Isotopic ratios in the feathers of birds hold evidence of their comings and goings, and may explain some of the mysteries of migration.

At the end of Hitchcock’s classic suspense thriller The Birds, a badly gashed Tippi Hedren is helped away from a white house on the shore of northern California’s Bodega Bay by a taciturn Rod Taylor. Every surface, every wall, every telegraph wire is covered by a menacing sea of birds, continually shifting but eerily silent. It is one of the most powerful images of 20th century cinema. 

The central MacGuffin of the film (Hitchcock’s own term for the dramatic question that drives a story) is: Why are these birds acting in such a hostile fashion? Are they seeking revenge for our mistreatment of nature, are they a presentiment of some apocalyptic Doomsday, or are they sent by God as a punishment for the evil of humankind? 

If Hitchcock were alive today he would perhaps be glad to know that if we still don’t know why the birds in the movie were attacking humankind we now at least have a method of determining where they came from. One of the greatest puzzles in ecology - the mystery of migration - is in the process of being solved. 

Migration, the regular seasonal movement of individuals to and from breeding locations, is a fundamental part of the life cycle of many species and its most conspicuous practitioners are birds. For decades migratory biology has attracted the attention of ecologists but attempts to track migrating individuals - by banding or radio-telemetry, for example - have almost always resulted in failure. 

Consider the problems: only a small subset of a very large population is ever tagged and the individual elements that are tagged typically weigh less than 50g. Is it any surprise that the recovery rate of tagged individuals is only between one-in-1000 and one-in-100,000? But in recent years all of that has changed through the use of isotopes - particularly the stable isotopes - as natural abundance tracers. The technique relies on the fact that the relative abundance of several species of stable isotopes in the tissues of animals is intimately linked to geographic location. 

The use of natural abundance isotopes as migration tracers has been stimulated partly by fears over the decline in certain species. For example, in the US, southern breeding populations of the black-throated blue warbler (Dendroica caerulescens) have been declining and the worry is that this may be due to overwintering habitat loss. 

In a recent issue of Science a group based at Dartmouth College in New Hampshire, Stanford University and the Smithsonian Institution, have managed to track the movements of populations of the black-throated blue warbler using the abundances of isotopes of hydrogen and carbon in feathers. The two stable isotopes of hydrogen (H and D) are strongly latitude dependent because the heavier D isotope tends to condense out first and fall as precipitation. 

Since water vapour enters the atmosphere in the low latitudes through evaporation, and the D-rich water condenses out first, the net result is a gradient of decreasing D abundance with increasing latitude. When this is expressed as a ratio of D to H we find that isotope ratios decline with increasing latitude (Table 1). This gross pattern can be locally modified by seasonal changes, distance from the sea, and the altitude dependence of precipitation (rain tends to fall as warm, moist air is forced over mountain ranges). 

Table 1. Isotropic species in migration and trophic studies

Isotope  Convention  Standard* Fractionation  
Hydrogen/Deuterium δ SMOW  Precipitation/condensation
Carbon-12/Carbon-13 δ13 C PDB Proportion of C3/C4 plants
Nitrogen-15/Nitrogen-14 δ15 N Air Marine or terrestrial plants + degree of aridity
Strontium-87/Strontium-86 δ87 Sr NBS-987 Source rock type

*SMOW = Standard Mean Ocean Water; PDB = Pee Dee Belemnite

In the continental US the overall effect of these interactions is that the hydrogen isotope ratios tend to decrease in a north-westerly direction. Since local vegetation reflects this unique isotopic signature, all higher elements in the food chain, including the migratory species under study do so too. 

C3 plants are an ancient group of carbon-fixing green plants. They use rubisco (the most abundant enzyme on Earth) to make a three-carbon compound which is the initial, stable product of photosynthesis. However, such plants may subsequently lose up to half of this recently fixed carbon through photorespiration at night. But since the onset of the global decline in atmospheric CO2 that started at the end of the Cretaceous period (65 million years ago) another type of photosynthetic pathway has evolved, the C4 pathway. 

C4 plants possess biochemical and anatomical mechanisms that serve to concentrate CO2 at the site of fixation, on the chloroplasts, which greatly reduces carbon loss through respiration. Their biochemistry is based on the enzyme PEP-carboxylase that makes an initial four-carbon compound. This is then transferred to specialised cells where the CO2 is re-released and refixed using rubisco. These different biochemistries of C3 and C4 plants result in very different stable isotope signatures. 

Stable isotope ratios are conventionally expressed in parts per thousand (per mil or o/oo) difference from an agreed standard. C3 plants tend to cluster around an average δ 13 C of -25o/oo, while C4 plants have a much heavier isotopic composition, with an average δ 13 C of about -12o/oo. Once again there are local complications but these are minor. Since C3 plants predominate at high latitudes and C4 plants are more common in the low latitudes there is a gradient of decreasing δ 13 C with increasing latitude. 

