Isolation And molecular characterization of xylella fastidiosa from different host plant species in Italy (Apulia region)

L'isolement en culture pure de la souche Xylella fastidiosa liée au syndrome de déclin rapide de l'olivier (OQDS), récemment observé dans les Pouilles (sud de l'Italie) a été tenté à partir des plantes symptomatiques naturellement infectée, principalement Olea europaea L., Polygala myrtifolia et Rosmarinus officinalis, en utilisant les méthodes d'impression et d'extraction de la sève. Avant l'isolement, RT-LAMP et qPCR ont été utilisés pour déterminer la présence de X. fastidiosa chez tous les hôtes. Des cultures bactériennes pures ont été obtenues à partir d'extraits d'Olea europaea L. et de Polygala myrtifolia étalés dans un milieu tamponné d'extrait de cystéine-levure (BCYE). Deux isolats d'olive ont ensuite été typés à l'aide du système de séquençage multilocus (MLST). Les résultats indiquent qu'Olea europaea a un grand potentiel d'isolement, et la méthode d'impression a présenté des résultats nettement meilleurs. MLST montre des points communs génétiques avec la souche De Donno (ST53) et confirme que nous avons toujours le même type de séquence ST53 dans la région.Pure culture isolation of the Xylella fastidiosa strain related to the olive quick declinesyndrome (OQDS), lately noticed in Apulia (southern Italy) was attempted fromnaturally infected symptomatic plants mainly Olea europaea L., Polygala myrtifolia and Rosmarinus officinalis using printing and sap extraction methods. Prior to isolation, RTLAMP and qPCR were used to determine the presence of Xylella fastidiosa in all hosts. Pure bacterial cultures were obtained from both Olea europaea L. and Polygala myrtifolia extracts plated in buffered cysteine-yeast extract (BCYE) media. Two olive isolates were subsequently typed using the Multilocus Sequencing System (MLST). Findings indicate that beside Olea europaea, Polygala myrtifolia seems to have a good potential for isolation, and printing method has presented significantly better results. MLST shows genetic commonality with De Donno (ST53) strain and ascertain that Apulia region still have the same sequence type ST53 in the region

Germination et viabilité du Bromus rigidus sous l’action de différents traitements chimiques

Bromus are annual herbaceous plants belonging to the Poaceae family. It’s about the weeds that infest cereals. The most harmful one is Bromus rigidus Roth. The purpose of this study is to find out the effect of different treatments on the germination and viability of ripgut brome. The first test in vitro germination was carried out on a collection, from 1993 to 2015, under controlled conditions in temperature (25°C), photoperiods (eight hours of darkness and sixteen hours of light) and relative humidity (70%). Several treatments were used: gibberellic acid (1g/l), hot water (ranging from 60°C to 90°C), sodium azide (1g/0,75l), potassium nitrate (10g/l) and sulfuric acid 5%. The results obtained showed that the bromus seeds collected on 2013 germinated at a rate of 60% without any treatment and of 100% with the different treatments. Gibberellic acid is the most element promoting germination. The germination rate of bromus rigidus depends closely on the year of seed harvest and also on the treatment carried out. The viability test in vitro under the effect of tetrazolium chloride (C19H15ClN4) which is used only on non-germinated seeds, showed that the seeds remain viable for seven years and they germinate only if favorable conditions are met.Les bromes sont des plantes herbacées annuelles appartenant à la famille des Poaceae ; il s’agit des mauvaises herbes qui infestent les céréales et dont le plus nuisible est le brome rigide (Bromus rigidus Roth.). Cette étude a pour but de savoir l’effet de différents traitements sur la germination et la viabilité du brome. Le test de germination in vitro a été réalisé sur des graines de brome issues d’une collection de 1993 à 2015 et sous des conditions contrôlées en température (25°C), en photopériodes (huit heures d’obscurité et seize heures de lumières) et en humidité relative (70%). Plusieurs traitements sont testés, à savoir : l’acide gibbérellique (1g/l), l’eau chaude (60°C à 90°C), l’azoture de sodium (1g/0,75l), le nitrate de potassium (10g/l) et l’acide sulfurique 5%. Les résultats obtenus ont montré que les semences du brome de l’année 2013 ont germé à un taux de 60% sans aucun traitement et de 100% avec les différents traitements et plus précisément l’acide gibbérellique comme élément favorisant la germination. En effet, Le taux de germination du brome dépend étroitement de l’année de récolte des semences et aussi du traitement effectué. Le test de la viabilité in vitro des graines par traitement avec le chlorure de tétrazolium (C19H15ClN4) a montré une prolongation de la durée de la viabilité des graines qui peut aller jusqu’à sept ans, ainsi que la germination ne peut s’activer qu’en présence des conditions favorables

Characterization of plant growth-promoting mechanisms of rhizobacteria from the rhizosphere of date palms in tafilalet oases

