p53 over expression in yeast results in cell death with typical markers of apoptosis such as DNA fragmentation and phosphatidylserine externalization. We aimed to substitute/supplement classical fluorescent techniques (TUNEL, Annexin V, ROS detection) usually used to detect biochemical changes occurring during yeast apoptosis mediated by p53 over expression and the effect of anti-apoptotic purified molecules from Nigel (Nigella sativa) extracts on these same yeasts by the label free technique of FTIR spectroscopy. The comparison of the entire IR spectra highlighted clear modifications between apoptotic p53-expressing yeasts and normal ones. More precisely, DNA damage was detected by the decrease of band intensities at 1079 and 1048 cm-1. While phosphatidylserine exposure was followed by the increase of νsCH2 and νasCH2 bands of unsaturated fatty acids that were exhibited at 2855 and 2926 cm-1, and the appearance of the C = O ester functional group band at 1740 cm-1. In a second step, this FTIR approach was used to estimate the effect of a purified fraction of the Nigel extract. The modulation of band intensities specific to DNA and membrane status was in agreement with apoptosis supression in presence of the Nigel extracts. FTIR spectroscopy is thus proven to be a very reliable technique to monitor the apoptotic cell death in yeast and to be used as a means of evaluating the biomolecules effect on yeast survival.

A novel bioconversion approach of Tunisian wastewater to low-cost Photorhabdus temperata bioinsecticide is presented in this study. Our results showed that when cultured on the food industry wastewater (WS4), P. temperata cells exhibited oral toxicity of about 42%, which is the same as those cultured in complex medium (CM), used as control. Moreover, variants small colony polymorphism (Vsm) of the strain K122 was completely avoided after a prolonged incubation. However, viable but non-culturable (VBNC) state was enhanced with the maximum colony-forming units (CFU) count of 9 × 106 cells/mL obtained after 48 h of incubation in the WS4. According to flow cytometry analysis, almost 100% of P. temperata cells were viable until 48 h of incubation. The appearance of propidium iodide (PI) positively stained cells was observed after a prolonged incubation with a maximum of 17% of damaged cells in WS1. In order to follow the progress of P. temperata fermentation process carried out in industrial wastewater, we established for the first time, the mathematical relationship between total cell counts, CFU counts and oral toxicity of P. temperata strain K122. Indeed, irrespective of the medium used, the relationship between CFU count and total cell count followed a power law. Additionally, when plotting CFU count, or total cell count against toxicity, a semi-log linear relationship was obtained. Our results proved the efficiency of this bioconversion approach to produce bioinsecticide based on the entomopathogenic bacterium P. temperata, with practical benefits in terms of cost production and wastewater management.

This study investigates the optimization of the culture conditions for enhancing Photorhabdus temperata biopesticide production using wastewater (WS4) as a raw material. Box-Behnken design (BBD) was used to evaluate the effects of carbon to nitrogen ratio (C/N), sodium chloride concentration and inoculum size on P. temperata biomass production and insecticidal activity. For an enhanced biopesticide production, the optimum operating conditions were as follows: inoculum size = 4%; C/N ratio = 12.5 and [NaCl] = 4 g/L for two responses. 1.95 and 2.75 fold improvements in oral toxicity and biomass production were respectively obtained in the cost-effective medium developed in this study (WS4 I) using the three variables at their optimal values. Under the optimized conditions, WS4 I-grown cells exhibited higher membrane integrity according to flow cytometry analysis since dead cells presented only 9.2% compared to 29.2% in WS4. From batch fermentations carried out in WS4 I and WS4, P. temperata kinetic parameters in terms of biomass production and substrate consumption rates were modeled. The obtained results showed that the maximum specific growth rate in WS4 I was of 0.43 h−1 while that obtained in WS4 was of 0.14 h−1. In addition, the efficiency of P. temperata to metabolize organic carbon was enhanced by optimizing the culture conditions. It reached 72.66% instead of 46.18% in the control fermentation after 10 h of incubation. Under the optimized conditions, P. temperata cells showed the highest specific consumption rate resulting in a toxin synthesis improvement.

Latent membrane protein 1 (LMP1) encoded by the Epstein-Barr virus (EBV) plays an important role in EBV-induced cell transformation. Down-regulation of the LMP1 expression had shown promising results on cancer cell therapy. In this study, we identified by Phage display a novel peptide called B1.12 (ACPLDLRSPCG) which selectively binds to the extracellular loop (B1) of the LMP1 oncoprotein as demonstrated by molecular docking, NMR and ITC. Using an LMP1 expressing cell line, we showed that B1.12 decreased cell viability, and induced G0/G1 cell cycle arrest. In addition, the expression of A20, pAkt, and pNFkb (pRelA536) in C666-1 cells treated with B1.12 decreased compared to the untreated cells. In conclusion, we selected a novel peptide able to bind specifically to the extracellular loop of LMP1 and thus modulate its oncogenic properties.

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