Materials and methods
Prey items were obtained from RodentPro in April 2023 and stored frozen
until analysis. Smaller prey including smaller sized rats, pup Guinea
pigs and all sizes of mice were sold in bulk in packs of 10-25, five
individuals of each species group were processed. Quail were sold in a
pack of five and all five were processed and larger prey (medium rabbits
and extra large rats) were sold individually, and we ordered three
individuals of each. Each individual was weighed, and then the hair was
removed using a
Remington
Ultimate Precision Detail Trimmer Black NE3160, and feathers were
plucked manually. The individual was reweighed to determine contribution
of keratin outer covering to total organism weight. The trimmer was
cleaned between individuals using water and 70% isopropyl alcohol. The
individual was then homogenized using a
OXO
Good Grips Meat Tenderizer and VEVOR Hand Operated 304 Stainless Steel
Multifunction Manual Meat Mincer (pup rats and large mice were too small
for the meat grinder and were only homogenized with the meat
tenderizer). Between individuals, the meat grinder was fully
disassembled and each component plus the meat tenderizer were cleaned
with soap and water, then sanitized in 95% EtOH. A subsample of hair or
feathers was taken from each individual to analyze separately, and the
remainder of this material was blended back into remaining homogenized
mass with the meat grinder. All samples were dried at 60°C for at least
24 hours, then ground into powder using a mortar and pestle or finely
diced with a scalpel blade.
We had three tissue types for analysis: 1) homogenized mass of whole
body with keratin included (WB, three replicates per individual); 2)
homogenized mass of whole body with keratin removed (KR, three
replicates per individual), and 3) keratin tissue only (K, one replicate
per individual). Isotopic values from the three replicates of WB and KR
were averaged, and the mean value was used in statistical analysis for
each tissue type.
Samples weighing between 0.400 and 0.600mg were placed into 5mm tin
capsules and analyzed at the Stable Isotope Mass Spec Lab at the
University of Florida on one of two systems. Samples were combusted on
N.C. Technologies 8020 Elemental Analyzer or a Costech 4010 Elemental
Analyzer at 1000°C interfaced to a ConFlo IV before passing to a Thermo
Electron Delta V Advantage isotope ratio mass spectrometer. Sample gas
was measured relative to laboratory reference N2 and
CO2 gases. All are expressed in standard delta notation
relative to Vienna Pee Dee Belemnite (carbon) or AIR (nitrogen).
Analytical precision was calibrated by the standard deviation of the
USGS40 replicates (n = 30, 0.08 ‰ [δ 13C]
and 0.07 ‰ [δ 15N]).
Some mixing model scenarios have demonstrated that lipid-correction can
substantially alter the estimated diet composition and interpretation of
diet reconstruction, and lipid extraction may not be appropriate when
using the isotopic composition of keratinous tissues to examine
diet21,22. Therefore, samples in this study were not
lipid extracted to best represent the isotopic values of the whole prey
item.