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bioRxiv preprint doi: https://doi.org/10.1101/086223; this version posted November 7, 2016. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license.

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! "! bioRxiv preprint doi: https://doi.org/10.1101/086223; this version posted November 7, 2016. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license.

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! -! bioRxiv preprint doi: https://doi.org/10.1101/086223; this version posted November 7, 2016. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license.

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! 1! bioRxiv preprint doi: https://doi.org/10.1101/086223; this version posted November 7, 2016. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license.

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! 2! bioRxiv preprint doi: https://doi.org/10.1101/086223; this version posted November 7, 2016. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license.

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! 3! bioRxiv preprint doi: https://doi.org/10.1101/086223; this version posted November 7, 2016. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license.

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! 4! bioRxiv preprint doi: https://doi.org/10.1101/086223; this version posted November 7, 2016. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license.

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! "5! bioRxiv preprint doi: https://doi.org/10.1101/086223; this version posted November 7, 2016. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license.

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! ""! bioRxiv preprint doi: https://doi.org/10.1101/086223; this version posted November 7, 2016. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license.

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! "-! bioRxiv preprint doi: https://doi.org/10.1101/086223; this version posted November 7, 2016. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license.

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! "0! bioRxiv preprint doi: https://doi.org/10.1101/086223; this version posted November 7, 2016. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license.

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! "2! bioRxiv preprint doi: https://doi.org/10.1101/086223; this version posted November 7, 2016. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license.

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! "3! bioRxiv preprint doi: https://doi.org/10.1101/086223; this version posted November 7, 2016. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license.

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! "4! bioRxiv preprint doi: https://doi.org/10.1101/086223; this version posted November 7, 2016. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license.

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! -/! bioRxiv preprint doi: https://doi.org/10.1101/086223; this version posted November 7, 2016. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license.

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! -0! bioRxiv preprint doi: https://doi.org/10.1101/086223; this version posted November 7, 2016. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license.

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! -1! bioRxiv preprint doi: https://doi.org/10.1101/086223; this version posted November 7, 2016. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license.

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! -2! bioRxiv preprint doi: https://doi.org/10.1101/086223; this version posted November 7, 2016. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license.

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[!3"+-,-!$1(%4($!"2$,4&'!#!$4&'+(!.#%,(04#+!"#1+2,51(! 12! 13! 3O.! 30! 05! [! \#%,(04#+! "#1+2,51($! 4&*(%,4&'! #! $4&'+(! 3"+-,-! 30! 2/! 33! 05! 34! $1(%4($! 3.-! !

! -4! bioRxiv preprint doi: https://doi.org/10.1101/086223; this version posted November 7, 2016. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license.

Host plant Buchnera Serratia Erwinia Type-X Sodalis Wolbachia HamiltonellaRickettsia EdwardsiellaAcinetobacterRegiella Unknown Enreobacteriaceae Tuberolachnus_salignus Tr ama_troglodytes Mindar us_sp Lachnus_roboris C_sp_3210 C_cf_costata C_englemanniensis_bonica C_togyuensis_smaragdina C_pectinatae C_todocola1 C_todocola2 C_occidentalis1 C_occidentalis2 C_occidentalis3 C_occidentalis4 C_cf_confinis C_confinis C_hattor i_longipennis C_sp_3415 C_grande C_tujafilina C_fresai2 C_fresai1 C_petersoni Cinara C_juniperi2 C_juniperi1 C_cedri2 C_cedri1 C_puerca C_laportei C_piceae C_cur tihirsuta C_curvipes C_cf_piceae C_cf_coloradensis1 C_coloradensis C_cf_coloradensis2 C_pseudotaxifoliae2 C_pseudotaxifoliae1 C_pseudotaxifoliae3 C_splendens_pseudotsugae C_pseudotsugae_sensu_Palmer C_splendens_sensu_Palmer C_cuneomaculata C_laricicola2 C_laricicola1 C_kochi C_kochiana C_laricis C_laricifex C_laricifoliae C_cf_spiculosa Schizolachnus C_glabra1 C_glabra3 C_glabra2 C_solitar ia C_watsoni C_pergandei1 C_pergandei2 C_strobi C_watanabei C_cembrae C_cembrae_mongolica C_shinjii C_anelia C_moketa C_apini C_wahtolca C_formosana C_gudaris C_piniformosana C_pinea C_neubergi C_brauni C_pinimaritimae C_pilicornis3 C_pilicornis1 C_pilicornis2 C_pruinosa4 C_pruinosa5 C_pruinosa3 C_pruinosa_eozana C_pruinosa2 C_pruinosa6 C_pruinosa1 C_wahluca C_sp_3274 C_sp_3348 C_sp_3017 C_obscura_pallidipes C_fornacula2 C_fornacula1 C_sp_3082 C_sp_3208 C_sp_3046 C_obscura_sitchensis C_sp_3011 C_arizonica C_edulis1 C_edulis2 C_schwartzi C_thatcheri C_sp_3328 C_nuda C_pini2 C_pini1 C_cf_guadarramae C_maghrebica C_palaestinensis C_neweli_cronarti C_parvicornis C_brevispinosa C_nigra2 C_nigra3 C_nigra1 C_gracilis C_nigra5 C_nigra6 C_nigra4 C_oregonensis C_pinivora C_atlantica_banksiana C_harmonia C_murrayanae C_medispinosa C_terminalis2 C_terminalis1 0 20 60 100 C_terminalis3 C_ponderosae1 C_ponderosae3 Percentage of reads C_ponderosae2 C_ponderosae4

