The novel bacteriocin BacYB1 produced by Leuconostoc mesenteroides YB1: From recent analytical characterization to biocontrol Verticillium dahliae and Agrobacterium tumefaciens.

Biocontrol of phytopathogens involving the use of bioactive compounds produced by lactic acid bacteria (LAB), is a promising approach to manage many diseases in agriculture. In this study, a lactic acid bacterium designated YB1 was isolated from fermented olives and selected for its antagonistic activity against Verticillium dahliae (V. dahliae) and Agrobacterium tumefaciens (A. tumefaciens). Based on the 16S rRNA gene nucleotide sequence analysis (1565 pb, accession number: OR714267), the new isolate YB1 bacterium was assigned as Leuconostoc mesenteroides YB1 (OR714267) strain. This bacterium produces an active peptide "bacteriocin" called BacYB1, which was purified in four steps. Matrix-assisted lasers desorption/ionization (MALDI) time-of-flight (TOF) mass spectrometry (MS) based approach was performed to identify and characterize BacYB1. The exact mass was 5470.75 Da, and the analysis of the N-terminal sequence (VTRASGASTPPGTASPFKTL) of BacYB1 revealed no significant similarity to currently available antimicrobial peptides. The BacYB1 displayed a bactericidal mode of action against A. tumefaciens. The potentiel role of BacYB1 to supress the growth of A. tumefaciens was confirmed by live-dead cells viability assay. In pot experiments, the biocontrol efficacy of BacYB1 against V. dahliae wilt on young olive trees was studied. The percentage of dead plants (PDP) and the final mean symptomes severity (FMS) of plants articifialy infected by V. dahliae and treated with the pre-purified peptide BacYB1 (preventive and curative treatments) were significantly inferior to untreated plants. Biochemical analysis of leaves of the plants has shown that polyophenols contents were highly detected in plants infected by V. dahliae and the highest contents of chlorophyl a, b and total chlorophyll were recorded in plants treated with the combination of BacYB1 with the biofertilisant Humivital. BacYB1 presents a promising alternative for the control of Verticillium wilt and crown gall diseases.

Bacillus methylotrophicus DCS1: Production of Different Lipopeptide Families, In Vitro Antifungal Activity and Suppression of Fusarium Wilt in Tomato Plants.

The present work aims to quantitatively and qualitatively monitor the production of lipopeptide mixtures by Bacillus methylotrophicus DCS1 strain in Landy medium and to investigate the antifungal activities of DCS1 strain and its produced lipopeptides. The in vitro activities were tested by the direct confrontation and agar well diffusion methods, while the in vivo study was carried out in order to test the efficiency of DCS1 bacterial suspension in the control of Fusarium wilt in tomato plants. Identification of lipopeptides by mass spectrometry (LC/MSD-TOF) showed that lipopeptide isoforms produced during the first 24 h and 48 h of fermentation are identical, belonging to bacillomycin D and fengycins A and B homologues with a difference in the yield of production. After 72 h of fermentation corresponding to the end of incubation period, B. methylotrophicus DCS1 is able to produce a mixture of surfactin, pumilacidin, iturin A/mycosubtilin, iturin C1, bacillomycin D and fengycins A and B isoforms. The results of in vitro antifungal experiments suggest that B. methylotrophicus DCS1 has a significant potential as a biocontrol agent, owing to lipopeptides produced, endowed with antifungal activity against several phytopathogenic fungi. The curative treatment of tomato plants with DCS1 bacterial suspension was more effective in the protection against Fusarium oxysporum f. sp. radicis-lycopersici (FORL) than the preventive treatment by comparing the average number of leaves remaining healthy after 30 days of each treatment and the appearance of tomato plants roots. The results indicate that B. methylotrophicus DCS1 exhibit a significant suppression of Fusarium wilt symptoms in tomato plants comparable to that of commercial fungicides and could be an alternative to chemically synthesized pesticides.

Bacillus subtilis SPB1 lipopeptide biosurfactant: antibacterial efficiency against the phytopathogenic bacteria Agrobacterium tumefaciens and compared production in submerged and solid state fermentation systems.

