Abderrazak Maaroufi

Rapid Aeromonas hydrophila identification by TaqMan PCR assay: comparison with a phenotypic method.

Aims:  Aeromonas hydrophila is recognized as a human pathogen following wound exposure or ingestion of contaminated water and food. For rapid identification of this bacterium, a TaqMan‐based real‐time PCR assay has been developed.

Anaerosalibacter bizertensis gen. nov., sp. nov., a halotolerant bacterium isolated from sludge

A strictly anaerobic, halotolerant and thermotolerant strain, designated C5BEL(T), was isolated in north Tunisia from storage tanks holding waste generated by the recycling of discarded motor oils. Cells of strain C5BEL(T) were Gram-stain-positive, motile by laterally inserted flagella, straight, and spore-forming. Their two major fatty acids were iso-C(15 : 0) and iso-C(15 : 0) dimethyl acetal. Growth was observed at temperatures of 25-55 °C (optimum, 40 °C) and at pH 6-9 (optimum, pH 7.5). The salinity range for growth was 0-100 g l(-1) NaCl (optimum, 5 g l(-1)). Yeast extract was required for growth. Strain C5BEL(T) was heterotrophic, able to use glucose, pyruvate, succinate, yeast extract, bio-trypticase and peptone, but unable to grow on Casamino acids. Sulfate, thiosulfate, sulfite, elemental sulfur, fumarate, nitrate and nitrite were not reduced. The DNA G+C content of strain C5BEL(T) was 31.1 mol%. 16S rRNA gene sequence analysis indicated that strain C5BEL(T) was a member of the family Clostridiaceae, class Clostridia, phylum Firmicutes and was most closely related to Sporanaerobacter acetigenes Lup33(T) ( = DSM 13106(T)) (92.4 % similarity). On the basis of 16S rRNA gene sequence comparisons and physiological characteristics, strain C5BEL(T) can be classified as a novel species in a new genus, for which the name Anaerosalibacter bizertensis gen. nov., sp. nov. is proposed. The type strain of the type species is C5BEL(T) ( = DSM 23801(T) = JCM 17239(T)).

Sporosalibacterium faouarense gen. nov., sp. nov., a moderately halophilic bacterium isolated from oil-contaminated soil.

A novel strictly anaerobic, moderately halophilic and mesophilic bacterium, designated strain SOL3f37(T), was isolated from a hydrocarbon-polluted soil surrounding a deep petroleum environment located in south Tunisia. Cells of strain SOL3f37(T) stained Gram-positive and were motile, straight and spore-forming. Strain SOL3f37(T) had a typical Gram-positive-type cell-wall structure, unlike the thick, multilayered cell wall of its closest relative Clostridiisalibacter paucivorans. The major fatty acids were iso-C(15 : 0) (41 %), iso-C(14 : 0) 3-OH and/or iso-C(15 : 0) dimethyl acetal (21.6 %), iso-C(13 : 0) (4.4 %), anteiso-C(15 : 0) (3.9 %) and iso-C(15 : 1) (2.8 %). Strain SOL3f37(T) grew between 20 and 48 °C (optimum 40 °C) and at pH 6.2-8.1 (optimum pH 6.9). Strain SOL3f37(T) required at least 0.5 NaCl l(-1) and grew in the presence of NaCl concentrations up to 150 g l(-1) (optimum 40 g l(-1)). Yeast extract (2 g l(-1)) was required for degradation of pyruvate, fumarate, fructose, glucose and mannitol. Also, strain SOL3f37(T) grew heterotrophically on yeast extract, peptone and bio-Trypticase, but was unable to grow on Casamino acids. Sulfate, thiosulfate, sulfite, elemental sulfur, fumarate, nitrate and nitrite were not reduced. The DNA G+C content was 30.7 mol%. Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain SOL3f37(T) was a member of the family Clostridiaceae in the order Clostridiales; strain SOL3f37(T) was related to members of various genera of the family Clostridiaceae. It exhibited highest 16S rRNA gene sequence similarity (93.4 %) with Clostridiisalibacter paucivorans 37HS60(T), 91.8 % with Thermohalobacter berrensis CTT3(T) and 91.7 % with Caloranaerobacter azorensis MV1087(T). On the basis of genotypic, phenotypic and phylogenetic data, it is suggested that strain SOL3f37(T) represents a novel species in a new genus. The name Sporosalibacterium faouarense gen. nov., sp. nov. is proposed, with SOL3f37(T) (=DSM 21485(T) =JCM 15487(T)) as the type strain of Sporosalibacterium faouarense.

