Phylogeny, Diversification Rate, and Divergence Time of Agave sensu lato (Asparagaceae), a Group of Recent Origin in the Process of Diversification.

Agave sensu lato is one of the most diverse and complex genera of Asparagaceae, with more than 250 species. The morphological, ecological, and evolutionary diversity of the group has complicated its taxonomical study. We conducted phylogenetic analyses of DNA sequence data to reconstruct the phylogenetic relationships of the Agave genus. We included 107 species of the Asparagaceae family from which 83 correspond to the Agave sensu lato clade (Agave sensu stricto + Polianthes + Manfreda and Prochnyanthes, which together represent 30% of the genus) and as outgroups the genera Dasylirion, Hesperoyucca, Chlorogalum, Camassia, Hesperaloe, Yucca, Beschorneria, and Furcraea, in order to estimate the age and propose the history of their diversification. Previous studies postulated the relevance of the Miocene in the speciation rates of the agaves, as well as the relevance of the type of inflorescence in its diversification. However, these assertions have not been well supported. The analysis of chloroplast regions resulted in low resolution, which could be the consequence of the few variable sites. On the other hand, the internal transcribed spacer (ITS) implemented in our analysis ensued in higher resolution and better support values. Our phylogenetic analyses recovered five groups; one is the Striatae group, which is the sister group to Agave sensu stricto clade. Within this clade, we found three main groups with high support; these groups are not related with previous morphological proposals. We also analyzed the dates of origin and diversification rates. A Bayesian analysis of macroevolutionary mixtures indicated two significant shifts; the first was identified at 6.18 Ma, where the speciation rate increased to 4.10 species/Mya, this shift occurred during the late Miocene period, characterized by the emergence of arid biomes in North America. The second was identified at a stem age of 2.68 Ma where the speciation rate increased to 6.04 species/Mya. Concerning the ancestral reconstruction state of the inflorescence type in the Agave sensu stricto clade, the spike inflorescence character was predominant in the early-diverging groups, whereas the late-diverging groups present panicle inflorescences as the predominant character and higher speciation rates.

Mayahuelin, a Type I Ribosome Inactivating Protein: Characterization, Evolution, and Utilization in Phylogenetic Analyses of Agave.

Agaves resist extreme heat and drought. In A. tequilana var. azul, the central spike of the rosette -containing the shoot apical meristem and folded leaves in early stages of development- is remarkably heat tolerant. We found that the most abundant protein in this organ is a 27 kDa protein. This protein was named mayahuelin to honor Mayáhuel, the agave goddess in the Aztec pantheon. LC-MS/MS analyses identified mayahuelin as a type I RIP (Ribosome Inactivating Protein). In addition to the spike, mayahuelin was expressed in the peduncle and in seeds, whereas in mature leaves, anthers, filaments, pistils, and tepals was absent. Anti-mayahuelin antibody raised against the A. tequilana var. azul protein revealed strong signals in spike leaves of A. angustifolia, A. bracteosa, A. rhodacantha, and A. vilmoriniana, and moderate signals in A. isthmensis, A. kerchovei, A. striata ssp. falcata, and A. titanota, indicating conservation at the protein level throughout the Agave genus. As in charybdin, a type I RIP characterized in Drimia maritima, mayahuelin from A. tequilana var. azul contains a natural aa substitution (Y76D) in one out of four aa comprising the active site. The RIP gene family in A. tequilana var. azul consists of at least 12 genes and Mayahuelin is the only member encoding active site substitutions. Unlike canonical plant RIPs, expression of Mayahuelin gene in S. cerevisiae did not compromise growth. The inhibitory activity of the purified protein on a wheat germ in vitro translation system was moderate. Mayahuelin orthologs from other Agave species displayed one of six alleles at Y76: (Y/Y, D/D, S/S, Y/D, Y/S, D/S) and proved to be useful markers for phylogenetic analysis. Homozygous alleles were more frequent in wild accessions whereas heterozygous alleles were more frequent in cultivars. Mayahuelin sequences from different wild populations of A. angustifolia and A. rhodacantha allowed the identification of accessions closely related to azul, manso, sigüín, mano larga, and bermejo varieties of A. tequilana and var. espadín of A. angustifolia. Four A. rhodacantha accessions and A. angustifolia var. espadín were closer relatives of A. tequilana var. azul than A. angustifolia wild accessions or other A. tequilana varieties.

