A tangible prospect for the treatment of gingivitis using a potentially probiotic strain Lactobacillus plantarum MK06 isolated from traditional dairy products: a triple blind randomized clinical trial.

BACKGROUND: Frequent bacterial plaque buildup at the gingival margin and crevice can provoke an inflammatory reaction in gingival tissues which manifests as gingivitis. Probiotics could serve as a beneficial complementary therapy for treating gingival inflammation. The main aim of this research was to investigate the effect of the Lactobacillus plantarum MK06 probiotic strain on the treatment of gingivitis. METHODS: Patients with gingivitis, who were referred to a private clinic and were systematically healthy, were included in this randomized, triple-blind, placebo-controlled trial. They were instructed to use either placebo or Lactobacillus plantarum suspensions for one minute two times a day after tooth-brushing for four weeks. Then, the clinical parameters of gingivitis, including plaque index (PI), gingival index (GI), bleeding on probing (BOP), and oral hygiene index (OHI-s), were measured in the first, second, and fourth weeks. A total of forty-two patients were randomly assigned to the experimental (n = 21) and control (n = 21) groups. The mean age of the experimental and control groups was 29.10 and 28.48, respectively. RESULTS: The mean scores of BOP, GI, PI, and OHI-s reduced over time in both the control and test groups. However, according to the Mann-Whitney test, the difference between the two groups was not significant at the same time intervals (P ≥ 0.05) and only GI showed a significant difference in the fourth week (GI-3, P = 0.006). Nevertheless, the experimental group experienced a higher overall reduction rate than the control group. The BOP, GI, PI, and OHI-s scores decreased by 0.081, 0.204, 0.186, and 0.172 times in the second week, respectively, resulting from the interaction of time and the intervention, which considerably diminished these indices. CONCLUSION: This study shows the potential of the probiotic Lactobacillus plantarum MK06 suspension as a promoting therapeutic adjuvant in the treatment of gingivitis.

Desertifilum sp. EAZ03 cell extract as a novel natural source for the biosynthesis of zinc oxide nanoparticles and antibacterial, anticancer and antibiofilm characteristics of synthesized zinc oxide nanoparticles.

AIMS: The use of cyanobacterial cell extracts for the synthesis of zinc oxide nanoparticles (ZnO NPs) seems to be superior to other methods of synthesis because of its a green, environmentally friendly and low-cost approach. In this study, the cell extract of a newly characterized cyanobacterial strain Desertifilum sp. EAZ03 was used for the biosynthesis of ZnO NPs. The antimicrobial, antibiofilm and anticancer activities of the biosynthesized ZnO NPs (hereinafter referred to as CED-ZnO NPs) were examined as well. METHODS AND RESULTS: UV-Vis spectroscopy analysis of CED-ZnO NPs showed an absorbance band at 364 nm, and powder X-ray diffraction analysis confirmed the purity of the synthesized nanoparticles. The analyses of scanning electron microscopy and transmission electron microscopy images revealed that CED-ZnO NPs were rod-shaped with a size of 88 nm. The study of the biological features of CED-ZnO NPs showed a significant antimicrobial potential against the bacterial strains tested. CED-ZnO NPs were able to impede the biofilm formation by Staphylococcus aureus, Escherichia coli and Pseudomonas aeruginosa up to 80%, 89% and 85%, respectively. The nanoparticles also showed 69%, 70% and 62% degrading activity against S. aureus, E. coli and P. aeruginosa 1-day-old biofilms, respectively. The antibiofilm activity of the synthesized nanoparticles was investigated by confocal laser scanning microscopy. The MTT assay showed that CED-ZnO NPs, at a concentration of 100 µg/ml, had less cytotoxicity towards normal lung (MRC-5) cells, at the half, compared to cancerous lung alveolar epithelial (A549) cells. The minimum inhibitory concentration and minimum bactericidal concentration values of CED-ZnO NPs against E. coli, P. aeruginosa and S. aureus were 1500, 2000 and 32 µg/ml, and 2500, 3500 and 64 µg/ml, respectively. CONCLUSIONS: The multifunctional CED-ZnO NPs seem to be promising for possible applications in the therapeutic and pharmaceutical industries. SIGNIFICANCE AND IMPACT OF THE STUDY: This study proposes a new approach for the biosynthesis of zinc oxide nanoparticles using a newly characterized cyanobacterial strain Desertifilum sp. EAZ03. The considerable antimicrobial, antibiofilm and anticancer activities of the biosynthesized zinc oxide nanoparticles further emphasize the emerging role of microbial systems in the green synthesis of metal oxide nanoparticles.