Fatty acids composition of Tunisian Ziziphus lotus L. (Desf.) fruits and variation in biological activities between leaf and fruit extracts

This study was conceived to evaluate the essential fatty acids, secondary metabolites, antiradical and antimicrobial activities of unexploited Tunisian Ziziphus lotus L. The obtained results indicated that the major components of fatty acids were oleic acid (88.12%) and elaidic acid (7.88%). Leaves contained higher amount of total phenols, flavonoids and tannins than fruits, although both methanolic extracts had significant antioxidant activities. Significant correlations were observed between the total phenol or flavonoid contents in methanolic extracts and antioxidant activity estimated by using both 2,2′-diphenyl-1-picrylhydrazyl and 2,2′-azino-bis-3-ethylbenzothiazoline-6-sulfonic radical-scavenging methods. In addition, both methanolic extracts exhibited strong antibacterial and antifungal activities. The inhibition zone diameters and the minimal inhibition concentration values were in the range of 10–17 mm and 3.1–50 mg/mL, respectively.

Optimization of nutrient-induced germination of Bacillus sporothermodurans spores using response surface methodology

Spores of Bacillus sporothermodurans are known to be contaminant of dairy products and to be extremely heat-resistant. The induction of endospore germination before a heat treatment could be an efficient method to inactivate these bacteria and ensure milk stability. In this study, the nutrient-induced germination of B. sporothermodurans LTIS27 spores was studied. Testing the effect of 23 nutrient elements to trigger an important germination rate of B. sporothermodurans spores, only D-glucose, L-alanine, and inosine were considered as strong independent germinants. Both inosine and L-alanine play major roles as co-germinants with several other amino acids. A central composite experimental design with three factors (L-alanine, D-glucose, and temperature) using response surface methodology was used to optimize the nutrient-induced germination. The optimal rate of nutrient-induced germination (100%) of B. sporothermodurans spores was obtained after incubation of spore for 60 min at 35 °C in presence of 9 and 60 mM of D-glucose and L-alanine, respectively. The results in this study can help to predict the effect of environmental factors and nutrients on spore germination, which will be beneficial for screening of B. sporothermodurans in milk after induction their germination. Moreover, the chosen method of optimization of the nutrient-induced germination was efficient in finding the optimum values of three factors.

In vitro antimicrobial activity of Pistacia lentiscus L. edible oil and phenolic extract

Pistacia lentiscus L. is known in some Tunisian forest area by its fixed oil used in traditional medicine as an antiseptic product. This investigation is the first to study the antimicrobial activity of P.lentiscus edible oil and its phenolic extract. Oil was extracted from fruits harvested from six provenances located in Tunisia. The antimicrobial activity was tested using disc diffusion assay and the broth dilution method. Kbouch and Sidi Zid oils were most efficient (p < 0.003) against, respectively, Staphylococcus aureus and Aspergillus niger with an inhibition zone of 9.33 mm. The phenolic extract had the largest spectrum of sensitive microorganisms. The minimum inhibitory concentration and minimum bactericidal concentration results showed that all strains were inhibited by both oil and extract.

The combined effect of nisin, moderate heating and high hydrostatic pressure on the inactivation of Bacillus sporothermodurans spores

To investigate the combined effect of hydrostatic pressure (HP), moderate temperature and nisin on the inactivation of Bacillus sporothermodurans spores which are known to be contaminant of dairy products and to be extremely heat‐resistant.

