Young Cladodes Research Articles

Comparison of total polyphenol content, antioxidant activity, and antimicrobial potential among nine cactus pear (Opuntia spp.) cultivars and their by-products

Nopal cacti (Opuntia spp.) are important natural resources in semi-arid zones throughout theworld, where their fruits and stems are used as forage or human food depending on the varietyand ripeness of the cladode. Members of the genus Opuntia show variable phytochemicalcomposition as well as a high capacity for adaptation to extreme environmental conditions. Onepart of the process to prepare and use of “nopalitos” for human consumption includes removalof modified leaves or thorns, which is approximately 20% of the fresh weight of cladodes isconsidered as a by-product (production residue). The objective of this study was to evaluatethe phytochemical composition, antioxidant capacity, and antibacterial potential of cladodesand cladode by-products from nine nopal cultivars. Young cladodes (<1 month, nopalitos) andtheir corresponding by-products were washed, dried, and macerated in ethanol. Ethanolextracts were isolated by evaporation and resuspended in water. Total phenols and flavonoidswere quantified, and antioxidant capacity was assessed by spectroscopic assays [troloxequivalent antioxidant capacity (TEAC) and percent inhibition of diphenylpicrylhydrazyl (DPPH)radicals]. Antimicrobial activities of the extracts against Vibrio cholerae and Clostridiumperfringens were determined by the well diffusion method and calculation of minimalbactericidal concentration (MBC) by agar plate assay. All extracts tested showed antioxidantand antibacterial activities. Extracts of cladodes and their corresponding by-products showedsimilar MBCs for C. perfringens (around 15 mg ml-1) and V. cholerae (4.0 mg ml-1). However,extracts of cladode by-products showed higher antioxidant capacity than those of cladodesaccording to both DPPH and TEAC assays. Higher concentrations of phenols and flavonoidswere also found in the extracts of by-products (ranging from 91.23 to 853.78 mg gallic acidequivalents per gram dry weight and from 17.10 to 39.42 mg quercetin equivalents per gramdry weight, respectively). This study demonstrates that nopal cactus species exhibit higherantioxidant activity and contents of both phenols and flavonoids in cladode by-products thanthat in their corresponding cladodes. Our data indicate that discarded cladode by-products maybe are a potentially valuable resource for nutraceutical and drug development.

First description of extrafloral nectaries in Opuntia robusta (Cactaceae): Anatomy and ultrastructure.

To our knowledge, there are no studies about the structure and ecological function of extrafloral nectaries (EFNs) in Opuntia robusta. This is the first description of EFNs in O. robusta, where young spines have an interesting structure and a secreting function, which are different from EFNs described in other Cactaceae species. We used light, scanning-electron, and transmission-electron microscopy to examine morphology, anatomy, and ultrastructure of the secretory spines in areoles in female and hermaphrodite individuals of O. robusta. Young cladodes develop areoles with modified and secretory spines as EFNs only active during the early growth phase. EFNs are non-vascularized structures, with no stomata, that consist of a basal meristematic tissue, a middle elongation region, and an apical secretory cone formed by large globular epidermal cells, containing nectar and medullar elongated cells. We observed the presence of Golgi apparatus, vesicles and plastids in the medullar and sup-epidermal cells of the spine. We propose that the nectar is stored in the globular cells at the apex of the spine and secreted by breaking through the globular cells or by pores. We recorded a more frequent presence of ants on younger cladode sprouts producing young secreting spines: this result is parallel with the predictions of Optimal Defense Hypothesis, which states that younger plant organs should be better defended than older ones because their loss produces a higher fitness impairment. Although Diaz-Castelazo’s hypothesis states that a more complex structure of EFNs correlates with their lower among-organs dispersion, comparing to less complex EFNs, non-vascularized structure of EFNs in O. robusta is not associated with their higher among-organs dispersion likened to O. stricta, which produces vascularized EFNs. We provide evidence that this characteristic is not a good taxonomic feature of Opuntia genus. Moreover, the comparison of EFNs of O. robusta and O. stricta suggests that the hypothesis of Diaz-Castelazo should be revised: it is rather a rule but not a law.

