Inhibition of Adherence and Biofilm Formation of Pseudomonas aeruginosa by Immobilized ZnO Nanoparticles on Silicone Urinary Catheter Grafted by Gamma Irradiation.

Nosocomial infections caused by microbial biofilm formation on biomaterial surfaces such as urinary catheters are complicated by antibiotic resistance, representing a common problem in hospitalized patients. Therefore, we aimed to modify silicone catheters to resist microbial adherence and biofilm formation by the tested microorganisms. This study used a simple direct method to graft poly-acrylic acid onto silicone rubber films using gamma irradiation to endow the silicone surface with hydrophilic carboxylic acid functional groups. This modification allowed the silicone to immobilize ZnO nanoparticles (ZnO NPs) as an anti-biofilm. The modified silicone films were characterized by FT-IR, SEM, and TGA. The anti-adherence ability of the modified silicone films was evidenced by the inhibition of biofilm formation by otherwise strong biofilm-producing Gram-positive, Gram-negative, and yeast clinical isolates. The modified ZnO NPs grafted silicone showed good cytocompatibility with the human epithelial cell line. Moreover, studying the molecular basis of the inhibitory effect of the modified silicone surface on biofilm-associated genes in a selected Pseudomonas aeruginosa isolate showed that anti-adherence activity might be due to the significant downregulation of the expression of lasR, lasI, and lecB genes by 2, 2, and 3.3-fold, respectively. In conclusion, the modified silicone catheters were low-cost, offering broad-spectrum anti-biofilm activity with possible future applications in hospital settings.

Comparative pathogenic potential of avian influenza H7N3 viruses isolated from wild birds in Egypt and their sensitivity to commercial antiviral drugs.

Active surveillance and studying the virological features of avian-origin influenza viruses are essential for early warning and preparedness for the next potential pandemic. During our active surveillance of avian influenza viruses in wild birds in Egypt in the period 2014-2017, multiple reassortant low-pathogenic avian influenza H7N3 viruses were isolated. In this study, we investigated and compared the infectivity, pathogenicity, and transmission of four different constellation forms of Egyptian H7N3 viruses in chickens and mice and assessed the sensitivity of these viruses to different commercial antiviral drugs in vitro. Considerable variation in virus pathogenicity was observed in mice infected with different H7N3 viruses. The mortality rate ranged from 20 to 100% in infected mice. Infected chickens showed only ocular clinical signs at three days postinfection as well as systemic viral infection in different organs. Efficient virus replication and transmission in chickens was observed within each group, indicating that these subtypes can spread easily from wild birds to poultry without prior adaptation. Mutations in the viral proteins associated with antiviral drug resistance were not detected, and all strains were sensitive to the antiviral drugs tested. In conclusion, all of the viruses studied had the ability to infect mice and chickens. H7N3 viruses circulating among wild birds in Egypt could threaten poultry production and public health.

Aspartate α-decarboxylase a new therapeutic target in the fight against Helicobacter pylori infection.

Effective eradication therapy for Helicobacter pylori is a worldwide demand. Aspartate α-decarboxylase (ADC) was reported as a drug target in H. pylori, in an in silico study, with malonic acid (MA) as its inhibitor. We evaluated eradicating H. pylori infection through ADC inhibition and the possibility of resistance development. MA binding to ADC was modeled via molecular docking. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of MA were determined against H. pylori ATCC 43504, and a clinical H. pylori isolate. To confirm selective ADC inhibition, we redetermined the MIC in the presence of products of the inhibited enzymatic pathway: ß-alanine and pantothenate. HPLC was used to assay the enzymatic activity of H. pylori 6x-his tagged ADC in the presence of different MA concentrations. H. pylori strains were serially exposed to MA for 14 passages, and the MICs were determined. Cytotoxicity in different cell lines was tested. The efficiency of ADC inhibition in treating H. pylori infections was evaluated using a Sprague-Dawley (SD) rat infection model. MA spectrum of activity was determined in different pathogens. MA binds to H. pylori ADC active site with a good docking score. The MIC of MA against H. pylori ranged from 0.5 to 0.75 mg/mL with MBC of 1.5 mg/mL. Increasing ß-alanine and pantothenate concentrations proportionally increased MA MIC. The 6x-his tagged ADC activity decreased by increasing MA concentration. No resistance to ADC inhibition was recorded after 14 passages; MA lacked cytotoxicity in all tested cell lines. ADC inhibition effectively eradicated H. pylori infection in SD rats. MA had MIC between 0.625 to 1.25 mg/mL against the tested bacterial pathogens. In conclusion, ADC is a promising target for effectively eradicating H. pylori infection that is not affected by resistance development, besides being of broad-spectrum presence in different pathogens. MA provides a lead molecule for the development of an anti-helicobacter ADC inhibitor. This provides hope for saving the lives of those at high risk of infection with the carcinogenic H. pylori.

