Phylogenetic Relationships of Santalales with Insights Into the Origins of Holoparasitic Balanophoraceae Huei-Jiun Su, Jer-Ming Hu, Frank E

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Vol. 64 (3) • June 2015

International Journal of Taxonomy, Phylogeny and Evolution

Electronic Supplement to

Phylogenetic relationships of Santalales with insights into the origins of holoparasitic Balanophoraceae Huei-Jiun Su, Jer-Ming Hu, Frank E. Anderson, Joshua P. Der & Daniel L. Nickrent

Taxon : – TAXON 64 (3) • June 2015 Electr. Suppl. to: Su & al. • Phylogenetic relationships of Santalales

APPENDIX S1. JUSTIFICATION FOR WHY parasite lineages, they found that Cynomoriaceae diverged ca. CYNOMORIUM WAS EXCLUDED FROM 100 Ma and that it was most closely related to Saxifragales. THIS STUDY In summary, the genes that support placing Cynomorium in (or sister to) Saxifragales include nuclear SSU rDNA and Historically, Cynomorium L. has either been included in mitochondrial atp1, coxI, matR and nad5. Because the mito- Balanophoraceae or not. For example, Eichler (1867) removed chondrial genes atp1 and coxI are known to be subjected to Cynomorium from Balanophoraceae as did Van Tieghem horizontal transfer, particularly in holoparasitic angiosperms, (1896). Interestingly, Eichler (1873) reversed his position on this it is not clear how different results were obtained in Qiu & al. in the Prodromus. Workers in the 20th century usually included (2010) vs. Naumann & al. (2013). Finally, one should be skeptical Cynomoriaceae in Balanophoraceae (Cronquist, 1981) or at least about phylogenetic results derived from plastid-encoded genes placed it proximally in the classification system (Takhtajan, from Cynomorium given the heteroplasmic nature of this organ- 1997, 2009). elle and the extreme reduction and modification its plastome Nickrent & al. (2005) were the first to use molecular data to experienced as the plant evolved the holoparasitic habit. show that Cynomorium and Balanophoraceae were not closely related. Support was strong for Balanophoraceae in or near Santalales and Cynomorium in Saxifragales. Curiously, APG LITERATURE CITED III (2009) stated that the evidence for placing Cynomoriaceae in Saxifragales “is not strong”. The MP plus BI tree shown in APG III 2009. An update of the Angiosperm Phylogeny Group classifi- Nickrent & al. (2005) utilized nuclear SSU rDNA, chloroplast cation for the orders and families of flowering plants. Bot. J. Linn. rbcL and atpB, and mitochondrial matR. That tree (their fig. 2) Soc. 161: 105–121. http://dx.doi.org/10.1111/j.1095-8339.2009.00996.x Barkman, T.J., McNeal, J.R., Lim, S.-H., Coat, G., Croom, H.B., shows Cynomorium as sister to Peridiscus (MPBS 98, BIPP Young, N.D. & dePamphilis, C.W. 2007. Mitochondrial DNA 0.78) and that clade sister to Hamamelis (MPBS 98, BIPP 100), suggests at least 11 origins of parasitism in angiosperms and reveals both members of Saxifragales. The tree that resulted from an genomic chimerism in parasitic plants. B. M. C. Evol. Biol. 7: 248. attempt to more precisely place Cynomorium within Saxifrag- http://dx.doi.org/10.1186/1471-2148-7-248 ales (their fig. 3) did not have strong support for any particular Cronquist, A. 1981. An integrated system of classification of flowering plants. Bronx, New York: Columbia University Press. relationship. Eichler, A.W. 1867. Sur la structure femelle de quelques Balanopho- Barkman & al. (2007) recovered a relationship with Saxi- racées. Pp. 137–155 in: Fournier, E. (ed.), Actes du congrès inter- fragales using matR but a combined atp1 and coxI analyses national de botanique tenu à Paris en août 1867. Paris: Germer supported a placement with Sapindales. This conflicting rela- Baillière. http://dx.doi.org/10.5962/bhl.title.11026 tionship is likely the result of horizontal gene transfer, known Eichler, A.W. 1873. Balanophoraceae. Pp. 117–150 in: Candolle, A. de to occur with both atp1 and coxI. (ed.), Prodromus systematis naturalis regni vegetabilis, vol. 17. Parisiis [Paris]: sumptibus G. Masson. Cynomorium was excluded from the study of Saxifragales http://dx.doi.org/10.5962/bhl.title.286 by Jian & al. (2008) because the authors report that it was placed García, M.A., Nicholson, E.H. & Nickrent, D.L. 2004. Extensive in Santalales in an unpublished analysis of 561 angiosperms. intra-individual variation in plastid rDNA sequences from the holo- Given that phylogenetic position, it is likely that the authors parasite Cynomorium coccineum (Cynomoriaceae). J. Molec. Evol. accidentally used a sample of Balanophoraceae; however, no 58: 322–332. http://dx.doi.org/10.1007/s00239-003-2554-y voucher was cited to verify the identity of the plant sampled. Jian, S., Soltis, P.S., Gitzendanner, M.A., Moore, M.J., Li, R., Hendry, T.A., Qiu, Y.-L., Dhingra, A., Bell, C.D. & Soltis, Zhang & al. (2009) conducted a phylogenetic study using D.E. 2008. Resolving an ancient, rapid radiation in Saxifragales. the inverted repeat of the plastid genome. Their data indicated Syst. Biol. 57: 38–57. http://dx.doi.org/10.1080/10635150801888871 that Cynomorium songaricum was sister to Rosaceae (Prunus Molina, J., Hazzouri, K.M., Nickrent, D., Geisler, M., Meyer, and Fragaria) with 99% BS, a result also seen by Moore & al. R.S., Pentony, M.M., Flowers, J.M., Pelser, P., Barcelona, J., (2011) using the same molecular marker. As was shown by Gar- Inovejas, S.A., Uy, I., Yuan, W., Wilkins, O., Michel, C.-I., LockLear, S., Concepcion, G.P. & Purugganan, M.D. 2014. Pos- cía & al. (2004), Cynomorium has extensive intraindividual sible loss of the chloroplast genome in the parasitic flowering plant variation in plastid rDNA, likely deriving from heteroplasmy. Rafflesia lagascae (Rafflesiaceae). Molec. Biol. Evol. 31: 793–803. For this reason, one must use extreme caution before using http://dx.doi.org/10.1093/molbev/msu051 phylogenetic data from the plastid genome of Cynomorium Moore, M.J., Hassan, N., Gitzendanner, M.A., Bruenn, R.A., Croley, or other extreme haustorial parasites (Naumann & al., 2013). M., Vandeventer, A., Horn, J.W., Dhingra, A., Brockington, In their study of angiosperm phylogeny using mitochon- S.F., Latvis, M., Ramdial, J., Alexandre, R., Peidrahita, A., Xi, Z., Davis, C.C., Soltis, P.S. & Soltis, D.E. 2011. Phylogenetic drial genes, Qiu & al. (2010) discuss but did not include Cyno- analysis of the plastid inverted repeat for 244 species: Insights into morium in their trees because it was placed in Saxifragales with deeper-level angiosperm relationships from a long, slowly evolving matR and nad5 but with Sapindales with atp1 and rps3. Again, sequence region. Int. J. Pl. Sci. 172: 541–558. it is well documented that holoparasite mitochondrial genes http://dx.doi.org/10.1086/658923 can derive from horizontal transfer, particularly with atp1 and Naumann, J., Salomo, K., Der, J.P., Wafula, E.K., Bolin, J.F., Maass, ribosomal protein genes (Molina & al., 2014). But interestingly, E., Frenzke, L., Samain, M.-S., Neinhuis, C., dePamphilis, C.W. & Wanke, S. 2013. Single-copy nuclear genes place hausto- the study by Naumann & al. (2013) also used mitochondrial rial Hydnoraceae within Piperales and reveal a Cretaceous origin atpA (= atp1 ?), coxI and matR. Using a relaxed molecular clock of multiple parasitic angiosperm lineages. PLoS ONE 8: e79204. approach to estimate the timing of origins for all angiosperm http://dx.doi.org/10.1371/journal.pone.0079204

