Search results for: microplate antimicrobial assay

Authors: Abdul Walusansa

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In this paper, paper notes and coins were collected from general public in Kampala City where ready-to-eat food can be served, in order to survey for bacterial contamination. The total bacterial number and potentially pathogenic organisms loading on currency were tested. All isolated potential pathogens were also tested for antibiotic resistance against four most commonly prescribed antibiotics. 1. The bacterial counts on one hundred paper notes sample were ranging between 6～10918/cm cm-2,the median was 141/ cm-2, according to the data it was much higher than credit cards and Australian notes which were made of polymer. The bacterial counts on sixty coin samples were ranging between 2～380/cm-2, much less than paper notes. 2. Coliform (65.6%), E. coli (45.9%), S. aureus (41.7%), B. cereus (67.7%), Salmonella (19.8%) were isolated on one hundred paper notes. Coliform (22.4%), E. coli (5.2%), S. aureus (24.1%), B. cereus (34.5%), Salmonella (10.3%) were isolated from sixty coin samples. These results suggested a high rate of potential pathogens contamination of paper notes than coins. 3. Antibiotic resistances are commonly in most of the pathogens isolated on currency. Ampicillin resistance was found in 60%of Staphylococcus aureus isolated on currency, as well as 76.6% of E. coil and 40% of Salmonella. Erythromycin resistance was detected in 56.6% of S. aureus and in 80.0% of E. coli. All the pathogens isolated were sensitive to Norfloxacin, Salmonella and S. aureus also sensitive to Cefaclor. In this paper, we also studied the antimicrobial capability of metal coins, coins collected from different countries were tested for the ability to inhibit the growth of E. sakazakii, S. aureus, E. coli, L. monocytogenes and S. typhimurium. 1) E. sakazakii appeared very sensitive to metal coins, the second is S. aureus, but E. coli, L. monocytogenes and S. typhimurium are more resistant to these metal coin samples. 2) Coins made of Nickel-brass alloy and Copper-nickel alloy showed a better effect in anti-microbe than other metal coins, especially the ability to inhibited the growth of E. sakazakii and S. aureus, all the inhibition zones produced on nutrient agar are more than 20.6 mm. Aluminium-bronze alloy revealed weak anti-microbe activity to S. aureus and no effect to kill other pathogens. Coins made of stainless steel also can’t resist bacteria growth. 3) Surprisingly, one cent coins of USA which were made of 97.5% Zinc and 2.5% Cu showed a significant antimicrobial capability, the average inhibition zone of these five pathogens is 45.5 mm.

Keywords: antibiotic sensitivity, bacteria, currency, coins, parasites

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Authors: M. Cortés, E. Vera, M. Avella

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Corrosion is an imminent process in nature, which affects all types of materials. In sewerage systems, the corrosion process caused by microorganisms, also known as biogenic sulfuric acid attack, has been studied. This affects the structural integrity of the concrete drainage pipes and the sewage treatment plants. This article is a review of research which focuses on the study of how to reduce the production of hydrogen sulfide, how to improve the resistance of concrete through the use of additives and the implementation of antimicrobial techniques to reduce bacterial growth.

Keywords: bactericides, biogenic sulfuric acid, corrosion, concrete, hydrogen sulphide, nano materials, zeolites

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Authors: Saifalarab A. Mohmmed, Morgana E. Vianna, Jonathan C. Knowles

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The issue of apical periodontitis has received considerable critical attention. Bacteria is integrated into communities, attached to surfaces and consequently form biofilm. The biofilm structure provides bacteria with a series protection skills against, antimicrobial agents and enhances pathogenicity (e.g. apical periodontitis). Sodium hypochlorite (NaOCl) has become the irrigant of choice for elimination of bacteria from the root canal system based on its antimicrobial findings. The aim of the study was to investigate the effect of different agitation techniques on the efficacy of 2.5% NaOCl to eliminate the biofilm from the surface of the lateral canal using the residual biofilm, and removal rate of biofilm as outcome measures. The effect of canal complexity (lateral canal) on the efficacy of the irrigation procedure was also assessed. Forty root canal models (n = 10 per group) were manufactured using 3D printing and resin materials. Each model consisted of two halves of an 18 mm length root canal with apical size 30 and taper 0.06, and a lateral canal of 3 mm length, 0.3 mm diameter located at 3 mm from the apical terminus. E. faecalis biofilms were grown on the apical 3 mm and lateral canal of the models for 10 days in Brain Heart Infusion broth. Biofilms were stained using crystal violet for visualisation. The model halves were reassembled, attached to an apparatus and tested under a fluorescence microscope. Syringe and needle irrigation protocol was performed using 9 mL of 2.5% NaOCl irrigant for 60 seconds. The irrigant was either left stagnant in the canal or activated for 30 seconds using manual (gutta-percha), sonic and ultrasonic methods. Images were then captured every second using an external camera. The percentages of residual biofilm were measured using image analysis software. The data were analysed using generalised linear mixed models. The greatest removal was associated with the ultrasonic group (66.76%) followed by sonic (45.49%), manual (43.97%), and passive irrigation group (control) (38.67%) respectively. No marked reduction in the efficiency of NaOCl to remove biofilm was found between the simple and complex anatomy models (p = 0.098). The removal efficacy of NaOCl on the biofilm was limited to the 1 mm level of the lateral canal. The agitation of NaOCl results in better penetration of the irrigant into the lateral canals. Ultrasonic agitation of NaOCl improved the removal of bacterial biofilm.

Keywords: 3D printing, biofilm, root canal irrigation, sodium hypochlorite

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Authors: M. Yakoubi, N. Belboukhari, A. Cheriti, K. Sekoum

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Launaea nudicaulis (L.) Hook.f. is a desert, spontaneous plant and endemic to northem Sahara, which belongs to the Asteraceae family. This species exists in the region of Bechar (Local name; El-Rghamma). In our knowledge, no work has been founded, except studies showing the antimicrobial and antifungal activity of methalonic extract of this plant. The present paper describes the chemical composition of the essential oil from Launaea nudicaulis and qualification of isolation and identification of some pure products by column chromatography. The essential oil from the aerial parts of Launaea nudicaulis (Asteraceae) was obtained by hydroditillation in 0.4% yield, led to isolation of four several new products. The isolation is made by column chromatography and followed by GC-IK and GC-MS analysis.

Keywords: Launaea nudicaulis, asteraceae, essential oil, column chromatography, GC-FID, GC-MS

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Authors: Muhammad Zeeshan, Rabia Nazir, Lubna Tahir

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Low cost filter based on iron doped zeolite (Fe-Z) and pottery clay was developed for an effective and efficient treatment of the drinking water contaminated with microbes. Fe-Z was characterized using powder XRD, SEM and EDX and shown to have average particle size of 49 nm with spongy appearance. The simulated samples of water self-contaminated with six microbes (S. typhi, B. subtilus, E. coli, S. aures, K. pneumoniae, and P. aeruginosa) after treatment with Fe-Z indicated effective removal of all the microbes in less than 30 min. Equally good results were obtained when actual drinking water samples, totally unfit for human consumption, were treated with Fe-Z.

