An aphid pest superclone benefits from a facultative bacterial endosymbiont in a host dependent-manner

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Rubio-Meléndez, and 2 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-4338445/v2 This work is licensed under a CC BY 4.0 License Status: Posted Version 2 posted You are reading this latest preprint version Show more versions Abstract The English grain aphid, Sitobion avenae , is a significant agricultural pest affecting wheat, barley, and oats. In Chile, the most prevalent and persistent clone (superclone) of S. avenae harbours the facultative endosymbiont bacterium Regiella insecticola . To determine the role of this bacteria in the ecological success of this superclone, the presence of R. insecticola was manipulated to evaluate the impact on 1) the reproductive performance of this clone in two host plant species (wheat and barley), 2) the production of winged morphs, 3) changes in the proteomic profile of insects, and 4) root/shoot ratio of plant. It was determined that this superclone of S. avenae proliferates differentially in the host plants studied depending on the presence of the facultative bacterial endosymbiont, observing that the clone develops better in wheat when it is infected with R. insecticola while the opposite occurs when it develops in barley. Aphid biomass was higher when harbouring R. insecticola , particularly in barley. Individuals infected with R. insecticola , in both host plants, showed higher proportion of winged individuals. The protein regulation of aphids on wheat was comparatively lower and stable than that on barley. A higher root/shoot biomass ratio was detected in wheat than in oats in plants attacked with aphids harbouring R. insecticola . R. insecticola significantly affects the reproductive and proteomic performance of the S. avenae superclone, changes influenced by the host plant, suggesting that the host plant x facultative endosymbiont interaction can drive host specialization intraclonally, partly the ecological success of the superclones. Entomology host specialization endosymbionts aphid population growth rate insect proteomic Figures Figure 1 Figure 2 Research Highlights The aphid endosymbiont R. insecticola affect the reproductive performance of S. avenae R. insecticola alter the aphid reproductive performance in a host-dependent manner R. insecticola showed a positive effect on wheat and a negative on barley 1. INTRODUCTION Aphids (Hemiptera: Aphididae) are important crop pests that amplify clonally under favourable conditions. There is an increasing number of studies describing the presence of few widespread multilocus genotypes (“clones” for simplicity) within aphid species, particularly invasive ones with outstanding ability to persist in time and space (called “superclones”) (Vorburger et al. 2003 ; Fenton et al. 2010 ; Harrison and Mondor 2011 ; Piffaretti et al. 2013 ; Chen et al. 2013 ; Nibouche et al. 2014 ; Harris-Shultz et al. 2017 , 2022 ; Zepeda-Paulo et al. 2017 a; Rubio-Meléndez et al. 2019 ). However, few studies have addressed the causes explaining this ability. Hypotheses explaining the success of superclones that have found support, states that they exhibit: 1) greater polyphenism enabling responses to variable environmental factors such as host plant quality (Castañeda et al. 2010a ), 2) obligate asexuality enabling superclones to do not spend energy on finding suitable mates or mating sites (Piffaretti et al. 2013 ), 3) adaptation to their host plants (Fenton et al. 2010 ), 4) the ability to tolerate allelochemicals (Castañeda et al. 2010b ), and 5) flexibility on feeding behaviour allowing a similar reproductive performance across different context (Barrios-SanMartin et al. 2016 a). Surprisingly, the role of facultative bacterial endosymbionts on the ecological success of these invasive superclones has not been studied yet. The grain aphid Sitobion avenae is a worldwide aphid-pest of cereals. While populations in UK, north of France and Romania are predominated by cyclical parthenogenetic asexual lineages and typically featured by a high genetic diversity (Sunnucks et al. 1997 ; Dedryver et al. 2001 ; Papura et al. 2003 ; Llewellyn et al. 2004 ), populations in Australia, Chile and China (the introduced range) are predominated by a dissimilar number of obligated parthenogenetic asexual lineages depending on the region (Wilson et al. 1999 ; Figueroa et al. 2005 ; Xin et al. 2014 ). In Chile, this aphid was most probably introduced in the middle of 1970s from Europe (Apablaza and Fernández 1982 ; Figueroa et al. 2005 ), and it has been shown that nearly 90% of its genotypic variation is accounted for by only four predominant asexual lineages (Figueroa et al. 2005 ; Zepeda-Paulo and Lavandero 2021 ). One of these asexual lineage dominates rapidly early in the season and displays low variance in performances on different hosts (Castañeda et al. 2010b ; Zepeda-Paulo and Lavandero 2021 ). Interestingly, asexual lineages of S. avenae in Chile varied in the presence of the facultative bacterial endosymbionts (Sepúlveda et al. 2014), with the most frequent asexual lineages found to be regularly harbouring Regiella insecticola (Enterobacteriales, Enterobacteriaceae)(Zepeda-Paulo et al. 2017 ). Since R. insecticola has been reported to improve the fitness in a host dependent manner in the pea aphid (Tsuchida et al. 2004 ), it is likely that this bacterium plays a role in the performance of S. avenae , determining ecological success on different host plants. Here we report a study with the most common genotype of the aphid S. avenae in Chile, in which the presence of the predominant facultative bacterial endosymbiont R. insecticola was manipulated to assess the impact of this endosymbiont on the reproductive performance of this aphid on these two host plants (wheat and barley), and the changes that this generates in the whole-body proteomic profile of aphids. The latter can shed light on the physiological mechanisms underlying reproductive performance in each host due to the presence of this facultative endosymbiont. These two hosts were chosen because they are contrasting in terms of the prevalence of S. avenae in Chile (Figueroa et al. 2005 ; Sepúlveda et al. 2017 ), since while it is very recurrent in wheat it is scarce in barley, a situation that allows evaluating, controlling for the genetic background, the impact of harbouring the facultative endosymbiont R. insecticola . Plant responses (root/shoot ratio) were also measured to assess the consequences of endosymbiont-dependent aphid herbivory. 2. MATERIAL AND METHODS 2.1 Aphids and plants Individuals of the most widely distributed genotype in cereal crops in the Maule to De Los Rios regions of Chile of S. avenae were kindly provided by Dr. Francisca Zepeda-Paulo. This genotype was characterized with eight microsatellite loci ( Sm11 , S3.43 , S16b , S30 , S4Σ , S5L , Sm17 and Sm10 ) and corresponds to the genotype G as described by Zepeda-Paulo et al. (2021) found to harbour R. insecticola (see more details in Zepeda-Paulo et al. 2021). In the present study we used this genotype as two asexual lineages: one infected (hereafter E+) composed by individuals naturally harbouring the facultative endosymbiont R. insecticola (as collected from the field), and the other asexual lineage was originated by individuals treated with antibiotics (ampicilin, cefotaxim and gentamicin) by using the method of (i) artificial diet (synthetic diet) and (ii) micro-injections with the methods of Koga et al. (2007) and Simon et al. (2011) resulting in R. insecticola -disinfected individuals (hereafter E-). Both lineages harboured the primary endosymbiont Buchnera aphidicola . Aphid colonies (E + and E-) were reared in separate mesh cages (50 cm x 42 cm x 31 cm) containing pots with wheat or barley seedlings. Thus, four distinct colonies of the genotype G1 were reared in the laboratory. Host plants used were barley ( Hordeum vulgare L., cultivar Sebastian) and wheat ( Triticum aestivum L., cultivar Pantera). Plants were grown in organic soil during one week in a greenhouse by sowing about twenty seeds per pot (bottle cap in pots). Every pot was disinfected (Virginia igenix) and fertilized (Nutrifeed follare) every week. The growth chamber was under controlled conditions (22 ± 1 ºC, 16:8 photoperiod). 2.2 Aphid reproductive performance and the effect on host plants To use age-synchronized aphids for the experiments with E + as E- aphids, firstly 50 adult aphids were placed on potted wheat or barley plants. After 12 h, all adults were removed, leaving only new nymphs on plants. Four days later, four wingless aphids were transferred to each individually potted wheat or barley seedlings. In total, 20 pots with wheat seedlings with E + aphids, 20 pots with wheat seedlings with E- aphids, 20 pots with barley seedlings with E + aphids, and 20 pots with barley with E- aphids were obtained, each selected for setting the experimental systems. After 14 days the final number aphids, including winged individuals was recorded. With the aim of determining relations in aphid weights, 25 wingless adults were collected in each of the four rearing boxes. They were weighted with an analytic balance (XP2U Mettler Toledo, Columbus, OH, USA, precision: 10 − 3 mg). In addition, once the reproductive performance experiment finished, the plants were removed from the pots and carefully separated the root and shoots, dried out for about one week at 60°C and (Memmert Beschickung-Loading Modell 100–800) and weighted with an analytical balance (Radwag as 220/c/2). 