Mohamed Ali Triki

Isolation and characterization of rhizosphere bacteria for the biocontrol of the damping-off disease of tomatoes in Tunisia.

Fluorescent Pseudomonas spp., isolated from tomato and pepper plants rhizosphere soil, was evaluated in vitro as a potential antagonist of fungal pathogens. Pseudomonas strains were tested against the causal agents of tomatoes damping-off (Sclerotinia sclerotiorum), root rot (Fusarium solani), and causal agents of stem canker and leaf blight (Alternaria alternata). For this purpose, dual culture antagonism assays were carried out on 25% tryptic soy agar, King B medium and potato dextrose agar to determine the effect of the strains on mycelial growth of the pathogens. In addition, strains were screened for their ability to produce exoenzymes and siderophores. All the strains significantly inhibited Alternaria alternata, particularly in 25% TSA medium. Antagonistic effect on Sclerotinia sclerotiorum and Fusarium solani was greater on King B medium. Protease was produced by 30% of the strains, but no strain produced cellulase or chitinase. Finally, the selected Pseudomonas strain, Psf5, was evaluated on tomato seedling development and as a potential candidate for controlling tomato damping-off caused by Sclerotinia sclerotiorum, under growth chamber conditions. In vivo studies resulted in significant increases in plant stand as well as in root dry weight. Psf5 was able to establish and survive in tomato plants rhizosphere after 40days following the planting of bacterized seeds.

Genetic diversity of Verticillium dahliae from olive trees in Tunisia based on RAMS and IGS-RFLP analyses

Abstract Forty-two isolates of Verticillium dahliae were recovered from stem and root samples of olive trees showing symptoms of verticillium wilt in various olive-growing regions in Tunisia. Each isolate was identified based on microscopic observations of morphological and cultural characteristics, pathogenicity tests, as well as PCR amplification using Vd1/Vd2 primers. Genetic diversity among the isolates was investigated using random amplified microsatellites (RAMS) and PCR-RFLP of intergenic spacer region (IGS) of ribosomal DNA (rDNA). A single fragment of approximately 1.7–2.1 kb was amplified from all isolates by PCR using primers CNL12 and CNS1. Digestion of the amplified IGS region with restriction enzyme RsaI produced similar banding patterns (1200 and 800 bp) for 40 isolates while individual and distinctive banding patterns (1100, 850 and 150 bp) were observed for two isolates. Using RAMS primers, nine, eight and four bands were produced when using primers CGA (2200, 1400, 1200, 1100, 1000, 650, 550, 500 and 350 bp), CCA (2000, 1200, 950, 850, 800, 550, 500 and 400 bp) and GT (2500, 2400, 700 and 500), respectively. A total of 21 polymorphic markers were scored when data from the RAMS experiments were combined. Overall, the results of this study revealed associations between the genetic diversity of the isolates and their pathogenicity phenotype, but not between genetic diversity and the geographic origin, suggesting that they are randomly spread across Tunisia.

Biocontrol of olive knot disease by Bacillus subtilis isolated from olive leaves

We evaluated the biocontrol efficacy of a Bacillus subtilis strain isolated from symptomless olive leaves against the olive knot disease pathogen Pseudomonas savastanoi pv. savastanoi. Bacillus subtilis F1 and F-4 displayed antibacterial activity against the pathogen. In planta, only B. subtilis F1 significantly reduced the weight of knots caused by P. savastanoi pv. savastanoi IVIA 1628 and Aw9, whereas copper treatment was effective only when using strain Aw9, thus a reduction of 50% was obtained. Preliminary characterisation of the active compound produced by B. subtilis F1 showed that it was proteinaceous in nature. These results suggest that B. subtilis strain F1 could be used to control, and prevent infection by, the causal agent of olive knot disease.

