Supplemental Information for: Introgression across evolutionary scales suggests reticulation contributes to Amazonian tree diversity Rowan J. Schley, R. Toby Pennington, Oscar Alejandro Pérez-Escobar, Andrew J. Helmstetter, Manuel de la Estrella, Isabel Larridon, Izai Alberto Bruno Sabino Kikuchi, Timothy Barraclough, Félix Forest, and Bente Klitgård Table of Contents: Fig. S1 Page 2 Fig. S2 Page 3 Fig. S3 Page 4 Fig. S4 Page 5 Fig. S5 Page 8 Table S1 Page 10 Table S2 Page 13 Table S3 Page 18 Methods S1 Page 19 Methods S2 Page 23 1 Fig. S1 PhyParts analysis based on the ASTRAL species tree, inferred from RAxML gene trees using the multi-species coalescent model. Pie charts at nodes show the number of gene trees with a congruent topology at a node (blue segments), the number which support a single alternate topology (green segments), the number which support all other conflicting bipartitions (red segments) and how many gene trees were uninformative at the node (grey segments). Numbers above branches show how many gene trees were congruent at a node, whereas numbers below branches show the number of conflicting gene trees at a node. 2 Fig. S2 Negative log pseudolikelihood profile for each number of hybridization events inferred using SNaQ!. The best-fitting number of hybridization events (h) is displayed as the value at which the rate of change in -log pseudolikelihood plateaus, which in this case was h=2. 3 Fig. S3 SPLITSTREE built using uncorrelated P distances based on the full dataset for all genotyped individuals. The three coloured zones represent groupings of the two species (B. jaramilloi and B. grandiceps) as well as hybrid individuals. Groupings in the coloured zones were defined based on the clustering apparent in this plot, not on the species delimitations present in Table S4. 4 Fig. S4 A) Difference in marginal likelihoods between sequential numbers of population clusters (K), as estimated by FastSTRUCTURE. The best value of K results in the largest increase in marginal likelihood (in this case, K=2). B) fastSTRUCTURE plots generated using different values of estimated clusters (K=3 to K=5). This plot indicates ancestry proportions from K inferred population clusters. Each individual accession is represented by a column, and the proportion of ancestry from each inferred population is proportional to the length of different coloured bars in each column. C) fastSTRUCTURE plot incorporating 40 individuals from each of the two species under study in order to reduce error associated with different sample sizes. This plot indicates ancestry proportions from K = 2 population clusters (which are, in this case, different species: B. grandiceps and B. jaramilloi). Each individual accession is represented by a column, and the proportion of ancestry from either species is proportional to the length of different coloured bars in each column. A) 5 B) 6 C) 7 Fig. S5 Assessment of MCMC chain mixing for runs used to make posterior estimates for the α and β parameters in bgc. A) Log-likelihood plot, showing log-likelihood plotted against number of MCMC iterations. B) Histogram of Geweke’s diagnostics for the MCMC chains used to estimate the α parameter, with the significance cut-offs for the Geweke’s diagnostic (Z = +/- 1.96) shown by blue dashed lines. C) Histogram of Geweke’s diagnostics for the MCMC chains used to estimate the β parameter, with the significance cut-offs for the Geweke’s diagnostic (Z = +/- 1.96) shown by blue dashed lines. 