Ten species of smelts (Osmeridae), including 1 introduced species, have been found in the eastern North Pacific (Love and others 2005). Of these 10 species, 5 have been recorded from the inland marine waters of the Salish Sea, including Puget Sound and the Straits of Juan de Fuca and Georgia (Hart 1973): Whitebait Smelt (Allosmerus elongatus), Surf Smelt (Hypomesus pretiosus), Capelin (Mallotus villosus), Longfin Smelt (Spirinchus thaleichthys), and Eulachon (Thaleichthys pacificus). Results from a study using molecular markers to distinguish smelt remains found in the stomach contents of fishes (Paquin and others 2014) indicated that a specimen collected from Discovery Bay, Washington, an embayment at the eastern terminus of the Strait of Juan de Fuca, was misidentified as S. thaleichthys in the online Barcode of Life Database (BOLD; Ratnasingham and Hebert 2007). Subsequent examination of the preserved specimen has confirmed its identity as Night Smelt (Spirinchus starksi) and constitutes a new record for this species in the Salish Sea. Since this discovery, additional specimens of S. starksi have been collected in the area for inclusion in this study. To confirm the identification of the Discovery Bay specimen, both morphological and nucleotide sequence data from S. starksi were compared with data from the closely related species S. thaleichthys. Whole specimens and tissue samples (frozen or preserved in ethanol) from 11 S. starksi and 5 S. thaleichthys were obtained from the Northwest Fisheries Science Center (NWFSC) and the University of Washington Fish Collection (UW). Specimens of S. starksi were collected from Discovery Bay, Washington (n 5 1), Green Point, Washington (n 5 5), and north of Monterey Bay off the California coast (n 5 5). Specimens of S. thaleichthys were collected from the Gulf of Alaska (n 5 1), San Juan Islands (n 5 1), and Puget Sound (n 5 3) (Table 1). DNA extractions were performed on fin clips using a QIAGEN DNeasy kit (QIAGEN, Valencia, CA) and the manufacturer’s animal tissue protocol. A 750 base-pair region of the mitochondrial genome, cytochrome c oxidase I (COI), was amplified using PCR with universal fish primer cocktail C_FishF1t1-C_FishR1t1 (Ivanova and others 2007). The COI gene region has been used as the barcode for biodiversity to distinguish among species, whether they are distantly related species such as mammals and insects, or closely related congenerics (Ward and others 2005, 2009; Holloway 2006; Ward and others 2009). Nucleotide sequence data were collected for 16 samples (including the misidentified specimen of S. starksi from Discovery Bay) in both forward and reverse directions with Big Dye chemistry using EXOSAP purified PCR products. Sequences were read using an ABI 3730 automated sequencer
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