Search results for: bacterial ring rot

Authors: Alan N. A. Heberle, Salatiel W. Da Silva, Valentin Perez-Herranz, Andrea M. Bernardes

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Contaminants of emerging concern are substances widely used, such as pharmaceutical products. These compounds represent risk for both wild and human life since they are not completely removed from wastewater by conventional wastewater treatment plants. In the environment, they can be harm even in low concentration (µ or ng/L), causing bacterial resistance, endocrine disruption, cancer, among other harmful effects. One of the most common taken medicine to treat cardiocirculatory diseases is the Atenolol (ATL), a β-Blocker, which is toxic to aquatic life. In this way, it is necessary to implement a methodology, which is capable to promote the degradation of the ATL, to avoid the environmental detriment. A very promising technology is the advanced electrochemical oxidation (AEO), which mechanisms are based on the electrogeneration of reactive radicals (mediated oxidation) and/or on the direct substance discharge by electron transfer from contaminant to electrode surface (direct oxidation). The hydroxyl (HO•) and sulfate (SO₄•⁻) radicals can be generated, depending on the reactional medium. Besides that, at some condition, the peroxydisulfate (S₂O₈²⁻) ion is also generated from the SO₄• reaction in pairs. Both radicals, ion, and the direct contaminant discharge can break down the molecule, resulting in the degradation and/or mineralization. However, ATL molecule and byproducts can still remain in the treated solution. On this wise, some efforts can be done to implement the AEO process, being one of them the use of a cationic membrane to separate the cathodic (reduction) from the anodic (oxidation) reactor compartment. The aim of this study is investigate the influence of the implementation of a cationic membrane (Nafion®-117) to separate both cathodic and anodic, AEO reactor compartments. The studied reactor was a filter-press, with bath recirculation mode, flow 60 L/h. The anode was an Nb/BDD2500 and the cathode a stainless steel, both bidimensional, geometric surface area 100 cm². The solution feeding the anodic compartment was prepared with ATL 100 mg/L using Na₂SO₄ 4 g/L as support electrolyte. In the cathodic compartment, it was used a solution containing Na₂SO₄ 71 g/L. Between both solutions was placed the membrane. The applied currents densities (iₐₚₚ) of 5, 20 and 40 mA/cm² were studied over 240 minutes treatment time. Besides that, the ATL decay was analyzed by ultraviolet spectroscopy (UV/Vis). The mineralization was determined performing total organic carbon (TOC) in TOC-L CPH Shimadzu. In the cases without membrane, the iₐₚₚ 5, 20 and 40 mA/cm² resulted in 55, 87 and 98 % ATL degradation at the end of treatment time, respectively. However, with membrane, the degradation, for the same iₐₚₚ, was 90, 100 and 100 %, spending 240, 120, 40 min for the maximum degradation, respectively. The mineralization, without membrane, for the same studied iₐₚₚ, was 40, 55 and 72 %, respectively at 240 min, but with membrane, all tested iₐₚₚ reached 80 % of mineralization, differing only in the time spent, 240, 150 and 120 min, for the maximum mineralization, respectively. The membrane increased the ATL oxidation, probably due to avoid oxidant ions (S₂O₈²⁻) reduction on the cathode surface.

Keywords: contaminants of emerging concern, advanced electrochemical oxidation, atenolol, cationic membrane, double compartment reactor

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Authors: Lena Lapidot, Amir Amnon, Rita Nosenko, Veitsman Ella, Cohen-Ezra Oranit, Davidov Yana, Segev Shlomo, Koren Omry, Safran Michal, Ben-Ari Ziv