Bacillus subtilis SPB1 derived biosurfactants (BioS) proved its bio-control activity against Agrobacterium tumefaciens using tomato plant. Almost 83% of disease symptoms triggered by Agrobacterium tumefaciens were reduced. Aiming potential application, we studied lipopeptide cost-effective production in both fermentations systems, namely the submerged fermentation (SmF) and the solid-state fermentation (SSF) as well as the use of Aleppo pine waste and confectionery effluent as cheap substrates. Optimization studies using Box-Behnken (BB) design followed by the analysis with response surface methodology were applied. When using an effluent/sea water ratio of 1, Aleppo pine waste of 14.08 g/L and an inoculum size of 0.2, a best production yield of 17.16 ± 0.91 mg/g was obtained for the SmF. While for the SSF, the best production yield of 27.59 ± 1.63 mg/g was achieved when the value of Aleppo pine waste, moisture, and inoculum size were, respectively, equal to 25 g, 75%, and 0.2. Hence, this work demonstrated the superiority of SSF over SmF.

Exploring Bacterial and Fungal Biodiversity in Eight Mediterranean Olive Orchards (Olea europaea L.) in Tunisia.

A wide array of bacteria and fungi are known for their association with pests that impact the health of the olive tree. The latter presents the most economically important cultivation in Tunisia. The microbial diversity associated with olive orchards in Tunisia remains unknown and undetermined. This study investigated microbial diversity to elucidate the microbial interactions that lead to olive disease, and the bio-prospects for potential microbial biocontrol agents associated with insect pests of economic relevance for olive cultivation in the Mediterranean area. Bacterial and fungal isolation was made from soil and olive tree pests. A total of 215 bacterial and fungal strains were randomly isolated from eight different biotopes situated in Sfax (Tunisia), with different management practices. 16S rRNA and ITS gene sequencing were used to identify the microbial community. The majority of the isolated bacteria, Staphylococcus, Bacillus, Alcaligenes, and Providencia, are typical of the olive ecosystem and the most common fungi are Penicillium, Aspergillus, and Cladosporium. The different olive orchards depicted distinct communities, and exhibited dissimilar amounts of bacteria and fungi with distinct ecological functions that could be considered as promising resources in biological control.

Desirable traits for a good biocontrol agent against fire blight disease.

AIMS: This study aimed to improve the screening strategy for the selection of biocontrol agents with high biocontrol efficacy against fire blight disease. METHODS AND RESULTS: A two-step screening procedure consisting of in-vitro laboratory tests and an ex-vivo test system using detached pear leaves was applied to 43 Bacillus strains originated from the rhizosphere and the aerial parts of apple and pear plants. The grouping of the studied strains and the tested traits based on the principal component analysis and the two-way hierarchical cluster analysis showed that siderophore production and biofilm formation are the most desirable traits in a Bacillus biocontrol agent to control fire blight disease and that rhizospheric originating strains are the most effective. CONCLUSIONS: In contrast to the previous screening strategies that are often insufficient to select the most suitable microorganisms, this study reported an improved strategy based on the microbial competition traits to select potential Bacillus biocontrol agents with high biocontrol efficacy against fire blight disease.

Detection of Urban and Environmental Changes via Remote Sensing.

Rapid climate and environmental change at limited, regional, and general scales have been a major concern for researchers in a number of fields, such as topography, economy, environment, and sustainable development. Changes in land cover and land use have taken into account due to potential impacts on soil depletion, amplified run-off, water balance, and climate change. A detailed understanding of the characteristics of land exploitation and land structure is indispensable for the study of their influences on life and nature. In addition, urban extension is a major form of land extraction and land transformation, as it relates to the rise in population and the availability of financial services. Remote sensing records have been shown to be important for reporting and perceiving urban development and transition, and for providing critical information for future growth. Transformation and shift identification are the tools used to recognise distinctions in a land cover by tracking them at various times. In addition, various change identification and detection approaches are routinely tested with the goal of providing the greatest change detection deductions for a particular appliance. This review would aim to establish a practical plan that combines remote sensing techniques, on the one hand, and modelling approaches, on the other, to track land use, to cover changes, and to predict future trends.

Identification of fungi in Tunisian olive orchards: characterization and biological control potential.