S1 TAXON 64 (3) • June 2015 Electr. Suppl. to: Su & al. • Phylogenetic relationships of Santalales

Nickrent, D.L., Der, J.P. & Anderson, F.E. 2005. Discovery of the Takhtajan, A. 2009. Flowering plants, ed. 2. Springer Netherlands. photosynthetic relatives of the “Maltese mushroom” Cynomorium. http://dx.doi.org/10.1007/978-1-4020-9609-9 B. M. C. Evol. Biol. 5: 38. http://dx.doi.org/10.1186/1471-2148-5-38 Van Tieghem, P. 1896. Sur l’organisation florale des Balanophoracées et Qiu, Y., Li, L., Wang, B., Xue, J.-Y., Hendry, T.A., Li, R.-Q., Brown, sur la place de cette famille dans la sous-classe des Dicotylédones J.W., Liu Y., Hudson, G.T. & Chen, Z.-D. 2010. Angiosperm inovulées ou Loranthinées. Bull. Soc. Bot. France 43: 295–310. phylogeny inferred from sequences of four mitochondrial genes. http://dx.doi.org/10.1080/00378941.1896.10830679 J. Syst. Evol. 48: 391–425. Zhang, Z.-H., Li, C.-Q. & Li, J. 2009. Phylogenetic placement of http://dx.doi.org/10.1111/j.1759-6831.2010.00097.x Cynomorium in Rosales inferred from sequences of the inverted Takhtajan, A. 1997. Diversity and classification of flowering plants. repeat region of the chloroplast genome. J. Syst. Evol. 47: 297–304. New York: Columbia University Press. http://dx.doi.org/10.1111/j.1759-6831.2009.00035.x

