A TonB-dependent outer membrane receptor of Pseudomonas fluorescens : virulence and vaccine potential
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A TonB-dependent outer membrane receptor of Pseudomonas fluorescens: virulence and vaccine potential Yong-hua Hu • Wei Dang • Li Sun
Received: 7 January 2012 / Revised: 18 March 2012 / Accepted: 26 March 2012 / Published online: 12 April 2012 Ó Springer-Verlag 2012
Abstract Pseudomonas fluorescens is a Gram-negative bacterium and a common aquaculture pathogen. In this study, we identified from a pathogenic P. fluorescens strain a TonB-dependent outer membrane receptor, TdrA, as a secreted protein and examined its function and vaccine potential. TdrA is composed of 746 residues and possesses conserved structural domains of TonB-dependent outer membrane receptors. Quantitative real-time reverse transcriptase-PCR analysis showed that expression of tdrA was upregulated under conditions of iron starvation and during infection of host cells. Consistently, iron depletion induced increased production of TdrA protein in the outer membrane. Compared to the wild type, a tdrA-knock out mutant (1) was unable to grow in the absence of iron, (2) exhibited drastically attenuated overall bacterial virulence, and (3) was impaired in the ability to establish lethal infection in host tissues. Purified recombinant TdrA (rTdrA), when used as a subunit vaccine to immunize flounder, was able to induce strong protective immunity, including production
Communicated by Djamel Drider. Y. Hu W. Dang L. Sun (&) Key Laboratory of Experimental Marine Biology, Institute of Oceanology, Chinese Academy of Sciences, 7 Nanhai Road, Qingdao 266071, People’s Republic of China e-mail: [email protected] W. Dang Hangzhou Key Laboratory for Animal Adaptation and Evolution, College of Life and Environmental Sciences, Hangzhou Normal University, Hangzhou 310036, People’s Republic of China W. Dang Graduate University of the Chinese Academy of Sciences, Beijing 100049, People’s Republic of China
of serum-specific antibodies that resulted in effective protection against lethal-dose P. fluorescens challenge. Together, these results indicate that TdrA is an outer membrane receptor and a protective immunogen that is likely to be involved in iron acquisition and, as a result, required for optimal bacterial virulence. Keywords Pseudomonas fluorescens TonB-dependent receptor Virulence Vaccine
Introduction Pseudomonas fluorescens is a Gram-negative, rod-shaped bacterium of the family Pseudomonadaceae. It is a common aquaculture pathogen that can infect a variety of farmed fish species including carp, tilapia, and flounder (Swain et al. 2003, 2007; Geng et al. 2006; Wang et al. 2009; Lo´pez et al. 2011). Fish infected by P. fluorescens develop Red Skin Disease with clinical signs of skin hemorrhage and ulceration. In China, P. fluorescens-associated Red Skin Disease has been reported to affect both freshwater and marine fish (Shu et al. 1997; Wang et al. 1999, 2009; Yan 2005). Control of P. fluorescens infection in aquaculture relies mainly on the use of antibiotic compounds, as no commercial vaccines have been developed. To date, very lit
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