The most commonly used direct assessment is using doubly labeled water
(Speakman 1997, 2000). The method depends on the fact that isotopes of oxygen
in body water are in complete equilibrium with oxygen in respiratory carbon
dioxide. An isotopic label of oxygen introduced into body water will be eliminated
as water and as carbon dioxide. In contrast, an isotopic label of hydrogen will be
eliminated only as water. If both isotopes are introduced as water (^2 H 218 O—
hence doubly labeled water) at the same time the relative difference in their elim-
ination will reflect production of carbon dioxide. In practice, a bird is injected
with doubly labeled water, a blood sample is taken at about the time that the
doubly labeled water has equilibrated with unlabeled body water (30–60 min)
and a further sample(s) taken later. The isotopic ratios in the samples are measured
by mass spectrometry.
In addition to metabolism, the nutritional status of a bird is important. This can
be assessed from body mass and size, and from fat score (see Chapter 4). Change in
mass or condition can be assessed only by re-capturing the birds and repeating the
measurements. Measuring metabolites of lipids in a single blood sample may prove
to be a useful indicator of change in body mass (Williams et al. 1999; Guglielmo
et al. 2002). Similarly, measurement of yolk precursors in the blood may be useful
to assess the reproductive status of female birds (Vanderkist et al. 2000).
9.6 Molecular genetics
Recent advances in molecular biology have resulted in methodologies that can be
used to determine the sex of individual birds, paternity and kinship, geographical
structuring within species, phylogenetic relationships among species and the
timing of speciation events. See Brown (2001) for further information on DNA
technologies.
DNA can be obtained from a wide range of sources. From dead birds, any tissue
can be used. From living birds, blood is the most practical, and this can be
obtained as a blood sample following venepuncture, or from the base of a plucked
feather. An advantage of birds over mammals is that their red blood cells are nucle-
ated and so only a small sample is required. DNA can also be obtained directly
from shed feathers (Eguchi and Eguchi 2000; Bello et al. 2001). DNA is extracted
from the tissue sample using protein-denaturing agents, salt and solvents.
Identifying individuals and relatives. In a major breakthrough that led to the
process of DNA fingerprinting or profiling, Jefferies et al. (1985a) discovered
that specific nucleotide sequences occurred in a repeated order, called tandem
repeats, with high levels of variation meaning that individuals differ in their
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