BACKGROUND: Olive production is the main agricultural activity in Tunisia. The diversity of fungi was explored in two different olive groves located in two distant geographical zones in Sfax (Tunisia) with different management practices. RESULTS: Fungal isolation was made from soil and the major olive tree pests, namely the Olive fly, Bactrocera oleae Gmelin (Diptera: Tephritidae), and the Olive psyllid, Euphyllura olivina Costa (Homoptera: Psyllidae). A total of 34 fungal isolates were identified according to their phenotypic, genotypic, biochemical and biological activities. Twenty fungal species were identified belonging to six different genera (Alternaria, Aspergillus, Cladosporium, Fusarium, Lecanicillium and Penicillium) by the analysis of their ITS1-5.8S-ITS2 ribosomal DNA region. Different bioassays performed in this work revealed that 25/34 (73.5%) of the identified fungal isolates showed an entomopathogenic and/or antagonistic activity, 9/34 (26.5%) of them displayed phytopathogenic features. CONCLUSIONS: Fungal species that showed entomopathogenic and/or antagonistic potentialities and that are non-phytopathogenic, (17/34; 50%) of our fungal isolates, could be explored for olive protection against fungal diseases and pests, and might have a future application as biocontrol agents.

Physico-chemical and spectroscopic quality assessment of compost from date palm (Phoenix dactylifera L.) waste valorization.

Organic matter (OM) is a vital component for a healthy soil, its lack arise a major problem for farmers who need to use commercialized fertilizers with high costs. Considering circular economy approach and for increasing OM availability, water soaked date palm waste was co-composted with goat manure in aerated windrow to produce a soil organic amendment. The OM biodegradation was mainly controlled based on biological parameters and spectroscopic techniques. The results showed a rapid temperature increase during the first week, and a relatively long compost maturity phase. The OM content reduction was of 36% and C/N ratio shifted from 60 to 20 at the process end. During the composting process, the specific ultraviolet absorbance SUVA254, SUVA269 and the SUVA280 values increase confirmed the OM and hydrophilic compounds degradation, as well as substrate content oxidation into aromatic compounds. The Fourier Transform Infrared Spectroscopy (FTIR) analyses of the different samples collected during the process exhibited both OM biodegradation and mineralization. The 3 absorption ratios 1650/2845, 1525/2925 and 2920/1640 confirmed an aromaticity increase by aromatic structures biosynthesis, such as humic-like and fulvic-like substances, with the decomposition/transformation of aliphatic components, polysaccharides, and alcohols. Fluorescence excitation-emission matrix (FEEM) spectroscopy coupled with parallel factor analysis (PARAFAC) evidenced the dissolved organic matter (DOM) humification. A four-component model was obtained, i.e. humic-like component (S1, S2 and S3) and fulvic-like component (S4). The produced compost didn't exhibit any phytotoxicity evidenced by cress seed germination index exceeding 80%. All the analyses confirmed the good quality of the compost issued from mixed date palm waste and goat manure.

Providencia entomophila sp. nov., a new bacterial species associated with major olive pests in Tunisia.

Bioprospection for potential microbial biocontrol agents associated with three major insect pests of economic relevance for olive cultivation in the Mediterranean area, namely the olive fly, Bactrocera oleae, the olive moth, Prays oleae, and the olive psyllid, Euphyllura olivina, led to the isolation of several strains of readily cultivable Gram-negative, rod-shaped bacteria from Tunisian olive orchards. Determination of 16S ribosomal RNA encoding sequences identified the bacteria as members of the taxonomic genus Providencia (Enterobacterales; Morganellaceae). A more detailed molecular taxonomic analysis based on a previously established set of protein-encoding marker genes together with DNA-DNA hybridization and metabolic profiling studies led to the conclusion that the new isolates should be organized in a new species within this genus. With reference to their original insect association, the designation "Providencia entomophila" is proposed here for this hypothetical new taxon.

Olea europaea L. Root Endophyte Bacillus velezensis OEE1 Counteracts Oomycete and Fungal Harmful Pathogens and Harbours a Large Repertoire of Secreted and Volatile Metabolites and Beneficial Functional Genes.

