Plasmid Characterization and Chromosome Analysis of Two netF+ Clostridium perfringens Isolates Associated with Foal and Canine Necrotizing Enteritis
The recent discovery of a novel beta pore forming toxin, NetF, which is strongly associated with canine and foal necrotizing enteritis should improve our understanding of the role of type A Clostridium perfringens associated disease in these animals. The current study presents the complete genome sequence of two netF positive strains, JFP55 and JFP838, which were recovered from cases of foal necrotizing enteritis and canine hemorrhagic gastroenteritis, respectively. Genome sequencing was done using Single Molecule, Real Time (SMRT) technology PacBio and Illumina Hiseq2000. The JFP55 and JFP838 genomes include a single 3.34 Mb and 3.53 Mb chromosome, respectively, and both genomes include five circular plasmids. Plasmid annotation revealed that three plasmids were shared by the two newly sequenced genomes, including a NetF/NetE toxins encoding tcp conjugative plasmid, a CPE/CPB2 toxins encoding tcp conjugative plasmid and a putative bacteriocin encoding plasmid. The putative beta pore forming toxin genes, netF, netE and netG, were located in unique pathogenicity loci on tcp conjugative plasmids. The C. perfringens JFP55 chromosome carries 2,825 protein coding genes whereas the chromosome of JFP838 contains 3,014 protein encoding genes. Comparison of these two chromosomes with three available reference C. perfringens chromosome sequences identified 48 ~247 kb) and 81 ~430 kb) regions unique to JFP55 and JFP838, respectively. Some of these divergent genomic regions in both chromosomes are phage and plasmid related segments. Sixteen of these unique chromosomal regions ~69 kb) were shared between the two isolates. Five of these shared regions formed a mosaic of plasmid integrated segments, suggesting that these elements were acquired early in a clonal lineage of netF positive C. perfringens strains. These results provide significant insight into the basis of canine and foal necrotizing enteritis and are the first to demonstrate that netF resides on a large and unique plasmid encoded locus.
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