Molecular systematics of Boraginaceae tribe Boragineae based on ITS1 and trnL sequences, with special reference to Anchusa s.l

Abstract

Background and aims: Boragineae is one of the main tribes of Boraginaceae, but delimitation and intergeneric classification of this group are unclear and have not yet been studied using DNA sequences. In particular, phylogenetic relationships in Anchusa s.l. still need to be elucidated in order to assess its taxonomic boundaries with respect to the controversial segregate genera Hormuzakia, Gastrocotyle, Phyllocara and Cynoglottis.

Methods: Phylogenetic relationships among 51 taxa of tribe Boragineae were investigated by comparative sequencing of the trnL(UAA) intron of the plastid genome and of the ITS1 region of the nuclear ribosomal DNA. Exemplar taxa from 16 genera of Boragineae and all subgenera of Anchusa s.l. were included, along with two selected outgroups from tribes Lithospermeae and Cynoglosseae.

Key results: Phylogenies generated by maximum parsimony and combined ITS1-trnL sequences support the monophyly of the tribe and a split into two clades, Pentaglottis and the remainder of Boragineae. The latter contains two large monophyletic groups. The first consists of three moderately to well-supported branches, Borago-Symphytum, Pulmonaria-Nonea and Brunnera. In the Pulmonaria-Nonea subclade, the rare endemic Paraskevia cesatiana is sister to Pulmonaria, and Nonea appears to be paraphyletic with respect to Elizaldia. The second main group corresponds to the well-supported clade of Anchusa s.l., with the megaphyllic, polyploid herb Trachystemon orientalis as sister taxon, although with low support. Anchusa s.l. is highly paraphyletic to its segregate genera and falls into four subclades: (1) Phyllocara, Hormuzakia, Anchusa subgenus Buglossum and A. subgenus Buglossoides; (2) Gastrocotyle; (3) A. subgenus Buglossellum and Cynoglottis; and (4) A. subgenus Anchusa, Lycopsis and Anchusella. All species of Anchusa subg. Anchusa, including the South African A. capensis, are included in a single unresolved clade. Anchusa subgenus Limbata is also included here despite marked divergence in floral morphology. The low nucleotide variation of ITS1 suggests a recent partly adaptive radiation within this group.

Conclusions: Molecular data show that nine of the usually accepted genera of the Boragineae consisting of two or more species are monophyletic: Anchusella, Borago, Brunnera, Cynoglottis, Gastrocotyle, Hormuzakia, Nonea, Pulmonaria and Symphytum. In addition, the tribe includes the four monotypic genera Paraskevia, Pentaglottis, Phyllocara and Trachystemon. The morphologically well-characterized segregate genera in Anchusa s.l. are all confirmed by DNA sequences and should be definitively accepted. Most of the traditionally recognized subgenera of Anchusa are also supported as monophyletic groups by both nuclear and plastid sequence data. In order to bring taxonomy in line with phylogeny, the institution of new, independent generic entities for subgenera Buglossum, Buglossellum and Buglossoides and a narrower but more natural concept of Anchusa are advocated.

Fig. 1.

Strict consensus of the 3380 most-parsimonious trees from trnL sequence data. Tree length (L) = 134, consistency index (CI) = 0·813, retention index (RI) = 0·903. The subgenera of Anchusa s.l. are indicated. The numbers above the branches are bootstrap percentages (percentages <50 % are not shown). Main polytomies are indicated by dotted lines and the main unresolved clades are numbered (1–8).

Fig. 2.

Condensed alignment scheme of trnL sequences (outgroups, identical and non-informative positions removed). Taxa are ordered according to the main insertions/deletions shared. Column numbers refer to nucleotide positions in the original alignment.

Fig. 3.

One of the six most-parsimonious trees from ITS1-trnL sequences. L = 740, CI = 0·605, RI = 0·787. The subgenera of Anchusa s.l. are indicated. Letters on the branches indicate the main clades discussed in the text; numbers indicate bootstrap percentages (percentages < 50 % are not shown). Branch lengths are estimated under ACCTRAN. The branches collapsing in the strict consensus tree (not shown) are indicated by dotted lines.

Fig. 4.

Distribution of 15 morphological characters plotted onto the nine major clades of ITS1-trnL phylogeny of the Boragineae. A square on the clade indicates bootstrap support >85 %. A cross indicates that all or the majority of the taxa of the clade possess the morphological character. Habit: (1) only perennial. Inflorescence and flower: (2) cymes always ebracteate; (3) corolla brachymorphic (with short tube and rotate to subrotate limb); (4) zygomorphy present in some taxa; (5) corolla scales in two series; (6) granular thickenings of cutin on the trichomes of the faucal scales; (7) stylar polymorphism present in some taxa; (8) stigma papillae with typical lageniform shape. Fruit: (9) mericarpids excentrically stalked at the base; (10) mericarpids erect or suberect in all or some of the taxa. Pollen: (11) grains small (P < 15 μm); (12) grains with prolate shape; (13) mesocolpia with supratectum gemmae; (14) grains with (3)-4-(5) apertures; (15) apertures rhomboidal with conical processes on the colpus membrane.