Cyclopropanation and membrane unsaturation improve antibiotic resistance of swarmer Pseudomonas and its sod mutants exposed to radiations, in vitro and in silico approch.

It was known that UVc irradiation increases the reactive oxygen species' (ROS) levels in bacteria hence the intervention of antioxidant enzymes and causes also changes in fatty acids (FAs) composition enabling bacteria to face antibiotics. Here, we intended to elucidate an interrelationship between SOD and susceptibility to antibiotics by studying FA membrane composition of UVc-treated P. aeruginosa PAO1 and its isogenic mutants (sodM, sodB and sod MB) membrane, after treatment with antibiotics. Swarmer mutants defective in genes encoding superoxide dismutase were pre-exposed to UVc radiations and then tested by disk diffusion method for their contribution to antibiotic tolerance in comparison with the P. aeruginosa wild type (WT). Moreover, fatty acid composition of untreated and UVc-treated WT and sod mutants was examined by Gaz chromatography and correlated to antibiotic resistance. Firstly, it has been demonstrated that after UVc exposure, swarmer WT strain, sodM and sodB mutants remain resistant to polymixin B, a membrane target antibiotic, through membrane unsaturation supported by the intervention of Mn-SOD after short UVc exposure and cyclopropanation of unsaturated FAs supported by the action of Fe-SOD after longer UVc exposure. However, resistance for ciprofloxacin is correlated with increase in saturated FAs. This correlation has been confirmed by a molecular docking approach showing that biotin carboxylase, involved in the initial stage of FA biosynthesis, exhibits a high affinity for ciprofloxacin. This investigation has explored the correlation of antibiotic resistance with FA content of swarmer P.aeruginosa pre-exposed to UVc radiations, confirmed to be antibiotic target dependant.

Proteomic and morphological insights into the exposure of Cupriavidus metallidurans CH34 planktonic cells and biofilms to aluminium.

Aluminium (Al) is one of the most popular materials for industrial and domestic use. Nevertheless, research has proven that this metal can be toxic to most organisms. This light metal has no known biological function and to date very few aluminium-specific biological pathways have been identified. In addition, information about the impact of this metal on microbial life is scarce. Here, we aimed to study the effect of aluminium on the metal-resistant soil bacterium Cupriavidus metallidurans CH34 in different growth modes, i.e. planktonic cells, adhered cells and mature biofilms. Our results indicated that despite a significant tolerance to aluminium (minimal inhibitory concentration of 6.25 mM Al2(SO4)3.18H2O), the exposure of C. metallidurans to a sub-inhibitory dose (0.78 mM) caused early oxidative stress and an increase in hydrolytic activity. Changes in the outer membrane surface of planktonic cells were observed, in addition to a rapid disruption of mature biofilms. On protein level, aluminium exposure increased the expression of proteins involved in metabolic activity such as pyruvate kinase, formate dehydrogenase and poly(3-hydroxybutyrate) polymerase, whereas proteins involved in chemotaxis, and the production and transport of iron scavenging siderophores were significantly downregulated.

Isolation and characterization of a newly chrysene-degrading Achromobacter aegrifaciens.

Polycyclic aromatic hydrocarbons (PAHs) are considered substances of potential human health hazards because of their resistance to biodegradation and carcinogenic index. Chrysene is a PAH with a high molecular weight (HMW) that poses challenges for its elimination from the environment. However, bacterial degradation is an effective, environmentally friendly, and cost-effective solution. In our study, we isolated a potential chrysene-degrading bacteria from crude oil-contaminated seawater (Bizerte, Tunisia). Based on 16SrRNA analysis, the isolate S5 was identified as Achromobacter aegrifaciens. Furthermore, the results revealed that A. aegrifaciens S5 produced a biofilm on polystyrene at 20 °C and 30 °C, as well as at the air-liquid (A-L) interface. Moreover, this isolate was able to swim and produce biosurfactants with an emulsification activity (E24%) over 53%. Chrysene biodegradation by isolate S5 was clearly assessed by an increase in the total viable count. Confirmation was obtained via gas chromatography-mass spectrometry (GC-MS) analyses. A. aegrifaciens S5 could use chrysene as its sole carbon and energy source, exhibiting an 86% degradation of chrysene on day 7. In addition, the bacterial counts reached their highest level, over 25 × 1020 CFU/mL, under the conditions of pH 7.0, a temperature of 30 °C, and a rotary speed of 120 rpm. Based on our findings, A. aegrifaciens S5 can be a potential candidate for bioremediation in HMW-PAH-contaminated environments.

