平均核苷酸同一性仅为93.83%。两株马尤尼伯氏疏螺旋体基因组均含有与狭义伯氏疏螺旋体lp54、lp36、lp28 - 3、lp28 - 4、lp25、lp17、lp5、5条cp32、cp26和cp9相似的质粒。马尤尼伯氏疏螺旋体MN14 - 1420的lp28 - 10上存在的可变主要表面蛋白（vls）位点非常长，由24个沉默vls盒组成。两株马尤尼伯氏疏螺旋体基因组之间的遗传差异有限，包括15个单核苷酸变异、MN14 - 1539中少7个沉默vls盒以及缺少lp5质粒。值得注意的是，马尤尼伯氏疏螺旋体基因组中似乎缺少68个与狭义伯氏疏螺旋体中存在的蛋白质同源的蛋白。这些包括补体抑制剂CspZ（BB_H06）、纤连蛋白结合蛋白BB_K32，以及多种脂蛋白和功能未知的蛋白质。本研究展示了长读长测序在Bbsl基因组全基因组组装中的实用性，确定了马尤尼伯氏疏螺旋体中可能与临床表现或组织嗜性相关的假定基因组区域，并为致病性、诊断和疫苗研究提供了宝贵资源。
