The heat-stable protein fraction from Opuntia ficus-indica seeds exhibits an enzyme protective effect against thermal denaturation and an antibacterial activity.

Desiccation tolerance in developing seeds occurs through several mechanisms among which, a common group of proteins named dehydrins has received considerable attention. So far, there is no information dealing with the accumulation of dehydrins in seeds of Opuntia ficus-indica. We have initiated here an extraction protocol based on two critical steps: heat and acid treatments, and the purity of this fraction was analyzed by FTIR spectroscopy. Western blot analysis of the heat-stable protein fraction (HSF) revealed two main bands of approximately 45 and 44 kDa, while three others of ∼40, 32, and 31 kDa were faintly visible, which were recognized by anti-dehydrin antibodies. This fraction exhibited a Cu2+ -dependent resistance to protease treatments. Next, we performed a series of assays to compare the functional properties of the HSF with those of the previously characterized wheat dehydrin (DHN-5). Antibacterial assays revealed that HSF exhibits only moderate antibacterial activities against gram-negative and gram-positive bacteria, with a minimum inhibition concentration ranging from 0.25 to 1 mg/ml. However, in vitro assays revealed that compared to DHN-5, HSF exhibits higher protective activities of the lactate dehydrogenase (LDH) when exposed to heat, freezing, and dehydration stresses. The protective role of HSF seems to be linked to its best ability to minimize protein aggregation.

Enhanced growth and metabolite production from a novel strain of Porphyridium sp.

Microalgae are capable of generating numerous metabolites that possess notable biological activities and hold substantial promise for various industrial applications. Nevertheless, the taxonomic diversity of these photosynthetic microorganisms has not received thorough investigation. Using the 18S rRNA encoding gene, a recently discovered strain originating from the Tunisian coast (the governorate of Mahdia) was identified as a member of the Porphyridium genus. The growth response as well as the metabolite accumulation of Porphyridium sp. to different culture media (Pm, F/2, and Hemerick) was investigated over a period of 52 days. The highest biomass production was recorded with Pm medium (2 × 107 cell/mL). The apparent growth rates (µ) and the doubling time (Dt) were about 0.081 day-1 and 12.34 days, respectively. The highest chlorophyll a (0.678 ± 0.005 pg/cell), total carotenoids (0.18 ± 0.003 pg/cell), phycoerythrin (3.88 ± 0.003 pg/cell), and proteins (14.58 ± 0.35 pg/cell) contents were observed with F/2 medium. Cultivating Porphyridium sp. in both F/2 and Hemerick media yielded similar levels of starch accumulation. The Hemerick medium has proven to be the most suitable for the production of lipids (2.23% DW) and exopolysaccharides (5.41 ± 0.56 pg/cell).

Lead removal from the aqueous solution by extracellular polymeric substances produced by the marine diatom Navicula salinicola.

Microbial extracellular polymeric substances (EPS) have recently emerged as significant contributors in diverse biotechnological applications. Extracellular polymeric substances (EPS), produced by a Navicula salinicola strain, have been studied for potential applications in a specific heavy metal (lead (Pb II)) removal from wastewater. The optimisation of operational parameters, mainly pH, Pb and EPS concentrations, using the Box-Behnken design (BBD) was undertaken to enhance lead uptake. The higher Pb adsorption capacity reached 2211.029 mg/g. Hydroxyl, carbonyl, carboxyl, phosphoric, and sulfhydryl groups were identified quantitatively as potential sites for Pb adsorption. EPS exhibited a notable flocculation rate of 70.20% in kaolin clay at a concentration of 15 mg/L. They demonstrated an emulsifying activity greater than 88%, showcasing their versatile potential for both sedimentation processes and stabilising liquid-liquid systems. EPS could be excellent nonconventional renewable biopolymers for treating water and wastewater.

Isolation and molecular characterization of an FSK2-type dehydrin from Atriplex halimus.