Erratum to: A Rapid and Reliable Method for Total Protein Extraction from Succulent Plants for Proteomic Analysis.

The original version of this article unfortunately contains a mistake. The authors have inadvertently incorrectly listed the concentration of TCA in the acetone/TCA/ß-ME solution in the materials and methods section of this paper. The TCA concentration in Sects. 2.3.2 and 2.3.5 should be 10% TCA, making the solution acetone/10% TCA/0.07% ß-ME. It is now corrected with this erratum.

A Rapid and Reliable Method for Total Protein Extraction from Succulent Plants for Proteomic Analysis.

Crassulacean acid metabolism plants have some morphological features, such as succulent and reduced leaves, thick cuticles, and sunken stomata that help them prevent excessive water loss and irradiation. As molecular constituents of these morphological adaptations to xeric environments, succulent plants produce a set of specific compounds such as complex polysaccharides, pigments, waxes, and terpenoids, to name a few, in addition to uncharacterized proteases. Since all these compounds interfere with the analysis of proteins by electrophoretic techniques, preparation of high quality samples from these sources represents a real challenge. The absence of adequate protocols for protein extraction has restrained the study of this class of plants at the molecular level. Here, we present a rapid and reliable protocol that could be accomplished in 1 h and applied to a broad range of plants with reproducible results. We were able to obtain well-resolved SDS/PAGE protein patterns in extracts from different members of the subfamilies Agavoideae (Agave, Yucca, Manfreda, and Furcraea), Nolinoideae (Dasylirion and Beucarnea), and the Cactaceae family. This method is based on the differential solubility of contaminants and proteins in the presence of acetone and pH-altered solutions. We speculate about the role of saponins and high molecular weight carbohydrates to produce electrophoretic-compatible samples. A modification of the basic protocol allowed the analysis of samples by bidimensional electrophoresis (2DE) for proteomic analysis. Furostanol glycoside 26-O-ß-glucosidase (an enzyme involved in steroid saponin synthesis) was successfully identified by mass spectrometry analysis and de novo sequencing of a 2DE spot from an Agave attenuata sample.

A GIS-based comparison of the Mexican national and IUCN methods for determining extinction risk.

The national systems used in the evaluation of extinction risk are often touted as more readily applied and somehow more regionally appropriate than the system of the International Union for Conservation of Nature (IUCN). We compared risk assessments of the Mexican national system (method for evaluation of risk of extinction of wild species [MER]) with the IUCN system for the 16 Polianthes taxa (Agavaceae), a genus of plants with marked variation in distribution sizes. We used a novel combination of herbarium data, geographic information systems (GIS), and species distribution models to provide rapid, repeatable estimates of extinction risk. Our GIS method showed that the MER and the IUCN system use similar data. Our comparison illustrates how the IUCN method can be applied even when all desirable data are not available, and that the MER offers no special regional advantage with respect to the IUCN regional system. Instead, our results coincided, with both systems identifying 14 taxa of conservation concern and the remaining two taxa of low risk, largely because both systems use similar information. An obstacle for the application of the MER is that there are no standards for quantifying the criteria of habitat condition and intrinsic biological vulnerability. If these impossible-to-quantify criteria are left out, what are left are geographical distribution and the impact of human activity, essentially the considerations we were able to assess for the IUCN method. Our method has the advantage of making the IUCN criteria easy to apply, and because each step can be standardized between studies, it ensures greater comparability of extinction risk estimates among taxa.

Yeasts associated with the production of Mexican alcoholic nondistilled and distilled Agave beverages.

The great variety of agaves and their multiple uses have played an important role in the cultural identification of Mexico. They have been exploited in many ways for over 10,000 years, and one of these applications is the production of alcoholic nondistilled and distilled beverages. Most of the production processes of these Mexican beverages involve a complex fermentation in which bacteria (mainly lactic and acetic acid) and yeasts (non-Saccharomyces and Saccharomyces) are present in stable mixed populations, or succeeding one another, and have a significant impact on the sensorial characteristics and nutritive value of the final product. This minireview focuses on several nondistilled and distilled Agave beverages, their production area, the Agave species used in their elaboration, the functional microbiota involved in the fermentation process, their fermentation products (when known), the biochemical changes of these unique fermentations, and their impact on the quality and sensorial characteristics of the product.