Phobalysin, a Small β-Pore-Forming Toxin of Photobacterium damselae subsp. damselae

Rivas, Amable J.; Hoven, Gisela von; Neukirch, Claudia; Meyenburg, Martina; Qin, Qianqian; Füser, Sabine; Boller, Klaus; Lemos, Manuel L.; Osorio, Carlos R.; Husmann, Matthias

doi:10.1128/iai.00277-15

Cited by 26 publications

(60 citation statements)

References 73 publications

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“…damselae for mammals and for fish and are responsible for hemolytic and cytotoxic activity (Kreger et al, 1987; Rivas et al, 2013a, 2015b). We have shown here that deletion of rstB strongly impairs virulence in the sea bass fish model.…”

Section: Discussionmentioning

confidence: 99%

“…dly gene encodes damselysin, a phospholipase- D active against sphingomyelin (Kreger et al, 1987). hlyA pl encodes the recently characterized pore-forming toxin phobalysin P (PhlyP) which stands for “photobacterial lysin encoded on a plasmid.” This toxin forms small membrane pores in the target cells causing efflux of K + and entry of vital dyes (Rivas et al, 2015b). In addition, all the hemolytic strains contain hlyA ch gene in chromosome I (Rivas et al, 2013a, 2014).…”

Section: Introductionmentioning

confidence: 99%

“…PhlyP and PhlyC exert an additive effect in hemolysis and virulence, whereas all three hemolysins act synergistically on erythrocytes, a phenomenon that accounts for maximum virulence in mice and fish (Rivas et al, 2013a). Recently, we have demonstrated that hemolysins increase bacterial adherence (Rivas et al, 2015b), which may be of paramount relevance in the early events of natural infection. In addition, tissue damage caused by hemolysins may provide a source of nutrients to further support bacterial infection.…”

Section: Introductionmentioning

confidence: 99%

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rstB Regulates Expression of the Photobacterium damselae subsp. damselae Major Virulence Factors Damselysin, Phobalysin P and Phobalysin C

et al. 2017

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The marine pathogenic bacterium Photobacterium damselae subsp. damselae causes septicemia in marine animals and in humans. The pPHDD1 plasmid-encoded hemolysins damselysin (Dly) and phobalysin P (PhlyP), and the chromosome-encoded hemolysin phobalysin C (PhlyC) constitute its main virulence factors. However, the mechanisms by which expression of these three hemolysins is regulated remain unknown. Here we report the isolation of a mini-Tn10 transposon mutant which showed a strong impairment in its hemolytic activity. The transposon disrupted a putative sensor histidine kinase gene vda_000600 (rstB), which together with vda_000601 (rstA) is predicted to encode a putative two-component regulatory system. This system showed to be homologous to the Vibrio cholerae CarSR/VprAB and Escherichia coli RstAB systems. Reconstruction of the mutant by allelic exchange of rstB showed equal impairment in hemolysis, and complementation with a plasmid expressing rstAB restored hemolysis to wild-type levels. Remarkably, we demonstrated by promoter expression analyses that the reduced hemolysis in the rstB mutant was accompanied by a strong decrease in transcription activities of the three hemolysin genes dly (damselysin), hlyApl (phobalysin P) and hlyAch (phobalysin C). Thus, RstB, encoded in the small chromosome, regulates plasmid and chromosomal virulence genes. We also found that reduced expression of the three virulence genes correlated with a strong decrease in virulence in a sea bass model, demonstrating that RstB constitutes a master regulator of the three P. damselae subsp. damselae hemolysins and plays critical roles in the pathogenicity of this bacterium. This study represents the first evidence of a direct role of a RstAB-like system in the regulation of bacterial toxins.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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rstB Regulates Expression of the Photobacterium damselae subsp. damselae Major Virulence Factors Damselysin, Phobalysin P and Phobalysin C

et al. 2017

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“…Currently, it is recognized that highly hemolytic isolates harbor a 153-kb virulence plasmid named pPHDD1, encoding the two hemolysins damselysin (Dly) and HlyA pl (12), with HlyA pl recently renamed phobalysin (PhlyP) due to its differential characteristics (13). In addition, the hemolytic isolates encode in chromosome I a hemolysin dubbed HlyA ch , which is responsible for the hemolytic activity of pPHDD1-negative strains (14).…”

