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The vascular wilt of date palm (Phoenix dactylifera L.), also known as Bayoud and caused by Fusarium oxysporum f.sp. albedinis (FOA), is the most destructive disease in North Africa. The disease has resulted in huge economic losses due to declining crop yield and quality. The use of potential biocontrol agents is a sustainable and environmentally friendly strategy compared to synthetic fungicides. The use of date palm-associated microflora for the screening of native antagonistic bacteria with potential applications is the most promising way to control this disease. Thus, the epidemic palm groves (in the valley of M'zab-Ghardaia-Algeria) were chosen for the isolation of rhizospheric bacteria and endophytes from the soil and roots of healthy or infected plants. A total of 8 bacterial isolates (83, 84, 300, 333, 322, 260, 249, and 227) selected from 75 FOA-active strains, showed strong activity against several strains of FOA and other major plant pathogens. Their phylogenetic identification proved they belong to the genus Bacillus (Bacillus sp., B. subtilis, B. atrophaeus, B. halotolerans, B. amyloliquefasiens, and B. paralicheformis). Bacterial cultures and a cell-free culture supernatant tested alone or in co-culture showed FOA inhibitory activity. However, the effect of the co-culture did not show any synergy in both cases. Unlike diffusible compounds, volatile organic compounds did not show a significant antifungal ability. The concept of selecting biocontrol agents in vitro does not always guarantee in vivo performance. In addition to antibiosis, other strategies such as competition and resistance induction are required for biocontrol agent efficacy. To evaluate the biocontrol effect in vivo, germinated seeds of date palm were treated with bacteria, infected with the pathogen, and then incubated for 45 days at room temperature in the dark. The majority of the strains (84, 300, and 333) tested showed moderate fungistatic effects and the protection rate reached an average of 60%. In particular, promising results were obtained with B. paralicheniformis strain 260 which significantly protected palm seeds up to 85%, compared to its in vitro test where a low inhibition rate (27.77%) was recorded. Screening methods should be equipped with multifactorial or plant-mediated control mechanisms. Furthermore, these bacteria have shown other potentialities to improve plant growth and resist stressful conditions. Thus, an effective biocontrol agent must combine several beneficial characteristics to be used successfully in situ. In this respect, Bacillus remains the best candidate for biological control. [ABSTRACT FROM AUTHOR]

To explore proteolytic activity of endophytic fungi inhabiting date palm roots, a Penicillium bilaiae isolate, displaying the highest level of protease production, has been recovered. Response surface methodology (RSM) was applied to optimize culture conditions for protease production by the fungus. Plackett-Burman design allowed for screening of variables effective in protease production. Results indicated that temperature, initial pH and glucose concentration dramatically affect protease yield. These factors were further optimized using a Box-Behnken design and RSM. A combination of initial pH (6.26), temperature (24.5 °C), glucose (13.75 g/L), NaNO3 (1.5 g/L), MgSO4 (0.2 g/L), KH2PO4 (0.5 g/L) and KCl (0.5 g/L) were optimum for maximum production of protease. A 1086-fold enhancement of protease production was gained after optimization. Biochemical properties of fungal protease including the effect of pH and temperature on the activity and the stability of proteolytic enzyme were determined. Moreover, the influence of carbon and nitrogen sources, metal ions, detergents as well as enzyme inhibitors was investigated. Our results highlighted that protease of Penicillium bilaiae isolate TDPEF30 could be considered as a promising candidate for industrial applications. [ABSTRACT FROM AUTHOR]

Endophytic fungi of healthy and brittle leaf diseased (BLD) date palm trees (Phoenix dactylifera L.) represent a promising source of bioactive compounds with biomedical, industrial, and pharmaceutical applications. The fungal endophytes Penicillium citrinum isolate TDPEF34, and Geotrichum candidum isolate TDPEF20 from healthy and BLD date palm trees, respectively, proved very effective in confrontation assays against three pathogenic bacteria, including two Gram-positive bacteria Bacillus thuringiensis (Bt) and Enterococcus faecalis (Ef), and one Gram-negative bacterium Salmonella enterica (St). They also inhibited the growth of three fungi Trichoderma sp. (Ti), Fusarium sporotrichioides (Fs), Trichoderma sp. (Ts). Additionally, their volatile organic compounds (VOCs) were shown to be in part responsible for the inhibition of Ti and Ts and could account for the full inhibition of Fs. Therefore, we have explored their potential as fungal cell factories for bioactive metabolites production. Four extracts of each endophyte were prepared using different solvent polarities, ethanol (EtOH), ethyl acetate (EtOAc), hexane (Hex), and methanol (MetOH). Both endophyte species showed varying degrees of inhibition of the bacterial and fungal pathogens according to the solvent used. These results suggest a good relationship between fungal bioactivities and their produced secondary metabolites. Targeting the discovery of potential anti-diabetic, anti-hemolysis, anti-inflammatory, anti-obesity, and cytotoxic activities, endophytic extracts showed promising results. The EtOAc extract of G. candidum displayed IC50 value comparable to the positive control diclofenac sodium in the anti-inflammatory assays. Antioxidant activity was evaluated using α,α-diphenyl-β-picrylhydrazyl (DPPH), β-carotene bleaching, reducing power (RP), and 2,2-azino-bis(3-ethylbenzothiazoline-6-sulphonique) (ABTS) radical scavenging assays. The findings revealed strong anti-oxidant power with an IC50 of 177.55 µg/mL for G. candidum EtOAc extract using DPPH assay, probably due to high polyphenol and flavonoid content in both fungal extracts. Finally, LC-HRMS (Liquid Chromatography–High Resolution Mass Spectrometry) and GC-MS (Gas Chromatography–Mass Spectrometry) analysis of G. candidum and P. citrinum extracts revealed an impressive arsenal of compounds with previously reported biological activities, partly explaining the obtained results. Finally, LC-HRMS analysis indicated the presence of new fungal metabolites that have never been reported, which represent good candidates to follow for the discovery of new bioactive molecules. [ABSTRACT FROM AUTHOR]