2.3 2D-DIGE and protein identifications Aphids hosting or not a facultative symbiont, namely R. insecticola , reared on two host, wheat or barley were collected to perform proteomic analysis. Whole body proteins from around 50 mg fresh S. avenae asexual lineages were extracted, purified, and quantified as described by Francis et al. (2010). Three Cy dyes (GE Healthcare) were used for labelling and protein samples of aphid asexual lineages were labelled either with Cy3 or Cy5 and mixed with an internal reference standard protein mixture (pooled from equal aliquots from all experimental samples) labelled with Cy2. Two replicates from each treatment with one dye (Cy3 or Cy5) and a third replicate with the other of the two Cy dyes were established for a conventional dye swap of DIGE. The first and second-dimensional electrophoresis, the excision of protein spots and the process of protein identification were performed following the description in Francis et al. (2010). The Mascot server 2.2.06 with BioToolsTM3.2 (Bruker Daltonics) and NCBI (National Center for Biotechnology Information) server accessed accessed in November 2022 were used for protein identification. The identified proteins were categorized according to metabolic function using the Kegg pathway database ( http://www.genome.jp/kegg/pathway.html , accessed in November 2022) and Expasy Proteomic tools ( http://www.expasy.org/tools/ , accessed in January 2022). 2.4 Statistical analysis The total number of aphids (i.e., nymphs + adults + adults winged) and adults winged were subjected to glm with Poisson distribution and log-link, whereas the aphid weight and plant root/shoot ratio were subjected glm function with a Gaussian distribution as the error structure. The glm included the factor “endosymbiont” with the levels E + and E-, and the factor “host plant” with the levels wheat and barley. After detecting a significant effect, the Tukey post hoc comparison with Šidák correction procedure was used to make pairwise contrasts between treatments using the emmeans package (Lenth R et al. 2021 ). Wilcoxon rank sum test was used to compare fold ratios of protein regulation. All statistical analyses were carried out with the R software, version 4.2.2 (2022; R Development Core Team). 3. RESULTS 3.1 Effect of the presence endosymbiont on aphids’ performance and plant growth The total number of aphids at the end of 14 days varied depending on the host plant and the presence of the endosymbiont, as revealed by the significant interaction between the endosymbiont and host plant factors (Deviance = 783.9; d.f.= 1, 76; p < 0.001). In wheat, the final number of aphids was higher in the E + aphids, while in barley, the opposite was observed, with a higher number of aphids in the E- aphids (Fig. 1 A). The final number of winged individuals was very low, but was affected only by the endosymbiont factor, with winged individuals only found in the E + aphids in both host plants (Wheat: E + = 0.65 ± 1.04 and E-: 0 ± 0; Barley: E + = 0.30 ± 0.57 and E-: 0.05 ± 0.22; Deviance = 19.3; d.f.= 1, 78; p < 0.001). Aphid final weight exhibited a significant effect of the endosymbiont factor (F = 214.7; d.f.= 1,96; p < 0.001; Fig. 1 B) with E + aphids being weightier than E-. There was also a significant effect to the host plant factor (F = 49.2; d.f.= 1,96; p < 0.001), with aphids being weightier on barley than on wheat. Additionally, there was a significant interaction between the endosymbiont and host factors (F = 5.0; d.f.= 1, 96; p < 0.05), with both E + and E- aphids being weightier on barley than on wheat. On the other hand, the root/shoot ratio also varied depending on the host plant and the presence of the endosymbiont, as revealed by the significant interaction between endosymbiont and host plant factor (F 1, 76 = 7.79; p < 0.001; Fig. 1 C). In wheat, the root/shoot ratio was higher on plant attacked by the E + aphids that by E- aphids, while in barley, no difference in the root/shoot was found (Fig. 1 C). 3.2 Identification of differentially expressed proteins In total, 43 proteins were identified from the 2D gel among which spots varied significantly (p < 0.05; Figure S1). The complete properties of over- and under‐expressed proteins in S. avenae asexual lineages with and without R. insecticola facultative symbiont on wheat or barley host plants were listed in Table 1 . We found that 38 proteins were related to aphids and six6 proteins were related to endosymbionts. The fold change ratios of these proteins between the aphid asexual lineages with or without R. insecticola on the same plant, either wheat or barley ranged from 0.27 to 7.7. The absolute range of protein regulation (considering the extremes of up and downregulation fold-ratios) in barley was 1.43 fold-ratio, while it was 0.47 fold-ratio in wheat. With regards to proteins in the aphid group, 58% were detected in A. pisum while fewer (8%) were detected in other species ( A. citricidus , A. gossypii and S. avenae ). With regards to the endosymbiont group, various proteins were detected in B. aphidicola (63%), the other were related to Regiella and Rickettsia facultative endosymbionts. Proteins with different expression levels were relate to several metabolic pathways (Table 1 ). We found that these proteins accounted for various roles in metabolic pathways such as genetic information processing (20%), cytoskeleton (18%), environmental information processing (16%), energy metabolism (11%), carbohydrates (9%), cellular process and stress responses (both 5%). The presence of R. insecticola in aphid developing in barley showed a greater average upregulation of proteins than in wheat (barley fold-ratio = 2.0 ± 1.4, wheat fold-ratio = 1.27 ± 0.28, p < 0.001, W = 419, Wilcoxon rank sum test), whereas on wheat infected aphids showed a greater average downregulation than on barley (wheat fold-ratio = 0.88 ± 0.17, barley fold-ratio = 0.57 ± 0.29, p < 0.001, W = 75, Wilcoxon rank sum test). On barley energy and carbohydrate metabolism, as well as cellular process and response to stress were highly downregulated. Contrastingly, downregulation in wheat was globally mild, except in the case of other protein group, particularly in the 1-acyl-sn-glycerol-3-phosphate acyltransferase. Upregulation was globally mild on both host plants, with a distinct high upregulation of the protein 1-acyl-sn-glycerol-3-phosphate acyltransferase on E + aphids developing on barley. 4. DISCUSSION In this work we have found that a single genotype of the aphid S. avenae proliferates differentially in two hosts depending on the presence of a facultative bacterial endosymbiont. The most striking result of our study was that among aphids sharing the same genetic background, those developing on wheat and harbouring the facultative endosymbiont R. insecticola exhibit larger colony development, while the opposite was true for the aphids developing in barley. A similar positive effect of this endosymbiont on reproduction has been found on the Vicia-specialized pea aphid populations (Tsuchida et al. 2004 ). However, that result was not reproduced in other pea aphid or other aphid species, with studies showing negative or neutral effects of R. insecticola on aphid reproduction (Ferrari et al. 2004 , 2007 ; Wang et al. 2016 ; Luo et al. 2017 ; Liu et al. 2019 ). Nevertheless, our results are consistent with field-based monitoring of S. avenae in wheat plantations showing R. insecticola increased the prevalence throughout time when compared with uninfected individuals (Zepeda-Paulo and Lavandero 2021 ). It should be noted that G1 genotype dominates early in the season, but a recent study showed that its predominance significantly decreased at mid-season (Zepeda-Paulo and Lavandero 2021 ). However, although G1 decreases its prevalence during the season studied, it persists over the years as describe by Figueroa et al. )2005). Ramírez-Caceres et al. (2019), studying another S. avenae genotype (G2) also described a higher population growth of E + aphids on wheat than on barley, while E- negatively affect the growth of aphids on barley. All this support that R. insecticola entail within-genotype difference on S. avenae performance across two common food plants, wheat and barley. This suggest that host plant x facultative endosymbiont interaction may drive host specialization even within a genotype. Thus, clones composed by E + aphid on wheat and E- aphids on barley, are expected to be positive selected and negatively selected, respectively, leading to ecological divergent populations dominated by asexual clones. However, given the temporal instability of the cereal plantations in Chile, these populations might not reach such a divergence (González U et al. 2013). The lack of host-based differentiation of S. avenae populations in Chile confirm this hypothesis (Figueroa et al. 2005 ). Nevertheless, it is surprising that the same facultative endosymbiont can generate such dissimilar effects on its host aphid, which raises several questions about the underlying mechanisms. Our result show that E + aphids on both host plants showed higher number of winged individuals, although in a very low number, this contrast with previous studies. For example, E + individuals of asexual lineages of S. avenae from China under crowed conditions, produced less winged offspring than E- aphids (Liu et al. 2019 ). This result was found at 25°C, which is slightly higher than our conditions (22 ± 1 ºC). They also found that winged morph production did not showed differences among E + and E- at higher temperatures, which suggested that winged morph production was dependent on environmental temperature and aphid density. On the other hand, R. insecticola negatively affect the production of winged offspring in the pea aphid (Leonardo and Mondor 2006 ). In our case, because the production of winged individuals in E + aphids was irrespective of the host plant, this could be an