Tafilalet is the largest oases in Morocco, but it is threatened by climatic and environmental stress like drought, soil degradation as well as plant diseases, in particular Fusarium wilt of date palm caused by Fusarium oxysporum f.sp albedinis (Foa). Soils are poor in terms of organic matter, nutrient concentrations and biological activity resulting in a dramatic decrease in date palm growth. Improving date palm productivity is regularly achieved by the excessive application of chemical fertilizers, which are harmful for human health and environment. In this work, we aimed at screening the plant growth-promoting rhizobacteria (PGPRs) and at the same time acting as biocontrol agents against Foa. Rhizobacteria associated with date palm and spontaneous plants in Tafilalet oases have been isolated and functionally characterized. Several plant growth promoting and biocontrol traits were investigated, including nutrients solubilization, and the production secondary metabolites and enzymes. Results showed that 97 % of strains was effective in N-fixation, 36 % in ammonia production, 90 % in P-solubilization. The Siderophores and Indol 3-acetic acid production were observed at 30 and 32 strains respectively. Besides, 78 % of strains had an inhibitory effect against Foa, in which 42 % completely inhibited the mycelia growth on PDA medium. Several strains produced cellulase, protease, amylase, chitinase and other secondary metabolites like hydrogen cyanide. Based on these results, consortia of efficient PGPRs could be selected and used as efficient microorganisms for future greenhouse and field experiments

Fertiledatepalm – a transdisciplinary collaboration project to ameliorate date palm cultivation via microbial inoculation, organic matter management and mixed cropping using nurse plants

Date palm is an important crop in Morocco, Tunisia and other drylands with a high agricultural, economic and cultural value. Harsh environmental conditions of those areas, further accelerated by climate change and the spread of root diseases, threaten date palm cultivation. To overcome limitations in productivity, high inputs of mineral fertilizers and pesticides are applied. However, these external inputs strongly affect the environment and livelihoods. The project aims at establishing an integrated microbe-assisted fertilization approach, combining the inoculation of native soil microbes, namely arbuscular mycorrhizal fungi (AMF) and plant growth-promoting rhizobacteria (PGPR) during the different date palm growth stages, with adapted agricultural management practices using organic amendments and mixed-cropping in Morocco and Tunisia. As initial step, we established a culture collection of native microbes, isolated from date palm roots and rhizosphere composed of 24 AMF isolates including eight species from six genera, twelve bacterial endophyte isolates composed of Paenibacillus, Mycobacterium, and Achromobacter species and 34 PGPR isolates. Functional characterization of PGPRs revealed that around 50 % can solubilize phosphorus and potassium and between 9 % and 68 % have the ability to produce siderophores, hydrogen cyanid, chitinase, cellulase, amylase and protease. Consortia of microbes were formed and used for inoculations. Experiments under nursery conditions revealed that inoculation with AMF and PGPR combined with compost significantly increased growth of date palms as compared to non-amended controls enabling farmers to decrease the time prior to field transplantation. On-farm trials performed in productive date palm groves have shown that PGPR inoculation with or without mixed-cropping with sorghum as nurse plants significantly increase fruit characteristics such as fruit flesh weight as well as fruit length and diameter for up to 14 % and leaf macronutrient concentrations for up to 200 % while in addition enhancing the mycorrhizal potential of the soil. Our integrated fertilization approach is developed in a participatory approach with key stakeholders in so-called innovation platforms, working at laboratory, on-station and on-farm scale to best tackle farmers’ needs in order to facilitate adoption and implementation

Effect of PGPR and mixed cropping on mycorrhizal status, soil fertility, and date palm productivity under organic farming system

A field study was carried out for two years at an organic farm under arid climate in Morocco to investigate the effect of an integrated biofertilization approach on Arbuscular Mycorrhizal Fungal (AMF) abundance and infectivity, soil fertility, yield, and fruit quality of date palm. The biofertilization approach included three management practices namely application of compost, inoculation with a consortium of native PGPR strains originally isolated from date palms of Drâa-Tafilalet region (Pseudomonas koreensis, Serratia nematodiphila, S. marcescens, and Klebsiella sp.) and using mixed-cropping with sorghum. Accordingly, four treatments were established in this study: 1) mixed-cropping with sorghum, 2) PGPR inoculation, 3) sorghum + PGPR, and 4) control (without sorghum or PGPR). All treatments received compost as organic amendment. Results revealed that mixed-cropping with sorghum significantly increased AMF colonization intensity and spore density by more than 50% and 29%, respectively. Sorghum association also resulted in a significant increase in organic matter concentrations of up to 2.95% against 2.45% in monocropping soils. The integrated biofertilization approach resulted in the highest yield with an increase rate of 10.6% and 12.1% in the first and the second year, respectively compared to date palms receiving compost alone. Similarly, the mineral composition and quality characteristics of date fruits were significantly improved. The enhancement of soil fertility and date palm productivity under harsh environmental conditions represents a first step towards the adoption of sustainable practices in the region and in similar areas