Implication of Mn-cofactored superoxide dismutase in the tolerance of swarmer Pseudomonas aeruginosa to polymixin, ciprofloxacin and meropenem antibiotics.

The protective role of superoxide dismutase (Sod) against oxidative stress, resulting from the common antibiotic pathway of action, has been studied in the wild type and mutant strains of swarmer Pseudomonas aeruginosa, lacking Cytosolic Mn-Sod (sodM), Fe-Sod (sodB) or both Sods (sodMB).Our results showed that inactivation of sodB genes leads to significant motility defects and tolerance to meropenem. This resistance is correlated with a greater membrane unsaturation as well as an effective intervention of Mn-Sod isoform, in antibiotic tolerance.Moreover, loss of Mn-Sod in sodM mutant, leads to polymixin intolerance and is correlated with membrane unsaturation. Effectivelty, sodM mutant showed an enhanced swarming motility and a conserved rhamnolipid production. Whereas, in the double mutant sodMB, ciprofloxacin tolerance would be linked to an increase in the percentage of saturated fatty acids in the membrane, even in the absence of superoxide dismutase activity.The overall results showed that Mn-Sod has a protective role in the tolerance to antibiotics, in swarmer P.aeruginosa strain. It has been further shown that Sod intervention in antibiotic tolerance is through change in membrane fatty acid composition.

Musico-bioremediation of seafood canning wastewater by Yarrowia lipolytica.

Due to the lack of water resources and the harmful effects of wastewater on environment and human health, treatment of wastewater becomes necessary. The present study explored the effect of musical sounds on the biological treatment of seafood canning wastewater by using Yarrowia lipolytica. Our results showed that low frequency (21 Hz to 1356 Hz) and high frequency (21 Hz to 16,214 Hz) musical sounds stimulated the growth of Y. lipolytica and increased the polluant removal efficiency. Such treatment decreased significantly the chemical oxygen demand (COD) and salinity as well as the color of this wastewater. Our study revealed that low frequency musical sounds are more effective in COD (87.5%) and salinity (44%) reduction as well as the decolorization (86.46%) of this effluent. Additionally, after 7 days of incubation significant yeast cell dry biomass (3.46 ± 0.22 g/L) and single cell proteins (46.45 ± 0.7 mg/g) were obtained under low frequency waves. Musico-bioremediation represents an innovative ecotechnological approach to wastewater treatment with low operating costs and significant environmental benefits.

Damage of the swarmer Pseudomonas soil isolate cell by UVc as revealed by transmission electron microscopy.

The modeling of the response of living organisms to a change in environment is an important issue of current interest. An example is the effect of ultraviolet radiation on biological systems. In this paper, molecular and analytical identification of Pseudomonas isolate were reported. Then, swarmer Pseudomonas cells were exposed to UVc radiations. The spatiotemporal response of swarmer Pseudomonas, to UVc exposure, was followed. Observing alterations in bacterial membrane integrity by electron microscopy can help to clarify the detailed mechanisms of resistance to UVc. The most evident changes were related to membrane structures. In the cytoplasm, the main finding was the appearance of round mesosomes as intracellular bilayered membranes. Another impact of UVc on Pseudomonas was evident from the appearance of additional membrane structures. In accordance with the viability results, UVc-induced ultrastructural changes of Pseudomonas membrane structures were identified, resulting in cell death, through a multistage model of UVc inactivation.

Growth and biofilm formation of Cupriavidus metallidurans CH34 on different metallic and polymeric materials used in spaceflight applications.

