Aberrant meiotic behavior in Agave tequilana Weber var. azul

Background  

Agave tequilana Weber var. azul, is the only one variety permitted by federal law in México to be used for tequila production which is the most popular contemporary alcoholic beverage made from agave and recognized worldwide. Despite the economic, genetic, and ornamental value of the plant, it has not been subjected to detailed cytogenetic research, which could lead to a better understanding of its reproduction for future genetic improvement.     

Spatial and temporal dynamics of blue agave (Agave tequilana Weber var. azul) wilt in Jalisco Mexico

Tequila is a Mexican alcoholic beverage made from the fermentation and distillation of the blue agave (Agave tequilana Weber var. azul) stem. This crop is affected by a wilt associated mainly with Fusarium oxysporum. This disease can produce considerable yield losses. Little is known about the spatial and temporal behaviour of blue agave wilt. In this work, the spatial and temporal dynamics of the disease in commercial blue agave plantations in the state of Jalisco, Mexico, were analysed. Four fields of approximately 1 ha were selected in the municipalities of Arandas and Magdalena, in which disease assessments were carried out over a year of evaluation. Each plant was categorized based on a scale with four severity classes (healthy plant, severity class 1, severity class 2 and severity class 3). Maps of disease distribution were made. The spatial pattern was analysed by means of four indicators of spatial variation for binomial data; the spatial and temporal variation was analysed by means of transition matrices. An aggregate spatial pattern was observed in all fields. The transitions in severity classes were not completely unidirectional; some plants showed symptom remission between the date of first and second disease assessment, while others remained at their original severity. Severity class 1 occurred most frequently in Arandas fields (from 12.9% to 40.3%). There was a notable increase in wilt severity to class 2 in the Magdalena fields (from 4% to 50.6%). The rates of disease development towards severity class 3 are low and do not suppose a significant loss for the crop; nevertheless, the rates of disease development towards the wilt severity class 2 do put in risk the health of the crop and the availability of the raw material for the making of tequila.     

Bioprospecting for fungi with potential pathogenic activity on leaves of Agave tequilana Weber var. Azul

Thirty different fungal strains were isolated from A. tequilana leaves showing disease symptoms such as wilt and curled leaves, black, red and chlorotic spots. Ten genera were identified and confirmed by using the LSU D1/D2 rDNA and ITS1‐5.8S‐ITS2 regions, mainly of the Ascomycota phylum, where the Lasiodiploidia and Neoscytalidium genera were the more (46.6%) abundant. The other genera identified were Cladosporium, Cytospora, Epicoccum, Flavodon, Lasiodiplodia, Myrmaecium, Neoscytalidium, Penicillium, Peniophora, Purpureocillium, Trametes and Fusarium. Five strains of Lasiodiplodia and one of Fusarium were selected based on their representativeness and pathogenic potential on Agaves. Pathogenic potential was analysed by both, an infection assay, evidenced as necrosis, and by pectinolytic activity. Specifically, necrosis infection assay was conducted by puncture (wounded) infection and by direct mycelium contact. In general, Lasiodiplodia strains exhibited different pathogenic profiles according to their necrosis percentages, regardless of the infection method used. Fusarium strain analysed also showed a high necrosis infection (> 99%). Pectinolytic activity used as an indirect measurement of pathogenesis presented a high Fusarium extract activity (peaking at 23.9 U). Lasiodiplodia strains exhibited up 6 times more enzymatic activity (peaking at 143.5) than Fusarium strain analysed. In addition, Agave leaf extracts used totally or partially as carbon source during fungal induction culture may induce different pathogenic activities in these strains. In general, the two pathogenicity assays implemented evidenced differences in the pathogenicity profile of these analysed strains.     