Comparison of Growth Performance, Pigment Synthesis, and Esterase Activity of Synechococcus sp. HS01 and Limnothrix sp. KO01 in Response to Cadmium Toxicity.

Various studies have been conducted to understand the impact of environmental pollutants on cyanobacteria due to their abundant presence in aquatic and terrestrial environments, specific morphological and physiological characteristics, and high ecological flexibility in response to environmental changes. Here, the effect of different concentrations of cadmium on two native strains of cyanobacteria, namely Synechococcus sp. HS01 and Limnothrix sp. KO01 was studied and compared with each other. In this regard, the cyanobacterial growth, pigment contents, and esterase enzyme activity were evaluated after exposure of the cells to different concentrations of cadmium (II). The toxic effects of Cd(II) on the growth rate of Limnothrix sp. KO01, even at low concentrations, tended to be higher than those for Synechococcus sp. HS01. The content of pigments decreased by an increase in Cd(II) concentration. In compliance with the cell growth, the changes occurred in pigment contents of Limnothrix sp. KO01 was more sensitive than Synechococcus sp. HS01 in the presence of different concentrations of cadmium. Flow cytometry analysis of Cd(II) effects on esterase activity of both strains after 6, 24, 48, and 72 h of exposure to Cd(II) concentrations of 9, 27, 63, and 90 µM showed that tolerance to Cd(II) toxicity in Limnothrix sp. KO01 is less than Synechococcus sp. HS01. The results obtained in this study suggest high potentials of Synechococcus sp. HS01 for heavy metal bioaccumulation due to its considerable tolerance to cadmium.

Multifunctional Acidocin 4356 Combats Pseudomonas aeruginosa through Membrane Perturbation and Virulence Attenuation: Experimental Results Confirm Molecular Dynamics Simulation.

A longstanding awareness in generating resistance to common antimicrobial therapies by Gram-negative bacteria has made them a major threat to global health. The application of antimicrobial peptides as a therapeutic agent would be a great opportunity to combat bacterial diseases. Here, we introduce a new antimicrobial peptide (∼8.3 kDa) from probiotic strain Lactobacillus acidophilus ATCC 4356, designated acidocin 4356 (ACD). This multifunctional peptide exerts its anti-infective ability against Pseudomonas aeruginosa through an inhibitory action on virulence factors, bacterial killing, and biofilm degradation. Reliable performance over tough physiological conditions and low hemolytic activity confirmed a new hope for the therapeutic setting. Antibacterial kinetic studies using flow cytometry technique showed that the ACD activity is related to the change in permeability of the membrane. The results obtained from molecular dynamic (MD) simulation were perfectly suited to the experimental data of ACD behavior. The structure-function relationship of this natural compound, along with the results of transmission electron microscopy analysis and MD simulation, confirmed the ability of the ACD aimed at enhancing bacterial membrane perturbation. The peptide was effective in the treatment of P. aeruginosa infection in mouse model. The results support the therapeutic potential of ACD for the treatment of Pseudomonas infections.IMPORTANCE Multidrug-resistant bacteria are a major threat to global health, and the Pseudomonas bacterium with the ability to form biofilms is considered one of the main causative agents of nosocomial infections. Traditional antibiotics have failed because of increased resistance. Thus, finding new biocompatible antibacterial drugs is essential. Antimicrobial peptides are produced by various organisms as a natural defense mechanism against pathogens, inspiring the possible design of the next generation of antibiotics. In this study, a new antimicrobial peptide was isolated from Lactobacillus acidophilus ATCC 4356, counteracting both biofilm and planktonic cells of Pseudomonas aeruginosa A detailed investigation was then conducted concerning the functional mechanism of this peptide by using fluorescence techniques, electron microscopy, and in silico methods. The antibacterial and antibiofilm properties of this peptide may be important in the treatment of Pseudomonas infections.

Nostoc entophytum cell response to cadmium exposure: A possible role of chaperon proteins GroEl and HtpG in cadmium-induced stress.