Phytochemical Composition and in Vitro Analysis of Nopal (O. Ficus-Indica) Cladodes at Different Stages of Maturity

ABSTRACTThis study aimed to determine the phytochemical profile and nutraceutical properties of nopal cladodes (Opuntia ficus-indica) at different stages of maturity. Medium-age cladodes showed the highest total saponins, phytosterols, and indigestible fiber, as well as the highest in vitro antioxidant capacity and digestive enzymes inhibitory activity. Furthermore, these cladodes presented the highest content of p-hydroxybenzoic acid, p-coumaric acid, rutin, narcissin, nicotiflorin, β-sitosterol, and sitosteryl-3-β-glucopyranoside, as well as several amino acids, organic acids, and fatty acids. Whereas young cladodes contained the highest concentration of condensed and hydrolyzable tannins. These results demonstrated that maturity affects the nutritional and nutraceutical properties of nopal cladodes.

Antioxidant properties and chemical characterization of Spanish Opuntia ficus-indica Mill. cladodes and fruits.

Recent studies have demonstrated that consumption of Opuntia ficus-indica Mill. has an important positive health benefit, mainly due to antioxidant properties, which justifies this research. This study examined antioxidant activity, organic acid and sugar profile, total phenolic, and physicochemical characteristics of six O. ficus-indica cultivars growing in the Spanish Mediterranean. It should be noted that, in this study, both cladodes (young and adult) and fruits (peel and pulp) were analyzed. The antioxidant activity (2,2-azinobis (3-ethylbenzothiazoline-6-sulfonic acid) and 2,2-diphenyl-1-picrylhydrazyl methods) was higher in fruit peel than in cladodes. The young cladodes presented an important antioxidant activity by the ferric-reducing ability of plasma method as well as a higher total phenolic content (18.90 g gallic acid equivalent per kilogram). High-performance liquid chromatography (HPLC) with diode-array detector analysis revealed the absence of sucrose and the presence of glucose and fructose, which the values were higher in pulp fruits. HPLC with refractive index detector analysis showed that citric, malic, and succinic acids were the main organic acids in all cultivars, with a significant higher content in old cladodes. These investigations valorize O. ficus-indica fruits in comparison with cladodes. In general, this plant can be considered as an ingredient for the production of health-promoting food, highlighting mainly in the antioxidant activity and total polyphenols content found in young cladodes and peel fruits. © 2017 Society of Chemical Industry.

Phenolic composition and antioxidant activities of cladodes of the two varieties of cactus pear (Opuntia ficus-indica) grown in Ethiopia

In this study, young cladodes corresponding to two (thorn-less and thorn-containing) varieties of cactus pear ( Opuntia ficus-indica ) from four different areas of Tigray region, Ethiopia, were investigated for their phenolic contents and antioxidant activities. Quantitative estimation of total phenolic contents and antioxidant activities in the cladodes was carried out using spectrophotometric methods. A 1,1-diphenyl-2-picrylhydrazyl (DPPH) scavenging assay was used for the determination of the antioxidant activities, while Folin-Ciocalteau reagent, aluminum chloride and protein (albumin) precipitation methods were used for the determination of total polyphenol, flavonoid and tannin contents, respectively. The cladodes were found to be rich in polyphenols, with values in the range 18.0-71.4 mg gallic acid equivalent (GAE)/g of dry weight (DW). The predominant part of the polyphenols is represented by tannins, 9.00-41.2 mg GAE/g of DW, followed by flavonoids, 6.40-25.0 mg catchin equivalent (CE)/g of DW. The cladodes showed strong antioxidant activities, 59.3-85.8% inhibition compared to ascorbic acid. Application of paired t-test on the overall mean total polyphenol contents of samples collected from the different areas revealed the presence of significant variation between the thorn-less (31.6 GAE/g of DW) and thorn-containing (47.7 GAE/g of DW) varieties. One-way ANOVA also revealed that there are significant variations among samples from the same variety but collected from different areas. KEY WORDS : Opuntia ficus-indica , Flavonoids, Tannins, Polyphenols, Antioxidant activities, Ethiopia Bull. Chem. Soc. Ethiop. 2016 , 30(3), 347-356. DOI: http://dx.doi.org/10.4314/bcse.v30i3.3

