Gonzalez-Escalona Narjol, Sharma Shashi K
Division of Microbiology, Office of Regulatory Science, Center for Food Safety and Applied Nutrition, Food and Drug Administration, College Park, MD, United States.
Front Microbiol. 2020 Feb 19;11:239. doi: 10.3389/fmicb.2020.00239. eCollection 2020.
is a Gram-positive, spore-forming anaerobic bacterium that produces botulinum neurotoxin (BoNT). Closing their genomes provides information about their neurotoxin clusters' arrangement(s) and their location (e.g., chromosome or plasmid) which cannot be assessed using draft genomes. Therefore, we tested the use of long-read sequencing (nanopore sequencing) in combination with short-read sequencing to close two toxin-producing strains. These genomes could be used by the Public Health Emergency Preparedness and Response staff during botulism outbreaks. The genomes of two toxin-producing strains, one from an environmental sample (83F_CFSAN034202) and the other from a clinical sample (CDC51232_CFSAN034200) were sequenced using MinION and MiSeq devices. The genomes, including the chromosomes and the plasmids, were closed by a combination of long-read and short-read sequencing. They belonged to different sequence types (STs), with 83F belonging to ST4 and CDC51232 to ST7. A whole genome single nucleotide polymorphism (SNP) analysis clustered these two strains with strains in lineage 2 (e.g., 6CDC297) and 4 (e.g., NCTC2916) from Group I, respectively. These two strains were also bivalent strains with the BoNTB and BoNTA4 clusters located in the larger plasmid for CDC51232, and the BoNTB and BoNTA1 clusters located both in the chromosome for 83F. Overall, this study showed the advantage of combining these two sequencing methods to obtain high quality closed genomes that could be used for SNP phylogenies (source tracking) as well as for fast identification of BoNT clusters and their gene arrangements.
是一种革兰氏阳性、形成芽孢的厌氧细菌，可产生肉毒杆菌神经毒素（BoNT）。完成它们的基因组测序可提供有关其神经毒素簇排列及其位置（例如，染色体或质粒）的信息，而这些信息无法通过草图基因组进行评估。因此，我们测试了使用长读长测序（纳米孔测序）与短读长测序相结合的方法来完成两个产毒素菌株的基因组测序。在肉毒中毒爆发期间，公共卫生应急准备和响应人员可以使用这些基因组。使用MinION和MiSeq设备对两个产毒素菌株的基因组进行了测序，其中一个来自环境样本（83F_CFSAN034202），另一个来自临床样本（CDC51232_CFSAN034200）。通过长读长和短读长测序相结合的方式完成了包括染色体和质粒在内的基因组测序。它们属于不同的序列类型（STs），83F属于ST4，CDC51232属于ST7。全基因组单核苷酸多态性（SNP）分析将这两个菌株分别与第一组中的2型菌株（例如6CDC297）和4型菌株（例如NCTC2916）聚类在一起。这两个菌株也是双价菌株，对于CDC51232，BoNTB和BoNTA4簇位于较大的质粒中，而对于83F，BoNTB和BoNTA1簇均位于染色体中。总体而言，这项研究显示了结合这两种测序方法以获得高质量封闭基因组的优势，这些基因组可用于SNP系统发育分析（溯源追踪）以及快速鉴定BoNT簇及其基因排列。