Heterologous expression of Fusarium oxysporum tomatinase in Saccharomyces cerevisiae increases its resistance to saponins and improves ethanol production during the fermentation of Agave tequilana Weber var. azul and Agave salmiana must

This paper describes the effect of the heterologous expression of tomatinase from Fusarium oxysporum f. sp lycopersici in Saccharomyces cerevisiae. The gene FoTom1 under the control of the S. cerevisiae phosphoglycerate kinase (PGK1) promoter was cloned into pYES2. S. cerevisiae strain Y45 was transformed with this vector and URA3 transformant strains were selected for resistance to α-tomatine. Two transformants were randomly selected for further study (designated Y45-1 and Y45-2). Control strain Y45 was inhibited at 50 μM α-tomatine, in contrast, transformants Y45-1 and Y45-2 did not show inhibition at 200 μM. Tomatinase activity was detected by HPLC monitoring tomatine disappearance and tomatidine appearance in the supernatants of culture medium. Maximum tomatinase activity was observed in the transformants after 6 h, remaining constant during the following 24 h. No tomatinase activity was detected in the parental strain. Moreover, the transformants were able to grow and produce ethanol in a mix of Agave tequilana Weber var. azul and Agave salmiana must, contrary to the Y45 strain which was unable to grow and ferment under these conditions.     

In vitro induction of a trisomic of Agave tequilana Weber var. Azul (Agavaceae) by para-fluorophenylalanine treatment

The effect of para-fluorophenylalanine (PFP) on the production of trisomic plants of Agave tequilana Weber var. Azul produced through somatic embryogenesis was investigated. Normal diploid plants with 2n = 2x = 60 were obtained in the control treatment and with 4 mg L⁻¹ PFP exposure, while use of 8 and 12 mg L⁻¹ PFP led to production of trisomics with 2n = 2x = 61. Normal diploid plants showed a bimodal karyotype with five pairs of large chromosomes and 25 pairs of small chromosomes. Trisomic plants also had a bimodal karyotype with a group of three chromosomes in position five of the chromosome set. More than 13 homologous chromosome pairs exhibited structural changes. Differences in chromosome arm ratio (long arm/short arm) were also found in eight chromosome pairs; all these aberrations in the chromosome complement of trisomic plants were probably caused by inversions, deletions, and/or duplications produced by high concentrations of PFP. The gross chromosome structural changes and the presence of a single extra chromosome could have been induced by the effect of PFP on the mitotic spindle by inducing nondisjunction of sister chromatids, resulting in hyperploids (2n + x) and hypoploids (2n − x). Flow cytometric analysis of nuclear DNA content was performed using nuclei isolated from young leaves of normal and trisomic plants. The 2C DNA content of 8.635 pg (1Cx = 4,223 Mbp of trisomic plants was different (p < 0.001) than that of normal plants (2C DNA = 8.389 pg (1Cx = 4,102 Mbp). The difference in genome size was correlated with the large structural changes in the trisomic plant genomes.     

Active telomerase during leaf growth and increase of age in plants from Agave tequilana var. Azul

In plants, previous studies show that telomerase activity contributes to the maintenance of telomeric length for the proper development of organs and tissues. In this work, we investigated telomerase activity in A. tequilana during several years of cultivation. We found that during growth of the leaf there are two crucial phases: (1) the onset of cell elongation in 3 years and (2) differentiation of vascular bundles in 6 years. This coincides with the ages where the highest telomerase activity is seen. Therefore indicates that telomerase is associated with cellular activities such as; elongation, division, and cell differentiation. Likewise, we detected high activity during the period of vegetative growth, indicating that telomerase also contributes to telomeric maintenance on the leaf in A. tequilana.     

Utilization of byproducts from the tequila industry: part 1: agave bagasse as a raw material for animal feeding and fiberboard production

Agave bagasse was successfully separated into fractions that were used in sheep feeding trials. Agave bagasse can be substituted for corn stubble in the sheep's diet which resulted in improved weight gain. Agave bagasse was also processed into long and short fiber fractions with a hammermill and fiberboards of medium and high specific gravities being produced. Medium specific gravity agave fiberboards had moisture and mechanical properties comparable to medium specific gravity fiberboards made using aspen fiber. All high specific gravity agave fiberboards made from short or long fibers were stronger in bending than the ANSI standard for hardboards.     

