A peer-reviewed open-access journal PhytoKeys 87: Molecular1–139 (2017) phylogenetics of cool-season grasses in the subtribes Agrostidinae... 1 doi: 10.3897/phytokeys.87.12774 RESEARCH ARTICLE http://phytokeys.pensoft.net Launched to accelerate biodiversity research Molecular phylogenetics of cool-season grasses in the subtribes Agrostidinae, Anthoxanthinae, Aveninae, Brizinae, Calothecinae, Koeleriinae and Phalaridinae (Poaceae, Pooideae, Poeae, Poeae chloroplast group 1) Jeffery M. Saarela1, Roger D. Bull1, Michel J. Paradis1, Sharon N. Ebata1, Paul M. Peterson2, Robert J. Soreng2, Beata Paszko3 1 Botany Section, Research and Collections, Canadian Museum of Nature, Ottawa, Ontario, Canada 2 De- partment of Botany, National Museum of Natural History, Smithsonian Institution, Washington, DC, United States of America 3 Department of Vascular Plant Systematics and Phytogeography, W. Szafer Institute of Bo- tany, Polish Academy of Sciences, Kraków, Poland Corresponding author: Jeffery M. Saarela ([email protected]) Academic editor: C. Morden | Received 17 March 2017 | Accepted 4 August 2017 | Published 9 October 2017 Citation: Saarela JM, Bull RD, Paradis MJ, Ebata SN, Peterson PM, Soreng RJ, Paszko B (2017) Molecular phylogenetics of cool-season grasses in the subtribes Agrostidinae, Anthoxanthinae, Aveninae, Brizinae, Calothecinae, Koeleriinae and Phalaridinae (Poaceae, Pooideae, Poeae, Poeae chloroplast group 1). PhytoKeys 87: 1–139. https://doi.org/10.3897/ phytokeys.87.12774 Abstract Circumscriptions of and relationships among many genera and suprageneric taxa of the diverse grass tribe Poeae remain controversial. In an attempt to clarify these, we conducted phylogenetic analyses of >2400 new DNA sequences from two nuclear ribosomal regions (ITS, including internal transcribed spacers 1 and 2 and the 5.8S gene, and the 3’-end of the external transcribed spacer (ETS)) and five plastid regions (matK, trnL–trnF, atpF–atpH, psbK–psbI, psbA–rps19–trnH), and of more than 1000 new and previously published ITS sequences, focused particularly on Poeae chloroplast group 1 and including broad and in- creased species sampling compared to previous studies. Deep branches in the combined plastid and com- bined ITS+ETS trees are generally well resolved, the trees are congruent in most aspects, branch support across the trees is stronger than in trees based on only ITS and fewer plastid regions, and there is evidence of conflict between data partitions in some taxa. In plastid trees, a strongly supported clade corresponds to Poeae chloroplast group 1 and includes Agrostidinae p.p., Anthoxanthinae, Aveninae s.str., Brizinae, Koeleriinae (sometimes included in Aveninae s.l.), Phalaridinae and Torreyochloinae. In the ITS+ETS tree, a supported clade includes these same tribes as well as Sesleriinae and Scolochloinae. Aveninae s.str. Copyright Jeffery M. Saarela et al. This is an open access article distributed under the terms of the Creative Commons Attribution License (CC BY 4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. 2 Jeffery M. Saarela et al. / PhytoKeys 87: 1–139 (2017) and Sesleriinae are sister taxa and form a clade with Koeleriinae in the ITS+ETS tree whereas Aveninae s.str. and Koeleriinae form a clade and Sesleriinae is part of Poeae chloroplast group 2 in the plastid tree. All species of Trisetum are part of Koeleriinae, but the genus is polyphyletic. Koeleriinae is divided into two major subclades: one comprises Avellinia, Gaudinia, Koeleria, Rostraria, Trisetaria and Trisetum subg. Trisetum, and the other Calamagrostis/Deyeuxia p.p. (multiple species from Mexico to South America), Peyritschia, Leptophyllochloa, Sphenopholis, Trisetopsis and Trisetum subg. Deschampsioidea. Graphephorum, Trisetum cernuum, T. irazuense and T. macbridei fall in different clades of Koeleriinae in plastid vs. nuclear ribosomal trees, and are likely of hybrid origin. ITS and matK trees identify a third lineage of Koeleriinae corresponding to Trisetum subsect. Sibirica, and affinities ofLagurus ovatus with respect to Aveninae s.str. and Koeleriinae are incongruent in nuclear ribosomal and plastid trees, supporting recognition of Lagurus in its own subtribe. A large clade comprises taxa of Agrostidinae, Brizinae and Calothecinae, but neither Agrostidinae nor Calothecinae are monophyletic as currently circumscribed and affinities of Brizinae differ in plastid and nuclear ribosomal trees. Within this clade, one newly identified lineage comprises Calamagrostis coarctata, Dichelachne, Echinopogon (Agrostidinae p.p.) and Relchela (Calothecinae p.p.), and another comprises Chascolytrum (Calothecinae p.p.) and Deyeuxia effusa (Agrostidinae p.p.). Within Agrostidinae p.p., the type species of Deyeuxia and