Potential Antigenic Candidates for the Development of Peptide-Based Vaccines to Induce Immunization against Helicobacter pylori Infection in BALB/c Mice.

Helicobacter pylori (H. pylori) has been identified as a group-1 definite carcinogen. As of yet, there is no available vaccine for this microorganism. Our study aimed to identify antigenic peptides in H. pylori using an in silico proteomic approach, and to evaluate their effectiveness as potential vaccine candidates. Four different peptide sequences were prioritized using the reverse vaccinology, namely, CagA1, CagA2, VacA, and SabA. Peptides emulsified with Freunde's adjuvant were used to immunize BALB/C mice. Subcutaneously immunized mice were challenged by oral administration of H. pylori. IgG, IgA, IL4, and IL17 were detected in mice sera. Histopathology of the dissected stomach of vaccinated and control mice were assessed using H&E stain. IgG was significantly higher in mice vaccinated with SabA. IL-4 was significantly increased in CagA1, CagA2, VacA, and SabA vaccinated mice compared to the adjuvant group. Additionally, histopathological examination of gastric tissue showed a protective effect in the vaccinated groups compared to adjuvant and PBS groups. Our findings indicate a promising effect of the tested epitopes, particularly the SabA antigen, to induce an immune response against H. pylori.

Effect of spdC gene expression on virulence and antibiotic resistance in clinical Staphylococcus aureus isolates / Efecto de la expresión del gen spdC sobre la virulencia y la resistencia a los antibióticos en aislamientos clínicos de Staphylococcus aureus

Surface protein display C (SpdC) protein was described as a novel virulence factor of Staphylococcus aureus that affects biofilm formation and pathogenesis and favors resistance to antimicrobials targeting cell wall. We evaluated the possible correlation between spdC gene expression level and virulence as well as antibiotic resistance phenotypes in S. aureus clinical isolates. The antimicrobial susceptibility of S. aureus clinical isolates (n = 100) was determined by the disk diffusion method. Vancomycin susceptibility was determined by the broth microdilution method. The level of the extracellular proteases and delta-hemolysin was evaluated by measuring the proteolysis and hemolysis zone diameters in skim milk and blood agar plates, respectively. Biofilm formation was assayed using the 96-well microtiter plate method. Most of the isolates (81%) were multidrug-resistant and about half of the isolates (49%) were methicillin-resistant S. aureus. Hemolysin, protease, and biofilm production were detectable in 79%, 71%, and 96% of the isolates. No significant correlation was detectable between the level of spdC gene expression and the activity of tested virulence factors or the antimicrobial resistance phenotype. Therefore, the role of SpdC protein as a virulence regulator in S. aureus needs further evaluation together with the determination of the predominant regulators for each virulence factor.(AU)

Biodegradation, Decolorization, and Detoxification of Di-Azo Dye Direct Red 81 by Halotolerant, Alkali-Thermo-Tolerant Bacterial Mixed Cultures.