Development of SCAR markers and a semi-selective medium for the quantification of strains Ach 1-1 and 1113-5, two Aureobasidium pullulans potential biocontrol agents.

peer reviewe

Rendement, caractérisation phytochimique et activité anti-Allorhizobium vitis des huiles essentielles de quatre espèces marocaines d'Eucalyptus

This study aims to investigate the yield, chemical composition and inhibitory effect of four essential oils of eucalyptus collected from Morocco on the pathogenic Allorhizobium vitis S4 causal agent of the crown gall of grapevine. The identification of the constituents of essential oils was performed by GC-MS analysis and the antibacterial activity was evaluated using an aromatogram test. The yield of different samples of eucalyptus ranged from 2-4.5%. The results of chemical compounds analysis show that the major compounds of eucalyptus essential oils are, 1,8-Cineol (Eucalyptol) (65.55-78.11%), α-Pinene (3-15.21%), 2-β-Pinene (2.33-5.27%) and p-Cymene (0.79-11.51%). All eucalyptus essential oil exhibits an antibacterial activity in vitro against Allorhizobium vitis strain S4 with a percentage of inhibition and minimal inhibitory concentration values in the range of 13.67-20.50 % and 20-40 mg/mL, respectively.Ce travail de recherche vise à étudier le rendement, la composition chimique et l'effet inhibiteur de quatre huiles essentielles d'eucalyptus collectées au Maroc sur l'agent phytopathogène Allorhizobium vitis la souche S4, responsable de la maladie de la galle du collet de la vigne. L'identification des constituants des huiles essentielles a été réalisée par analyse GC-MS et l'activité antibactérienne a été évaluée à l'aide d'un test de chromatogramme. Le rendement de différents échantillons d'eucalyptus variait de 2 à 4,5 %. Les résultats de l'analyse des composés chimiques montrent que les principaux composés des huiles essentielles d'eucalyptus sont le 1,8-Cinéol (Eucalyptol) (65,55-78,11%), l'α-Pinène (3-15,21%), le 2-β-Pinène (2,33-5,27 %) et le p-Cymène (0,79-1,51 %). Toutes les huiles essentielles d'eucalyptus présentent une activité antibactérienne in vitro contre la souche S4 d'Allorhizobium vitis avec un pourcentage d'inhibition et des valeurs de concentration minimales inhibitrices comprises entre 13,67-20,50 % et 20-40 mg/mL, respectivement

Assessment of Ionomic, Phenolic and Flavonoid Compounds for a Sustainable Management of Xylella fastidiosa in Morocco

Morocco belongs to the countries ranked at a high-risk level for entry, establishment, and spread of Xylella fastidiosa, which has recently re-emerged as a plant pathogen of global importance causing olive quick decline syndrome (OQDS). Symptomatic infection by X. fastidiosa leads to devastating diseases and important economic losses. To prevent such losses and damages, countries without current outbreaks like Morocco need to first understand their host plant responses to X. fastidiosa. The assessment of the macro and micro-elements content (ionome) in leaves can give basic and useful information along with being a powerful tool for the sustainable management of diseases caused by this devastating pathogen. Herein, we compare the leaf ionome of four important autochthonous Moroccan olive cultivars (‘Picholine Marocaine’, ‘Haouzia’, ‘Menara’, and ‘Meslalla’), and eight Mediterranean varieties introduced in Morocco (‘Arbequina’, ‘Arbosana’, ‘Leccino’, ‘Ogliarola salentina’, ‘Cellina di Nardo’, ‘Frantoio’, ‘Leucocarpa’, and ‘Picholine de Languedoc’), to develop hypotheses related to the resistance or susceptibility of the Moroccan olive trees to X. fastidiosa infection. Leaf ionomes, mainly Ca, Cu, Fe, Mg, Mn, Na, Zn, and P, were determined using inductively coupled plasma optical emission spectroscopy (ICP-OES). These varieties were also screened for their total phenolics and flavonoids content. Data were then involved in a comparative scheme to determine the plasticity of the pathogen. Our results showed that the varieties ‘Leccino’, ’Arbosana’, ‘Arbequina’ consistently contained higher Mn, Cu, and Zn and lower Ca and Na levels compared with the higher pathogen-sensitive ‘Ogliarola salentina’ and ‘Cellina di Nardò’. Our findings suggest that ‘Arbozana’, ‘Arbiquina’, ‘Menara’, and ‘Haouzia’ may tolerate the infection by X. fastidiosa to varying degrees, provides additional support for ‘Leccino’ having resistance to X. fastidiosa, and suggests that both ‘Ogliarola salentina’ and ‘Cellina di Nardö’ are likely sensitive to X. fastidiosa infection