Received: 14 December 2018 | Revised: 28 February 2019 | Accepted: 1 March 2019 DOI: 10.1111/jfd.12998 ORIGINAL ARTICLE The Three‐spined Stickleback, Gasterosteus aculeatus Linnaeus 1758, plays a minor role as a host of Lepeophtheirus salmonis (Krøyer 1837) in the Gulf of Maine Michael Pietrak1 | Alexander Jensen2,3 | Gayle Barbin Zydlewski3 | Ian Bricknell3,4 1 USDA, Agricultural Research Service, National Cold Water Marine Aquaculture Center, Franklin, Maine Department of Fisheries and Wildlife, Michigan State University, East Lansing, Michigan 2 3 School of Marine Sciences, University of Maine, Orono, Maine 4 USA Aquaculture Research Institute, University of Maine, Orono, Maine Correspondence Ian Bricknell, School of Marine Sciences, University of Maine, Orono, ME. Email: Ian.Bricknell@maine.edu Funding information NSF EPSCoR SEANET Program, Grant/ Award Number: Federal Grant Number: 1355457; Maine Sea Grant development funds, Grant/Award Number: DV‐12‐08 Abstract The sea louse, Lepeophtheirus salmonis (Krøyer 1837), is a significant parasite of farmed salmon throughout the Northern Hemisphere. Management of on‐farm louse populations can be improved by understanding the role that wild fish play in sustain‐ ing and providing refuge for the local population of sea lice. In this study, 1,064 stick‐ lebacks were captured. Of these animals, 176 individuals were carrying a total of 238 sea lice, yielding a prevalence and intensity of 16.5% and 1.4 lice per fish, respec‐ tively. Detailed examination of the sea lice on the three‐spined sticklebacks captured in Cobscook Bay found two L. salmonis individuals using three‐spined sticklebacks as hosts. A 2012 survey of wild fish in Cobscook Bay, Maine, found multiple wild hosts for Caligus elongatus (von Nordmann 1832), including three‐spined sticklebacks (Gasterosteus aculeatus L.), but no L. salmonis were found in this earlier study. KEYWORDS Gasterosteus aculeatus, Lepeophtheirus salmonis, sea lice, three‐spined stickleback 1 | I NTRO D U C TI O N continued development and optimization of control measures is im‐ portant as it is unlikely that any single measure will be completely The salmon louse (Lepeophtheirus salmonis (Krøyer 1837)) is a signif‐ effective; instead, successful management will depend on an effec‐ icant parasite of farmed Atlantic salmon (Salmo salar Linnaeus 1758) tive IPMS. throughout the Northern Hemisphere costing the salmon industry As IPMSs increase in complexity through the inclusion of a variety an estimated $742 million in 2012 (Roth, 2015). The development of of ecological and veterinary control measures, each targeting differ‐ resistance to the primary antiparasitic drugs used to for lice (Aaen, ent life stages or vulnerabilities, it will be important to consider host– Helgesen, Bakke, Kaur, & Horsberg, 2015; Horsberg, 2012; Jones, parasite relationships both on the farm and within the environment. Hammell, Dohoo, & Revie, 2012; Roth, 2015) is shifting management Lice often utilize wild populations as hosts where they are not ex‐ strategies from drugs to integrated pest management strategies posed to various veterinary control measures. Additionally, L. salmo- (IPMSs). These often incorporate other strategies such as “all‐in– nis have been found on non‐salmonid hosts such as saithe (Pollachius all‐out” management, fallowing and the use of cleaner fish (Bron, virens Linnaeus 1758) in the UK (Bricknell, Bron, & Bowden, 2006) Sommerville, Wootten, & Rae, 1993; Costello, 2006; Imsland et al., and three‐spined sticklebacks (Gasterosteus aculeatus Linnaeus 2014), integrated multitrophic aquaculture and selective breeding or 1758) in British Columbia and Newfoundland, Canada (Eaves, Ang, new engineering solutions (Bartsch et al., 2013; Gjerde, Odegard, & Murray, 2014; Jones, Prosperi‐Porta, Kim, Callow, & Hargreaves, & Thorland, 2011; Imsland et al., 2014; Molloy, Pietrak, Bouchard, 2006). Knowing what potential wild fish populations can serve as & Bricknell, 2011; Stien et al., 2016,2012; Webb et al., 2013). The hosts for various life stages of L. salmonis will help identify risk. J Fish Dis. 2019;1–5. wileyonlinelibrary.com/journal/jfd © 2019 John Wiley & Sons Ltd | 1 2 | PIETRAK ET Al. In Cobscook Bay, Maine, USA, as part of a site rotation and “all‐ each fish in the field (see below) prevented the loss of any mobile in–all‐out” area management plan, growers stock their sites once stages of lice during transportation. All individuals selected for ex‐ every three years in an effort to fallow the bay to reduce sea lice, amination were weighed, measured and then preserved in 95% eth‐ other pathogens and potential benthic impact. Recent sentinel cage anol (EtOH; Fisher Scientific) and brought back to the laboratory for studies conducted by the University of Maine show a decline in sea microscopic examination (VWR VistaVision). louse infectious pressure in the spring following a fallow (Bricknell & All observed lice were visually identified to species (Caligus elon- Frederick, Pietrak & Bricknell unpublished data). Few studies have gatus (von Nordmann 1832), L. salmonis or unknown) and life his‐ examined wild populations of fish in the Gulf of Maine for sea lice, tory stage (chalimus, pre‐adult (for L. salmonis) or the adult louse), resulting in a poor understanding of how lice interact with wild and removed from the fish and then preserved in 95% EtOH for DNA se‐ farmed fish in this system (Jensen, Zydlewski, Barker, & Pietrak, quencing. All of the undetermined lice (n = 44) and L. salmonis (n = 1) 2016; Shaw & Opitz, 1996). Developing a better understanding of and a minimum 10% of the C. elongatus from each sample (total se‐ this interaction is important to future louse management strategies lected n = 71) were identified to the species level via the mitochon‐ in Maine (USA) given that wild Atlantic salmon hosts are very de‐ drial cytochrome c oxidase I (COI) PCR developed by McBeath et al. pleted (Anon., 2014). In particular, understanding whether non‐sal‐ (2006) and subsequent DNA sequencing. monid hosts such as three‐spined sticklebacks serve an important Methods for the DNA isolation and PCR followed those of Jensen role as temporary hosts as they appear to do in British Columbia et al. (2016). DNA was extracted using DNeasy® Blood and Tissue (Jones et al., 2006) would be important. Kit (Qiagen, MD, USA). The louse tissue was lysed by removing the In 2012, Jensen et al. (2016) examined 35 species of fish caught EtOH from each individual louse and then incubating the louse over‐ in 2012 in Cobscook Bay for the presence of sea lice and found no night at 56°C in 180 µl of Buffer ATL and 20 µl of proteinase K. The L. salmonis on potential hosts. Three‐spined sticklebacks were the remaining steps of the manufacturer's protocol for animal tissues most dominant species caught, whereas known potential hosts such were followed until the elution of the DNA from the spin column. as saithe and pollock were rare (Jensen et al., 2016). To confirm the DNA was eluted from the spin column twice into separate 1.5‐ml results of the 2012 sampling (Jensen et al., 2016) and validate the tubes, as recommended by the manufacturer, using only 50 µl of AE fact that three‐spined sticklebacks are not hosts for L. salmonis in buffer instead of the 200 µl called for in the instructions. Cobscook Bay, an additional survey was conducted in 2013. The COI gene was amplified using the universal primers LCO1490 and HCO2198 (Folmer, Black, Hoeh, Lutz, & Vrijenhoek, 1994). PCR products were then purified using the QIAquick PCR purification kit 2 | M E TH O DS (Qiagen, Maryland, USA) and were sequenced at the University of Maine DNA Sequencing Facility with the HCO2198 primer. Edited Cobscook Bay is a macrotidal estuary located near the border be‐ sequences were run through Standard Nucleotide BLAST for tween Maine and Canada in the Western North Atlantic Ocean. The matches with known sequences. bay is subdivided into three regions: the inner bay, central bay and outer bay, with commercial Atlantic salmon farm leases primarily in the outer bay and one lease site in the central bay (http://www. maine.gov/dmr/aquaculture/leaseinventory/cobscookbay.htm, 3 | R E S U LT S ac‐ cessed on 10 March 2016). The bay has an average depth of 10 m A total of 1,064 fish were examined for lice in 2013. Among the and maintains salinities above 30 ppt with little freshwater input, 1,064 fish, 176 individuals were infested by a total of 238 sea lice, and temperatures that typically range between 0 and 12°C (Larsen, yielding an observed louse infestation prevalence and intensity of 2004). 