Calamagrostis s.str. are closely related, supporting clas- sification of Deyeuxia as a synonym of Calamagrostis s.str. Furthermore, the two species of Ammophila are not sister taxa and are nested among different groups of Calamagrostis s.str., supporting their classifica- tion in Calamagrostis. Agrostis, Lachnagrostis and Polypogon form a clade and species of each are variously intermixed in plastid and nuclear ribosomal trees. Additionally, all but one species from South America classified inDeyeuxia sect. Stylagrostis resolve in Holcinae p.p. (Deschampsia). The current phylogenetic results support recognition of the latter species in Deschampsia, and we also demonstrate Scribneria is part of this clade. Moreover, Holcinae is not monophyletic in its current circumscription because Deschampsia does not form a clade with Holcus and Vahlodea, which are sister taxa. The results support recognition of Deschampsia in its own subtribe Aristaveninae. Substantial further changes to the classification of these grasses will be needed to produce generic circumscriptions consistent with phylogenetic evidence. The following 15 new combinations are made: Calamagrostis ×calammophila, C. breviligulata, C. breviligulata subsp. champlainensis, C. ×don-hensonii, Deschampsia aurea, D. bolanderi, D. chrysantha, D. chrysantha var. phalaroides, D. eminens, D. eminens var. fulva, D. eminens var. inclusa, D. hackelii, D. ovata, and D. ovata var. nivalis. D. podophora; Deschampsia parodiana is proposed; the new subtribe Lagurinae is de- scribed; and a second-step lectotype is designated for the name Deyeuxia phalaroides. Keywords grasses, phylogenetics, ETS, systematics, taxonomy, classification Table of contents Introduction ............................................................................................................. 4 Methods ................................................................................................................... 7 Taxon and genome sampling ................................................................................ 7 DNA sequencing and alignment .......................................................................... 8 Phylogenetic analyses ......................................................................................... 10 Results .................................................................................................................... 12 ITS+ETS and ITS analyses ................................................................................. 12 Combined plastid analyses ................................................................................. 21 Molecular phylogenetics of cool-season grasses in the subtribes Agrostidinae... 3 Indels ................................................................................................................. 40 Discussion .............................................................................................................. 41 Aveninae and Koeleriinae ................................................................................... 43 Koeleriinae Clade A ....................................................................................... 45 Trisetum p.p., Trisetaria, Koeleria, Rostraria, Avellinia and Gaudinia ........... 45 Lagurus ...................................................................................................... 51 Koeleriinae Clade B ....................................................................................... 52 Calamagrostis/Deyeuxia ............................................................................... 52 Leptophyllochloa .......................................................................................... 54 Peyritschia .................................................................................................. 54 Sphenopholis ............................................................................................... 55 Trisetopsis ................................................................................................... 56 Generic classification in Koeleriinae ............................................................... 57 Agrostidinae + Calothecinae + Brizinae .............................................................. 58 Chascolytrum (Calothecinae p.p.) + Deyeuxia effusa (Agrostidinae p.p.) ........... 58 Relchela (Calothecinae p.p.) + Echinopogon + Calamagrostis coarctata + Di- chelachne (Agrostidinae p.p.) .................................................................. 59 Brizinae ...................................................................................................... 60 Agrostidinae p.p. ...........................................................................................