Isolation, identification and quantitation of hydroxycinnamic acid conjugates, potential platform chemicals, in the leaves and stems of Miscanthus × giganteus using LC-ESI-MSn

Miscanthus × giganteus is a source of platform chemicals and bioethanol through fermentation. Cinnamates in leaves and stems were analysed by LC–ESI-MSⁿ. Free phenols were extracted and separated chromatographically. More than 20 hydroxycinnamates were identified by UV and LC–ESI-MSⁿ. Comparative LC–MS studies on the leaf extract showed isomers of O-caffeoylquinic acid (3-CQA, 4-CQA and 5-CQA), O-feruloylquinic acid (3-FQA, 4-FQA and 5-FQA) and para-coumaroylquinic acid (3-pCoQA and 5-pCoQA). Excepting 3-pCoQA, all were also detected in stem. 5-CQA dominated in leaf; a mandelonitrile–caffeoylquinic acid dominated in stem. Three minor leaf components were distinguished by fragmentation patterns in a targetted MS² experiment as dicaffeoylquinic acid isomers. Others (M_r 516) were tentatively identified as hexosylcaffeoyl-quinates. Three positional isomers of O-caffeoylshikimic acid were minor components. p-Hydroxybenzaldehyde was also a major component in stem. This is the first report of the hydroxycinnamic acid profile of leaves and stems of M. × giganteus.     

Photooxidative Stress Modulation of Endogenous Phytohormone and Antioxidant Accumulations and Fruit Maturity in Date Palm (Phoenix dactylifera L.)

A common modern cultivation practice is bagging the fruit bunch if date palm (Phoenix dactylifera L.) which may influence fruit maturity and nutraceutical quality. Exposure of fruits to photooxidative stress induces changes in the endogenous concentrations of plant hormones and other metabolites, which may cause accelerated fruit maturity. This study was conducted to examine the effect of exposure to direct and indirect sunlight on date palm fruit development. The indirect sunlight treatment was simulated by fruit bunch bagging, a common practice in modern date production. The exposure of date palm fruits to direct sunlight-induced photooxidative stress causing an increased concentration of ascorbic acid and decreased content of chlorophyll, anthocyanins, carotenoids, and phenols compared to the fruit bagging treatment. Direct sunlight also reduced the concentration of phytohormones, including indoleacetic acid, gibberellin, and zeatin, but increased abscisic acid accumulation. The directly-exposed fruits reached a partially-mature stage (Rutab) in August, whereas the bagged fruits remained at the immature stage (Khalal). This study is the first to describe the biochemical basis of the observed improvement of date palm fruit development in response to reduced light intensity. Besides, it provides insights into controlling date palm fruit maturity and subsequently prolonging the shelf life dates on the tree; thus, extending the marketing period for the benefit of the farmers.

     

Relationship between Cell Wall Susceptibility to Cellulases and Pectinases of Fusarium oxysporum and Susceptibility of Date Palm Cultivars

Fusarium oxysporum f. sp. albedinis, the bayoud disease agent of date palm, grows on a mineral medium containing the cell walls of date palm roots as a sole carbohydrate source. The growth and development of pathogen under these conditions was related to the production of extracellular cell wall-degrading enzymes (CWDE): cellulases, polygalacturonases, polygalacturonate transeliminases, and pectinmethylesterases. The mycelial growth and the sporulation of pathogen were higher in the presence of cell walls of susceptible cultivars (BFG, JHL, BSK) than in the presence of those of resistant cultivars (IKL, SLY, BSTN). After 8 d of fungal culture, the activity of CWDE was equal whatever is the origin of the cell walls (resistant or susceptible cultivars). After 16 d of culture, the activity of these CWDE was higher when the parasite was cultivated on the cell walls of the susceptible cultivars than on those of the resistant cultivars. A positive correlation was observed between CWDE activities and the growth and the sporulation of F. oxysporum after 16 d of culture. These results clearly show a relation between the susceptibility/resistance of the cell walls of the roots of the date palm to the parasitic CWDE and the susceptibility/resistance of the cultivars.     