S2 TAXON 64 (3) • June 2015 Electr. Suppl. to: Su & al. • Phylogenetic relationships of Santalales Individual Gene Trees

ML_SSU MP_SSU ML_RPB2 MP_RPB2 55 Viscaceae Viscaceae 66 Viscaceae 90 Viscaceae Amphorogynaceae Nanodeaceae 70 Amphorogynaceae 54 Amphorogynaceae Nanodeaceae Amphorogynaceae 69 Nanodeaceae Santalaceae Santalaceae Santalaceae Santalaceae Nanodeaceae 58 Cervantesiaceae Cervantesiaceae Thesieaceae Thesieaceae Comandraceae 72 75 Thesieaceae Opiliaceae Opiliaceae Thesieaceae Balanophoraceae A 68 Cervantesiaceae Cervantesiaceae Opiliaceae Comandraceae 76 Comandraceae 60 Loranthaceae* Comandraceae 58 Opiliaceae 72 Loranthaceae Balanophoraceae A 57 Misodendraceae 58 Loranthaceae* 73 Balanophoraceae B Misodendraceae 61 77 Schoepfiaceae Balanophoraceae B 70 Schoepfiaceae Balanophoraceae B 60 Balanophoraceae B Misodendraceae 78 Misodendraceae Schoepfiaceae Loranthaceae* Schoepfiaceae Balanophoraceae A Loranthaceae 62 59 Balanophoraceae A Loranthaceae* 79 Octoknemaceae Octoknemaceae Olacaceae Aptandraceae 78 Olacaceae Olacaceae 80 60 Olacaceae Aptandraceae Aptandraceae Ximeniaceae 62 Ximeniaceae Octoknemaceae 81 79 Ximeniaceae Coulaceae Octoknemaceae Ximeniaceae Coulaceae Erythropalaceae Erythropalaceae Erythropalaceae 87 Strombosiaceae Strombosiaceae Strombosiaceae Strombosiaceae Coulaceae Erythropalaceae Aptandraceae Coulaceae Outgroups Outgroups Outgroups Outgroups

ML_LSU MP_LSU ML_matR MP_matR Viscaceae Balanophoraceae A Nanodeaceae Nanodeaceae Amphorogynaceae Viscaceae Opiliaceae 69 67 Opiliaceae Cervantesiaceae Cervantesiaceae Thesiesiaceae Cervantesiaceae Comandraceae Thesieaceae Santalaceae 58 Thesieaceae Santalaceae Ximeniaceae Cervantesiaceae Amphorogynaceae Santalaceae Comandraceae Loranthaceae Santalaceae Loranthaceae Opiliaceae Schoepfiaceae Thesiaceae 64 Schoepfiaceae Misodendraceae 68 Loranthaceae Olacaceae 61 Misodendraceae Balanophoraceae B 88 Balanophoraceae B Loranthaceae Balanophoraceae A2 & B 80 Misodendraceae Misodendraceae Balanophoraceae A2 Balanophoraceae A1 Schoepfiaceae Schoepfiaceae 55 Olacaceae Olacaceae Balanophoraceae B Balanophoraceae A1 Coulaceae Balanophoraceae A Coulaceae Opiliaceae Ximeniaceae Ximeniaceae 54 Octoknemaceae Erythropalaceae Erythropalaceae Octoknemaceae Ximeniaceae 74 Olacaceae Outgroups* Aptandraceae Aptandraceae Aptandraceae Strombosiaceae Strombosiaceae Strombosiaceae Strombosiaceae Aptandraceae Erythropalaceae Erythropalaceae Outgroups Outgroups* Outgroups Outgroups

ML_rbcL MP_rbcL ML_accD MP_accD Amphorogynaceae Amphorogynaceae 84 57 Amphorogynaceae 95 Amphorogynaceae Viscaceae 100 Viscaceae Viscaceae Viscaceae Santalaceae Santalaceae 68 Santalaceae Santalaceae Nanodeaceae Nanodeaceae Nanodeaceae Nanodeaceae Cervantesiaceae Cervantesiaceae 79 Cervantesiaceae 60 Cervantesiaceae 54 57 Thesiaceae 77 Thesiaceae Opiliaceae Comandraceae Comandraceae Opiliaceae 87 98 Thesieaceae Opiliaceae Opiliaceae 99 Thesieaceae Comandraceae Loranthaceae Comandraceae 76 70 Loranthaceae 100 Loranthaceae 92 Loranthaceae 97 Schoepfiaceae 79 Schoepfiaceae 99 84 Schoepfiaceae Misodendraceae Misodendraceae 81 Schoepfiaceae 51 71 Misodendraceae 51 Olacaceae Olacaceae Misodendraceae Ximeniaceae Ximeniaceae Olacaceae Olacaceae Aptandraceae Aptandraceae Aptandraceae Aptandraceae Coulaceae Octoknemaceae Ximeniaceae 76 Ximeniaceae Octoknemaceae Coulaceae 95 Coulaceae Coulaceae Erythropalaceae Erythropalaceae Erythropalaceae Erythropalaceae 67 84 Strombosiaceae Strombosiaceae Strombosiaceae Strombosiaceae Outgroups Outgroups Outgroups Outgroups