Bacteria biofilm formation and its complications are of special concern in isolated structures, such as offshore stations, manned submarines and space habitats, as maintenance and technical support are poorly accessible due to costs and/or logistical challenges. In addition, considering that future exploration missions are planned to adventure farther and longer in space, unlocking biofilm formation mechanisms and developing new antifouling solutions are key goals in order to ensure spacecraft's efficiency, crew's safety and mission success. In this work, we explored the interactions between Cupriavidus metallidurans, a prevalently identified contaminant onboard the International Space Station, and aerospace grade materials such as the titanium alloy TiAl6V4, the stainless steel AISI 316 (SS316) and Polytetrafluoroethylene (PTFE) or Teflon. Borosilicate glass was used as a control and all surfaces were investigated at two different pH values (5.0 and 7.0). Biofilms were almost absent on stainless steel and the titanium alloy contrary to Teflon and glass that were covered by an extensive biofilm formed via monolayers of scattered matrix-free cells and complex multilayered clusters or communities. Filamentous extracellular DNA structures were observed specifically in the complex multilayered clusters adherent to Teflon, indicating that the employed attachment machinery might depend on the physicochemical characteristics of the surface.

Nanotechnology in drug and gene delivery.

Over the last decade, nanotechnology has widely addressed many nanomaterials in the biomedical area with an opportunity to achieve better-targeted delivery, effective treatment, and an improved safety profile. Nanocarriers have the potential property to protect the active molecule during drug delivery. Depending on the employing nanosystem, the delivery of drugs and genes has enhanced the bioavailability of the molecule at the disease site and exercised an excellent control of the molecule release. Herein, the chapter discusses various advanced nanomaterials designed to develop better nanocarrier systems used to face different diseases such as cancer, heart failure, and malaria. Furthermore, we demonstrate the great attention to the promising role of nanocarriers in ease diagnostic and biodistribution for successful clinical cancer therapy.

High-performance biological treatment of tuna wash processing wastewater using Yarrowia lipolytica.

It is well known that the lack of an effective treatment of tuna wash processing wastewater may pose substantial environmental and public health hazards. The present work investigates the performance of biological treatment of tuna wash processing wastewater (TWPW) by using Yarrowia lipolytica. Under optimized experimental conditions (pH "6.40-6.50" and 29 °C), Y. lipolytica reduced the pollution level of the crude and the diluted TWPW after only 7 days of incubation. The Yarrowia treatment leaded to a reduction of 66% chemical oxygen demand, 69.8% total organic carbon, 66% salinity, and phosphorus total (100%) removal of the crude TWPW, while the treated-diluted TWPW revealed significant reductions in chemical oxygen demand and total organic carbon (75% and 74%, respectively), as well as salinity (68%). Interestingly, a total removal of nitrogen and phosphorus from the diluted TWPW was obtained. Under high salinity, an important Y. lipolytica biomass of 5 g L-1 is produced with high levels of lipids and protein contents at around 336 ± 12.2 mg g-1 and 302.15 ± 5.44 mg g-1, respectively. The phytotoxicity assessment of the treated TWPW on fenugreek seeds shows promising results, which reveals the good performance of Yarrowia treatment in reducing the toxicity of this wastewater.

A new practical approach for the biological treatment of a mixture of cheese whey and white wastewaters using Kefir grains.

Kefir grains are a microbial consortium of different genera of bacteria and yeasts. In this study, the performance of Tunisian Kefir grains during the biological treatment of a mixture of Gouda cheese whey and white wastewaters (GCW) in ratio 1:1 with very high organic matter concentration is investigated. The biological process was evaluated and optimized through the response surface methodology. Under the optimum conditions, Kefir grains concentration of 1.02%, temperature at 36.68 °C, and incubation time of 5.14 days, the removal efficiencies of COD, PO43-, and NO3- were 87, 37.48, and 39.5%, respectively. Interestingly, the reusability tests of the grains proved not only their high resistance to harsh environmental conditions but also their great potential for more practical applications. Particularly, different strains were isolated from the grains and identified as Kluyveromyces marxianus, Lactoccocus lactis, Lactobacillus kefiri, and Bacillus spp. using 16S rDNA sequence analysis and rep-PCR fingerprinting. At the biological level, the raw GCW (RGCW) has a negative impact on the Hordeum vulgare both on seed germination, and on the growth parameters of seedlings. Interestingly, after Kefir grains treatment, the treated GCW (TGCW) allow a seedlings growth and germination rate similar to those soaked in water.