The present study is pursuing our previous research, focused on some aspects of Nostoc entophytum ISC32 cell response to the stress caused by exposure to cadmium at the cellular and molecular levels. Variations in the antioxidant system (catalase and ascorbate peroxidase activity) of N. entophytum ISC32 exposed to varying concentrations of Cd (2, and 5 mg/L) resulted in a significant increase in the activity of both catalase and peroxidase. Activity of these enzymes was, however, not significantly changed in the presence of Cd concentrations of 10 and 20 mg/L. Levels of lipid peroxidation, as measured by malondialdehyde (MDA) assay, were observed in response to exposure to Cd (20 mg/L). There was, however, a sharp drop in both antioxidant and lipid peroxidation activities of Cd treated cells after 5 days exposure, likely in consequence of cellular damage. The content of chlorophyll a and phycobiliproteins of living cells were altered under Cd-induced conditions. TEM images of cyanobacterial cells treated with Cd showed cell surface alteration and modification along with altered cellular microcompartments. Cyanobacterial cells treated with Cd at concentrations below the minimum inhibitory concentration (MIC) remained with no apparent structural changes. However, at a higher concentration of Cd (30 mg/L), a clear detachment effect was observed between the mucilage external layer and cell membrane which may be attributed to cell plasmolysis due to toxic effects of Cd. Subsequently, the thickness of the ring-shaped mucilage external layer increased likely as a result of the cell defense mechanisms against toxic concentrations of Cd. Characterization of cells treated with Cd (30 and 150 mg/L) by scanning electron microscopy (SEM) indicated cell shrinkage with varying degrees of distortion and surface wrinkling. Energy-dispersive X-ray spectrometry (EDS) analysis suggested that Cd was not present as nanoparticles within the cell, but in the form of salt or other molecular structures. The up-regulation of chaperons was confirmed for GroEL and HtpG using real-time PCR and northern blot analyses. Interestingly, the expression of GroEL was markedly increased at lower Cd concentration (5 mg/L). However, the ISC32 strain accrued higher levels of HtpG transcript in response to an elevated concentration of Cd (15 mg/L). This pattern seems to be related to the fast and early induction of GroEL, which may be necessary for induction of other factors and heat shock proteins such as HtpG in Cd-treated Nostoc cells. The result of this study paves the way for a more detailed exploration of Cd effects on the defense mechanisms of cyanobacteria. Our research also shed some light on how cyanobacterial cells have evolved to respond to the heavy metal toxicity at the cellular, molecular and ultrastructural levels.

High phenol degradation capacity of a newly characterized Acinetobacter sp. SA01: Bacterial cell viability and membrane impairment in respect to the phenol toxicity.

An efficient phenol-degrading bacterial strain, belonging to Acinetobacter genus, was isolated and selected to study the impact of different environmentally relevant phenol concentrations on the degradation process. The bacterial isolate, labeled as Acinetobacter sp. SA01 was able to degrade the maximum phenol concentration of 1 g/l during 60 h at optimum condition of pH 7, 30 °C and 180 rpm. Aeration and initial cell density, the two important factors, were carefully examined in the optimal growth conditions. The results showed that these two variables related proportionally with phenol degradation rate. Further investigations showed no effect of inoculum size on the enhancement of degradation of phenol at over 1 g/l. Flow cytometry (FCM) study was performed to find out the relationship between phenol-induced damages and phenol degradation process. Single staining using propidium iodide (PI) showed increased cell membrane permeability with an increase of phenol concentration, while single staining with carboxyfluorescein diacetate (cFDA) demonstrated a considerable reduction in esterase activity of the cells treated with phenol at more than 1 g/l. A detailed investigation of cellular viability using concurrent double staining of cFDA/PI revealed that the cell death increases in cells exposed to phenol at more than 1 g/l. The rate of cell death was low but noticeable in the presence of phenol concentration of 2 g/l, over time. Phenol at concentrations of 3 and 4 g/l caused strong toxicity in living cells of Acinetobacter sp. SA01. The plate count method and microscopy analysis of the cells treated with phenol at 1.5 and 2 g/l confirmed an apparent reduction in cell number over time. It was assumed that the phenol concentrations higher than 1 g/l have destructive effects on membrane integrity of Acinetobacter sp. SA01. Our results also revealed that the toxicity did not reduce by increasing initial cell density. Scanning electron microscopy (SEM) examination of bacterial cells revealed the surface morphological changes following exposure to phenol. The bacterial cells, with wizened appearance and wrinkled surface, were observed by exposing to phenol (1 g/l) at lag phase. A morphological change occurred in the mid-logarithmic phase as the bacterial cells demonstrated coccobacilli form as well as elongated filamentous shape. The wrinkled cell surface were totally disappeared in mid-stationary phase, suggesting that the complete degradation of phenol relieve the stress and direct bacterial cells toward possessing smoother cell membrane.