16.5% and 1.4 lice per fish, respectively. All 238 observed lice were Three‐spined Sticklebacks were collected in May, June, visually identified as either C. elongatus or unknown, except for a sin‐ August, September and November 2013 using beach seine nets, gle louse that was visually identified as a chalimus‐stage L. salmonis 30.48 m x 1.83 m in dimension with 0.64 cm diamond mesh, from (Table 2). The visual identifications of all 71 tested C. elongatus and two sites in the outer bay (Carrying Place Cove and Broad Cove), the solitary L. salmonis individual were confirmed through DNA se‐ three sites in the central bay (East Bay, Pennamaquan River and quencing. Of the 44 sea lice unidentified via visual examinations, 35 South Bay) and two sites in the inner bay (Denny's Bay and Cobscook were identified as C. elongatus, an additional specimen was identi‐ Bay State Park; Figure 1). Sampling was conducted at slack high tide fied as L. salmonis, and 8 remained unidentified, not giving a positive during the day with some sites also being sampled at slack high at PCR result to either the L. salmonis or C. elongatus primers, and were night (Table 1). presumed to be other members of the Siphonostomatoida clade. All At each site and collection time, up to 30 individual three‐spined genetically identified C. elongatus were further confirmed as C. elon- sticklebacks were haphazardly selected per seine, based on the first gatus genotype 1 (Øines & Heuch, 2005). The two sequences identi‐ 30 fish removed from the bucket of fish collected in the sampling net, fied as L. salmonis via DNA sequencing were 647 and 674 base pairs, and killed with an overdose of MS222 (250 mg/L; Argent Chemical respectively. They had 86% and 87% coverage and were 99% identi‐ Laboratories) for later examination. The individual preservation of cal with an E value of 0.0 to L. salmonis isolate Ls CM12 (accession | PIETRAK ET Al. 3 F I G U R E 1 (a) The location of the research area in relation to Maine. (b) The 2 stickleback collection sites in the outer bay (Carrying Place Cove and Broad Cove), the 3 sites in the central bay (East Bay, Pennamaquan River and South Bay) and the 2 sites in the inner bay (Denny's Bay and Cobscook Bay State Park) TA B L E 1 Summary of sampling effort during each sampling period. DS = sampled at high slack tide during the day; NS = sampled at high slack tide at night. When a site was not sampled either during the day or at night, it is labelled as “Not Sampled” Sampling site May June August September November Denny's Bay DS DS DS DS Sampled Whiting Bay Not sampled DS DS DS Not sampled East Bay DS DS DS DS Sampled South Bay DS DS DS Not sampled Not sampled Pennamaquan River DS DS DS DS Not sampled Carrying Place Cove DS DS DS DS Sampled Broad Cove DS DS DS DS Not sampled AY602748.1). Both of the identified L. salmonis were in a chalimus Three‐spined Sticklebacks as a host in the Gulf of Maine ecosystem. stage and were found in Denny's Bay and East Bay in September and Jensen et al. (2016) examined a total of 1,996 three‐spined stick‐ November 2013, respectively. lebacks in 2012 for C. elongatus or L. salmonis infections with all of the lice examined both visually and through DNA sequencing being C. elongatus. The uncommon use of Three‐spined Sticklebacks as 4 | D I S CU S S I O N hosts in the Gulf of Maine is consistent with the only other report of L. salmonis on Three‐spined Sticklebacks in the Atlantic (Eaves et al., The finding of the two L. salmonis chalimi on three‐spined stick‐ 2014). They examined 822 Three‐spined Sticklebacks from the Bay lebacks in 2013 represents the first report of L. salmonis using d'Espoir in Newfoundland and found 3 chalimus‐stage L. salmonis. 