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Introduction: Hepatocellular carcinoma (HCC) is one of the leading causes of cancer related mortality worldwide. Liver Cirrhosis is the main predisposing risk factor for the development of HCC. The factor(s) influencing disease progression from Cirrhosis to HCC remain unknown. Gut microbiota has recently emerged as a major player in different liver diseases, however its association with HCC is still a mystery. Moreover, there might be an important association between the gut microbiota, nutrition, life style and the progression of Cirrhosis and HCC. The aim of our study was to characterize the gut microbial signature in association with life style and nutrition of patients with Cirrhosis, HCC-Cirrhosis and healthy controls. Design: Stool samples were collected from 95 individuals (30 patients with HCC, 38 patients with Cirrhosis and 27 age, gender and BMI-matched healthy volunteers). All participants answered lifestyle and Food Frequency Questionnaires. 16S rRNA sequencing of fecal DNA was performed (MiSeq Illumina). Results: There was a significant decrease in alpha diversity in patients with Cirrhosis (qvalue=0.033) and in patients with HCC-Cirrhosis (qvalue=0.032) compared to healthy controls. The microbiota of patients with HCC-cirrhosis compared to patients with Cirrhosis, was characterized by a significant overrepresentation of Clostridium (pvalue=0.024) and CF231 (pvalue=0.010) and lower expression of Alphaproteobacteria (pvalue=0.039) and Verrucomicrobia (pvalue=0.036) in several taxonomic levels: Verrucomicrobiae, Verrucomicrobiales, Verrucomicrobiaceae and the genus Akkermansia (pvalue=0.039). Furthermore, we performed an analysis of predicted metabolic pathways (Kegg level 2) that resulted in a significant decrease in the diversity of metabolic pathways in patients with HCC-Cirrhosis (qvalue=0.015) compared to controls, one of which was amino acid metabolism. Furthermore, investigating the life style and nutrition habits of patients with HCC-Cirrhosis, we found significant correlations between intake of artificial sweeteners and Verrucomicrobia (qvalue=0.12), High sugar intake and Synergistetes (qvalue=0.021) and High BMI and the pathogen Campylobacter (qvalue=0.066). Furthermore, overweight in patients with HCC-Cirrhosis modified bacterial diversity (qvalue=0.023) and composition (qvalue=0.033). Conclusions: To the best of the our knowledge, we present the first report of the gut microbial composition in patients with HCC-Cirrhosis, compared with Cirrhotic patients and healthy controls. We have demonstrated in our study that there are significant differences in the gut microbiome of patients with HCC-cirrhosis compared to Cirrhotic patients and healthy controls. Our findings are even more pronounced because the significantly increased bacteria Clostridium and CF231 in HCC-Cirrhosis weren't influenced by diet and lifestyle, implying this change is due to the development of HCC. Further studies are needed to confirm these findings and assess causality.

Keywords: Cirrhosis, Hepatocellular carcinoma, life style, liver disease, microbiome, nutrition

