Comparative genomic analyses reveal broad diversity in botulinum-toxin-producing Clostridia

Abstract

BackgroundClostridium botulinum is a diverse group of bacteria characterized by the production of botulinum neurotoxin. Botulinum neurotoxins are classified into serotypes (BoNT/A–G), which are produced by six species/Groups of Clostridia, but the genetic background of the bacteria remains poorly understood. The purpose of this study was to use comparative genomics to provide insights into the genetic diversity and evolutionary history of bacteria that produce the potent botulinum neurotoxin.ResultsComparative genomic analyses of over 170 Clostridia genomes, including our draft genome assemblies for 59 newly sequenced Clostridia strains from six continents and publicly available genomic data, provided in-depth insights into the diversity and distribution of BoNT-producing bacteria. These newly sequenced strains included Group I and II strains that express BoNT/A,/B,/E, or/F as well as bivalent strains. BoNT-producing Clostridia and closely related Clostridia species were delineated with a variety of methods including 16S rRNA gene, concatenated marker genes, core genome and concatenated multi-locus sequencing typing (MLST) gene phylogenies that related whole genome sequenced strains to publicly available strains and sequence types. These analyses illustrated the phylogenetic diversity in each Group and the diversity of genomic backgrounds that express the same toxin type or subtype. Comparisons of the botulinum neurotoxin genes did not identify novel toxin types or variants.ConclusionsThis study represents one of the most comprehensive analyses of whole genome sequence data for Group I and II BoNT-producing strains. Read data and draft genome assemblies generated for 59 isolates will be a resource to the research community. Core genome phylogenies proved to be a powerful tool for differentiating BoNT-producing strains and can provide a framework for the study of these bacteria. Comparative genomic analyses of Clostridia species illustrate the diversity of botulinum-neurotoxin-producing strains and the plasticity of the genomic backgrounds in which bont genes are found.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-016-2502-z) contains supplementary material, which is available to authorized users.

Impact of Clostridium botulinum genomic diversity on food safety

“…Genes encoding the neurotoxin and accessory proteins (e.g. non-toxic-non-haemagglutinin (NTNH)) are co-located in one of two conserved neurotoxin complex clusters ( ha cluster or orf-X cluster) at one of several specific insertion sites on the chromosome or a plasmid [7•, 26•, 37, 38, 39•]. All type B and some type A neurotoxin genes are located in the ha cluster, that comprises genes encoding the neurotoxin, NTNH, three haemagglutinins, and a positive regulator ( botR ).…”

“…C. botulinum Groups I and II are associated with foodborne botulism and other forms of human botulism, C. botulinum Group III with botulism in animals, while C. botulinum Group IV has not been strongly associated with botulism. Some strains of C. baratii and C. butyricum also form botulinum neurotoxin and are associated with human botulism [1, 2, 3, 4, 5, 6•, 7•]. …”

Diversity of the Germination Apparatus in Clostridium botulinum Groups I, II, III, and IV

“…There are seven confirmed botulinum neurotoxins (types A to G), and approximately forty different subtypes (Carter and Peck, 2015; Hill et al, 2015; Williamson et al, 2016). The botulinum neurotoxin is the most powerful toxin known, with as little as 30–100 ng sufficient to cause human botulism.…”

“…C. sporogenes is a significant cause of food spoilage (McClure, 2006), and due to its close physiological similarity to C. botulinum Group I is widely used as a surrogate in demonstrating the effectiveness of food preservation processes (Brown et al, 2012; Taylor et al, 2013). Recent analysis indicates that several strains that form type B neurotoxin, and were previously classified as C. botulinum Group I, appear more like strains of C. sporogenes that have acquired a neurotoxin gene (Carter et al, 2009; Weigand et al, 2015; Williamson et al, 2016). C. botulinum Group II (non-proteolytic C. botulinum ) is an important cause of foodborne botulism in humans, and is a concern for the safe production of minimally heat-processed refrigerated foods (Peck and Stringer, 2005; Peck, 2006; Peck et al, 2008).…”

An Integrative Approach to Computational Modelling of the Gene Regulatory Network Controlling Clostridium botulinum Type A1 Toxin Production

“…We use biological data from the literature that relates to five strains of C . botulinum Group I type A1 (ATCC19397, 62A, Hall A, Hall A- hyper and ATCC3502); the close relationship between these strains having been established by whole genome sequencing, microarray analysis and MLST [53–56]. Additionally, these five strains all possess identical or very similar bont and botR genes [52,57,58].…”