Biochemical properties of Glu-SH3 as a family 13 glycoside hydrolase with remarkable substrate specificity for trehalose: Implications to sequence-based classification of CAZymes.

A novel glycoside hydrolase from Exiguobacterium sp. SH3 was characterized. The enzyme, designated as Glu-SH3, was predicted by in silico analysis to have structural similarity with members of oligo-1,6-glucosidase and trehalose-6-phosphate hydrolase subfamilies in the GH-13 family of glycoside hydrolases. The gene was expressed in Escherichia coli and the recombinant enzyme was purified as a His-tagged protein of about 60 kDa. The enzyme was shown to have remarkable substrate specificity for trehalose. The characteristic ability of Glu-SH3 to hydrolyze trehalose was ascertained by zymography, thin layer chromatography, and NMR spectroscopy. The maximum activity of Glu-SH3 was obtained at 35 °C and pH 7, but it was able to exhibit more than 90% of the activity within the pH range of 5-8. The Vmax and Km values were estimated to be 170 U and 4.5 mg ml(-1), respectively. By comparison with trehalases, Glu-SH3 with Kcat and Kcat/Km values of 1552 s(-1) and 119.4 mM(-1) s(-1) can be recognized as a very efficient trehalose-hydrolyzing glycosidase. Given the phylogeny and the substrate specificity of Glu-SH3, it may be assumed that the enzyme shares a common ancestor with oligo-1,6-glucosidases but have evolved distinctly to serve a physiological function in trehalose metabolism.

Lipid production and mixotrophic growth features of cyanobacterial strains isolated from various aquatic sites.

The present study was conducted to determine the potential of five cyanobacteria strains isolated from aquatic zones to induce lipid production. The phylogenetic affiliation of the isolates was determined by 16S rRNA gene sequencing. Amongst the isolates, an efficient cyanobacterium, Synechococcus sp. HS01 showing maximal biomass and lipid productivity, was selected for further studies. In order to compare lipid productivity, the HS01 strain was grown in different media to screen potential significant culture ingredients and to evaluate mixotrophic cultivation. Mixotrophic cultivation of the strain using ostrich oil as a carbon source resulted in the best lipid productivity. GC analysis of fatty acid methyl esters of the selected cyanobacterial strain grown in media supplemented with ostrich oil showed a high content of C16 (palmitoleic acid and palmitic acid) and C18 (linoleic acid, oleic acid and linolenic acid) fatty acids of 42.7 and 42.8 %, respectively. Transmission electron micrographs showed that the HS01 cells exhibited an elongated rod-shaped appearance, either isolated, paired, linearly connected or in small clusters. According to initial experiments, ostrich oil, NaNO3 and NaCl were recognized as potential essential nutrients and selected for optimization of media with the goal of maximizing lipid productivity. A culture optimization technique using the response surface method demonstrated a maximum lipid productivity of 56.5 mg l(-1) day(-1). This value was 2.82-fold higher than that for the control, and was achieved in medium containing 1.12 g l(-1) NaNO3, 1 % (v/v) ostrich oil and 0.09 % (w/v) NaCl.

Characterization of a pH and detergent-tolerant, cold-adapted type I pullulanase from Exiguobacterium sp. SH3.