The Dartmouth study used these isotopic gradients to identify in detail where birds from breeding populations of the black-throated blue warbler spent their breeding season and the winter. By analysing feather keratin (which preserves the isotopic ratios at the time of moult) they found that birds overwintering on the Caribbean islands from Cuba in the west to Puerto Rico in the east came from two latitudinally distinct breeding populations on the US mainland. 

Those overwintering in the west of the Greater Antilles (Cuba and Jamaica) came predominantly from northern populations ranging from Lake Michigan to as far north as New Brunswick, Canada, while those overwintering to the east of the range (the islands of Hispaniola and Puerto Rico) came predominantly from the southern part of the warbler’s breeding range in Georgia and Virginia. Interestingly, although there is latitudinal segregation within these overwintering populations there was no longitudinal segregation in the northern populations overwintering on Cuba and Jamaica. (The southern population came from too restricted a longitudinal range to test for segregation.) 

The study hinted at an explanation for why the southern breeding populations of the black-throated blue warbler are declining in abundance. These are the birds that overwinter in the eastern portion of the Greater Antilles and it may be significant that the island of Hispaniola has undergone massive deforestation in the past few years with consequent habitat destruction. 

This approach is not restricted to migratory species that live in the US. A similar study has used nitrogen isotopes together with carbon isotopes to show that two subspecies of the Scandinavian Willow Warbler (Phylloscopus trochilus trochilus and Phylloscopus trochilus acredula), which occupy distinct latitudinal ranges in Sweden, also occupy different overwintering grounds when they migrate to Africa. 

Nitrogen isotopes, like carbon isotopes vary geographically, although here the control is more complicated. The largest differences are between marine and terrestrial plants. Marine plants have more positive δ 15 N values than terrestrial plants and these differences are reflected further up the food chain, for example in migrating song birds. But there also appears to be an effect that is rather enigmatically linked to aridity. 

With increasing dryness δ 15 N ratios tend to increase (spanning a range from <10o/oo in regions with >800mm of rainfall a year to >15o/oo in areas withThe ratio of strontium-87 to Sr-86 can also be used as a migratory tracer (although note that Sr-87 can be formed radiogenically by the decay of Rb-87 so it is not a stable isotope in the sense of, for example, C-13). In an earlier study of the black-throated blue warbler the Dartmouth team found that breeding populations could be separated according to the δ 87 Sr ratio preserved in their bones. Birds breeding near the core of the Appalachian Mountains in Georgia and Virginia, had high isotopic ratios of around 9o/oo, while those that lived further north near the Great Lakes region and southern Canada had lower values of around 4o/oo

This contrast is because of the higher proportion of geologically ancient, igneous, rocks in the Appalachians relative to the low δ 87 Sr sedimentary rocks of the Great Lakes and southern Canadian regions. It seems that even the basic geology of a region has an isotopic spoor that can reflected in the animals that live there. 

So we can see then that to use stable isotopes as tracers of migratory animals various conditions must be satisfied. In the case of annually migrating animals the tissue to be analysed must be renewed seasonally otherwise the analysis will be an average of isotopic ratios incorporated into the animal throughout its lifetime. Feathers of course are ideal. 

Secondly, the isotope ratios must be an accurate reflection of their local levels. In practice carbon, nitrogen and strontium tend to be true reflections because they do not exchange with the atmosphere after incorporation with feather keratin. Hydrogen isotopes tend to be more exchangeable. Thirdly, the geographic variability of the isotope system in question needs to have been carefully mapped if small scale differences in regional origin are to be discerned. 

The use of Sr isotopes as migratory tracers, however, is more limited than the other isotope systems so far discussed for several reasons. First, geographic Sr variability has not been as well mapped as other isotope systems; secondly its areal distribution is not as straightforward as, for example, hydrogen or carbon; and thirdly, the technique demands the sacrifice of the animal in order that its bone tissue may be sampled, which rather defeats the underlying emphasis on conservation. 

Birds are not the only animal group that is being studied using natural abundance isotopes. In some cases isotope tracking can help preserve even severely endangered species. Today isotopes are offering undreamt of insights into the ecology of several animal species. These tools will undoubtedly be refined in the future and offer the hope that we can at last understand the complex web of feeding and migratory relationships among animals and use this information to help in their conservation. 

The final scene envisaged by Hitchcock for the end of The Birds was cut for budgetary reasons. It was to be a composite matte of the Golden Gate Bridge covered in a seething mass of birds preparing to assault San Francisco. Their place of origin - the ’where’ of the movie might now be solved through the use of isotopes - but the master of suspense can rest easy; his ’why’ remains as enigmatic as ever. 