Size Fractionation of Titania Nanoparticles in Wild Dittrichia viscosa Grown in a Native Environment.

We report a size fractionation of titania (TiO2) nanoparticles absorbed from the environment and found within wild Dittrichia viscosa plants. The nanoparticles were isolated by extraction and isolation from distinct plant organs, as well as from the corresponding rhizosphere of wild, adult plants. The collected nanoparticles were characterized by scanning transmission electron microscopy coupled with energy dispersive X-ray spectroscopy (STEM-EDS). More than 1200 TiO2 nanoparticles were analyzed by these techniques. The results indicated the presence of TiO2 nanoparticles with a wide range of sizes within the inspected plant organs and rhizospheres. Interestingly, a size selective process occurs during the internalization and translocation of these nanoparticles (e.g., foliar and root uptake), which favors the accumulation of mainly TiO2 nanoparticles with diameters <50 nm in the leaves, stems, and roots. In fact, our findings indicate that among the total number of TiO2 nanoparticles analyzed, the fraction of the particles with dimensions <50 nm were 52% of those within the rhizospheres, 88.5% of those within the roots, 90% of those within the stems, and 53% of those within the leaves. This significant difference observed in the size distribution of the TiO2 nanoparticles among the rhizosphere and the plant organs could have impacts on the food chain and further biologicals effects that are dependent on the size of the TiO2.

Mixed culture of Lactococcus lactis and Kluyveromyces marxianus isolated from kefir grains for pollutants load removal from Jebel Chakir leachate.

The wastewater from the dumping site usually contains high pollutant levels. Biological process and physico-chemical treatments are among several technologies for wastewater treatment. Using microorganisms in the treatment of landfill leachate is an emerging research issue. Furthermore, bioremediation is a feasible approach for pollutants removal from landfill leachate which would provide an efficient way to resolve the issue of landfill leachate. In this study, the performance of yeast and bacteria isolated from kefir grains was assessed for landfill leachate treatment. Kefir grains microbial composition was evaluated by molecular approaches (Rep-PCR and 16S rRNA gene sequencing). The obtained outcomes denoted that high concentrations of lactic acid bacteria and yeast populations (over 107  CFU/ml) were found in the kefir grains and were essentially composed of Lactococcus lactis, Lactobaccillus kefirien, bacillus sp., L. lactis, and Kluyveromyces marxianus. The co-culture with 1% of inoculum size was demonstrated as the most efficient in the degradation of different contaminants. The overall abatement rate of chemical oxygen demand (COD), ammonium nitrogen ( NH 4 + - N ), and salinity were 75.8%, 85.9%, and 75.13%, respectively. The bioremediation process resulted in up of 75% removal efficiency of Ni and Cd, and a 73.45%, 68.53%, and a 58.17% removal rates of Cu, Pb, and Fe, respectively. The research findings indicate the performance of L. lactis and K. marxianus co-culture isolated from kefir grains for the bioremediation of LFL. PRACTITIONER POINTS: Isolation and identification of microorganisms from kefir grains was carried out. Biological treatment of LFL using monoculture of (Lactoccocus lactis; Kluyveromyces marxianus) and co-culture (5% of L. lactis and 5% K. marxianus) has been performed. Biological treatment using co-culture strain is an effective approach to remove organic matter, NH 4 + - N and heavy metals.

Aloe sp. leaf gel and water glass for municipal wastewater sludge treatment and odour removal.

Aloe gel (Alg), which is a natural extract from the Aloe sp. plant, was evaluated in this study for its potential use as a bioflocculant to treat urban wastewater sewage sludge. The gel was used alone and combined with water glass (WG) under controlled conditions in laboratory experiments. Alg was found effective to settle the flocculated sludge rapidly and remove distinctive unpleasant odours of the sludge as highlighted by gas chromatography-mass spectrometry (GC/MS) analysis. Furthermore, Alg was pH tolerant and had no effect in changing the pH of the wastewater. The optimum dose of Alg was 3% at which a sludge volume index (SVI) of 45.4 mL/g was obtained within 30 min settling time. To enhance the treatment performances of Alg, WG was also evaluated as an alkali agent to further reduce the chemical oxygen demand (COD) and ammonia (NH4-N) in the wastewater. At equal doses of 3% of WG and Alg each, the combined treatment outcomes showed high turbidity and NH4-N removals of 83 and 89%, respectively, but the overall COD removal was at best 25%. The settling rate of treated sludge with combined Alg/WG was very rapid giving an SVI of 25.4 mL/g within only 5 min.