Procedia PDF Downloads 127

Authors: Sima E. Rugarabamu, Mecky I. Matee, Elison N. M. Simon

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Background: In Tanzania bacteriological studies of etiological agents of oro-facial infections are very limited, and very few have investigated anaerobes. The aim of this study was to determine the spectrum of bacterial agents involved in oral and maxillofacial infections in patients attending Muhimbili National Hospital, Dar-es-salaam, Tanzania. Method: This was a hospital based descriptive cross-sectional study that was conducted in the Department of Oral and Maxillofacial Surgery of the Muhimbili National Hospital in Dar es Salaam, Tanzania from 1st January 2014 to 31st August 2014. Seventy (70) patients with various forms of oral and maxillofacial infections who were recruited for the study. The study participants were interviewed using a prepared questionnaire after getting their consent. Pus aspirate was cultured on Blood agar, Chocolate Agar, MacConkey agar and incubated aerobically at 37°C. Imported blood agar was used for anaerobic culture whereby they were incubated at 37°Cin anaerobic jars in an atmosphere of generated using commercial gas-generating kits in accordance with manufacturer’s instructions. Plates were incubated at 37°C for 24 hours (For aerobic culture and 48 hours for anaerobic cultures). Gram negative rods were identified using API 20E while all other isolates were identified by conventional biochemical tests. Antibiotic sensitivity testing for isolated aerobic and anaerobic bacteria was detected by the disk diffusion, agar dilution and E-test using routine and commercially available antibiotics used to treat oral facial infections. Results: This study comprised of 41 (58.5%) males and 29 (41.5%) females with a mean age of 32 years SD +/-15.1 and a range of 19 to 70 years. A total of 161 bacteria strains were isolated from specimens obtained from 70 patients which were an average of 2.3 isolates per patient. Of these 103 were aerobic organism and 58 were strict anaerobes. A complex mix of strict anaerobes and facultative anaerobes accounted for 87% of all infections.The most frequent aerobes isolated was streptococcus spp 70 (70%) followed by Staphylococcus spp 18 (18%). Other organisms such as Klebsiella spp 4 (4%), Proteus spp 5 (5%) and Pseudomonas spp 2 (2%) were also seen. The anaerobic group was dominated by Prevotella spp 25 (43%) followed by Peptostreptococcus spp 18 (31%); other isolates were Pseudomonas spp 2 (1%), black pigmented Pophyromonas spp 4 (5%), Fusobacterium spp 3 (3%) and Bacteroides spp 5 (8%). Majority of these organisms were sensitive to Amoxicillin (98%), Gentamycin (89%), and Ciprofloxacin (100%). A 40% resistance to metronidazole was observed in Bacteroides spp otherwise this drug and others displayed good activity against anaerobes. Conclusions: Oral and maxillofacial facial infections at Muhimbili National Hospital are mostly caused by streptococcus spp and Prevotella spp. Strict anaerobes accounted for 36% of all isolates. The profile of isolates should assist in selecting empiric therapy for infections of the oral and maxillofacial region. Inclusion of antimicrobial agents against anaerobic bacteria is highly recommended.

Keywords: bacteria, oral and maxillofacial infections, antibiotic susceptibility, Tanzania

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Authors: Shi-Bo Feng, Sheng-Jie Zhang, Jin Zhou

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The relationship between microbial diversity and algal bloom has received considerable attention for decades. Microbes undoubtedly affect annual bloom events and impact the physiology of both partners, as well as shape ecosystem diversity. However, knowledge about interactions and network correlations among broader-spectrum microbes that lead to the dynamics in a complete bloom cycle are limited. In this study, pyrosequencing and network approaches simultaneously assessed the associate patterns among bacteria, archaea, and microeukaryotes in surface water and sediments in response to a natural dinoflagellate (Alexandrium sp.) bloom. In surface water, among the bacterial community, Gamma-Proteobacteria and Bacteroidetes dominated in the initial bloom stage, while Alpha-Proteobacteria, Cyanobacteria, and Actinobacteria become the most abundant taxa during the post-stage. In the archaea biosphere, it clustered predominantly with Methanogenic members in the early pre-bloom period while the majority of species identified in the later-bloom stage were ammonia-oxidizing archaea and Halobacteriales. In eukaryotes, dinoflagellate (Alexandrium sp.) was dominated in the onset stage, whereas multiply species (such as microzooplankton, diatom, green algae, and rotifera) coexistence in bloom collapse stag. In sediments, the microbial species biomass and richness are much higher than the water body. Only Flavobacteriales and Rhodobacterales showed a slight response to bloom stages. Unlike the bacteria, there are small fluctuations of archaeal and eukaryotic structure in the sediment. The network analyses among the inter-specific associations show that bacteria (Alteromonadaceae, Oceanospirillaceae, Cryomorphaceae, and Piscirickettsiaceae) and some zooplankton (Mediophyceae, Mamiellophyceae, Dictyochophyceae and Trebouxiophyceae) have a stronger impact on the structuring of phytoplankton communities than archaeal effects. The changes in population were also significantly shaped by water temperature and substrate availability (N & P resources). The results suggest that clades are specialized at different time-periods and that the pre-bloom succession was mainly a bottom-up controlled, and late-bloom period was controlled by top-down patterns. Additionally, phytoplankton and prokaryotic communities correlated better with each other, which indicate interactions among microorganisms are critical in controlling plankton dynamics and fates. Our results supplied a wider view (temporal and spatial scales) to understand the microbial ecological responses and their network association during algal blooming. It gives us a potential multidisciplinary explanation for algal-microbe interaction and helps us beyond the traditional view linked to patterns of algal bloom initiation, development, decline, and biogeochemistry.