Description of the Fruit of Nopalea cochenillifera (L.) Salm-Dyck Sold in the Traditional Market of Rioverde, San Luis Potosí, Mexico

There are nine species of Nopalea and the most widely distributed is N. cochenillifera (L.) Salm-Dyck. This species and its wild congeners inhabit dry and subhumid tropical and subtropical regions in western and eastern Mexico down to Panama. In Mexico, its tender, young cladodes (known locally as nopalitos) and flower buds (tunitas) are eaten as vegetables, its developed cladodes are used as forage in different parts of the world, and the plants are used as living fences and also as ornamentals. The fruits of N. cochenillifera have received little attention in the specialized literature; therefore, the purpose of this note is to describe a set of fruits that was bought in a traditional market in San Luis Potosí, México. The fruits of N. cochenillifera were, on average, two centimeters shorter than those reported in the literature; by weight, the pulp accounted for 45.70 % on average, the peel 48.07 % and the seeds 6.23 %. Mean sweetness was 10.62 oBrix, somewhat lower than that of Opuntia cultivars, which is 12.95. Due to the lack of abortive seeds, we infer that the fruit of Nopalea have been of little interest to humans.

Efficacy of cactus pear (Opuntia ficus-indica) varieties as a source of food and feed in Endamehoni district, northern Ethiopia.

Cactus Pear (Opuntia ficus-indica) serves as a source of food, feed, as well as a means of additional income to the people in northern Ethiopia. The species has different varieties with varied rate of productivity and palatability. This study was conducted to assess the food and feed value of the Cactus Pear varieties in Endamehoni District, northern Ethiopia. It was carried out in three “Tabias” (small administrative subdistricts) and nine sub-Tabias (also known as “Kushets”). These areas were purposively selected as they are well known for their good diversity of Cactus varieties. A total of one hundred and twenty households (n = 120) were randomly selected for interviews using a semi-structured interview. This was supplemented by information obtained through key informants’ interviews and field visits. The local farmers made the identification and characterization of cactus varieties traditionally based on the outstanding phenotypes like fruit characteristics, seed size and content, cladode descriptors and plant height. A total of 13 Cactus varieties were identified and recorded. To assess the food and feed value and to select the best used varieties in the area, preference rankings were made. It was found that not all varieties were equally edible and palatable. The most serious constraints in palatability were associated with fruit taste, fruit size, presence of spines, and hardness of seeds. The ethnobotanical study on palatability was supplemented by chemical analysis on Dry Matter (DM), Ash, Organic Matter (OM) and Crude Protein (CP) content from two-year-old young cladodes of four Cactus varieties (Kille, Wadwada, Magalla and Limo) that are more preferred for food and feed. The analysis of variance (ANOVA) showed significant differences in DM, OM and Ash content at p<0.05 (not at p<0.10). There was no variation in crude protein content amongst the varieties tested. From the results, it is concluded that propagation and use of varieties Kille, Limo, Magalla and Wadwada by farmers of the area is advisable.

Comparación morfológica entre poblaciones silvestres y manejadas de Opuntia atropes (Cactaceae) en Michoacán, México

Mexico has an ancient tradition of consumption of stems and fruits of Opuntia, of wild plants or plants in different degree of domestication. Even though Opuntia ficus-indica is the most cultivated species, there are other Opuntia species locally consumed and less well studied. This is the case of O. atropes native of Mexico and appreciated for its edible young cladodes “nopalitos”. This study evaluated the morphological variability of wild, tolerated, of home gardens and commercial plantations, in order to know whether O. atropes is in the process of domestication and to explore which characteristics have been modified. Three study sites were selected in the state of Michoacán where a total of 120 plants were sampled. Fourteen morphological variables were analyzed using univariate and multivariate methods. A morphological divergence between individuals from managed and non-managed populations was observed. Inmature and mature cladodes from cultivated cactus pears were bigger, with more areoles and with less spines and glochids. These differences as well as its wide cultivated extension in Michoacán suggests that O. atropes is going through an evolutionary process of domestication.