8 9 Table S1 DNA numbers, species identifications, sampled vouchers, collector information and collection localities for Brownea species (and Brownea clade outgroup species) used in this study. Herbaria from which accessions were collected are cited after the collector name and number. Outgroup taxa are marked with an asterisk (*). DNA Identification Voucher Collection locality Number Iganci 886 S1 Macrolobium discolor Benth.* Brazil (MO) Edwards 328 S2 Brownea grandiceps Jacq. Aragua, Venezuela (K) Macrolobium montanum var. Andel 5548 S3 Suriname potaroanum R.S. Cowan* (K) Macrolobium colombianum (Britton & Klitgaard 682 S4 Napo, Ecuador Killip) Killip ex L. Uribe* (K) Klitgaard 99504 S5 Brownea grandiceps Jacq. Napo, Ecuador (AAU) Iganci 875 S6 Heterostemon mimosoides Desf.* Brazil (MO) Klitgaard 662 S7 Brownea jaramilloi A.J. Pérez & Klitg. Yasuní, Napo, Ecuador (K) Daly 6167 S8 Brownea grandiceps Jacq. Rio Marañon, Loreto, Peru (NY) Edwards 372 S18 Brownea coccinea Jacq. Aragua, Venezuela (K) Brownea coccinea subsp. coccinea Steyermark 126503 S19 Morrocoy, Venezuela Jacq. (K) Brownea coccinea subsp. capitella Clark 6062 S20 Cuyuni-Mazaruni, Guyana (Jacq.) D. Velazquez (K) Pennington 15642 S21 Brownea multijuga Britton & Killip Pichincha, Ecuador (K) Trujillo 19314 S22 Brownea coccinea Jacq. Falcon, Venezuela (K) Brownea coccinea subsp. capitella Gutierrez S23 Venezuela (Jacq.) D. Velazquez (K) Marshall 392 S24 Macrolobium herrerae Zarucchi* Izabal, Guatemala (K) Macrolobium parvifolium (Huber) R.S. de Lima 6814 S25 Acre, Brazil Cowan* (K) Macrolobium longipedicellatum de Lima 2790 S26 Amazonas, Brazil Ducke* (K) Martins 54 S27 Paloue speciosa (Ducke) Redden* Amazonas, Brazil (K) Paloue princeps (Schomb. Ex Benth.) Sabatier S28 French Guiana Redden* (K) Klitgaard 684 S29 Browneopsis ucayalina Huber* Morona-Santiago, Ecuador (K) Klitgaard Rio de Janeiro Botanical Garden, S30 Brownea longipedicellata Huber (Silica) Brazil 10 Villa 1645 S31 Brownea jaramilloi A.J. Pérez & Klitg. Yasuní, Napo, Ecuador (MO) Valencia 68473 S32 Brownea ariza Benth. Sucumbios, Ecuador (AAU) Browneopsis peruviana (J.F. Macbr.) Castro MHT-01_2 S33 Peru Klitg.* (Silica) Baker HD1402 S34 Brownea grandiceps Jacq. Ecuador (Silica) Brownea coccinea subsp. capitella Hollowell 485 Hossororo, Barima-Waini, S35 (Jacq.) D. Velazquez (US) Guyana Klitgaard 67032 S36 Browneopsis disepala (Little) Klitg.* Rio Palenque, Los Rios, Ecuador (AAU) Villa 1855 S37 Brownea grandiceps Jacq. Yasuní, Napo, Ecuador (Silica) Villa 747 S38 Brownea "rosada" Jacq. Yasuní, Napo, Ecuador (MO) Villa 1866 S39 Brownea grandiceps Jacq. Yasuní, Napo, Ecuador (Silica) Villa 1859 S40 Brownea grandiceps Jacq. Yasuní, Napo, Ecuador (Silica) Villa 1868 S41 Brownea "rosada" Jacq. Yasuní, Napo, Ecuador (Silica) Villa 1609 S42 Brownea jaramilloi A.J. Pérez & Klitg. Yasuní, Napo, Ecuador (MO) Killip 34351 S43 Brownea