A pullulanase-encoding gene from psychrotrophic Exiguobacterium sp. SH3 was cloned and expressed in both E. coli and Bacillus subtilis. The functional recombinant enzyme (Pul-SH3) was purified as a His-tagged protein. Pul-SH3 was characterized to be a cold-adapted type I pullulanase with maximum activity at 45 °C. Using fluorescence spectroscopy, the melting temperature of Pul-SH3 was determined to be about 52 °C. The enzyme was able to hydrolyze pullulan, soluble starch, potato starch, and rice flour. It was exceptionally tolerant in the pH range of 4-11, exhibiting maximum activity at pH 8.5 and more than 60% of the activity in the pH range of 5-11. Being a detergent-tolerant pullulanase, Pul-SH3 retained 99, 89, and 54% of its activity at 10% concentration of Triton-X100, Tween 20, and SDS, respectively. The enzyme also exhibited an activity of 80.4 and 93.7% in the presence of two commercial detergents, Rika (7.5% v/v) and Fadisheh (2.5% w/v), respectively. The enzyme was even able to remain active by 54.5 and 85% after 10-day holding with the commercial detergents. Thermal stability of the enzyme could w on silica. Pul-SH3 with several industrially important characteristics seems desirable for cold hydrolysis of starch.

Calcium carbonate precipitation by strain Bacillus licheniformis AK01, newly isolated from loamy soil: a promising alternative for sealing cement-based materials.

The relevant experiments were designed to determine the ability of indigenous bacterial strains isolated from limestone caves, mineral springs, and loamy soils to induce calcium carbonate precipitation. Among all isolates examined in this study, an efficient carbonate-precipitating soil bacterium was selected from among the isolates and identified by 16S rRNA gene sequences as Bacillus licheniformis AK01. The ureolytic isolate was able to grow well on alkaline carbonate-precipitation medium and precipitate calcium carbonate more than 1 g L(-1). Fourier transform infrared (FTIR) spectroscopy, X-ray diffraction (XRD) analyses, and scanning electron microscopy (SEM)/energy-dispersive X-ray spectroscopy (EDX) examinations were performed in order to confirm the presence of calcium carbonate in the precipitate and to determine which polymorphs were present. The selected isolate was determined to be an appropriate candidate for application in a surface treatment of cement-based material to improve the properties of the mortar. Biodeposition of a layer of calcite on the surface of cement specimens resulted in filling in pore spaces. This could be an alternative method to improve the durability of the mortar. The kind of bacterial culture and medium composition had a profound impact on the resultant CaCO(3) crystal morphology.

A bacterial monorhamnolipid alters the biophysical properties of phosphatidylethanolamine model membranes.

This work presents a biophysical study on the interactions of a monorhamnolipid (monoRL) produced by Pseudomonas aeruginosa MA01 with model dielaidoylphosphatidylethanolamine (DEPE) membranes. Incorporation of monoRL into DEPE shifts the onset temperature of the Lß-to-Lα and the Lα-to-HII phase transitions toward lower values. Incorporation of monoRL into DEPE indicates the coexistence of lamellar and hexagonal-HII phases in rhamnolipid-containing samples at 60°C, at which pure DEPE is lamellar. Thus, both techniques show that monoRL facilitates formation of the hexagonal-HII phase in DEPE, i.e. it destabilizes the bilayer organization. The phase diagram for the phospholipid component indicates a near-ideal behavior, with better miscibility of monoRL into DEPE in the fluid phase than in the gel phase. The various vibrational mode bands of the acyl chains of DEPE were studied by FTIR spectroscopy, focusing on the CH2 symmetric stretching mode. Incorporation of monoRL into DEPE shifts the frequency of this band to higher wavenumbers, at temperatures both below and above the main gel to liquid-crystalline phase transition. Examination of the CO stretching band of DEPE indicates that monoRL/DEPE interactions result in an overall dehydration effect on the polar headgroup of DEPE. These results are discussed in light of the possible role of rhamnolipids as bilayer stabilizers/destabilizers during cell membrane fluctuation events.

Optimization of extracellular truncated staphylococcal protein A expression in Escherichia coli BL21 (DE3).

Staphylococcal protein A (SpA) plays an important role in Staphylococcus aureus pathogenesis. The recombinant SpA is also widely used in biotechnology to purify polyclonal and monoclonal immunoglobulin G antibodies. In this study, expression and secretion of a truncated form of SpA containing five immunoglobulin-binding domains using its own native signal sequence were optimized in Escherichia coli. Optimization was carried out using response surface method (RSM), making use of the interaction between five variables. The initial results revealed that the signal peptide from S. aureus was recognized in E. coli and the resulting SpA was expressed and secreted into the medium. Compounds, such as glycine, affected the secretion of SpA into the culture medium. The central composite design experiment showed that the optimum conditions for the maximum expression of recombinant truncated SpA in E. coli included 10% (w/v) lactose, 1.77% (w/v) glycine, induction time of 11 H, an optical density (600) of 1.1, and a temperature of 33 °C. Optimization using RSM resulted in a fivefold increase in the secretion of SpA. To date, this is the first study of its kind regarding the definite influence of glycine concentration and duration of the cultivation period on the secretion of SpA.

