Evaluating environmental DNA detection of a rare fish in turbid water using field and experimental approaches.

Detection sensitivity of aquatic species using environmental DNA (eDNA) generally decreases in turbid water but is poorly characterized. In this study, eDNA detection targeted delta smelt (Hypomesus transpacificus), a critically endangered estuarine fish associated with turbid water. eDNA sampling in the field was first paired with a trawl survey. Species-specific detection using a Taqman qPCR assay showed concordance between the methods, but a weak eDNA signal. Informed by the results of field sampling, an experiment was designed to assess how turbidity and filtration methods influence detection of a rare target. Water from non-turbid (5 NTU) and turbid (50 NTU) estuarine sites was spiked with small volumes (0.5 and 1 mL) of water from a delta smelt tank to generate low eDNA concentrations. Samples were filtered using four filter types: cartridge filters (pore size 0.45 µm) and 47 mm filters (glass fiber, pore size 1.6 µm and polycarbonate, pore sizes 5 and 10 µm). Prefiltration was also tested as an addition to the filtration protocol for turbid water samples. eDNA copy numbers were analyzed using a censored data method for qPCR data. The assay limits and lack of PCR inhibition indicated an optimized assay. Glass fiber filters yielded the highest detection rates and eDNA copies in non-turbid and turbid water. Prefiltration improved detection in turbid water only when used with cartridge and polycarbonate filters. Statistical analysis identified turbidity as a significant effect on detection probability and eDNA copies detected; filter type and an interaction between filter type and prefilter were significant effects on eDNA copies detected, suggesting that particulate-filter interactions can affect detection sensitivity. Pilot experiments and transparent criteria for positive detection could improve eDNA surveys of rare species in turbid environments.

Molecular Systematics of Redband Trout from Genome-Wide DNA Sequencing Substantiates the Description of a New Taxon (Salmonidae: Oncorhynchus mykiss calisulat) from the McCloud River.

Rainbow Trout, Oncorhynchus mykiss, exhibit high levels of phenotypic diversity leading to the recognition of numerous subspecies. A major distinction among Rainbow Trout subspecies exists between Coastal Rainbow Trout (O. m. irideus), which occurs west of the Cascade and Sierra Nevada mountain ranges, and interior Redband Trout (O. mykiss sspp.), largely distributed to the east. Interior Redband Trout are composed of three primary lineages and can share various outward, anatomical or physiological characteristics that are often symplesiomorphies or examples of convergence. We examine high-throughput DNA sequence data from Sacramento Redband Trout O. m. stonei from the Upper Pit and Upper McCloud Rivers along with representatives of Rainbow Trout and Golden Trout lineages to clarify the composition and relationships of the Sacramento Redband Trout. We find O. m. stonei to be polyphyletic, divided between populations in the Pit River and the Upper McCloud River. Redband Trout obtained from the Pit River are most-closely related to Great Basin Redband Trout O. m. newberrii and to fish of the Warner Lakes Basin and Surprise Valley within the O. m. newberrii lineage. The type specimen of O. m. stonei, collected from the Lower McCloud River, is phenotypically similar to Great Basin Redband Trout. We find as well that the isolated populations of trout in the Upper McCloud River Basin represent a lineage of Rainbow Trout now restricted to that region, are monophyletic and are not most closely related to the interior Redband Trout lineages of O. m. newberrii or O. m. gairdnerii. Furthermore, they are not represented by the type specimens of O. m. stonei or O. m. shasta. Consequently, we formally describe the McCloud River Redband Trout O. mykiss calisulat, new subspecies.

UrsaPlex: An STR multiplex for forensic identification of North American black bear (Ursus americanus).

UrsaPlex is a forensic quality 5-dye multiplexed tetranucleotide STR and sex identification panel for individual genetic identification of North American black bears (Ursus americanus). The panel is validated for the identification of black bears involved in human-wildlife conflict events and poaching investigations. This is the first single multiplex panel composed solely of tetranucleotide STRs derived from black bear and bear-specific sex markers. UrsaPlex produces complete genetic profiles from as little as 78 pg of DNA template and has a probability of identity of 2.63 × 10-13. The panel has also been tested for utility in other ursids, and our results indicate with minor modifications, UrsaPlex should prove valuable in identification investigations involving these species as well.

