Olfa Frikha-Gargouri

Plant-based nanotechnology has emerged as a sustainable solution that minimizes environmental impact while increasing therapeutic effectiveness. This study aimed to synthesize silver nanoparticles (AgNPs) using seeds of Coriandrum sativum (Coriander) and stem bark of Cinnamomum verum (Cinnamon) essential oils (EOs) and to evaluate their antibacterial activity against bacteria isolated from diabetic foot ulcers (DFUs) in comparison to individual EOs and their mixture. The biosynthesized AgNPs (Cor-AgNPs and Cin-AgNPs) were characterized using various analytical techniques. The ultraviolet–visible spectroscopy spectral analysis showed absorption peaks at around 435 nm, confirming the formation of stable AgNPs. Field emission scanning electron microscopy analysis showed that Cor-AgNPs and Cin-AgNPs had average particle sizes of 25.56 ± 12.5 nm and 26.70 ± 8.6 nm, and zeta potentials of −24.2 ± 2.8 mV and −13.6 ± 2.3 mV, respectively. The x-ray diffraction crystallography test indicated that the crystalline size of the Cor-AgNPs was 24.52 nm, while that of Cin-AgNPs was 23.64 nm. The minimal inhibitory concentration (MIC) and minimal bactericidal concentration values were determined for the EOs, their mixture, and their synthesized nanoparticles. Cor-AgNPs and Cin-AgNPs exhibited significantly enhanced antibacterial activity when compared to their respective EOs with MIC values reaching as low as 0.6 µ g ml. Only a marginal improvement in antibacterial activity was observed for Cor-AgNPs and Cin-AgNPs when compared to those obtained with the EO mixture. This study highlights coriander and cinnamon EOs as promising agents for the green synthesis of AgNPs. The EO mixture and their nanoparticles may serve as potential alternatives to conventional antibiotics for the treatment of DFU bacteria.

Essential oils, known for their antimicrobial properties, are being investigated as natural alternatives to synthetic fungicides in agriculture. This study aimed to assess the chemical composition of six commercial essential oils (clove, tea tree, rosemary, thyme, oregano, and garlic) and to evaluate their fungistatic and/or fungicidal activity against six phytopathogenic fungi that cause significant damage to olive trees in Tunisia. For this purpose, the essential oils qualitative and quantitative chemical compositions were analyzed using gas chromatography-mass spectrometry. The antifungal activity was assessed using the poisoned substrate method at different concentrations (250, 500, 1000, and 4000 ppm). Results showed that the chemical composition analysis revealed that monoterpenoids were the dominant fraction in all oils except clove and garlic, which were primarily composed of eugenol (96.28%) and trisulfide (31.97%), respectively. The antifungal activity results showed that lower concentrations (250, 500, 1000 ppm) of tea tree, rosemary, thyme, and oregano oils had limited inhibitory effects on the tested fungi. However, Biscogniauxia mediterranea was highly sensitive to clove, garlic, and rosemary oils at 4000 ppm. Fusarium oxysporum, Fusarium solani, Verticillium dahliae, and Lasiodiplodia theobromae were mainly inhibited by clove oil at concentrations ranging from 500 to 4000 ppm, while Rhizoctonia bataticola was inhibited by clove and garlic oils at high concentrations. In conclusion, among the tested essential oils, clove oil demonstrated the highest antifungal efficacy, making it a promising natural alternative to synthetic fungicides for controlling olive tree phytopathogenic fungi.

BACKGROUND: Fusarium maize ear and root rot disease caused by Fusarium verticillioides has become one of the most serious fungal diseases associated with maize production. Due to their abilities to promote plant development and manage diseases, bacterial endophytes provide a more promising approach for treating this vascular disease.RESULTS: This work was undertaken for the selection and identification of promising isolates as plant growth promoters and biocontrol agents against F. verticillioides in maize agroecosystems. A screening procedure consisting of in vitro and in situ tests was applied to 27 endophytic strains originating from desert plants: Euphorbia antiquorum, Calotropis procera, and Alcasia albida. In vitro studies indicated that the bacteria exhibited variable results in biocontrol, endophytism, and plant growth‐promoting traits. In addition, in situ plant growth promotion and biocontrol experiments allowed the identification of the most promising bacterial endophytes. In vitro and in situ comparative study results indicated a low correlation. Our data revealed that in situ screening must be used as the method of selection of biocontrol agents against Fusarium ear and root rot disease. Based on in situ results, seven potent strains were selected and identified as Bacillus subtilis, Bacillus velezensis, Bacillus tequilensis, Stenotrophomonas maltophilia, and Klebsiella pneumoniae.CONCLUSION: The results of this study showed that the selected strains seem to be promising candidates to be exploited as biofertilizers and biocontrol agents against Fusarium maize ear and root rot disease. © 2023 Society of Chemical Industry.

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.

This study aimed to improve the screening strategy for the selection of biocontrol agents with high biocontrol efficacy against fire blight disease. 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. 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.

