Dataset Information

Complete DNA sequence and analysis of the transferable multiple-drug resistance plasmids (R Plasmids) from Photobacterium damselae subsp. piscicida isolates collected in Japan and the United States.

ABSTRACT: Photobacterium damselae subsp. piscicida is a bacterial fish pathogen that causes a disease known as pasteurellosis. Two transferable multiple-drug resistance (R) plasmids, pP99-018 (carrying resistance to kanamycin, chloramphenicol, tetracycline, and sulfonamide) and pP91278 (carrying resistance to tetracycline, trimethoprim, and sulfonamide), isolated from P. damselae subsp. piscicida strains from Japan (P99-018) and the United States (P91278), respectively, were completely sequenced and analyzed, along with the multiple-drug resistance regions of three other R plasmids also from P. damselae subsp. piscicida strains from Japan. The sequence structures of pP99-018 (150,057 bp) and pP91278 (131,520 bp) were highly conserved, with differences due to variation in the drug resistance and conjugative transfer regions. These plasmids, shown to be closely related to the IncJ element R391 (a conjugative, self-transmitting, integrating element, or constin), were divided into the conjugative transfer, replication, partition, and multiple-drug resistance regions. Each of the five multiple-drug resistance regions sequenced exhibited unique drug resistance marker composition and arrangement.

SUBMITTER: Kim MJ

PROVIDER: S-EPMC2224721 | biostudies-literature | 2008 Feb

REPOSITORIES: biostudies-literature

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Complete DNA sequence and analysis of the transferable multiple-drug resistance plasmids (R Plasmids) from Photobacterium damselae subsp. piscicida isolates collected in Japan and the United States.

Kim Mi-Jung MJ Hirono Ikuo I Kurokawa Ken K Maki Takeshi T Hawke John J Kondo Hidehiro H Santos Mudjekeewis D MD Aoki Takashi T

Antimicrobial agents and chemotherapy 20071210 2

Photobacterium damselae subsp. piscicida is a bacterial fish pathogen that causes a disease known as pasteurellosis. Two transferable multiple-drug resistance (R) plasmids, pP99-018 (carrying resistance to kanamycin, chloramphenicol, tetracycline, and sulfonamide) and pP91278 (carrying resistance to tetracycline, trimethoprim, and sulfonamide), isolated from P. damselae subsp. piscicida strains from Japan (P99-018) and the United States (P91278), respectively, were completely sequenced and analy ...[more]

PMID: 18070959

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Project description:AIP56 (apoptosis inducing protein of 56 kDa) is a key virulence factor secreted by virulent strains of Photobacterium damselae subsp. piscicida (Phdp), a Gram-negative bacterium that causes septicemic infections in several warm water marine fish species. AIP56 is systemically disseminated during infection and induces massive apoptosis of host macrophages and neutrophils, playing a decisive role in the disease outcome. AIP56 is a single-chain AB-type toxin, being composed by a metalloprotease A domain located at the N-terminal region connected to a C-terminal B domain, required for internalization of the toxin into susceptible cells. After binding to a still unidentified surface receptor, AIP56 is internalised through clathrin-mediated endocytosis, reaches early endosomes and translocates into the cytosol through a mechanism requiring endosomal acidification and involving low pH-induced unfolding of the toxin. At the cytosol, the catalytic domain of AIP56 cleaves NF-κB p65, leading to the apoptotic death of the intoxicated cells. It has been reported that host cytosolic factors, including host cell chaperones such as heat shock protein 90 (Hsp90) and peptidyl-prolyl cis/trans isomerases (PPIases), namely cyclophilin A/D (Cyp) and FK506-binding proteins (FKBP) are involved in the uptake of several bacterial AB toxins with ADP-ribosylating activity, but are dispensable for the uptake of other AB toxins with different enzymatic activities, such as Bacillus anthracis lethal toxin (a metalloprotease) or the large glycosylating toxins A and B of Clostridium difficile. Based on these findings, it has been proposed that the requirement for Hsp90/PPIases is a common and specific characteristic of ADP-ribosylating toxins. In the present work, we demonstrate that Hsp90 and the PPIases cyclophilin A/D are required for efficient intoxication by the metalloprotease toxin AIP56. We further show that those host cell factors interact with AIP56 in vitro and that the interactions increase when AIP56 is unfolded. The interaction with Hsp90 was also demonstrated in intact cells, at 30 min post-treatment with AIP56, suggesting that it occurs during or shortly after translocation of the toxin from endosomes into the cytosol. Based on these findings, we propose that the participation of Hsp90 and Cyp in bacterial toxin entry may be more disseminated than initially expected, and may include toxins with different catalytic activities.

Project description:Photobacterium damselae subsp. damselae is a pathogen of marine animals, including fish of importance in aquaculture. The virulence plasmid pPHDD1, characteristic of highly hemolytic isolates, encodes the hemolysins damselysin (Dly) and phobalysin (PhlyP). Strains lacking pPHDD1 constitute the vast majority of the isolates from fish outbreaks, but genetic studies to identify virulence factors in plasmidless strains are scarce. Here, we show that the chromosome I-encoded hemolysin PhlyC plays roles in virulence and cell toxicity in pPHDD1-negative isolates of this pathogen. By combining the analyses of whole genomes and of gene deletion mutants, we identified two hitherto uncharacterized chromosomal loci encoding a phospholipase (PlpV) and a collagenase (ColP). PlpV was ubiquitous in the subspecies and exerted hemolytic activity against fish erythrocytes, which was enhanced in the presence of lecithin. ColP was restricted to a fraction of the isolates and was responsible for the collagen-degrading activity in this subspecies. Consistent with the presence of signal peptides in PlpV and ColP sequences, mutants for the type II secretion system (T2SS) genes epsL and pilD exhibited impairments in phospholipase and collagenase activities. Sea bass virulence experiments and cell culture assays demonstrated major contributions of PhlyC and PlpV to virulence and toxicity.IMPORTANCE This study constitutes genetic and genomic analyses of plasmidless strains of an emerging pathogen in marine aquaculture, Photobacterium damselae subsp. damselae To date, studies on the genetic basis of virulence were restricted to the pPHDD1 plasmid-encoded toxins Dly and PhlyP. However, the vast majority of the recent isolates of this pathogen from fish farm outbreaks lack this plasmid. Here we demonstrate that the plasmidless strains produce two hitherto uncharacterized ubiquitous toxins encoded in chromosome I, namely, the hemolysin PhlyC and the phospholipase PlpV. We report the main roles of these two toxins in fish virulence and in cell toxicity. Our results constitute the basis for a better understanding of the virulence of a widespread marine pathogen.

Dataset Information

Complete DNA sequence and analysis of the transferable multiple-drug resistance plasmids (R Plasmids) from Photobacterium damselae subsp. piscicida isolates collected in Japan and the United States.

Publications

Complete DNA sequence and analysis of the transferable multiple-drug resistance plasmids (R Plasmids) from Photobacterium damselae subsp. piscicida isolates collected in Japan and the United States.

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