In our study, we evaluated the role of the facultative endosymbiont of aphids R. insecticola on intra and interclonal competitive interactions of aphids. Our experiments revealed that, regardless the presence of R. insecticola, in the cases of single colonies the clone G1 exhibited a higher PGR than clone G2. However, in competitive interclonal coexistence, G2 outperformed clone G1. These results suggest that aphid clones have differential features that allow more efficient performance as coexisting or solitary colonies, but not in both contexts. Surprisingly, clone G2 exhibited a higher PGR when coexisting with G1 than when developing alone or intraclonal coexistence. Variation in competitive abilities among aphid clones has been observed among clones for other aphid species such as the pea aphid, Acyrthosiphon pisum (Hazell et al. 2005), but not including the effect of facultative endosymbionts. Similar results were also observed between a mutant colour (yellow) and its original (green) aphid clone which regulated its reproductive rate depending on the relative density of self and non-self-clones, but in this study only one aphid genotype was studied (Li and Akimoto 2021), which involved only intraclonal competition. In this study, authors have suggested that aphid clones exposed to clonal coexistence and single rearing could alter their reproduction by discriminating between closely related and unrelated clones in mixed colonies. In Chile, S. avenae aphid shows strong signatures of obligated parthenogenesis and aphid clones are frequently observe over several seasons (Figueroa et al. 2005). Interestingly, the clone G1 and G2 of S. avenae used in our study were found to be common in wheat and oat plantations of centre-south of Chile (G1 and G4 respectively in Zepeda-Paulo et al. (2021). In the field, the clone G1 has been observed as the dominant clone at the beginning of the season compared to the other clones (including G2), but this decreased from the mid-season, on the contrary the clone G2 increased its prevalence throughout the season being dominant at the end of the season (Zepeda-Paulo et al. 2021). In light of our results, temporal dynamic and dominance of aphid clones throughout the growing season could be explained in part by a differential response of aphid clones, by example when they are exposed to lesser competitive (e.g., early season) and highly competitive scenarios (i.e., growing season), which would contribute to maintaining clonal variation within natural populations. The greater competitive ability of the clone G2 can be explained by a rapid occupation of the resource, developing a niche pre-emption strategy. This result supports Silvertown`s hypothesis (2004) for niche pre-emption limiting the diversification of late-arriving lineages. Evidence supporting this is the higher number of individuals of clone G2 than G1 at day 15 developing with or without harbouring R. insecticola, with clone G2 reaching an asymptote at a stage when clone G1 growing alone was initiating an exponential type of growth (Fig. 2).
Consistent with our data, a similar study found that one clone of aphid species Myzus persicae outgrew another clone only under coexistence (Turcotte et al. 2011a, b). Thus, aphid clones appear interact when mixed, leading to an asymmetrical competitive interaction, which in turn can drive eco-evolutionary feedback loops. However, in that work Turcotte et al. (2011a, b) did not explore the mechanism underpinning clonal interaction. In our study, unlike our initial prediction, we can state that the asymmetric interaction between clone G1 and G2 appear not to be related with the presence of endosymbionts. Although competitive interactions between aphid clones was not significantly influenced by endosymbiont-infection, we have found that individual of S. avenae aphids harbouring the facultative endosymbiont R. insecticola exhibited a greater PGR than endosymbiont-free aphids. Furthermore, the positive effect on PGR that R. insecticola inflicts on the different aphid clones studied here agrees with the fact that positive selection of aphid clones harbouring R. insecticola in S. avenae natural populations (Zepeda-Paulo et al. 2021). Since high Regiella-infection levels were reached (87%-93%) over the course of a season on different cereal plants (wheat and oat) (Zepeda-Paulo et al. 2021), which shed some light on how facultative endosymbionts could drive different eco-evolutionary pathways of aphid clones at field level.
For instance, the fact the clone G1 harbouring endosymbiont (+ G1) showed a higher fresh weight growing as solitary clone than -G1 and also than + G2 and -G2, suggest that R. insecticola positively affects the traits of this clone. This positive effect could be associated with an adaptation to host plants, similar to when the reproduction of infected pea aphids on clover is enhanced (Tsuchida et al. 2004), as a defensive effect of Regiella in the aphid S. avenae has already been ruled out (Zepeda-Paulo et al., 2017)
Another interesting result is the effect of endosymbionts on the proportion of winged morphs in aphid clones exposed to intraclonal interactions. Herein we found that harbouring R. insecticola affects the proportion of winged individuals in a clone-dependent manner. While R. insecticola resulted in a lower final proportion of winged individuals in clone G1, this response changed when aphids were exposed to interclonal interactions, where clone G2 produced a higher proportion of winged morphs in response to clonal competence. Previous studies have confirmed that main factors inducing winged individuals in aphids are overcrowding and the presence of predation risk (Mehrparvar et al. 2013) and parasitism (Braendle et al. 2006). There is also genetic variation between clones in the winged morphs (Braendle et al., 2005). Mixed effects on the production of winged individuals due to the presence of R. insecticola have been described in the aphid pea, with evidence that support positive (Leonardo and Mondor 2006) and negative (Reyes et al. 2019), differences that have been attributed in part to the Regiella genotype (Reyes et al. 2019). A reduced production of winged individuals in aphids harbouring R. insecticola in a temperature-dependent manner was also described in S. avenea (Liu et al. 2019). Here we add that variation in winged production in aphids harbouring facultative endosymbionts also appears to depend on coexistence with other clones.
We also inquired about the impact of the study clones, either with or without harbouring R. insecticola, on the relative change of root/shoot ratio in the host plant as a measured of plant stress induced by aphids. We observed that aphid infestation significantly increased the root/shoot ratio in wheat plants compared aphid-free plants. Feeding by aphids can affect plant growth and structure, mainly by reducing root tissue density, likely by a carbohydrate depletion in plants due to translocation from roots to shoots (Smith and Schowalter 2001). Interestingly, our results showed differences in the impact of aphids on the root/shoot ratio between aphid clones, indicating different levels of damage produced by the aphid clones. For instance, clone G1 showed a greater damaging capacity on host plants compared to clone G2, which could indicate for the aphid genotypes studied that clonal competition could be beneficial for plants as clone G2 outperformed clone G1 in competitive interaction. Although R. insecticola appears to do not induce significant changes on the root/shoot ratio, aphids harbouring R. insecticola showed a lesser damaging capacity on the host plants than endosymbiont-free aphids, which was true for both aphid clones studied. A similar tendency to show a lower root/shoot ratio in plants infected with aphids that harbour facultative endosymbionts was reported in the pea aphid-Hamiltonella defensa-broad bean system (Serteyn et al. 2020). Further studies are needed to understand how variation in the aphid genotype x endosymbiont interaction in different ecological contexts (e.g., intra and interclonal competition) affect growth, photosynthesis, and reproduction of host plants. .