A High Throughput Viability Screening Method for the Marine Ectoparasite Neoparamoeba perurans

Abstract

The marine protozoan parasite Neoparamoeba perurans has been established as the causative agent for amoebic gill disease (AGD) in Atlantic salmon (Salmo salar). Freshwater bathing is the only routinely used treatment for AGD in Australia while hydrogen peroxide (H₂O₂) is used in countries with cooler water temperatures. The identification of new treatments that do not rely on either freshwater or H₂O₂ bathing is highly sought. However, in vitro based methods for high throughput screening of antiparasitic compounds have not been established for this parasite. To this end the present study evaluated two in vitro bioassays based on metabolic energy production and cellular membrane integrity to distinguish between amoebistatic (crenated or pseudocyst forms with recovery possible) and amoebicidal (death) activity. Amoebae were subject to either freshwater, H₂O₂ or chloramine-T for 4h treatment and assessed 24h after recovery. Visualization by microscopy and bioassay assessment 24h post-treatment confirmed that H₂O₂ and freshwater are 95% amoebicidal albeit due to different mechanisms of action. These data are consistent with other studies where amoebae have been observed to recover following exposure to these compounds and provide evidence for the inclusion of a recovery component to differentiate between the mechanism of action of amoebicidal and amoebistatic treatments. Together these bioassays are a critical tool for high throughput screening of novel and more effective treatments against AGD.

Keywords: AGD; Atlantic salmon; Neoparamoeba perurans; high throughput; screening.