4 | PIETRAK ET Al. Visual sea lice identifica‐ tions DNA‐based verification of visually identified C. elongatus DNA‐based verification of visually identified L. salmonis DNA‐based verification of visually identified “unknown” lice C. elongatus 193 71 0 35 L. salmonis 1 0 1 1 Unknown 44 0 0 8 1 44 Total a 238 71 a TA B L E 2 Summary of louse observations and identifications from the 1,064 three‐spined sticklebacks examined as part of the 2013 survey Only a subset of the visually identified C. elongatus were verified using DNA sequencing. There are no additional reports of L. salmonis utilizing Three‐spined trutta L.) and rainbow trout Oncorhynchus mykiss (Walbaum, 1792) Sticklebacks as hosts from the North Atlantic Ocean. to run to sea in the area too. These salmonids may act as a reservoir The infrequent use of Three‐spined Sticklebacks as an alterna‐ for sea lice. A very small extant population of Atlantic salmon is also tive host in the Atlantic Ocean differs from the more widespread reported in the Denny's River (Anon., 2014). Given the small size of use of Three‐spined Sticklebacks as hosts for L. salmonis in the the wild salmonid populations, it is not likely that they are a major Northeast Pacific Ocean. Jones and Prosperi‐Porta (2011) reported source of L. salmonis in the area. that during a 4‐year period, the prevalence of Lepeophtheirus spp. The finding of two L. salmonis chalimus stages attached to Three‐ on Three‐spined Sticklebacks ranged from 11% to 50%. They col‐ spined Sticklebacks from Cobscook Bay confirms the potential use of lected 10,128 individual Lepeophtheirus spp. with 71% being L. sal- this species as an alternative host in Cobscook Bay. It appears that the monis and the remainder being Lepeophtheirus cuneifer (Kabata 1974; use of Three‐spined Sticklebacks is much less prevalent in the Atlantic Jones & Prosperi‐Porta, 2011). The apparent difference in the use Ocean than their use in the Pacific Ocean, which possibly reflects an of Three‐spined Sticklebacks in the Atlantic and Pacific may reflect adaptation of the subspecies found in the Pacific Ocean. The rare use fundamental host choice preferences and supports the recent split of wild fish as hosts for L. salmonis, observed both in this study and in of L. salmonis into two sub species: Lepeophtheirus salmonis salmo- Jensen et al. (2016), would further support the use of fallowing as an nis in the Atlantic and L. salmonis oncorhynchi in the Pacific (Skern‐ important tool to manage sea louse populations in the area. This is an Mauritzen, Torrissen, & Glover, 2014). The decline in wild Atlantic important finding as it will inform the current IPMS and future develop‐ salmon in Maine is also a major consideration. The lack of a signifi‐ ment of improved IPMS for the Cobscook Bay region. As Three‐spined cant wild Atlantic salmon population in the study area that could act Sticklebacks are not a significant wild reservoir for L. salmonis mobile as hosts for L. salmonis has generated several hypotheses to the ori‐ or sessile stages, they should not be seen as a key indicator for the in‐ gin of the pioneer sea louse populations that establish infections on teractions of sea lice between wild and farmed L. salmonis populations. the farmed Atlantic salmon populations after the whole of Cobscook Bay's commercial salmon farms have been fallowed. This study has demonstrated that the prevalence of L. salmonis on the stickleback AC K N OW L E D G E M E N T S population is low (0.002), and it is unlikely that stickleback are a major We would like to thank J. McCleave, G. Staines, M. Altenritter and source of lice. Hypothetically, it is possible that this low prevalence B. Fleenor who assisted in the fish survey efforts and D. Noyes, G. level could introduce lice onto a farm. Hypothetically, it is possible Andrews, T. Van Kirk and B. Smith for their assistance in process‐ that this low prevalence level could introduce lice onto a farm. But ing sticklebacks and lice. Fish sampling was part of a larger project, when compared to prevalence of other diseases that have been stud‐ led by G. Zydlewski with J. McCleave and J. Vieser and supported ies in wild fish when a higher prevalence was found compared to sea in part by the US Department of Energy. The views expressed lice (such