idiosyncratic capacity of this asexual lineage (G1) to respond to the infection with R. insecticola . Remarkably, this asexual lineage is one the most persistent and predominant in the wheat field of Chile (Figueroa et al. 2005 ), and frequently found harbouring R. insecticola (Zepeda-Paulo et al. 2017 ) and thus the evolution of this lineage with R. insecticola might have reached a fixed pattern of response to this endosymbiont. Surprisingly, the body weight of E + and E- aphids did not fallowed the same trend as reproductive performance. Instead, regardless host plant, aphid showed greater weight when they harboured R. insecticola . This indicates that the production of offspring is uncoupled with the weight of each individual. This contrast with the results found in Rhopalosiphum maidis (Fitch) feeding on barley, where R. insecticola -infected aphids performed poorly in weight (Liu et al. 2023 ). Regarding the distinct effect of R. insecticola on weight and reproductive performance in aphids of S. avenae sharing the same genetical background, again open questions about the undelaying mechanisms of these responses. Thus, we subsequently develop a proteomic study with the infected and non-infected aphids after their development in wheat or barley, to obtain some light on what the underlying mechanisms are. Our findings suggest that the presence of R. insecticola generates different changes in the proteomic profile of S. avenae depending in a host-dependent manner, that could account for the difference in reproductive performance. Interestingly, the fact that protein regulation of aphids developing on wheat was comparatively milder and steadier than on barley, suggest that E + reared on wheat inflict lower impact of their physiology. In this regard, due to expansion of gene families associated with resistance to insecticides and plant chemical defenses described in the genome of S. avenae (Villarroel et al. 2022 ), if R. insecticola would have a beneficial effect on confronting those compounds, a larger regulation of those proteins would be expected when comparing E + and E- aphids. However, no differential regulation of those proteins was found in our study. Interestingly, this is consistent with the lack of association between the presence of R. insecticola and sensitivity to pyrethroids in S. avenae population from Germany (Leybourne et al. 2023 ). Since populations of S. avenae in Chile are most predominant in wheat than on barley (Apablaza and Fernández 1982 ; Figueroa et al. 2005 ) and that the genotype G1 is also predominant (Zepeda-Paulo et al. 2017 ; Zepeda-Paulo and Lavandero 2021 ), the comparative lower protein regulation on wheat suggests that much steady physiological response as compared with that on barley, is probably due to a recent adaptation of S. avenae to wheat after introduction. Indeed, it has been described that S. avenae superclones exhibit a broad host range, flat energetic costs for non-induced detoxification enzymes, and low variation in their reproductive performance on different host plants (Castañeda et al. 2010b ; Barrios-SanMartin et al. 2016 b). The facultative aphid endosymbiont Serratia symbiotica manipulates the expression of specific proteins in the pea aphid impairing plant defence response and improve feeding (Wang et al. 2020 ), a mechanism that may be occurring in E + in S. avenae aphids. The lower reproductive performance of E + aphids on barley could be link to the higher number of proteins that were upregulated in these aphids, which could be an indicative of a reaction to a bacterium infection. The upregulation of the 1-acyl-sn-glycerol-3-phosphate acyltransferase enzyme (plsC), which participates several lipid biosynthetic pathways (Chen et al. 2011 ), and the elongation factor 1-alpha, may both be related with active enzymes delivery as a reaction to R. insecticola . However, the functional role of these upregulated protein on E + aphids remain to be deciphered. On the other hand, downregulation of proteins in E + on barley such as such as murein hydrolase activator EnvC, a protein produced by R. insecticola and normally associated with bacteria proliferation within aphids (Cook et al. 2020 ), could be aphid defensive response against R. insecticola induced by feeding on barley. It remains to be deciphered what are the host-dependent cues triggering such a striking difference in the aphid physiology. Regarding the effects of E + and E- aphids on the plants, the higher root/shoot ratio exhibited by E + aphids compared to E- aphids on wheat plants but not on barley, suggests that these plants experienced a greater response stress. Since plants usually show a higher root/shoot ratio under water deficiency, it seems that E + aphids on wheat were more damaging than on E-. It is likely that aphids harbouring R. insecticola obtain and specific benefit on wheat which increase their efficiency in the use of this specific plant. This result contrasts with what was found in Medicago truncatula , where plants treated with pea aphids free or infected with R. insecticola showed no difference on the dry weight of plant shoots (Pandharikar et al. 2020 ). 5. CONCLUSION In summary, our laboratory study we detected important effects of R. insecticola on S. avenae reproductive performance and proteomic which were highly influenced by the host plant. Since these results were detected within one single aphid genotype, the direct impact of the facultative endosymbionts x host plants interaction it is highlighted. Accordingly, as the presence of facultative endosymbionts can alter aphid reproductive performance in a host-dependent manner, we found that the prevalence of facultative endosymbionts, larger on wheat and lower on barley, is also dependent of the host plants. Declarations AUTHOR CONTRIBUTIONS Claudio C. Ramírez and Frederic Francis conceptualized the research and designed the experiments. Leandro Mahieu and Algélica González-González conducted the aphid performance experiments. María Eugenia Rubio-Meléndez performed the genotyping and endosymbiont detection. Frederic Francis performed the proteomic analysis. Claudio C. Ramírez analyzed the data, prepared the graphic design and wrote the original draft. All authors edited the manuscript. CONFLICT OF INTEREST The authors do not have any conflict of interest. DATA AVAILABILITY STATEMENT The data supporting this study's findings are available from the corresponding author upon request. ACKNOWLEDGMENTS The authors would like to thank the staff of the Laboratorio Interacciones Insecto-Planta (Universidad de Talca, Chile) for providing the aphids used in this study and their help performing the performance experiment. LM internship in Chile was possible thanks to the “Fonds d’Aide à la Mobilité Etudiante” of the Federation Wallonie-Bruxelles (FAME) completed by the funds of the University of Liège (Ulg) via “Fond de mobilité”. Funding of this research was also provided by the Chilean Iniciativa Científica Milenio NC120027. References Apablaza, J.U., Fernández, J.E. 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Available from: https://doi.org/10.1098/rspb.2005.3408 Leybourne, D.J., Melloh, P., Martin, E.A. (2023) Common facultative endosymbionts do not influence sensitivity of cereal aphids to pyrethroids. Agricultural and Forest Entomology , 25,344-354. Available from: https://doi.org/https://doi.org/10.1111/afe.12539 Liu, S., Liu, X., Zhang. T., et al (2023) Secondary symbionts affect aphid fitness and the titer of primary symbiont. Frontiers in Plant Science , 14. Available from: https://doi.org/10.3389/fpls.2023.1096750 Liu, X-D., Lei, H-X., Chen, F-F. (2019) Infection pattern and negative effects of a facultative endosymbiont on its insect host are environment-dependent. Scientific Reports 9, 4013. Available from: https://doi.org/10.1038/s41598-019-40607-5 Llewellyn, K.S., Loxdale, H.D., Harrington, R., et al (2004) Evidence for gene flow and local clonal selection in field populations of the grain aphid ( Sitobion avenae ) in Britain revealed using microsatellites. Heredity , 93, 143-153. Available from: https://doi.org/10.1038/sj.hdy.6800466 Luo, C., Luo, K., Meng, L., et al (2017) Ecological impact of a secondary bacterial symbiont on the clones of Sitobion avenae (Fabricius) (Hemiptera: Aphididae). Scientific Report , 7, 40754. Available from: https://doi.org/10.1038/srep40754 Nibouche, S., Fartek, B., Mississipi, S., et al (2014) Low genetic diversity in Melanaphis sacchari aphid populations at the worldwide scale. PLoS One , 9, e106067. Available from: https://doi.org/10.1371/journal.pone.0106067 Pandharikar, G., Gatti, J-L., Simon, J-C., et al (2020) Aphid infestation differently affects the defences of nitrate-fed and nitrogen-fixing Medicago truncatula and alters symbiotic nitrogen fixation. Proceedings of the Royal Society B: Biological Sciences , 287, 20201493. Available from: https://doi.org/10.1098/rspb.2020.1493 Papura, D,, Simon, J-C., Halkett, F., et al (2003) Predominance of sexual reproduction in, Romanian populations of the aphid Sitobion avenae inferred from phenotypic and genetic structure. Heredity , 90, 397-404. Available from: https://doi.org/10.1038/sj.hdy.6800262 Piffaretti, J., Clamens, A.L., Vanlerberghe-masutti, F., et al (2013) Regular or covert sex defines two lineages and worldwide superclones within the leaf-curl plum aphid ( Brachycaudus helichrysi , Kaltenbach). Molecular Ecology , 22, 3916-3932. Available from: https://doi.org/10.1111/mec.12371 Rubio-Meléndez, M.E., Barrios-SanMartin, J., Pina-Castro, F.E., et al (2019) Asexual reproduction of a few genotypes favored the invasion of the cereal aphid Rhopalosiphum padi in Chile. PeerJ , 26, 7, e7366. Available from: https://doi.org/10.7717/peerj.7366 