Lactobacillus paracasei BEJ01 prevents immunotoxic effects during chronic zearalenone exposure in Balb/c mice.

BACKGROUND AND AIM: Zearalenone (ZEN) is an estrogenic mycotoxin produced by numerous Fusarium species in pre- or post-harvest cereals. ZEN displays a potent estrogenicity in livestock and also causes severe immunological problems. The aims of this study were to isolate a new ZEN-degrading micro-organism for biological detoxification, to examine its ability to degrade ZEN in liquid medium, and to evaluate its potential for in vivo preventitive effects against ZEN (as would occur with contaminated feed)-induced immunomodulation in mice. MATERIALS AND METHODS: Lactobacillus paracasei BEJ01 (LP) isolated from Tunisian artisanal butter was found to display significant binding ability to ZEN in phosphate-buffered saline (i.e. 96.6%) within 24 h of incubation. The in vivo study was conducted using Balb/c mice that received either vehicle (control), LP only (at 2 × 10(9 )cfu/l, ∼2 mg/kg BW), ZEN alone (at 40 mg/kg BW), or ZEN + LP daily for 15 d. RESULTS: Compared to control mice, ZEN treatment led to significantly decreased body weight gains and decrements in all immune parameters assessed. The addition of LP to ZEN strongly reduced the adverse effects of ZEN on each parameter. In fact, mice receiving ZEN + LP co-treatment displayed no significant differences in the assayed parameters as compared to the control mice. The exposures to the bacteria alone had no adverse effects in the mice. CONCLUSION: From these data, we conclude that LP bacteria could be beneficial in human and animals for protection against immunotoxicity from ZEN at high levels and during chronic exposures.

Response surface optimization of biosurfactant produced by Pseudomonas aeruginosa MA01 isolated from spoiled apples.

A potent biosurfactant-producing bacterial strain isolated from spoiled apples was identified by 16S rRNA as Pseudomonas aeruginosa MA01. Compositional analysis revealed that the extracted biosurfactant was composed of high percentages of lipid (66%, w/w) and carbohydrate (32%, w/w). The surface tension of pure water decreased gradually with increasing biosurfactant concentration to 32.5 mN m(-1) with critical micelle concentration (CMC) value of 10.1 mg L(-1). The Fourier transform infrared spectrum of extracted biosurfactant confirmed the glycolipid nature of this natural product. Response surface methodology (RSM) was employed to optimize the biosynthesis medium for the production of MA01 biosurfactant. Nineteen carbon sources and 11 nitrogen sources were examined, with soybean oil and sodium nitrate being the most effective carbon and nitrogen sources on biosurfactant production, respectively. Among the organic nitrogen sources examined, yeast extract was necessary as a complementary nitrogen source for high production yield. Biosurfactant production at the optimum value of fermentation processing factor (15.68 g/L) was 29.5% higher than the biosurfactant concentration obtained before the RSM optimization (12.1 g/L). A central composite design algorithm was used to optimize the levels of key medium components, and it was concluded that two stages of optimization using RSM could increase biosurfactant production by 1.46 times, as compared to the values obtained before optimization.

New bacterial strain of the genus Ochrobactrum with glyphosate-degrading activity.

Thirty bacterial strains with various abilities to utilize glyphosate as the sole phosphorus source were isolated from farm soils using the glyphosate enrichment cultivation technique. Among them, a strain showing a remarkable glyphosate-degrading activity was identified by biochemical features and 16S rRNA sequence analysis as Ochrobactrum sp. (GDOS). Herbicide (3 mM) degradation was induced by phosphate starvation, and was completed within 60 h. Aminomethylphosphonic acid was detected in the exhausted medium, suggesting glyphosate oxidoreductase as the enzyme responsible for herbicide breakdown. As it grew even in the presence of glyphosate concentrations as high as 200 mM, Ochrobactrum sp. could be used for bioremediation purposes and treatment of heavily contaminated soils.