Azo dyes impact the environment and deserve attention due to their widespread use in textile and tanning industries and challenging degradation. The high temperature, pH, and salinity used in these industries render industrial effluent decolorization and detoxification a challenging process. An enrichment technique was employed to screen for cost-effective biodegraders of Direct Red 81 (DR81) as a model for diazo dye recalcitrant to degradation. Our results showed that three mixed bacterial cultures achieved ≥80% decolorization within 8 h of 40 mg/L dye in a minimal salt medium with 0.1% yeast extract (MSM-Y) and real wastewater. Moreover, these mixed cultures showed ≥70% decolorization within 24 h when challenged with dye up to 600 mg/L in real wastewater and tolerated temperatures up to 60 °C, pH 10, and 5% salinity in MSM-Y. Azoreductase was the main contributor to DR81 decolorization based on crude oxidative and reductive enzymatic activity of cell-free supernatants and was stable at a wide range of pH and temperatures. Molecular identification of azoreductase genes suggested multiple AzoR genes per mixed culture with a possible novel azoreductase gene. Metabolite analysis using hyphenated techniques suggested two reductive pathways for DR81 biodegradation involving symmetric and asymmetric azo-bond cleavage. The DR81 metabolites were non-toxic to Artemia salina nauplii and Lepidium sativum seeds. This study provided evidence for DR81 degradation using robust stress-tolerant mixed cultures with potential use in azo dye wastewater treatment.

Effect of spdC gene expression on virulence and antibiotic resistance in clinical Staphylococcus aureus isolates.

Surface protein display C (SpdC) protein was described as a novel virulence factor of Staphylococcus aureus that affects biofilm formation and pathogenesis and favors resistance to antimicrobials targeting cell wall. We evaluated the possible correlation between spdC gene expression level and virulence as well as antibiotic resistance phenotypes in S. aureus clinical isolates. The antimicrobial susceptibility of S. aureus clinical isolates (n = 100) was determined by the disk diffusion method. Vancomycin susceptibility was determined by the broth microdilution method. The level of the extracellular proteases and delta-hemolysin was evaluated by measuring the proteolysis and hemolysis zone diameters in skim milk and blood agar plates, respectively. Biofilm formation was assayed using the 96-well microtiter plate method. Most of the isolates (81%) were multidrug-resistant and about half of the isolates (49%) were methicillin-resistant S. aureus. Hemolysin, protease, and biofilm production were detectable in 79%, 71%, and 96% of the isolates. No significant correlation was detectable between the level of spdC gene expression and the activity of tested virulence factors or the antimicrobial resistance phenotype. Therefore, the role of SpdC protein as a virulence regulator in S. aureus needs further evaluation together with the determination of the predominant regulators for each virulence factor.

In Vitro and In Vivo Antiviral Studies of New Heteroannulated 1,2,3-Triazole Glycosides Targeting the Neuraminidase of Influenza A Viruses.

There is an urgent need to develop and synthesize new anti-influenza drugs with activity against different strains, resistance to mutations, and suitability for various populations. Herein, we tested in vitro and in vivo the antiviral activity of new 1,2,3-triazole glycosides incorporating benzimidazole, benzooxazole, or benzotriazole cores synthesized by using a click approach. The Cu-catalyzation strategy consisted of 1,3-dipolar cycloaddition of the azidoalkyl derivative of the respective heterocyclic and different glycosyl acetylenes with five or six carbon sugar moieties. The antiviral activity of the synthesized glycosides against wild-type and neuraminidase inhibitor resistant strains of the avian influenza H5N1 and human influenza H1N1 viruses was high in vitro and in mice. Structure-activity relationship studies showed that varying the glycosyl moiety in the synthesized glycosides enhanced antiviral activity. The compound (2R,3R,4S,5R)-2-((1-(Benzo[d]thiazol-2-ylmethyl)-1H-1,2,3-triazol-4-yl)methoxy)tetrahydro-2H-pyran-3,4,5-triyl triacetate (Compound 9c) had a 50% inhibitory concentration (IC50) = 2.280 µM and a ligand lipophilic efficiency (LLE) of 6.84. The compound (2R,3R,4S,5R)-2-((1-((1H-Benzo[d]imidazol-2-yl)methyl)-1H-1,2,3-triazol-4-yl)methoxy)tetrahydro-2H-pyran-3,4,5-triyl triacetate had IC50 = 2.75 µM and LLE = 7.3 after docking analysis with the H5N1 virus neuraminidase. Compound 9c achieved full protection from H1N1 infection and 80% protection from H5N1 in addition to a high binding energy with neuraminidase and was safe in vitro and in vivo. This compound is suitable for further clinical studies as a new neuraminidase inhibitor.

Epidemiological features of nosocomial Klebsiella pneumoniae: virulence and resistance determinants.