Sepúlveda, D.A., Zepeda-Paulo, F., Ramírez, C.C., et al (2017) Diversity, frequency, and geographic distribution of facultative bacterial endosymbionts in introduced aphid pests. Insect Science , 24, 511-521. Available from: https://doi.org/10.1111/1744-7917.12313 Sunnucks, P., De Barro, P.J., Lushai, G., et al (1997) Genetic structure of an aphid studied using microsatellites: cyclic parthenogenesis, differentiated lineages, and host specialization. Molecular Ecology , 6, 1059-1073. Available from: https://doi.org/10.1046/j.1365-294X.1997.00280.x Tsuchida, T., Koga, R., Fukatsu, T. (2004) Host plant specialization governed by facultative symbiont. Science , 303,1989. Available from: https://doi.org/10.1126/science.1094611 Villarroel, C.A., González-González, A., Alvarez-Baca, J.K., Villarreal, P., et al (2022) Genome sequencing of a predominant clonal lineage of the grain aphid Sitobion avenae . Insect Biochemistry and Molecular Biology , 143, 103742. Available from: https://doi.org/10.1016/j.ibmb.2022.103742 Vorburger, C., Lancaster, M., Sunnucks, P. (2003) Environmentally related patterns of reproductive modes in the aphid Myzus persicae and the predominance of two ‘superclones’ in Victoria, Australia. Molecular Ecology , 12, 3493-3504. Available from: https://doi.org/10.1046/j.1365-294X.2003.01998.x Wang, D., Shi, X., Dai, P., et al (2016) Comparison of fitness traits and their plasticity on multiple plants for Sitobion avenae infected and cured of a secondary endosymbiont. Scientific Report , 6, 23177. Available from: https://doi.org/10.1038/srep23177 Wang, Q., Yuan, E., Ling, X., et al (2020) An aphid facultative symbiont suppresses plant defence by manipulating aphid gene expression in salivary glands. Plant Cell and Environment , 43, 2311–2322. Available from: https://doi.org/10.1111/pce.13836 Wilson, A.C.C., Sunnucks, P., Hales, D.F. (1999) Microevolution, low clonal diversity and genetic affinities of parthenogenetic Sitobion aphids in New Zealand. Molecular Ecology , 8, 1655-1666. Available from: https://doi.org/10.1046/j.1365-294x.1999.00751.x Xin, J-J., Shang, Q-L., Desneux, N., Gao, X-W. (2014) Genetic diversity of Sitobion avenae (Homoptera: Aphididae) populations from different geographic regions in China. PLoS One , 9, e109349. Available from: https://doi.org/10.1371/journal.pone.0109349 Zepeda-Paulo, F., Lavandero, B. (2021) Effect of the genotypic variation of an aphid host on the endosymbiont associations in natural host populations. Insects , 12: Available from: https://doi.org/10.3390/insects12030217 Zepeda-Paulo, F., Villegas, C., Lavandero, B. (2017) Host genotype–endosymbiont associations and their relationship with aphid parasitism at the field level. Ecological Entomology , 42, 86-95. Available from: https://doi.org/10.1111/een.12361 Tables Table 1 is available in the Supplementary Files section. Additional Declarations The authors declare no competing interests. Supplementary Files TABLE1.docx FigureS1.docx Cite Share Download PDF Status: Posted Version 2 posted You are reading this latest preprint version Show more versions Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-4338445","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":297770094,"identity":"8ccb308c-fa49-4e86-b91d-116525934524","order_by":0,"name":"Leandro Mahieu","email":"","orcid":"","institution":"University of Liege","correspondingAuthor":false,"prefix":"","firstName":"Leandro","middleName":"","lastName":"Mahieu","suffix":""},{"id":297770095,"identity":"6ca88a47-f79e-49cf-956b-6593e3f496bd","order_by":1,"name":"Angélica González-González","email":"","orcid":"https://orcid.org/0000-0001-7377-6545","institution":"University of Talca","correspondingAuthor":false,"prefix":"","firstName":"Angélica","middleName":"","lastName":"González-González","suffix":""},{"id":297770096,"identity":"511e7837-d038-48cd-a546-766fca417ffe","order_by":2,"name":"María E. Rubio-Meléndez","email":"","orcid":"https://orcid.org/0000-0002-9852-427X","institution":"University of Talca","correspondingAuthor":false,"prefix":"","firstName":"María","middleName":"E.","lastName":"Rubio-Meléndez","suffix":""},{"id":297770097,"identity":"54c56ee8-5f63-4150-b5a5-a23450ed55d4","order_by":3,"name":"Frederic Francis","email":"","orcid":"https://orcid.org/0000-0001-7731-0849","institution":"University of Liege","correspondingAuthor":false,"prefix":"","firstName":"Frederic","middleName":"","lastName":"Francis","suffix":""},{"id":297386243,"identity":"580c4006-a508-48b4-87f8-d75c32094e15","order_by":4,"name":"Claudio C. Ramirez","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA00lEQVRIiWNgGAWjYDACdhBhAGI0AIkCEI+5Ab8WZpgWngOMDWAGAyMxWkBAIoFILfzNvA8/3Si4Jyc/8435gx8GdvIM7Afxa5E4zG4snWNQbMw4O8ewsccg2bCBJxG/FgNmNgagloTEZukcoGoDoIckCDgMqIX5N1BLfZvkGcPGPwYH7InRwgayJYFHgsewGWhLIkEtEofZ2KyBWgxn8KQVzpYxSE5uI+QX/vY25ts5fxLk5dsPb/j4psLOtp/98AG8WjABG4nqR8EoGAWjYBRgAQC4sTsvNzMqQAAAAABJRU5ErkJggg==","orcid":"https://orcid.org/0000-0002-6110-3973","institution":"University of Talca","correspondingAuthor":true,"prefix":"","firstName":"Claudio","middleName":"C.","lastName":"Ramirez","suffix":""}],"badges":[],"createdAt":"2024-04-28 15:02:04","currentVersionCode":2,"declarations":{"humanSubjects":false,"vertebrateSubjects":true,"conflictsOfInterestStatement":false,"humanSubjectEthicalGuidelines":false,"humanSubjectConsent":false,"humanSubjectClinicalTrial":false,"humanSubjectCaseReport":false,"vertebrateSubjectEthicalGuidelines":true},"doi":"10.21203/rs.3.rs-4338445/v2","doiUrl":"https://doi.org/10.21203/rs.3.rs-4338445/v2","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":55821884,"identity":"515da409-8f24-4d54-8ef5-dc6f808b2202","added_by":"auto","created_at":"2024-05-03 23:48:42","extension":"jpg","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":1577461,"visible":true,"origin":"","legend":"\u003cp\u003eA) Total number of aphids (mean ± SE) and B) weight reached (mean ± SE) by \u003cem\u003eS. avenae\u003c/em\u003e individuals after 14 days of development on wheat and barley seedlings as a function of the presence or absence of the secondary endosymbiont \u003cem\u003eR. insecticola.\u003c/em\u003e C) Root/shoot ratio of dry weight of wheat and barley plants after treated with \u003cem\u003eS. avenae\u003c/em\u003e individuals depending on the presence or absence of the secondary endosymbiont \u003cem\u003eR. insecticola.\u003c/em\u003e\u003c/p\u003e","description":"","filename":"Figure1Leandro.jpg","url":"https://assets-eu.researchsquare.com/files/rs-4338445/v2/370460516f7821ee945b7909.jpg"},{"id":55821881,"identity":"7d15e526-d2cd-4faa-97cd-35d5235938d6","added_by":"auto","created_at":"2024-05-03 23:48:42","extension":"jpg","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":708103,"visible":true,"origin":"","legend":"\u003cp\u003eDistribution of metabolic pathways for proteins with different expression levels in \u003cem\u003eS. avenae\u003c/em\u003edepending on the occurrence of \u003cem\u003eR. insecticola\u003c/em\u003e secondary symbiont on barley or wheat host plants.\u003c/p\u003e","description":"","filename":"Figure2AIPB.jpg","url":"https://assets-eu.researchsquare.com/files/rs-4338445/v2/91e9635b39582f538229913b.jpg"},{"id":55822458,"identity":"471b00c6-9d93-40b6-b68b-926c825712fb","added_by":"auto","created_at":"2024-05-04 00:12:43","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":541471,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-4338445/v2/cb341226-3fea-49c5-81f7-ee3a41731b16.pdf"},{"id":55822162,"identity":"ecafe6b4-452b-411f-8000-7c0f5fbb5eda","added_by":"auto","created_at":"2024-05-03 23:56:42","extension":"docx","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":44120,"visible":true,"origin":"","legend":"","description":"","filename":"TABLE1.docx","url":"https://assets-eu.researchsquare.com/files/rs-4338445/v2/c235b51cd84a6f502460131a.docx"},{"id":55822201,"identity":"c3d2d293-b9a8-46b0-ab15-4c8c971a9ef1","added_by":"auto","created_at":"2024-05-04 00:04:42","extension":"docx","order_by":2,"title":"","display":"","copyAsset":false,"role":"supplement","size":1055070,"visible":true,"origin":"","legend":"","description":"","filename":"FigureS1.docx","url":"https://assets-eu.researchsquare.com/files/rs-4338445/v2/11ab4199fba6cc9cb89c4586.docx"}],"financialInterests":"The authors declare no competing interests.","formattedTitle":"An aphid pest superclone benefits from a facultative bacterial endosymbiont in a host dependent-manner","fulltext":[{"header":"Research Highlights","content":"\u003cul\u003e\n \u003cli\u003eThe aphid endosymbiont \u003cem\u003eR. insecticola\u003c/em\u003e affect the reproductive performance of \u003cem\u003eS. avenae\u003c/em\u003e\u003c/li\u003e\n \u003cli\u003e\u003cem\u003eR. insecticola\u003c/em\u003e alter the aphid reproductive performance in a host-dependent manner\u003c/li\u003e\n \u003cli\u003e\u003cem\u003eR. insecticola\u003c/em\u003e showed a positive effect on wheat and a negative on barley\u003c/li\u003e\n\u003c/ul\u003e"},{"header":"1. INTRODUCTION","content":"\u003cp\u003eAphids (Hemiptera: Aphididae) are important crop pests that amplify clonally under favourable conditions. There is an increasing number of studies describing the presence of few widespread multilocus genotypes (\u0026ldquo;clones\u0026rdquo; for simplicity) within aphid species, particularly invasive ones with outstanding ability to persist in time and space (called \u0026ldquo;superclones\u0026rdquo;) (Vorburger et al. \u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e2003\u003c/span\u003e; Fenton et al. \u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e2010\u003c/span\u003e; Harrison and Mondor \u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e2011\u003c/span\u003e; Piffaretti et