Salmo Salar) Siblings Following Experimental Infection with Neoparamoeba Perurans, Causative Agent of Amoebic Gill Disease
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1229 A comparison of disease susceptibility and innate immune response between diploid and triploid Atlantic salmon (Salmo salar) siblings following experimental infection with Neoparamoeba perurans, causative agent of amoebic gill disease LYNN CHALMERS*, JOHN F. TAYLOR, WILLIAM ROY, ANDREW C. PRESTON, HERVE MIGAUD and ALEXANDRA ADAMS Institute of Aquaculture, University of Stirling, Stirling, FK9 4LA, UK (Received 27 September 2016; revised 5 April 2017; accepted 7 April 2017; first published online 11 May 2017) SUMMARY Few studies have focussed on the health and immunity of triploid Atlantic salmon and therefore much is still unknown about their response to commercially significant pathogens. This is important if triploid stocks are to be considered for full-scale commercial production. This study aimed to investigate and compare the response of triploid and diploid Atlantic salmon to an experimental challenge with Neoparamoeba perurans, causative agent of amoebic gill disease (AGD). This disease is economically significant for the aquaculture industry. The results indicated that ploidy had no significant effect on gross gill score or gill filaments affected, while infection and time had significant effects. Ploidy, infec- tion and time did not affect complement or anti-protease activities. Ploidy had a significant effect on lysozyme activity at 21 days post-infection (while infection and time did not), although activity was within the ranges previously recorded for sal- monids. Stock did not significantly affect any of the parameters measured. Based on the study results, it can be suggested that ploidy does not affect the manifestation or severity of AGD pathology or the serum innate immune response. Additionally, the serum immune response of diploid and triploid Atlantic salmon may not be significantly affected by amoebic gill disease. Key words: triploid, amoebic gill disease, AGD, cohabitation, immune response. INTRODUCTION Atlantic salmon (Salmo salar), rainbow trout (Oncorhynchus mykiss), brown trout (Salmo trutta), Sexual maturation in fish causes the transfer of turbot (Scophthalmus maximus) and grass carp energy from normal somatic growth to gonadal (Ctenopharyngodon idella) (Tiwary et al. 2004; development. This can have adverse effects on Maxime, 2008; Piferrer et al. 2009; Preston et al. body growth rates and flesh quality, and may 2013). This ‘shock’ treatment prevents second increase incidences of disease and mortality meiotic division and causes the retention of the (Ojolick et al. 1995; Leclercq et al. 2011). Sexually second polar body, which results in three sets of mature fish that escape from production sites also chromosomes rather than two, and in turn sterility have the potential to interact with wild fish, impact- in triploid fish (Tiwary et al. 2004; Maxime, 2008). ing on the genetics and fitness of wild populations However, despite the clear advantages of being (Glover et al. 2013). As such, sexual maturation is sterile, if triploid Atlantic salmon are to be consid- of serious concern for the salmonid aquaculture ered for commercial production, they must industry and work continues to derive a solution to perform as well as diploids in all aspects of their these problems. Triploidy is the only commercially biology and physiology. Recent studies have contin- available and acceptable means of achieving sterility ued to explore and elucidate the physiology and per- in fish, and is increasingly being used as a method to formance of triploid salmon. These results show that control sexual maturation (Oppedal et al. 2003; the commonly reported problems of deformity, poor Taylor et al. 2011). Triploidy can be readily survival and reduced growth in triploids can be induced through the application of a hydrostatic or addressed through refined husbandry, feeds and temperature ‘shock’ to newly-fertilized eggs and management, thus further supporting the applica- the process has been optimized for several commer- tion of triploids in the aquaculture industry (Burke cially important species in aquaculture including et al. 2010; Fjelldal and Hansen, 2010; Leclercq * Corresponding author: Institute of Aquaculture, et al. 2011; Taylor et al. 2011, 2012). However, University of Stirling, Stirling, FK9 4LA, UK. E-mail: very few studies have focussed on triploid Atlantic [email protected] salmon health and immunity. Parasitology (2017), 144, 1229–1242. © Cambridge University Press 2017. This is an Open Access article, distributed under the terms of the Creative Commons Attribution licence (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted re-use, distribu- tion, and reproduction in any medium, provided the original work is properly cited. Downloaded fromdoi:10.1017/S0031182017000622 https://www.cambridge.org/core. IP address: 170.106.40.40, on 25 Sep 2021 at 10:13:24, subject to the Cambridge Core terms of use, available at https://www.cambridge.org/core/terms. https://doi.org/10.1017/S0031182017000622 Lynn Chalmers and others 1230 Understanding how