Aim: The authors aimed to examine antibiotic resistance genes and representative virulence determinants among 100 Klebsiella pneumoniae isolates with an emphasis on capsular serotypes and clonality of some of the isolates. Methods: PCR amplification of (rmpA, rmpA2, iutA, iroN and IncHI1B plasmid) and (NDM, OXA-48, KPC, CTX-M-15, VIM, IMP, SPM) was conducted. Wzi sequencing and multilocus sequence typing (MLST) were performed. Results: K2 was the only detected serotype in the authors' collection. RMPA2 was the most common capsule-associated virulence gene detected. All studied isolates harbored OXA-48-like (100%) and NDM (43%) (n = 43). ST147 was the most common sequence type. Conclusion: This work provides insight into the evolution of the coexistence of virulence and resistance genes in a tertiary healthcare setting in Cairo, Egypt.

Optimization strategy of Bacillus subtilis MT453867 levansucrase and evaluation of levan role in pancreatic cancer treatment.

Pancreatic cancer is the fourth most lethal cancer type worldwide. Due to multiple levan applications including anticancer activities, studies related to levansucrase production are of interest. To our knowledge, levan effect on pancreatic cancer cells has not been tested previously. In this work, among eighteen bacterial honey isolates, Bacillus subtilis MT453867 showed the highest levan yield (33 g/L) and levansucrase production (8.31 U/mL). One-factor-at-a-time technique increased levansucrase activity by 60% when MgSO4 was eliminated. The addition of 60 g/L banana peels enhanced the enzyme activity (192 U/mL). Placket Burman design determined the media composition for maximum levan yield (54.8 g/L) and levansucrase production (505 U/mL). The identification of levan was confirmed by thin-layer chromatography, Fourier-Transform Infrared spectrometric analysis, 13C-nuclear-magnetic resonance, and 1H-nuclear-magnetic resonance. Both crude and dialyzed levan completely inhibited the pancreatic cancer cell line at 100 ppm with no cytotoxicity on the normal retinal cell line. The LD50 of crude levan was 4833 mg/kg body weight. Levan had strong antioxidant activity and significantly reduced the expression of CXCR4 and MCM7 genes in pancreatic cancer cells with significant DNA fragmentation. In conclusion, Bacillus subtilis MT453867 levan is a promising adjunct to pancreatic-anticancer agents with both anti-cancer and chemoprotective effects.

Identification of Aspergillus species in human blood plasma by infrared spectroscopy and machine learning.

Invasive Aspergillosis is a challenging infection that requires convenient, efficient, and cost-effective diagnostics. This study addresses the potential of infrared spectroscopy to satisfy this clinical need with the aid of machine learning. Two models, based on Partial Least Squares-Discriminant Analysis (PLS-DA), have been trained by a set of infrared spectral data of 9 Aspergillus-spiked and 7 Aspergillus-free plasma samples, and a set of 200 spectral data simulated by oversampling these 16 samples. Two further models have also been trained by the same sets but with auto-scaling performed prior to PLS-DA. These models were assessed using 45 mock samples, simulating the challenging samples of patients at risk of Invasive Aspergillosis, including the presence of drugs (9 tested) and other common pathogens (5 tested) as potential confounders. The simple model shows good prediction performance, yielding a total accuracy of 84.4%, while oversampling and autoscaling improved this accuracy to 93.3%. The results of this study have shown that infrared spectroscopy can identify Aspergillus species in blood plasma even in presence of potential confounders commonly present in blood of patients at risk of Invasive Aspergillosis.

Identification of Potential Drug Targets in Helicobacter pylori Using In Silico Subtractive Proteomics Approaches and Their Possible Inhibition through Drug Repurposing.

The class 1 carcinogen, Helicobacter pylori, is one of the World Health Organization's high priority pathogens for antimicrobial development. We used three subtractive proteomics approaches using protein pools retrieved from: chokepoint reactions in the BIOCYC database, the Kyoto Encyclopedia of Genes and Genomes, and the database of essential genes (DEG), to find putative drug targets and their inhibition by drug repurposing. The subtractive channels included non-homology to human proteome, essentiality analysis, sub-cellular localization prediction, conservation, lack of similarity to gut flora, druggability, and broad-spectrum activity. The minimum inhibitory concentration (MIC) of three selected ligands was determined to confirm anti-helicobacter activity. Seventeen protein targets were retrieved. They are involved in motility, cell wall biosynthesis, processing of environmental and genetic information, and synthesis and metabolism of secondary metabolites, amino acids, vitamins, and cofactors. The DEG protein pool approach was superior, as it retrieved all drug targets identified by the other two approaches. Binding ligands (n = 42) were mostly small non-antibiotic compounds. Citric, dipicolinic, and pyrophosphoric acid inhibited H. pylori at an MIC of 1.5-2.5 mg/mL. In conclusion, we identified potential drug targets in H. pylori, and repurposed their binding ligands as possible anti-helicobacter agents, saving time and effort required for the development of new antimicrobial compounds.

