Clostridium botulinum

(CbMT-PLST is currently in beta-testing phase; contact MicrobiType to inquire about no-cost typing.) 

The gram positive, spore-forming bacterium Clostridium botulinum sporadically contaminates various foods (typically, low acid foods that were improperly canned or stored), where it may propagate anaerobically and produce a potent, life-threatening neurotoxin. For example, a recent botulism outbreak with 10 cases and 1 death was traced to an inadequately heated nacho cheese dispenser in a California convenience store (1). Less commonly, C. botulinum is also responsible for infant gastrointestinal infection. To identify outbreaks and track down their source, rapid epidemiological analysis is required. Strain typing can be an important tool in these investigations, particularly if the cases are geographically dispersed. Multiple methods have been developed for C. botulinum typing, including multilocus variable number of tandem repeats analysis which examines length variation in 15 tandem repeats (2), and more recently whole genome sequencing which examines single nucleotide polymorphisms in the core genome (3,4). To varying extents these methods have limitations relating to turnaround time, data portability, technical complexity, and cost. Furthermore, these methods rely on viable pure cultures as starting material; this is problematic since growing and transporting these cultures raise concerns over safety and security (botulinum toxin is a Category A biothreat agent). Also, the trend in clinical laboratories is towards culture-independent diagnostic tests which do not yield pure cultures. 

 

To address the need for a C. botulinum typing system that is compatible with crude, non-viable samples, the tandem repeat-containing locus CbMT2 was identified at MicrobiType as a promising target for polymorphic locus sequence typing (PLST) of Group I strains, the primary culprits in food poisoning (5). This locus was evaluated by analyzing its sequence from 116 C. botulinum strains represented in GenBank databases. As shown in the dendrogram (poster presented at the 2018 International Association for Food Protection Annual Meeting) dendrogram, CbMT2 resolved 87 alleles for these strains, with several clusters representing strains known or likely to be epidemiologically related. Diversity index (following removal of epidemiological replicates) was 0.99. 

Results reported include CbMT2 sequence for your submitted (inactivated) C. botulinum sample, a clustal alignment with closest GenBank match and with any additional samples submitted from your lab, and (upon request) a dendrogram showing relationship to other strains.

 

(1) Choudhry S.A. et al., 2017, Foodborne botulism, I only had nacho cheese: a case report. Cureus 9:e1666.
(2) Fillo S. et al., 2011, Clostridium botulinum group I strain genotyping by 15-locus multilocus variable-number tandem-repeat analysis. J. Clin. Microbiol. 49:4252.
(3) Raphael B.H. et al., 2014, Distinguishing highly-related outbreak-associated
Clostridium botulinum type A(B) strains. BMC Microbiology 14:192.
(4) Williamson C.H. et al., 2016, Comparative genomic analyses reveal broad diversity in botulinum-toxin-producing Clostridia. BMC Genomics 17:180.
(5) Peck M.W. and van Vliet A.H., 2016, Impact of
Clostridium botulinum genomic diversity on food safety. Curr Opin Food Sci 10:52.

 

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