Methylene blue (MB) has been shown to photoinduce the direct inactivation of Gram-positive bacteria Staphylococcus aureus (S. aureus) in water. We have investigated the mechanism of S. aureus photoinactivation conducting firstly sodium azide (reactive ion N3−), as a good physical singlet oxygen quencher, then the amino acids tryptophan (Trp), as a non-specific singlet oxygen quencher and the mannitol, as an hydroxyl free radical scavenger. Inactivation of MB photosensitization is the antioxidants type dependent. When the bacteria was treated with MB (20 μM) under light during 10 min of exposure, it was found that survival fraction had decreased dramatically to about 31.27±5.39%. The presence of sodium azide and Trp failed to shown any protection from the MB photodynamic activity. In the presence of mannitol, S. aureus could be protected, reaching a protection level of about 27%. It is possible that the photodynamic activity of MB occurred in part, via a Type I mechanism in which •OH was produced. The interactions between MB and S. aureus were studied spectrophotometrically. This demonstrated that a metachromatic reaction took place between MB and S. aureus bacteria. Furthermore, S. aureus bacteria induced additional dimerization of MB.

Wastewater sludge represents an important resource for reuse in agriculture. However, potentially harmful pathogens are a main threat in this context. Thus, the aim of this study was to examine the presence of helminth ova and protozoan cysts in dried sewage sludge samples collected from ten wastewater treatment plants (WWTPs) located in eight governorates in Tunisia. Based on morphological criteria, protozoan cysts of Giardia spp., Entamoeba histolytica/dispar/moshkovskii, and Entamoeba coli, were detected in all dried sludge composite samples (N = 116) from the investigated WWTPs. The mean concentration ranged from 1.4 to 10.7 cysts per 100 g dry matter (DM). The identified helminth eggs were Ascaris spp., Strongyles, Taeniid eggs, Hymenolepis nana, Enterobius vermicularis, and hookworm species. Ascaris spp. and Taeniid eggs were detected in 56.9 and 74.1% of analyzed samples, respectively. The presence of Trichuris spp., Hymenolepis diminuta, and Toxocara spp. eggs in dried sewage sludge samples was low (0.9, 1.7, and 2.6%, respectively). The mean concentration of helminth eggs during the three-year study was less than 1 egg/100 g DM. All examined dried sewage sludge sample contents were below the WHO (2006) and US EPA (2003) recommendations, and thus, the sludge can potentially be reused in agriculture.

In recent years, farmers in Beja, an agricultural governorate in northwestern Tunisia, have expressed their willingness to use urban sewage sludge as agricultural fertilizer, especially with the unavailability of chemical fertilizers and the soil type of the region that is poor in organic matter. However, there is an imbalance between the important farmers’ demand versus the limited quantity of sludge produced by the Beja wastewater treatment plants (WWTPs). In the face of this, this study aims to identify the problems related to the agricultural reuse of sludge in Beja and propose solutions to solve them. The quality of the sludge produced by the five Beja WWTPs was assessed based on physicochemical and microbiological parameters. The data were collected using the Delphi method, with 15 experts representing different positions on the issue treated. The SWOT-AHP methodology was used to define the strategies promoting the sustainable use and management of urban sewage sludge for sustainable agricultural development in Beja. Results showed that there were no problems with compliance with the Tunisian standards NT 106.20 for the sludge produced. A set of twelve practical conclusions was identified, constituting the strategies of Strengths–Opportunities, Strengths–Threats, Weaknesses–Opportunities, and Weaknesses–Threats deduced from the SWOT-AHP.

This study aims to follow the photodynamic and spectroscopic properties of dianionic rose Bengal disodium salt (RB) on Staphylococus aureus (S. aureus) in phosphate buffer solution (PBS) at pH 7.3. It focused on: (1) the effect of several reactive oxygen species (ROS) antioxidants used [such as sodium azide (NaN₃), l-tryptophan (l-Trp) and d-mannitol] on the RB photodynamic efficiency as a mean to identify the main ROS attributed, and (2) the possible interactions of the RB with the important singlet oxygen quencher used namely tryptophan and/or between the dye and the bacteria S. aureus thanks to a spectroscopic study. The results showed that 20 µM of RB and 10 min of visible light (50 mW/cm²) with a light fluence dose of 30 J/cm² are crucial for a good photodynamic action, achieving a reduction of 79.4% in the viability. Rose Bengal photodynamic action was in part inhibited by D-mannitol and l-Trp, indicating the mediation by.OH and ¹O₂, respectively. The high inhibition of the RB activity against S. aureus by l-Trp is not due only to its singlet oxygen quencher ability but it is mainly due to the interaction between RB and l-Trp as shown spectrophotometrically.

The aim of this study was to evaluate the photobactericidal effect of four photosensitizers (PSs) with different structural and physico-photochemical properties, namely mesotetracationic porphyrin (T4MPyP), dianionic rose Bengal (RB), monocationic methylene blue (MB) and neutral red (NR). Their photokilling activity was tested in vitro on pathogenic bacteria such as Pseudomonas aeruginosa (P. aeruginosa) and Staphylococcus aureus (S. aureus) suspended in nutrient broth (NB) and in phosphate buffered saline (PBS) through following their influence on the PSs antimicrobial efficacy. Photodynamic inactivation (PDI) experiments were performed using visible light (L) and different PSs concentrations (20–70 μM). The ability of these PSs to mediate bacterial photodynamic inactivation was investigated as a function of type of PS and its concentrations, spectral and physico-chemical properties, bacterial strain, irradiation time and suspending medium. Indeed, they showed antibacterial effects against S. aureus and P. aeruginosa with significant difference in potency. Staphylococcus aureus suspended in NB showed 0.92 log units reduction in viable count in the presence of T4MPyP at 20 μM. Changing the suspending medium from NB to PBS, S. aureus was successfully photoinactivated by T4MPyP (20 μM) when suspended in PBS at least time exposure (10 and 30 min), followed by MB and RB.

The present study aimed to: (i) identify helminth eggs and protozoan cysts in wastewater samples to which both human and animals could be exposed when they are reused in agriculture, and (ii) evaluate the efficiency of their removal by two wastewater treatment systems (activated sludge and lagoon). For these purposes, 234 wastewater samples (117 raw, 117 treated) were collected from 20 wastewater treatment plants (WWTPs) in Tunisia over a one-year sampling period. Parasitic load was enumerated by the modified Bailenger method. The results showed that helminth eggs found were Nematodes mainly represented by Ascaris sp. (95%), Hookworm species (50%), and Enterobius vermicularis (45%). For Cestodes, species identified were Taeniid eggs (85%), Hymenolepis nana (70%), Hymenolepis diminuta (55%) and digestive Strongyles. Among protozoa, Giardia sp., Entamoeba histolytica/dispar/moshkovskii and Entamoeba coli cysts were found in 100% of raw wastewater samples. The overall removal efficiency of helminth eggs and protozoan cysts in the treatment plants ranged from 56.5 to 100% and from 50.4 to 95.5%, respectively. The result from Multiple Correspondence Analysis (MCA) shows that the close clustering of the parasitic mean removal levels indicates that there is little difference in wastewater treatment processes between the WWTPs examined.