Olive Leaf Extract Modulates Quorum Sensing Genes and Biofilm Formation in Multi-Drug Resistant Pseudomonas aeruginosa.

Biofilm acts as a complex barrier against antibiotics. In this study, we investigated the inhibitory activities of Olea europaea (olive) leaves Camellia sinensis (green tea), Styrax benzoin, Ocimum basilicum, Humulus lupulus, Ruta graveolens, and Propolis extracts on the biofilm formation, pyocyanin production, and twitching motility of Pseudomonas aeruginosa isolates. Moreover, we investigated the effect of olive leaf extract on the transcription of some biofilm related genes. A total of 204 isolates of Pseudomonas were collected from different Egyptian hospitals. A susceptibility test, carried out using the disc diffusion method, revealed that 49% of the isolates were multidrug-resistant. More than 90% of the isolates were biofilm-forming, of which 26% were strong biofilm producers. At subinhibitory concentrations, green tea and olive leaf extracts had the highest biofilm inhibitory effects with 84.8% and 82.2%, respectively. The expression levels of lasI, lasR, rhlI, and rhlR treated with these extracts were significantly reduced (p < 0.05) by around 97-99% compared to untreated isolates. This study suggests the ability of olive leaf extract to reduce the biofilm formation and virulence factor production of P. aeruginosa through the down regulation of quorum sensing (QS) genes. This may help in reducing our dependence on antibiotics and to handle biofilm-related infections of opportunistic pathogens more efficiently.

The Antimicrobial Activity of Ciprofloxacin-Loaded Niosomes against Ciprofloxacin-Resistant and Biofilm-Forming Staphylococcus aureus.

PURPOSE: The threat of Staphylococcus aureus antimicrobial resistance is increasing worldwide. Niosomes are a new drug delivery system that enhances the antimicrobial potential of antibiotics. We hereby aim to evaluate the antimicrobial and antibiofilm activity of ciprofloxacin-loaded niosomes. METHODS: The antimicrobial susceptibility of clinical S. aureus isolates (n=59) was determined by Kirby-Bauer disk diffusion method. Their biofilm formation activity was tested by Christensen's method. Two ciprofloxacin-loaded niosomal formulations were prepared by thin-film hydration method, and their minimum inhibitory concentrations (MIC) were determined by agar dilution method, against ciprofloxacin-resistant and biofilm-forming isolates (n=24). Their ability to inhibit biofilm formation and eradicate already formed biofilms was evaluated and further confirmed by scanning electron microscope images. Non-synonymous mutations, in a quinolone resistance-determining regions of S. aureus isolates, were detected by polymerase chain reaction. RESULTS: Most of the isolates were methicillin- (47/59) and ciprofloxacin-resistant (45/59). All except two isolates were capable of biofilm production. Niosomal preparation I reduced ciprofloxacin MIC by twofold in four isolates, whereas preparation II reduced ciprofloxacin MIC of most isolates by 8- to 32-fold, with three isolates that became ciprofloxacin-susceptible. Non-synonymous mutations were detected in isolates that maintained phenotypic ciprofloxacin resistance against ciprofloxacin-loaded niosomal preparation II. Ciprofloxacin-loaded niosomes reduced the minimum biofilm inhibitory concentration and the minimum biofilm eradication concentration in 58% and 62% of the tested isolates, respectively. CONCLUSION: Ciprofloxacin-loaded niosomes can restore ciprofloxacin activity against resistant S. aureus isolates. To our knowledge, this is the first report on the inhibition of biofilm formation and eradication of formed biofilms by ciprofloxacin-loaded niosomes.