Keywords: iron doped zeolite, biological and chemical treatment, drinking water

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Authors: Nima Samie, Batoul Sadat Haerian, Sekaran Muniandy, M. S. Kanthimathi

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Colon and breast cancers are categorized as the most prevalent types of cancer worldwide. Recently, the broad clinical application of metal complex compounds has led to the discovery of potential therapeutic drugs. The aim of this study was to evaluate the cytotoxic action of a selected nickel complex compound (NCC) against human colon and breast cancer cells. In this context, we determined the potency of the compound in the induction of apoptosis, cell cycle arrest, and cytoskeleton rearrangement. HT-29, WiDr, CCD-18Co, MCF-7 and Hs 190.T cell lines were used to determine the IC50 of the compound using the MTT assay. Analysis of apoptosis was carried out using immunofluorescence, acridine orange/ propidium iodide double staining, Annexin-V-FITC assay, evaluation of the translocation of NF-kB, oxygen radical antioxidant capacity, quenching of reactive oxygen species content , measurement of LDH release, caspase-3/-7, -8 and -9 assays and western blotting. The cell cycle arrest was examined using flowcytometry and gene expression was assessed using qPCR array. Results showed that our nickel complex compound displayed a potent suppressive effect on HT-29, WiDr, MCF-7 and Hs 190.T after 24 h of treatment with IC50 value of 2.02±0.54, 2.13±0.65, 3.76±015 and 3.14±0.45 µM respectively. This cytotoxic effect on normal cells was insignificant. Dipping in the mitochondrial membrane potential and increased release of cytochrome c from the mitochondria indicated induction of the intrinsic apoptosis pathway by the nickel complex compound. Activation of this pathway was further evidenced by significant activation of caspase 9 and 3/7.The nickel complex compound (NCC) was also shown activate the extrinsic pathways of apoptosis by activation of caspase-8 which is linked to the suppression of NF-kB translocation to the nucleus. Cell cycle arrest in the G1 phase and up-regulation of glutathione reductase, based on excessive ROS production were also observed. The results of this study suggest that the nickel complex compound is a potent anti-cancer agent inducing both intrinsic and extrinsic pathways as well as cell cycle arrest in colon and breast cancer cells.

Keywords: nickel complex, apoptosis, cytoskeletal rearrangement, colon cancer, breast cancer

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Authors: Ghebremedhin Tsegay, Weldu Tesfagaber, Yuanmao Zhu, Xijun He, Wan Wang, Zhenjiang Zhang, Encheng Sun, Jinya Zhang, Yuntao Guan, Fang Li, Renqiang Liu, Zhigao Bu, Dongming Zhao*

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African swine fever (ASF) is a highly infectious viral disease of pigs, resulting in significant economic loss worldwide. As there is no approved vaccines and treatments, the control of ASF entirely depends on early diagnosis and culling of infected pigs. Thus, highly specific and sensitive diagnostic assays are required for accurate and early diagnosis of ASF virus (ASFV). Currently, only a few recombinant proteins have been tested and validated for use as reagents in ASF diagnostic assays. The most promising ones for ASFV antibody detection were p72, p30, p54, and pp62. So far, three ELISA kits based on these recombinant proteins have been commercialized. Due to the complex nature of the virus and variety forms of the disease, robust serodiagnostic assays are still required. ASFV p22 protein, encoded by KP177R gene, is located in the inner membrane of viral particle and appeared transiently in the plasma membrane early after virus infection. The p22 protein interacts with numerous cellular proteins, involved in processes of phagocytosis and endocytosis through different cellular pathways. However, p22 does not seem to be involved in virus replication or swine pathogenicity. In this study, E.coli expressed recombinant p22 protein was used to generate a monoclonal antibody (mAb), and its potential use for the development of blocking ELISA (bELISA) was evaluated. A total of 806 pig serum samples were tested to evaluate the bELISA. Acording the ROC (Reciever operating chracteristic) analysis, 100% sensitivity and 98.10% of specificity was recorded when the PI cut-off value was set at 47%. The novel assay was able to detect the antibodies as early as 9 days post infection. Finaly, a highly sensitive, specific and rapid novel p22-mAb based bELISA assay was developed, and optimized for detection of antibodies against genotype I and II ASFVs. It is a promising candidate for an early and acurate detection of the antibodies and is highly expected to have a valuable role in the containment and prevention of ASF.

Keywords: ASFV, blocking ELISA, diagnosis, monoclonal antibodies, sensitivity, specificity

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Authors: Recep Kesli, Huseyin Bilgin, Yasar Unlu, Gokhan Gungor

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Aim: The aim of this study, was to investigate the presence of Helicobacter pylori (H. pylori) infection by culture, histology, and RUT (Rapid Urease Test) in gastric antrum biopsy samples taken from patients presented with dyspeptic complaints and to determine resistance rates of amoxicillin, clarithromycin, levofloxacin and metronidazole against the H. pylori strains by E-test. Material and Methods: A total of 278 patients who admitted to Konya Education and Research Hospital Department of Gastroenterology with dyspeptic complaints, between January 2011-July 2013, were included in the study. Microbiological and histopathological examinations of biopsy specimens taken from antrum and corpus regions were performed. The presence of H. pylori in biopsy samples was investigated by culture (Portagerm pylori-PORT PYL, Pylori agar-PYL, GENbox microaer, bioMerieux, France), histology (Giemsa, Hematoxylin and Eosin staining), and RUT(CLOtest, Cimberly-Clark, USA). Antimicrobial resistance of isolates against amoxicillin, clarithromycin, levofloxacin, and metronidazole was determined by E-test method (bioMerieux, France). As a gold standard in the diagnosis of H. pylori; it was accepted that the culture method alone was positive or both histology and RUT were positive together. Sensitivity and specificity for histology and RUT were calculated by taking the culture as a gold standard. Sensitivity and specificity for culture were also calculated by taking the co-positivity of both histology and RUT as a gold standard. Results: H. pylori was detected in 140 of 278 of patients with culture and 174 of 278 of patients with histology in the study. H. pylori positivity was also found in 191 patients with RUT. According to the gold standard criteria, a false negative result was found in 39 cases by culture method, 17 cases by histology, and 8 cases by RUT. Sensitivity and specificity of the culture, histology, and RUT methods of the patients were 76.5 % and 88.3 %, 87.8 % and 63 %, 94.2 % and 57.2 %, respectively. Antibiotic resistance was investigated by E-test in 140 H. pylori strains isolated from culture. The resistance rates of H. pylori strains to the amoxicillin, clarithromycin, levofloxacin, and metronidazole was detected as 9 (6.4 %), 22 (15.7 %), 17 (12.1 %), 57 (40.7 %), respectively. Conclusion: In our study, RUT was found to be the most sensitive, culture was the most specific test between culture, histology, and RUT methods. Although we detected the specificity of the culture method as high, its sensitivity was found to be quite low compared to other methods. The low sensitivity of H. pylori culture may be caused by the factors affect the chances of direct isolation such as spoild bacterium, difficult-to-breed microorganism, clinical sample retrieval, and transport conditions.