Keywords: microbial community, harmful algal bloom, ecological process, network

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Authors: Elena Iacob, Marie-Louise Ionescu, Elena Ionescu, Carmen Elena Tebrencu, Oana Teodora Ciuperca

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Infrared spectroscopy (FT-IR) is an advanced technique frequently used to authenticate both raw materials and final products using their specific fingerprints and to determine plant extracts biomarkers based on their functional groups. In recent years the market for Hypericum has grown rapidly and also has grown the cases of adultery/replacement, especially for Hypericum perforatum L.specie. Presence/absence of same biomarkers provides preliminary identification of Hypericum species in safe use in the manufacture of food supplements. The main objective of the work was to characterize the main biomarkers of Hypericum perforatum L. (St. John's wort) and identify this species in herbal food supplements after specific FT-IR fingerprint. An experimental program has been designed in order to test: (1) raw material (St. John's wort); (2)intermediate raw materials (St. John's wort dry extract ); (3) the finished products: tablets based on powders, on extracts, on powder and extract, hydroalcoholic solution from herbal mixture based on St. John's wort. The analyze using FTIR infrared spectroscopy were obtained raw materials, intermediates and finished products spectra, respectively absorption bands corresponding and similar with aliphatic and aromatic structures; examination was done individually and through comparison between Hypericum perforatum L. plant species and finished product The tests were done in correlation with phytochemical markers for authenticating the specie Hypericum perforatum L.: hyperoside, rutin, quercetin, isoquercetin, luteolin, apigenin, hypericin, hyperforin, chlorogenic acid. Samples were analyzed using a Shimatzu FTIR spectrometer and the infrared spectrum of each sample was recorded in the MIR region, from 4000 to 1000 cm-1 and then the fingerprint region was selected for data analysis. The following functional groups were identified -stretching vibrations suggests existing groups in the compounds of interest (flavones–rutin, hyperoside, polyphenolcarboxilic acids - chlorogenic acid, naphtodianthrones- hypericin): oxidril groups (OH) free alcohol type: rutin, hyperoside, chlorogenic acid; C = O bond from structures with free carbonyl groups of aldehyde, ketone, carboxylic, ester: hypericin; C = O structure with the free carbonyl of the aldehyde groups, ketone, carboxylic acid, esteric/C = O free bonds present in chlorogenic acid; C = C bonds of the aromatic ring (condensed aromatic hydrocarbons, heterocyclic compounds) present in all compounds of interest; OH phenolic groups: present in all compounds of interest, C-O-C groups from glycoside structures: rutin, hyperoside, chlorogenic acid. The experimental results show that: (I)The six fingerprint region analysis indicated the presence of specific functional groups: (1) 1000 - 1130 cm-1 (C-O–C of glycoside structures); (2) 1200-1380 cm-1 (carbonyl C-O or O-H phenolic); (3) 1400-1450 cm-1 (C=C aromatic); (4) 1600- 1730 cm-1 (C=O carbonyl); (5) 2850 - 2930 cm-1 (–CH3, -CH2-, =CH-); (6) 338-3920 cm-1 (OH free alcohol type); (II)Comparative FT-IR spectral analysis indicate the authenticity of the finished products ( tablets) in terms of Hypericum perforatum L. content; (III)The infrared spectroscopy is an adequate technique for identification and authentication of the medicinal herbs , intermediate raw material and in the food supplements less in the form of solutions where the results are not conclusive.