Dehydrins form the group II LEA protein family and are known to play multiple roles in plant stress tolerance and enzyme protection. They harbor a variable number of conserved lysine rich motifs (K-segments) and may also contain three additional conserved motifs (Y-, F- and S-segments). In this work, we report the isolation and characterization of an FSK2-type dehydrin from the halophytic species Atriplex halimus, which we designate as AhDHN1. In silico analysis of the protein sequence revealed that AhDHN1 contains large number of hydrophilic residues, and is predicted to be intrinsically disordered. In addition, it has an FSK2 architecture with one F-segment, one S-segment, and two K-segments. The expression analysis showed that the AhDHN1 transcript is induced by salt and water stress treatments in the leaves of Atriplex seedlings. Moreover, circular dichroism spectrum performed on recombinant AhDHN1 showed that the dehydrin lacks any secondary structure, confirming its intrinsic disorder nature. However, there is a gain of α-helicity in the presence of membrane-like SDS micelles. In vitro assays revealed that AhDHN1 is able to effectively protect enzymatic activity of the lactate dehydrogenase against cold, heat and dehydration stresses. Our findings strongly suggest that AhDHN1 can be involved in the adaptation mechanisms of halophytes to adverse environments.

Influence of the sulfate content of the exopolysaccharides from Porphyridium sordidum on their elicitor activities on date palm vitroplants.

Given the increasing interest that is being paid to polysaccharides derived from algae as plant natural defense stimulators, the degree of sulfation of exopolysaccharides produced by P. sordidum for inducing defense responses in date palm vitroplants was investigated. Firstly, the culture parameters of P. sordidum were optimized to maximize the amount of sulfate in EPS using a Box-Behnken experimental design and the elicitor effects of two EPS which differ in the sulfation degrees were compared. Results demonstrated that the concentrations of NaCl, NaNO3 and MgSO4 set at 28, 0.54 and 16.31 g/L, respectively yielded the best sulfate contents. To elucidate defense-inducing activities in date palm vitroplants, EPS with the highest sulfate content (EPS1) were prepared for comparison with those obtained under standard conditions (EPS0). A fucoidan extracted from Cystoseira compressa was used as positive control and MgSO4 as negative control. Both EPS and the fucoidan displayed H2O2 accumulation and expression of PR1, SOD, PAL and WRKY genes. Interestingly, EPS1 was significantly more bioactive than EPS0 and the fucoidan suggesting that the elicitor activity is positively correlated with the sulfate groups content of this polysaccharide.

Potential of three local marine microalgae from Tunisian coasts for cadmium, lead and chromium removals.

Metal elements are widely used in various industrial activities and are considered as common water source contaminants. Thus, the development of cost-effective, simple design and efficient processes for trace metal elements removal from contaminated water sources is of great interest. The effects of cadmium, lead and chromium on growth, biomolecules accumulation and metabolic responses of Amphora coffaeiformis, Navicula salinicola and Dunaliella salina isolated from Tunisian coasts were tested. The bioremediation capacities of the three microalgae strains and the mechanisms involved in ions metal removal were also investigated. N. salinicola and D. salina seem to be better tolerating to Cr, while A. coffaeiformis and N. salinicola showed high resistance to Pb. The expression profile analyses by qRT-PCR of the antioxidant defense-related genes revealed that Cd, Pb and Cr treatments induce the up-regulation of catalase and superoxide dismutase coding genes for A. coffaeiformis and D. salina. Regarding N. salinicola, the catalase coding gene seems to be overexpressed after Cd, Pb and Cr exposure while only Cd and Cr induce superoxide dismutase gene overexpression. Moreover, the phytochelatin synthase (a metal chelator synthesis-related gene) was up-regulated in N. salinicola, A. coffaeiformis and D. salina after Cr exposure and also in A. coffaeiformis and D. salina after Cd exposure. While Pb treatments induce overexpression of phytochelatin synthase coding gene only for D. salina. Studied strains showed promising metal removal efficiencies for both Pb and Cr ions metals reached 95% for D. salina. Ion metal removal mechanisms study revealed that intracellular bioaccumulation process is used by D. salina for Cr up-taking. However, both intracellular and extracellular removal mechanisms are involved for Pb and Cr removal using A. coffaeiformis, N. salinicola and for Pb removal using D. salina. FTIR analysis demonstrated that several functional groups as carboxyl, hydroxyl, amino, phosphate and sulfate may participate in the bioadsorption process.

Optimization of Exopolysaccharides Production by Porphyridium sordidum and Their Potential to Induce Defense Responses in Arabidopsis thaliana against Fusarium oxysporum.