Two novel Algerian field‐collected isolates were selected for their antifungal activity against Zymoseptoria tritici (teleomorph Mycosphaerella graminicola). The novel strains, termed Alg.24B1 and Alg.24B2, were identified as Bacillus subtilis and Bacillus simplex since their respective nucleotide sequences of the 16S rRNA gene were 100% and 99.93% identical to those of B. subtilis and B. simplex, respectively. The antifungal activities of Alg.24B1 and Alg.24B2 were evaluated by the well diffusion method and compared to those of other Bacillus species. The maximum activity was obtained after two days of confrontation of the bacterial strain supernatants with the fungus for Alg.24B1 and three days for Alg.24B2. Furthermore, the metabolites responsible for the antifungal activity of both strains were detected by the investigation of either gene presence (PCR) or molecule production (activity detection of lytic enzymes and HPLC detection of lipopeptides). Overall, this study showed that in addition to their ability to produce lytic enzymes (protease and β‐glucanase), both strains coproduce three types of lipopeptides viz. surfactin, iturin, and fengycin. Thus, the biofungicide activity of both strains may be a result of a combination of different mechanisms. Therefore, they had a great potential to be used as biocontrol agents to effectively manage septoria tritici blotch of wheat (STB).

The persistence of pathogenic Agrobacterium strains as soil-associated saprophytes may cause an inconsistency in the efficacy of the biocontrol inoculants under field condition. The study of the interaction occurring in the rhizosphere between the beneficial and the pathogenic microbes is thus interesting for the development of effective biopesticides for the management of crown gall disease. However, very little is still known about the influence of these complex interactions on the biocontrol determinants of beneficial bacteria, especially Bacillus strains. This study aimed to evaluate the effect of the soil borne pathogen Agrobacterium tumefaciens C58 on root colonization and lipopeptide production by Bacillus velezensis strain 32a during interaction with tomato plants. Results show that the presence of A. tumefaciens C58 positively impacted the root colonization level of the Bacillus strain. However, negative impact on surfactin production was observed in Agrobacterium-treated seedling, compared with control. Further investigation suggests that these modulations are due to a modified tomato root exudate composition during the tripartite interaction. Thus, this work contributes to enhance the knowledge on the impact of interspecies interaction on the ecological fitness of Bacillus cells living in the rhizosphere.

Despite the potential biological importance of lipopeptides from Bacillus amyloliquefaciens as antimicrobial compounds, their effects on Agrobacterium tumefaciens biofilms have not been previously studied. These latter are important virulence factors for the development and re-occurrence of crown gall disease. As part of the development of a new biopesticide acting as anti-biofilm and biocontrol agent, we investigated for the first time the ability of a mixture of lipopeptides produced by B. amyloliquefaciens strain 32a to inhibit the tumor formation on plants and to reduce the formation of biofilms by the phytopathogenic A. tumefaciens strains C58 and B6. The mixture was found to display a strong biosurfactant activity as well as bactericidal activity against planktonic Agrobacterium cells. Moreover, the lipopeptide treatment inhibited biofilm formation in the range of 79.58 ± 0.60–100.00 ± 0.00% and dislodged 43.42 ± 0.91–93.89 ± 2.70% of preformed biofilm. For these assays, fluorescence microscopy did not show any adherent cell in the anti-adhesive assay and only few ones in the cell-dislodging assay. More importantly, lipopeptide-enriched extract inhibits tumor formation on tomato stem when treatments were applied after pathogen adhesion to wounded tissues. By virtue of its ability to inhibit biofilms formed on biotic and abiotic surfaces and to control efficiently tumor development, the 32a lipopeptide mixture may represent an excellent new tool for an efficient biocontrol of crown gall disease.

This study aimed to improve the screening method for the selection of Bacillus biocontrol agents against crown gall disease. The relationship between the strain biocontrol ability and their in vitro studied traits was investigated to identify the most important factors to be considered for the selection of effective biocontrol agents. In fact, previous selection procedure relying only on in vitro antibacterial activity was shown to be not suitable in some cases. A direct plant-protection strategy was performed to screen the 32 Bacillus biocontrol agent candidates. Moreover, potential in vitro biocontrol traits were investigated including biofilm formation, motility, hemolytic activity, detection of lipopeptide biosynthetic genes (sfp, ituC and bmyB) and production of antibacterial compounds. The obtained results indicated high correlations of the efficiency of the biocontrol with the reduction of gall weight (p=0.000) and the antibacterial activity in vitro (p=0.000). Moreover, there was strong correlations of the efficiency of the biocontrol (p=0.004) and the reduction in gall weight (p=0.000) with the presence of the bmyB gene. This gene directs the synthesis of the lipopeptide bacillomycin belonging to the iturinic family of lipopeptides. These results were also confirmed by the two-way hierarchical cluster analysis and the correspondence analysis showing the relatedness of these four variables. According to the obtained results a new screening procedure of Bacillus biocontrol agents against crown gall disease could be advanced consisting on two step selection procedure. The first consists on selecting strains with high antibacterial activity in vitro or those harboring the bmyB gene. Further selection has to be performed on tomato plants in vivo. Moreover, based on the results of the biocontrol assay, five potent strains exhibiting high biocontrol abilities were selected. They were identified as Bacillus subtilis or Bacillus amyloliquefaciens. These strains were found to produce either surfactin or surfactin and iturin lipopeptides. In conclusion, our study presented a new and effective method to evaluate the biocontrol ability of antagonistic Bacillus strains against crown gall disease that could increase the efficiency of screening method of biocontrol agents. Besides, the selected strains could be used as novel biocontrol agents against pathogenic A. tumefaciens strains.