Determination of MIC, MPC, and MSW of Ilex paraguariensis against non-typhoidal Salmonella with identification of the mechanisms of resistance and pathogenicity factors.

This study investigated the antibacterial activity of the aqueous extract of Ilex paraguariensis against 32 different strains of nontyphoidal Salmonella (NTS) through the determination of the minimum inhibitory concentration (MIC), mutant prevention concentration (MPC), and mutant selection window (MSW) and the detection of virulence genes by multiplex PCR assays. The MIC values of Ilex paraguariensis against Salmonella spp. strains varied between 0.78 mg/ml and 6.25 mg/ml with a MIC90 of 3.12 mg/ml. The highest MPC in this study was 48 mg/ml yielding a mutant selection window of 41.75 mg/ml. The MSW values of the remaining strains varied between 1.56 and 8.87 mg/ml. Genes of pathogenicity detected in Salmonella spp. isolates were most commonly the stn, sdiA, invA, sopB, invH, and sopE genes. The antibacterial activity of yerba mate extract was not affected by the antimicrobial resistance patterns or pathogenicity genes expressed. More work is needed to identify the active antibacterial compound(s) responsible for the antibacterial activity.

Citrus aurantium L. Active Constituents, Biological Effects and Extraction Methods. An Updated Review.

Citrus genus is a prominent staple crop globally. Long-term breeding and much hybridization engendered a myriad of species, each characterized by a specific metabolism generating different secondary metabolites. Citrus aurantium L., commonly recognized as sour or bitter orange, can exceptionally be distinguished from other Citrus species by unique characteristics. It is a fruit with distinctive flavor, rich in nutrients and phytochemicals which possess different health benefits. This paper presents an overview of the most recent studies done on the matter. It intends to provide an in-depth understanding of the biological activities and medicinal uses of active constituents existing in C. aurantium. Every plant part is first discussed separately with regards to its content in active constituents. All extraction methods, their concepts and yields, used to recover these valuable molecules from their original plant matrix are thoroughly reported.

Glucosinolates, a natural chemical arsenal: More to tell than the myrosinase story.

Glucosinolates are a group of thioglucosides that belong to the class of plant nitrogen-containing natural products. So far, very little biological activity has been associated with intact glucosinolates. The hydrolysis of glucosinolates has, for long, attracted attention because of the potent biological activity of the hydrolysis products. From allelopathic to antiparasitic, antimicrobial and antineoplastic effects, the activity spectrum of the degradation products of typical glucosinolates has been the subject of much research. The present review seeks to address the various means of glucosinolate degradation (thermal, enzymatic, or chemical degradation) and the ensuing products. It also aims to draw a comparative profile of the various antimicrobial effects of these degradation products to provide a further understanding of the biological function of these important compounds.

Mechanical damage and thermal effect induced by ultrasonic treatment in olive leaf tissue. Impact on polyphenols recovery.

The influence of ultrasound treatment (US) on cellular damage of olive leaf tissue was studied. Mechanical damage and thermal effect of US were characterized. The level of tissue damage was defined by the diffusivity disintegration index ZD based on the diffusivity of solutes extracted from olive leaves differently treated. The Arrhenius form using the temperature dependences of the thermal treatment time within the temperature interval 20-90 °C was observed for the thermal process. The corresponding activation energy ΔUT was estimated as 57 kJ/mol. The temperature dependences of electrical conductivity were measured for extracts of intact and maximally treated olive leaves. Then the diffusivity disintegration index ZD and total phenolic compounds recovery for three studied US powers were calculated (100, 200, and 400 W). The results evidenced that the mechanically stimulated damage in olive leaf tissue can occur even at a low US power of 100 W if treatment time is long enough (t = 3.5 h). The US treatment noticeably accelerated the diffusion process mechanically in addition to its thermal effect. Trials in aqueous solution revealed the dependence of polyphenols extraction on damage level with respect to the US power applied.

The antioxidant and anti-proliferative activity of the Lebanese Olea europaea extract.