Potential of Opuntia ficus-indica for air pollution biomonitoring: a lead isotopic study.

Opuntia ficus-indica (Ofi) is a long-domesticated cactus that is widespread throughout arid and semiarid regions. Ofi is grown for both its fruits and edible cladodes, which are flattened photosynthetic stems. Young cladodes develop from mother cladodes, thus forming series of cladodes of different ages. Therefore, successive cladodes may hold some potential for biomonitoring over several years the local atmospheric pollution. In this study, cladodes, roots, dust deposited onto the cladodes, and soil samples were collected in the vicinity of three heavily polluted sites, i.e., a fertilizer industry, the road side of a highway, and mine tailings. The lead content was analyzed using atomic absorption spectroscopy (AAS) and inductively coupled plasma-mass spectrometry (ICP-MS). Scanning electron microscopy coupled with energy dispersive X-ray spectroscopy (SEM-EDX) was used to characterize the cladode surfaces and the nature of dust deposit, and the lead isotopes were analyzed to identify the origin of Pb. The results show that (i) Ofi readily bioaccumulates Pb, (ii) the lead isotopic composition of cladodes evidences a foliar pathway of lead into Ofi and identifies the relative contributions of local Pb sources, and (iii) an evolution of air quality is recorded with successive cladodes, which makes Ofi a potential biomonitor to be used in environmental and health studies.

Efficacy of cactus pear (Opuntia ficus-indica) varieties as a source of food and feed in Endamehoni District, northern Ethiopia.

Cactus Pear ( Opuntia ficus-indica ) serves as a source of food, feed, as well as a means of additional income to the people in northern Ethiopia. The species has different varieties with varied rate of productivity and palatability. This study was conducted to assess the food and feed value of the Cactus Pear varieties in Endamehoni District, northern Ethiopia. It was carried out in three “Tabias” (small administrative sub-districts) and nine sub-Tabias (also known as “Kushets”). These areas were purposively selected as they are well known for their good diversity of Cactus varieties. A total of one hundred and twenty households (n = 120) were randomly selected for interviews using a semi-structured interview. This was supplemented by information obtained through key informants’ interviews and field visits. The local farmers made the identification and characterization of cactus varieties traditionally based on the outstanding phenotypes like fruit characteristics, seed size and content, cladode descriptors and plant height. A total of 13 Cactus varieties were identified and recorded. To assess the food and feed value and to select the best used varieties in the area, preference rankings were made. It was found that not all varieties were equally edible and palatable. The most serious constraints in palatability were associated with fruit taste, fruit size, presence of spines, and hardness of seeds. The ethnobotanical study on palatability was supplemented by chemical analysis on Dry Matter (DM), Ash, Organic Matter (OM) and Crude Protein (CP) content from two-year-old young cladodes of four Cactus varieties (Kille, Wadwada, Magalla and Limo) that are more preferred for food and feed. The analysis of variance (ANOVA) showed significant differences in DM, OM and Ash content at p<0.05 (not at p<0.10). There was no variation in crude protein content amongst the varieties tested. From the results, it is concluded that propagation and use of varieties Kille, Limo, Magalla and Wadwada by farmers of the area is advisable. Keywords: Characterization, Cladode, Community, ethnobotany, fruit, palatability, preference, propagation, traditional knowledge

Biophysical and physiological characteristics of nopalitos (Opuntia spp., Cactaceae) as influenced by domestication

