Assessment of N-glycan heterogeneity of cactus glycoproteins by one-dimensional gel electrophoresis and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

Artificial environmental conditions in tissue culture, such as elevated relative humidity and rich nutrient medium, can influence and modify tissue growth and induce spontaneous changes from characteristic organization pattern to unorganized callus. As succulent plants with crassulacean acid metabolism, cacti are particularly susceptible to this altered growth environment. Glycosylated proteins of Mammillaria gracillis tissues cultivated in vitro, separated by SDS-PAGE, were detected with Con A after the transfer of proteins onto the nitrocellulose membrane. The glycan components were further characterized by affinity blotting with different lectins (GNA, DSA, PNA, and RCA(120)). The results revealed significant differences in glycoprotein pattern among the investigated cactus tissues (shoot, callus, hyperhydric regenerant, and tumor). To test whether the N-glycosylation of the same protein can vary in different developmental stages of cactus tissue, the N-glycans were analyzed by MALDI-TOF MS after in-gel deglycosylation of the excised 38-kDa protein band. Paucimannosidic-type N-glycans were detected in oligosaccharide mixtures from shoot and callus, while the hyperhydric regenerant and tumor shared glycans of complex type. The hybrid oligosaccharide structures were found only in tumor tissue. These results indicate that the adaptation of plant cells to artificial environment in tissue culture is reflected in N-glycosylation, and structures of N-linked glycans vary with different developmental stages of Mammillaria gracillis tissues.     

DNA Isolation and Amplification from Cacti

The cacti family is a morphologically heterogeneous group comprising 100 genera and about 1500 species (Hernandez and Barcenas, 1996). With the exception of one genus, all members of this family are native to America (Hernandez and Barcenas, 1996). There are three subfamilies, Opuntioideae, Cactoideae, and Pereskioideae (Gibson and Nobel, 1986). DNA isolation from cacti is notoriously difficult because they contain high amounts of polysaccharides and secondary metabolites which form insoluble complexes with nucleic acids during extraction (Guillemaut and Marechal-Drouard, 1992). Like in other groups of plants, the secondary metabolites and polysaccharides in cacti inhibit enzyme action (Porebski et al., 1997). The polysaccharides are visually evident by their viscous, glue-like texture and they make the DNA unmanageable when pipeting and hard to amplify by the polymerase chain reaction (PCR) (Poresbski et al., 1997). We report an easy and inexpensive protocol to isolate DNA from cacti. We used this method to isolate DNA from 85 species (170 individuals) of 39 genera of the subfamilies Pereskioideae, Opuntioidea, and Cactoideae. This procedure is a modification of a protocol described by De la Cruz et al. (1995) for the Cacti family. It requires only a few grams of tissue and does not require destruction of the whole plant to produce high molecular weight genomic DNA. The DNA from this procedure can be amplified consistently by PCR and used for RAPD analysis.     

Association between Opuntia species invasion and changes in land-cover in the Mediterranean region

In Mediterranean regions, biological invasions pose a major threat to the conservation of native species and the integrity of ecosystems. In addition, changes in land‐cover are a widespread phenomenon in Mediterranean regions, where an increase in urban areas and major changes from agricultural abandonment to shrub encroachment and afforestation are occurring. However, the link between biological invasions and changes in land‐cover has scarcely been analyzed. We conducted a regional survey of the distribution of the two alien prickly‐pear cacti Opuntia maxima and O. stricta in Cap de Creus (Catalonia, Spain) and related patterns of invasion to spatially explicit data on land‐cover/change from 1973 to 1993 to test the hypotheses that the two Opuntia species invade areas that have experienced large land‐cover transformations. We found that Opuntia invasion is particularly high in shrublands and woodlands located near urban areas. O. maxima are over‐represented in the shrublands and O. stricta in the woodlands that were former crops. Crop coverage has dropped by 71% in this 20‐year period. This study highlights the role of past land‐cover in understanding the present distribution of plant invasions.     