Oomycete and fungal pathogens, mainly Phytophthora and Fusarium species, are notorious causal agents of huge economic losses and environmental damages. For instance, Phytophthora ramorum, Phytophthora cryptogea, Phytophthora plurivora and Fusarium solani cause significant losses in nurseries and in forest ecosystems. Chemical treatments, while harmful to the environment and human health, have been proved to have little or no impact on these species. Recently, biocontrol bacterial species were used to cope with these pathogens and have shown promising prospects towards sustainable and eco-friendly agricultural practices. Olive trees prone to Phytophthora and Fusarium disease outbreaks are suitable for habitat-adapted symbiotic strategies, to recover oomycetes and fungal pathogen biocontrol agents. Using this strategy, we showed that olive trees-associated microbiome represents a valuable source for microorganisms, promoting plant growth and healthy benefits in addition to being biocontrol agents against oomycete and fungal diseases. Isolation, characterization and screening of root microbiome of olive trees against numerous Phytophthora and other fungal pathogens have led to the identification of the Bacillus velezensis OEE1, with plant growth promotion (PGP) abilities and strong activity against major oomycete and fungal pathogens. Phylogenomic analysis of the strain OEE1 showed that B. velezensis suffers taxonomic imprecision that blurs species delimitation, impacting their biofertilizers' practical use. Genome mining of several B. velezensis strains available in the GenBank have highlighted a wide array of plant growth promoting rhizobacteria (PGPR) features, metals and antibiotics resistance and the degradation ability of phytotoxic aromatic compounds. Strain OEE1 harbours a large repertoire of secreted and volatile secondary metabolites. Rarefaction analysis of secondary metabolites richness in the B. velezenis genomes, unambiguously documented new secondary metabolites from ongoing genome sequencing efforts that warrants more efforts in order to assess the huge diversity in the species. Comparative genomics indicated that B. velezensis harbours a core genome endowed with PGP features and accessory genome encoding diverse secondary metabolites. Gas Chromatography-Mass Spectrometry (GC-MS) analysis of OEE1 Volatile Organic Compounds (VOCs) and Liquid Chromatography High Resolution Mas Spectrometry (LC-HRMS) analysis of secondary metabolites identified numerous molecules with PGP abilities that are known to interfere with pathogen development. Moreover, B. velezensis OEE1 proved effective in protecting olive trees against F. solani in greenhouse experiments and are able to inhabit olive tree roots. Our strategy provides an effective means for isolation of biocontrol agents against recalcitrant pathogens. Their genomic analysis provides necessary clues towards their efficient implementation as biofertilizers.

Screening of the High-Rhizosphere Competent Limoniastrum monopetalum' Culturable Endophyte Microbiota Allows the Recovery of Multifaceted and Versatile Biocontrol Agents.

Halophyte Limoniastrum monopetalum, an evergreen shrub inhabiting the Mediterranean region, has well-documented phytoremediation potential for metal removal from polluted sites. It is also considered to be a medicinal halophyte with potent activity against plant pathogens. Therefore, L. monopetalum may be a suitable candidate for isolating endophytic microbiota members that provide plant growth promotion (PGP) and resistance to abiotic stresses. Selected for biocontrol abilities, these endophytes may represent multifaceted and versatile biocontrol agents, combining pathogen biocontrol in addition to PGP and plant protection against abiotic stresses. In this study 117 root culturable bacterial endophytes, including Gram-positive (Bacillus and Brevibacillus), Gram-negative (Proteus, Providencia, Serratia, Pantoea, Klebsiella, Enterobacter and Pectobacterium) and actinomycete Nocardiopsis genera have been recovered from L. monopetalum. The collection exhibited high levels of biocontrol abilities against bacterial (Agrobacterium tumefaciens MAT2 and Pectobacterium carotovorum MAT3) and fungal (Alternaria alternata XSZJY-1, Rhizoctonia bataticola MAT1 and Fusarium oxysporum f. sp. radicis lycopersici FORL) pathogens. Several bacteria also showed PGP capacity and resistance to antibiotics and metals. A highly promising candidate Bacillus licheniformis LMRE 36 with high PGP, biocontrol, metal and antibiotic, resistance was subsequently tested in planta (potato and olive trees) for biocontrol of a collection of 14 highly damaging Fusarium species. LMRE 36 proved very effective against the collection in both species and against an emerging Fusarium sp. threatening olive trees culture in nurseries. These findings provide a demonstration of our pyramiding strategy. Our strategy was effective in combining desirable traits in biocontrol agents towards broad-spectrum resistance against pathogens and protection of crops from abiotic stresses. Stacking multiple desirable traits into a single biocontrol agent is achieved by first, careful selection of a host for endophytic microbiota recovery; second, stringent in vitro selection of candidates from the collection; and third, application of the selected biocontrol agents in planta experiments. That pyramiding strategy could be successfully used to mitigate effects of diverse biotic and abiotic stresses on plant growth and productivity. It is anticipated that the strategy will provide a new generation of biocontrol agents by targeting the microbiota of plants in hostile environments.

