Introduction
Invasive ants often reach extremely high densities, outcompete and prey upon native species, disrupt mutualisms, and lower ecosystem biodiversity (Berman, Andersen, Hély, & Gaucherel, 2013; Holway, 1998; Holway, Lach, Suarez, Tsutsui, & Case, 2002; LeBrun, Abbott, & Gilbert, 2013; McGlynn, 1999; Porter & Savignano, 1990). Understanding the factors that promote the success of invasive ants is critical to defining and managing their ecological impacts. One factor thought to play a role in invasive ant success is their proclivity for intraspecific cooperation (Holway et al., 2002; Passera, 1994). Many invasive ant species are unicolonial in their introduced range, meaning that populations are characterized by an absence of behavioral boundaries among nests (Helanterä, Strassmann, Carrillo, & Queller, 2009; Holway et al., 2002; Krushelnycky, Holway, & LeBrun, 2010). By cooperating instead of competing with conspecifics, unicolonial ants avoid the costs of intraspecific competition, which is thought to allow them to reach higher densities (Giraud, Pedersen, & Keller, 2002; Porter, Fowler, & Mackay, 1992) and achieve greater ecological dominance by more effectively outcompeting other species (Holway et al., 2002; Holway & Suarez, 2004; LeBrun et al., 2013). For example, the number of Argentine ant workers (Linepithema humile ) was approximately 50-fold higher in sites where nests cooperated compared with sites where nests competed (Holway & Suarez, 2004). Consequently, cooperative Argentine ants more effectively outcompeted native ants, as native ant species richness was reduced by over 50% when Argentine ant nests cooperated compared with when they competed with each other (Holway & Suarez, 2004). The positive correlation between instraspecific cooperation and ecological impact has also been implicated in other social insects (Hanna et al., 2014; Korb & Foster, 2010; Perdereau et al., 2015, Wilson, Mullen, & Holway, 2009).
However, unicoloniality does not explain the success of all invasive ant species (Holway et al., 2002; Passera, 1994). The dark rover ant (Brachymyrmex patagonicus ), for example, has invaded large portions of the southern USA (MacGown, Hill, & Deyrup, 2007), but introduced populations of this species are multicolonial, with distinct nests usually headed by a single egg-laying queen (monogyne; Eyer, Espinoza, Blumenfeld, & Vargo, 2020). Moreover, despite the ecological benefits, when many unrelated queens cooperate and produce workers within a single colony, relatedness between nestmates can decrease to effectively zero (Crozier & Pamilo, 1996; Keller, 1995; Queller & Strassmann, 1998). As a consequence, polygyny and especially unicoloniality increase the likelihood of workers interacting with and caring for less related individuals, which in turn reduces their inclusive fitness (Helanterä et al., 2009; Schmid-Hempel, 1997; Schmid-Hempel & Crozier, 1999). Inclusive fitness may be maintained, however, if workers discern and preferentially direct care towards kin over non-kin (Hamilton, 1964; Keller, 1995). For example, although Argentine ant supercolonies are characterized by cooperative, interconnected nests (Giraud et al., 2002; Tsutsui, Suarez, Holway, & Case, 2000), workers spend more time investigating (i.e., antennating) non-nestmates than nestmates from the same supercolony (Björkman-Chiswell, Van Wilgenburg, Thomas, Swearer, & Elgar, 2008), indicating nestmate recognition despite a lack of aggression within the same supercolony in this species. Nestmate recognition may reduce or eliminate indiscriminate sharing between Argentine ant nests, as sharing was consistently limited to distinct clusters of nests within a single supercolony over a three-year-period (Heller, Ingram, & Gordon, 2008). Kin recognition can also impact sharing between individuals within the nest in what is known as nepotism. For example, in polygyne ant colonies of Formica argentea , workers preferentially care for brood that are more closely related to them (Snyder, 1993). By discriminately sharing food resources with more related individuals from neighboring mounds or from within the same mound, workers may increase their inclusive fitness in polygyne colonies (Hamilton, 1964; Helanterä et al., 2009). Despite the evolutionary benefits of preferential care towards kin over non-kin, it has only rarely been documented in polygyne or unicolonial ant species and may be less common than previously thought (Boomsma & d’Ettorre, 2013; Keller, 1997).
We tested for unicoloniality and within-colony nepotism in polygyne red imported fire ants (Solenopsis invicta ; hereafter fire ants). Fire ants occur in two social forms: the polygyne form (i.e., colonies with multiple egg-laying queens) and the monogyne form (i.e., colonies with only a single egg-laying queen; Gotzek, Shoemaker, & Ross, 2007; Ross, 1993; Ross, Vargo, & Keller, 1996; Tschinkel, 2006). Polygyne fire ant colonies are considered highly cooperative, and they are often referred to as unicolonial throughout their invaded range (e.g., Greenberg, Vinson, & Ellison, 1992; Holway et al., 2002; Morel, Vander Meer, & Lofgren, 1990; Plowes, Dunn, & Gilbert, 2007; Porter et al., 1992; Vander Meer, Obin, & Morel, 1990). Cooperative behavior in polygyne fire ants is thought to correspond with a greater abundance compared with the monogyne form due to reduced intraspecific competition (Porter, Bhatkar, Mulder, Vinson, & Clair, 1991). For example, polygyne mounds were over twice as abundant on average compared with monogyne mounds in Texas (mean ± SE: 680 ± 475 polygyne mounds/ha vs. 295 ± 240 monogyne mounds/ha; Porter et al., 1991). The greater abundance of the polygyne form may increase the likelihood of ants interacting with and preying upon native species, thereby increasing their ecological impact (Allen, Epperson, & Garmestani, 2004; Porter & Savignano, 1990). Despite the assumption that polygyne mounds cooperate and are highly interconnected (see Bhatkar & Vinson, 1987), the physical exchange of workers and resources between polygyne mounds in the field is actually poorly documented. Moreover, although polygyne workers from different nests do not aggressively attack each other outside of the nest (Vander Meer et al., 1990), their interactions within the nest are relatively unknown (but see DeHeer & Ross, 1997), particularly in the case of worker-brood interactions. Within-colony relatedness between polygyne workers is often near zero throughout their invaded range (DeHeer & Ross, 1997; Goodisman, Sankovich, & Kovacs, 2007; Ross, 1993; Ross & Fletcher, 1985; Ross et al., 1996), but workers may increase their inclusive fitness by preferentially caring for more related brood.
Our study tests fundamental assumptions about inter- and intracolonial cooperation in introduced populations of polygyne fire ants. To delimit boundaries between colonies, we quantified the exchange of a15N-glycine tracer dissolved in a sucrose solution and correlated this exchange with colony genetic structure. Using a labeled resource in combination with genetics data allows for two different ways to define colony boundaries (Ellis, Procter, Buckham-Bonnett, & Robinson, 2017). We also quantified within-colony conflict between workers and larvae by comparing brood-tending behaviors towards close siblings (i.e., from the same mother) and non-siblings (i.e., from a different mother). Studies that actually document cooperation between and within colonies are critical to understanding the ecology and success of invasive species.