Leonard I. Wassenaar

Global application of stable hydrogen and oxygen isotopes to wildlife forensics.

Stable isotopes are being increasingly used in wildlife forensics as means of determining the origin and movement of animals. The heavy isotope content of precipitated water and snow (δDp, δ18Op) varies widely and systematically across the globe, providing a label that is incorporated through diet into animal tissue. As a result, these isotopes are potentially ideal tracers of geographic origin. The hydrogen and oxygen isotope tracer method has excellent potential where (1) spatial variation of precipitation isotopes exist, and (2) strong, mechanistic relationships link precipitation and isotope ratios in biological tissue. Here, we present a method for interpolation of precipitation isotope values and use it to create global basemaps of growing-season (GS) and mean annual (MA) δDp and δ18Op. The use of these maps for forensic application is demonstrated using previously published isotope data for bird feathers (δDf) in North America and Europe. The precipitation maps show that the greatest potential for applying hydrogen and oxygen isotope forensics exists in mid- to high-latitude continental regions, where strong spatial isotope gradients exist. We demonstrate that δDf/δDp relationships have significant predictive power both in North America and Europe, and show how zones of confidence for the assignment of origin can be described using these predictive relationships. Our analysis focuses on wildlife forensics, but the maps and approaches presented here will be equally applicable to criminal forensic studies involving biological materials. These maps are available in GIS format at http://www.waterisotopes.org.

Linking breeding and wintering grounds of neotropical migrant songbirds using stable hydrogen isotopic analysis of feathers

Abstract Recent studies have shown that stable hydrogen isotope ratios (δD) in the tissues of animals often correlate with δD of local precipitation. Here we examined the relationship between δD in feathers and growing season precipitation for neotropical migrant songbirds breeding over a continent-wide isotopic gradient. δD values were determined on feathers of 140 individuals of 6 species of wild insectivorous forest songbirds (Setophaga ruticilla, Empidonax minimus, Vermivora peregrinus, Catharus ustulatus, Seiurus aurocapillus, Hylocichla mustelina) taken from 14 breeding locations across North America. The δD of feathers was strongly correlated with the δD of growing season precipitation at breeding sites across North America. As feather hydrogen is metabolically inert after growth, this relationship was then used to assess the breeding origins of wintering migrants. Deuterium values of feathers from 64 individuals representing 5 species of migrants (Helmitheros vermivorus, Wilsonia citrina, Hylocichla mustelina, Dumetella carolinensis, Seirus aurocapillus) at a wintering site in Guatemala were consistent with those predicted from the known breeding ranges of these species. This study demonstrates hydrogen isotopes may become a powerful tool for linking breeding and wintering grounds of neotropical migrant songbirds, as well as other migratory species moving between isotopically distinct regions.

Evaluation of the origin and fate of nitrate in the Abbotsford Aquifer using the isotopes of15N and18O in NO3

Extensive NO3− contamination of groundwater in the Abbotsford aquifer to levels above drinking water limits is a major problem in the Fraser Lowlands of southwestern British Columbia, Canada. Nitrate concentrations in the aquifer ranged from 0 to 151 mg/l NO3−, with a median concentration of 46 mg/l NO3−. Of 117 wells sampled, 54% had NO3− concentrations exceeding the drinking water limit of 45 mg/1. Approximately 80% of the study area had groundwater NO3− concentrations exceeding 40 mg/1 NO3−. Potential NO3− source materials were poultry manure N and synthetic NH4 based fertilizers. Theδ15N of solid poultry manure samples ranged between + 7.9 and + 8.6‰ (AIR). Four brands of synthetic fertilizers commonly used hadδ15N values between −1.5 and −0.6‰. Ammonia volatilization caused theδ15N of groundwater NO3− produced from poultry manure N to range between +8 and +16‰. Theδ18O values of groundwater NO3−, by contrast, mostly ranged between +2 and +5‰ (SMOW). This narrow range ofδ18O values fell within the expected range of NO3− produced by nitrification of reduced N forms such as poultry manure N and NH4 fertilizers, and had a similar range ofδ18O values as NO3− in the upper part of the unsaturated zone below raspberry fields and beneath former manure piles. Theδ15N-NO3− andδ18O-NO3− data confirmed that NO3− in the aquifer was predominantly derived from poultry manure and to a lesser extent from synthetic fertilizers. Theδ18O-NO3− data further suggested the nitrification process occurred mainly in the summer months, with the soil NO3− produced subsequently flushed into the aquifer during fall recharge. Theδ15N-NO3−andδ18O-NO3− data conclusively indicated that no significant bacterial denitrification is taking place in the Abbotsford aquifer.

