Sequence and Organization of pXO1, the Large Bacillus anthracis Plasmid Harboring the Anthrax Toxin Genes

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Journal of Bacteriology, October 1999, p. 6509-6515, Vol. 181, No. 20
0021-9193/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

Sequence and Organization of pXO1, the Large Bacillus anthracis Plasmid Harboring the Anthrax Toxin Genes

R. T. Okinaka,¹^,^* K. Cloud,¹^, O. Hampton,¹ A. R. Hoffmaster,² K. K. Hill,¹ P. Keim,³ T. M. Koehler,² G. Lamke,¹^, S. Kumano,¹^,^§ J. Mahillon,⁴ D. Manter,¹^, Y. Martinez,¹ D. Ricke,¹ # R. Svensson,¹ and P. J. Jackson¹

Life Sciences Division, Los Alamos National Laboratory, Los Alamos, New Mexico 87545¹; Department of Biology, Northern Arizona University, Flagstaff, Arizona 86011-5640³; Department of Microbiology and Molecular Genetics, University of Texas $---$ Houston Medical School, Houston, Texas 77030²; and Laboratoire de Génétique Microbienne $---$ UCL, Louvain-la-Neuve, Belgium⁴

Received 12 April 1999/Accepted 30 July 1999

The Bacillus anthracis Sterne plasmid pXO1 was sequenced by random, "shotgun" cloning. A circular sequence of 181,654 bp wasgenerated. One hundred forty-three open reading frames (ORFs)were predicted using GeneMark and GeneMark.hmm, comprising only61% (110,817 bp) of the pXO1 DNA sequence. The overall guanine-plus-cytosinecontent of the plasmid is 32.5%. The most recognizable featureof the plasmid is a "pathogenicity island," defined by a 44.8-kbregion that is bordered by inverted IS1627 elements at each end.This region contains the three toxin genes (cya, lef, and pagA),regulatory elements controlling the toxin genes, three germinationresponse genes, and 19 additional ORFs. Nearly 70% of the ORFson pXO1 do not have significant similarity to sequences availablein open databases. Absent from the pXO1 sequence are homologsto genes that are typically required to drive theta replicationand to maintain stability of large plasmids in Bacillus spp. Amongthe ORFs with a high degree of similarity to known sequences area collection of putative transposases, resolvases, and integrases,suggesting an evolution involving lateral movement of DNA amongspecies. Among the remaining ORFs, there are three sequences thatmay encode enzymes responsible for the synthesis of a polysaccharidecapsule usually associated with serotype-specific virulentstreptococci.

^* Corresponding author. Mailing address: Life Sciences Division, Los Alamos National Laboratory, M888, Los Alamos, NM 87545.Phone: (505) 667-2743. Fax: (505) 665-3024. E-mail: okinaka{at}telomere.lanl.gov.

Present address: University of Colorado Health Science Center, Denver, CO80220.

Present address: Mayo Medical School, Rochester, MN55905.

^§ Present address: Department of Molecular and Cell Genetics, Tottori University, Yonago, Tottori 683,Japan.

Present address: College of Forestry, Oregon State University, Corvallis, OR97331.

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