A gradient of domestication based on morphological characteristics has been observed in species of Opuntia genus; but, a biophysical and physiological gradient have been not described. This study aimed to quantify biophysical and physiological characteristics in nopalitos (edible young cladodes of flat-stemmed spiny cacti) of 15 variants of five Opuntia species in a domestication gradient, including wild (O. streptacantha Lem.), semi-wild (O. hyptiacantha F.A.C. Weber), semi-domesticated (O. megacantha Salm-Dyck), more domesticated (O. albicarpa Scheinvar), and that recognized with the highest degree of domestication [O. ficus-indica (L.) Miller]. We hypothesized that biophysical and physiological characteristics of nopalitos are modified throughout the domestication process and that such changes will be evident when variants with different degree of domestication are grown under drought conditions. Tissue firmness, water potential (ψW), osmotic potential (ψπ), content of total soluble solids (TSS), content of malic acid, and membranes permeability were evaluated in nopalitos harvested from plants after 60 days without watering. A randomized design, with six replications and a plant as experimental unit by treatment, was used. The data were analysed by ANOVA, multiple comparisons of means by Tukey test (P ≤ 0.05) among variants and species, and multivariate analysis of principal components. Firmness (from 4.6 ± 0.082 to 6.8 ± 0.053 kgf/cm2), TSS (from 4.86 ± 0.105 to 6.33 ± 0.142 °Brix), membrane permeability (from 26 ± 0.712 to 31 ± 0.474 (µS cm−1)/g−1) and malic acid content (from 0.61 ± 0.008 to 0.82 ± 0.014 g 100 g−1 wet matter) increased during domestication, and simultaneously ψW and ψS decreased (from −0.85 ± 0.034 to −1.1 ± 0.030 MPa and −1.01 ± 0.011 to −1.15 ± 0.011 MPa). The firmness, SST and ψS are the main characteristics that contributed to separate the Opuntia species on the gradient of domestication. Opuntia species on a domestication gradient based on morphometric characteristics of plants (cladodes, nopalitos and cactus pear fruits) correlate to biophysical and physiological characteristics of nopalitos.

Phytochemical characterization and antioxidant activity of the cladodes of Opuntia macrorhiza (Engelm.) and Opuntia microdasys (Lehm.).

The genus Opuntia (Cactaceae) includes different plants well adapted to arid and semi-arid zones. These species are cultivated under restricted growth conditions, not suitable for the growth of other fruits/vegetables. The cactus pear is a well-known example inside Opuntia genus. Its young cladodes, also known as nopalitos, are frequently consumed and used in folk medicine due to their beneficial effects and phytochemical composition. Herein, hydrophilic and lipophilic extracts from cladodes of Opuntia microdasys and Opuntia macrorhiza were characterized. Furthermore, their antioxidant properties were compared to the corresponding phytochemical profile. Despite the phylogenetic proximity and similar geographical origin O. microdasys and O. macrorhiza showed significant differences in sugars, organic acids, phenolic compounds, fatty acids and tocopherols profiles. In particular, O. microdasys distinguished for having high contents in fructose, glucose, C6:0, C8:0, C12:0, C14:0, C14:1, C16:0, C18:3, C20:0, C22:0, C23:0, C24:0, SFA and tocopherols, and also for its higher DPPH EC50 values. O. microdasys by its side proved to have significantly higher amounts of trehalose, organic acids, C13:0, C16:1, C17:0, C18:0, C18:1, C20:1, C20:2 and MUFA. The obtained phytochemical profiles might be considered as useful information to select the best Opuntia species regarding a determined application of its natural extracts/isolated compounds.

Micropropagation of tuna (Opuntia ficus – indica ) and effect of medium composition on proliferation and rooting