Micropropagation of Turbinicarpus laui Glass et Foster, an endemic and endangered species

Summary In a short period of time, a large number of adventitious shoots were regenerated from longitudinal sections of in vitro-germinated seedlings of the endangered Mexican cactus, Turbinicarpus laui. The induction medium consisted of Murashige and Skoog salts, supplemented with 6-benzylaminopurine (BA) and α-naphthaleneacetic acid (NAA) in a wide range of combinations. The most effective concentrations of growth regulators for shoots initiation were 8.8–13.32 μM BA with 0–2.68 μM NAA. After 3–4 mo., individualized shoots were rooted on half-strength Murashige and Skoog medium and then transferred to soil to acclimatize under greenhouse conditions. In vitro strategies play a key role in the conservation and propagation of this commercially important endangered species of cactus.     

Evolutionary trends in the columnar cactus genus Eulychnia (Cactaceae) based on molecular phylogenetics,morphology, distribution,and habitat

Populations of the columnar cactus genus Eulychnia (Cactaceae) are an iconic sight in the Chilean Atacama Desert. The most recent taxonomic treatment of the genus suggested to accept up to seven taxa at species level based on morphological data. To date, species boundaries and infrageneric relationships in Eulychnia have not been investigated using a molecular approach. In this study, sequence data were generated for six chloroplast markers (rpl32-trnL, trnH-psbA, trnL-trnF, trnQ-rps16, trnS-trnG, and ycf1) for the seven species. Where possible, samples were collected from the south and north of the distribution range of widely distributed species, as well as plants from two morphologically distinct populations in the Atacama and Coquimbo Regions. Evolutionary trends of morphological characters were investigated using ancestral state reconstruction, and the habitat of the Chilean taxa was taken into account based on latitudinal and altitudinal distribution, precipitation regime, and vegetation zones. Two major clades were retrieved in the molecular phylogenetic hypotheses, a northern clade and a southern clade, which can easily be distinguished morphologically by differences in rib shape and type of the indumentum of the pericarpel and the hypanthium. The only Eulychnia taxon found outside Chile is most commonly accepted as Eulychnia iquiquensis subsp. ritteri. However, its isolated geographic distribution and the fact that this taxon is not most closely related to E. iquiquensis but was retrieved as sister to the rest of the northern clade in our molecular phylogenetic results support the recognition of E. ritteri at species level. Our results also provide some support for the two recently published species, E. chorosensis (previously placed in E. acida s.l.) and E. taltalensis (previously considered to be part of E. breviflora s.l.). The relationships in the southern clade need further study.     

Molecular evolution and population genetics of two Drosophila mettleri cytochrome P450 genes involved in host plant utilization

Understanding the genetic basis of adaptation is one of the primary goals of evolutionary biology. The evolution of xenobiotic resistance in insects has proven to be an especially suitable arena for studying the genetics of adaptation, and resistant phenotypes are known to result from both coding and regulatory changes. In this study, we examine the evolutionary history and population genetics of two Drosophila mettleri cytochrome P450 genes that are putatively involved in the detoxification of alkaloids present in two of its cactus hosts: saguaro (Carnegiea gigantea) and senita (Lophocereus schottii). Previous studies demonstrated that Cyp28A1 was highly up-regulated following exposure to rotting senita tissue while Cyp4D10 was highly up-regulated following exposure to rotting saguaro tissue. Here, we show that a subset of sites in Cyp28A1 experienced adaptive evolution specifically in the D. mettleri lineage. Moreover, neutrality tests in several populations were also consistent with a history of selection on Cyp28A1. In contrast, we did not find evidence for positive selection on Cyp4D10, although this certainly does not preclude its involvement in host plant use. A surprising result that emerged from our population genetic analyses was the presence of significant genetic differentiation between flies collected from different host plant species (saguaro and senita) at Organ Pipe National Monument, Arizona, USA. This preliminary evidence suggests that D. mettleri may have evolved into distinctive host races that specialize on different hosts, a possibility that warrants further investigation.     