ML_matK MP_matK

100 Amphorogynaceae 90 Amphorogynaceae Viscaceae 57 Viscaceae Santalaceae Cervantesiaceae 56 Nanodeaceae Nanodeaceae Cervantesiaceae Santalaceae 86 Thesieaceae 61 Thesieceae Comandraceae Comandraceae 100 100 Opiliaceae Opiliaceae Loranthaceae Loranthaceae 74 88 Misodendraceae 83 Schoepfiaceae Schoepfiaceae Misodendraceae Fig. S1. Trees obtained following maximum likelihood (ML) and Octoknemaceae Octoknemaceae Olacaceae Olacaceae maximum parsimony (MP) analyses of individual gene parti- 96 58 Aptandraceae 57 Aptandraceae tions. Bootstrap support greater than 50% are shown above the Ximeniaceae Ximeniaceae 72 nodes. Balanophoraceae clade A: fast-evolving lineages, of which Coulaceae 99 Coulaceae 99 Erythropalaceae Erythropalaceae A1 contains Balanophora and Thonningea and A2 conains all of Strombosiaceae Strombosiaceae the remaining. Balanophoraceae clade B: Mystropetalaceae. The Outgroups Outgroups groups that break into two branches are marked with an asterik.

S3 TAXON 64 (3) • June 2015 Electr. Suppl. to: Su & al. • Phylogenetic relationships of Santalales

59 LeptSpi 100 SpirRub 100 LeptAph 100 ChorPau 73 DendLed 100 DendVar 100 98 DufrSph Amphorogynaceae PhacRig 100 AmphCel DaenCor 99 100 PhorCal 100 PhorLeu 96 DendCla GinaArn 99 KortLin Viscaceae 88 ArceVer 88 NotoSub 100 ViscAlb ViscArt 100 SantAlb 100 81 SantMac AntiVis 85 EubrAmb LepiChi 100 ColpCom 80 RhoiCap Santalaceae 100 NestUmb 100 OsyrLan OsyrQua 100 MyosObl 89 ExocAph OmphAce ExocBid 79 100 MidaSal NanoMus Nanodeaceae 80 OkouAub 100 SclePen 99 StauCap 100 PilgMad PyruPub 100 89 AcanAsi 94 AcanFal Cervantesiaceae 100 CervTom JodiRho 100 ComaUmb 88 GeocLiv Comandraceae 67 KunkSub ThesFra 100 ThesChi 100 ThesFru 98 OsyrSch Thesiaceae BuckDis 87 CansLep 91 UrobSia 79 100 OpilAme 73 PentMar 95 ChamMan Opiliaceae 100 AgonMac AnthLep LepiSyl 100 OnceAmb 75 OncoSul AgelSan 100 EnglRam 57 TapiCon ActiMen BerhSen OediPen BakeSpe PlicSag 83 SocrBem VanwRem 67 EmelPan GlobDin 87 67 83 EriaDre OlivRub MoquRub PhraCra 50 DendLon 72 HeliCoc DendCur 97 ScurFer ScurPar 96 ScurPul 83 TaxiChi TaxiPse 100 HeliCyl 100 AmyeGla DiplFur 91 AmyeQue 59 BentAly DactNov SogeSes BaraAxa 100 100 IleoMic MuelEuc 98 LoraDel 98 LoraKao 100 LoraEur 98 CecaObt Loranthaceae TupeAnt DesmMut NotaHet GaiaPun 97 LepiFor 84 MacrCoc 59 DecaTri 97 LysiFil 73 94 AmylDut LoxaSpe 72 LepeLan 100 AlepFla PeraTet 100 DendBic 81 OrycOcc PassPyr 98 AetaNod 62 PsitCal 100 81 StruOer 100 100 StruWoo CladGra 100 TripAcu 74 LigaCun TrisCor AtkiLig NuytFlo 100 CoryCra 100 HeloCay 100 LophLea 100 OmbrSub 73 100 BalaFun 100 BalaLax Balanophoraceae A 68 ThonSan SarcSan 100 DactTay 100 HachAus Balanophoraceae B 64 MystTho 100 SchoChi 100 SchoJas 93 SchoSch 100 ArjoTub Schoepfiaceae 54 QuinChi 100 MisoLin MisoPun Misodendraceae OctoSpe 91 CoulEdu Octoknemaceae 83 100 MinqGui 52 OchaAme Coulaceae 100 CuruTef 100 XimeAme MalaOle Ximeniaceae 100 AptaTub 100 OngoGor 88 HarmMek 83 HondUrc 100 ChauKap Aptandraceae 97 AnacPap 100 PhanCap CathAcu 62 DulaCan 67 OlaxEmi 100 OlaxImb 100 100 OlaxAph Olacaceae OlaxBen PtycPet 80 HeisCau 57 91 HeisPar 99 HeisCon 100 HeisDen 90 HeisAcu Erythropalaceae 100 MabuTri ErytSca DiogZen 90 EngoGor StroTet 100 TetrPer 77 StroGra Strombosiaceae 100 100 StroPhi StroPus ScorBor 94 AntiMaj 81 SpinOle 82 CornFlo CameJap Outgroups 96 ArabTha MyrtCom