<i>In Vitro</i>-Propagation of <i>Agave tequilana</i> Weber cv. azul in a Temporary Immersion System

In Mexico, there is a need to produce large quantities of plantlets for the establishment and replanting of blue (cv. azul) agave production areas. Most of these plots are within the origin denomination area (DOT, Spanish acronym) of the distilled product of this plant, known as tequila. The objective of this study was to develop an in vitro-propagation protocol for Agave tequilana Weber cv. azul using segmented stems in both: solid and liquid media. A disinfection and in vitro technique were developed to obtain shoots, through plantlets collected in commercial plots, which attained 100% surface-disinfection and budding rate. At the multiplication stage, the effects of 6-Benzylaminopurine (BA) (0.0, 4.4 and 13.2 μM) and kinetin (0.0, 9.4, 18.8 and 37.6 μM) were evaluated on lateral-shoot production of segmented sagittal stems. These were cultivated on Murashige & Skoog (MS) medium, with the addition of 3.0% sucrose and 8 g L⁻¹ agar. It was observed that BA and kinetin increased the number of shoots per explant, obtaining up to 18 and 26, respectively. Furthermore, it was found that just the sagittal segmentation of explants increased axillary budding. On the other hand, segmented-stem bases were grown in MS liquid medium with 3.0% sucrose, inside a RITA® system, programmed by a 5 min immersion step with a frequency of every 4 h. The effect of Indole−3-Acetic acid (IAA) (0.57, 2.9, 5.7 μM) was evaluated, while maintaining a concentration of BA (13.2 μM). It was observed that the greatest concentration of IAA led to the formation of more than 20 buds per explant. These results offer a new methodology to increase the efficiency of A. tequilana Weber cv. azul-in vitro multiplication by sagittal segmentation of stems and the addition of BA and/or IAA.     

Potential of Plants from the Genus Agave as Bioenergy Crops

Agave is a succulent genus within the monocot family Agavaceae. The plants have a large rosette of thick fleshy leaves, each ending generally in a sharp point, and are native to arid and semi-arid regions from the southern USA to northern South America. The most important commercial species is Agave tequilana grown for production of tequila. Several cultivated species of Agave such as Agave sislana and Agave salmiana can perform well in areas where rainfall is insufficient for the cultivation of many C₃ and C₄ crops. The key feature of the crassulacean acid metabolism photosynthetic pathway used by agaves is the stomata opening and CO₂ uptake during the night, thus allowing less water to be lost by transpiration. Alcoholic beverages, sweeteners, fibers, and some specialty chemicals are currently the main products coming from agave plants. The recovered information related to productivity, biofuel processability, by-products, etc. suggests that some Agave species have a real potential to compete economically with other bioenergy crops. But more than compete, it could complement the list of bioenergy crops due to its capacity to grow with very little rainfall and/or inputs and still reach good amount of biomass, so unused semi-arid land could be productive. Although Agave has great potential to be developed as a bioenergy crop, more laboratory and field research are needed.     

Utilization of salt-rich by-products from the dairy industry as feedstock for recombinant protein production by Debaryomyces hansenii

The dairy industry processes vast amounts of milk and generates high amounts of secondary by-products, which are still rich in nutrients (high Chemical Oxygen Demand (COD) and Biochemical Oxygen Demand (BOD) levels) but contain high concentrations of salt. The current European legislation only allows disposing of these effluents directly into the waterways with previous treatment, which is laborious and expensive. Therefore, as much as possible, these by-products are reutilized as animal feed material and, if not applicable, used as fertilizers adding phosphorus, potassium, nitrogen, and other nutrients to the soil. Finding biological alternatives to revalue dairy by-products is of crucial interest in order to improve the utilization of dry dairy matter and reduce the environmental impact of every litre of milk produced. Debaryomyces hansenii is a halotolerant non-conventional yeast with high potential for this purpose. It presents some beneficial traits – capacity to metabolize a variety of sugars, tolerance to high osmotic environments, resistance to extreme temperatures and pHs – that make this yeast a well-suited option to grow using complex feedstock, such as industrial waste, instead of the traditional commercial media. In this work, we study for the first time D. hansenii's ability to grow and produce a recombinant protein (YFP) from dairy saline whey by-products. Cultivations at different scales (1.5, 100 and 500 ml) were performed without neither sterilizing the medium nor using pure water. Our results conclude that D. hansenii is able to perform well and produce YFP in the aforementioned salty substrate. Interestingly, it is able to outcompete other microorganisms present in the waste without altering its cell performance or protein production capacity.     