It is becoming increasingly evident that certain phytochemicals possess cancer chemopreventive properties. In this study, the anti-proliferative activity of plant extracts from olive (Olea europaea L.) leaves was tested on human leukemic cell line (Jurkat). Cytotoxicity of various concentrations of plant extracts was examined and the IC(50) was determined. Olive leaf extracts showed concentration-dependent anti-proliferative effect as determined by the WST-1 proliferation kit and [(3)H]-thymidine incorporation method. To study whether cell death was due to apoptosis, cells were stained with Annexin V-FITC and PI and the expression of important regulatory proteins (Bcl-2, Bax, and p53) involved in apoptosis were examined by Western blot. The antioxidant activity of olive leaves (SC(50) = 0.1 mg dry weight) was studied using the DPPH scavenging method. Present findings suggest that olive leaves extracts exhibit anti-proliferative effect on leukemic cells by inducing apoptosis.

The antibacterial activity of Libanstin from Ilex paraguariensis (Yerba Mate).

Infectious diseases are reported to be one of the major causes of death in the world. The World Health Organization (WHO) warns of an increase in the deaths number because of antibacterial resistance. Lately, a trend towards searching for new active antibacterial compounds in plants has been observed. Ilex paraguariensis, known as Yerba Mate, is a plant known to be rich in numerous bioactive compounds that have an important role in human health. In this study, Yerba Mate was extracted with acetone: water (1:1) and further fractionated with hexane, chloroform and ethyl acetate. The obtained fractions were tested for antibacterial activity against Staphylococcus aureus and Salmonella species. The minimum inhibitory concentration (MIC) values on S. aureus ranged from 1.56 to 3.12 mg/mL for both the chloroform and ethyl acetate fractions. Whereas for the water fraction, the MIC values ranged from 0.78 to 3.12 mg/mL on S. aureus and ranged from 1.56 mg/mL to 3.12 mg/mL on Salmonella species. The aqueous fraction was further treated with different enzymes to mimic in vivo digestion and the fractions obtained were then tested for antibacterial activity. Furthermore, the Yerba Mate aqueous fraction was run on High Performance Liquid Chromatography (HPLC) and collected fractions were tested for antibacterial activity, to identify the active metabolite. Fraction 3 was tested on different strains of S. aureus and the MIC values ranged from 0.19 to 1.56 µg/mL. A novel pyrazinone, Libanstin, from Ilex paraguariensis was identified using NMR spectroscopy.

"Intensification of Vaporization by Decompression to the Vacuum" (IVDV), a novel technology applied as a pretreatment to improve polyphenols extraction from olive leaves.

Impact of the "Intensification of Vaporization by Decompression to the Vacuum" (IVDV) on extraction of polyphenols from olive leaves was investigated. Using Response Surface Methodology, the effect of three variables were studied: initial water content of leaves, processing time and steam pressure on total phenolic content (TPC). Extractions of TPC from leaves were achieved either using 100% water as a solvent (w100), or 50% (v/v) aqueous ethanol (w50). Following IVDV pretreatment, TPC yields were enhanced with both solvents by approximately 3 times compared to the negative controls. Furthermore, oleuropein and hydroxytyrosol were intensified by up to 600% and 238% respectively. Antioxidant-antiradical assays revealed higher activities, up to 3.5 times, in extracts from IVDV-treated leaves. Calculation of the extraction indices Zp, reflecting cellular damage, confirmed the beneficial effect of IVDV on the extraction yield. Finally, Scanning Electron Microscopy (SEM) permitted the morphological observation of IVDV-treated as compared to untreated leaves.

Modulation of membrane properties by silver nanoparticles probed by curcumin embedded in 1,2-Dimyristoyl-sn-glycero-3-phosphocholine liposomes.