as infectious viral haemorrhagic septicaemia (VHS) where herein are those of the authors and do not necessarily reflect the prevalence was found to be 0.006 (Sandlund et al., 2014)) there was view of the Ocean Renewable Power Company or any of its sub‐ been no evidence of direct transmission at this level to farmed fish. agencies. Sea louse identification and laboratory‐based procedures It is unlikely, given the current information, that Three‐spined were made possible through Maine Sea Grant development funds Sticklebacks are an important host in the Atlantic though they may (DV‐12‐08), the NOAA Sea Grant Aquaculture Research Program present a viable wild host for L. salmonis in the Pacific Northwest. and the NSF EPSCoR SEANET programme. Fish sampling was con‐ It is not possible to say how common the use of wild salmonids as a ducted under the University of Maine Institutional Animal Care and host in Cobscook Bay is because the fish survey did not capture any Use Committee Protocol # A2010‐03‐01. Mention of trade name, salmonids in the area; however, they are known to exist in the bay, proprietary product, or specific equipment does not constitute a presumably in very low numbers. There are small populations of sea‐ guarantee or warranty by the US Department of Agriculture and run brook trout (Salvelinus fontinalis (Mitchell 1814)) and there is the does not imply approval to the exclusion of other products that may potential for stocked introduced populations of brown trout (Salmo be suitable. | PIETRAK ET Al. C O N FL I C T O F I N T E R E S T The authors declare that they have no conflicts of interest in this research. ORCID Ian Bricknell https://orcid.org/0000‐0001‐6042‐7143 REFERENCES Aaen, S. M., Helgesen, K. O., Bakke, M. J., Kaur, K., & Horsberg, T. E. (2015). Drug resistance in sea lice: a threat to salmonid aquacul‐ ture. Trends in Parasitology, 31, 72–81. https://doi.org/10.1016/j. pt.2014.12.006 Anon. (2014). Annual report of the U.S. Atlantic Salmon Assessment Committee. In (ed. by U.S.a.S.A. Committee), p. 282, Kittery, ME. Bartsch, A., Robinson, S. M. C., Liutkus, M., Ang, K. P., Webb, J., & Pearce, C. M. (2013). Filtration of sea louse, Lepeophtheirus salmonis, copepodids by the blue mussel, Mytilus edulis, and the Atlantic sea scallop, Placopecten magellanicus, under different flow, light and copepodid‐density regimes. Journal of Fish Diseases, 36, 361–370. Bricknell, I. R., Bron, J. E., & Bowden, T. J. (2006). Diseases of gadoid fish in cultivation: a review. ICES Journal of Marine Science, 63, 253–266. https://doi.org/10.1016/j.icesjms.2005.10.009 Bron, J. E., Sommerville, C., Wootten, R., & Rae, G. H. (1993). Fallowing of marine Atlantic Salmon, Salmo salar L, farms as a method for the con‐ trol of sea lice, Lepeophtheirus salmonis (Kroyer, 1837). Journal of Fish Diseases, 16, 487–493. https://doi.org/10.1111/j.1365‐2761.1993. tb00882.x Costello, M. J. (2006). Ecology of sea lice parasitic on farmed and wild fish. Trends in Parasitology, 22, 475–483. https://doi.org/10.1016/j. pt.2006.08.006 Eaves, A. A., Ang, K. P., & Murray, H. M. (2014). Occurrence of the par‐ asitic copepod Ergasilus labracis on threespine sticklebacks from the South Coast of Newfoundland. Journal of Aquatic Animal Health, 26, 233–242. Folmer, O., Black, M., Hoeh, W., Lutz, R., & Vrijenhoek, R. (1994). DNA primers for amplification of mitochondrial cytochrome C oxidase subunit I from diverse metazoan invertebrates. Molecular Marine Biology and Biotechnology, 3, 294–299. Gjerde, B., Odegard, J., & Thorland, I. (2011). Estimates of genetic varia‐ tion in the susceptibility of Atlantic salmon (Salmo salar) to the salmon louse Lepeophtheirus salmonis. Aquaculture, 314, 66–72. https://doi. org/10.1016/j.aquaculture.2011.01.026 Horsberg, T. E. (2012). Avermectin use in aquaculture. Current Pharmaceutical Biotechnology, 13, 1095–1102. Imsland, A. K., Reynolds, P., Eliassen, G., Hangstad, T. A., Foss, A., Vikingstad, E., & Elvegård, T. A. (2014). The use of lumpfish (Cyclopterus lumpus L.) to control sea lice (Lepeophtheirus salmonis Krøyer) infestations in intensively farmed Atlantic salmon (Salmo salar L.). Aquaculture, 