Polysaccharides from marine algae are one novel source of plant defense elicitors for alternative and eco-friendly plant protection against phytopathogens. The effect of exopolysaccharides (EPS) produced by Porphyridium sordidum on elicitation of Arabidopsis thaliana defense responses against Fusarium oxysporum was evaluated. Firstly, in order to enhance EPS production, a Box-Behnken experimental design was carried out to optimize NaCl, NaNO3 and MgSO4 concentrations in the culture medium of microalgae. A maximum EPS production (2.45 g/L) higher than that of the control (0.7 g/L) was observed for 41.62 g/L NaCl, 0.63 g/L NaNO3 and 7.2 g/L MgSO4 concentrations. Structurally, the EPS contained mainly galactose, xylose and glucose. Secondly, the elicitor effect of EPS was evaluated by investigating the plant defense-related signaling pathways that include activation of Salicylic or Jasmonic Acid-dependent pathway genes. A solution of 2 mg/mL of EPS has led to the control of fungal growth by the plant. Results showed that EPS foliar application induced phenylalaline ammonia lyase and H2O2 accumulation. Expression profile analysis of the defense-related genes using qRT-PCR revealed the up-regulation of Superoxide dismutases (SOD), Peroxidase (POD), Pathogenesis-related protein 1 (PR-1) and Cytochrome P450 monooxyge-nase (CYP), while Catalase (CAT) and Plant defensin 1.2 (PDF1.2) were not induced. Results suggest that EPS may induce the elicitation of A. thaliana's defense response against F. oxysporum, activating the Salicylic Acid pathway.

Comparison of full-length and conserved segments of wheat dehydrin DHN-5 overexpressed in Arabidopsis thaliana showed different responses to abiotic and biotic stress.

Dehydrins (DHNs) are among the most common proteins accumulated in plants under water-related stress. They typically contain at least three conserved sequences designated as the Y-, S- and K-segments. The present work aims to highlight the role of the K-segments in plant tolerance to biotic and abiotic stresses. For this purpose, transgenic Arabidopsis thaliana (L.) Heyhn. lines expressing distinct wheat (Triticum aestivum L.) DHN-5 truncated constructs with or without the K-segments were generated. Our results showed that unlike the derivative lacking a K-segment, constructs containing only one or two K-segments enhanced the tolerance of A. thaliana to diverse stresses and were similar to the full-length wheat DHN-5. Moreover, compared with the wild-type and the YS form, the transgenic plants overexpressing wheat DHN-5 with K-segments maintained higher superoxide dismutase, catalase and peroxide dismutase enzymatic activity, and accumulated lower levels of H2O2 and malondialdehyde. In addition, we demonstrated that lines like A. thaliana overexpressing wheat DHN-5 showed increased resistance to fungal infections caused by Botrytis cinerea and Alternaria solani. Finally, the overexpression of the different forms of wheat DHN-5 led to the regulation of the expression of several genes involved in the jasmonic acid signalling pathway.

Wheat dehydrin K-segments ensure bacterial stress tolerance, antiaggregation and antimicrobial effects.

Dehydrins are a group of plant proteins that have been shown to be involved in the tolerance of various abiotic stresses such as dehydration, salinity, and low temperature. We have previously shown that the K-segments of the wheat dehydrin DHN-5 are essential for the protection of enzyme activities in vitro. In this study, we further investigate the role of the K-segments in the growth of Escherichia coli under various stresses, and we tested their antibacterial and antifungal activities. Our results showed that the truncated forms of DHN-5 containing the two K-segments enhanced tolerance of E. coli against diverse stresses by protecting proteins against aggregation. In addition, we demonstrated that the K-segments have antibacterial and antifungal activities against Gram-positive and Gram-negative bacteria and fungi. Based on these results, we propose that the K-segments may play a protective role in plants not only under abiotic stress conditions but also most likely during defense mechanisms.

The K-segments of the wheat dehydrin DHN-5 are essential for the protection of lactate dehydrogenase and ß-glucosidase activities in vitro.

The wheat dehydrin DHN-5 has been previously shown to exhibit heat protecting effect on enzymatic activities. In order to understand the molecular mechanism by which DHN-5 exerts its protective function, we performed an approach to dissect the functional domains of DHN-5 responsible for this feature. In two distinct enzymatic assays, we found that the truncated forms of DHN-5 containing only one K- or two K-segments are able to protect albeit to less extent than the wild type protein, lactate dehydrogenase and ß-glucosidase against damage induced by various stresses in vitro. However, the YS- and Φ-segments alone have no protective effects on these enzymes. Therefore, our study provides the evidence that the protective function of DHN-5 seems to be directly linked to its K-segments which through their amphipatic α-helical structure, may act to prevent protein aggregation.