The goal of this study was to determine micropropagation system for a mass production of Tuna (Opuntia ficus – indica). For this reason, explants dissected from strilled young cladodes successfully established on Murashige and Skoog (MS) medium supplemented with 5 mg l -1 Benzyl amino purine (BAP). MS medium containing different combinations of BAP (5 mg l-1) and Indole acetic acid (IAA) (0, 0.25, 0.5, 1, 2 mg l-1) , BAP (5 mg l-1) and Naphtalene acetic acid (NAA) ( 0, 0.25, 0.5, 1, 2 mg l -1 ) and BAP (0.5 and 1 mg l -1 ) and Kinetin ( 0.5 and 1 mg l 1 ) were tested for shoot development . The best results for shoot development and elongation were obtained in media containing 5 mg l-1 and 0.25 mg l-1 NAA. The highest multiplication rate (3.9) was observed in media supplemented with 5 mg l -1 BAP and 2 mg l -1 of NAA. Satisfactory rooting was achieved in MS Basal medium (5-6 cm length) without callus formation. The percentage of rooting was 100% and Plants were successfully established in a mix of pit and perlite (2:1) (100%) and acclimatization accomplished under greenhouse condition. In this study, total concentration of carbohydrates and proteins were measured in in vivo cultured (control) and in vitro propagated tuna after 3 months. As a result, no significant differences were observed between control and micropropagated tuna in protein concentration. Whereas carbohydrate content in micropropagated plants (3.24 mg g -1 ) was 2- fold that of the control plants (1.52 mg g -1 ).

Diversity of Unavailable Polysaccharides and Dietary Fiber in Domesticated Nopalito and Cactus Pear Fruit (Opuntia spp.)

The aim of this study was to quantify mucilages, pectins, hemicelluloses, and cellulose of nopalitos (edible, as vegetable, young cladodes of flat-stemmed spiny cacti) of most consumed Mexican cultivars, and sweet and acid cactus pear fruits of Opuntia spp. The hypothesis is that, regardless of their unavailable polysaccharides diversity, nopalitos and cactus pear fruits are rich sources of soluble and insoluble dietary fiber. Twelve cultivars of Opuntia spp. were used. Nopalitos had a significant variation in structural polysaccharides among the cultivars: mucilages (from 3.8 to 8.6% dry matter (DM)) averaged near a half of pectins content (from 6.1 to 14.2% DM) and tightly bound hemicelluloses (from 2.2 to 4.7% DM), which were the less abundant polysaccharides, amounted 50% of the loosely bound hemicelluloses (from 4.3 to 10.7% DM). Acid fruits (or 'xoconostle') had significantly higher unavailable polysaccharides content than sweet fruit, and contain similar proportions than nopalitos. Unavailable polysaccharides represent a high proportion of dry tissues of nopalitos and cactus pear fruits, composition of both of these soluble and insoluble polysaccharides (total dietary fiber) widely vary among cultivars without an evident pattern. Nopalitos and cactus pear fruit can be considered an excellent source of dietary fiber.

Effects of domestication on structural polysaccharides and dietary fiber in nopalitos (Opuntia spp.)

Domestication is an evolutionary process that modifies morphological, physiological, chemical and genetic features of wild plants, and is a product of artificial selection. A gradient of domestication can be appreciated in species of the Opuntia genus. There are wild species like O. streptacantha, others semi-domesticated like O. hyptiacanthaO. megacantha and O. albicarpa, or other like O. ficus-indica with the highest degree of domestication. At the same time, some structural polysaccharides have been associated with plant drought and freeze resistance. This study aimed at quantify five groups of structural polysaccharides in 14 variants of nopalitos (edible young cladodes of flat-stemmed spiny cacti consumed as vegetables) of Opuntia spp. in a domestication gradient. Given that extreme environment is less frequent under cultivation than in the wild, we hypothesized that structural polysaccharides in nopalitos are reduced throughout the domestication process. The experiment was conducted as a completely randomized design with a 5 × 14 two-factor factorial treatment structure, with four replications. Mucilages, pectins, hemicelluloses and cellulose were extracted and dietary fiber content was calculated. ANOVA of data was performed, pair wise comparisons of species and variants were conducted by Tukey test and principal components analysis was carried out. Mucilages were higher in O. ficus-indica (12% of total dry mass) and pectins, loosely and tightly bound hemicelluloses in O. streptacantha (2.5, 9 and 3%, respectively). All five species had similar content of cellulose (5.1% dry mass); but, O. streptacantha presented more dietary fiber. Higher pectins and both loosely and tightly bound hemicelluloses content in wild species can be related to genotype and the persistence of reaction to a wild environment. Mucilages which are one of the main characteristics of consumer preference and loosely bound hemicelluloses could have impact for selection during Opuntia domestication.

