title: Genome sequencing and analysis of Yersina pestis KIM D27, an avirulent strain exempt from select agent regulation.
abstract: Yersinia pestis is the causative agent of the plague. Y. pestis KIM 10+ strain was passaged and selected for loss of the 102 kb pgm locus, resulting in an attenuated strain, KIM D27. In this study, whole genome sequencing was performed on KIM D27 in order to identify any additional differences. Initial assemblies of 454 data were highly fragmented, and various bioinformatic tools detected between 15 and 465 SNPs and INDELs when comparing both strains, the vast majority associated with A or T homopolymer sequences. Consequently, Illumina sequencing was performed to improve the quality of the assembly. Hybrid sequence assemblies were performed and a total of 56 validated SNP/INDELs and 5 repeat differences were identified in the D27 strain relative to published KIM 10+ sequence. However, further analysis showed that 55 of these SNP/INDELs and 3 repeats were errors in the KIM 10+ reference sequence. We conclude that both 454 and Illumina sequencing were required to obtain the most accurate and rapid sequence results for Y. pestis KIMD27. SNP and INDELS calls were most accurate when both Newbler and CLC Genomics Workbench were employed. For purposes of obtaining high quality genome sequence differences between strains, any identified differences should be verified in both the new and reference genomes.
authors.affiliation: J. Craig Venter Institute, Rockville, Maryland, United States of America. llosada@jcvi.org
authors.forename: Liliana
authors.initials: L
authors.lastname: Losada
authors.forename: John J
authors.initials: JJ
authors.lastname: Varga
authors.forename: Jessica
authors.initials: J
authors.lastname: Hostetler
authors.forename: Diana
authors.initials: D
authors.lastname: Radune
authors.forename: Maria
authors.initials: M
authors.lastname: Kim
authors.forename: Scott
authors.initials: S
authors.lastname: Durkin
authors.forename: Olaf
authors.initials: O
authors.lastname: Schneewind
authors.forename: William C
authors.initials: WC
authors.lastname: Nierman
mesh_terms: D019295:Computational Biology / Q000379:methods*; D017931:DNA Primers / Q000235:genetics; D016680:Genome, Bacterial; D006801:Humans; D020641:Polymorphism, Single Nucleotide; D012091:Repetitive Sequences, Nucleic Acid; D015203:Reproducibility of Results; D017422:Sequence Analysis, DNA / Q000379:methods*; D013045:Species Specificity; D014774:Virulence; D015010:Yersinia pestis / Q000235:genetics* / Q000378:metabolism*
chemical_list: D017931:DNA Primers