Development of nanomaterials has drawn interest on silver nanoparticles (AgNPs), which are being incorporated in several biomedical and environmental applications, especially anti-bacterial properties of AgNPs has intense excitement for their commercial use. However, the impact of AgNPs on cell membranes, such as phospholipid membrane properties, is not clearly understood yet. By applying curcumin as a probe molecule, this work was done for the first time to investigate the effect of AgNPs on membrane properties, such as permeability and phase transition temperature using 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) liposomes as a model for phospholipid membranes. We concluded that AgNPs at low concentration decrease the partition of curcumin into DMPC liposomes by ∼4-fold. In the presence of AgNPs, curcumin was found to be located close to the stern layer of DMPC liposomes by using a hydrophobic quencher, cetylpyridinium bromide (CPB). In addition, AgNPs broadened the phase transition temperature of DMPC liposomes, which ranged from 20 °C to 35 °C.

Immunomodulatory effect of natural and modified Citrus pectin on cytokine levels in the spleen of BALB/c mice.

Pectin is present in the cell wall of different vegetables and fruits. Beside its importance in the plant cell wall, pectin has enticed great attention for its beneficial effects on human health. It was shown to decrease cholesterol levels, to possess anti-oxidative, anti-bacterial and anti-cancer activity. The immunomodulatory activity of pectin and its mechanism of action is recently being investigated. In this study, the differential immunomodulatory activities of both CP (citrus pectin) and MCP (modified citrus pectin) were investigated. Females BALB/c mice (20-25 g) were randomly divided into 7 groups and different concentrations of CP and MCP (0%, 1.5%, 3% and 5%) were added to their drinking water for 21 days. Then, the splenic level of IL-1ß, IL-4, IL-10, IL-17, IFN-γ and TNF-α were evaluated using ELISA. Both CP and MCP exhibited immunomodulatory activities by increasing the levels of the pro-inflammatory cytokines IL-17, IFN-γ and TNF-α levels. This tendency seems to be regulated by the up-regulation of IL-4 levels but with no major effect on those of IL-10. Therefore, CP and especially MCP have potential immunomodulatory effects which might be highly beneficial in immunotherapy.

Antimicrobial Activity of Polyphenols and Alkaloids in Middle Eastern Plants.

Antibiotic-resistant microorganisms have been an ever-growing concern over the past years. This has led researchers to direct their attention onto plants to be able to discover new possible antimicrobial compounds. The Middle East encompasses a wide spectrum of plant diversity with over 20,000 different species in habitats ranging from deserts to snow-capped mountains. Several plant secondary metabolites and their derivatives have been identified as possible antimicrobial agents. Among the secondary metabolites studied, alkaloids and polyphenols have shown strong antimicrobial activity. Polyphenols are one of the most numerous and diverse group of secondary metabolites; their antioxidant properties provide the basis for antimicrobial effects. Alkaloids provided the underlying structure for the development of several antibiotics with a diverse range of action. The ability of some plant secondary metabolites to act as resistance-modifying agents is a promising field in mitigating the spread of bacterial resistance.

Intensification of Polyphenol Extraction from Olive Leaves Using Ired-Irrad®, an Environmentally-Friendly Innovative Technology.

Optimization of infrared-assisted extraction was conducted using Response Surface Methodology (RSM) in order to intensify polyphenol recovery from olive leaves. The extraction efficiency using Ired-Irrad®, a newly-patented infrared apparatus (IR), was compared to water bath (WB) conventional extraction. Under optimal conditions, as suggested by the model and confirmed experimentally, the total phenolic content yield was enhanced by more than 30% using IR as contrasted to WB, which even required 27% more ethanol consumption. High Performance Liquid Chromatography analyses quantified the two major phenolic compounds of the leaves: Oleuropein and hydroxytyrosol, which were both intensified by 18% and 21%, respectively. IR extracts increased the antiradical activity by 25% and the antioxidant capacity by 51% compared to WB extracts. On the other hand, extracts of olive leaves obtained by both techniques exhibited equal effects regarding the inhibition of 20 strains of Staphylococcus aureus, with a minimum inhibitory concentration (MIC) varying between 3.125 and 12.5 mg/mL. Similarly, both extracts inhibited Aflatoxin B1 (AFB1) secretion by Aspergillus flavus, with no growth inhibition of the fungus. Finally, optimization using RSM allowed us to suggest other IR operating conditions aiming at significantly reducing the consumption of energy and solvent, while maintaining similar quantity and quality of phenolic compounds as what is optimally obtained using WB.