Keywords: antimicrobial resistance, culture, histology, H. pylori, RUT

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Authors: Hawraa Zbeeb, Hala Khalifeh, Mohamad Khalil, Francesca Storace, Francesca Baldini, Giulio Lupidi, Laura Vergani

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Sarcopoterium spinosum, a widely distributed spiny shrub belonging to the Rosaceae family, is rich in essential and beneficial constituents. In fact, S. spinosum fruits and roots are traditionally used as herbal medicine in the eastern Mediterranean landscape, and this shrub is mentioned as a medicinal plant in a large number of ethnobotanical surveys. Aqueous root extracts from S. spinosum are used by traditional medicinal practitioners for weight loss treatment of diabetes and pain. Moreover, the anti-diabetic activity of S. spinosum root extract has been reported in different studies, but the beneficial effects of aerial parts, especially fruits, have not been elucidated yet. The aim of the present study was to investigate the in vitro antioxidant and lipid-lowering properties of an ethanolic extract from S. spinosum fruits using both hepatic (FaO) and endothelial (HECV) cells in an attempt to evaluate its possible employment as a nutraceutical supplement. First of all, in vitro spectrophotometric assays were employed to characterize the extract. The total phenol content (TPC) was evaluated by Folin–Ciocalteu spectrophotometric method and the radical scavenging activity was tested by 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 2, 2'-azino-bis-3-ethylbenzothiazoline-6-sulfonic acid (ABTS) assays. After that, the beneficial effects of the extract were tested on cells. FaO cells treated for 3 hours with 0.75 mM oleate/palmitate mix (1:2 molar ratio) mimic in vitro a moderate hepato-steatosis. HECV cells exposed for 1 hour to 100 µM H₂O₂ mimic an oxidative insult leading to oxidative stress conditions. After the metabolic and oxidative insult, both cell lines were treated with increasing concentrations of the S. spinosum extract (1, 10, 25 µg/mL) for 24 hours. The results showed the S. spinosum ethanolic extract is rather rich in phenols (TPC of 18.6 mgGAE/g dry extracts). Moreover, the extract showed a good scavenging ability in vitro (IC₅₀ 15.9 µg/ml and 10.9 µg/ml measured by DPPH and ABTS assays, respectively). When the extract was tested on cells, the results showed that it could ameliorate some markers of cell dysfunction. The three concentrations of the extract led to a significant decrease in the intracellular triglyceride (TG) content in steatotic FaO cells measured by spectrophotometric assay. On the other hand, HECV cells treated with increasing concentrations of the extract did not result in a significant decrease in both lipid peroxidation measured by the Thiobarbituric Acid Reactive Substances (TBARS) assay, and in reactive oxygen species (ROS) production measured by fluorometric analysis after DCF staining. Interestingly, the ethanolic extract was able to accelerate the wound repair of confluent HECV cells with respect to H₂O₂-insulted cells as measured by T-scratch assay. Taken together, these results seem to indicate that the ethanol extract from S. spinosum fruits is rich in phenol compounds and plays considerable lipid-lowering activity in vitro on steatotic hepatocytes and accelerates wound healing repair on endothelial cells. In light of that, the ethanolic extract from S. spinosum fruits could be a potential candidate for nutraceutical applications.

Keywords: antioxidant activity, ethanolic extract, lipid-lowering activity, phenolic compounds, Sarcopoterium spinosum fruits

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Authors: Mansi Patel, Priti Mehta

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Background: Ionizing radiations have detrimental effects on humans, and the growing technological encroachment has increased human exposure to it enormously. So, the safety issues have emphasized researchers to develop radioprotector from natural resources having minimal toxicity. A substance having anti-inflammatory, antioxidant, and immunomodulatory activity can be a potential candidate for radioprotection. One such plant with immense potential i.e. Bamboo was selected for the present study. Purpose: The study aims to evaluate the potential of Indian bamboo leaves for protection against the clastogenic effect of gamma radiation. Methods: The protective effect of bamboo leaf extract against gamma radiation-induced genetic damage in human peripheral blood lymphocytes (HPBLs) was evaluated in vitro using Cytokinesis blocked micronuclei assay (CBMN). The blood samples were pretreated with varying concentration of extract 30 min before the radiation exposure (4Gy & 6Gy). The reduction in the frequency of micronuclei was observed for the irradiated and control groups. The effect of various concentration of bamboo leaf extract (400,600,800 mg/kg) on the development of radiation induced sickness and altered mortality in mice exposed to 8 Gy of whole-body gamma radiation was studied. The developed symptoms were clinically scored by multiple endpoints for 30 days. Results: Treatment of HPBLs with varying concentration of extract before exposure to a different dose of γ- radiation resulted in significant (P < 0.0001) decline of radiation induced micronuclei. It showed dose dependent and concentration driven activity. The maximum protection ~ 70% was achieved at nine µg/ml concentration. Extract treated whole body irradiated mice showed 50%, 83.3% and 100% survival for 400, 600, and 800mg/kg with 1.05, 0.43 and 0 clinical score respectively when compared to Irradiated mice having 6.03 clinical score and 0% survival. Conclusion: Our findings indicate bamboo leaf extract reduced the radiation induced cytogenetic damage. It has also increased the survival ratio and reduced the radiation induced sickness and mortality when exposed to a lethal dose of gamma radiation.

Keywords: bamboo leaf extract, Cytokinesis blocked micronuclei (CBMN) assay, ionizing radiation, radio protector

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Authors: Manuela Villani, Anila Xhahysa, Christopher Brooks, Marco Pagani

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The geological structure of Western Balkans is strongly affected by the collision between Adria microplate and the southwestern Euroasia margin, resulting in a considerably active seismic region. The Harmonization of Seismic Hazard Maps in the Western Balkan Countries Project (BSHAP) (2007-2011, 2012-2015) by NATO supported the preparation of new seismic hazard maps of the Western Balkan, but when inspecting the seismic hazard models produced later by these countries on a national scale, significant differences in design PGA values are observed in the border, for instance, North Albania-Montenegro, South Albania- Greece, etc. Considering the fact that the catalogues were unified and seismic sources were defined within BSHAP framework, obviously, the differences arise from the Ground Motion Prediction Equations selection, which are generally the component with highest impact on the seismic hazard assessment. At the time of the project, a modest database was present, namely 672 three-component records, whereas nowadays, this strong motion database has increased considerably up to 20,939 records with Mw ranging in the interval 3.7-7 and epicentral distance distribution from 0.47km to 490km. Statistical analysis of the strong motion database showed the lack of recordings in the moderate-to-large magnitude and short distance ranges; therefore, there is need to re-evaluate the Ground Motion Prediction Equation in light of the recently updated database and the new generations of GMMs. In some cases, it was observed that some events were more extensively documented in one database than the other, like the 1979 Montenegro earthquake, with a considerably larger number of records in the BSHAP Analogue SM database when compared to ESM23. Therefore, the strong motion flat-file provided from the Harmonization of Seismic Hazard Maps in the Western Balkan Countries Project was merged with the ESM23 database for the polygon studied in this project. After performing the preliminary residual analysis, the candidate GMPE-s were identified. This process was done using the GMPE performance metrics available within the SMT in the OpenQuake Platform. The Likelihood Model and Euclidean Distance Based Ranking (EDR) were used. Finally, for this study, a GMPE logic tree was selected and following the selection of candidate GMPEs, model weights were assigned using the average sample log-likelihood approach of Scherbaum.

Keywords: residual analysis, GMPE, western balkan, strong motion, openquake

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Authors: Sandeep K. Shukla, Roli Jain, Soumitra S. Pande, Archna Pandey

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Sulfur nanoparticles were prepared by different methods with different sizes and shapes. When the sulfur is present as nanoparticles they have many practical applications in our life. This research discusses sulfur nanoparticles synthesis, characterizations and applications. With dandruff being a common everyday problem and the market is loaded with antidandruff shampoos and such skin care products, it is obvious to assume resourceful research into this area would be both objective to present scenario and potentially lucrative. Nanoparticles are frequently in use in some very powerful antimicrobial, antifungal cosmetics nowadays, especially silver. To check its antidandruff activity, experiments have been conducted on Malassezia furfur the causal organism for seborrheaic dermatitis or dandruff, which have been cultured for such study in our lab.