Section: Importancementioning

confidence: 99%

Photobacterium damselae subsp. damselae, an Emerging Fish Pathogen in the Black Sea: Evidence of a Multiclonal Origin

Terceti

Öğüt

Osorio

2016

Appl Environ Microbiol

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Photobacterium damselae subsp. damselae is considered to be an emerging pathogen of marine fish of importance in aquaculture, with a notable increase in its geographical distribution during the last several years. In this study, we carried out for the first time to our knowledge a genetic and pathobiological characterization of 14 strains isolated from sea bass (Dicentrarchus labrax) reared in the Southeastern Black Sea, where high mortalities were observed at two aquaculture farms during the summer and autumn of 2011. Heterogeneity was evidenced among strains in phenotypical traits, such as sucrose fermentation, motility, and hemolysis. Although 11 of 14 isolates were hemolytic, we found that all of the isolates lacked the pPHDD1 virulence plasmid that encodes the phospholipase-D damselysin (Dly) and the pore-forming toxin PhlyP, two hemolysins previously reported to constitute major virulence factors for turbot. Subsequent PCR and sequencing analyses demonstrated that the 11 hemolytic isolates harbored a complete hlyA ch gene, a chromosome I-borne gene that encodes HlyA ch hemolysin, whereas the three nonhemolytic isolates contained hlyA ch pseudogenes caused by insertion sequence elements. Virulence challenges with two representative strains revealed that, albeit less virulent than the pPHDD1-harboring strain RM-71, the plasmidless hlyA ch -positive and hlyA chnegative Black Sea isolates were pathogenic for sea bass. A phylogenetic analysis based on the toxR gene sequence uncovered a greater diversity in the isolates, indicating that the presence of this pathogen in the Black Sea was not caused by the introduction and spread of a single virulent clone but by the proliferation of different clones. IMPORTANCEThe geographical distribution of marine bacterial pathogens is undergoing a worldwide increase. In particular, bacteria of the group vibrios are increasingly being isolated as the causative agents of disease in novel species of cultivated fish in areas where they had not been previously reported. Here we characterize for the first time to our knowledge a collection of isolates of the fish and human pathogen Photobacterium damselae subsp. damselae from diseased sea bass reared in the Black Sea. We uncovered great genetic diversity in the Black Sea isolates of this pathogen, suggesting a multiclonal origin. We also demonstrate for the first time that these isolates bear pathogenic potential for sea bass cultures by virulence challenges. P hotobacterium damselae subsp. damselae is a marine bacterium of the family Vibrionaceae that is recognized as a pathogen for a wide variety of aquatic animals, including fish, molluscs, and crustaceans. In addition, it is a pathogen of concern for humans, as it is capable of causing fatal infections (1). Most of the reported infections in humans originated from wounds inflicted during the handling of fish and fishing tools or from exposure to marine animals or seawater. Notably, it is a primary pathogen of fish species of economical importance in aquaculture. During ...

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“…Damselysin (Dly), the product of dly, is a phospholipase D (12). Phobalysin P (PhlyP, "photobacterial lysin encoded on a plasmid"), the product of hlyA pl , is a small ␤-pore-forming toxin (13). The amino acid sequence of phobalysin C (PhlyC), the product of hlyA ch , shares 92% identity with that of PhlyP.…”

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confidence: 99%

Chromosome-Encoded Hemolysin, Phospholipase, and Collagenase in Plasmidless Isolates of Photobacterium damselae subsp. damselae Contribute to Virulence for Fish

Vences

Rivas

Lemos

et al. 2017

Appl Environ Microbiol

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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.

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Phobalysin, a Small β-Pore-Forming Toxin of Photobacterium damselae subsp. damselae

Cited by 26 publications

References 73 publications

rstB Regulates Expression of the Photobacterium damselae subsp. damselae Major Virulence Factors Damselysin, Phobalysin P and Phobalysin C

rstB Regulates Expression of the Photobacterium damselae subsp. damselae Major Virulence Factors Damselysin, Phobalysin P and Phobalysin C

Photobacterium damselae subsp. damselae, an Emerging Fish Pathogen in the Black Sea: Evidence of a Multiclonal Origin

Chromosome-Encoded Hemolysin, Phospholipase, and Collagenase in Plasmidless Isolates of Photobacterium damselae subsp. damselae Contribute to Virulence for Fish

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