Pinus Abies Cupre. Picea Larix Pseud. Cedrus other

Figure 1. Bacterial community (phylum and frequency as estimated by read abundances) of Cinara species mapped onto the tips of the ultrametric species tree of Cinara. For the total number of reads found in an aphid specimen, we have calculated the percent- age of reads belonging to each bacterium. In this figure, only endosymbionts that are fixed within Cinara species (bacteria found in all specimens of a given species) are plotted. Therefore, the frequency of each endosymbiont (the red bars) in a species represents the mean percentage of reads obtained in all the specimens of that species (including PCR replicates). Colours in the figure corre- spond to the colour circles and conifer silhouettes in the inset legend. Black circles in the inset map show the collection sites of the specimens of Cinara (N=371). The photo shows Cinara ponderosae (col: Coeur d’acier). bioRxiv preprint was notcertifiedbypeerreview)istheauthor/funder,whohasgrantedbioRxivalicensetodisplaypreprintinperpetuity.Itmade and the riboflavin biosynthesis take-over/rescue between Figure 2. different species are: CLCA= are: species of different relationships phylogenetic the of representation schematic a left, of systems endosymbiotic the in symbionts obligate from Gosalbesetal(2008),Lamelasal.(2011a,b) andManzano-Marín&Latorre(2014). = Cced ; Cpse = Cpse cedri; C. Diagram representing the metabolic complementation found in the biosynthesis of tryptophan, biotin CLCA doi: Cced Cpse Ccon Cfor Cstr Ctuj https://doi.org/10.1101/086223

trpE tryptophan trpG

; Cfor = Cfor pseudotaxifoliae; C. trpD trpC available undera Cinara trpA trpB

last common ancestors; Ccon = Ccon ancestors; common last bioC

fabB(F) ; this versionpostedNovember7,2016. fabG CC-BY-NC-ND 4.0Internationallicense

biotin fabZ fabI* bioH . Gene names are used as column names. On the On names. column as used are names Gene Cinara. . Data from Data fornacula. C. bioF

Buchnera bioA bioD bioB ribA and–

riboflavin ribD pce.Abeitos o the for Abbreviations species. Cinara

, Ctuj = Ctuj confinis, Cinara yigB(ybjI)* The copyrightholderforthispreprint(which Serratia, Erwinia, Sodalis or Type-X ribB . C. ribE tujhafilina ribC ribF Serratia Buchnera Sodalis Buchnera Erwinia Buchnera Serratia Buchnera Serratia Buchnera Buchnera Serratia Buchnera X-type and C. cedri C. C. tujhafilina; C. Redundancy Missing Both Alternative Present comes bioRxiv preprint doi: https://doi.org/10.1101/086223; this version posted November 7, 2016. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license.

Serratia Erwinia Sodalis Type-X

100 Habit Feeding-site Feeding-range Feeding-plant Distribution

50 harbouring symboints Number of aphid species

0 WP EP EN WN Abies Larix single colony woody Pinus Picea Pseudot. Cupres. Cedrus no-woody polypagous monopagous Figure 3. Distribution of cosymbionts across aphid’s ecological contexts. For several aphids characters, the histograms show the number of species hosting each cosymbionts. Abbreviations: Pseudot.= Pseudotsuga, Cupre.=Cupressaceae, WP= Western Palearctic, EP= Eastern Palearctic, WN= Western Nearctic, EN= Eastern Nearctic. bioRxiv preprint doi: https://doi.org/10.1101/086223; this version posted November 7, 2016. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license. Serratia Erwinia Sodalis Type-X

bio5

bio1

bio6 2

4 1 3

bio13 axis 2 33%

bio12

bio14

axis 1 44.2%

Figure 4. Circle of correlation and factor scores obtained from a principal component analysis performed on bioclimatic variables. Ellipses delimit the climatic envelopes of aphids hosting the differ- ent cosymbionts. Climatic values were extracted from the localities where aphid specimens carrying the essential symbionts were found. The percentages of variance explained by the first two principal components are indicated in the axis labels. Abbreviations; BIO1 = Annual Mean Temperature; BIO5 = Max Temperature of Warmest Month; BIO6 = Min Temperature of Coldest Month; BIO12 = Annual Precipitation; BIO13 = Precipitation of Wettest Month; BIO14 = Precipitation of Driest Month