Keywords: CTAB surfactant SEM, sulfur nanoparticles (S-NPs), XRD, polymeric surfactant

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Authors: Ibrahim Isah Lakan, Nasiru Ibrahim

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These Phytochemical screening, Antimicrobial activities and mineral Determination of aqueous extract of Stachtarpheta indica were assessed. The result reveals the presence of flavonoids, tannins, saponins, alkaloids, glycosides and anthraquinones. The disc diffusion of aqueous extract showed Escherichia coli, 13 and antibiotic, 19 mm; Bacillus subtilis, 10 and anti –biotic, 17 mm; Klebsiller pnemuoniae , 14 and antibiotic, 24mm and Pseudmonas aeruginosa, 24 and antibiotic, 36 mm which are all comparable with the standard antibiotic cyprotomycin. The mineral content determination by flame photometer revealed that 1.25 (Na+), 0.85 (K +), 1.75 (Ca 2+) % which is a clear indication of the safety of the extract for the hypertensive patients and could be used to lower blood pressure.

Keywords: microbials, mineral, phytochemicals, stachtarpheta indica extracts

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Authors: Ahmed F. Ghanem, Marwa I. Wahba, Asmaa N. El-Dein, Mohamed A. EL-Raey, Ghada E. A. Awad

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Numerous attempts are being performed in order to formulate suitable packaging materials for the meat products. However, to the best of our knowledge, the incorporation of the free hibiscus extract or its microcapsules in the pure polyvinyl alcohol (PVA) matrix as packaging materials for the meats is seldom reported. Therefore, this study aims at the protection of the aqueous crude extract of the hibiscus flowers utilizing the spry drying encapsulation technique. Results of the Fourier transform infrared (FTIR), the scanning electron microscope (SEM), and the particle size analyzer confirmed the successful formation of the assembled capsules via strong interactions, the spherical rough microparticles, and the particle size of ~ 235 nm, respectively. Also, the obtained microcapsules enjoy higher thermal stability than the free extract. Then, the obtained spray-dried particles were incorporated into the casting solution of the pure PVA film with a concentration of 10 wt. %. The segregated free-standing composite films were investigated, compared to the neat matrix, with several characterization techniques such as FTIR, SEM, thermal gravimetric analysis (TGA), mechanical tester, contact angle, water vapor permeability, and oxygen transmission. The results demonstrated variations in the physicochemical properties of the PVA film after the inclusion of the free and the extract microcapsules. Moreover, biological studies emphasized the biocidal potential of the hybrid films against the microorganisms contaminating the meat. Specifically, the microcapsules imparted not only antimicrobial but also antioxidant activities to the PVA matrix. Application of the prepared films on the real meat samples displayed a low bacterial growth with a slight increase in the pH over the storage time which continued up to 10 days at 4 oC, as further evidence to the meat safety. Moreover, the colors of the films did not significantly changed except after 21 days indicating the spoilage of the meat samples. No doubt, the dual-functional of the prepared composite films pave the way towards combined active and smart food packaging applications. This would play a vital role in the food hygiene, including also the quality control and the assurance.

Keywords: PVA, hibiscus, extraction, encapsulation, active packaging, smart and intelligent packaging, meat spoilage

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Authors: Agnieszka E. Laudy, Karolina Stępien, Sergiusz Lulinski, Krzysztof Durka, Stefan Tyski

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1,3-dihydro-1-hydroxy-2,1-benzoxaborole and its derivatives are a particularly interesting group of synthetic agents and were successfully employed in supramolecular chemistry medicine. The first important compounds, 5-fluoro-1,3-dihydro-1-hydroxy-2,1-benzoxaborole and 5-chloro-1,3-dihydro-1-hydroxy-2,1-benzoxaborole were identified as potent antifungal agents. In contrast, (S)-3-(aminomethyl)-7-(3-hydroxypropoxy)-1-hydroxy-1,3-dihydro-2,1-benzoxaborole hydrochloride is in the second phase of clinical trials as a drug for the treatment of Gram-negative bacterial infections of the Enterobacteriaceae family and Pseudomonas aeruginosa. Equally important and difficult task is to search for compounds active against Gram-negative bacilli, which have multi-drug-resistance efflux pumps actively removing many of the antibiotics from bacterial cells. We have examined whether halogen-substituted benzoxaborole-based derivatives and their analogues possess antibacterial activity and are substrates for multi-drug-resistance efflux pumps. The antibacterial activity of 1,3-dihydro-3-hydroxy-1,1-dimethyl-1,2,3-benzosiloxaborole and 10 halogen-substituted its derivatives, as well as 1,2-phenylenediboronic acid and 3 synthesised fluoro-substituted its analogs, were evaluated. The activity against the reference strains of Gram-positive (n=5) and Gram-negative bacteria (n=10) was screened by the disc-diffusion test (0.4 mg of tested compounds was applied onto paper disc). The minimal inhibitory concentration values and the minimal bactericidal concentration values were estimated according to The Clinical and Laboratory Standards Institute and The European Committee on Antimicrobial Susceptibility Testing recommendations. During the minimal inhibitory concentration values determination with or without phenylalanine-arginine beta-naphthylamide (50 mg/L) efflux pump inhibitor, the concentrations of tested compounds ranged 0.39-400 mg/L in the broth medium supplemented with 1 mM magnesium sulfate. Generally, the studied benzosiloxaboroles and benzoxadiboroles showed a higher activity against Gram-positive cocci than against Gram-negative rods. Moreover, benzosiloxaboroles have the higher activity than benzoxadiboroles compounds. In this study, we demonstrated that substitution (mono-, di- or tetra-) of 1,3-dihydro-3-hydroxy-1,1-dimethyl-1,2,3-benzosiloxaborole with halogen groups resulted in an increase in antimicrobial activity as compared to the parent substance. Interestingly, the 6,7-dichloro-substituted parent substance was found to be the most potent against Gram-positive cocci: Staphylococcus sp. (minimal inhibitory concentration 6.25 mg/L) and Enterococcus sp. (minimal inhibitory concentration 25 mg/L). On the other hand, mono- and dichloro-substituted compounds were the most actively removed by efflux pumps present in Gram-negative bacteria mainly from Enterobacteriaceae family. In the presence of efflux pump inhibitor the minimal inhibitory concentration values of chloro-substituted benzosiloxaboroles decreased from 400 mg/L to 3.12 mg/L. Of note, the highest increase in bacterial susceptibility to tested compounds in the presence of phenylalanine-arginine beta-naphthylamide was observed for 6-chloro-, 6,7-dichloro- and 6,7-difluoro-substituted benzosiloxaboroles. In the case of Escherichia coli, Enterobacter cloacae and P. aeruginosa strains at least a 32-fold decrease in the minimal inhibitory concentration values of these agents were observed. These data demonstrate structure-activity relationships of the tested derivatives and highlight the need for further search for benzoxaboroles and related compounds with significant antimicrobial properties. Moreover, the influence of phenylalanine-arginine beta-naphthylamide on the susceptibility of Gram-negative rods to studied benzosiloxaboroles indicate that some tested agents are substrates for efflux pumps in Gram-negative rods.

