Mutualistic symbioses are widespread and of crucial importance in many ecosystems
]. Although evolutionary theory to explain the stability of mutualistic interactions has progressed considerably (see
] for a review), consensus on the general underlying mechanisms that keep these interactions stable and cooperative has not been achieved
]. While further theoretical work might alleviate this problem, these difficulties also illustrate that mutualistic interactions are highly variable in their ecological contexts
] and degrees of commitment
], and that very few of them have been studied in considerable depth (reviewed in
]). Two aspects are thought to have important implications for the interaction stability of host-symbiont mutualisms:
1. The level of sexual reproduction and the degree of independent dispersal of the symbionts, and
2. Genetic diversity among symbionts of a single host
]. In a previous study we investigated the first aspect in the hitherto poorly studied mutualism of Lasius flavus
ants farming root-aphids
]. The present study focuses on the second aspect.
In cultivation (farming) mutualisms, the host partner promotes the growth of a symbiont that it consumes, either individually or as somatic modules
]. While scenarios of ‘enslavement domestication’ have been suggested for the early evolution of such mutualisms
], it remains difficult to understand how symbionts would be actively selected to make the transition from free-living to being domesticated. The latter state would imply becoming reproductively isolated from free-living relatives which would require consistent direct benefits to be sustainable. Domestication often also implies losing options for horizontal transmission, having many offspring consumed by the host, and potentially being mixed with other symbiont lineages, consequences that could all discourage life as a symbiont. Domestication mutualisms would thus seem most likely to evolve if symbiont services ultimately benefit the reproduction of close symbiont relatives and if the productivity of domesticated reproduction consistently exceeds the fitness that can be obtained from a free-living life-style. When symbionts are already clonal before domestication, one would therefore expect symbioses to elaborate this form of propagation when making symbionts commit irreversibly to a dependent life-style, which requires new host-serving adaptations that impede survival and reproduction without the host. The ‘trophobiotic organs’ evolved in the aphids of our present study
] are examples of such adaptations.
While symbiont interests in being cultivated would be expected to benefit from monopolizing host attention to a group of close relatives, hosts should not necessarily favor the same tendencies towards rearing monocultures, as a more variable community of symbionts might offer a broader spectrum of services or be less vulnerable to parasites (e.g.
]). As outlined by in earlier studies
], hosts would be selected to enforce monocultures only if scramble competition between multiple symbiont strains would decrease the overall productivity of the symbiotic interaction, i.e. if different symbiont strains would compete for the same limited resource provided by the host. Similar selection pressure towards monoculture farming would apply if coexistence of multiple strains within the same host would allow free-riding by underperforming strains, leading to a direct reduction in overall productivity (e.g.
Incentives for competition or cheating would destabilize mutualistic interactions between symbionts and hosts, unless specific mechanisms of symbiont screening upon admission
] or symbiont rewarding/sanctioning in proportion to performance
] can evolve. The relative importance of these mechanisms is controversial, but available data suggest that monocultures are commonly found in the cultivation mutualisms that have been studied, from the gardens of algae-growing damselfish
] to those of fungus-growing termites and ants
]. In fungus-farming leaf-cutting ants, monocultures appear to be enforced by a combination of incompatibility between genetically different symbiont strains and active symbiont policing by the hosts
], whereas a simple mechanism of positive frequency-dependent propagation within established colonies appears sufficient to enforce life-time commitment between a termite host colony and a single symbiont clone
]. However, more studies are needed to establish the generality of this principle, particularly for cultivation mutualisms where hosts are able to segregate symbionts in space or time to avoid competition
], so that the benefits of polyculture might surpass the costs.
In the present study we focus on a farming symbiosis that has been known for decades but has rarely been studied: the root aphid husbandry for sugar (honeydew, “milk”) and nitrogen (“meat”) of the Yellow meadow ant Lasius flavus
, which is likely to be essential for ant colony growth and reproduction, and involves an entire array of root aphid species
]. These root aphid species have a number of distinct traits that improve performance as ant symbionts but are never found in free-living aphids, such as the ‘trophobiotic organ’ to hold honeydew for the ants
]. The most common species have further lost most if not all sexual reproduction in Northwest Europe, but have maintained low frequencies of winged morphs that may disperse between colonies
]. In the present study we use a newly developed set of DNA microsatellite markers
] to assess aphid species number and clonal diversity at the level of single ant nest mounds.
The objectives of our study were to use hierarchical sampling (Figure
) and DNA microsatellite analysis to: 1. Estimate species- and clone diversity for three focal species of root aphids (Geoica utricularia, Tetraneura ulmi, Forda marginata) within L. flavus nests, soil samples within nests, and single aphid chambers (Figure
a) within these soil samples, 2. Evaluate whether the observed distributions are consistent with the expectation that symbiont diversity within nests is low, 3. Analyze the extent to which diversity patterns change across sampling levels and years, and 4. Infer which potential mechanisms can lead to the observed diversity patterns.
Figure 1 The sampling scheme for root aphids in nest mounds of the ant Lasius flavus. a. A representative large aphid chamber with many, mostly adult, Geoica utricularia, b. Aphids were sampled from ant mounds on the island of Schiermonnikoog (The Netherlands) (more ...)