Dual Valorization of Olive Mill Wastewater by Bio-Nanosynthesis of Magnesium Oxide and Yarrowia lipolytica Biomass Production.

This research investigates an efficient dual valorization of olive mill wastewater in the biosynthesis of magnesium oxide nanoparticles and in the depollution of the effluent by Yarrowia lipolytica growth evaluation. After removal of polyphenols, the recovered biophenols were reacted with the magnesium precursor to provide magnesium oxide nanoparticles. In order to confirm the biosynthesized magnesium oxide nanoparticles, several analyses were undertaken. The Fourier transform infrared spectrum gives a broad absorption at 658 cm-1 confirming the presence of the magnesium oxide nanoparticles, while the UV/VIS absorption spectroscopy reveals an intense transition with a maximum absorption at 300 nm. The X-ray diffraction and transmission electron microscopy analyses show that nanoparticles are in pure cubic crystalline with spherical and hexagonal shapes (average size is 19.4 nm). The zeta potential analysis illustrates a negative potential proving a good stability of the biosynthesized nanoparticles. Nanoparticles were assigned for their in vitro antibacterial activity against Escherichia coli, Enterobacter aerogenes, Salmonella typhimurium, Staphylococcus cohnii, and Bacillus niacini. The evaluation of the growth of Yarrowia lipolytica on the recovered olive mill wastewater after removal of polyphenols yielded 3.2 g/L of the Yarrowia biomass in 72 h without nutriment additions, providing an important decrease of chemical oxygen demand (73 %).

A new insight into highly contaminated landfill leachate treatment using Kefir grains pre-treatment combined with Ag-doped TiO2 photocatalytic process.

This study investigates the performance of the combination of biological pre-treatment with Kefir grains (KGs) and photocatalytic process using Ag-doped TiO2 nanoparticles (NPs) for the simultaneous removal of toxic pollutants from landfill leachate (LFL). After 5 days of 1% (w/v) KGs pre-treatment at 37 °C, TOC, COD, NH4+-N, and PO43- removal rates were 93, 83.33, 70 and 88.25%, respectively. The removal efficiencies were found to be 100, 94, 62.5, 53.16 and 47.52 % for Cd, Ni, Zn, Mn and Cu, respectively. The optimal conditions of Ag-doped TiO2 photocatalytic process were optimized using Box-Behnken design and response surface methodology (BBD-RSM) to enhance the quality of pre-treated LFL. Interestingly, Ag-doped TiO2 photocatalytic process increases the overall removal efficiencies to 98, 96, 85 and 93% of TOC, COD, NH4+-N, and PO43-, respectively. Furthermore, the removal efficiency of toxic heavy metals was gradually improved. In addition, KGs and Ag-doped TiO2 exhibited excellent recyclability showing the potential of combined biological/photocatalytic process to treat hazardous LFL.

Phosphorus and ammonium removal characteristics from aqueous solutions by a newly isolated plant growth-promoting bacterium.

An indigenous plant growth-promoting bacterium isolated from Peganum Harmala rhizosphere in the arid ecosystem was found to solubilize and accumulate phosphates. This isolate was identified as Pseudomonas sp. (PHR6) by partial 16S rRNA gene sequence analysis. Controlled batch experiments on nutrients removal by this isolate in mineral medium showed relatively high efficiencies after 24 h of aerobic incubation with average values of 117.59 and 335.38 mg gVSS-1 for phosphorus (P-PO4) and nitrogen (N-NH4), respectively. Furthermore, the strain performed heterotrophic nitrification ranging from 48.81% to 84.24% of the total removed nitrogen. On the other hand, the experimental results showed that a short idle period (24 h) significantly enhanced P accumulation (up to 95%) and N assimilation (up to 50%) of the total removed amounts. However, long idle period (20 days) revealed firstly aerobic phosphorous release phase succeeded by another removal one within 24 h of incubation. Overall, the idle treatment enhances P removal efficiency from the mineral liquid medium without significant effects on N-NH4 removal performance. The isolated strain showed also significant nutrient removal ability from synthetic wastewater providing an accumulated fraction of 98% from the total removed phosphorus amount. This study highlights the potential contribution of the selected rhizobacterium PHR6 to both environmental nutrient recycling and pollution control especially regarding phosphorus.