Fig. S2. The strict consensus of 6912 MP trees inferred from the concatenated 7-gene dataset. Bootstrap support values greater than 50% are shown above the nodes. Tree length = 28,736, consistency index = 0.380, retention index = 0.641.

S4 TAXON 64 (3) • June 2015 Electr. Suppl. to: Su & al. • Phylogenetic relationships of Santalales

LeptAph LeptSpi SpirRub ChorPau DendLed DendVar Amphorogynaceae DufrSph PhacRig AmphCel DaenCor PhorCal PhorLeu DendCla GinaArn KortLin Viscaceae ArceVer ViscAlb ViscArt NotoSub SantAlb SantMac AntiVis EubrAmb LepiChi MyosObl ColpCom Santalaceae RhoiCap NestUmb OsyrLan OsyrQua ExocAph OmphAce ExocBid MidaSal NanoMus Nanodeaceae OkouAub SclePen StauCap PilgMad PyruPub Cervantesiaceae AcanAsi AcanFal CervTom JodiRho 89/1.0/- KunkSub ThesFra ThesChi ThesFru Thesiaceae OsyrSch BuckDis ComaUmb GeocLiv Comandraceae CansLep UrobSia OpilAme PentMar ChamMan AgonMac Opiliaceae AnthLep LepiSyl OnceAmb OncoSul AgelSan ActiMen BerhSen EnglRam TapiCon OediPen EmelPan GlobDin EriaDre OlivRub MoquRub PhraCra BakeSpe PlicSag SocrBem VanwRem DendLon HeliCoc DendCur ScurPar ScurPul ScurFer TaxiChi TaxiPse HeliCyl AmyeGla DiplFur DactNov AmyeQue BentAly SogeSes BaraAxa IleoMic MuelEuc Loranthaceae LoraDel LoraKao LoraEur CecaObt LepiFor MacrCoc DecaTri AmylDut LoxaSpe LysiFil LepeLan AlepFla PeraTet TupeAnt DesmMut StruOer StruWoo CladGra DendBic OrycOcc PassPyr AetaNod PsitCal TripAcu NotaHet LigaCun TrisCor GaiaPun AtkiLig NuytFlo DactTay HachAus MystTho Balanophoraceae B SchoChi SchoJas SchoSch Schoepfiaceae ArjoTub QuinChi MisoLin MisoPun Misodendraceae OctoSpe Octoknemaceae AptaTub OngoGor HarmMek HondUrc ChauKap Aptandraceae AnacPap PhanCap CathAcu DulaCan OlaxEmi OlaxImb OlaxAph Olacaceae OlaxBen PtycPet CuruTef XimeAme Ximeniaceae MalaOle CoulEdu MinqGui OchaAme HeisCau Coulaceae HeisPar HeisCon HeisDen HeisAcu MabuTri Erythropalaceae ErytSca StroTet TetrPer EngoGor DiogZen StroGra Strombosiaceae StroPhi StroPus ScorBor AntiMaj CornFlo CameJap ArabTha Outgroups MyrtCom SpinOle

0.01 substitutions/site Fig. S. The ML tree inferred from the concatenated 7-gene dataset that excluded Balanophoraceae clade A (fast-evolving). Bootstrap support greater than 80% and Bayesian posterior probabilities greater than 0.95 are shown with bold lines.

S5 TAXON 64 (3) • June 2015 Electr. Suppl. to: Su & al. • Phylogenetic relationships of Santalales