Source: Chemistry in Britain

Richard Corfield is an isotope geochemist and science writer in the department of earth sciences, Oxford University, where he directs the stable isotope laboratory.

1. Tracking wildlife

African elephants have long been at risk from ivory poachers and one way that this trade can be discouraged is by tracing where a piece of ivory originated. Studies using carbon, nitrogen, strontium and radiogenic lead isotopes in South African elephant populations have shown promise particularly when the data are combined using multivariate statistics to generate ’composite variables’. 

Using this approach a team based in South Africa has shown that a combination of carbon and nitrogen data, and lead and strontium data, was sufficient to distinguish where elephant bone - and ivory - had originated. The technique is still in its infancy however and more recent studies hint that there may be much more geographic variability in isotope concentrations than originally thought. Once again though, use of a sufficiently large number of isotopic species - when combined statistically - may be the way forward. 

Currently a project is under way at the University of Cape Town to map the distribution of isotopes of strontium, lead and neodymium throughout sub-Saharan Africa so that an isotopic database can be produced with which poached ivory can be compared. In this way regions prone to poaching can be identified and steps taken to eradicate the threat. 

However, stable isotopes vary not only as a consequence of geographic region, they often vary as a result of food source as well. Scientists working for the US Geological Survey are using ratios of carbon, nitrogen and sulphur isotopes to investigate whether a change in the basic diet of the US Grizzly bear in the area around Yellowstone National Park will have any long term effects on bear populations. 

In recent years the native cut-throat trout of the lakes in Yellowstone National Park in Wyoming have been under threat from an invasion of lake trout. By calibrating the stable isotope composition of Grizzly bear food sources and then tracking the changing composition in bear hair, scientists are monitoring whether the bears are able to change their diet without any damage to their population structure. 

The diet of Arctic foxes has also been investigated using the ratio of carbon isotopes. Arctic foxes usually feed on lemmings which have the distinctive carbon isotope of a terrestrial herbivore that feeds predominantly on C3 plants (approximately -25o/oo). Research has shown that following winters where the lemming population has been low, Arctic fox hair has the significantly more positive carbon isotope signature indicative of a marine diet. 

It seems that in low-lemming abundance winters Arctic foxes follow polar bears out onto the ice and feed on seal carrion or pups. A concern must be that as global warming results in a shorter sea-ice season Arctic fox populations may become more vulnerable to times of low lemming abundance.  

2. Measuring up

Most stable isotope measurements (C, N, O, S, H) are routinely made on a magnetic sector mass spectrometer. There are two variants: ’dual-inlet’ and ’continuous-flow’ machines. 

In dual-inlet MS, the sample gas (liberated by digestion in acid or by combustion) is frozen in a low-volume cold-trap where it is then sublimated back to gas. The pressure of this sample gas is then balanced against that of a reference gas of known isotopic composition before multiple measurements of each are made. This allows very accurate measurement of the differences in the isotopic ratios of the sample and reference gases. 

In continuous-flow MS, the gas is carried in a helium carrier-flow to the analyser, where the isotopic abundance is measured directly from the height of the peak in the mass spectrum. (Until recently, this technique was considerably less accurate than the dual inlet approach.) 

Subsequently, in both cases, the isotopic ratios (of D to H, or C-13 to C-12, for example), are then corrected for machine error and normalised to an internationally agreed standard (eg PDB or SMOW). The final δ values are not absolute isotope abundances, therefore, but differences between samples and one or other of several natural abundance standards. For example, if a sample is found to have a C-13/C-12 ratio greater than the standard PDB’s by 10 parts per thousand, this value is reported as δ C = +10o/oo. 

Strontium isotope measurements are made on a thermal ionisation magnetic sector mass spectrometer. Samples are not introduced into the machine from an on-line device but rather are laboriously evaporated onto rhenium filaments before measurement in the machine. Although attempts have been made to express strontium isotope ratios as delta values (δ 87Sr) it is still more common to see them expressed as an absolute ratio of Sr-87 to Sr-86, normally, for terrestrial rocks, in the region of 0.47. 

Before using stable isotopes as tracers of migratory animals, three conditions must be satisfied. 

1. The tissue to be analysed must be renewed seasonally otherwise the analysis will be an average of isotopic ratios incorporated into the animal throughout its lifetime. Feathers are ideal because the primary (most complete) moult for most temperate song birds occurs at the end of the breeding season and before migration. 

2. The isotope ratios must be an accurate reflection of their local levels. In practice carbon, nitrogen and strontium tend to be true reflections because they do not exchange with the atmosphere after incorporation with the feather keratin. Hydrogen isotopes tend to be more exchangeable but this does not appear to affect their value as a geographic tool. 

3. The geographic variability of the isotopes in question needs to be mapped if small-scale differences in regional origin are to be discerned in migrating populations.