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The five most common species of Pacific salmon, Rainbow Trout (steelhead) Oncorhynchus spp., and Atlantic Salmon Salmo salar intermingle in the North Pacific Ocean and its freshwater tributaries. Efficient morphological methods for distinguishing among these species are sometimes limited by condition of the specimen (degraded or missing morphology), life history stage, or training of the observer. Researchers have successfully applied various genetic methods to distinguish among these species when morphological analyses are not possible, but they cannot easily incorporate these methods into standard fish and wildlife population monitoring analysis workflows. Here we test five 5′–3′ exonuclease (TaqMan) assays developed from mitochondrial genes and provide novel methods that take advantage of TaqMan output to distinguish among these species. We found that combinations of as few as two of the five assays were adequate to distinguish all species. TaqMan chemistry is designed to interrogate a single nucleotide locus. We also explore the basis for the variation in the observed scatter plot distributions (variation in florescent signals) and show that this variation is due to nucleotide diversity in and near the probe site. Because the SNPs underlying the assays developed here are all physically close to one another along the mitochondrial genome, the potential exists to develop a single DNA sequence-based assay to discriminate among salmon species. This single assay can be added to a genotyping-by-sequencing panel to identify and exclude nontarget species from analyses.

Genotypic sex identification assays can provide valuable information about fish populations when phenotypic sex determination is difficult. Here we describe the development of a TaqMan® assay (Ots_SexID) designed to identify the genotypic sex by targeting a region previously examined in the growth hormone pseudogene for winter-run Chinook salmon (Oncorhynchus tshawytscha) collected from the Sacramento River and spawned at the Livingston Stone National Fish Hatchery. Accuracy of the marker was assessed by comparing genotypic sex assignments for Chinook salmon spawned at Livingston Stone National Fish hatchery in 2012 (n = 84) with phenotypic sex recorded during spawning. Genotypic sex was observed to be concordant with phenotypic sex identified using Ots_SexID in 83/84 individuals, suggesting that the assay could be used to predict phenotypic sex with ∼︀99% accuracy. To evaluate the utility of the TaqMan assay in other parts of the species’ range, we examined collections from 29 other populations ranging from Alaska to California. Genotypic sex assignments based on the assay were generally concordant with observed phenotypes, but there were some strong exceptions. These results suggest that the new assay will be very useful for Sacramento River winter-run Chinook salmon, but also highlight the importance of thoroughly testing any genotypic sex identification assay before application in a population of interest.

Most coho salmon Oncorhynchus kisutch in Washington state spawn at 3 y of age, creating the potential for three temporal populations or “broodlines” at each spawning site. This is generally prevented by a portion of males in each site that mature and reproduce at 2 y of age, resulting in population structure in which the geographic component is stronger than the temporal component. The Quilcene National Fish Hatchery, located on Big Quilcene River in the Hood Canal region of Washington state, selected against late returning coho salmon by excluding all but the earliest returning fish from its broodstock for an unknown number of generations, and restricted gene flow among broodlines by excluding 2-y-old males for 27 generations. The resulting hatchery population exhibited three distinct broodlines that returned in alternating years: an “early” broodline that arrived 1 mo before the wild fish, a “late” broodline that arrived at the same time as the wild fish, and a “middle” broodline that arrived in between these two broodlines. We evaluated temporal and geographic components of population genetic structure in coho salmon from the Quilcene National Fish Hatchery and nine other sites from Puget Sound and the Strait of Juan de Fuca using 10 microsatellite loci. Genetic diversity at the Quilcene National Fish Hatchery was lowest in the early broodline and highest in the late broodline. Divergence among broodlines at the Quilcene National Fish Hatchery was greater than that observed at any other site, and was also greater than that observed between any of the sites. This apparent reversal of the relative magnitudes of temporal and geographic components for this species emphasizes the importance of variable age-at-maturity in shaping population genetic structure.

The Modoc sucker Catostomus microps received legal protection in the United States based partially on concerns that anthropogenic environmental changes had restricted migration among populations and catalyzed hybridization with a more abundant congener, the Sacramento sucker Catostomus occidentalis. We applied eight microsatellite markers to samples of both species collected from two tributaries to the Pit River, California (Ash Creek and Turner Creek), and one tributary of Goose Lake, Oregon (Thomas Creek). Modoc sucker populations in these three tributaries seemed to be largely isolated from one another: gene flow between Ash Creek and Turner Creek was no greater than that among these two creeks and Thomas Creek. In contrast, divergence estimates among collections of Sacramento suckers indicated greater gene flow between Ash Creek and Turner Creek than between either of these creeks and Thomas Creek. Samples collected at a single site (Ash Valley) were identified based on morphology as Modoc suckers, but genetic data suggested they were much more similar to Sacramento suckers. Interspecific hybrids were detected in all three tributaries. Collections of Modoc suckers yielded 0.0–3.9% hybrids, and collections of Sacramento suckers yielded 0.0–80.0% hybrids. The two collections with the greatest proportions of hybrids (54.5 and 80.0%) were both from tributaries to lower Thomas Creek, and neither of these tributaries is thought to have upstream populations of Modoc suckers. Based on 1) low levels of hybrid detection in all three tributaries, 2) the absence of hybrids from typical parental habitats (upstream habitats for Modoc suckers and Pit River mainstem for Sacramento suckers), and 3) highly significant RST (variance in allele size) values between the species, we conclude that hybridization is common but that significant introgression (i.e., loss of parental genotypes) has not occurred. We also note that hybridization, and subsequent introgression, may become a conservation concern in such cases when the habitat of one or both of these species is eliminated or modified.