Differentiation of Nanoparticles Isolated from Distinct Plant Species Naturally Growing in a Heavy Metal Polluted Site.

Leaves harvested from the plants of two different species (Dittrichia viscosa and Cichorium intybus) grown in their autogenous environment near a steel manufacturing company were characterized for naturally accumulated nanoparticles. These plant species are known to accumulate heavy metals. It was, however, unknown if these species would also accumulate these heavy metals in the form of nanoparticles. The isolated solid fractions were analyzed using dynamic light scattering, X-ray fluorescence, and transmission electron microscopy. These analyses revealed the presence of nanoparticles within the plants. The composition of nanoparticles found in each plant species is distinct: (i) for Dittrichia viscosa, the nanoparticle composition matched the heavy metal pollution anticipated from the surrounding industries; (ii) for Cichorium intybus, the nanoparticle composition was similar to the most abundant elements in the soil. The different behavior is a reflection of the phytoaccumulator characteristics of both species. This study provides the first evidence of sequestration of heavy metals in the form of nanoparticles by plants grown autogenously in polluted areas and will have implications in waste management of phytoremediation systems and in understanding the heavy metal life-cycle in the environment.

Pseudomonas aeruginosa Swarmer Cells Adaptation Toward UVc Radiations.

Swarming is the most rapid surface motility allowing Pseudomonas aeruginosa bacteria to rapidly colonize new surfaces. However, swarming behavior is affected by environmental factors like ultraviolet irradiation (UVc). UVc radiation is the most disinfection technology usually applied for wastewater and proven to be effective to inactivate microorganisms. However, efficiency against motile bacteria is not yet studied. This study aims to explain the mechanisms of resistance of swarmer P. aeruginosa cells toward UVc exposure. P. aeruginosa liquid cultures were allowed to swarm across a semisolid surface for 18 h and directly exposed to UVc radiations. Emergent swarmer colonies, revealed after re-incubation, were selected to study biofilm formation, fatty acid (FA) composition, and ultrastructure. Our results showed that membrane adaptation to UVc radiations was seen in Pseudomonas cells by an increase of cyclic fatty acid (CFA) content, confirming the role of cyclopropane in radio-resistance of swarmer cells. Furthermore, electron microscopic study confirmed that over production of S-layer is believed to be a protective form adopted by P. aeruginosa swarmer cells to resist after 5 min of UVc exposure. Moreover, membrane disintegration is the lethal effect observed after 15 min of UVc exposure. In the other hand, study of biofilm production showed an enhancement of biofilm formation, of swarmer cells mainly after 15 min of UVc exposure. There results confirmed that swarming process is highly correlated with particular FA composition of P. aeruginosa membrane and that radio-resistance of swarmer cells is highly supported by CFA biosynthesis and S-layer overproduction.

UV-C Adaptation of Shigella: Morphological, Outer Membrane Proteins, Secreted Proteins, and Lipopolysaccharides Effects.

Water UV disinfection remains extremely important, particularly in developing countries where drinking and reclaimed crop irrigation water may spread devastating infectious diseases. Enteric bacterial pathogens, among which Shigella, are possible contaminants of drinking and bathing water and foods. To study the effect of UV light on Shigella, four strains were exposed to different doses in a laboratory-made irradiation device, given that the ultraviolet radiation degree of inactivation is directly related to the UV dose applied to water. Our results showed that the UV-C rays are effective against all the tested Shigella strains. However, UV-C doses appeared as determinant factors for Shigella eradication. On the other hand, Shigella-survived strains changed their outer membrane protein profiles, secreted proteins, and lipopolysaccharides. Also, as shown by electron microscopy transmission, morphological alterations were manifested by an internal cytoplasm disorganized and membrane envelope breaks. Taken together, the focus of interest of our study is to know the adaptive mechanism of UV-C resistance of Shigella strains.