Table_3_No Assembly Required: Using BTyper3 to Assess the Congruency of a Proposed Taxonomic Framework for the Bacillus cereus Group With Historical Typing Methods.XLSX

The Bacillus cereus group, also known as B. cereus sensu lato (s.l.), is a species complex comprising numerous closely related lineages, which vary in their ability to cause illness in humans and animals. The classification of B. cereus s.l. isolates into species-level taxonomic units is essential for facilitating communication between and among microbiologists, clinicians, public health officials, and industry professionals, but is not always straightforward. A recently proposed genomospecies-subspecies-biovar taxonomic framework aims to provide a standardized nomenclature for this species complex but relies heavily on whole-genome sequencing (WGS). It thus is unclear whether popular, low-cost typing methods (e.g., single- and multi-locus sequence typing) remain congruent with the proposed taxonomy. Here, we characterize 2,231 B. cereus s.l. genomes using a combination of in silico (i) average-nucleotide identity (ANI)-based genomospecies assignment, (ii) ANI-based subspecies assignment, (iii) seven-gene multi-locus sequence typing (MLST), (iv) single-locus panC group assignment, (v) rpoB allelic typing, and (vi) virulence factor detection. We show that sequence types (STs) assigned using MLST can be used for genomospecies assignment, and we provide a comprehensive list of ST/genomospecies associations. For panC group assignment, we show that an adjusted, eight-group framework is largely, albeit not perfectly, congruent with the proposed eight-genomospecies taxonomy, as panC alone may not distinguish (i) B. luti from Group II B. mosaicus and (ii) B. paramycoides from Group VI B. mycoides. We additionally provide a list of loci that capture the topology of the whole-genome B. cereus s.l. phylogeny that may be used in future sequence typing efforts. For researchers with access to WGS, MLST, and/or panC data, we showcase how our recently released software, BTyper3 (https://github.com/lmc297/BTyper3), can be used to assign B. cereus s.l. isolates to taxonomic units within this proposed framework with little-to-no user intervention or domain-specific knowledge of B. cereus s.l. taxonomy. We additionally outline a novel method for assigning B. cereus s.l. genomes to pseudo-gene flow units within proposed genomospecies. The results presented here highlight the backward-compatibility and accessibility of the recently proposed genomospecies-subspecies-biovar taxonomic framework and illustrate that WGS is not a necessity for microbiologists who want to use the proposed nomenclature effectively.

枯草芽孢杆菌属，亦称B. cereus sensu lato（广义B. cereus），是一个包含众多密切相关谱系的物种复合体，其致病能力在人类和动物中存在差异。将B. cereus s.l.分离株分类至物种水平的分类单元对于促进微生物学家、临床医生、公共卫生官员及产业专业人士之间的沟通至关重要，但这一过程并非总是简单直接。近期提出的一套基因组物种-亚基因组物种-生物变种分类框架旨在为该物种复合体提供标准化的命名法，然而其高度依赖全基因组测序（WGS）。因此，尚不清楚常见的低成本分型方法（如单核苷酸多态性分析和多位点序列分型）是否与所提出的分类学保持一致。在本研究中，我们采用结合了模拟方法（i）基于平均核苷酸一致性（ANI）的基因组物种分配，（ii）基于ANI的亚基因组物种分配，（iii）七基因多位点序列分型（MLST），（iv）单基因panC群分配，（v）rpoB等位基因分型，以及（vi）致病因子检测等手段，对2,231个B. cereus s.l.基因组进行了表征。我们发现，通过MLST分配的序列型（STs）可以用于基因组物种分配，并提供了全面的ST/基因组物种关联列表。对于panC群分配，我们发现经过调整的八群框架与所提出的八基因组物种分类学在很大程度上是一致的，尽管并非完美无缺，因为panC本身可能无法区分（i）B. luti与第二组B. mosaicus以及（ii）B. paramycoides与第六组B. mycoides。此外，我们还提供了一份能够捕捉全基因组B. cereus s.l.系统发育拓扑结构的位点列表，这些位点可用于未来的序列分型工作。对于能够获取WGS、MLST和/或panC数据的科研人员，我们展示了如何使用我们最近发布的软件BTyper3（https://github.com/lmc297/BTyper3），在几乎无需用户干预或对B. cereus s.l.分类学有特定领域知识的情况下，将B. cereus s.l.分离株分配至所提出的分类单元中。我们还概述了一种将B. cereus s.l.基因组分配至所提出基因组物种内的伪基因流单元的新方法。本研究结果突出了近期提出的基因组物种-亚基因组物种-生物变种分类框架的向后兼容性和易用性，并说明了全基因组测序并非微生物学家有效使用所提议命名法的必要条件。