Differential effects of light and fungal elicitor on chlorogenic acid and caffeoylshikimic acid metabolism

Upon irradiation an increase in the extractable activity of hydroxycinnamoyl-CoA:d-quinate hydroxycinnamoyl transferase (CQT) and a decrease in the activity of hydroxycinnamoyl-CoA:shikimate hydroxycinnamoyl transferase (CST) was observed in cell suspension cultures and seedling of carrot (Daucus carota L). Conversely, CST was induced and CQT repressed in the cell cultures upon treatment with fungal elicitor (i.e. cell wall preparations from Phytophthora megasperma). In the cell cultures irradiation led to a continuous accumulation of 5-O-caffeoyl-d-quinic (chlorogenic) acid, while a transient accumulation of 5-O-caffeoylshikimic acid took place in response to elicitor treatment. Cell wall bound 4-hydroxybenzoic, 4-coumaric and also ferulic acid were increased after treatment with Pmg elicitor. These wall bound phenolics may be involved in protection against microbil attack.     

Webbing life type and behavioral response of the date palm mite, <Emphasis Type="Italic">Oligonychus afrasiaticus</Emphasis>, to webbing residues on leaves and fruits of date palm

The present study examined the webbing life type and behavioral response of date palm mite (DPM) Oligonychus afrasiaticus (McGregor) toward its webbing residues, in presence or absence of immature individuals, on leaves and fruits of date palm. DPM exhibited a complicated web life type on both the leaves and fruits of date palm, which is characterized by the following features: complicated irregular web structure; webbing density on the leaves is medium and on the fruits extremely high; eggs are always laid on the threads of web without web cover; fecal pellets are deposited on the substrate surface (both leaf and fruit); quiescent stages are on the silken web threads; and the preferred site for feeding and walking is under the web. Behavioral observation of DPM females revealed that the midrib of leaves and the base of fruits of date palm were the preferred sites for feeding and web construction. The number and development stage of DPM affected the behavioral response of females on date palm leaves but not on the fruits. DPM spent most time feeding under the web on both the leaves and fruits of date palm.     

The effect of biofungicide Mycostop on Ceratocystis radicicola, the causal agent of black scorch on date palm

In vitro studies were conducted to assess the efficacy of the biofungicide Mycostop on the fungus Ceratocystis radicicola (= Ceratostomella radicicola), which causes black scorch on date palm (Phoenix dactylifera) and the effect salinity on the growth of Streptomyces griseoviridis, the activecomponent of Mycostop. C. radicicola wasisolated from roots of diseased date palm inlandscape settings, nurseries, andplantations. In vitro, the effectiveconcentration (EC₅₀) of Mycostop thatreduced the mycelial growth by 50% was 0.35 g/l of PDA. Mycostop at the rate of 0.35 g/lor greater reduced spore germination,plasmolyzed germlings and reduced sporulationof C. radicicola. In essence it reducedthe inoculum potential of C. radicicola. In sterile soil treated with Mycostop, rootpieces of date palm inoculated with C.radicicola were less necrotic than those inuntreated soil. Streptomycesgriseoviridis in Mycostop did not grow on PDAamended with 5.0% NaCl (–4.28 MPa), but thenumber of colony formed units on PDA amendedwith 1.0 to 4.0% NaCl was not significantlydifferent from a factor that was considered ifMycostop is to be used to manage black scorchin Kuwait. The observations showed the effectsof Mycostop on C. radicicola inlaboratory studies but its efficacy on blackscorch disease of date palms in nurseries andplantations is yet to be determined.     