Expression of P53, BAX, and BCL-2 in human malignant melanoma and squamous cell carcinoma cells after tea tree oil treatment in vitro.

Tea tree oil (TTO) is an essential oil obtained by steam distillation from the leaves of Melaleuca alternifolia (Myrtaceae). This oil has traditionally been used for the treatment of various skin infections. The present study aimed to investigate the cytotoxic effects of TTO against two representative types of human skin cancer, namely malignant melanoma (A-375) and squamous cell carcinoma (HEp-2).To outline the basic molecular mechanism involved in apoptosis induction in A-375 and HEp-2 cell lines, Annexin V/PI staining for apoptosis detection, cell cycle analysis were monitored using flow cytometry and mRNA expression levels of the apoptosis-regulatory genes P53, BAX, and BCL-2 were determined by real-time PCR and western blot after treatment with TTO. Results showed that TTO exhibited a strong cytotoxicity towards A-375 and HEp-2 cell lines, with IC50 values of 0.038% (v/v) and 0.024% (v/v) respectively. This cytotoxicity resulted from TTO induced apoptosis in both A-375 and HEp-2 cell lines as evidenced by morphological features of apoptosis and Annexin V/PI staining results in addition to the activation of caspase-3/7 and -9, upregulation of pro-apoptotic genes (P53 and BAX) and downregulation of the anti-apoptotic gene BCL-2. Additionally, cell cycle analysis showed that TTO caused cell cycle arrest mainly at G2/M phase. Taken together, the results of this study reveal that TTO is an effective apoptosis inducer in A-375 and HEp-2 cancer cell lines, indicating that it could be a promising chemopreventive candidate to be used in topical formulations against melanoma and squamous cell cancers; however, further in vivo studies may be warranted.

Origanum vulgare L. Essential Oil as a Potential Anti-Acne Topical Nanoemulsion-In Vitro and In Vivo Study.

Antibiotics are often prescribed in acne treatment; however, Propionibacterium acnes and Staphylococcus epidermidis, the two of the major acne-associated bacteria, developed antibiotic resistance. Essential oils (EOs) present a natural, safe, efficacious and multifunctional alternative treatment. This study aimed to assess the potential anti-acne activity of selected seven EOs commonly used in Mediterranean folk medicine. Antimicrobial activity screening of these oils showed oregano to exhibit the strongest antimicrobial activity with minimum inhibitory concentration (MIC) of 0.34 mg/mL and minimum bactericidal concentration (MBC) of 0.67 mg/mL against P. acnes; and MIC of 0.67 mg/mL and MBC of 1.34 mg/mL against S. epidermidis. The composition of the most effective EOs (oregano and thyme) was determined using gas chromatography-mass spectrometry (GC-MS). Monoterpenoid phenols predominated oregano and thyme EO with thymol percentile 99 and 72, respectively. Thymol showed MIC 0.70 mg/mL against both P. acnes and S. epidermidis whereas MBC was 1.40 and 2.80 mg/mL against P. acnes and S. epidermidis, respectively. Moreover, oregano exhibited the strongest anti-biofilm effect against S. epidermidis with MBIC 1.34 mg/mL and killing dynamic time of 12 and 8 h against P. acnes and S. epidermidis, respectively. Oregano, the most effective EO, was formulated and tested as a nanoemulsion in an acne animal mouse model. The formulation showed superior healing and antimicrobial effects compared to the reference antibiotic. Collectively, our data suggested that oregano oil nanoemulsion is a potential natural and effective alternative for treating acne and overcoming the emerging antibiotic resistance.

Prevalence of CagA and antimicrobial sensitivity of H. pylori isolates of patients with gastric cancer in Egypt.