FIRST REPORT OF CYTOSPORA PUNICAE ASSOCIATED WITH WOOD CANKER AND BRANCH DIEBACK DISEASE OF POMEGRANATE IN TUNISIA

Pomegranate (Punica granatum L.) is an economically important crop in Tunisia with an annual production of 23.000 tons. During May and June of 2014, severe branch dieback was observed in pomegranate plants cv. Gabsi located in Gabes region with about 8% of disease incidence. Symptoms of the disease included leaf yellowing, wood lesion and canker formation. Isolation of the pathogen was performed from 10 samples of active cankers plated onto PDA medium amended with 100 mg l-1 of tetracycline hydrochloride. Fungal colonies were then sub-cultured onto PDA medium at 22°C. All isolates were identified as Cytospora sp. based on colony morphology, conidial characteristics and pycnidia formation (Palavouzis et al., 2015). The isolates developed white mycelium, which turned green to dark brown with hyaline, allantoid, aseptate conidia (average 4-5 μm x 1.75 μm) and production of dark coloured pycnidia 300 to 450 μm in diameter after 15 days (Peduto Hand et al., 2014). Identity of these isolates was confirmed by sequencing the internal transcribed spacer region. The ITS sequences were deposited in GenBank (Accession No. KT272402). These sequences revealed 99% genetic identity with those of Cytospora punicae species available in GenBank (KJ621689; KJ621688). Pathogenicity of Cytospora punicae was evaluated by inoculation of two isolates in 1-year-old shoots of pomegranate cv. Gabsi (Palavouzis et al., 2015). The inoculated shoots developed necrotic spots with vascular discoloration spreading downward and upward from the inoculation site. Cytospora punicae was recovered from 100% of the inoculated shoots. This is the first report of Cytospora punicae causing wood canker and branch dieback of pomegranate in Tunisia.

Ruta montana L. leaf essential oil and extracts: characterization of bioactive compounds and suppression of crown gall disease.

The aims of this study were to assess the antimicrobial efficacy of the leaf essential oil and the leaf extracts of R. montana against Botrytis cinerea, Fusarium oxysporum, Verticillium dahliae, Aspergillus oryzae and Fusarium solani. The oil (1.000 µg/disk) and the extracts (1.500 µg/disk) revealed a remarkable antifungal effect against the tested plant pathogenic fungi with a radial growth inhibition percentage of 40.0-80.0 % and 5.0-58.0 %, respectively along with their respective MIC values ranging from 100 to 1100 µg/mL and 250 to 3000 µg/mL. The oil had a strong detrimental effect on spore germination of all the tested plant pathogens along with the concentration as well as time-dependent kinetic inhibition of Fusarium oxysporum. Also, the oil exhibited a potent in vivo antifungal effect against Botrytis cinerea on tomato plants. Experiments carried out in plant revealed that the essential oil was slightly effective in suppression of gall formation induced by Agrobacterium tumefaciens on bitter almond. The results of this study indicate that the oil and extracts of R. montana leaves could become natural alternatives to synthetic fungicides to control certain important plant microbial diseases. The GC-MS analysis determined that 28 compounds, which represented 89.03 % of total oil, were present in the oil containing mainly 1-butene, methylcyclopropane, 2-butene and caryophyllene oxide.