Landscape genetics of California mule deer (Odocoileus hemionus): the roles of ecological and historical factors in generating differentiation.

Landscape genetics is an emerging discipline that utilizes environmental and historical data to understand geographic patterns of genetic diversity. Niche modelling has added a new dimension to such efforts by allowing species-environmental associations to be projected into the past so that hypotheses about historical vicariance can be generated and tested independently with genetic data. However, previous approaches have primarily utilized DNA sequence data to test inferences about historical isolation and may have missed very recent episodes of environmentally mediated divergence. We type 15 microsatellite loci in California mule deer and identify five genetic groupings through a Structure analysis that are also well predicted by environmental data. We project the niches of these five deer ecotypes to the last glacial maximum (LGM) and show they overlap to a much greater extent than today, suggesting that vicariance associated with the LGM cannot explain the present-day genetic patterns. Further, we analyse mitochondrial DNA (mtDNA) sequence trees to search for evidence of historical vicariance and find only two well-supported clades. A coalescence-based analysis of mtDNA data shows that the genetic divergence of the mule deer genetic clusters in California is recent and appears to be mediated by ecological factors. The importance of environmental factors in explaining the genetic diversity of California mule deer is unexpected given that they are highly mobile species and have a broad habitat distribution. Geographic differences in the timing of reproduction and peak vegetation as well as habitat choice reflecting natal origin may explain the persistence of genetic subdivision.

Can fatty acid and mineral compositions of sturgeon eggs distinguish between farm-raised versus wild white (Acipenser transmontanus) sturgeon origins in California? Preliminary report.

The objective was to investigate the potential of using fatty acid and mineral compositions of sturgeon eggs to distinguish their source, either farm-raised or wild fish. Trafficking of illegally obtained wild white sturgeon eggs is a major concern to the California Department of Fish and Game, but there is no forensic method to separate wild and farm-raised white sturgeon eggs. The extension of these findings in future work will be to use the fatty acid and mineral compositions as forensic indicators of caviar produced legally from farm raised sturgeon compared with illegal caviar produced from sturgeon poached from the wild. Samples (10) of sturgeon eggs were collected from a commercial aquaculture facility in the Sacramento Valley. Eggs from wild sturgeon (9) were obtained primarily from confiscations of illegally caught sturgeon by fish and game law enforcement personnel. The total lipid content of sturgeon eggs was analyzed for fatty acid composition. The most notable difference was the higher concentration (P<0.001) of C18:2n6 in farm raised eggs (6.5 mg/100g total lipid) than wild eggs (0.6 mg/100g total lipid) while other differences between fatty acids were smaller. Eicosapentaenoic acid (C20:5n3) was higher (P<0.02) in farm-raised (5.56 mg/100g) than wild (4.49 mg/100g). Docosahexaenoic acid (C22:6n3), C18:1 cis 9&10, and C20:4n6 were not different for origin of the eggs. Concentration of selenium was markedly higher (P<0.001) in eggs from wild sturgeon (10.0 mg/kg dry weight) than farm-raised sturgeon (2.7 mg/kg dry weight). Concentrations of iron, zinc, copper, phosphorus, sulfur, calcium, and potassium did not differ between farm-raised and wild eggs. Arsenic concentration in wild eggs was 3.3mg/kg dry weight whereas arsenic was not detected in the farm-raised eggs. Fatty acid and mineral compositions of eggs differed significantly between farm-raised and wild sturgeon and these should be investigated further as biological markers for forensic identification of caviar origin.

Inheritance of microsatellite loci in the white sturgeon (Acipenser transmontanus).

Nine tetramer motif (GATA)n microsatellite systems were developed for use in the white sturgeon, Acipenser transmontanus. We report inheritance patterns for these nine systems, which range from one possible disomic system to tetrasomy and octosomy, with some systems containing null alleles. Because of the complex modes of inheritance underlying these systems and the highly duplicated nature of the genome, we propose each allele be scored as its own dominant marker, similar to AFLPs or RAPDs. The utility of this method is validated by the observation that individual alleles within a microsatellite system generally fit the expectation for independent transmission and fit the expected transmission frequency for single copy nuclear markers.