Enhancement of Defence Responses against Bayoud Disease by Treatment of Date Palm Seedlings with an Hypoaggressive Fusarium oxysporum Isolate

Pretreatment of date palm seedlings with an hypoaggressive Fusarium isolate (AHD) protected them partially from further infection by Fusarium oxysporum f.sp. albedinis (Foa), the Bayoud disease pathogen. No mortality occurred during 2–3 months of incubation in plants pretreated with AHD, as opposed to aggressive isolate (ZAG) inoculated controls where up to 100% mortality was observed 15–30 days after inoculation. Such protection involved biochemical interactions between the host plant and AHD since no direct competition or antagonism was revealed between AHD and ZAG. The examination of the accumulation of phenolics and peroxidase activity, two parameters previously reported to be involved in date palm resistance to Foa, indicated that the response to AHD was correlated with the ability of pretreated date palm tissues to establish a faster defence response in the roots of both susceptible and resistant cultivars. Plants pretreated with AHD accumulated higher amounts of phenolics, mainly non‐constitutive hydroxycinnamic acid derivatives, which play a crucial role in date palm defence against Foa, as previously described by our group. These compounds were accumulated along with the constitutive caffeoylshikimic acids. A faster induction of peroxidase activity in response to Foa was also recorded in plants pretreated with AHD. Given the multi‐component nature of these induced responses, AHD could be part of integrated disease management strategies for a sustainable control of the palm tree Bayoud disease.     

Monitoring of cultivar identity in tissue culture-derived date palms using RAPD and AFLP analysis

In the present study, two polymerase chain reaction (PCR)-based methods namely, randomly amplified polymophic DNA (RAPD) and amplification fragment length polymorphism (AFLP) were employed to assess genetic variations, which may appeared, in tissue culture-derived date palm (Phoenix dactylifera) offshoots. Analysis of RAPD banding patterns generated by PCR amplification using 37 random primers gave no evidences for somaclonal variations and the percentage of polymorphic bands in a total of 259 scored bands was zero. Meanwhile, analysis of AFLP banding patterns generated using 13 primer combinations pointed to minor genetic variations in the AFLP banding patterns. The percentage of genetic variations (polymorphism) in tissue culture-derived date palm offshoots belonging to cultivars Sakkoty, Gandila and Bertamoda was 2.6, 0.79 and 1 %, respectively, as revealed by AFLP analysis. The low percentage of genetic variations confirms the genetic stability of tissue culture-derived dry date palm cultivars.     

4-O-Caffeoylshikimic and 4-O-(p-Coumaroyl)shikimic Acids from the Dwarf Tree Fern,Dicksonia antarctica

Two derivatives of shikimic acid were isolated from croziers of the dwarf tree fern, Dicksonia antarctica, and their structures were elucidated as 4-O-caffeoylshikimic acid and 4-O-(p-coumaroyl)-shikimic acid on the basis of mass spectrometric and NMR spectroscopic evidence.     

Oil palm (Elaeis guineensis) protoplasts: isolation, culture and microcallus formation

Procedures are deseribed for the efficient isolation of protoplasts from a variety of oil palm (Elaeis guineensis Jacq.) tissues. Various factors including donor source, composition of enzyme mixture and culture medium affected the yield and viability of the protoplasts Polyembryogenic cultures of oil palm were the most suitable starting material in terms of yield, viability and metabolic competence. Pectolyase Y-23 in association with cellulase and hemicellulase was required for the efficient release of protoplasts from the oil palm tissues. Limited cell division to form microcallus was observed at very low frequency (<0.01%) when glutathione and catalase were incorporated in the culture medium.Abbreviations 2,4-d dichlorophenoxyacetic acid - DTT dithiothreitol - MES 2[N-morpholino] ethanesulphonic acid - NAA 1-naphthalene acetic acid - PVP polyvinylpyrrolidone     

Callogenesis and rhizogenesis in date palm leaf segments: are there similarities between the two auxin-induced pathways?

