RESUMEN
Plants living above and around caves represent an important, albeit poorly studied, resource within cave ecosystems. The presence of plant material (root-like structures or rhizothemes, saplings, seeds, and seedlings) correlates positively with the biodiversity of the cave dwelling animals as shown for iron-ore caves in Carajás, Pará, Brazil. Plant material collected in caves has proven to be difficult to identify by traditional botanical methods, thus this research aims to provide a qualitative insight into the taxonomy and morphology of rhizothemes and other plant fragments found in the caves. The identification process used a combination of different molecular markers (ITS2, rbcL, and trnH-psbA) followed by a comparison of the sequences obtained against publicly available databases. The rhizothemes were submitted to micromorphological analysis to ascertain their putative root or stem origin and to compare their anatomy with known patterns found in the plant families or genera recovered through molecular matches. All studied samples were Angiosperms, mostly belonging to subclass Rosideae, within four orders: Malpighiales (Euphorbiaceae, Hypericaceae), Sapindales (Anacardiaceae and Sapindaceae), Myrtales (Myrtaceae), Fabales (Fabaceae), and only two belonging to subclass Asteridae, order Gentianales (Apocynaceae). Some of the samples were matched to generic level, with ITS2 being the best marker to identify the fragments because it shows high degree of sequence variation even at specific level and result reliability. All rhizothemes turned out to be roots, and correspondence was found between the existing literature and the individual anatomical patterns for the families and genera retrieved. DNA barcode has proved to be a useful tool to identify plant fragments found in this challenging environment. However, the existence of well curated, authoritatively named collections with ample biological information has proven to be essential to achieve a reliable identification.
RESUMEN
Habenaria is a large genus of terrestrial orchids distributed throughout the tropical and subtropical regions of the world. The integrity and monophyly of this genus have been under discussion for many years, and at one time or another, several genera have been either included in a broadly defined Habenaria or segregated from it. In this study, the phylogenetic relationships of the Neotropical members of the genus and selected groups of African Habenaria were investigated using DNA sequences from the nuclear internal transcribed spacer (ITS) region and the plastid matK gene sampled from 151 taxa of Habenaria from the Neotropics (ca. 51% of the total) as well as 20 species of Habenaria and Bonatea from the Old World. Bayesian and parsimony trees were congruent with each other, and in all analyses, the Neotropical species formed a highly supported group. African species of Habenaria in sections Dolichostachyae, Podandria, Diphyllae, Ceratopetalae and Bilabrellae, and the Neotropical clade formed a highly supported "core Habenaria clade", which includes the type species of the genus from the New World. The topology of the trees indicates an African origin for the Neotropical clade and the low sequence divergence among the Neotropical species suggests a recent radiation of the genus in the New World. Species of Bonatea and Habenaria sections Chlorinae and Multipartitae formed a well-supported clade that was sister to the "core Habenaria clade". The Neotropical clade consists of at least 21 well-supported subgroups, but all Neotropical sections of the current sectional classification are paraphyletic or polyphyletic and will need extensive revision and recircumscription. Most of the Neotropical subgroups formed morphologically uniform assemblage of species, but some cases of morphological divergence within subgroups and convergence between subgroups indicated that morphology alone can be misleading for inferring relationships within the genus. The genera Bertauxia, Kusibabella and Habenella, segregated from New World Habenaria, are not monophyletic and a revision of the sectional classification rather than a generic division seems most appropriate. Our results do not support an extensive generic fragmentation of Habenaria as previously suggested and will provide a framework for revising the infrageneric classification and investigating the patterns of morphological evolution and geographical distribution of the genus in the New World.