57 100 LeptSpi 100 SpirRub LeptAph 100 ChorPau 74 DendLed 100 DendVar 100 99 DufrSph Amphorogynaceae PhacRig AmphCel DaenCor 100 100 PhorCal 100 PhorLeu 96 DendCla GinaArn Viscaceae 100 KortLin 88 ArceVer 99 NotoSub ViscAlb ViscArt 100 SantAlb 81 SantMac AntiVis 86 EubrAmb LepiChi 100 ColpCom 80 RhoiCap 50 100 NestUmb Santalaceae 100 OsyrLan 100 OsyrQua MyosObl 88 ExocAph 93 OmphAce ExocBid 100 MidaSal NanoMus Nanodeaceae 100 ComaUmb GeocLiv Comandraceae 87 81 OkouAub 100 99 SclePen 100 StauCap PilgMad PyruPub 100 87 93 AcanAsi Cervantesiaceae 100 AcanFal CervTom JodiRho 68 KunkSub 100 99 ThesFra 100 ThesChi ThesFru Thesiaceae 100 OsyrSch BuckDis 88 CansLep 91 UrobSia 77 99 OpilAme 73 PentMar Opiliaceae 95 ChamMan 100 AgonMac AnthLep LepiSyl 100 OnceAmb 77 OncoSul AgelSan 100 EnglRam 59 TapiCon ActiMen BerhSen OediPen BakeSpe PlicSag 83 SocrBem VanwRem 63 EmelPan GlobDin 88 69 83 EriaDre OlivRub MoquRub PhraCra DendLon 72 HeliCoc 57 DendCur 65 ScurFer ScurPar 97 ScurPul 82 TaxiChi TaxiPse 100 HeliCyl 100 AmyeGla DiplFur 92 AmyeQue 58 BentAly DactNov SogeSes BaraAxa 100 100 100 IleoMic Loranthaceae MuelEuc 98 LoraDel 98 LoraKao 100 LoraEur CecaObt 96 LepiFor 86 MacrCoc 61 DecaTri 98 LysiFil 73 95 AmylDut LoxaSpe 73 LepeLan 100 AlepFla PeraTet 100 100 DendBic 80 OrycOcc Pass Pyr 97 AetaNod 61 PsitCal 80 StruOer 100 100 StruWoo CladGra 100 TripAcu 72 LigaCun TrisCor AtkiLig GaiaPun TupeAnt 58 DesmMut 100 NotaHet NuytFlo 100 SchoChi 100 SchoJas 95 SchoSch 100 ArjoTub Schoepfiaceae 65 QuinChi 100 MisoLin 96 MisoPun Misodendraceae 100 100 DactTay HachAus MystTho Balanophoraceae B OctoSpe 92 CoulEdu Octoknemaceae 100 MinqGui 84 53 OchaAme Coulaceae 100 CuruTef 100 XimeAme MalaOle Ximeniaceae 100 AptaTub 100 OngoGor 88 HarmMek 83 HondUrc 100 ChauKap 97 AnacPap 100 PhanCap Aptandraceae 58 CathAcu 68 DulaCan OlaxEmi 100 OlaxImb 100 100 OlaxAph Olacaceae OlaxBen PtycPet 80 HeisCau 58 HeisPar 90 100 100 HeisCon 92 HeisDen 86 HeisAcu Erythropalaceae MabuTri ErytSca DiogZen 91 EngoGor StroTet 100 TetrPer 100 77 StroGra 100 StroPhi Strombosiaceae StroPus ScorBor 94 AntiMaj 78 SpinOle 80 CornFlo CameJap Outgroups 97 ArabTha MyrtCom

Fig. S4. The strict consensus of 15,552 MP trees inferred from the concatenated 7-gene dataset that excluded Balanophoraceae clade A (fast- evolving). Bootstrap support greater than 50% are shown above the nodes. Tree length = 25,219, consistency index = 0.384, retention index = 0.657.

S6 TAXON 64 (3) • June 2015 Electr. Suppl. to: Su & al. • Phylogenetic relationships of Santalales

Nonplastid Genes With & Without 3rd Codons

MP strict With 3rd codons 3rd codons removed Amphorogynaceae Viscaceae Nanodeaceae 52 Santalaceae Thesiaceae 70 Cervantesiaceae Comandraceae 84 98 Opiliaceae Loranthaceae 74 65 62 Misodendraceae 92 52 Schoepfiaceae 51 Balanophoraceae B Balanophoraceae A Olacaceae Coulaceae Ximeniaceae Octoknemaceae 79 Aptandraceae 66 Erythropalaceae Strombosiaceae Outgroups

ML With 3rd codons 3rd codons removed 84 Amphorogynaceae 54 Viscaceae Santalaceae 52 Nanodeaceae 55 98 98 Thesiaceae 91 Cervantesiaceae 100 Comandraceae 74 Opiliaceae 89 Loranthaceae 70 98 Balanophoraceae B 97 89 100 Schoepfiaceae 99 59 Misodendraceae Balanophoraceae A Olacaceae Coulaceae Ximeniaceae Octoknemaceae 54 Aptandraceae 54 Erythropalaceae Strombosiaceae Outgroups

Fig. S5. Santalales tree topologies using the four-non-plastid gene partition: SSU and LSU rDNA, RPB2, and matR. Trees on the left included third codons whereas trees on the right excluded third codons from RPB2 and matR. Strict consensus maximum parsimony trees (MP) shown on the top, maximum likelihood trees (ML) on the bottom. Support values greater than 50% are shown above the nodes.

