Shrimp microsporidian EHP spores in culture water lose activity in 10 days or can be inactivated quickly with chlorine

Abstract Hepatopancreatic microsporidiosis (HPM) caused by Enterocytozoon hepatopenaei (EHP) is a highly prevalent and hard to eradicate parasite in major shrimp aquaculture hubs. Laboratory models to study the spread of EHP have revealed horizontal transmission via spores released from infected shrimp into rearing water. However, infectivity duration for the released spores is unknown. Thus, we carried out experiments using aquaria in which shrimp heavily-infected with EHP were held for 4 weeks after which the presence of spores in the water was confirmed by SybrGreen qPCR, and spore activity was determined by polar tube extrusion assay. After the shrimp were removed, the EHP-water was used directly (100%), diluted 50% or diluted 75% before adding naive PL-12 and rearing for 16 days. Infection was determined by PCR and in situ hybridization (ISH) and compared to the results obtained using a standard EHP-cohabitation model. Infections developed at all dilutions by day 12 (100, 66 and 33% positive samples respectively). During the second experiment, the EHP-water was subjected to chemical (20 ppm calcium hypochlorite) and to resting with aeration for 5 days and 10 days before addition of naive PL-12 to determine its infectivity. Within 16 days, the PL-12 in untreated EHP-water and in 5-days rested water developed EHP infections (50% and 10%, respectively), while the PL-12 cultured in EHP-water treated with 20 ppm calcium hypochlorite and in 10-days rested water remained uninfected. Altogether results showed that EHP contaminated water is a potential transmission risk but that chemical disinfection or resting for a suitable period in the absence of carriers can substantially reduce its infectivity.