Interaction of Lactobacillus plantarum MON03 with Tunisian montmorillonite clay and ability of the composite to immobilize Zearalenone in vitro and counteract immunotoxicity in vivo.

BACKGROUND AND AIM: The present study was conducted to determine the abilities of the living Lactobacillus plantarum MON03 (LP) cells, Tunisian montmorillonite clay and their composites to accumulate Zearalenone (ZEA) from a liquid medium and elucidate the preventive effect of their composite in ZEA-contaminated balb/c mice showing immunotoxicity disorders. MATERIALS AND METHODS: In the in vitro study, LP (2 × 10(9) CFU/mL), TM (0.5 mg) and LP+TM were incubated with 50 µg mL(-1) ZEA for 0, 12 and 24 h. For the in vivo study, the composite MT+LP was evaluated also for possible protection regarding ZEA-immunotoxicity in Balb/c mice as a sensitive model. RESULTS: Results indicated that TM and LP+TM had a high capacity of adsorbing ZEA 87.2 ± 2.1 and 94.2 ± 2.1%, respectively. However, LP alone able to remove only 78% after 24 h of incubation. The quantity of adsorbed ZEA by LP, TM and LP+TM were 39, 43,5 and 47 µg mL(-1) of PBS, respectively. The in vivo results indicated that mice orally exposed to ZEA- (40 mg/kg bw) for 2 weeks showed severe immunotoxicity typical of fusarotoxicosis regarding thymocytes and splenocytes cell viability count, IFN-γ, IL-12, TNF-α production and B-cell activation. Mice treated with LP and TM alone, and LP+MT in combination with ZEA were comparable to the control. CONCLUSION: Both LP and TM are safe by themselves and their composite succeeded to exert a potential prevention by counteracting ZEA-immunotoxicity and can be implicated in the biotechnology of ZEA removal in human food and animal feed.

Type 2 IDI performs better than type 1 for improving lycopene production in metabolically engineered E. coli strains.

In this study a comparison was made between type 1 and type 2 isopentenyl diphosphate isomerases (IDI) in improving lycopene production in Escherichia coli. The corresponding genes of Bacillus licheniformis and the host (i(Bl) and i(Ec), respectively) were expressed in lycopene producing E. coli strains by pTlyci(Bl) and pTlyci(Ec) plasmids, under the control of tac promoter. The results showed that the overexpression of i(Ec) improved the lycopene production from 33 ± 1 in E. coli Tlyc to 68 ± 3 mg/gDCW in E. coli Tlyci(Ec). In contrast, the expression of i(Bl) increased the lycopene production more efficiently up to 80 ± 9 mg/gDCW in E. coli Tlyci(Bl). The introduction of a heterologous mevalonate pathway to elevate the IPP abundance resulted in a lycopene production up to 132 ± 5 mg/gDCW with i(Ec) in E. coli Tlyci(Ec)-mev and 181 ± 9 mg/gDCW with i(Bl) in E. coli Tlyci(Bl)-mev, that is, 4 and 5.6 times respectively. When fructose, mannose, arabinose, and acetate were each used as an auxiliary substrate with glycerol, lycopene production was inhibited by different extents. Among auxiliary substrates tested, only citrate was an improving one for lycopene production in all strains with a maximum of 198 ± 3 mg/gDCW in E. coli Tlyci(Bl)-mev. It may be concluded that the type 2 IDI performs better than the type 1 in metabolic engineering attempts for isoprenoid production in E. coli. In addition, the metabolic engineering of citrate pathway seems a promising approach to have more isoprenoid accumulation in E. coli.

Metagenomic psychrohalophilic xylanase from camel rumen investigated for bioethanol production from wheat bran using Bacillus subtilis AP.