Study of carbohydrates present in the cladodes of Opuntia ficus-indica (fodder palm), according to age and season

Types and content of carbohydrates were evaluated by chemical (spectrophotometric assay) and physicochemical (Thin Layer Chromatography - TLC and High Performance Liquid Chromatography - HPLC) methods in some Opuntia ficus-indica varieties according to age and season. The samples comprised four varieties of palm (giant, copena F1, clone 20, and round palm). The results demonstrated that the four varieties of palm contain a good quantity of neutral and acid sugars in both summer and winter seasons. However, samples collected in the summer presented a higher content of carbohydrates specially glucose, fructose, galactose, xylose, and arabinose. The tertiary cladode (old cladode) presented almost the double content of sugar found in the quaternary cladodes (young cladodes). Pectic polysaccharides were sequentially extracted with water at 60 oC, and EDTA at 60 oC solution resulting in Water-Soluble Pectin (WSP) and a Chelating-Soluble Pectin (CSP) respectively. Galacturonic acid was detected in the fractions WSP and CSP. However, the fraction CSP presented the highest content of sugar acid.

Salt stress increases the expression of p5cs gene and induces proline accumulation in cactus pear

Proline (Pro) is one of the most accumulated osmolytes in salinity and water deficit conditions in plants. In the present study, we measured the Pro content, the activity and the expression level of delta 1-pyrroline-5-carboxylate synthetase (P5CS: γ-glutamyl kinase, EC 2.7.2.11 and glutamate-5-semialdehyde dehydrogenase, EC 1.2.1.41), a key regulatory enzyme involved in the biosynthesis of Pro, in cactus pear ( Opuntia streptacantha) subjected to 6, 9 and 11 days of salt stress. Treatment with NaCl of O. streptacantha young plants resulted in a decrease in the cladode thickness and root length, and in a significant and gradual accumulation of Pro in young cladodes, in a time- and concentration-dependent manner. P5CS activity, studied as γ-glutamyl kinase, was reduced at all times as a consequence of salt treatment, except at the sixth day at 75 and 150 mM of NaCl, where a slight increase was observed. We isolated an open reading frame (ORF) fragment of p5cs gene. The deduced amino acid sequence of the P5CS protein exhibited 90.4% of identity with the P5CS protein from Mesembryanthemum crystallinum. RT-PCR analysis revealed that the Osp5cs gene of O. streptacantha was induced by salt stress at 9 and 11 days of treatment. Furthermore, ABA-induced Osp5cs gene expression was observed in cladodes of cactus pear young plants. We observed an evident correlation between the transcript up-regulation and the Pro accumulation under salt stress; however, these results do not parallel with the changes in P5CS enzymatic activity. This Pro accumulation might function as an osmolyte for the intracellular osmotic adjustment and might be playing a critical role in protecting photosynthetic activity in O. streptacantha plants under salt stress.

On the nature of facultative and constitutive CAM: environmental and developmental control of CAM expression during early growth of Clusia, Kalanchoe, and Opuntia

The capacity to induce crassulacean acid metabolism developmentally (constitutive CAM) and to up-regulate CAM expression in response to drought stress (facultative CAM) was studied in whole shoots of seven species by measuring net CO(2) gas exchange for up to 120 day-night cycles during early growth. In Clusia rosea, CAM was largely induced developmentally. Well-watered seedlings began their life cycle as C(3) plants and developed net dark CO(2) fixation indicative of CAM after the initiation of the fourth leaf pair following the cotyledons. Thereafter, CAM activity increased progressively and drought stress led to only small additional, reversible increases in dark CO(2) fixation. In contrast, CAM expression was overwhelmingly under environmental control in seedlings and mature plants of Clusia pratensis. C(3)-type CO(2) exchange was maintained under well-watered conditions, but upon drought stress, CO(2) exchange shifted, in a fully reversible manner, to a CAM-type pattern. Clusia minor showed CO(2) exchange reponses intermediate to those of C. rosea and C. pratensis. Clusia cretosa operated in the C(3) mode at all times. Notably, reversible stress-induced increases of dark CO(2) fixation were also observed during the developmental progression to pronounced CAM in young Kalanchoë daigremontiana and Kalanchoë pinnata, two species considered constitutive CAM species. Drought-induced up-regulation of CAM was even detected in young cladodes of a cactus, Opuntia ficus-indica, an archetypal constitutive CAM species. Evidently, the defining characteristics of constitutive and facultative CAM are shared, to variable degrees, by all CAM species.

