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A High-throughput-compatible FRET-based Platform for Identification and Characterization of Botulinum Neurotoxin Light Chain Modulators
Published on: December 27, 2013
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Clostridium botulinum strains producing BoNT/F4 or BoNT/F5
Brian H Raphael1, Marite Bradshaw, Suzanne R Kalb
1Centers for Disease Control and Prevention, Atlanta, Georgia, USA.
Applied and Environmental Microbiology
|March 18, 2014
Summary
Botulinum neurotoxin type F (BoNT/F) subtypes, including bont/F4 and bont/F5, were analyzed in Clostridium botulinum strains. Data suggest that bont/F5 is carried on a mobile plasmid, facilitating its spread among diverse bacterial strains.
Area of Science:
- Microbiology
- Molecular Biology
- Genetics
Background:
- Botulinum neurotoxin type F (BoNT/F) is produced by Clostridium botulinum, with seven identified subtypes.
- A specific strain, Af84, possesses three neurotoxin genes: bont/F4, bont/F5, and bont/A2.
Purpose of the Study:
- To investigate the genetic basis and distribution of BoNT/F subtypes (bont/F4 and bont/F5) in Clostridium botulinum.
- To determine the mechanisms of BoNT/F subtype dissemination among bacterial strains.
Main Methods:
- Analysis of eight strains with bont/F4 and seven with bont/F5 using culture supernatants and BoNT/F-specific peptide substrates.
- Mass spectral analysis of peptide cleavage products to detect BoNT/F subtypes.
- PCR, Southern hybridization, and whole-genome sequencing to identify genetic elements and genomic backgrounds.
Main Results:
- BoNT/F subtypes were detected in culture supernatants, confirming the presence of bont/F4 and bont/F5.
- A plasmid-specific marker (PL-6) and an approximately 242-kb plasmid carrying bont/F5 were identified in specific strains.
- Genome sequencing revealed diverse genomic backgrounds for strains harboring either bont/F4 or bont/F5.
- No strains simultaneously produced both BoNT/F4 and BoNT/F5, indicating strain Af84 is atypical.
Conclusions:
- The findings support the mobility of a bont/F5-carrying plasmid across diverse Clostridium botulinum genomic backgrounds.
- BoNT/F subtype distribution is influenced by horizontal gene transfer mechanisms, particularly plasmid mobility.
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