Resistance and virulence determinants of faecal Salmonella spp. isolated from slaughter animals in Benin.

OBJECTIVE: Salmonella spp. are one of the leading foodborne pathogens worldwide naturally found in the intestines of many animals. People that are in direct contact with the infected animals or their cages may become ill. The aim of this study was to determine the prevalence, antibiogram and virulence genes associated with Salmonella serovars from fecal samples of animals intended for consumption in Southern Benin. RESULTS: Out of a total of 406 samples, 2.46% were positive. The isolates identified were multidrug-resistant Salmonella spp. to penicillins, first generation cephalosporins and some aminoglycosides. All Salmonella isolates produced invA gene of 284 bp, fimA of 85 bp and stn of 260 bp. The spvC gene (571 bp) was present in 10% of the isolates whereas the spvR gene (310 bp) was found in 20% of the isolates. The control strain possessed all the tested genes. The invA gene implies that strains are able to invade epithelial cells. The fimA and stn genes present in all isolates show that they are capable of causing gastrointestinal illness in humans. The presence of spvC and spvR genes suggests the possibility of these strains to produce toxins.

Antibacterial activity of Ilex paraguariensis (Yerba Mate) against Gram-positive and Gram-negative bacteria.

INTRODUCTION: Limited research was performed on the antibacterial activity of the aqueous extract of Yerba Mate. The purpose of this study is to evaluate the anti-bacterial activity of Yerba Mate against Gram-positive and Gram-negative bacterial strains and its action against some resistant bacteria with genotypic molecular testing of resistance profiles. METHODOLOGY: Commercial Ilex paraguariensis stems and leaves were purchased and extracts were prepared by adding water at 70oC for 2 hrs. ATCC bacterial strains and clinical strains from Centre Hospitalier Du Nord (CHN) were used for testing. Macro dilution method was used to determine the minimal inhibitory. Minimal bactericidal concentration (MBC) was determined by sub-culturing the tubes with clear broth. For phenotypic and genotypic detection of ß-lactamases, Double Disk Synergy method, E-test, phenylboronic acid disc method and multiplex PCR were performed for the identification of the mechanisms of resistance. RESULTS: Antibacterial activity was observed against all tested strains, with a greater activity against Gram-positive bacteria. This study showed mostly a greater antibacterial activity of aqueous extract of Yerba Mate in comparison to different extraction methods published. In general, the MIC and MBC values ranged between 0.468 mg/mL and 15 mg/mL. No correlation was found between the different molecular resistance profiles and the antibacterial activity. CONCLUSION: More studies are needed to determine the molecule or molecules responsible for this activity. Moreover, testing a wider range of bacterial isolates is important for a better understanding of the potential role of Yerba Mate.

Multidrug-Resistant Enterobacteriaceae in Lebanese Hospital Wastewater: Implication in the One Health Concept.

Extended-spectrum beta-lactamases (ESBL) and carbapenemase-producing organisms pose severe problems for hospitalized patients. In the absence of efficient sanitation and sewage disposal, the risks for transmission of hospital organisms into the community are high. Our objectives were to study the occurrence and mechanisms of resistance of multidrug-resistant gram-negative bacilli in two Lebanese hospital sewage treatment plants. Wastewater samples were collected, filtered, and cultivated on MacConckey+cefotaxime agars. ESBL, AmpC, metallo-ß-lactamases (MBL), and Klebsiella pneumoniae Carbapenemases (KPC) production were phenotypically detected using plain Mueller Hinton agar plates, and plates impregnated with 5 mM EDTA, 10 mg/mL phenyl boronic acid, and 250 µg/mL cloxacillin (embedded). Temocillin discs were used for the presumptive detection of OXAs. ESBL, carbapenemase, outer membrane protein F (OMPF), and outer membrane protein C (OMPC) genes were detected using polymerase chain reaction. Pulsed-field gel electrophoresis (PFGE) was performed to study the clonality of Enterobacter cloacae isolates. In total, 32 and 38 Enterobacteriaceae were detected from Hospital 1 and Hospital 2, respectively. All Escherichia coli and Klebsiella spp. isolates were ESBL producers. AmpC reached 25% and 28.9% of all isolates. Only one Enterobacter cloacae isolate from one hospital showed full resistance to carbapenems. Molecular tests, however, detected NDM-1 in two strains of Enterobacter cloacae. PFGE results showed 80% similarity between these two strains. The isolation of NDM-1-producing Enterobacter cloacae from hospital wastewater occurred almost a year before the first case of carbapenem-resistant Enterobacter spp. was detected from a patient sample in the laboratory hospital. Understanding the local epidemiology of resistance in hospitals should include areas of potential resistance, such as wastewater and hospital environment.