In date palm (Phoenix dactylifera L. cv. Ahmar, Arecaceae), as for many monocotyledons, callogenesis is a prerequisite for the initiation of somatic embryogenesis, and requires the presence of auxin in the medium. Immature leaf explants were cultivated in medium supplemented with either 1 or 54 μM 1-naphtaleneacetic acid in order to induce either rhizogenesis or callogenesis. Histological studies performed throughout the culture period established that precocious cell reactivation is similar in both morphogenetic pathways. Early cytological modifications are associated with cell reactivation and are observed in the pluripotent cells of perivascular sheaths. Divergence between the callogenesis and rhizogenesis pathways is observed later, during the subsequent determination and morphological differentiation phases. We established that in date palm, the rhizogenesis and callogenesis pathways are initiated from the same cell type, the ultimate developmental fate depending upon auxin concentration.     

A rapid and efficient method for the extraction of total DNA from mature leaves of the data palm (Phoenix dactylifera L.)

A method is presented for the rapid isolation of high-molecular-weight DNA from mature leaves of date palm (Phoenix dactylifera L.), using a CTAB-based buffer. The method yields up to 800 μg of DNA from 1 g of leaf tissues. The procedure was also suitable for DNA extraction from callus or buds from tissue culture. The DNA obtained through this method was a good substrate for at least seventeen restriction endonucleases. This method was also used to extract DNA from mature leaves of coconut and may be applicable to other species of palms.     

Chromatographic Characterization and Phytotoxic Activity of Fusarium oxysporum f. sp. albedinis and Saprophytic Strain Toxins

The bayoud disease, vascular fusariosis of date palm tree (Phoenix dactylifera L.), is caused by the pathogenic fungus Fusarium oxysporum f. sp. albedinis. The characteristic symptoms of the bayoud disease were elicited on detached leaves of F. oxysporum f. sp. albedinis‐susceptible cultivars of date palm trees, which were treated either with the F_II (F. oxysporum f. sp. albedinis) fraction purified from the organic extracts of a F. oxysporum f. sp. albedinis liquid culture, or with a solution of fusaric acid. Enniatins, which are secreted by several Fusarium species, were tested at different concentrations and were not capable of inducing symptoms on such detached leaves. The F_II (F. oxysporum f. sp. albedinis) fraction was unable to induce necrosis of potato slices, which indicates that it does not contain significant amounts of enniatins. The high‐performance liquid chromatography (HPLC) profiles of the F_II (F. oxysporum f. sp. albedinis) fraction showed toxic peaks different from fusaric acid. A fraction, named F_II (AZ₄), was obtained from culture filtrates of a saprophytic Fusarium strain maintained in the same cultural conditions as for the F. oxysporum f. sp. albedinis. The HPLC profile of the F_II (AZ₄) fraction did not show the characteristic phytotoxic peaks present in the F_II (F. oxysporum f. sp. albedinis) fraction. This finding well agrees with the fact that the F_II (AZ₄) fraction is not toxic to detached date palm leaves. Moreover, the HPLC profiles of F_II fractions obtained from other special forms of F. oxysporum are different the F_II (F. oxysporum f. sp. albedinis) profile. The phytotoxic compounds purified from the F_II (F. oxysporum f. sp. albedinis) fraction are probably new molecules that may help in understanding the pathogenesis of bayoud disease.     

Effect of wastewater stabilization ponds on antimicrobial susceptibility and haemolysin occurrence among motile Aeromonas strains