Keywords: Authentication, FT-IR fingerprint, Herbal supplements, Hypericum perforatum L.

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Authors: Alvaro Valera Sosa

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Background: In recent years, childhood overweight and obesity have become an increasing and challenging phenomenon in Berlin and Germany in general. The highest shares of childhood overweight in Berlin are district localities within the inner city ring with lowest socio-economic levels and the highest number of migration background populations. Most factors explaining overweight and obesity are linked to individual dispositions and nutrition balances. Among various strategies, to target drinking behaviors of children and adolescents has been proven to be effective. On the one hand, encouraging the intake of water – which does not contain energy and thus may support a healthy weight status – on the other hand, reducing the consumption of sugar-containing beverages – which are linked to weight gain and obesity. Anyhow, these preventive approaches have mostly developed into individual or educational interventions widely neglecting environmental modifications. Therefore, little is known on how urban physical environment patterns and features can act as influence factors for childhood overweight. Aiming the development of a physical environment intervention tackling children overweight, this study evaluated urban situations surrounding public playgrounds in Berlin where the issue is evident. It verified the presence and state of physical environmental conditions that can be conducive for children to engage physical activity and water intake. Methods: The study included public playgrounds for children from 0-12 y/o within district localities with the highest prevalence of childhood overweight, highest population density, and highest mixed uses. A systematic observation was realized to describe physical environment patterns and features that may affect children health behavior leading to overweight. Neighborhood walkability for all age groups was assessed using the Walkability for Health framework (TU-Berlin). Playground physical environment conditions were evaluated using Active Living Research assessment sheets. Finally, the food environment in the playground’s pedestrian catchment areas was reviewed focusing on: proximity to suppliers offering sugar-containing beverages, and physical access for 5 y/o children and up to drinking water following the Drinking Fountains and Public Health guidelines of the Pacific Institute. Findings: Out of 114 locations, only 7 had a child population over 3.000. Three with the lowest socio-economic index and highest percentage of migration background were selected. All three urban situations presented similar walkability: large trafficked avenues without buffer bordering at least one side of the playground, and important block to block disconnections for active travel. All three playgrounds rated equipment conditions from good to very good. None had water fountains at the reach of a 5 y/o. and all presented convenience stores and/or fast food outlets selling sugar-containing beverages nearby the perimeter. Conclusion: The three playground situations selected are representative of Berlin locations where most factors that influence children overweight are found. The results delivered urban and architectural design directives for an environmental intervention, used to study children health-related behavior. A post-intervention evaluation could prove associations between designed spaces and children overweight rate reduction creating a precedent in public health interventions and providing novel strategies for the health sector.

Keywords: children overweight, evaluation research, public playgrounds, urban design, urban health

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Authors: Danilo F. Rodrigues, Hérida Regina N. Salgado