Keywords: antibacterial activity, benzosiloxaboroles, efflux pumps, phenylalanine-arginine beta-naphthylamide

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Authors: Haba Hamada, Lavaud Cathrine, Benkhaled Mohammed

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The Silene genus is the most representative of the caryophyllaceae family for their rich content in secondary metabolites; saponins, flavonoids and flavonoids glycosides, phytoecdysones, oligosaccharides have been isolated and identified. The Silene genus represented by about 700 species in the temrerate region of the word, the main concentration of spcies is Europe, Asia and North Africa. Three known compounds 1-3 were isolated from the aerial parts of Silene arenarioides Desf. by using different chromatographic methods. The structures of the isolated compounds were determined as stigmasterolglycoside, Soyacerebroside, maltol glycoside. The structures of the isolated compounds were determined by using the NMR (1H-NMR, 13C-NMR, COSY, HSQC, and HMBC) techniques and mass spectroscopy. The antimicrobial and antioxydant activities of the different extracts and compound have been reported.

Keywords: caryophyllaceae, flavonoids, saponosids, flavonoids glycosides

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Authors: M. Vijaya Bhaskar Reddy

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The air-dried and powdered methanol solvent extraction of the rhizomes of Alpinia galangal is subjected to bio-assay guided fractionation and isolation yielded a known compound namely, 1'-S-1'-Acetoxychavicol acetate (1). The isolated known compound has been identified based on the physical, spectral data (IR, ¹H, ¹³C, NMR and mass spectroscopy) and comparison with an authentic sample. Finally isolated 1'-S-1'-Acetoxychavicol acetate (1) was confirmed by synthesis. The crude methanol extract and identified known compound (1) were tested for antidandruff property against Malassezia furfur showed with MIC 1000 µg/mL and 7.81 µg/mL, respectively.

Keywords: Alpinia galanga, isolation, 1'-S-1'-Acetoxychavicol acetate, antidandruff activity, Malassezia furfur

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Authors: Rabah Iratni, Husain El Hasasna, Khawlah Athamneh, Halima Al Sameri, Nehla Benhalilou, Asma Al Rashedi

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Background: Cancer therapies have witnessed great advances in the recent past, however, cancer continues to be a leading cause of death, with colorectal cancer being the fourth cause of cancer-related deaths. Colorectal cancer affects both sexes equally with poor survival rate once it metastasizes. Phytochemicals, which are plant derived compounds, have been on a steady rise as anti-cancer drugs due to the accumulation of evidences that support their potential. Here, we investigated the anticancer effect of Rhus coriaria on colon cancer cells. Material and Method: Human colon cancer HT-29 cell line was used. Protein expression and protein phosphorylation were examined using Western blotting. Transcription activity was measure using Quantitative RT-PCR. Human tumoral clonogenic assay was used to assess cell survival. Senescence was assessed by the senescence-associated beta-galactosidase assay. Results: Rhus coriaria extract (RCE) was found to significantly inhibit the viability and colony growth of human HT-29 colon cancer cells. RCE induced senescence and cell cycle arrest at G1 phase. These changes were concomitant with upregulation of p21, p16, downregulation of cyclin D1, p27, c-myc and expression of Senescence-associated-β-Galactosidase activity. Moreover, RCE induced non-canonical beclin-1independent autophagy and subsequent apoptotic cell death through activation of activation caspase 8 and caspase 7. The blocking of autophagy by 3-methyladenine (3-MA) or chloroquine (CQ) reduced RCE-induced cell death. Further, RCE induced DNA damage, reduced mutant p53 protein level and downregulated phospho-AKT and phospho-mTOR, events that preceded autophagy. Mechanistically, we found that RCE inhibited the AKT and mTOR pathway, a regulator of autophagy, by promoting the proteasome-dependent degradation of both AKT and mTOR proteins. Conclusion: Our findings provide strong evidence that Rhus coriaria possesses strong anti-colon cancer activity through induction of senescence and autophagic cell death, making it a promising alternative or adjunct therapeutic candidate against colon cancer.

Keywords: autophagy, proteasome degradation, senescence, mTOR, apoptosis, Beclin-1

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Authors: Kiran Nawaz, Ahmad Ali Shahid, Sehrish Iftikhar, Waheed Anwar, Muhammad Nasir Subhani

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Chili (Solanum annum L.) attacks by many fungal pathogens, including members of Oomycetes which are responsible for root rot in different chili growing areas of the world. Oomycetes pathogens cause economic losses in different regions of the Pakistan. Most of the plant tissues, including roots, crowns, fruit, and leaves, are vulnerable to Phytophthora capsici. It is very difficult to manage the Phytophthora root rot of chili as many commercial varieties are tremendously vulnerable to P. capsici. The causal agent of the disease was isolated on corn meal agar (CMA) and identified on a morphological basis by using available taxonomic keys. The pathogen was also confirmed on the molecular basis through internal transcribed spacer region and with other molecular markers.The Blastn results showed 100% homology with already reported sequences of P. capsici in NCBI database. Most of the farmers have conventionally relied on foliar fungicide applications to control Phytophthora root rot in spite of their incomplete effectiveness. In this study, in vitro plate assay, seed soaking and foliar applications of 6 fungicides were evaluated against root rot of chili. In vitro assay revealed that significant inhibition of linear growth was obtained with Triflumizole at 7.0%, followed by Thiophanate methyl (8.9%), Etridiazole (6.0%), Propamocarb (5.9%) and 7.5% with Mefenoxam and Iprodione for P. capsici. The promising treatments of in vitro plate bioassay were evaluated in pot experiments under controlled conditions in the greenhouse. All fungicides were applied after at 6-day intervals. Results of pot experiment showed that all treatments considerably inhibited the percentage of P. capsici root rot incidence. In addition, application of seed soaking with all six fungicides combined with the foliar spray of the same components showed the significant reduction in root rot incidence. The combine treatments of all fungicides as in vitro bioassay, seed soaking followed by foliar spray is considered non-harmful control methods which have advantages and limitation. Hence, these applications proved effective and harmless for the management of soil-borne plant pathogens.

Keywords: blastn, bioassay, corn meal agar(CMA), oomycetes

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Authors: Kinnsley Travis, Camerron M. Crowder

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Mapk8ip3 (Mitogen-Activated Protein Kinase 8 Interacting Protein 3) is a gene that codes for the JIP3 protein, which is a part of the JIP scaffolding protein family. This protein is involved in axonal vesicle transport, elongation and regeneration. Variants in the Mapk8ip3 gene are associated with a rare-genetic condition that results in a neurodevelopmental disorder that can cause a range of phenotypes including global developmental delay and intellectual disability. Currently, there are 18 known individuals diagnosed to have sequenced confirmed Mapk8ip3 genetic disorders. This project focuses on examining the impact of a subset of missense patient variants on the Jip3 protein function by overexpressing the mRNA of these variants in a zebrafish knockout model for Jip3. Plasmids containing cDNA with individual missense variants were reverse transcribed, purified, and injected into single-cell zebrafish embryos (Wild Type, Jip3 -/+, and Jip3 -/-). At 6-days post mRNA microinjection, morphological, behavioral, and microscopic phenotypes were examined in zebrafish larvae. Morphologically, we compared the size and shape of the zebrafish during their development over a 5-day period. Total locomotive activity was assessed using the Microtracker assay and patterns of movement over time were examined using the DanioVision assay. Lastly, we used confocal microscopy to examine sensory axons for swelling and shortened length, which are phenotypes observed in the loss-of-function knockout Jip3 zebrafish model. Using these assays during embryonic development, we determined the impact of various missense variants on Jip3 protein function, compared to knockout and wild-type zebrafish embryo models. Variants in the gene Mapk8ip3 cause rare-neurodevelopmental disorders due to an essential role in axonal vesicle transport, elongation and regeneration. A subset of missense variants was examined by overexpressing the mRNA of these variants in a Jip3 knock-out zebrafish. Morphological, behavioral, and microscopic phenotypes were examined in zebrafish larvae. Using these assays, the spectrum of disorders can be phenotypically determined and the impact of variant location can be compared to knockout and wild-type zebrafish embryo models.

