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Infection and Immunity, September 2006, p. 5272-5283, Vol. 74, No. 9
0019-9567/06/$08.00+0     doi:10.1128/IAI.00546-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

Extensive Genomic Plasticity in Pseudomonas aeruginosa Revealed by Identification and Distribution Studies of Novel Genes among Clinical Isolates

Kai Shen,¹ Sameera Sayeed,^1, Patricia Antalis,¹ John Gladitz,¹ Azad Ahmed,¹ Bethany Dice,¹ Benjamin Janto,¹ Richard Dopico,¹ Randy Keefe,¹ Jay Hayes,¹ Sandra Johnson,¹ Sujun Yu,¹ Nathan Ehrlich,^1, Jennifer Jocz,^1, Laura Kropp,^1, Ray Wong,^1, Robert M. Wadowsky,³ Malcolm Slifkin,^4,¶ Robert A. Preston,¹ Geza Erdos,^1, J. Christopher Post,^1,2 Garth D. Ehrlich,^1,2* and Fen Z. Hu^1,2

Center for Genomic Sciences, Allegheny-Singer Research Institute, Pittsburgh, Pennsylvania 15212,¹ Department of Microbiology and Immunology, Drexel University College of Medicine, Allegheny Campus, Pittsburgh, Pennsylvania 15212,² Children's Hospital of Pittsburgh, Pittsburgh, Pennsylvania 15213,³ Clinical Microbiology Laboratory, Department of Pathology, Allegheny General Hospital, Pittsburgh, Pennsylvania 15212⁴

Received 4 April 2006/ Returned for modification 25 May 2006/ Accepted 23 June 2006

The distributed genome hypothesis (DGH) states that each strain within a bacterial species receives a unique distribution of genes from a population-based supragenome that is many times larger than the genome of any given strain. The observations that natural infecting populations are often polyclonal and that most chronic bacterial pathogens have highly developed mechanisms for horizontal gene transfer suggested the DGH and provided the means and the mechanisms to explain how chronic infections persist in the face of a mammalian host's adaptive defense mechanisms. Having previously established the validity of the DGH for obligate pathogens, we wished to evaluate its applicability to an opportunistic bacterial pathogen. This was accomplished by construction and analysis of a highly redundant pooled genomic library containing approximately 216,000 functional clones that was constructed from 12 low-passage clinical isolates of Pseudomonas aeruginosa, 6 otorrheic isolates and 6 from other body sites. Sequence analysis of 3,214 randomly picked clones (mean insert size, 1.4 kb) from this library demonstrated that 348 (10.8%) of the clones were unique with respect to all genomic sequences of the P. aeruginosa prototype strain, PAO1. Hypothetical translations of the open reading frames within these unique sequences demonstrated protein homologies to a number of bacterial virulence factors and other proteins not previously identified in P. aeruginosa. PCR and reverse transcription-PCR-based assays were performed to analyze the distribution and expression patterns of a 70-open reading frame subset of these sequences among 11 of the clinical strains. These sequences were unevenly distributed among the clinical isolates, with nearly half (34/70) of the novel sequences being present in only one or two of the individual strains. Expression profiling revealed that a vast majority of these sequences are expressed, strongly suggesting they encode functional proteins.

Editor: J. T. Barbieri

Present address: University of Pittsburgh, Pittsburgh, PA 15213.

Present address: Carnegie-Mellon University, Pittsburgh, PA 15213.

Present address: Pennsylvania State University, State College, PA 16802.

^¶ Deceased.