triploid fish cope with health MATERIALS AND METHODS challenges is an important milestone to characterize Ethical approval their robustness, especially as disease and subsequent health issues continue to restrict the development Experimental procedures were approved by the and success of the aquaculture industry (Weber Animal Welfare and Ethical Review Body et al. 2013). Anecdotal evidence suggests no differ- (AWERB) of the University of Stirling and were ences in mortality of triploid salmon compared completed under UK Government Home Office with diploid siblings when challenged with disease project licence 60/4189. The euthanisation of fish in commercial settings although no scientific assess- for sampling was carried out according to the UK ments were carried out. A study by Frenzl et al. Government Home Office Schedule 1 regulations. (2014) showed similar infection levels between ploidy when challenged with parasitic sea lice, Fish stock & history Lepeophtheirus salmonis, in both experimental and natural challenge conditions. In contrast, higher Fish used in this study were obtained from two com- Gyrodactylus salaris infection levels in triploids mercial breeding companies (Stock A and Stock B) were reported by Ozerov et al. (2010). Additionally, and supplied as eyed ova (Stock A: 372°D, 21 triploid goldfish (Carassius auratus) showed higher December 2012; Stock B: 380°D, 4 January 2013) loads of Metagonnimus sp. than diploids (Hakoyama to the University of Stirling Freshwater Research et al. 2001) and findings from rainbow trout Facility (56°N 4°W). Triploidy was induced at showed triploids were more susceptible to infection both company sites using the same protocol, by Ergasilus sieboldin (Tildesley, 2008). However, whereby post-fertilization, half of each egg batch with an overall lack of conclusive data, the response (Stock A, 26 unrelated dams and 6 sires; Stock B, of triploid Atlantic salmon to economically signifi- 20 unrelated dams and 5 sires) was exposed to a cant parasites remains to be fully elucidated. hydrostatic pressure shock of 655 bar applied for One such parasite is Neoparamoeba perurans,a 6·25 min, 37·5 min post fertilization at 8 °C. Ova free-living, marine amoebae and the causative (3 tanks per ploidy per stock with 2500 ova per agent of amoebic gill disease (AGD) (Young et al. tank) were incubated in complete darkness at 2008a; Crosbie et al. 2012). AGD has represented 7·1 ± 0·3 °C until hatching, where temperature was a significant problem for the marine salmonid indus- gradually increased to 10 °C for first feeding (Stock try since the mid-1980’s, particularly in Tasmania, A: 25 February 2013; Stock B: 4 March 2013). At and has now become a persistent problem for a first feeding, fish were reared under constant light growing number of countries including Ireland, and fed a commercial diet (Diploids – Inicio Plus; Scotland and Norway (Oldham et al. 2016). Triploids – Inicio-TriX, BioMar UK), distributed Clinical signs of AGD include respiratory distress, by automatic feeders (Arvo-Tec Oy, Finland). flared opercula and lethargy, and macroscopic mani- Specific feeding rates (% tank biomass per day) festation of white mucoid patches on the gills were adjusted automatically according to predicted (Munday, 1986; Munday et al. 1990). Histological growth and daily temperature, and pellet size (0·5 examination shows profound changes to gill archi- to 2 mm) increased with fish size. From August tecture including extensive hyperplasia and lamellar 2013, fish were maintained under ambient tempera- fusion (Munday, 1986; Roubal et al. 1989; Adams ture (min: 3 °C, max: 15 °C) and photoperiod to and Nowak, 2003). It is now recognized that AGD produce S1+ smolts for sea transfer on 22 April is the most significant disease caused by gill parasites 2014 (Marine Environmental Research Laboratory in terms of economic impact (Shinn et al. 2015). (MERL), Institute of Aquaculture, Machrihanish, While numerous studies have been undertaken to UK, 55·4°N 5·7°W). Fish were vaccinated in elucidate the interaction of N. perurans with the November 2013 with Alphaject 2·2 (PHARMAQ, host fish immune system (Gross et al. 2005; UK). Mortality between first feeding and sea trans- Pennacchi et al. 2014, 2016), further research is fer was 1·18 and 1·99% in Stock A, and 2·8 and 3·5% required to fully explain the overall immune in Stock B, respectively for diploids and triploids. response of Atlantic salmon to AGD. This is par- Following sea transfer, fish were maintained under ticularly important in relation to triploids with a pre- ambient temperature (min: 9 °C, max: 11 °C) in 1 vious study showing reduced survival in triploids m stock tanks (400 L; 1 L kg biomass−1 min−1 flow when challenged with N. perurans but fewer gill rate) with aeration provided by air stones for 24 lesions than diploids and no ploidy differences in days until allocation to challenge tanks. During immune response (Powell et al. 2008). this period, mortality was 1·32% for diploids and The aim of this study was therefore to investigate 1·79% for triploids in Stock A, and 4·9% for diploids and compare the response of triploid and diploid and 0·3% triploids for Stock B.