Economic production and biocontrol efficiency of lipopeptide biosurfactants from Bacillus mojavenis A21.

In the wake of an increased attention on the eco-friendly biopesticidal products and the rising market requirements for organic agents, lipopeptides compounds have been described as biological control agents which improve the overall health growth and development of plants. Nevertheless, their high production cost constitue the major flaw in their wide use to control plant diseases. The present article aims to formulate an economic media for lipopeptides production by Bacillus mojavensis A21 for application as natural fungicides for plant disease treatment. We herein demonstrated the suitability of the potato waste, as low cost substrate, for lipopeptides production. Moreover, sea water  was found to be a good mineral salts sources. In the second part of this study, we investigate the inhibitory activity of A21 lipopeptides against the phtopathogenic Fusarium sp. The in vitro test showed a minimal inhibitory concentration of about 0.3 mg/ml. The microscopic examination, of the treated Fusarium revealed an excessive lysis of the mycelia ultrastructure with destructed spores. The in vivo antagonist activity was confirmed towards the infected potato tubers. A21 lipopeptides are effective in decreasing by about 78.26% and 60.68% when applied as preventive and curative treatments, respectively, as compared to the untreated tubers.

MIP diversity from Trichoderma: Structural considerations and transcriptional modulation during mycoparasitic association with Fusarium solani olive trees.

Major intrinsic proteins (MIP) are characterized by a transmembrane pore-type architecture that facilitates transport across biomembranes of water and a variety of low molecular weight solutes. They are found in all parts of life, with remarkable protein diversity. Very little is known about MIP from fungi. And yet, it can legitimately be stated that MIP are pivotal molecular components in the privileged relationships fungi enjoy with plants or soil fauna in various environments. To date, MIP have never been studied in a mycoparasitism situation. In this study, the diversity, expression and functional prediction of MIP from the genus Trichoderma were investigated. Trichoderma spp. genomes have at least seven aquaporin genes. Based on a phylogenetic analysis of the translated sequences, members were assigned to the AQP, AQGP and XIP subfamilies. In in vitro and in planta assays with T. harzianum strain Ths97, expression analyses showed that four genes were constitutively expressed. In a mycoparasitic context with Fusarium solani, the causative agent of fusarium dieback on olive tree roots, these genes were up-regulated. This response is of particular interest in analyzing the MIP promoter cis-regulatory motifs, most of which are involved in various carbon and nitrogen metabolisms. Structural analyses provide new insights into the possible role of structural checkpoints by which these members transport water, H2O2, glycerol and, more generally, linear polyols across the membranes. Taken together, these results provide the first evidence that MIP may play a key role in Trichoderma mycoparasitism lifestyle.

Screening for Fusarium Antagonistic Bacteria From Contrasting Niches Designated the Endophyte Bacillus halotolerans as Plant Warden Against Fusarium.

Date palm (Phoenix dactylifera L.) plantations in North Africa are nowadays threatened with the spread of the Bayoud disease caused by Fusarium oxysporum f. sp. albedinis, already responsible for destroying date production in other infected areas, mainly in Morocco. Biological control holds great promise for sustainable and environmental-friendly management of the disease. In this study, the additional benefits to agricultural ecosystems of using plant growth promoting rhizobacteria (PGPR) or endophytes are addressed. First, PGPR or endophytes can offer an interesting bio-fertilization, meaning that it can add another layer to the sustainability of the approach. Additionally, screening of contrasting niches can yield bacterial actors that could represent wardens against whole genera or groups of plant pathogenic agents thriving in semi-arid to arid ecosystems. Using this strategy, we recovered four bacterial isolates, designated BFOA1, BFOA2, BFOA3 and BFOA4, that proved very active against F. oxysporum f. sp. albedinis. BFOA1-BFOA4 proved also active against 16 Fusarium isolates belonging to four species: F. oxysporum (with strains phytopathogenic of Olea europaea and tomato), F. solani (with different strains attacking O. europaea and potato), F. acuminatum (pathogenic on O. europaea) and F. chlamydosporum (phytopathogenic of O. europaea). BFOA1-BFOA4 bacterial isolates exhibited strong activities against another four major phytopathogens: Botrytis cinerea, Alternaria alternata, Phytophthora infestans, and Rhizoctonia bataticola. Isolates BFOA1-BFOA4 had the ability to grow at temperatures up to 35°C, pH range of 5-10, and tolerate high concentrations of NaCl and up to 30% PEG. The isolates also showed relevant direct and indirect PGP features, including growth on nitrogen-free medium, phosphate solubilization and auxin biosynthesis, as well as resistance to metal and xenobiotic stress. Phylogenomic analysis of BFOA1-BFOA4 isolates indicated that they all belong to Bacillus halotolerans, which could therefore considered as a warden against Fusarium infection in plants. Comparative genomics allowed us to functionally describe the open pan genome of B. halotolerans and LC-HRMS and GCMS analyses, enabling the description of diverse secondary metabolites including pulegone, 2-undecanone, and germacrene D, with important antimicrobial and insecticidal properties. In conclusion, B. halotolerans could be used as an efficient bio-fertilizer and bio-control agent in semi-arid and arid ecosystems.