First detection of ?Candidatus Phytoplasma australiense? in Liquidambar styraciflua in Australia

The Christmas cactus ( Zygocactus truncatus ) is a perennial succulent and is native to the South American tropics of Brazil. It is one of the most widely cultivated plants and healthy Christmas cacti in full bloom are often sold as holiday plants around the Christmas season. A pot containing a Christmas cactus with cladodes showing conspicuous malformed branches giving witches’ broom symptoms and stunted growth was found at the Agricultural University, Kunming. Based on the symptomatology, a phytoplasma was suspected to be associated with the disease. Samples from the plant exhibiting witches’ broom and symptomless plants were analyzed for the presence of phytoplasmas using 16SrDNA PCR assays. Total nucleic acids were extracted from young cladodes using the DNeasy Plant Mini kit (QIAGEN) and were used as the template in a nested PCR reaction employing primers R16mF2/R16mR1 followed by R16F2/R16R2 (Gundersen & Lee, 1996). A water control was also included in all the PCR reactions to check for any contamination. A DNA fragment of the expected size (1·2 kb) was amplified by nested PCR from the diseased plant but not from symptomless plants, nor from the water control. After cloning and sequencing, the DNA sequence of 1249 bp was deposited in GenBank (Accession No. AY647459). Phylogenetic analysis of the 16S rDNA sequence delineated the Christmas cactus witches’ broom phytoplasma as a Peanut witches’-broom (16SrII) group member (putative Candidatus Phytoplasma aurantifolia), most closely related to the faba bean phyllody phytoplasma (Lee et al ., 1998). This is the first report of a phytoplasma disease of the Christmas cactus.

First report of a phytoplasma associated with Christmas cactus witches’ broom

The Christmas cactus (Zygocactus truncatus) is a perennial succulent and is native to the South American tropics of Brazil. It is one of the most widely cultivated plants and healthy Christmas cacti in full bloom are often sold as holiday plants around the Christmas season. A pot containing a Christmas cactus with cladodes showing conspicuous malformed branches giving witches’ broom symptoms and stunted growth was found at the Agricultural University, Kunming. Based on the symptomatology, a phytoplasma was suspected to be associated with the disease. Samples from the plant exhibiting witches’ broom and symptomless plants were analyzed for the presence of phytoplasmas using 16SrDNA PCR assays. Total nucleic acids were extracted from young cladodes using the DNeasy Plant Mini kit (QIAGEN) and were used as the template in a nested PCR reaction employing primers R16mF2/R16mR1 followed by R16F2/R16R2 (Gundersen & Lee, 1996). A water control was also included in all the PCR reactions to check for any contamination. A DNA fragment of the expected size (1·2 kb) was amplified by nested PCR from the diseased plant but not from symptomless plants, nor from the water control. After cloning and sequencing, the DNA sequence of 1249 bp was deposited in GenBank (Accession No.AY647459). Phylogenetic analysis of the 16S rDNA sequence delineated the Christmas cactus witches’ broom phytoplasma as a Peanut witches’-broom (16SrII) group member (putative Candidatus Phytoplasma aurantifolia), most closely related to the faba bean phyllody phytoplasma (Lee et al., 1998). This is the first report of a phytoplasma disease of the Christmas cactus. This work was supported by the National Natural Science Foundation of China (grant no. 30260064) and the Science Fund of Yunnan Province (grant no. 2000C0014Q).