Abstract
We report the draft genome of the human pathogen Corynebacterium diphtheriae bv. mitis NCTC 3529. This is the first C. diphtheriae bv. mitis strain to be sequenced and reveals significant differences from the other primary biovar, C. diphtheriae bv. gravis.
GENOME ANNOUNCEMENT
Corynebacterium diphtheriae is the causative agent of diphtheria, a toxemic localized infection of the upper respiratory tract (6). Currently, four biovars are recognized (C. diphtheriae bv. gravis, C. diphtheriae bv. mitis, C. diphtheriae bv. intermedius, and C. diphtheriae bv. belfanti), based on their morphogenic and biochemical properties (5), and yet the molecular and genetic bases for these differences are poorly understood. We undertook the sequencing of C. diphtheriae bv. mitis NCTC 3529, a strain isolated prior to mass vaccination in the United Kingdom, to better understand the genetic basis that differentiates this biovar from the other main disease-causing biovar, C. diphtheriae bv. gravis.
Genome sequencing of C. diphtheriae bv. mitis strain NCTC 3529 was carried out using a whole-genome shotgun sequencing approach performed on a Roche GS-Junior 454 apparatus at the University of Strathclyde Genomics Facility. Using Rapid library-prepared single-ended runs, we obtained 175,879 reads with an average length of 446 bp. The reads were assembled using GS de novo Assembler (Roche), which led to a final assembly of 43 contigs of >200 bp. The total size of the assembly was 2,483,675 bp, with a mean contig size of 57,760 bp (an average of 47× coverage) and a G+C content of 53.57%. The contigs were ordered onto C. diphtheriae bv. gravis NCTC 13129 (1) using Mauve (4), and the merged sequence was annotated using xBASE (2).
The draft genome of C. diphtheriae bv. mitis NCTC 3529 is estimated to have a total of 2,272 protein coding genes. The overall genome size is smaller than that of C. diphtheriae bv. gravis by 4,960 bp. Analysis using mGenomeSubtractor (7) indicated that 1,982 coding sequences (CDS) were present in both strains, with 318 CDS being present in C. diphtheriae bv. mitis NCTC 3529 that were not present in C. diphtheriae bv. gravis NCTC 13129. These differences were largely confined to the presence of transposons and clustered, regularly interspaced, short palindromic repeat (CRISPR) elements. The biochemical tests used to distinguish between the gravis and mitis biovars in the laboratory include the inability of C. diphtheriae bv. mitis to ferment starch and reduce nitrates (3, 5). Genomic analysis shows that the lack of starch fermentation in NCTC 3529 is likely to be linked to the putative aldose-1-epimerase (DIP1011) that is present in NCTC 13129 but absent in NCTC 3529. In C. diphtheriae bv. mitis NCTC 3529, the region that encodes the molybdopterin biosynthesis machinery and the narIJHGK operon (DIP0492 to DIP0507 in C. diphtheriae bv. gravis NCTC 13129) is absent and likely to account for the lack of nitrate reduction by this biovar. Overall, the genome sequence of C. diphtheriae bv. mitis NCTC 3529 should advance our knowledge of the biology of pathogenic corynebacteria and the molecular basis for strain differentiation.
Nucleotide sequence accession numbers.
The C. diphtheriae bv. mitis (NCTC 3529) Whole Genome Shotgun project has been deposited at DDBJ/EMBL/GenBank under the accession no. AJGI00000000. The version described in this paper is the first version, AJGI01000000.
ACKNOWLEDGMENTS
Work in the P.A.H. laboratory is supported by Medical Research Scotland (grant 422 FRG) and the University of Strathclyde. A.B. is supported by the Deutsche Forschungsgemeinschaft (SFB796, B5).
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