Multiple linear regression and artificial neural networks for delta-endotoxin and protease yields modelling of Bacillus thuringiensis.

The aim of the present work was to develop a model that supplies accurate predictions of the yields of delta-endotoxins and proteases produced by B. thuringiensis var. kurstaki HD-1. Using available medium ingredients as variables, a mathematical method, based on Plackett-Burman design (PB), was employed to analyze and compare data generated by the Bootstrap method and processed by multiple linear regressions (MLR) and artificial neural networks (ANN) including multilayer perceptron (MLP) and radial basis function (RBF) models. The predictive ability of these models was evaluated by comparison of output data through the determination of coefficient (R 2) and mean square error (MSE) values. The results demonstrate that the prediction of the yields of delta-endotoxin and protease was more accurate by ANN technique (87 and 89% for delta-endotoxin and protease determination coefficients, respectively) when compared with MLR method (73.1 and 77.2% for delta-endotoxin and protease determination coefficients, respectively), suggesting that the proposed ANNs, especially MLP, is a suitable new approach for determining yields of bacterial products that allow us to make more appropriate predictions in a shorter time and with less engineering effort.

Bayesian and Phylogenic Approaches for Studying Relationships among Table Olive Cultivars.

To enhance table olive tree authentication, relationship, and productivity, we consider the analysis of 18 worldwide table olive cultivars (Olea europaea L.) based on morphological, biological, and physicochemical markers analyzed by bioinformatic and biostatistic tools. Accordingly, we assess the relationships between the studied varieties, on the one hand, and the potential productivity-quantitative parameter links on the other hand. The bioinformatic analysis based on the graphical representation of the matrix of Euclidean distances, the principal components analysis, unweighted pair group method with arithmetic mean, and principal coordinate analysis (PCoA) revealed three major clusters which were not correlated with the geographic origin. The statistical analysis based on Kendall's and Spearman correlation coefficients suggests two highly significant associations with both fruit color and pollinization and the productivity character. These results are confirmed by the multiple linear regression prediction models. In fact, based on the coefficient of determination (R 2) value, the best model demonstrated the power of the pollinization on the tree productivity (R 2 = 0.846). Moreover, the derived directed acyclic graph showed that only two direct influences are detected: effect of tolerance on fruit and stone symmetry on side and effect of tolerance on stone form and oil content on the other side. This work provides better understanding of the diversity available in worldwide table olive cultivars and supplies an important contribution for olive breeding and authenticity.

Morphological and molecular characterization of Fusarium spp. associated with olive trees dieback in Tunisia.

Dieback and wilting symptoms caused by complex soilborne fungi are nowadays the most serious threatening disease affecting olive trees (Olea europaea) in Tunisia and presumably in many Mediterranean basin countries. Fusarium is one of the important phytopathogenic genera associated with dieback symptoms of olive trees. The objective of the present study was to confirm the pathogenicity of Fusarium spp. isolated from several olive-growing areas in Tunisia. According to the pathogenic test done on young olive trees (cv. Chemlali), 23 out of 104 isolates of Fusarium spp. were found to be pathogenic and the others were weakly or not pathogenic. The pathogenic Fusarium spp. isolates were characterized using molecular methods based on ITS PCR. Isolation results revealed the predominance of Fusarium solani (56.5%) and F. oxysporum species (21.7%) compared to F. chalmydosporum (8.7%), F. brachygibbosum (8.7%) and F. acuminatum (4.34%). Based on pathogenicity test, disease severity was highly variable among the 23 pathogenic isolates tested (P < 0.05) where F. solani was the most aggressive dieback agent. To the best of our knowledge, this is the first work that shows that Fusarium spp. might be a major agent causing dieback disease of olive trees in Tunisia.

