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Journal of Bacteriology, June 2008, p. 3969-3978, Vol. 190, No. 11
0021-9193/08/$08.00+0     doi:10.1128/JB.02004-07
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

Opr86 Is Essential for Viability and Is a Potential Candidate for a Protective Antigen against Biofilm Formation by Pseudomonas aeruginosa{triangledown}

Yosuke Tashiro,1,{dagger} Nobuhiko Nomura,1,{dagger}* Ryoma Nakao,2 Hidenobu Senpuku,2 Reiko Kariyama,3 Hiromi Kumon,3 Saori Kosono,4 Haruo Watanabe,2 Toshiaki Nakajima,1 and Hiroo Uchiyama1

Graduate School of Life and Environmental Sciences, University of Tsukuba, Tsukuba, Ibaraki 305-8572,1 Department of Bacteriology, National Institute of Infectious Diseases, Tokyo 162-8640,2 Department of Urology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama, Okayama 700-8558,3 Environmental Molecular Biology Laboratory, RIKEN, Wako, Saitama 351-0198, Japan4

Received 26 December 2007/ Accepted 25 March 2008

Pseudomonas aeruginosa is an opportunistic bacterial pathogen that is one of the most refractory to therapy when it forms biofilms in the airways of cystic fibrosis patients. To date, studies regarding the production of an immunogenic and protective antigen to inhibit biofilm formation by P. aeruginosa have been superficial. The previously uncharacterized outer membrane protein (OMP) Opr86 (PA3648) of P. aeruginosa is a member of the Omp85 family, of which homologs have been found in all gram-negative bacteria. Here we verify the availability of Opr86 as a protective antigen to inhibit biofilm formation by P. aeruginosa PAO1 and several other isolates. A mutant was constructed in which Opr86 expression could be switched on or off through a tac promoter-controlled opr86 gene. The result, consistent with previous Omp85 studies, showed that Opr86 is essential for viability and plays a role in OMP assembly. Depletion of Opr86 resulted in streptococci-like morphological changes and liberation of excess membrane vesicles. A polyclonal antibody against Opr86 which showed reactivity to PAO1 cells was obtained. The antibody inhibited biofilm formation by PAO1 and the other clinical strains tested. Closer examination of early attachment revealed that cells treated with the antibody were unable to attach to the surface. Our data suggest that Opr86 is a critical OMP and a potential candidate as a protective antigen against biofilm formation by P. aeruginosa.

* Corresponding author. Mailing address: Graduate School of Life and Environmental Sciences, University of Tsukuba, Tsukuba, Ibaraki 305-8572, Japan. Phone and fax: 81-29-853-6627. E-mail: nomunobu{at}sakura.cc.tsukuba.ac.jp

{triangledown} Published ahead of print on 4 April 2008.

{dagger} These authors contributed equally.

Journal of Bacteriology, June 2008, p. 3969-3978, Vol. 190, No. 11
0021-9193/08/$08.00+0     doi:10.1128/JB.02004-07
Copyright © 2008, American Society for Microbiology. All Rights Reserved.



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