2.3 | Stable isotope analyses
Workers were placed in an oven at 60˚C until dry (approx. 24-48 hours)
and then stored in airtight vials prior to processing. The abdomens of
all ants were removed prior to weighing to avoid the effects of stomach
contents on isotopic signatures (Tillberg et al., 2006). To achieve
appropriate weights for each sample, five to ten workers per sample were
pooled and chopped in glass vials to fine homogeneous powders using
small scissors. Approximately 0.400 mg of each sample was weighed into
tin capsules (Costech Analytical Technologies Inc., Valencia, CA, USA)
using a microbalance (Mettler Toledo, Columbus, OH, USA). All samples
were analyzed at the Texas A&M University Stable Isotopes for Biosphere
Science Laboratory (https://sibs.tamu.edu/) using a Delta V Advantage
Isotope Ratio Mass Spectrometer coupled with a Costech Elemental
Analyzer and Thermo ConFlo IV Universal Interface (Thermo Fisher
Scientific, Waltham, MA, USA). All baseline samples (collected before
nests were fed the tracer) were analyzed before any post-feeding samples
to ensure that natural abundance values were not influenced by memory
effects from the high levels of 15N in spiked samples.
Nitrogen isotope ratios are presented in \(\delta\) notation:
δ15N (‰) = [(Rsample –
Rstandard)/Rstandard] x
103
where Rsample is the15N/14N ratio of the sample and
Rstandard is the15N/14N ratio of the atmospheric N
standard (Mariotti, 1983; Coplen, 2011). Precision was 0.1‰.
To verify that our isotope tracer methods could detect resource sharing
over 30-m distances, we conducted a proof-of-concept experiment using a
population of tawny crazy ants (Nylanderia fulva ) near College
Station, Texas. This species forms a single supercolony throughout its
invaded range in North America, in which workers regularly share
collected resources with each other and occupy transitory nests (Eyer et
al., 2018; Wang et al., 2016). Our results from this experiment
confirmed that the isotope tracer is highly successful at detecting
sharing in unicolonial populations at distances that were relevant for
our study (i.e., up to 28.4m from the treated area; Kjeldgaard 2020)
using the same biomass of workers for isotope analysis.