FIRST REPORT OF DIEBACK OF OLIVE TREES CAUSED BY PHOMA FUNGICOLA IN TUNISIA

During spring 2013, a new disease was observed on olive (Olea europea) cv. Chemlali in orchard in Sahlin (Sahel of Tunisia). Symptoms included abundant dead branches and wilted leaves and shoot necrosis. Shoots showing dieback symptoms were disinfected with 2% sodium hypochlorite, rinsed in sterile distilled water and air dried. Several fragments (3 x 3 mm) of infected shoots were cut and placed on potato dextrose agar medium (PDA). All plates were incubated at 25°C for 4 days under continuous fluorescent light. A pycnidial fungus was consistently isolated from branch cankers and identified as Phoma fungicola Aveskamp, Gruyter et Verkley, based on morphological characteristics and analysis of the ITS gene region (White et al., 1990). The sequence showed high identity (99%) with a reference sequence (strain H11 H10; accession No. KF29376 3.1). Pathogenicity tests were conducted on 2-year old olive plants (cv. Chemlali). A mycelial plug cut from the margin of an actively growing colony of the fungus was placed into a shallow wound (0.4 cm2) inferted with a sterilized scalpel on the stem base. Inoculated wounds were wrapped with Parafilm. In control plants, sterile PDA plugs were placed into artificial wounds. Ten replicate inoculated plants were used and maintained in a greenhouse at 25°C. Two months after the inoculation, the inoculated trees reproduced stem browning symptoms observed in the field, while control plants remained healthy. Koch’s postulates were then verified and P. fungicola was isolated from inoculated stems, whereas the controls were free of the fungus. Phoma sp. and P. incompta have been reported as responsible for branch dieback of olive tree in Tunisia and Italy, respectively (Rhouma et al., 2010; Ivic et al., 2010). To the best of our knowledge, this is the first report of P. fungicola as a causal agent of dieback of olive trees in Tunisia.

Preparers’ and users’ perception of intellectual capital information usefulness: a Tunisian exploratory study

Purpose The purpose of this paper is to explore preparers’ and users’ perception of intellectual capital (IC) usefulness and to examine the significant differences in the usefulness perceptions of IC information and its categories according to sex, age, function, educational level, specialty and professional experience of respondents. Design/methodology/approach This exploratory study drew on a questionnaire survey sent to five groups of preparers and users who were asked to provide their usefulness perception about information on IC and its categories. Findings This paper found that the five preparers and users groups perceive information on IC as well as its three categories as useful for their decision-making purposes. In addition, it concluded that the usefulness perception of IC information does not differ by sex, age, function, educational level and specialty of the respondents, but it differs according to the professional experience. Practical implications To the extent that users perceive IC information as useful, managers are encouraged to disclose more information about this hidden capital in order to improve their transparency. As there are no generally accepted IC reporting guidelines and in order to fill informational gaps between companies and their stakeholders, accounting standards bodies could regulate the IC information disclosure by developing relevant communication standards in accordance with stakeholders’ expectations. They may identify information items that should be considered as a priority by making them mandatory for disclosure purposes, and other items voluntary. Originality/value The paper can be regarded as the first exploratory study to investigate the IC information usefulness from the perspectives of five preparers and users groups in Tunisia, as an example of a developing economy in Africa.

Fusarium Brachygibbosum and Fusarium Chlamydosporum Causing Wilt and Die-back of Olive in Tunisia

The dieback and wilting symptoms induced by complex soilborne fungi has caused considerable economic losses in olive orchards 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.

FIRST REPORT OF NECTRIA HAEMATOCOCCA ASSOCIATED WITH DIEBACK OF OLIVE TREES IN TUNISIA

During a routine survey for olive diseases conducted in autumn 2013 in southern Tunisia (Bir Ali region), wilting and chlorosis of the leaves accompanied by brown-to-black discoloration of the wood in cross-sectioned twigs were observed on 3- to 10-year-old olive trees. On potato dextrose agar (PDA), a fungus was isolated from symptomatic twigs and branches with an initially white mycelium that over time became light gray-brown. This fungus was identified as Nectria sp. based on morphological characteristics and analysis of the ITS gene region (White et al., 1990). A BLAST search of GenBank database revealed 99% homology of the amplified product with a reference sequence of Nectria haematococca (strain HLJ_14, accession No. JN088237.1). Pathogenicity tests were conducted on 10 two-year old olive trees of cv. Chemlali, by placing a mycelial plug in a shallow wound on the stem of each plant. Control plants were inoculated with sterile PDA plugs. Two months post inoculation, symptoms appeared, with stems showing brown discolorations and necrotic lesions. Controls remained healthy. N. haematococca was recovered from necrotic lesions, thus fulfilling Kochs postulates. N. haematococca was held responsible for root rot of olive trees in Argentina (Barreto et al., 2003). To our knowledge this is the first report of N. haematococca as a causal agent of dieback of olive trees in Tunisia.