grandiceps Jacq. Meta, Colombia (MO) Brownea coccinea subsp. capitella Maguire 46987 S44 El Foco, Bolivar, Venezuela (Jacq.) D. Velazquez (NY) Brownea coccinea subsp. capitella Gentry 14773 S45 Los Caracas, Venezuela (Jacq.) D. Velazquez (NY) Brownea coccinea subsp. coccinea Gonzalez 29 S46 Churuguara-coro, Venezuela Jacq. (US) Villa 2004 Joya de los Sachas, Succumbios, S47 Brownea macrophylla hort. Ex Mast. (K) Ecuador Brownea coccinea subsp. angustiflora Klitgaard 99488 S48 Esmeraldas, Ecuador (Little) Klitg. (AAU) Polak 44 S50 Brownea latifolia Jacq. Guyana (K) Hoffman 2656 S51 Brownea latifolia Jacq. Pomeroon-Supenaam, Guyana (US) Brownea coccinea subsp. angustiflora Klitgaard 67044 S52 Esmeraldas, Ecuador (Little) Klitg. (AAU) Philcox 8078 S53 Brownea latifolia Jacq. Trinidad (K) Dorr 7792 S57 Brownea grandiceps Jacq. Barinas, Venezuela (NY) Brownea tillettiana D. Velásquez & G. Davidse 18640 S60 Perija, Venezuela Agostini (NY) Garcia-Barriga 17222 S64 Brownea enricii L.M. Quiñones Boyaca, Colombia (US) Gentry 15470 S65 Brownea coccinea Jacq. Cimitarra, Santander, Colombia (NY) 11 Callejas 3209 S66 Brownea rosa-de-monte P.J. Bergius Antioquia, Colombia (NY) Duke 11126 (3) S67 Brownea rosa-de-monte P.J. Bergius Rio Truando, Choco, Colombia (NY) Callejas 4503 S68 Brownea santanderensis L.M. Quiñones Antioquia, Colombia (NY) Stern 180 S71 Brownea stenantha Britton & Killip Paya, Darien, Panama (US) Berry 5413 S72 Brownea multijuga Britton & Killip Darien, Panama (NY) Klitgaard 1802 S74 Brownea macrophylla hort. ex. Mast. Boyaca, Colombia (K) Birschel s.n. S76 Brownea birschelli Hook. F. Rio de La Guaira, Venezuela (K) Schunke 0011 S77 Brownea grandiceps Jacq. (USM 14269) Maynas, Peru (K) Pennell 4698 S79 Brownea stenantha Britton & Killip Rio Sinu, Bolivar, Colombia (K) Little 6281 S80 Brownea puberula Little Esmeraldas, Ecuador (K) Killip 37047 S81 Brownea leucantha Jacq. Miranda, Venezuela (K) Brownea cf. santanderensis L.M. Klitgaard 1800 S87 Boyaca, Colombia Quiñones (K) Browneopsis cauliflora (Poepp.) Klug 4163 S88 San Martin, Peru Huber* (K) Haught 4788 S89 Brownea stenantha Britton & Killip Antioquia, Colombia (K) Hanbury-Tracy 509 Sierra de Santa Marta, S90 Brownea ariza Benth. (K) Magdalena, Colombia Allen 931 S91 Brownea grandiceps Jacq. Panama province, Panama (K) Dawe 883 S92 Brownea multijuga Britton & Killip Darien, Colombia (K) Fendler 340 Colonia Tovar, Aragua, S93 Brownea grandiceps Jacq. (K) Venezuela Grandez 1625 S94 Brownea grandiceps Jacq. Loreto, Peru (K) Fonnegra 8088 S96 Brownea santanderensis L.M. Quiñones Santander, Colombia (K) Castellanos 448 S97 Brownea santanderensis L.M. Quiñones Santander, Colombia (K) Whitmore 885 S100 Brownea grandiceps Jacq. Napo/Pastaza, Ecuador (K) Edwards 327 S101 Brownea grandiceps Jacq. Aragua, Venezuela (K) Diaz 4228 S103 Browneopsis ucayalina Huber* Amazonas, Peru (K) Fuchs 22384 S104 Brownea chocoana (L.M. Quiñones) Choco, Colombia (K) 12 Table S2 DNA numbers, species, coordinates for subplots, tree tag and habitat type for individuals of B. grandiceps, B. jaramilloi and B. “rosada” collected from the 50-hectare forest plot in Yasuní National Park, Ecuador. The ‘Subplot X’ and ‘Subplot