al. \u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e2013\u003c/span\u003e; Chen et al. \u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e2013\u003c/span\u003e; Nibouche et al. \u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e2014\u003c/span\u003e; Harris-Shultz et al. \u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e2017\u003c/span\u003e, \u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e2022\u003c/span\u003e; Zepeda-Paulo et al. \u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e2017\u003c/span\u003ea; Rubio-Mel\u0026eacute;ndez et al. \u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e2019\u003c/span\u003e). However, few studies have addressed the causes explaining this ability. Hypotheses explaining the success of superclones that have found support, states that they exhibit: 1) greater polyphenism enabling responses to variable environmental factors such as host plant quality (Casta\u0026ntilde;eda et al. \u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e2010a\u003c/span\u003e), 2) obligate asexuality enabling superclones to do not spend energy on finding suitable mates or mating sites (Piffaretti et al. \u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e2013\u003c/span\u003e), 3) adaptation to their host plants (Fenton et al. \u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e2010\u003c/span\u003e), 4) the ability to tolerate allelochemicals (Casta\u0026ntilde;eda et al. \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e2010b\u003c/span\u003e), and 5) flexibility on feeding behaviour allowing a similar reproductive performance across different context (Barrios-SanMartin et al. \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2016\u003c/span\u003ea). Surprisingly, the role of facultative bacterial endosymbionts on the ecological success of these invasive superclones has not been studied yet.\u003c/p\u003e \u003cp\u003eThe grain aphid \u003cem\u003eSitobion avenae\u003c/em\u003e is a worldwide aphid-pest of cereals. While populations in UK, north of France and Romania are predominated by cyclical parthenogenetic asexual lineages and typically featured by a high genetic diversity (Sunnucks et al. \u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e1997\u003c/span\u003e; Dedryver et al. \u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e2001\u003c/span\u003e; Papura et al. \u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e2003\u003c/span\u003e; Llewellyn et al. \u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e2004\u003c/span\u003e), populations in Australia, Chile and China (the introduced range) are predominated by a dissimilar number of obligated parthenogenetic asexual lineages depending on the region (Wilson et al. \u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e1999\u003c/span\u003e; Figueroa et al. \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e2005\u003c/span\u003e; Xin et al. \u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e2014\u003c/span\u003e). In Chile, this aphid was most probably introduced in the middle of 1970s from Europe (Apablaza and Fern\u0026aacute;ndez \u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1982\u003c/span\u003e; Figueroa et al. \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e2005\u003c/span\u003e), and it has been shown that nearly 90% of its genotypic variation is accounted for by only four predominant asexual lineages (Figueroa et al. \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e2005\u003c/span\u003e; Zepeda-Paulo and Lavandero \u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). One of these asexual lineage dominates rapidly early in the season and displays low variance in performances on different hosts (Casta\u0026ntilde;eda et al. \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e2010b\u003c/span\u003e; Zepeda-Paulo and Lavandero \u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). Interestingly, asexual lineages of \u003cem\u003eS. avenae\u003c/em\u003e in Chile varied in the presence of the facultative bacterial endosymbionts (Sep\u0026uacute;lveda et al. 2014), with the most frequent asexual lineages found to be regularly harbouring \u003cem\u003eRegiella insecticola\u003c/em\u003e (Enterobacteriales, Enterobacteriaceae)(Zepeda-Paulo et al. \u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e2017\u003c/span\u003e). Since \u003cem\u003eR. insecticola\u003c/em\u003e has been reported to improve the fitness in a host dependent manner in the pea aphid (Tsuchida et al. \u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e2004\u003c/span\u003e), it is likely that this bacterium plays a role in the performance of \u003cem\u003eS. avenae\u003c/em\u003e, determining ecological success on different host plants.\u003c/p\u003e \u003cp\u003eHere we report a study with the most common genotype of the aphid \u003cem\u003eS. avenae\u003c/em\u003e in Chile, in which the presence of the predominant facultative bacterial endosymbiont \u003cem\u003eR. insecticola\u003c/em\u003e was manipulated to assess the impact of this endosymbiont on the reproductive performance of this aphid on these two host plants (wheat and barley), and the changes that this generates in the whole-body proteomic profile of aphids. The latter can shed light on the physiological mechanisms underlying reproductive performance in each host due to the presence of this facultative endosymbiont. These two hosts were chosen because they are contrasting in terms of the prevalence of \u003cem\u003eS. avenae\u003c/em\u003e in Chile (Figueroa et al. \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e2005\u003c/span\u003e; Sep\u0026uacute;lveda et al. \u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e2017\u003c/span\u003e), since while it is very recurrent in wheat it is scarce in barley, a situation that allows evaluating, controlling for the genetic background, the impact of harbouring the facultative endosymbiont \u003cem\u003eR. insecticola\u003c/em\u003e. Plant responses (root/shoot ratio) were also measured to assess the consequences of endosymbiont-dependent aphid herbivory.\u003c/p\u003e"},{"header":"2. MATERIAL AND METHODS","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003e2.1 Aphids and plants\u003c/h2\u003e \u003cp\u003eIndividuals of the most widely distributed genotype in cereal crops in the Maule to De Los Rios regions of Chile of \u003cem\u003eS. avenae\u003c/em\u003e were kindly provided by Dr. Francisca Zepeda-Paulo. This genotype was characterized with eight microsatellite loci (\u003cem\u003eSm11\u003c/em\u003e, \u003cem\u003eS3.43\u003c/em\u003e, \u003cem\u003eS16b\u003c/em\u003e, \u003cem\u003eS30\u003c/em\u003e, \u003cem\u003eS4Σ\u003c/em\u003e, \u003cem\u003eS5L\u003c/em\u003e, \u003cem\u003eSm17\u003c/em\u003e and \u003cem\u003eSm10\u003c/em\u003e) and corresponds to the genotype G as described by Zepeda-Paulo et al. (2021) found to harbour \u003cem\u003eR. insecticola\u003c/em\u003e (see more details in Zepeda-Paulo et al. 2021). In the present study we used this genotype as two asexual lineages: one infected (hereafter E+) composed by individuals naturally harbouring the facultative endosymbiont \u003cem\u003eR. insecticola\u003c/em\u003e (as collected from the field), and the other asexual lineage was originated by individuals treated with antibiotics (ampicilin, cefotaxim and gentamicin) by using the method of (i) artificial diet (synthetic diet) and (ii) micro-injections with the methods of Koga et al. (2007) and Simon et al. (2011) resulting in \u003cem\u003eR. insecticola\u003c/em\u003e-disinfected individuals (hereafter E-). Both lineages harboured the primary endosymbiont \u003cem\u003eBuchnera aphidicola\u003c/em\u003e. Aphid colonies (E\u0026thinsp;+\u0026thinsp;and E-) were reared in separate mesh cages (50 cm x 42 cm x 31 cm) containing pots with wheat or barley seedlings. Thus, four distinct colonies of the genotype G1 were reared in the laboratory.\u003c/p\u003e \u003cp\u003eHost plants used were barley (\u003cem\u003eHordeum vulgare\u003c/em\u003e L., cultivar Sebastian) and wheat (\u003cem\u003eTriticum aestivum\u003c/em\u003e L., cultivar Pantera). Plants were grown in organic soil during one week in a greenhouse by sowing about twenty seeds per pot (bottle cap in pots). Every pot was disinfected (Virginia igenix) and fertilized (Nutrifeed follare) every week. The growth chamber was under controlled conditions (22\u0026thinsp;\u0026plusmn;\u0026thinsp;1 \u0026ordm;C, 16:8 photoperiod).\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec4\" class=\"Section2\"\u003e \u003ch2\u003e2.2 Aphid reproductive performance and the effect on host plants\u003c/h2\u003e \u003cp\u003eTo use age-synchronized aphids for the experiments with E\u0026thinsp;+\u0026thinsp;as E- aphids, firstly 50 adult aphids were placed on potted wheat or barley plants. After 12 h, all adults were removed, leaving only new nymphs on plants. Four days later, four wingless aphids were transferred to each individually potted wheat or barley seedlings. In total, 20 pots with wheat seedlings with E\u0026thinsp;+\u0026thinsp;aphids, 20 pots with wheat seedlings with E- aphids, 20 pots with barley seedlings with E\u0026thinsp;+\u0026thinsp;aphids, and 20 pots with barley with E- aphids were obtained, each selected for setting the experimental systems. After 14 days the final number aphids, including winged individuals was recorded. With the aim of determining relations in aphid weights, 25 wingless adults were collected in each of the four rearing boxes. They were weighted with an analytic balance (XP2U Mettler Toledo, Columbus, OH, USA, precision: 10\u003csup\u003e\u0026minus;\u0026thinsp;3\u003c/sup\u003e mg). In addition, once the reproductive performance experiment finished, the plants were removed from the pots and carefully separated the root and shoots, dried out for about one week at 60\u0026deg;C and (Memmert Beschickung-Loading Modell 100\u0026ndash;800) and weighted with an analytical balance (Radwag as 220/c/2).