Genetic diversity and relationships among wild and cultivated Stenocereus queretaroensis populations in western Mexico

Stenocereus queretaroensis is an endemic, chiropterophilous, columnar cactus of economic importance in Mexico. To investigate the effect of artificial selection on genetic diversity, inter-simple sequence repeat (ISSR) markers were used to estimate the genetic variation of wild, orchard, and backyard populations of S. queretaroensis from two regions in western Mexico. Six primers were used to generate 62 bands, of which 39 were polymorphic (62.9%). The total genetic diversity was similar in the wild (H_T = 0.296) and orchard (H_T = 0.291) groups, and slightly lower in the backyard group (H_T = 0.281). Wild populations (F_ST = 0.13) were less differentiated than backyard (F_ST = 0.17) and orchard (F_ST = 0.21) populations. The wild and backyard groups were genetically closer (0.015) than the wild and orchard (0.018) groups. A Mantel test revealed a positive correlation between genetic and geographic distances (r = 0.433, p = 0.002). In conclusion, gene flow and the prevailing management system have efficiently maintained genetic diversity and facilitated inter-population differentiation in S. queretaroensis.     

Population dynamics of the cactus Mammillaria gaumeri: an integral projection model approach

The Cactaceae family in Mexico is particularly important because members of this family exhibit a high degree of endemism. Unfortunately, many species of the Cactaceae are threatened or endangered. We employed an integral projection model for studies of the population dynamics of Mammillaria gaumeri, an endemic cactus of the Yucatán characterized by a small population size. The integral projection model provides estimates of the asymptotic growth rate, stable size distribution, reproductive values, and sensitivities and elasticities of the growth rate to changes in vital rates. Nine locations of this species were studied along the Yucatan coast over a 9-year period. Individuals were classified by plant volume. Most population growth rate (λ) values were below unity. The highest elasticity values corresponded to the survival of intermediate size individuals. The percentage of germination in the field was low, and consequently, fecundity values were also low. Reproductive values were observed to increase with plant volume. The stable size distribution of M. gaumeri was skewed toward small individuals. For all years, the kernel showed that individual survival determined the population growth rate.     

Effects of natural and artificial selection on survival of columnar cacti seedlings: the role of adaptation to xeric and mesic environments

Escontria chiotilla, Polaskia chichipe, and Stenocereus pruinosus are species of Mexican columnar cacti that are economically important because of their edible fruits. These species are managed by gathering fruits from the wild, silvicultural management in agroforestry systems, and cultivation in home gardens. Previous studies reported that artificial selection favored individuals that produced larger fruits, which indirectly led to the production of larger seeds and seedlings, with possible effects on survival. We hypothesized that seedlings from managed populations would be larger but more susceptible to xeric conditions than those from wild populations. We evaluated the effects of artificial and natural selection on seedling survival of the three species in wild and managed populations, which were managed with low and high intensity, respectively. We tested seedling performance in gradients of shade (0, 40, and 80%) and humidity (low and high). A GLM of seedling survival showed significant differences among species, shade, and humidity treatments, with each species having environmental requirements associated with their particular adaptations. High humidity decreased seedling survival of all species, and high solar radiation decreased survival of S. pruinosus and P. chichipe. The effect of management type was significant only in S. pruinosus. Significant differences in the initial growth of seedlings among species were detected with ANOVA. In optimal conditions, the hypocotyl and the cotyledons decreased in size and the epicotyl grew, whereas under stress, these structures remained unchanged. The optimum conditions of shade and humidity varied among species and management types. The seedlings of S. pruinosus were the largest and the most susceptible, but in all species, seedlings from managed populations were more susceptible to environmental conditions. Thus, artificial selection influenced the susceptibility of these cacti to xeric environments.