Effects of partial enzymic degradation of sugar beet pectin on oxidative coupling of pectin-linked ferulates in vitro.

Pectins were extracted from roots and petioles of sugar beet, and treated with alpha-arabinosidase, 1,4-beta-galactanase or polygalacturonase. They were then cross-linked using hydrogen peroxide and peroxidase. The effects on pectin molecular size were monitored by size-exclusion chromatography and viscometry. A decrease in apparent molecular size was observed after alpha-arabinosidase and polygalacturonase treatment, and all three enzymes caused a decrease in viscosity. The pectins were then cross-linked using hydrogen peroxide and peroxidase, and the effects on dehydrodiferulate formation were monitored by HPLC. Pretreatment with polygalacturonase caused no significant change in subsequent dehydrodiferulate cross-linking, while pretreatment with alpha-arabinosidase caused a slight change in the ratios of the different dehydrodiferulates formed. Pretreatment with 1,4-beta-d-galactanase caused a more significant change in the ratios of the different dehydrodiferulates formed, and also greatly increased the overall recovery of total ferulates (monomers plus dehydrodiferulates), both in root pectin and petiole pectin. The possible effects of polysaccharide microstructure on the dimerisation and further polymerisation of pectin-linked ferulates are discussed.

Nascent pectin formed in Golgi apparatus of pea epicotyls by addition of uronic acids has different properties from nascent pectin at the stage of galactan elongation.

Microsomal membranes were prepared from etiolated pea (Pisum sativum L.) epicotyls and used to form nascent [Uronic acid-14C]pectin. The enzyme products were characterized by selective enzymic degradation, gel permeation chromatography and analysis of cellulose binding properties. The product obtained had a molecular weight of around 40 kDa, which was significantly lower than that of nascent [Gal-14C]pectin prepared from the same tissues. It is composed mainly of polygalacturonan and perhaps also rhamnogalacturonan (RG-I). Evidence was obtained for the presence of a protein attached to the nascent [Uronic acid-14C]pectin, but it was unaffected by endoglucanase and did not bind to cellulose. Hence, no xyloglucan appeared to be attached to the nascent [Uronic acid-14C]pectin. A model is proposed in which xyloglucan is attached to nascent pectin after formation of homogalacturonan, but before the pectin leaves the Golgi apparatus.

Additive Effect of MCP in Combination with Cefotaxime Against Staphylococcus aureus.

BACKGROUND: Pectin is a heterogeneous polysaccharide present in plants and citrus fruits. It exhibits different beneficial biological activities. Conflicting reports exist about the antimicrobial effect of pectin and its derivatives. OBJECTIVE: In this study, we investigate the antimicrobial effect of Modified Citrus Pectin (MCP) against Staphylococcus aureus, a pathogen showing increasing rates of antimicrobial resistance worldwide. METHOD: Forty-three clinical isolates of S. aureus were obtained from a hospital in North Lebanon. Minimum Inhibitory Concentrations (MICs) and Minimum Bactericidal Concentrations (MBCs) were determined using MCP after determining its optimum pH activity. The combination between MCP and cefotaxime was then investigated for S. aureus isolates using the checkerboard technique. RESULTS AND DISCUSSION: The optimum pH for the activity of MCP was 6.0. MIC and MBC values against S. aureus ranged between 0.39-50 µg/µl and 3.13-50 µg/µl, respectively. These values are promising for using MCP in the inhibition of some S. aureus isolates at relatively low concentrations. Combination experiments showed an additive effect in most S. aureus strains between MCP and cefotaxime, and a synergistic effect in two strains. These preliminary findings open the way for further investigation into the therapeutic potential of MCP in the treatment of S. aureus infections. CONCLUSION: MCP demonstrates in vitro antimicrobial activity alone and in combination with cefotaxime against S. aureus.