Keywords: rare disease, neurodevelopmental disorders, mrna overexpression, zebrafish research

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Authors: Seyedahmad Hosseini, Mohammadjavad Sotoudeheian

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Introduction: Allergic asthma is a heterogeneous immuno-inflammatory disease based on Th-2-mediated inflammation. Histopathologic abnormalities of the airways characteristic of asthma include epithelial damage and subepithelial collagen deposition. Objectives: Human bronchial epithelial cell genome expression of TNF‑α, IL‑6, ICAM‑1, VCAM‑1, nuclear factor (NF)‑κB signaling pathways up-regulate during inflammatory cascades. Moreover, immunofluorescence assays confirmed the nuclear translocation of NF‑κB p65 during inflammatory responses. An absolute LDH leakage assays suggestedLPS-inducedcells injury, and the associated mechanisms are co-incident events. LPS-induced phosphorylation of ERKand JNK causes inflammation in epithelial cells through inhibition of ERK and JNK activation and NF-κB signaling pathway. Furthermore, the inhibition of NF-κB mRNA expression and the nuclear translocation of NF-κB lead to anti-inflammatory events. Likewise, activation of SUMF2 which inhibits IL-13 and reduces Th2-cytokines, NF-κB, and IgE levels to ameliorate asthma. On the other hand, TNFα-induced mucus production reduced NF-κB activation through inhibition of the activation status of Rac1 and IκBα phosphorylation. In addition, bradykinin B2 receptor (B2R), which mediates airway remodeling, regulates through NF-κB. Bronchial B2R expression is constitutively elevated in allergic asthma. In addition, certain NF-κB -dependent chemokines function to recruit eosinophils in the airway. Besides, bromodomain containing 4 (BRD4) plays a significant role in mediating innate immune response in human small airway epithelial cells as well as transglutaminase 2 (TG2), which is detectable in saliva. So, the guanine nucleotide-binding regulatory protein α-subunit, Gα16, expresses a κB-driven luciferase reporter. This response was accompanied by phosphorylation of IκBα. Furthermore, expression of Gα16 in saliva markedly enhanced TNF-α-induced κB reporter activity. Methods: The applied method to form NF-κB activation is the electromobility shift assay (EMSA). Also, B2R-BRD4-TG2 complex detection by immunoassay method within saliva with EMSA of NF-κB activation may be a novel biomarker for asthma diagnosis and follow up. Conclusion: This concept introduces NF-κB signaling pathway as potential asthma biomarkers and promising targets for the development of new therapeutic strategies against asthma.

Keywords: NF-κB, asthma, saliva, T-helper

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Authors: Madhusudana S. N., Reeta Mani, Ashwini S. Satishchandra P., Netravati, Udhani V., Fiaz A., Karande S.

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Rabies is an acute encephalitis which is considered 100% fatal despite occasional reports of survivors. However, in recent times more cases of human rabies survivors are being reported. In the last 5 years, there are six laboratories confirmed human rabies survivors in India alone. All cases were children below 15 years and all contracted the disease by dog bites. All of them also had received the full or partial course of rabies vaccination and 4 out of 6 had also received rabies immunoglobulin. All cases were treated in intensive care units in hospitals at Bangalore, Mumbai, Chandigarh, Lucknow and Goa. We report here the results of immunological and virological studies conducted at our laboratory on these patients. The clinical samples that were obtained from these patients were Serum, CSF, nuchal skin biopsy and saliva. Serum and CSF samples were subjected to standard RFFIT for estimation of rabies neutralizing antibodies. Skin biopsy, CSF and saliva were processed by TaqMan real-time PCR for detection of viral RNA. CSF, saliva and skin homogenates were also processed for virus isolation by inoculation of suckling mice. The PBMCs isolated from fresh blood was subjected to ELISPOT assay to determine the type of immune response (Th1/Th2). Both CSF and serum were also investigated for selected cytokines by Luminex assay. The level of antibodies to virus G protein and N protein were determined by ELISA. All survivors had very high titers of RVNA in serum and CSF 100 fold higher than non-survivors and vaccine controls. A five-fold rise in titer could be demonstrated in 4 out of 6 patients. All survivors had a significant increase in antibodies to G protein in both CSF and serum when compared to non-survivors. There was a profound and robust Th1 response in all survivors indicating that interferon gamma could play an important factor in virus clearance. We could isolate viral RNA in only one patient four years after he had developed symptoms. The partial N gene sequencing revealed 99% homology to species I strain prevalent in India. Levels of selected cytokines in CSF and serum did not reveal any difference between survivors and non-survivors. To conclude, survival from rabies is mediated by virus-specific immune responses of the host and clearance of rabies virus from CNS may involve the participation of both Th2 and Th1 immune responses.

Keywords: rabies, rabies treatment, rabies survivors, immune reponse in rabies encephalitis

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Authors: Tayyaba Bari, Muhammad Hamza Anjum, Samra Kanwal, Fakhera Ikram

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Osteoarthritis, a degenerative condition, affects more than 213 million individuals globally. Since articular cartilage has no or limited vessels, therefore, after deteriorating, it is unable to rejuvenate. Traditional approaches for cartilage repair, like autologous chondrocyte implantation, microfracture and cartilage transplantation are often associated with postoperative complications and lead to further degradation. Decellularized human umbilical cord has gained interest as a viable treatment for cartilage repair. Decellularization removes all cellular contents as well as debris, leaving a biologically active 3D network known as extracellular matrix (ECM). This matrix is biodegradable, non-immunogenic and provides a microenvironment for homeostasis, growth and repair. UC derived bioink function as 3D scaffolding material, not only mediates cell-matrix interactions but also adherence, proliferation and propagation of cells for 3D organoids. This study comprises different physical, chemical and biological approaches to optimize the decellularization of human umbilical cord (UC) tissues followed by the solubilization of these tissues to bioink formation. The decellularization process consisted of two cycles of freeze thaw where the umbilical cord at -20˚C was thawed at room temperature followed by dissection in small sections from 0.5 to 1cm. Similarly decellularization with ionic and non-ionic detergents Sodium dodecyl sulfate (SDS) and Triton-X 100 revealed that both concentrations of SDS i.e 0.1% and 1% were effective in complete removal of cells from the small UC tissues. The results of decellularization was further confirmed by running them on 1% agarose gel. Histological analysis revealed the efficacy of decellularization, which involves paraffin embedded samples of 4μm processed for Hematoxylin-eosin-safran and 4,6-diamidino-2-phenylindole (DAPI). ECM preservation was confirmed by Alcian Blue, and Masson’s trichrome staining on consecutive sections and images were obtained. Sulfated GAG’s content were determined by 1,9-dimethyl-methylene blue (DMMB) assay, similarly collagen quantification was done by hydroxy proline assay. This 3D bioengineered scaffold will provide a typical atmosphere as in the extracellular matrix of the tissue, which would be seeded with the mesenchymal cells to generate the desired 3D ink for in vitro and in vivo cartilage regeneration applications.