Aeromonas strains (total=953) isolated from raw wastewater, stabilization pond effluent and sediments were evaluated for their susceptibilities to 17 antibiotics and for their ability to produce haemolysins. Stabilization ponds did not seem to select highly resistant strains of aeromonads. There were no differences in the resistance patterns of isolates from raw sewage, stabilization pond effluent and sediments. All strains were found to possess multiple resistance, most commonly to ampicillin, amoxicillin and novobiocin. Almost 90% of the strains of A. hydrophila and A. caviae were resistant to cephalothin, whereas more than 80% of A. sobria isolates were found to be susceptible to this antibiotic. Resistance to trimethoprim, oxytetracycline, nalidixic acid, chloramphenicol, tetracycline, trimethoprim-sulphamethoxazol, polymyxin B, kanamycin or erythromycin among all isolates did not exceed 10%. Moreover, no strain was found to be resistant to gentamycin and only 9 of the 953 isolates exhibited resistance to cefotaxim. The percentage of haemolytic strains was significantly higher in the stabilization pond effluent than in raw sewage. This high incidence of haemolytic activity was connected with a high proportion of A. sobria whereas, in samples from the raw sewage or stabilization pond sediments a high proportion of A. caviae decreased the total amount of haemolytic aeromonads. The high incidence of haemolytic activity (+) was associated particularly with A. sobria (93.3%) and A. hydrophila (88.7%) whereas A. caviae was found to be the lowest haemolytic species (16.3%).B. Imziln and Y.M.O. Lafdal are with Cadi Ayyad University, Faculty of Sciences Semialia, Department of Biology, Laboratory of Microbiology, BP S/15 Marrakech, Morocco. M. Jana is with the Hôpital Millitaire Avicenne Marrakech, Morocco.     

Endophytic fungi of teak leaves Tectona grandis L. and rain tree leaves Samanea saman Merr.

Summary In order to study the foliar endophytes from teak (Tectona grandis L.) and rain tree (Samanea saman Merr.) growing in the campus of Chulalongkorn University, healthy leaves were collected at two-monthly intervals during January to December. The number of genera and species, together with their colonization frequency (CF%) in mature teak and rain tree leaves were greater than those in the young leaves. More endophytic isolates in the leaves of both trees were recovered during the rainy season. The fungal genera found in both young and mature teak leaves were Alternaria, Colletotrichum, Nigrospora, Phomopsis and mycelia sterilia. Phomopsis was the dominant genus in both young (newly emerged) and mature leaves. Fusarium, Penicillium, Schizophyllum commune and members of the Xylariaceae were found only in mature leaves. For the rain tree leaves, species of Phomopsis and mycelia sterilia were found in both young newly emerged and mature leaves. Colletotrichum and Penicillium were found only in mature leaves, whereas Nigrospora was found only in young newly emerged leaves. In this study, Phomopsis was the dominant genus in the leaves of both tree species. A total of 37 isolates of endophytic fungi isolated from teak and rain tree leaves were tested for the production of antimicrobial activities. Out of these, 18 isolates could produce inhibitory substances effective against Bacillus subtilis, Staphylococcus aureus and Escherichia coli and 3 isolates inhibited growth of Candida albicans in vitro.     

Flow cytometric analysis of the cell cycle in different coconut palm (<Emphasis Type="Italic">Cocos nucifera</Emphasis> L.) tissues cultured in vitro

We conducted a study of the cell cycle of coconut palm tissues cultured in vitro in order to regulate regeneration. Coconut palm is a plant for which it is difficult to monitor the ability of the meristematic cells to actively divide. Cell nuclei were isolated from various types of coconut palm tissues with and without in vitro culture. After the nuclei were stained with propidium iodide, relative fluorescence intensity was estimated by flow cytometry. Characterization of the cell cycle reinforced the hypothesis of a block in the G₀/G₁ and G₁/S phases of the coconut cells. A time-course study carried out on immature leaves revealed that this block takes place gradually, following the introduction of the material in vitro. Synchronization of in vitro-cultured leaves cells using 60 µM aphidicholin revealed an increase in the number of nuclei in the S phase after 108 h of treatment. The significance of these results is discussed in relation with the ability of coconut tissue cultured in vitro to divide.Communicated by P. Debergh     

Cultivation of oyster mushroom Pleurotus ostreatus on date-palm leaves mixed with other agro-wastes in Saudi Arabia