\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec5\" class=\"Section2\"\u003e \u003ch2\u003e2.3 2D-DIGE and protein identifications\u003c/h2\u003e \u003cp\u003eAphids hosting or not a facultative symbiont, namely \u003cem\u003eR. insecticola\u003c/em\u003e, reared on two host, wheat or barley were collected to perform proteomic analysis. Whole body proteins from around 50 mg fresh \u003cem\u003eS. avenae\u003c/em\u003e asexual lineages were extracted, purified, and quantified as described by Francis et al. (2010). Three Cy dyes (GE Healthcare) were used for labelling and protein samples of aphid asexual lineages were labelled either with Cy3 or Cy5 and mixed with an internal reference standard protein mixture (pooled from equal aliquots from all experimental samples) labelled with Cy2. Two replicates from each treatment with one dye (Cy3 or Cy5) and a third replicate with the other of the two Cy dyes were established for a conventional dye swap of DIGE. The first and second-dimensional electrophoresis, the excision of protein spots and the process of protein identification were performed following the description in Francis et al. (2010). The Mascot server 2.2.06 with BioToolsTM3.2 (Bruker Daltonics) and NCBI (National Center for Biotechnology Information) server accessed accessed in November 2022 were used for protein identification. The identified proteins were categorized according to metabolic function using the Kegg pathway database (\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttp://www.genome.jp/kegg/pathway.html\u003c/span\u003e\u003cspan address=\"http://www.genome.jp/kegg/pathway.html\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e, accessed in November 2022) and Expasy Proteomic tools (\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttp://www.expasy.org/tools/\u003c/span\u003e\u003cspan address=\"http://www.expasy.org/tools/\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e, accessed in January 2022).\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec6\" class=\"Section2\"\u003e \u003ch2\u003e2.4 Statistical analysis\u003c/h2\u003e \u003cp\u003eThe total number of aphids (i.e., nymphs\u0026thinsp;+\u0026thinsp;adults\u0026thinsp;+\u0026thinsp;adults winged) and adults winged were subjected to \u003cem\u003eglm\u003c/em\u003e with Poisson distribution and log-link, whereas the aphid weight and plant root/shoot ratio were subjected \u003cem\u003eglm\u003c/em\u003e function with a Gaussian distribution as the error structure. The \u003cem\u003eglm\u003c/em\u003e included the factor \u0026ldquo;endosymbiont\u0026rdquo; with the levels E\u0026thinsp;+\u0026thinsp;and E-, and the factor \u0026ldquo;host plant\u0026rdquo; with the levels wheat and barley. After detecting a significant effect, the Tukey \u003cem\u003epost hoc\u003c/em\u003e comparison with Šid\u0026aacute;k correction procedure was used to make pairwise contrasts between treatments using the \u003cem\u003eemmeans\u003c/em\u003e package (Lenth R et al. \u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). Wilcoxon rank sum test was used to compare fold ratios of protein regulation. All statistical analyses were carried out with the R software, version 4.2.2 (2022; R Development Core Team).\u003c/p\u003e \u003c/div\u003e"},{"header":"3. RESULTS","content":"\u003cdiv id=\"Sec8\" class=\"Section2\"\u003e\n \u003ch2\u003e3.1 Effect of the presence endosymbiont on aphids\u0026rsquo; performance and plant growth\u003c/h2\u003e\n \u003cp\u003eThe total number of aphids at the end of 14 days varied depending on the host plant and the presence of the endosymbiont, as revealed by the significant interaction between the endosymbiont and host plant factors (Deviance\u0026thinsp;=\u0026thinsp;783.9; d.f.= 1, 76; p\u0026thinsp;\u0026lt;\u0026thinsp;0.001). In wheat, the final number of aphids was higher in the E\u0026thinsp;+\u0026thinsp;aphids, while in barley, the opposite was observed, with a higher number of aphids in the E- aphids (Fig. \u003cspan class=\"InternalRef\"\u003e1\u003c/span\u003eA). The final number of winged individuals was very low, but was affected only by the endosymbiont factor, with winged individuals only found in the E\u0026thinsp;+\u0026thinsp;aphids in both host plants (Wheat: E\u0026thinsp;+\u0026thinsp;=\u0026thinsp;0.65 \u0026plusmn; 1.04 and E-: 0 \u0026plusmn; 0; Barley: E\u0026thinsp;+\u0026thinsp;=\u0026thinsp;0.30 \u0026plusmn; 0.57 and E-: 0.05 \u0026plusmn; 0.22; Deviance\u0026thinsp;=\u0026thinsp;19.3; d.f.= 1, 78; p\u0026thinsp;\u0026lt;\u0026thinsp;0.001). Aphid final weight exhibited a significant effect of the endosymbiont factor (F\u0026thinsp;=\u0026thinsp;214.7; d.f.= 1,96; p\u0026thinsp;\u0026lt;\u0026thinsp;0.001; Fig. \u003cspan class=\"InternalRef\"\u003e1\u003c/span\u003eB) with E\u0026thinsp;+\u0026thinsp;aphids being weightier than E-. There was also a significant effect to the host plant factor (F\u0026thinsp;=\u0026thinsp;49.2; d.f.= 1,96; p\u0026thinsp;\u0026lt;\u0026thinsp;0.001), with aphids being weightier on barley than on wheat. Additionally, there was a significant interaction between the endosymbiont and host factors (F\u0026thinsp;=\u0026thinsp;5.0; d.f.= 1, 96; p\u0026thinsp;\u0026lt;\u0026thinsp;0.05), with both E\u0026thinsp;+\u0026thinsp;and E- aphids being weightier on barley than on wheat. On the other hand, the root/shoot ratio also varied depending on the host plant and the presence of the endosymbiont, as revealed by the significant interaction between endosymbiont and host plant factor (F 1, 76\u0026thinsp;=\u0026thinsp;7.79; p\u0026thinsp;\u0026lt;\u0026thinsp;0.001; Fig. \u003cspan class=\"InternalRef\"\u003e1\u003c/span\u003eC). In wheat, the root/shoot ratio was higher on plant attacked by the E\u0026thinsp;+\u0026thinsp;aphids that by E- aphids, while in barley, no difference in the root/shoot was found (Fig. \u003cspan class=\"InternalRef\"\u003e1\u003c/span\u003eC).\u003c/p\u003e\n\u003c/div\u003e\n\u003cdiv id=\"Sec9\" class=\"Section2\"\u003e\n \u003ch2\u003e3.2 Identification of differentially expressed proteins\u003c/h2\u003e\n \u003cp\u003eIn total, 43 proteins were identified from the 2D gel among which spots varied significantly (p\u0026thinsp;\u0026lt;\u0026thinsp;0.05; Figure S1). The complete properties of over- and under‐expressed proteins in \u003cem\u003eS. avenae\u003c/em\u003e asexual lineages with and without \u003cem\u003eR. insecticola\u003c/em\u003e facultative symbiont on wheat or barley host plants were listed in Table \u003cspan class=\"InternalRef\"\u003e1\u003c/span\u003e. We found that 38 proteins were related to aphids and six6 proteins were related to endosymbionts. The fold change ratios of these proteins between the aphid asexual lineages with or without \u003cem\u003eR. insecticola\u003c/em\u003e on the same plant, either wheat or barley ranged from 0.27 to 7.7. The absolute range of protein regulation (considering the extremes of up and downregulation fold-ratios) in barley was 1.43 fold-ratio, while it was 0.47 fold-ratio in wheat. With regards to proteins in the aphid group, 58% were detected in \u003cem\u003eA. pisum\u003c/em\u003e while fewer (8%) were detected in other species (\u003cem\u003eA. citricidus\u003c/em\u003e, \u003cem\u003eA. gossypii\u003c/em\u003e and \u003cem\u003eS. avenae\u003c/em\u003e). With regards to the endosymbiont group, various proteins were detected in \u003cem\u003eB. aphidicola\u003c/em\u003e (63%), the other were related to Regiella and Rickettsia facultative endosymbionts.\u003c/p\u003e\n \u003cp\u003eProteins with different expression levels were relate to several metabolic pathways (Table \u003cspan class=\"InternalRef\"\u003e1\u003c/span\u003e). We found that these proteins accounted for various roles in metabolic pathways such as genetic information processing (20%), cytoskeleton (18%), environmental information processing (16%), energy metabolism (11%), carbohydrates (9%), cellular process and stress responses (both 5%). The presence of \u003cem\u003eR. insecticola\u003c/em\u003e in aphid developing in barley showed a greater average upregulation of proteins than in wheat (barley fold-ratio\u0026thinsp;=\u0026thinsp;2.0 \u0026plusmn; 1.4, wheat fold-ratio\u0026thinsp;=\u0026thinsp;1.27 \u0026plusmn; 0.28, p\u0026thinsp;\u0026lt;\u0026thinsp;0.001, W\u0026thinsp;=\u0026thinsp;419, Wilcoxon rank sum test), whereas on wheat infected aphids showed a greater average downregulation than on barley (wheat fold-ratio\u0026thinsp;=\u0026thinsp;0.88 \u0026plusmn; 0.17, barley fold-ratio\u0026thinsp;=\u0026thinsp;0.57 \u0026plusmn; 0.29, p\u0026thinsp;\u0026lt;\u0026thinsp;0.001, W\u0026thinsp;=\u0026thinsp;75, Wilcoxon rank sum test).\u003c/p\u003e\n \u003cp\u003eOn barley energy and carbohydrate metabolism, as well as cellular process and response to stress were highly downregulated. Contrastingly, downregulation in wheat was globally mild, except in the case of other protein group, particularly in the 1-acyl-sn-glycerol-3-phosphate acyltransferase. Upregulation was globally mild on both host plants, with a distinct high upregulation of the protein 1-acyl-sn-glycerol-3-phosphate acyltransferase on E\u0026thinsp;+\u0026thinsp;aphids developing on barley.