2.1 | Between-nest interactions
To quantify sharing between nests in the field, we treated selected
nests in each site with a stable isotope tracer and quantified its
movement into neighboring nests (Fig 1). Stable isotope tracers employ
naturally occurring, non-radioactive forms of biologically relevant
elements, such as nitrogen. The heavier isotope of nitrogen
(15N) occurs rarely in nature, so by artificially
‘spiking’ a food with an appropriate concentration of this heavy
isotope, we can trace the movement of this isotope through consumers and
identify the flow of nutrients through an ecosystem (Fry, 2006). We
ensured that only the treated nests had access to the isotope tracer, so
if a neighboring untreated nest showed unnaturally high levels of15N, this would indicate an exchange of either workers
or resources between the treated and untreated nests (i.e., no
boundaries between nests). In a preliminary laboratory experiment, we
detected highly elevated levels of 15N as quickly as
24 hours after feeding fire ant workers an isotope tracer, and these
isotope values decreased steadily over a 72-hour period (Kjeldgaard
2020). These preliminary results indicate that the tracer must have been
relatively recently consumed to be detected in workers, which would
allow for an effective measure of the movement of marked food.
Sampling was conducted between August and October 2019 in six field
sites in Texas, USA (Appendix S1). Habitats ranged from restored
grasslands (sites O, A, and B) to mowed fields (sites C, T, and T2).
Mounds were used as a proxy for individual nests. At this stage of the
experiment, we could not determine if distinct mounds/ nests belonged to
the same polydomous colony. After searching each field site and
identifying all active fire ant nests, we selected three clusters of
four to five nests at each site. One nest within each cluster was
selected as the treatment nest. Clusters were separated by at least 50 m
within each site to avoid potential sharing between clusters (Fig 1).
Mounds were selected to represent varying distances within each cluster
(between 0.4m-29.07m from the treated nest, with an average distance of
7.65m ± 0.72m) to determine any effect of distance on sharing between
nests.
Similar to other studies (Goodisman et al., 2007), several fire ant
nests disappeared or moved over the course of the sampling period. As a
consequence, we were unable to find three nests (one nest from site T,
one from O, and one from T2) after the treatment period. Each of these
were untreated nests within different clusters, so their removal did not
affect the number of clusters analyzed in each location. In total, we
sampled from 73 fire ant nests across six sites, with 12 nests in Site
A, 13 in Site B, 12 in site C, 11 in Site O, 13 in Site T, and 12 in
Site T2.