Promoting the use of agricultural waste is one of the newly prepared water and environment friendly agriculture strategies in the Kingdom of Saudi Arabia (KSA). The objective of this research was to study the efficiency of cultivating oyster mushroom (Pleurotus ostreatus) on date palm wastes mixed with other agricultural wastes available in KSA. Four agricultural wastes were mixed with date palm leaves at different ratios, with two supplements and three spawn rates were used. Wheat straw mixed with date palm at ratio of 25 (date palm): 75 (agro-waste) showed the best results in most of the parameters measured. Corn meal was superior over wheat bran as a supplement in all treatments. Parameter values increased with the increase of the spawn rate of P. ostreatus. Treatments with date palm leave wastes contained higher carbohydrates and fibers. No significant differences were found among the fruiting bodies produced on the different agro-wastes studied for the different proximates analyzed. Analyses of metal concentration showed that potassium was the highest in all the treatments tested followed by Na, Mg, Ca, and Zn. This is the first study that reported the success of growing oyster mushroom on date palm leaf wastes mixed with other agro-wastes obtainable in KSA.     

Biosynthesis and Characterization of Poly(3-hydroxybutyrate-co-3- hydroxyhexanoate) from Palm Oil Products in a Wautersia eutropha Mutant

Palm kernel oil, palm olein, crude palm oil and palm acid oil were used for the synthesis of poly (3-hydroxybutyrate-co-3-hydroxyhexanoate) [P(3HB-co-3HHx)] by a mutant strain of Wautersia eutropha (formerly Ralstonia eutropha) harboring the Aeromonas caviae polyhydroxyalkanoate (PHA) synthase gene. Palm kernel oil was an excellent carbon source for the production of cell biomass and P(3HB-co-3HHx). About 87% (w/w) of the cell dry weight as P(3HB-co-3HHx) was obtained using 5 g palm kernel oil/l. Gravimetric and microscopic analyses further confirmed the high PHA content in the recombinant cells. The molar fraction of 3HHx remained constant at 5 mol % regardless of the type and concentration of palm oil products used. The small amount of 3HHx units was confirmed by ¹³C NMR analysis. The number average molecular weight (M_n) of the PHA copolymer produced from the various palm oil products ranged from 27 0000 to 46 0000 Da. The polydispersity was in the range of 2.6–3.9.     

Role of native and exotic mycorrhizal symbiosis to develop morphological,physiological and biochemical responses coping with water drought of date palm,Phoenix dactylifera

Key Message

Arbuscular mycorrhizal (AM) symbiosis can improve date palm growth and alleviate drought-related impacts than non-mycorrhizal plants due to the ability of AMF for modifying plant metabolism and physiology.

Abstract

Date palm (Phoenix dactylifera L.) is an important agricultural and commercial crop in the North of Africa and Middle Eastern countries. During the last decade, date palm plantations were subjected to degradation due to an extensive exploitation and to drastic environmental conditions such as drought. Currently, there is a growing interest in the valorization of water due to environmental problems and economic aspects. The use of arbuscular mycorrhizal fungi (AMF) can offer a possibility to overcome these problems. The objective of this study was to study the influence of different Glomus species—Glomus intraradices, G. mosseae and Complex Aoufous (native AMF)—on the development of date palm grown under two water regimes (optimal irrigation, 75 % of field capacity or water deficit, 25 % of field capacity). Our results revealed that the beneficial effect of mycorrhizal symbiosis on plant growth depended on the fungal species and the water regime applied to the palm date seedling. While the native Complex Aoufous was the most effective in increasing the shoot height and biomass under well-watered conditions, G. intraradices was the most beneficial fungus for improving growth of plants that undergo restricted water supply. This positive effect of G. intraradices under drought conditions was not related to an enhancement of the antioxidant enzymatic activities in leaves; the association of palm date with G. intradices caused an increase in the elasticity of cell walls in leaves and allowed maintaining high water content in leaves without lowering leaf water potential under stressful conditions. The adequate selection of the AMF species is crucial for improving growth of palm date seedlings, and it must be in accordance with the water regime that will be applied to plants.