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Published on: April 2, 2013
Generation of Paramoeba perurans clonal cultures using flow cytometry and confirmation of virulence
Abstract:
Amoebic gill disease (AGD) in farmed Atlantic salmon is caused by the amoeba Paramoeba perurans. The recent establishment of in vitro culture techniques for P. perurans has provided a valuable tool for studying the parasite in detail. In this study, flow cytometry was used to generate clonal cultures from single-sorted amoeba, and these were used to successfully establish AGD in experimental Atlantic salmon. The clonal cultures displayed differences in virulence, based on gill scores. The P. perurans load on gills, determined by qPCR analysis, showed a positive relationship with gill score, and with clonal virulence, indicating that the ability of amoebae to proliferate and/or remain attached on gills may play a role in virulence. Gill scores based on gross signs and histopathological analysis were in agreement. No association between level of gill score and specific gill arch was observed. It was found that for fish with lower gill scores based on histopathological examination, gross examination and qPCR analysis of gills from the same fish were less successful in detecting lesions and amoebae, respectively.
Insights
Amoebic gill disease in Atlantic salmon, caused by Paramoeba perurans, showed varying virulence in clonal cultures. Higher parasite loads correlated with disease severity, suggesting proliferation and attachment are key to virulence.
Area of Science:
- Aquaculture
- Fish Pathology
- Parasitology
Background:
- Amoebic gill disease (AGD) significantly impacts farmed Atlantic salmon.
- Paramoeba perurans is the causative agent of AGD.
- In vitro culture of P. perurans enables detailed parasite study.
Purpose of the Study:
- To investigate the virulence of clonal isolates of Paramoeba perurans.
- To establish a link between P. perurans load and AGD severity.
- To evaluate diagnostic methods for AGD detection.
Main Methods:
- Single-cell sorting and clonal culture of P. perurans using flow cytometry.
- Experimental infection of Atlantic salmon to establish AGD.
- Quantification of P. perurans load via qPCR and assessment of gill pathology.
Main Results:
- Clonal P. perurans cultures exhibited differential virulence, leading to varied gill scores.
- A positive correlation was observed between P. perurans gill load, gill score, and clonal virulence.
- Gross and histopathological gill scoring methods showed agreement; qPCR accurately reflected P. perurans load.
Conclusions:
- The proliferation and/or attachment of P. perurans on salmon gills are critical factors in AGD virulence.
- qPCR and histopathology are reliable for detecting AGD, especially in cases with higher parasite loads.
- Gross examination may be less sensitive for detecting early or mild AGD lesions.