The use of newly isolated Streptomyces strain TN258 as potential biocontrol agent of potato tubers leak caused by Pythium ultimum.

An actinomycete strain designated TN258, was isolated from Tunisian Sahara soil and selected for its antagonistic activity, especially against Pythium ultimum (P. ultimum) causing potato tubers leak. Based on the results of cultural characteristic of TN258 strain, the 16S rRNA gene nucleotide sequence (1433 bp, accession n° HE600071) and the phylogenetic analysis, we propose the assignment of our new isolate bacterium as Streptomyces TN258 strain. After culture optimization, the inhibitory effect of TN258 free cell supernatant against P. ultimum was evaluated. As result, by application of 50% (v/v) from 25 mg ml-1 of concentration, mycelial growth was totally inhibited with hyphal destruction. At the same concentration, the oospores were distorted and the germination was completely stopped. In potato tubers, Streptomyces TN258 filtrated supernatant, applied 24 h before inoculation by P. ultimum (preventive treatment group) was able to significantly decrease pathogen penetration by 62% and to reduce the percentage of weight loss by 59.43%, in comparison with non-treated group.

Lipopeptides from a novel Bacillus methylotrophicus 39b strain suppress Agrobacterium crown gall tumours on tomato plants.

BACKGROUND: This study aims to characterise the antibacterial activity of a novel Bacillus methylotrophicus strain named 39b against tumourigenic Agrobacterium tumefaciens C58 and B6 strains. It also aims to identify the compound that is responsible for its activity and to evaluate its efficiency to control crown gall disease in tomato plants. RESULTS: B. methylotrophicus strain 39b was found to stop the growth of phytopathogenic A. tumefaciens strains in in vitro experiments. Lipopeptides - surfactins, iturins and fengycins - were detected under various isoforms by mass spectrometry analysis of the methanolic extract. The active principle acting against Agrobacterium strains was isolated from TLC plates and identified by mass spectrometry as surfactin. The strain was effective in reducing the weight and the number of galls induced by A. tumefaciens strains on tomato plants. Total inhibition of gall formation was observed using the antibacterial compounds. CONCLUSION: B. methylotrophicus strain 39b exhibited antibacterial activity against phytopathogenic A. tumefaciens C58 and B6 both in vitro and in vivo. Lipopeptides are the main compounds that confer the biocontrol ability. This strain has the potential to be developed as a biological control agent for crown gall disease. © 2016 Society of Chemical Industry.

Purification and identification of Bacillus subtilis SPB1 lipopeptide biosurfactant exhibiting antifungal activity against Rhizoctonia bataticola and Rhizoctonia solani.

This study reports the potential of a soil bacterium, Bacillus subtilis strain SPB1, to produce lipopeptide biosurfactants. Firstly, the crude lipopeptide mixture was tested for its inhibitory activity against phytopathogenic fungi. A minimal inhibitory concentration (MIC), an inhibitory concentration at 50% (IC50%), and an inhibitory concentration at 90% (IC90%) values were determined to be 0.04, 0.012, and 0.02 mg/ml, respectively, for Rhizoctonia bataticola with a fungistatic mode of action. For Rhizoctonia solani, a MIC, an IC50%, and IC90% values were determined to be 4, 0.25, and 3.3 mg/ml, respectively, with a fungicidal mode of action. For both of the fungi, a loss of sclerotial integrity, granulation and fragmentation of hyphal mycelia, followed by hyphal shriveling and cell lysis were observed with the treatment with SPB1 biosurfactant fraction. After extraction, separation, and purification, different lipopeptide compounds were identified in the culture filtrate of strain SPB1. Mass spectroscopic analysis confirmed the presence of different lipopeptide compounds consisting of surfactin isoforms with molecular weights of 1007, 1021, and 1035 Da; iturin isoforms with molecular weights of 1028, 1042, and 1056 Da; and fengycin isoforms with molecular weights of 1432 and 1446 Da. Two new clusters of lipopeptide isoforms with molecular weights of 1410 and 1424 Da and 973 and 987 Da, respectively, were also detected. This study reported the ability of a B. subtilis strain to co-produce lipopeptide isoforms with potential use as antifungal compounds.