Keywords: umbilical cord, 3d printing, bioink, tissue engineering, cartilage regeneration

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Authors: Renata Swislocka, Grzegorz Swiderski, Agata Jablonska-Trypuc, Wlodzimierz Lewandowski

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Forming a complex of a phenolic compound with a metal not only alters the physicochemical properties of the ligand (including increase in stability or changes in lipophilicity), but also its biological activity, including antioxidant, antimicrobial and many others. As part of our previous projects, we examined the physicochemical and antimicrobial properties of phenolic acids and their complexes with metals naturally occurring in foods. Previously we studied the complexes of manganese(II), copper(II), cadmium(II) and alkali metals with ferulic, caffeic and p-coumaric acids. In the framework of this study, the physicochemical and biological properties of cicoric acid, its sodium salt, and complexes with copper and zinc were investigated. Cichoric acid is a derivative of both caffeic acid and tartaric acid. It has first been isolated from Cichorium intybus (chicory) but also it occurs in significant amounts in Echinacea, particularly E. purpurea, dandelion leaves, basil, lemon balm and in aquatic plants, including algae and sea grasses. For the study of spectroscopic and biological properties of cicoric acid, its sodium salt, and complexes with zinc and copper a variety of methods were used. Studies of antioxidant properties were carried out in relation to selected stable radicals (method of reduction of DPPH and reduction of FRAP). As a result, the structure and spectroscopic properties of cicoric acid and its complexes with selected metals in the solid state and in the solutions were defined. The IR and Raman spectra of cicoric acid displayed a number of bands that were derived from vibrations of caffeic and tartaric acids moieties. At 1746 and 1716 cm-1 the bands assigned to the vibrations of the carbonyl group of tartaric acid occurred. In the spectra of metal complexes with cichoric these bands disappeared what indicated that metal ion was coordinated by the carboxylic groups of tartaric acid. In the spectra of the sodium salt, a characteristic wide-band vibrations of carboxylate anion occurred. In the spectra of cicoric acid and its salt and complexes, a number of bands derived from the vibrations of the aromatic ring (caffeic acid) were assigned. Upon metal-ligand attachment, the changes in the values of the wavenumbers of these bands occurred. The impact of metals on the antioxidant properties of cicoric acid was also examined. Cichoric acid has a high antioxidant potential. Complexation by metals (zinc, copper) did not significantly affect its antioxidant capacity. The work was supported by the National Science Centre, Poland (grant no. 2015/17/B/NZ9/03581).

Keywords: chicoric acid, metal complexes, natural antioxidant, phenolic acids

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Authors: Manar Y. Abd El-Aziz, Alyaa E. Morgham, Amira A. El-Fallal, Heba Tolla E. Abo El Naga

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Sport wears almost preferred with knitted fabrics, specially interlock construction. But, there is a need for higher comfortability and functional properties for fabrics to be more fitted to this application. This study compared cotton and polyester microfibers and blended them to improve the functional activity of interlock in sport wear by mercerized and non-mercerized. also, fabric dyeing and dyeability are affected by mercerization. Many functional properties, such as UV protection as well as antimicrobial activity. The changes in different mechanical as well as physical properties were investigated. The washing fastness properties of the dyed fabrics are also given. The changes in moisture regain, loss in weight, and burst and burst elongation for all mercerized fabrics have been studied.

Keywords: interlock, mercirization, dyeability, function properties, moisture management

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Authors: Qiaoyue Kuang, Yang Li, Mizuki Endo, Takeaki Ozawa

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G protein-coupled receptors (GPCR) are the largest family of receptor proteins that detect molecules outside the cell and activate cellular responses. Of the GPCRs, dopamine receptors, which recognize extracellular dopamine, are essential to mammals due to their roles in numerous physiological events, including autonomic movement, hormonal regulation, emotions, and the reward system in the brain. To precisely understand the physiological roles of dopamine receptors, it is important to spatiotemporally control the signaling mediated by dopamine receptors, which is strongly dependent on their surface expression. Conventionally, chemical-induced interactions were applied to trigger the endocytosis of cell surface receptors. However, these methods were subjected to diffusion and therefore lacked temporal and special precision. To further understand the receptor-mediated signaling and to control the plasma membrane expression of receptors, an optogenetic tool called E-fragment was developed. The C-terminus of a light-sensitive photosensory protein cyptochrome2 (CRY2) was attached to β-Arrestin, and the E-fragment was generated by fusing the C-terminal peptide of vasopressin receptor (V2R) to CRY2’s binding partner protein CIB. The CRY2-CIB heterodimerization triggered by blue light stimulation brings β-Arrestin to the vicinity of membrane receptors and results in receptor endocytosis. In this study, the E-fragment system was applied to dopamine receptors 1 and 2 (DRD1 and DRD2) to control dopamine signaling. First, confocal fluorescence microscope observation qualitatively confirmed the light-induced endocytosis of E-fragment fused receptors. Second, NanoBiT bioluminescence assay verified quantitatively that the surface amount of E-fragment labeled receptors decreased after light treatment. Finally, GloSensor bioluminescence assay results suggested that the E-fragment-dependent receptor light-induced endocytosis decreased cAMP production in DRD1 signaling and attenuated the inhibition effect of DRD2 on cAMP production. The developed optogenetic tool was able to induce receptor endocytosis by external light, providing opportunities to further understand numerous physiological activities by controlling receptor-mediated signaling spatiotemporally.

Keywords: dopamine receptors, endocytosis, G protein-coupled receptors, optogenetics

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Authors: Sanjay Shukla

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Excessive use of antibiotics is a main problem in the treatment of wounds and other chronic infections and antibiotic treatment is frequently non-curative, thus alternative treatment is necessary. Phage therapy is considered one of the most effective approaches to treat multi-drug resistant bacterial pathogens. Infections caused by Staphylococcus aureus are very efficiently controlled with phage cocktails, containing a different individual phages lysate infecting a majority of known pathogenic S. aureus strains. The aim of current study was to investigate the efficiency of a purified phage cocktail for prophylactic as well as therapeutic application in mouse model and in large animals with chronic septic infection of wounds. A total of 150 sewage samples were collected from various livestock farms. These samples were subjected for the isolation of bacteriophage by double agar layer method. A total of 27 sewage samples showed plaque formation by producing lytic activity against S. aureus in double agar overlay method out of 150 sewage samples. In TEM recovered isolates of bacteriophages showed hexagonal structure with tail fiber. In the bacteriophage (ØVS) had an icosahedral symmetry with the head size 52.20 nm in diameter and long tail of 109 nm. Head and tail were held together by connector and can be classified as a member of the Myoviridae family under the order of Caudovirale. Recovered bacteriophage had shown the antibacterial activity against the S. aureus in vitro. Cocktail (ØVS1, ØVS5, ØVS9 and ØVS 27) of phage lysate were tested to know in vivo antibacterial activity as well as the safety profile. Result of mice experiment indicated that the bacteriophage lysate was very safe, did not show any appearance of abscess formation which indicates its safety in living system. The mice were also prophylactically protected against S. aureus when administered with cocktail of bacteriophage lysate just before the administration of S. aureus which indicates that they are good prophylactic agent. The S. aureus inoculated mice were completely recovered by bacteriophage administration with 100% recovery which was very good as compere to conventional therapy. In present study ten chronic cases of wound were treated with phage lysate and follow up of these cases was done regularly up to ten days (at 0, 5 and 10 d). Result indicated that the six cases out of ten showed complete recovery of wounds within 10 d. The efficacy of bacteriophage therapy was found to be 60% which was very good as compared to the conventional antibiotic therapy in chronic septic wounds infections. Thus, the application of lytic phage in single dose proved to be innovative and effective therapy for treatment of septic chronic wounds.