Individual specialization and trophic adaptability of northern pike (Esox lucius): an isotope and dietary analysis

Abstract Northern pike (Esox lucius) are often considered to be specialist piscivores, but under some circumstances will continue to eat invertebrates as adults. To examine effects of fish assemblage composition on the trophic ecology of pike, we combined stable isotope analysis (SIA) of carbon and nitrogen and stomach content analysis (SCA) on pike from five lakes in northern Alberta, three of which contain only pike (“pike-only”) and two that also contain yellow perch (Perca flavescens) or white sucker (Catostomus commersoni) (“pike-other”). Fish were more important as prey and empty stomachs, which often characterize piscivores, were significantly more frequent in pike-other than in pike-only lakes. However, even though invertebrates were more important for pike in pike-only lakes, SIA and SCA indicated that invertebrates were also an important component of pike diets in pike-other lakes. SIA and SCA also revealed considerable intrapopulation variation in trophic ecology, with individuals in some populations differing by as much as two trophic levels. Comparisons of stomach contents and isotope signatures of the same fish suggested that within these variable populations, specialization on invertebrates or fish was a long-term trait of some individuals. SIA indicated that trophic position increased and diets shifted to a greater importance of littoral prey as pike grew in pike-only lakes, but not in lakes with other fish present. Trophic adaptability in northern pike is expressed at both the population level, where the trophic ecology is sensitive to differences in prey regimes, and at the organismal level, in the form of intrapopulation variation and individual specialization.

Using stable hydrogen and oxygen isotope measurements of feathers to infer geographical origins of migrating European birds

Successful application of stable-hydrogen isotope measurements (δDf) of feathers to track origins of migratory birds and other wildlife requires a fundamental understanding of the correlation between δDf and deuterium patterns in rainfall (δDp) over continental scales. A strong correlation between δDp and δDf has been confirmed for birds and insects in North America, but not yet for other continents. Here, we compare δDf data from resident European birds to new δDp basemaps for Europe. Three maps, representing growing-season and mean annual δDp estimates from an elevation-explicit, detrended interpolation model and growing-season δDp estimates from simple Kriging, all indicate that strong isotope gradients occur across Europe with a general depletion occurring in a northeast direction. The feather data, representing 141 individuals of 25 avian species from 38 sites, ranged from −131 to −38‰. Regression analysis showed that strong correlations existed between both mean annual and growing-season δDp estimated by detrended interpolation and δDf of non-aquatic and non-corvid birds (r2=0.66 and 0.65, respectively). We also examined mean annual and growing-season δ18Op vs. δ18Of for our samples. Both oxygen regressions were similar (r2=0.56 and 0.57, respectively) but poorer than for deuterium. Our study reveals that δD measurements of feathers from migratory birds in Europe may be used to track their origin and movements, and so provide a powerful investigative tool for avian migration research in Europe.

Stable isotopes (δD and δ13C) are geographic indicators of natal origins of monarch butterflies in eastern North America

Abstract Wing membranes of laboratory and field-reared monarch butterflies (Danaus plexippus) were analyzed for their stable-hydrogen (δD) and carbon (δ13C) isotope ratios to determine whether this technique could be used to identify their natal origins. We hypothesized that the hydrogen isotopic composition of monarch butterfly wing keratin would reflect the hydrogen isotope patterns of rainfall in areas of natal origin where wings were formed. Monarchs were reared in the laboratory on milkweed plants (Asclepias sp.) grown with water of known deuterium content, and, with the assistance of volunteers, on native milkweeds throughout eastern North America. The results show that the stable hydrogen isotopic composition of monarch butterflies is highly correlated with the isotopic composition of the milkweed host plants, which in turn corresponds closely with the long-term geographic patterns of deuterium in rainfall. Stable-carbon isotope values in milkweed host plants were similarly correlated with those values in monarch butterflies and showed a general pattern of enrichment along a southwest to northeast gradient bisecting the Great Lakes. These findings indicate that natal origins of migratory and wintering monarchs in Mexico can be inferred from the combined δD and δ13C isotopic signatures in their wings. This relationship establishes that analysis of hydrogen and carbon isotopes can be used to answer questions concerning the biology of migratory monarch butterflies and provides a new approach to tracking similar migratory movements of other organisms.

STABLE-CARBON AND HYDROGEN ISOTOPE RATIOS REVEAL BREEDING ORIGINS OF RED-WINGED BLACKBIRDS

We investigated the potential of using stable-carbon ( 13 C) and hydrogen (D) isotope ratios in feathers of Red-winged Blackbirds (Agelaius phoeniceus) to delineate geographic origins where feathers were grown. We used outer primary feathers from ter- ritorial male blackbirds along a broad latitudinal transect from Louisiana, USA to Sas- katchewan, Canada. Feather D values showed an expected depletion with latitude, and were correlated strongly (r 2 0.83) with the mean growing season D for precipitation at collection sites. Feather 13 C values revealed that blackbirds fed on both C3 and C4 based foodwebs. Inputs of C4-based carbon likely related to consumption of agricultural crops such as corn and sorghum. Overall, our dual isotope approach showed great potential as a means of delineating breeding origins of blackbirds throughout the continental Midwest, and supports previous findings that D measurements of feathers are strongly correlated with precipitation and latitude throughout the central region of the continent.