424–425, 18–23. Jensen, A. J., Zydlewski, G. B., Barker, S., & Pietrak, M. (2016). Sea lice in‐ festations of a wild fish assemblage in the Northwest Atlantic Ocean. Transactions of the American Fisheries Society, 145, 7–16. https://doi. org/10.1080/00028487.2015.1091381 Jones, P. G., Hammell, K. L., Dohoo, I. R., & Revie, C. W. (2012). Effectiveness of emamectin benzoate for treatment of Lepeophtheirus salmonis on farmed Atlantic salmon Salmo salar in the Bay of Fundy, Canada. Diseases of Aquatic Organisms, 102, 53–64. https://doi. org/10.3354/dao02517 5 Jones, S. R. M., & Prosperi‐Porta, G. (2011). The diversity of sea lice (Copepoda: Caligidae) parasitic on threespine stickleback (Gasterosteus Aculeatus) in Coastal British Columbia. Journal of Parasitology, 97, 399–405. https://doi.org/10.1645/GE‐2617.1 Jones, S. R. M., Prosperi‐Porta, G., Kim, E., Callow, P., & Hargreaves, N. B. (2006). The occurrence of Lepeophtheirus salmonis and Caligus clemensi (Copepoda: Caligidae) on three‐spine stickleback Gasterosteus aculeatus in coastal British Columbia. Journal of Parasitology, 92, 473– 480. https://doi.org/10.1645/GE‐685R1.1 Larsen, P. F. (2004). Introduction to ecosystem modeling in Cobscook Bay, Maine: a boreal, macrotidal estuary. Northeastern Naturalist, 11, 1– 12. https://doi.org/10.1656/1092‐6194(2004)11[1:ITEMIC]2.0.CO;2 Mcbeath, A. J. A., Penston, M. J., Snow, M., Cook, P. F., Bricknell, I. R., & Cunningham, C. O. (2006). Development and application of real‐time PCR for specific detection of Lepeophtheirus salmonis and Caligus elongatus larvae in Scottish plankton samples. Diseases of Aquatic Organisms, 73, 141–150. https://doi.org/10.3354/ dao073141 Molloy, S. D., Pietrak, M. R., Bouchard, D. A., & Bricknell, I. (2011). Ingestion of Lepeophtheirus salmonis by the blue mussel Mytilus edulis. Aquaculture, 311, 61–64. https://doi.org/10.1016/j. aquaculture.2010.11.038 Øines, Ø., & Heuch, P. A. (2005). Identification of sea louse species of the genus Caligus using mtDNA. Journal of the Marine Biological Association of the United Kingdom, 85, 73–79. https://doi.org/10.1017/ S0025315405010854h Roth, M. (2015). Management of sea lice on farmed salmon with veteri‐ nary medicines and biological control strategies. International Animal Health Journal, 2, 32–37. Sandlund, N., Gjerset, B., Bergh, O., Modahl, I., Olesen, N. J., & Johansen, R. (2014). Screening for viral hemorrhagic septicemia virus in marine fish along the Norwegian Coastal Line. PLoS One, 9, e108529. Shaw, R., & Opitz, M. (1996). Abundance of the parasitic copepod Caligus elongatus on wild pollock near commercial salmonid net‐pens. Journal of Aquatic Animal Health, 8, 75–77. Skern‐Mauritzen, R., Torrissen, O., & Glover, K. A. (2014). Pacific and Atlantic Lepeophtheirus salmonis (Krøyer, 1838) are allopat‐ ric subspecies: Lepeophtheirus salmonis salmonis and L. salmonis oncorhynchi subspecies novo. BMC Genetics, 15, 32. https://doi. org/10.1186/1471‐2156‐15‐32 Stien, L. H., Dempster, T., Bui, S., Glaropoulos, A., Fosseidengen, J. E., Wright, D. W., & Oppedal, F. (2016). ‘Snorkel’ sea lice barrier tech‐ nology reduces sea lice loads on harvest‐sized Atlantic salmon with minimal welfare impacts. Aquaculture, 458, 29–37. Stien, L. H., Nilsson, J., Hevroy, E. M., Oppedal, F., Kristiansen, T. S., Lien, A. M., & Folkedal, O. (2012). Skirt around a salmon sea cage to reduce infestation of salmon lice resulted in low oxygen levels. Aquacultural Engineering, 51, 21–25. https://doi.org/10.1016/j. aquaeng.2012.06.002 Webb, J. L., Vandenbor, J., Pirie, B., Robinson, S. M. C., Cross, S. F., Jones, S. R. M., & Pearce, C. M. (2013). Effects of temperature, diet, and bivalve size on the ingestion of sea lice (Lepeophtheirus salmonis) lar‐ vae by various filter‐feeding shellfish. Aquaculture, 406–407, 9–17. https://doi.org/10.1016/j.aquaculture.2013.04.010 How to cite this article: Pietrak M, Jensen A, Barbin Zydlewski G, Bricknell I. The Three‐spined Stickleback, Gasterosteus aculeatus Linnaeus 1758, plays a minor role as a host of Lepeophtheirus salmonis (Krøyer 1837) in the Gulf of Maine. J Fish Dis. 2019;00:1–5. https://doi.org/10.1111/ jfd.12998