Repeated sublethal freshwater exposures reduce the amoebic gill disease parasite, Neoparamoeba perurans, on Atlantic salmon

Abstract

Freshwater bathing is one of the main treatment options available against amoebic gill disease (AGD) affecting multiple fish hosts in mariculture systems. Prevailing freshwater treatments are designed to be long enough to kill Neoparamoeba perurans, the ectoparasite causing AGD, which may select for freshwater tolerance. Here, we tested whether using shorter, sublethal freshwater treatment durations are a viable alternative to lethal ones for N. perurans (2-4 hr). Under in vitro conditions, gill-isolated N. perurans attached to plastic substrate in sea water lifted off after ≥2 min in freshwater, but survival was not impacted until 60 min. In an in vivo experiment, AGD-affected Atlantic salmon Salmo salar subjected daily to 30 min (sublethal to N. perurans) and 120 min (lethal to N. perurans) freshwater treatments for 6 days consistently reduced N. perurans cell numbers on gills (based on qPCR analysis) compared to daily 3 min freshwater or seawater treatments for 6 days. Our results suggest that targeting cell detachment rather than cell death with repeated freshwater treatments of shorter duration than typical baths could be used in AGD management. However, the consequences of modifying the intensity of freshwater treatment regimes on freshwater tolerance evolution in N. perurans populations require careful consideration.

Evaluation of the Infectious Potential of Neoparamoeba perurans Following Freshwater Bathing Treatments

“…Lima et al [ 33 ] reported that amoeba survival in vitro is inversely related to freshwater exposure time, with survival after 30 min, 60 min, 2 h, 3 h and 4 h of 87.3%, 77.3%, 16.0%, 4.3% and 0%, respectively. These results differed from Wright et al [ 34 ], who found that pseudocysts do not survive for ≥1 h exposure in deionized water. The discrepancy between the two studies may be due to cultured strain differences or to dissimilar freshwater chemistry, which was not specified in either study.…”

“…Previous in vitro studies have shown that freshwater exposed N. perurans rapidly detach as floating pseudocysts, which recover to trophozoite form when returned to seawater and subsequently attach to plastic substrates. According to Wright et al [ 34 ] losses are negligible following short-term exposure to freshwater (2 to 30 min) when assessed 1 h after return to seawater, but no amoebae recovered after 1 h at 35 ppt following a 1 or 2 h freshwater exposure. However, it is probable that pseudocysts surviving an extended freshwater treatment will take a longer time to recover when returned to seawater.…”

“…Not only does the release of viable amoebae into the marine environment pose a localized risk of infection, there is concern that freshwater resistant N. perurans strains may develop over time because freshwater resistant strains may outcompete susceptible amoebae [ 34 ]. Evidence of increased frequency of freshwater bathing over the first 20 years of the Tasmanian salmon industry may suggest that freshwater exposure was becoming less effective [ 26 , 34 ]. However, this is likely also related to increased salmon biomass and farm proximity along with a shift to more proactive bath treatments at lower average gill score.…”

“…Over many years of liner-based freshwater treatment, the disposal of used AGD bathwater on farm sites was not considered to be a significant risk of reinfection, because it was understood that freshwater killed the majority of neoparamoebae [ 32 ]. However, in vitro studies have shown that N. perurans cells acutely exposed to freshwater will rapidly detach from the culture plate to form rounded pseudocysts [ 32 , 33 , 34 ] and that, following return to seawater conditions, many of these pseudocysts gradually return to amoeboid form and reattach to surfaces. Lima et al [ 33 ] reported that amoeba survival in vitro is inversely related to freshwater exposure time, with survival after 30 min, 60 min, 2 h, 3 h and 4 h of 87.3%, 77.3%, 16.0%, 4.3% and 0%, respectively.…”

Gill diseases in marine salmon aquaculture with an emphasis on amoebic gill disease.

“…Recently, Wright et al [120] have shown that targeting cell detachment rather than cell death with repeated freshwater treatments of shorter duration than typical baths could be used in AGD management. Thus, AGD-affected Atlantic salmon subjected daily to 30 min (sublethal) and 120 min (lethal) freshwater treatments for 6 days consistently reduced the N. perurans cell numbers on gills compared with daily 3 min freshwater or seawater treatments for 6 days.…”