BACKGROUND: Helicobacter pylori (H. pylori) infection has been recognized as a significant threat for gastric cancer. However, studies that investigated the oncogenic factors and antimicrobial resistance of H. pylori in Egyptian isolates with gastric cancer are rare. The current study aimed to examine: (1) The pattern of antimicrobial resistance of H. pylori isolates of Egyptian gastric cancer patients, and (2) the prevalence of Cytotoxin-associated gene A (CagA). METHODS: Samples were collected from patients with gastric cancer. Isolation of H. pylori was performed using Columbia blood agar supplemented with 10% horse blood, and selective supplement of H. pylori for 3 to 5 days at 37 °C under microaerophilic condition. Isolates were identified by biochemical traits of H. pylori: oxidase, urease and catalase tests. Antimicrobial susceptibility of H. pylori isolates was examined against five antimicrobial agents using disc diffusion method. After that, extraction of DNA and Polymerase Chain Reaction (PCR) were performed to amplify the target genes. RESULTS: Twelve samples were collected from six males and six females Egyptian patients with cancer with an age range from 22 to 65 years. These cases are scarce and samples were collected over a period of almost eleven months. All isolates were confirmed as positive H. pylori through colony morphology and biochemical tests. The most effective antibiotic found was ciprofloxacin whereas all isolates showed resistance to metronidazole and erythromycin. The target CagA oncogene gene with expected product size was reported and seven (out of twelve) isolates (58%) were identified as CagA positive. CONCLUSION: The current study is unique in two main aspects. First, it reported the pattern of antimicrobial susceptibility and prevalence of CagA gene in H. pylori from Egyptian patients. Second, it exclusively recruited isolates from gastric cancer patients which were confirmed by clinical and laparoscopic examination. The moderately high prevalence of CagA gene in Egyptian cancer patients calls for more vigilance against that oncogene.

Quercetin 3-O-glucoside recovered from the wild Egyptian Sahara plant, Euphorbia paralias L., inhibits glutamine synthetase and has antimycobacterial activity.

Tuberculosis remains a major health problem accentuated by the rise of resistance to all available drugs. Therefore, this study was launched to discover a novel antituberculosis agent from wild Egyptian Sahara plants. Twelve such plants were screened, in vitro, for their activity against various Mycobacterium species. The most active plant, Euphorbia paralias, was further fractionated with different organic solvents, and the activity of the obtained fractions was determined by the agar diffusion and broth microdilution methods. The methanol fraction was the most active against Mycobacterium spp., and was non-toxic in doses up to 10 g/kg of animal weight. Its main component was separated by column chromatography, and then identified by ultraviolet spectroscopy and nuclear magnetic resonance analysis as quercetin-3-O-ß-D-glucoside. Docking analysis suggested that quercetin-3-O-ß-D-glucoside inhibits the glutamine synthetase enzyme, a promising target for the development of antituberculosis drugs. This prediction was confirmed by an in vitro glutamine synthetase biosynthetic assay. To the best of our knowledge, and based on bioinformatics mining of the BioPhytMol database, this is the first report on the antimycobacterial activity of Euphorbia paralias plant. It is also the first report on the inhibition of mycobacterial glutamine synthetase by the flavonoid quercetin.

A Degradome-Based Polymerase Chain Reaction to Resolve the Potential of Environmental Samples for 2,4-Dichlorophenol Biodegradation.

A clean way to overcome environmental pollution is biodegradation. In this perspective, at the intersection of biodegradation and metagenomics, the degradome is defined as the totality of genes related to the biodegradation of a certain compound. It includes the genetic elements from both culturable and uncultured microorganisms. The possibility of assessing the biodegradation potential of an environmental samples, using a degradome-based polymerase chain reaction, was explored. 2,4-Dichlorophenol (2,4-DCP) was chosen as a model and the use of tfdB gene as a biodegradation marker was confirmed by bioinformatics study of TfdB protein. Five primer pairs were designed for the detection of different tfdB gene families. A total of 16 environmental samples were collected from Egyptian agricultural soils and wastewaters and tested for the presence of 2,4-DCP. The biodegradation capacity of 2,4-DCP was determined, for all isolated consortia, to reach up to 350 mg/l. Metagenomic DNA was extracted directly from the soil samples while successive 2,4-DCP-degrading microbial communities were enriched, with increasing concentrations of 2,4-DCP, then their DNA was extracted. The extracted DNA was tested for the distribution of the tfdB gene using a degradome-based polymerase chain reaction. tfdB-1 and tfdB-2 were detected in 5 and 9 samples, respectively. However, the co-existence of both genes was detected only in five samples. All tfdB positive samples were capable of 2,4-DCP degradation. The developed approach of assessing the potential of different environments for degrading 2,4-DCP was successfully measured in terms of accuracy (81.25%) and specificity (100%).