Dissolved organic carbon and methane in a regional confined aquifer, southern Ontario, Canada: Carbon isotope evidence for associated subsurface sources

Information regarding the origin, composition and transport of natural dissolved organic carbon (DOC) in groundwater is necessary to understand the transport of metals and organic pollutants, as well as for the use of14C in DOC as an isotopic groundwater dating method. Previous research in several groundwater systems has suggested soil organic C is the predominant source of high molecular weight DOC to the subsurface. Through the use of stable isotopes,14C and geochemical analyses, this study shows that significant concentrations of DOC and CH4 in a regional confined aquifer can be generated in situ from subsurface sedimentary organic sources. The DOC and CH4 produced is a combined result of degradation of buried peats and bacterial action, resulting in high DOC concentrations and strongly methanogenic conditions in the aquifer. The DOC and CH4 comprise, on average, nearly 50% of the total dissolved C pool in the central part of the aquifer. Methanogenic conditions complicate isotopic groundwater dating by the conventional dissolved inorganic carbon (DIC) method. Estimates of isotopic groundwater residence time using DOC14C data are proposed by the application of14C isotope and mass balance corrections.

Linking hydrogen (δ2H) isotopes in feathers and precipitation: sources of variance and consequences for assignment to isoscapes.

Background Tracking small migrant organisms worldwide has been hampered by technological and recovery limitations and sampling bias inherent in exogenous markers. Naturally occurring stable isotopes of H (δ2H) in feathers provide an alternative intrinsic marker of animal origin due to the predictable spatial linkage to underlying hydrologically driven flow of H isotopes into foodwebs. This approach can assess the likelihood that a migrant animal originated from a given location(s) within a continent but requires a robust algorithm linking H isotopes in tissues of interest to an appropriate hydrological isotopic spatio-temporal pattern, such as weighted-annual rainfall. However, a number of factors contribute to or alter expected isotopic patterns in animals. We present results of an extensive investigation into taxonomic and environmental factors influencing feather δ 2H patterns across North America. Principal Findings Stable isotope data were measured from 544 feathers from 40 species and 140 known locations. For δ 2H, the most parsimonious model explaining 83% of the isotopic variance was found with amount-weighted growing-season precipitation δ 2H, foraging substrate and migratory strategy. Conclusions/Significance This extensive H isotopic analysis of known-origin feathers of songbirds in North America and elsewhere reconfirmed the strong coupling between tissue δ 2H and global hydrologic δ 2H patterns, and accounting for variance associated with foraging substrate and migratory strategy, can be used in conservation and research for the purpose of assigning birds and other species to their approximate origin.

A method for investigating population declines of migratory birds using stable isotopes: origins of harvested lesser scaup in North America.

Background Elucidating geographic locations from where migratory birds are recruited into adult breeding populations is a fundamental but largely elusive goal in conservation biology. This is especially true for species that breed in remote northern areas where field-based demographic assessments are logistically challenging. Methodology/Findings Here we used hydrogen isotopes (δD) to determine natal origins of migrating hatch-year lesser scaup (Aythya affinis) harvested by hunters in the United States from all North American flyways during the hunting seasons of 1999–2000 (n = 412) and 2000–2001 (n = 455). We combined geospatial, observational, and analytical data sources, including known scaup breeding range, δD values of feathers from juveniles at natal sites, models of δD for growing-season precipitation, and scaup band-recovery data to generate probabilistic natal origin landscapes for individual scaup. We then used Monte Carlo integration to model assignment uncertainty from among individual δD variance estimates from birds of known molt origin and also from band-return data summarized at the flyway level. We compared the distribution of scaup natal origin with the distribution of breeding population counts obtained from systematic long-term surveys. Conclusions/Significance Our analysis revealed that the proportion of young scaup produced in the northern (above 60°N) versus the southern boreal and Prairie-Parkland region was inversely related to the proportions of breeding adults using these regions, suggesting that despite having a higher relative abundance of breeding adults, the northern boreal region was less productive for scaup recruitment into the harvest than more southern biomes. Our approach for evaluating population declines of migratory birds (particularly game birds) synthesizes all available distributional data and exploits the advantages of intrinsic isotopic markers that link individuals to geography.