\u003c/p\u003e\n\u003c/div\u003e"},{"header":"4. DISCUSSION","content":"\u003cp\u003eIn this work we have found that a single genotype of the aphid \u003cem\u003eS. avenae\u003c/em\u003e proliferates differentially in two hosts depending on the presence of a facultative bacterial endosymbiont. The most striking result of our study was that among aphids sharing the same genetic background, those developing on wheat and harbouring the facultative endosymbiont \u003cem\u003eR. insecticola\u003c/em\u003e exhibit larger colony development, while the opposite was true for the aphids developing in barley. A similar positive effect of this endosymbiont on reproduction has been found on the Vicia-specialized pea aphid populations (Tsuchida et al. \u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e2004\u003c/span\u003e). However, that result was not reproduced in other pea aphid or other aphid species, with studies showing negative or neutral effects of \u003cem\u003eR. insecticola\u003c/em\u003e on aphid reproduction (Ferrari et al. \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e2004\u003c/span\u003e, \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e2007\u003c/span\u003e; Wang et al. \u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e2016\u003c/span\u003e; Luo et al. \u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e2017\u003c/span\u003e; Liu et al. \u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e2019\u003c/span\u003e). Nevertheless, our results are consistent with field-based monitoring of \u003cem\u003eS. avenae\u003c/em\u003e in wheat plantations showing \u003cem\u003eR. insecticola\u003c/em\u003e increased the prevalence throughout time when compared with uninfected individuals (Zepeda-Paulo and Lavandero \u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). It should be noted that G1 genotype dominates early in the season, but a recent study showed that its predominance significantly decreased at mid-season (Zepeda-Paulo and Lavandero \u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). However, although G1 decreases its prevalence during the season studied, it persists over the years as describe by Figueroa et al. )2005).\u003c/p\u003e \u003cp\u003eRam\u0026iacute;rez-Caceres et al. (2019), studying another \u003cem\u003eS. avenae\u003c/em\u003e genotype (G2) also described a higher population growth of E\u0026thinsp;+\u0026thinsp;aphids on wheat than on barley, while E- negatively affect the growth of aphids on barley. All this support that \u003cem\u003eR. insecticola\u003c/em\u003e entail within-genotype difference on \u003cem\u003eS. avenae\u003c/em\u003e performance across two common food plants, wheat and barley. This suggest that host plant x facultative endosymbiont interaction may drive host specialization even within a genotype. Thus, clones composed by E\u0026thinsp;+\u0026thinsp;aphid on wheat and E- aphids on barley, are expected to be positive selected and negatively selected, respectively, leading to ecological divergent populations dominated by asexual clones. However, given the temporal instability of the cereal plantations in Chile, these populations might not reach such a divergence (Gonz\u0026aacute;lez U et al. 2013). The lack of host-based differentiation of \u003cem\u003eS. avenae\u003c/em\u003e populations in Chile confirm this hypothesis (Figueroa et al. \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e2005\u003c/span\u003e). Nevertheless, it is surprising that the same facultative endosymbiont can generate such dissimilar effects on its host aphid, which raises several questions about the underlying mechanisms.\u003c/p\u003e \u003cp\u003eOur result show that E\u0026thinsp;+\u0026thinsp;aphids on both host plants showed higher number of winged individuals, although in a very low number, this contrast with previous studies. For example, E\u0026thinsp;+\u0026thinsp;individuals of asexual lineages of \u003cem\u003eS. avenae\u003c/em\u003e from China under crowed conditions, produced less winged offspring than E- aphids (Liu et al. \u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e2019\u003c/span\u003e). This result was found at 25\u0026deg;C, which is slightly higher than our conditions (22\u0026thinsp;\u0026plusmn;\u0026thinsp;1 \u0026ordm;C). They also found that winged morph production did not showed differences among E\u0026thinsp;+\u0026thinsp;and E- at higher temperatures, which suggested that winged morph production was dependent on environmental temperature and aphid density. On the other hand, \u003cem\u003eR. insecticola\u003c/em\u003e negatively affect the production of winged offspring in the pea aphid (Leonardo and Mondor \u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e2006\u003c/span\u003e). In our case, because the production of winged individuals in E\u0026thinsp;+\u0026thinsp;aphids was irrespective of the host plant, this could be an idiosyncratic capacity of this asexual lineage (G1) to respond to the infection with \u003cem\u003eR. insecticola\u003c/em\u003e. Remarkably, this asexual lineage is one the most persistent and predominant in the wheat field of Chile (Figueroa et al. \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e2005\u003c/span\u003e), and frequently found harbouring \u003cem\u003eR. insecticola\u003c/em\u003e (Zepeda-Paulo et al. \u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e2017\u003c/span\u003e) and thus the evolution of this lineage with \u003cem\u003eR. insecticola\u003c/em\u003e might have reached a fixed pattern of response to this endosymbiont.\u003c/p\u003e \u003cp\u003eSurprisingly, the body weight of E\u0026thinsp;+\u0026thinsp;and E- aphids did not fallowed the same trend as reproductive performance. Instead, regardless host plant, aphid showed greater weight when they harboured \u003cem\u003eR. insecticola\u003c/em\u003e. This indicates that the production of offspring is uncoupled with the weight of each individual. This contrast with the results found in \u003cem\u003eRhopalosiphum maidis\u003c/em\u003e (Fitch) feeding on barley, where \u003cem\u003eR. insecticola\u003c/em\u003e-infected aphids performed poorly in weight (Liu et al. \u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e2023\u003c/span\u003e). Regarding the distinct effect of \u003cem\u003eR. insecticola\u003c/em\u003e on weight and reproductive performance in aphids of \u003cem\u003eS. avenae\u003c/em\u003e sharing the same genetical background, again open questions about the undelaying mechanisms of these responses. Thus, we subsequently develop a proteomic study with the infected and non-infected aphids after their development in wheat or barley, to obtain some light on what the underlying mechanisms are.\u003c/p\u003e \u003cp\u003eOur findings suggest that the presence of \u003cem\u003eR. insecticola\u003c/em\u003e generates different changes in the proteomic profile of \u003cem\u003eS. avenae\u003c/em\u003e depending in a host-dependent manner, that could account for the difference in reproductive performance. Interestingly, the fact that protein regulation of aphids developing on wheat was comparatively milder and steadier than on barley, suggest that E\u0026thinsp;+\u0026thinsp;reared on wheat inflict lower impact of their physiology. In this regard, due to expansion of gene families associated with resistance to insecticides and plant chemical defenses described in the genome of \u003cem\u003eS. avenae\u003c/em\u003e (Villarroel et al. \u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e2022\u003c/span\u003e), if \u003cem\u003eR. insecticola\u003c/em\u003e would have a beneficial effect on confronting those compounds, a larger regulation of those proteins would be expected when comparing E\u0026thinsp;+\u0026thinsp;and E- aphids. However, no differential regulation of those proteins was found in our study. Interestingly, this is consistent with the lack of association between the presence of \u003cem\u003eR. insecticola\u003c/em\u003e and sensitivity to pyrethroids in \u003cem\u003eS. avenae\u003c/em\u003e population from Germany (Leybourne et al. \u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e2023\u003c/span\u003e). Since populations of \u003cem\u003eS. avenae\u003c/em\u003e in Chile are most predominant in wheat than on barley (Apablaza and Fern\u0026aacute;ndez \u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1982\u003c/span\u003e; Figueroa et al. \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e2005\u003c/span\u003e) and that the genotype G1 is also predominant (Zepeda-Paulo et al. \u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e2017\u003c/span\u003e; Zepeda-Paulo and Lavandero \u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e2021\u003c/span\u003e), the comparative lower protein regulation on wheat suggests that much steady physiological response as compared with that on barley, is probably due to a recent adaptation of \u003cem\u003eS. avenae\u003c/em\u003e to wheat after introduction. Indeed, it has been described that \u003cem\u003eS. avenae\u003c/em\u003e superclones exhibit a broad host range, flat energetic costs for non-induced detoxification enzymes, and low variation in their reproductive performance on different host plants (Casta\u0026ntilde;eda et al. \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e2010b\u003c/span\u003e; Barrios-SanMartin et al. \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2016\u003c/span\u003eb). The facultative aphid endosymbiont \u003cem\u003eSerratia symbiotica\u003c/em\u003e manipulates the expression of specific proteins in the pea aphid impairing plant defence response and improve feeding (Wang et al. \u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e2020\u003c/span\u003e), a mechanism that may be occurring in E\u0026thinsp;+\u0026thinsp;in \u003cem\u003eS. avenae\u003c/em\u003e aphids.