S7 TAXON 64 (3) • June 2015 Electr. Suppl. to: Su & al. • Phylogenetic relationships of Santalales

100 AmphCel DaenCor 100 52 DufrSph Fig. S. The ML tree inferred from the concatenated 7-gene dataset 9840 DendLed PhacRig 39 DendVar that excluded fast-evolving sites. Bootstrap supports are shown above 99 ChorPau Amphorogynaceae 99 100 LeptAph 47 LeptSpi the branches. The fast-evolving sites were identified by using TIGER SpirRub NotoSub 100 100 ViscAlb (Cummins CA, McInerney JO (2011) Syst Biol 60: 833–844), which ViscArt 97 93 ArceVer Viscaceae 100 DendCla removed 2%–30% sites in the fastest-evolving category from each of 94 100 PhorLeu PhorCal 79 100 GinaArn the individual dataset. KortLin 100 OsyrQua 100 100 OsyrLan 89 RhoiCap ColpCom 83 100NestUmb 95 SantAlb 68 SantMac Santalaceae 51 AntiVis 100 69 99 LepiChi EubrAmb MyosObl 100 ExocBid 94 ExocAph 27 OmphAce 100 MidaSal NanoMus Nanodeaceae 96 BuckDis 97 76 OsyrSch 44 ThesFra 97 KunkSub ThesFru Thesiaceae 37 ThesChi 100 ComaUmb GeocLiv 52 SclePen Comandraceae 57 100 95 84 StauCap 100 OkouAub PilgMad 100 84 PyruPub 98 AcanAsi 100 AcanFal Cervantesiaceae CervTom JodiRho 99 OpilAme 83 PentMar 7991 UrobSia 76 CansLep Opiliaceae 97 ChamMan 100 AgonMac AnthLep LepiSyl100 90 OncoSul 41 OnceAmb 32 AgelSan BerhSen 7398 TapiCon 81 EnglRam 74 ActiMen OediPen 89 SocrBem 25 VanwRem 74 BakeSpe 95 PlicSag 26 EriaDre 33 OlivRub 866991 EmelPan 94 GlobDin PhraCra 44 MoquRub 7388 DendCur DendLon 40 HeliCoc 91 ScurPar 54 ScurPul 97 ScurFer 58 TaxiPse TaxiChi 29 100 HeliCyl 85 100DactNov 74 AmyeGla DiplFur 67 6759 AmyeQue 38 BentAly SogeSes BaraAxa 100 100 IleoMic 100MuelEuc Loranthaceae 100 LoraDel 100 LoraKao LoraEur CecaObt 21 96 TripAcu 100100 StruWoo 20 StruOer 98 CladGra 45 AetaNod 97 PsitCal 40 49 DendBic OrycOcc 45 PhthPyr 4937 NotaHet 35 TrisCor LigaCun 16 58 DesmMut 37 MacrCoc 20 LepiFor 99 DecaTri 54 LysiFil 95 84 AmylDut LoxaSpe 75100 94 AlepFla PeraTet 94 TupeAnt 93 LepeLan GaiaPun 57 AtkiLig NuytFlo 100 100 MystTho 50 DactTay 6 HachAus Balanophoraceae B 100 MisoLin 100 MisoPun Misodendraceae 97 100 SchoChi 100 SchoJas 100 SchoSch Schoepfiaceae ArjoTub QuinChi 100 ThonSan 3 16 100 BalaFun BalaLax 20 SarcSan 100 CoryCra 100 HeloCay 100 LophLea Balanophoraceae A OmbrSub 58 OctoSpe Octoknemaceae 15100 CoulEdu MinqGui Coulaceae OchaAme 100 OlaxBen 100 86 OlaxAph 48 DulaCan Olacaceae 100 OlaxEmi OlaxImb 79 100PtycPet 10 100 AptaTub 92 OngoGor 94 HarmMek HondUrc Aptandraceae 100 ChauKap 100 CathAcu 97 PhanCap AnacPap 67 100 100 MalaOle CuruTef Ximeniaceae 49 XimeAme 18 HeisAcu 23 HeisCon 100 HeisCau 75 HeisPar 97100 HeisDen Erythropalaceae MabuTri ErytSca 57 StroTet 9859 DiogZen TetrPer 97 Strombosiaceae 92 EngoGor 100 100 StroGra StroPhi StroPus ScorBor 71 SpinOle 78 CameJap 97 58 CornFlo AntiMaj Outgroups 100 ArabTha MyrtCom