Efficiency of refinery sludge biodegradation using municipal wastewater and activated sludge and effect of hydrocarbon concentration on culturable bacteria community

The efficiency of activated sludge as inoculum and municipal wastewater as diluent for the biodegradation of refinery sludge was investigated. At a laboratory scale, biodegradation experiments carried out in an aerobic batch reactor showed that toxic oily sludge was biodegradable after dilution. Compared with freshwater, the use of municipal wastewater for dilution clearly enhanced the biodegradation of this toxic pollutant. Using municipal wastewater, experiments showed that, starting from an initial total hydrocarbon concentration (THC) of 750 mg L−1, we could reach a final THC of 5 mg L−1 and a COD removal of 97% in only 19 days. In addition, we found that bacterial dominance was affected by hydrocarbon load. An absolute dominance of Gram-positive strains with high hydrocarbon load (7500 mg L−1) was found. However, with a hydrocarbon load of 750 mg L−1, some Gram-negative strains were also active. This bacterial dominance also changed in the course of treatment. We conclude that municipal wastewater and activated sludge, less often used nowadays in oily sludge biodegradation processes, contained bacterial strains that are efficient and able to work in a community. Municipal wastewater is also strongly recommended as a diluent as it offers an effective low-cost solution.

Evaluation of Pistacia lentiscus seed oil and phenolic compounds for in vitro antiproliferative effects against BHK21 cells

Abstract Context: Within the global context of increasing cancer diseases, natural products are important in devising new drugs and providing unique ideas in cancer therapy. In Tunisian folk medicine, Pistacia lentiscus L. (Anacardiaceae) fixed oil is used for cancer treatment. Objective: This investigation studied, for the first time, the antiproliferative effect of Pistacia lentiscus fixed oil and its phenolic extract on BHK21 cancer cells. Materials and methods: Oil was extracted from fruits harvested in northwest Tunisia and the phenolic fraction was obtained by mixing with methanol. The anti-proliferative activity of the two tested substances on BHK 21 cells were investigated in vitro using trypan blue assays. Cells were treated with different concentrations of P. lentiscus oil (0.009, 0.018, 0.036, and 0.09 g/mL) and the phenolic extract (0.007, 0.014, 0.03, and 0.07 g/mL) for 24, 48, and 72 h. Results: The inhibitory effect of Pistacia lentiscus fixed oil increases with the increase in dose. The IC50 value was estimated at 0.029 g/mL. The percentage of cell viability was 42.46 ± 3.4% at a dose of 0.09 g/mL and was significantly lower than that of the untreated control (96.24 ± 2.5%, p<0.01). The phenolic extract demonstrated a dose- and time-dependent inhibitory effect on BHK21 cell growth. After 48 h of incubation, the IC50 value was estimated at 0.15 g/mL. Discussion and conclusion: The results demonstrated the potential of Pistacia lentiscus fixed oil in treating cancer, as it is used in traditional medicine.

Chemical Composition of Ruta chalepensis Leaves Essential Oil and Variation in Biological Activities between Leaves, Stems and Roots Methanolic Extracts

Abstract: To our knowledge, this study is the first to investigate the biological activities of the methanolic extracts of leaves, roots and stems of Ruta chalepensis L. (Rutaceae) in Tunisia. The chemical composition of the essential oil extracted from R. chalepensis leaves was determined by GC-MS. The antioxidant activities of tested essential oil and the methanolic extracts of leaves, stems and roots of R. chalepensis were evaluated using DPPH and ABTS radicals scavenging tests. Their antimicrobial activities were achieved by disc diffusion method and broth dilution assay. The main components of the selected essential oil were menthol (43.92 %) and linalool (42.10 %). Significant differences were found between the total phenol contents and biological activities of the different parts of R. chalepensis. In addition, significant correlations were observed between antioxidant activities and phenolic contents for methanol extracts. In fact, leaves extract presented higher total phenol content than stems and roots extracts. The tested oil showed better antioxidant activity than methanolic extracts. The highest antioxidant activity was recorded in leaves methanolic extract. The latter showed strong antibacterial activity against tested microorganisms, whereas the evaluated oil remained almost inactive. The inhibition zones diameters and minimal inhibitrice concentration values for tested bacteria, which were sensible to different methanolic extracts, were in the range of 8-35 mm and 0.78-25 mg/ml, respectively. This study demonstrated that the R. chalepensis extract can be regarded as promising candidate for natural plant source of antioxidant and antimicrobial agents.