PERACETIC ACID AS A TREATMENT FOR AMOEBIC GILL DISEASE: EFFICACY AND PHYSIOLOGICAL RESPONSES OF ATLANTIC SALMON

Introduction

 Amoebic gill disease (AGD) is a parasitic infestation affecting Atlantic salmon, particularly during  the seawater phase of production. It is c aused by the amphizoic marine amoeba Neoparamoeba perurans. Upon attachment on the gills, the  amoeba trophozoites elicit a localis ed tissue response including epithelial hyperplasia, hypertrophy and lamellar fusion and the gross pathology is characterised by raised white mucoid lesions . These white plaques are used to grade the severity of AGD infection through visual inspection. Freshwater and oxidative therapeutic s are used to treat AGD, the former being the  most commonly used. Despite offering alternative options for freshwater bathing, oxidative therapeutics often have conflicting treatment resolutions. In addition, there is a significant knowledge gap on  the underlying physiological processes on  how salmon respond to these treatments. In this study, we explored the efficacy of peracetic acid (PAA) in treating AGD and  the physiological responses of salmon smolts during the treatment.

Materials and methods

AGD in smolts was induced by bath exposure to the parasites for 1 hr. Thereafter, the disease was allowed to develop with routine monitoring of the gill score (GS 0 = clear of lesions → GS 5 = extensive lesions). When the gill score (GS) reached 1-2, fish were treated with PAA via bath exposure. Three commercial PAA products were tested (Perfektoxid , AQUADes and ADDIAqua) at a concentration of 5 ppm. Two exposure du rations were evaluated – 30 mins and 60 mins.   Uninfected fish were likewise treated similarly. Sampling was performed 1 day and 2 weeks after treatment. The disease status of the fish was assessed by visual pathology of the gills and qPCR quantification of the parasites (gill swabs and water samples). The gills and olfactory organs were subjected to histological evaluation and gene expression analysis of key biomarkers for oxidative stress. Systemic oxidative stress was determined by quantifying the level of reactive oxygen species and total antioxidant capacity in plasma.

 Results  and Discussion

Gill score and parasite load: The infected-untreated group had an average GS of 2.8 two weeks post-treatment . Except for  the  group treated with ADDIAqua, groups treated with PAA for 1 hr had an almost similar GS with the infected-untreated group.  Fish treated  with PAA for 30 mins registered lower GS than the infected-untreated and 1 hr-treated groups . The lowest GS  (1.9)  was in AQUADes treated group, followed by ADDIAqua (2.1) , then with Perfektoxid (2.4) for 30 mins . qPCR analysis of the parasite on the gills did not provide a clear tendency either among the groups or between sampling points. The level of the parasite in the rearing water increased  through time in all PAA-treated groups, which might indicate an ongoing parasite shedding post-treatment.

Histology: AGD-infected fish displayed the typical pathology, which was increased hyperplasia. Two weeks post-treatment, t he number of lamellar hyperplasia cases in  the  30-min treated group was lower than the infected-untreated and 1-hr PAA-treated groups. This increased hyperplasia likewise resulted in increased lamellar fusion. Epithelial lifting was prevalent in all groups. The mucous cell number in the lamella but not in the filament increased following treatment, and neither inter-treatment nor temporal differences were found to influence this profile . PAA-treated fish exhibited a compromised olfactory organ architecture two weeks post-treatment. There was a tendency for the mucosal tip of the olfactory lamella  to  increase in size especially in Perfektoxid -treated group, regardless of the exposure duration. The thickness of the lamellar epithelium and  lamina propia did not demonstrate substantial changes among treatment groups and through time.

 Gene expression:  Thirteen biomarkers for oxidative stress were evaluated in the gills and olfactory organs. Overall, PAA treatments resulted in the modulation of the expression of the marker genes and the changes were pronounced 1 day after than at 2 weeks post-treatment. This indicates that mucosal oxidative stress was triggered, though not chronic. Unlike in other response variables where treatment duration elicited strong differential responses among the groups, the gene expression profile in the gills and olfactory organ revealed no such general distinction.

Systemic oxidative stress: ROS levels in the plasma increased through time, especially in the group treated for 30 mins. The groups exposed to AQUADes and Perfektoxid for 30 mins had higher ROS levels than the infected-untreated group two weeks post-treatment.  There was a large fish-to-fish variation in the total antioxidant capacity (TAC) in the plasma of the experimental fish . No significant differences in plasma TAC were found among t reatments and  between  sampling points .

Conclusions

 This study demonstrated that PAA treatment of AGD  in salmon smolts  resulted in quite varied response profiles. Disease resolution was not fully established as assessed  by visual pathology, histopathology and qPCR, though there were indications that the type of PAA product might play a role in its treatment efficacy . Future studies should be directed at standardising the  PAA  treatment protocol. From physiological aspects, salmon were able to respond to PAA treatments despite being in a diseased state. AGD however, could trigger changes in the expression  PAA was demonstrated to trigger local oxidative stress, though the heightened state did not last long.

Acknowledgment

 This research is part of the PERAGILL project funded by the Norwegian Seafood Research Fund (ref. 901472).