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Toxicity to bivalve hemocytes of pathogenic vibrio cytoplasmic extract
C Lambert1, J L Nicolas, V Bultel
1Laboratoire de Physiologie des Invertébrés Marins, IFREMER, Plouzané, 29280, France.
Abstract:
Using a chemiluminescence (CL) test, it had been previously demonstrated that Vibrio pectenicida, which is pathogenic to Pecten maximus larvae, was able to inhibit completely the CL activity of P. maximus hemocytes and partially inhibit those of Crassostrea gigas. Conversely, a Vibrio sp. strain, S322, pathogenic to C. gigas larvae was more active in reducing the CL activity of oyster hemocytes than of scallop hemocytes. Using this same CL biotest, V. pectenicida and S322 cytoplasmic extracts were shown to reproduce CL inhibition while the cytoplasmic extract of a nonpathogenic strain (U1, Pseudoalteromonas) was without effect. Moreover, cytoplasmic extract as well as live V. pectenicida cells provoked, within a few hours, death of P. maximus hemocytes adhering to a glass slide. After partial purification, it was shown that toxic activities of V. pectenicida cytoplasmic extract was due to a toxin, named VHKT (for vibrio hemocyte-killer toxin), which is heat stable, acid and protease resistant, and less than 3 kDa in molecular weight. Attempts to purify VHKT by reverse-phase (C18) HPLC separated activity into the fraction eluted by water at a retention time of 4.02 min.
Insights
A novel Vibrio toxin, vibrio hemocyte-killer toxin (VHKT), kills scallop hemocytes. This heat-stable toxin, less than 3 kDa, is isolated from Vibrio pectenicida, a pathogen of Pecten maximus larvae.
Area of Science:
- Marine microbiology
- Aquaculture pathology
- Invertebrate immunology
Background:
- Vibrio species are significant pathogens in bivalve aquaculture.
- Hemocytes play a crucial role in the innate immune response of bivalves.
- Previous studies indicated differential effects of Vibrio strains on hemocyte activity in scallops and oysters.
Purpose of the Study:
- To investigate the mechanism of hemocyte inhibition by pathogenic Vibrio strains.
- To identify and characterize the toxic component from Vibrio pectenicida responsible for hemocyte cytotoxicity.
- To assess the potential of chemiluminescence (CL) as a biotest for evaluating Vibrio pathogenicity.
Main Methods:
- Chemiluminescence (CL) biotest to measure hemocyte activity.
- Preparation and application of cytoplasmic extracts from Vibrio strains.
- Microscopic observation of hemocyte viability.
- Partial purification of the toxic factor using chromatography.
Main Results:
- Vibrio pectenicida cytoplasmic extract inhibited Pecten maximus hemocyte CL activity and caused hemocyte death.
- A Vibrio sp. strain S322 showed differential effects on hemocyte CL activity between scallops and oysters.
- A heat-stable, acid and protease-resistant toxin, VHKT (<3 kDa), was identified as the cytotoxic agent in V. pectenicida.
- VHKT was partially purified using reverse-phase HPLC, eluting with water.
Conclusions:
- Vibrio pectenicida produces a potent hemocyte-killing toxin (VHKT).
- VHKT contributes to the pathogenicity of V. pectenicida in Pecten maximus larvae.
- The CL biotest is a valuable tool for assessing the impact of bacterial pathogens on bivalve hemocytes.