Keywords: phage therapy, phage lysate, antimicrobial resistance, S. aureus

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Authors: Melanie Ostermann, Eivind Birkeland, Ying Xue, Alexander Sauter, Mihaela R. Cimpan

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Background: New reliable and robust techniques to assess biological effects of nanomaterials (NMs) in vitro are needed to speed up safety analysis and to identify key physicochemical parameters of NMs, which are responsible for their acute cytotoxicity. The central aim of this study was to validate and evaluate the applicability and reliability of an impedance-based flow cytometry (IFC) technique for the high-throughput screening of NMs. Methods: Eight inorganic NMs from the European Commission Joint Research Centre Repository were used: NM-302 and NM-300k (Ag: 200 nm rods and 16.7 nm spheres, respectively), NM-200 and NM- 203 (SiO₂: 18.3 nm and 24.7 nm amorphous, respectively), NM-100 and NM-101 (TiO₂: 100 nm and 6 nm anatase, respectively), and NM-110 and NM-111 (ZnO: 147 nm and 141 nm, respectively). The aim was to assess the biological effects of these materials on human monoblastoid (U937) cells. Dispersions of NMs were prepared as described in the NANOGENOTOX dispersion protocol and cells were exposed to NMs at relevant concentrations (2, 10, 20, 50, and 100 µg/mL) for 24 hrs. The change in electrical impedance was measured at 0.5, 2, 6, and 12 MHz using the IFC AmphaZ30 (Amphasys AG, Switzerland). A traditional toxicity assay, Trypan Blue Dye Exclusion assay, and dark-field microscopy were used to validate the IFC method. Results: Spherical Ag particles (NM-300K) showed the highest toxic effect on U937 cells followed by ZnO (NM-111 ≥ NM-110) particles. Silica particles were moderate to non-toxic at all used concentrations under these conditions. A higher toxic effect was seen with smaller sized TiO2 particles (NM-101) compared to their larger analogues (NM-100). No interferences between the IFC and the used NMs were seen. Uptake and internalization of NMs were observed after 24 hours exposure, confirming actual NM-cell interactions. Conclusion: Results collected with the IFC demonstrate the applicability of this method for rapid nanotoxicity assessment, which proved to be less prone to nano-related interference issues compared to some traditional toxicity assays. Furthermore, this label-free and novel technique shows good potential for up-scaling in directions of an automated high-throughput screening and for future NM toxicity assessment. This work was supported by the EC FP7 NANoREG (Grant Agreement NMP4-LA-2013-310584), the Research Council of Norway, project NorNANoREG (239199/O70), the EuroNanoMed II 'GEMN' project (246672), and the UH-Nett Vest project.

Keywords: cytotoxicity, high-throughput, impedance, nanomaterials

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Authors: Hanaa M. Abu El Einin, Ahmed T. Sharaf El- Din

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Biomphalaria snails play an integral role in the transmission of Schistosoma mansoni, the causative agent for human schistosomiasis. Two species of Biomphalaria were reported from Egypt, Biomphalaria alexandrina and Biomphalaria glabrata, and later on a hybrid of B. alexandrina and B. glabrata was reported in streams at Nile Delta. All were known to be excellent hosts of S. mansoni. Host-parasite relationship can be viewed in terms of snail susceptibility and parasite infectivity. The objective of this study will highlight the progress that has been made in using molecular approaches to describe the correct identification of snail species that participating in transmission of schistosomiasis, rapid diagnose of infection in addition to susceptibility and resistance type. Snails were identified using of molecular methods involving Randomly Amplified Polymorphic DNA (RAPD), Polymerase Chain Reaction, Restriction Fragment Length Polymorphisms (PCR-RFLP) and Species - specific- PCR. Molecular approaches to diagnose parasite in snails from Egypt: Nested PCR assay and small subunit (SSU) rRNA gene. Also RAPD PCR for study susceptible and resistance phenotype. The results showed that RAPD- PCR, PCR-RFLP and species-specific-PCR techniques were confirmed that: no evidence for the presence of B. glabrata in Egypt, All Biomphalaria snails collected identified as B. alexandrina snail i-e B alexandrinia is a common and no evidence for hybridization with B. glabrata. The adopted specific nested PCR assay revealed much higher sensitivity which enables the detection of S. mansoni infected snails down to 3 days post infection. Nested PCR method for detection of infected snails using S. mansoni fructose -1,6- bisphosphate aldolase (SMALDO) primer, these primers are specific only for S. mansoni and not cross reactive with other schistosomes or molluscan aldolases Nested PCR for such gene is sensitive enough to detect one cercariae. Genetic variations between B. alexandrina strains that are susceptible and resistant to Schistosoma infec¬tion using a RAPD-PCR showed that 39.8% of the examined snails collected from the field were resistant, while 60.2% of these snails showed high infection rates. In conclusion the genetics of the intermediate host plays a more important role in the epidemiological control of schistosomiasis.

Keywords: biomphalaria, molecular differentiation, parasite detection, schistosomiasis

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Authors: Douglas N. Simoes, Michele Pittol, Vanda F. Ribeiro, Daiane Tomacheski, Ruth M. C. Santana

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Thermoplastic elastomers (TPE) compounds are used in a wide range of applications, like home appliances, automotive components, medical devices, footwear, and others. These materials are susceptible to microbial attack, causing a crack in polymer chains compounds based on SEBS copolymers, poly (styrene-b-(ethylene-co-butylene)-b-styrene, are a class of TPE, largely used in domestic appliances like refrigerator seals (gaskets), bath mats and sink squeegee. Moisture present in some areas (such as shower area and sink) in addition to organic matter provides favorable conditions for microbial survival and proliferation, contributing to the spread of diseases besides the reduction of product life cycle due the biodegradation process. Zinc oxide (ZnO) has been studied as an alternative antibacterial additive due its biocidal effect. It is important to know the influence of these additives in the properties of the compounds, both at the beginning and during the life cycle. In that sense, the aim of this study was to evaluate the effect of accelerated aging in oven on antibacterial and mechanical properties of ZnO loaded SEBS based TPE compounds. Two different comercial zinc oxide, named as WR and Pe were used in proportion of 1%. A compound with no antimicrobial additive (standard) was also tested. The compounds were prepared using a co-rotating double screw extruder (L/D ratio of 40/1 and 16 mm screw diameter). The extrusion parameters were kept constant for all materials, screw rotation rate was set at 226 rpm, with a temperature profile from 150 to 190 ºC. Test specimens were prepared using the injection molding machine at 190 ºC. The Standard Test Method for Rubber Property—Effect of Liquids was applied in order to simulate the exposition of TPE samples to detergent ingredients during service. For this purpose, ZnO loaded TPE samples were immersed in a 3.0% w/v detergent (neutral) and accelerated aging in oven at 70°C for 7 days. Compounds were characterized by changes in mechanical (hardness and tension properties) and mass. The Japan Industrial Standard (JIS) Z 2801:2010 was applied to evaluate antibacterial properties against Staphylococcus aureus (S. aureus) and Escherichia coli (E. coli). The microbiological tests showed a reduction up to 42% in E. coli and up to 49% in S. aureus population in non-aged samples. There were observed variations in elongation and hardness values with the addition of zinc The changes in tensile at rupture and mass were not significant between non-aged and aged samples.

Keywords: antimicrobial, domestic appliance, sebs, zinc oxide

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