Study design: The focus of our study was Hidden Lake, Banff National Park, Canada (Table 1), a high elevation mountain lake in the Canadian Rockies that received rotenone treatment during the summers of 2018 and 2019 to eradicate non-native brook trout (Salvelinus fontinalis) that were introduced and established in the 1970s, leading to the extirpation of a population of Westslope cutthroat trout (Oncorhynchus clarkii lewisi). The rotenone treatment was done accordingly to Montana State (USA) rotenone policy in the absence of a Canadian equivalent. This policy recommends two rotenone treatment for brook trout eradication because their spawning is not perfectly synchronous and because brook trout eggs in the gravel are not susceptible to rotenone (MFWP 2017). Moreover, several fish and traces of environmental DNA from brook trout were detected between the two rotenone treatments in summer 2018 and 2019 (Derry A, unpubl. data). The rotenone formulation applied to Hidden Lake (21-22 Aug 2018 and 13 Aug 2019) was Nusyn-Noxfish® and contained 2.5 % rotenone active ingredient. The theoretical rotenone concentration of Hidden Lake once it penetrated the thermocline by pumping was 30 ppb and 25 ppb in 2018 and 2019 respectively (Parks Canada 2020).
For crustacean zooplankton, we assessed the effects of rotenone on the density of three dominant species: the cyclopoid Acanthocyclops vernalis, the calanoid Leptodiaptomus tyrelli, and the cladoceran Daphnia pulicaria. Long-term patterns were examined with historical data on the density of the three dominant crustacean zooplankton species in Hidden Lake collected in early August from 2014-2018 by the Fisher lab, and these were compared with post-rotenone treatment samples from early August 2019 (11 months after the first rotenone application) and early August 2020 (11 months after the second rotenone application). A second dataset was collected to understand short-term impacts of rotenone on the developmental stage of the three focal crustacean zooplankton species, which would affect differences in the potential for reproduction in relation to the timing of rotenone treatment. For the second dataset, crustacean zooplankton were sampled in Hidden Lake in mid-July 2018 (5 weeks before the first rotenone application), early September 2018 (3 weeks after the first rotenone application) and early August 2019 (11 months after the first rotenone application).
Field and laboratory: For the long-term zooplankton dataset from the Fisher lab, crustacean zooplankton were collected each year (2014-2020) during an index period between the 4 and 11 of August using full water column tows at the deepest point of the lake with a 30 cm diameter, 243 µm mesh plankton net. Samples were preserved immediately with 95% ethanol and later enumerated under a dissecting microscope. We generated subsamples using a Folsam plankton splitter and enumerated at least 200 individuals per sample. Samples with less than 200 individuals were counted in their entirety. Taxonomic keys used included Brooks (1957), Wilson (1957), Smith and Fernando (1978), Thorp and Covich (2010), and Haney et al. (2013). Since members of the Daphnia pulex species complex cannot be reliably distinguished using morphological characteristics, we verified the identification of D. pulicaria using sequences from two mitochondrial DNA loci, cytochrome oxidase I (COI) and ND5, following the protocol of Miner et al. (2013) (J. Fisher, unpublished data).
For the second zooplankton dataset from the Derry lab, crustacean zooplankton were collected by whole water column vertical tows with a 35 cm diameter, 54 µm mesh Wisconsin net from 0.5 m off the bottom to the surface. Ideally the same net should have been used for both zooplankton datasets, but the long-term dataset (2014-2020) was part of another project. However, our findings for rotenone impacts on the density of sexually mature individuals of the three focal species of crustacean zooplankton were the same for both types of net. For the second zooplankton dataset, the zooplankton were sampled from four sampling stations along an open-water transect across each lake, anaesthetized with bromoseltzer, and then preserved with 95% ethanol. These four samples subsequently were pooled into a single sample per lake for enumeration of developmental stages in the focal species, using taxonomic keys described above. The maturity of copepods was determined by the presence of eggs, or by using the fifth leg shape, as described by Wilson (1957) and Smith and Fernando (1978). The maturity of daphniids was determined by the presence of eggs or embryos, or by the examination of the abdominal processes according to Brooks (1957). Individuals were considered mature adults if the first process was longer than the second. Maturity rate was calculated as the ratio of mature individuals divided by the total density of each species (excluding nauplii) and represents the proportion of sexually-mature adult individuals in the population.
References:
Brooks JL. 1957. The systematics of North American Daphnia. Mem Conn Acad Arts Sci. 13:1-180.
Haney JF, et al. 2013. An-image-based key to the zooplankton of north america version 5.0. University of New Hampshire. Center for Freshwater Biology [cited 11 May 2020] Available from http://cfb.unh.edu/cfbkey/html/
Miner BE, Knapp RA, Colbourne JK, Pfrender ME. 2013. Evolutionary history of alpine and subalpine Daphnia in western North America. Freshw Biol. 58(7):1512-1522.
[MFWP] Montana Fish, Wildlife and Parks: Fisheries Division. 2017. Montana Rotenone Policy. Approved by: Eileen Ryce, Fisheries Division Administrator. Issued April 18, 1996. Revised April 5, 2017.
Parks Canada. 2020. Summary report for the chemical removal of brook trout from Hidden Lake, Upper Corral Creek and Tributaries. Radium Hot-Springs (BC): Internal report
Smith KE, Fernando CH. 1978. A guide to the freshwater calanoid and cyclopoid Copepoda. Crustacea of Ontario. University of Waterloo Biology Series. 18:1-76
Thorp JH, Covich AP. 2010. Ecology and classification of north american freshwater invertebrates. 3rd ed. London (UK): Elsevier Academic Press.
Wilson MS. 1957. Calanoida, In: W.T. Edmondson (ed.). Fresh-Water Biology. 2nd ed. New York (NY). John Wiley & Sons. p. 738-794.