Physicochemical characteristics, mutagenicity and genotoxicity of airborne particles under industrial and rural influences in Northern Lebanon.

In this work, the main objectives were to assess the mutagenic and genotoxic effects of fine particulate matter collected in an industrial influenced site in comparison with a non-industrial influenced one (rural site) and to relate the particulate matter (PM) composition to the observed genotoxic effects. At the industrial influenced site, higher concentrations of phosphates, trace metals, and polycyclic aromatic hydrocarbons (PAHs) in particles could be related to the contributions of quarries, fertilizer producer, cement plants, and tires burning. Gasoline and diesel combustion contributions were evidenced in particles collected at both sites. Particles collected under industrial influence showed a higher mutagenic potential on three tested strains of Salmonella typhimurium (TA98, YG1041, and TA102), and especially on the YG1041, compared to particles from the rural site. Furthermore, only particles collected in the vicinity of the industrial site showed a tendency to activate the SOS responses in Escherichia coli PQ37, which is indicative of DNA damage as a result of exposure of the bacteria cells to the action of mutagenic samples. The mutagenicity and genotoxicity of the industrial PM2.5-0.3 particulates may be attributed to its composition especially in organic compounds. This study showed that proximity of industries can affect local PM composition as well as PM genotoxic and mutagenic potential.

The apoptotic and anti-proliferative activity of Origanum majorana extracts on human leukemic cell line.

Scientists are constantly searching for phytochemicals and compounds with anti-cancer and antioxidant activity. In this study, the anti-proliferative activity of plant extracts from Origanum majorana (marjoram) was tested on human lymphoblastic leukemia cell line Jurkat. Cytotoxicity was examined using non-radioactive cytotoxicity assay and the IC(50) was calculated. At non-cytotoxic concentrations, the viability of cells decreased with increase of concentration of plant extract. The anti-proliferative effect was also found to be dose-dependent. Analysis via flow cytometry shows that marjoram extracts stimulated apoptosis. Induction of apoptosis was caused by an up-regulation of p53 protein levels and down-regulation of Bcl-2alpha. Marjoram exhibited a strong scavenging activity (SC(50)=0.03mg dry weight). The conclusions from this study suggest that marjoram extracts exhibit anti-proliferative effect and high antioxidant activity. For that it merits further investigation as a potential therapeutic agent.

The effect of PEG-coated gold nanoparticles on the anti-proliferative potential of Specific Nutrient Synergy.

The role of PEG-coated gold nanoparticles (Au NPs) on the anti-proliferative effect of Specific Nutrient Synergy (SNS) on HTLV-1 infected (C91-PL and HuT-102) and non-infected (CEM and Jurkat) malignant T-lymphocytes cells, was investigated. When PEG-coated Au NPs (of different molecular weights) were added alone, there was no effect on either viability or proliferation of the leukemic cell lines studied. Treatment of cells with SNS and PEG (5 or 10 kDa) coated Au NP reduced significantly the proliferation in all cell lines tested; this reached more than 50% reduction as compared to the control for cells treated for 96 h. Data showed that the best anti-proliferative effect was obtained using SNS and Au NP coated with PEG of molecular weights of 5 and 10 kDa with almost no effect of PEG of lower molecular weights (0.75 and 2 kDa) or higher ones (20 kDa). This was true as well for HTLV-1 infected as for non-infected malignant T-lymphocytes. Electron microscopy results showed uptake of the gold particles to Jurkat cells. All described effects are specific to leukemia cell lines, and no effects were observed with freshly activated human mononuclear lymphocytes as control.