Composition and ultrastructure of the suberized cell wall of isolated crystal idioblasts from Agave americana L. leaves

Styloid-calcium-oxalate-crystal-containing idioblasts possess an interior cell-wall layer which has a lamellar ultrastructure. Idioblasts were isolated by centrifugation of an Agave americana leaf homogenate through 2M sucrose. The aliphatic monomers of the polymeric material from an idioblast fraction were primarily -hydroxy acids (32%) and dicarboxylic acids (35%), with C_18:1 dicarboxylic acid being the most dominant monomer (25%). Nitrobenzene oxidation of the idioblasts yielded syringaldehyde and vanillin in a ratio of 0.46:1. The major class of wax associated with the idioblasts was free fatty acids (34%). A major homologue of both the fatty acid and fatty alcohol fractions of this wax was C₂₂. The hydrocarbon fraction of the wax had a broad chainlength distribution with a large amount of even-numbered (47%) and shorter-chain homologues. The ultrastructure, the composition of the aliphatic and aromatic components of the polymeric material as well as the composition of the wax show that the idioblast cell wall is suberized. The wax and cutin polymer of the epidermis of A. americana leaves were chemically characterized for comparative purposes.Scientific paper No. 6115, Project 2001, College of Agriculture Research Center, Washington State University, Pullman, WA 99164, USA     

Potential of Agave lechuguilla biomass for Cr(III) removal from aqueous solutions: thermodynamic studies

Thermodynamic studies on the bioadsorption of Cr(III) onto Agave lechuguilla biomass were conduced. The experimental results at different temperatures were modeled using the Langmuir and Freundlich isotherms to obtain the characteristic parameters of each model. Both the Freundlich and Langmuir models were found to represent the bioadsorption process. The average adsorption capacities calculated from Freundlich (4.7 mg/g) and Langmuir (14.2 mg/g) isotherms showed A. lechuguilla to be an effective biomass in the removal of Cr(III) from an aqueous solution. Thermodynamic parameters (deltaG0, deltaH0 and deltaS0) determined in the temperature range from 10 to 40 degrees C along with the parameters of the Dubinin-Radushkevick equation support the idea that the binding of Cr(III) may be caused by interactions with functional groups such as carboxyl groups located on the outer surface of the cell tissue of the bioadsorbent.     

叶表面角质层在贝母属植物叶鉴定中的意义

叶表面角质层在贝母属植物叶鉴定中的意义李萍，濮祖茂，蒋鑫，刘惠娟，徐国钧（中国药科大学生药学教研室；分析中心电镜室南京２１０００９）ＴｈｅｄｉａｇｎｏｓｔｉｃｖａｌｕｅｏｆｔｈｅｃｕｔｉｃｌｅｉｎｔｈｅｌｅａｖｅｓｆｒｏｍｇｅｎｕｓＦｒｉｔｉｌｌａｒ...     

Effect of ultrasound and addition of bacterial product on hydrolysis of by-products from the meat-processing industry

The hydrolysis efficiency of ultrasound (US) and bacterial product (BP; Liquid Certizyme 5™) was studied as a pre-treatment for anaerobic digestion (AD) of four animal by-products (ABP) from the meat-processing industry (digestive tract content + drumsieve waste, dissolved air flotation (DAF) sludge, grease trap sludge and the mixture of these). The change in volatile solids (VS) based hydrolysis parameters was screened using different specific US energy (Es) inputs (1000-14,000 kJ/kg of total solids (TS)) and durations of BP treatment (3-48 h). The Es of 6000-8500 kJ/kg TS increased most extensively the soluble chemical oxygen demand (COD_sol) and VS ratio, by 45-290%, and soluble nitrogen (N_sol) by 44-99%. Simultaneously, the average particle size (APS) decreased by 45-75%, from the original value. With BP, the highest increases in COD_sol/VS (29-130%), N_sol (35-63%) and decline in APS (40-70%) depended on the raw material (digestive tract content + drumsieve waste: 3 and 24 h; DAF sludge: 3 and 6 h, mixture: 12 and 48 h). BP apparently was more effective at hydrolyzing smaller cellulose particles, while US appeared to enhance the degradation of grease-containing cells.