\u003c/p\u003e \u003cp\u003eThe lower reproductive performance of E\u0026thinsp;+\u0026thinsp;aphids on barley could be link to the higher number of proteins that were upregulated in these aphids, which could be an indicative of a reaction to a bacterium infection. The upregulation of the 1-acyl-sn-glycerol-3-phosphate acyltransferase enzyme (plsC), which participates several lipid biosynthetic pathways (Chen et al. \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e2011\u003c/span\u003e), and the elongation factor 1-alpha, may both be related with active enzymes delivery as a reaction to \u003cem\u003eR. insecticola\u003c/em\u003e. However, the functional role of these upregulated protein on E\u0026thinsp;+\u0026thinsp;aphids remain to be deciphered. On the other hand, downregulation of proteins in E\u0026thinsp;+\u0026thinsp;on barley such as such as murein hydrolase activator EnvC, a protein produced by \u003cem\u003eR. insecticola\u003c/em\u003e and normally associated with bacteria proliferation within aphids (Cook et al. \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e2020\u003c/span\u003e), could be aphid defensive response against \u003cem\u003eR. insecticola\u003c/em\u003e induced by feeding on barley. It remains to be deciphered what are the host-dependent cues triggering such a striking difference in the aphid physiology.\u003c/p\u003e \u003cp\u003eRegarding the effects of E\u0026thinsp;+\u0026thinsp;and E- aphids on the plants, the higher root/shoot ratio exhibited by E\u0026thinsp;+\u0026thinsp;aphids compared to E- aphids on wheat plants but not on barley, suggests that these plants experienced a greater response stress. Since plants usually show a higher root/shoot ratio under water deficiency, it seems that E\u0026thinsp;+\u0026thinsp;aphids on wheat were more damaging than on E-. It is likely that aphids harbouring \u003cem\u003eR. insecticola\u003c/em\u003e obtain and specific benefit on wheat which increase their efficiency in the use of this specific plant. This result contrasts with what was found in \u003cem\u003eMedicago truncatula\u003c/em\u003e, where plants treated with pea aphids free or infected with \u003cem\u003eR. insecticola\u003c/em\u003e showed no difference on the dry weight of plant shoots (Pandharikar et al. \u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e2020\u003c/span\u003e).\u003c/p\u003e"},{"header":"5. CONCLUSION","content":"\u003cp\u003eIn summary, our laboratory study we detected important effects of \u003cem\u003eR. insecticola\u003c/em\u003e on \u003cem\u003eS. avenae\u003c/em\u003e reproductive performance and proteomic which were highly influenced by the host plant. Since these results were detected within one single aphid genotype, the direct impact of the facultative endosymbionts x host plants interaction it is highlighted. Accordingly, as the presence of facultative endosymbionts can alter aphid reproductive performance in a host-dependent manner, we found that the prevalence of facultative endosymbionts, larger on wheat and lower on barley, is also dependent of the host plants.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eAUTHOR CONTRIBUTIONS\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eClaudio C. Ram\u0026iacute;rez and Frederic Francis conceptualized the research and designed the experiments. Leandro Mahieu and Alg\u0026eacute;lica Gonz\u0026aacute;lez-Gonz\u0026aacute;lez conducted the aphid performance experiments. Mar\u0026iacute;a Eugenia Rubio-Mel\u0026eacute;ndez performed the genotyping and endosymbiont detection. Frederic Francis performed the proteomic analysis. Claudio C. Ram\u0026iacute;rez analyzed the data, prepared the graphic design and wrote the original draft. All authors edited the manuscript.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCONFLICT OF INTEREST\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors do not have any conflict of interest.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eDATA AVAILABILITY STATEMENT\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe data supporting this study\u0026apos;s findings are available from the corresponding author upon request.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eACKNOWLEDGMENTS\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors would like to thank the staff of the Laboratorio Interacciones Insecto-Planta (Universidad de Talca, Chile) for providing the aphids used in this study and their help performing the performance experiment. LM internship in Chile was possible thanks to the \u0026ldquo;Fonds d\u0026rsquo;Aide \u0026agrave; la Mobilit\u0026eacute; Etudiante\u0026rdquo; of the Federation Wallonie-Bruxelles (FAME) completed by the funds of the University of Li\u0026egrave;ge (Ulg) via \u0026ldquo;Fond de mobilit\u0026eacute;\u0026rdquo;. Funding of this research was also provided by the Chilean Iniciativa Cient\u0026iacute;fica Milenio NC120027.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eApablaza, J.U., Fern\u0026aacute;ndez, J.E. (1982) Preferencias de los afidos \u003cem\u003eMetopolophium dirhodum\u003c/em\u003e (Walker) y \u003cem\u003eSitobion avenae\u003c/em\u003e (Fabricius) entre plantulas de avena, cebada y trigo y triticale. \u003cem\u003eInternational Journal of Agriculture and Natural Resources,\u003c/em\u003e 9, 37-41. Available from: http://dx.doi.org/10.7764/rcia.v9i1.708\u003c/li\u003e\n\u003cli\u003eBarrios-SanMartin, J., Figueroa, C.C., Ramirez, C.C. 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Available from: https://doi.org/10.1371/journal.pone.0109349\u003c/li\u003e\n\u003cli\u003eZepeda-Paulo, F., Lavandero, B. (2021) Effect of the genotypic variation of an aphid host on the endosymbiont associations in natural host populations. \u003cem\u003eInsects\u003c/em\u003e, 12: Available from: https://doi.org/10.3390/insects12030217\u003c/li\u003e\n\u003cli\u003eZepeda-Paulo, F., Villegas, C., Lavandero, B. (2017) Host genotype\u0026ndash;endosymbiont associations and their relationship with aphid parasitism at the field level. \u003cem\u003eEcological Entomology\u003c/em\u003e, 42, 86-95. Available from: https://doi.org/10.1111/een.12361\u003c/li\u003e\n\u003c/ol\u003e"},{"header":"Tables","content":"\u003cp\u003eTable 1 is available in the Supplementary Files section.\u003c/p\u003e"}],"fulltextSource":"","fullText":"","funders":[{"identity":"e33e7bbf-41c2-4a65-a43f-7192bbd37a89","identifier":"10.13039/100008725","name":"Agencia Nacional de Investigación e Innovación","awardNumber":"Chilean Iniciativa Científica Milenio NC120027.","order_by":0}],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":true,"hideJournal":true,"highlight":"","institution":"University of Talca","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":true,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"host specialization, endosymbionts, aphid population growth rate, insect proteomic","lastPublishedDoi":"10.21203/rs.3.rs-4338445/v2","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-4338445/v2","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eThe English grain aphid, \u003cem\u003eSitobion avenae\u003c/em\u003e, is a significant agricultural pest affecting wheat, barley, and oats. In Chile, the most prevalent and persistent clone (superclone) of \u003cem\u003eS. avenae\u003c/em\u003e harbours the facultative endosymbiont bacterium \u003cem\u003eRegiella insecticola\u003c/em\u003e. To determine the role of this bacteria in the ecological success of this superclone, the presence of \u003cem\u003eR. insecticola\u003c/em\u003e was manipulated to evaluate the impact on 1) the reproductive performance of this clone in two host plant species (wheat and barley), 2) the production of winged morphs, 3) changes in the proteomic profile of insects, and 4) root/shoot ratio of plant. It was determined that this superclone of \u003cem\u003eS. avenae\u003c/em\u003e proliferates differentially in the host plants studied depending on the presence of the facultative bacterial endosymbiont, observing that the clone develops better in wheat when it is infected with \u003cem\u003eR. insecticola\u003c/em\u003e while the opposite occurs when it develops in barley. Aphid biomass was higher when harbouring \u003cem\u003eR. insecticola\u003c/em\u003e, particularly in barley. Individuals infected with \u003cem\u003eR. insecticola\u003c/em\u003e, in both host plants, showed higher proportion of winged individuals. The protein regulation of aphids on wheat was comparatively lower and stable than that on barley. A higher root/shoot biomass ratio was detected in wheat than in oats in plants attacked with aphids harbouring \u003cem\u003eR. insecticola\u003c/em\u003e. \u003cem\u003eR. insecticola\u003c/em\u003e significantly affects the reproductive and proteomic performance of the \u003cem\u003eS. avenae\u003c/em\u003e superclone, changes influenced by the host plant, suggesting that the host plant x facultative endosymbiont interaction can drive host specialization intraclonally, partly the ecological success of the superclones.\u003c/p\u003e","manuscriptTitle":"An aphid pest superclone benefits from a facultative bacterial endosymbiont in a host dependent-manner","msid":"","msnumber":"","nonDraftVersions":[{"code":2,"date":"2024-05-03 23:48:37","doi":"10.21203/rs.3.rs-4338445/v2","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}},{"code":1,"date":"2024-04-30 20:31:02","doi":"10.21203/rs.3.rs-4338445/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"ea935dd5-4aef-47b8-bb35-40ac282df7e0","owner":[],"postedDate":"May 3rd, 2024","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[{"id":31374117,"name":"Entomology"}],"tags":[],"updatedAt":"2024-04-30T20:31:02+00:00","versionOfRecord":[],"versionCreatedAt":"2024-05-03 23:48:37","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v2","identity":"rs-4338445","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-4338445","identity":"rs-4338445","version":["v2"]},"buildId":"qtupq5eGEP_6zYnWcrvyt","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}