title: Analysis of Clostridium botulinum serotype E strains by using multilocus sequence typing, amplified fragment length polymorphism, variable-number tandem-repeat analysis, and botulinum neurotoxin gene sequencing.
abstract: A total of 41 Clostridium botulinum serotype E strains from different geographic regions, including Canada, Denmark, Finland, France, Greenland, Japan, and the United States, were compared by multilocus sequence typing (MLST), amplified fragment length polymorphism (AFLP) analysis, variable-number tandem-repeat (VNTR) analysis, and botulinum neurotoxin (bont) E gene sequencing. The strains, representing environmental, food-borne, and infant botulism samples collected from 1932 to 2007, were analyzed to compare serotype E strains from different geographic regions and types of botulism and to determine whether each of the strains contained the transposon-associated recombinase rarA, involved with bont/E insertion. MLST examination using 15 genes clustered the strains into several clades, with most members within a cluster sharing the same BoNT/E subtype (BoNT/E1, E2, E3, or E6). Sequencing of the bont/E gene identified two new variants (E7, E8) that showed regions of recombination with other E subtypes. The AFLP dendrogram clustered the 41 strains similarly to the MLST dendrogram. Strains that could not be differentiated by AFLP, MLST, or bont gene sequencing were further examined using three VNTR regions. Both intact and split rarA genes were amplified by PCR in each of the strains, and their identities were confirmed in 11 strains by amplicon sequencing. The findings suggest that (i) the C. botulinum serotype E strains result from the targeted insertion of the bont/E gene into genetically conserved bacteria and (ii) recombination events (not random mutations) within bont/E result in toxin variants or subtypes within strains.
authors: Bioscience Division, Los Alamos National Laboratory, Los Alamos, New Mexico 87545, USA.
authors: Thomas E
authors: TE
authors: Macdonald
authors: Charles H
authors: CH
authors: Helma
authors: Yulin
authors: Y
authors: Shou
authors: Yolanda E
authors: YE
authors: Valdez
authors: Lawrence O
authors: LO
authors: Ticknor
authors: Brian T
authors: BT
authors: Foley
authors: Stephen W
authors: SW
authors: Davis
authors: George E
authors: GE
authors: Hannett
authors: Cassandra D
authors: CD
authors: Kelly-Cirino
authors: Jason R
authors: JR
authors: Barash
authors: Stephen S
authors: SS
authors: Arnon
authors: Miia
authors: M
authors: Lindström
authors: Hannu
authors: H
authors: Korkeala
authors: Leonard A
authors: LA
authors: Smith
authors: Theresa J
authors: TJ
authors: Smith
authors: Karen K
authors: KK
authors: Hill
mesh_terms: D001905:Botulinum Toxins / Q000235:genetics; D001906:Botulism / Q000382:microbiology; D047970:Clostridium botulinum type E / Q000145:classification* / Q000235:genetics* / Q000302:isolation & purification; D016000:Cluster Analysis; D004251:DNA Transposable Elements; D004269:DNA, Bacterial / Q000235:genetics*; D004783:Environmental Microbiology; D005516:Food Microbiology; D005838:Genotype; D006801:Humans; D008969:Molecular Sequence Data; D058889:Molecular Typing / Q000379:methods*; D011110:Polymorphism, Genetic; D011995:Recombination, Genetic; D017422:Sequence Analysis, DNA
chemical_list: D004251:DNA Transposable Elements; D004269:DNA, Bacterial; D001905:Botulinum Toxins; C096326:botulinum toxin type E