0.06 S8 TAXON 64 (3) • June 2015 Electr. Suppl. to: Su & al. • Phylogenetic relationships of Santalales

2 8 NotoSub 8 9 ViscArt Fig. S. The ML tree inferred from amino acid sequences of the five 7 5 ViscAlb 8 8 KortLin Viscaceae 4 4 GinaArn protein-codon genes (matR, RPB2, matK, rbcL, accD). The tree was 7 ArceVer 9 9 6 7 DendCla 5 5 PhorCal consructed by RAxML v.8.1.7 and the best protein model was auto- PhorLeu 100 AmphCel DaenCor 9 3 matically determined directly by the program. Bootstrap supports 9 6 DendVa5 3 r 4 2 PhacRig 2 5 8 6 DufrSph computed by 500 replicates are shown above the branches. DendLed 9 3 ChorPau Amphorogynaceae 9 4 LeptAph 8 2 LeptSpi 100 SpirRub 2 6 MidaSal NanoMus Nanodeaceae MyosObl 3 7 7 8 6 6 EubrAmb 2 3 LepiChi AntiVis 100 SantMac 7 8 SantAlb NestUmb 9 6 9 6 Santalaceae 5 2 OsyrLan 100 OsyrQua 5 6 2 4 ColpCom RhoiCa7 6 p 9 6 ExocAph OmphAce ExocBid 3 9 ThesChi 7 3 BuckDis 100 5 4 OsyrSch 9 7 KunkSub Thesiaceae ThesFra ThesFru 2 8 8 2 5 0 OkouAub 6 8 8 8 StauCap 100 SclePen PilgMad 100 PyruPub 100 JodiRho Cervantesiaceae 4 3 7 9 CervTom 6 9 AcanAsi AcanFal 9 8 ComaUmb GeocLiv Comandraceae 7 4 3 9 OpilAme 4 2 PentMar 100 CansLep UrobSia 1 7 ChamMan Opiliaceae 2 3 AgonMac 1 0 LepiSyl AnthLep 9 0 EmelPan 4 0 GlobDin 6 2 MoquRub PhraCra 9 2 EriaDre 5 8 OlivRub 7 3 DendLon 4 9 HeliCoc 3406 DendCur 1 5 BakeSpe VanwRem 2 6 PlicSag 1 6 SocrBem 3 7 OediPen 5 86 6 ActiMen 5 2 BerhSen 9 7 EnglRam 2 6 TapiCon 7 4 AgelSan 9 9 OnceAmb OncoSul 9 1 100 TaxiChi 100 ScurFer 3 8 6 3 ScurPar ScurPul 2 9 4 8 HeliCyl BaraAxa 6 6 5 4 SogeSes AmyeQue 3991 DactNov 6 2 100 AmyeGla DiplFur BentAly 1 4 9 9 IleoMic Loranthaceae 2 4 8 8 MuelEuc 9 2 LoraDel 5 2 LoraKao 1 LoraEur CecaObt GaiaPun 7 8 LigaCun 33 6 TrisCor 3 3 4 3 AetaNod 1 NotaHet DesmMut TaxiPse AtkiLig 7 PsitCal 9 9 100 100 CladGra 5 2 StruOer 2 2 StruWoo 3 5 5 4 TripAcu 9 5 9 9 PhthPyr 9 5 OrycOcc DendBic 8 2 LepeLan TupeAnt 9 3 PeraTet 9 8 3 9 AlepFla 8 0 LoxaSpe 2 4 AmylDut 6 79 1 LysiFil 2 3 8 5 MacrCoc LepiFor DecaTri NuytFlo 100 100 HachAus 9 DactTay 4 3 MystTho Balanophoraceae B 100 SchoSch 100 100 SchoChi 100 SchoJas Schoepfiaceae 7 5 ArjoTub QuinChi 100 MisoPun MisoLin Misodendraceae OctoSpe Octoknemaceae 100 100 CoryCra 9 8 HeloCay 5 6 LophLea Balanophoraceae A2 100 OmbrSub 9 9 XimeAme 2 6 CuruTef 4 1 MalaOle Ximeniaceae 9 7 CathAcu 5 5 AnacPap 9 6 PhanCap 6 9 HondUr100 c 100 AptaTub 5 2 OngoGor Aptandraceae HarmMek ChauKap 8 6 ThonSan 4 6 100 BalaLax 2 5 BalaFun 8 9 PtycPet 100 9O9 laxImb 1 5 OlaxAph 5 5 OlaxBen Olacaceae 1 6 7 DulaCan Balanophoraceae A OlaxEmi 9 0 OchaAme 6 4 CoulEdu Coulaceae MinqGui 8 8 SarcSan 3 3 9 ScorBor Balanophoraceae A 8 5 4 37 8 EngoGor 7 8 StroGra 9 6 StroPhi StroPus 5 8 Strombosiaceae 4 5 TetrPer DiogZen StroTet MabuTri HeisAcu 7 7 100 7 5 7 1 HeisPar 5 6 6 6 HeisCau HeisDen Erythropalaceae HeisCon ErytSca 5 8 SpinOle 5 1 CameJap 8 8 4 7 CornFlo AntiMaj Outgroups 100 ArabTha MyrtCom

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