Draft Genome Sequence of Clostridium botulinum Type B Strain Osaka05, Isolated from an Infant Patient with Botulism in Japan.

Clostridium botulinum strain Osaka05, which has been isolated from an infant patient with botulism in Japan, is the first strain producing botulinum neurotoxin subtype B6. Here, we report the draft genome sequence of C. botulinum Osaka05.

GENOME ANNOUNCEMENT

Clostridium botulinum is a Gram-positive, spore-forming, and anaerobic bacterium. It produces botulinum neurotoxins (BoNTXs), and it causes intoxication in humans and animals (1, 2). Bacterial type and toxin type are important in C. botulinum because they are strongly related to the type of intoxication and host animal species. C. botulinum can be classified not only by culture characteristics (groups I to IV) but also by the antigenic properties of BoNTX (serotypes A to G) (3). BoNTX serotype B (BoNTX/B) can be further classified into five subtypes (B1, B2, B3, nonproteolytic B, and bivalent B), and more recently B6, due to the diversity of the amino acid sequences within serotype B (4, 5).

C. botulinum type B1 is generally isolated in infant and food-borne botulism. C. botulinum strain Osaka05, which has been isolated from an infant patient with botulism in Japan and identified as a group I strain, is the first identified C. botulinum producing BoNTX/B6 (5). According to pulsed-field gel electrophoresis and Southern blot analyses, the extrachromosomal DNAs have been revealed to contain the BoNTX/B6 gene in strain Osaka05 (5). Genomic investigation will extend our knowledge of C. botulinum. Here, we report for the first time the draft genome sequence of C. botulinum producing BoNTX/B6, strain Osaka05.

The DNA of strain Osaka05 was sequenced using the Roche 454 GS FLX titanium platform. A single-end library (77,710 reads) and an 8-kb paired-end library (178,004 reads) were assembled with GS Assembler software into one scaffold containing 67 gaps. Then, 22 gaps were manually closed by an in silico analysis using GenoFinisher software and resulted in 55 contigs (46 contigs for a chromosome and 9 contigs for extra chromosomes) (6). The draft sequence was 4,004,744 bp in total length, which is similar to the genome sizes of the other C. botulinum strains. The G+C content of the draft sequence (27.8%) was also similar. By use of the Microbial Genome Annotation Pipeline (MiGAP) (7), the total numbers of coding sequences (CDS) (3,671 chromosomes and 545 extrachromosomes) were predicted and at least one rRNA and at least 49 tRNAs were identified.

In the present study, the BoNTX/B6 gene was found on the Osaka05p2_contig004 (accession no. AP014517), which was considered to be a part of the extrachromosomal DNAs. Unfortunately, the complete nucleotide sequences of the extrachromosomal DNAs were not obtained in this study. Currently, it is not certain whether the extrachromosomal DNA harboring the BoNTX/B6 gene is a plasmid or bacteriophage in strain Osaka05. In the future, the complete nucleotide sequence of the extrachromosomal DNA harboring the BoNTX/B6 gene is expected to be obtained, and the evolutionary traits of the extrachromosomal DNA harboring the BoNTX/B6 gene will be analyzed.

Nucleotide sequence accession number.

The genome sequence of C. botulinum strain Osaka05 was deposited in DDBJ under the accession no. BAUF00000000.