Bioethanol produced from lignocellulosic biomass is regarded as a clean and sustainable energy source. The recalcitrant structure of lignocellulose is a major drawback to affordable bioethanol production from plant biomass. In this study, a novel endo-1,4-xylanase, named Xyn-2, from the camel rumen metagenome, was characterized and evaluated for hydrolysis of agricultural wastes. The enzyme was identified as a psychrohalophilic xylanase with maximum activity at 20 °C, keeping 58% of the activity at 0 °C, and exhibiting twice as much activity in 0.5-4 M NaCl concentrations. Xyn-2 was able to hydrolyze wheat bran (100%), sunflower-seed shell (70%), wheat straw (56%), rice straw (56%), and rice bran (41%), in the relative order of efficiency. Besides, the ethanologenic B. subtilis AP was evaluated without and with Xyn-2 for bioethanol production from wheat bran. The strain was able to produce 5.5 g/L ethanol with a yield of 22.6% in consolidated bioprocessing (CBP). The contribution of Xyn-2 to ethanol production of B. subtilis AP was studied in an SSF system (simultaneous saccharification and fermentation) giving rise to a significant increase in ethanol production (p ≤ 0.001) to a final concentration of 7.3 g/L with a yield of 26.8%. The results revealed that the camel rumen metagenome might be an invaluable source of novel xylanolytic enzymes with potential application in lignocellulosic biomass valorization. At the same time, the results suggest that B. subtilis with a diverse carbon-source preference and sophisticated systems for production and secretion of enzymes might be a promising candidate for strain development for bioethanol production from plant biomass. It might be assumed that the fortification of B. subtilis enzymatic arsenal with select xylanolytic enzymes from camel rumen metagenome may have a great impact on bioethanol production.

Effect of MA01 rhamnolipid on cell viability and expression of quorum-sensing (QS) genes involved in biofilm formation by methicillin-resistant Staphylococcus aureus.

A group of biosurfactants, called rhamnolipids, have been shown to have antibacterial and antibiofilm activity against multidrug-resistant bacteria. Here, we examined the effect of rhamnolipid biosurfactants extracted from Pseudomonas aeruginosa MA01 on cell growth/viability, biofilm formation, and membrane permeability of methicillin-resistant Staphylococcus aureus (MRSA) ATCC6538 bacterial cells. The results obtained from flow cytometry analysis showed that by increasing the concentration of rhamnolipid from 30 to 120 mg/mL, the cell viability decreased by about 70%, and the cell membrane permeability increased by approximately 20%. In fact, increasing rhamnolipid concentration was directly related to cell membrane permeability and inversely related to cell survival. Microtiter plate biofilm assay and laser scanning confocal microscopy analysis revealed that rhamnolipid, at a concentration of 60 mg/mL, exerts a reducing effect on the biofilm formation of Staphylococcus aureus. Real-time PCR analysis for monitoring the relative changes in the expression of agrA, agrC, icaA, and icaD genes involved in biofilm formation and related to the quorum-sensing pathway after treatment with rhamnolipid indicated a reduced expression level of these genes, as well as sortase A gene. The results of the present study deepen our knowledge regarding the use of microbial natural products as promising candidates for therapeutic applications.

Characterization of a chitinase with antifungal activity from a native Serratia marcescens B4A.

Chitinases have the ability of chitin digestion that constitutes a main compound of the cell wall in many of the phytopathogens such as fungi. In the following investigation, a novel chitinase with antifungal activity was characterized from a native Serratia marcescens B4A. Partially purified enzyme had an apparent molecular mass of 54 kDa. It indicated an optimum activity in pH 5 at 45°C. Enzyme was stable in 55°C for 20 min and at a pH range of 3-9 for 90 min at 25°C. When the temperature was raised to 60°C, it might affect the structure of enzymes lead to reduction of chitinase activity. Moreover, the Km and Vmax values for chitin were 8.3 mg/ml and 2.4 mmol/min, respectively. Additionally, the effect of some cations and chemical compounds were found to stimulate the chitinase activity. In addition, Iodoacetamide and Idoacetic acid did not inhibit enzyme activity, indicating that cysteine residues are not part of the catalytic site of chitinase. Finally, chitinase activity was further monitored by scanning electronic microscopy data in which progressive changes in chitin porosity appeared upon treatment with chitinase. This enzyme exhibited antifungal activity against Rhizoctonia solani, Bipolaris sp, Alternaria raphani, Alternaria brassicicola, revealing a potential application for the industry with potentially exploitable significance. Fungal chitin shows some special features, in particular with respect to chemical structure. Difference in chitinolytic ability must result from the subsite structure in the enzyme binding cleft. This implies that why the enzyme didn't have significant antifungal activity against other Fungi.