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Introduction: The emergence of resistant microorganisms to a large number of clinically approved antimicrobials has been increasing, which restrict the options for the treatment of bacterial infections. As a strategy, drugs with high antimicrobial activities are in evidence. Stands out a class of antimicrobial, the cephalosporins, having as fourth generation cefepime (CEF) a semi-synthetic product which has activity against various Gram-positive bacteria (e.g. oxacillin resistant Staphylococcus aureus) and Gram-negative (e.g. Pseudomonas aeruginosa) aerobic. There are few studies in the literature regarding the development of microbiological methodologies for the analysis of this antimicrobial, so researches in this area are highly relevant to optimize the analysis of this drug in the industry and ensure the quality of the marketed product. The development of microbiological methods for the analysis of antimicrobials has gained strength in recent years and has been highlighted in relation to physicochemical methods, especially because they make possible to determine the bioactivity of the drug against a microorganism. In this context, the aim of this work was the development and validation of a microbiological method for quantitative analysis of CEF in powder lyophilized for injectable solution by turbidimetric assay. Method: For performing the method, Staphylococcus aureus ATCC 6538 IAL 2082 was used as the test microorganism and the culture medium chosen was the Casoy broth. The test was performed using temperature control (35.0 °C ± 2.0 °C) and incubated for 4 hours in shaker. The readings of the results were made at a wavelength of 530 nm through a spectrophotometer. The turbidimetric microbiological method was validated by determining the following parameters: linearity, precision (repeatability and intermediate precision), accuracy and robustness, according to ICH guidelines. Results and discussion: Among the parameters evaluated for method validation, the linearity showed results suitable for both statistical analyses as the correlation coefficients (r) that went 0.9990 for CEF reference standard and 0.9997 for CEF sample. The precision presented the following values 1.86% (intraday), 0.84% (interday) and 0.71% (between analyst). The accuracy of the method has been proven through the recovery test where the mean value obtained was 99.92%. The robustness was verified by the parameters changing volume of culture medium, brand of culture medium, incubation time in shaker and wavelength. The potency of CEF present in the samples of lyophilized powder for injectable solution was 102.46%. Conclusion: The turbidimetric microbiological method proposed for quantification of CEF in lyophilized powder for solution for injectable showed being fast, linear, precise, accurate and robust, being in accordance with all the requirements, which can be used in routine analysis of quality control in the pharmaceutical industry as an option for microbiological analysis.

Keywords: cefepime hydrochloride, quality control, turbidimetric assay, validation

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Authors: Hyohun Kim, Dongwha Shin, Yeonseok Kim, Ji-Su Ahn, Kensuke Nakamura, Dongeun Choi, Byung-Woo Hong

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Artefacts are commonly encountered in the imaging process of clinical computed tomography (CT) where the artefact refers to any systematic discrepancy between the reconstructed observation and the true attenuation coefficient of the object. It is known that CT images are inherently more prone to artefacts due to its image formation process where a large number of independent detectors are involved, and they are assumed to yield consistent measurements. There are a number of different artefact types including noise, beam hardening, scatter, pseudo-enhancement, motion, helical, ring, and metal artefacts, which cause serious difficulties in reading images. Thus, it is desired to remove nuisance factors from the degraded image leaving the fundamental intrinsic information that can provide better interpretation of the anatomical and pathological characteristics. However, it is considered as a difficult task due to the high dimensionality and variability of data to be recovered, which naturally motivates the use of machine learning techniques. We propose an image restoration algorithm based on the deep neural network framework where the denoising auto-encoders are stacked building multiple layers. The denoising auto-encoder is a variant of a classical auto-encoder that takes an input data and maps it to a hidden representation through a deterministic mapping using a non-linear activation function. The latent representation is then mapped back into a reconstruction the size of which is the same as the size of the input data. The reconstruction error can be measured by the traditional squared error assuming the residual follows a normal distribution. In addition to the designed loss function, an effective regularization scheme using residual-driven dropout determined based on the gradient at each layer. The optimal weights are computed by the classical stochastic gradient descent algorithm combined with the back-propagation algorithm. In our algorithm, we initially decompose an input image into its intrinsic representation and the nuisance factors including artefacts based on the classical Total Variation problem that can be efficiently optimized by the convex optimization algorithm such as primal-dual method. The intrinsic forms of the input images are provided to the deep denosing auto-encoders with their original forms in the training phase. In the testing phase, a given image is first decomposed into the intrinsic form and then provided to the trained network to obtain its reconstruction. We apply our algorithm to the restoration of the corrupted CT images by the artefacts. It is shown that our algorithm improves the readability and enhances the anatomical and pathological properties of the object. The quantitative evaluation is performed in terms of the PSNR, and the qualitative evaluation provides significant improvement in reading images despite degrading artefacts. The experimental results indicate the potential of our algorithm as a prior solution to the image interpretation tasks in a variety of medical imaging applications. This work was supported by the MISP(Ministry of Science and ICT), Korea, under the National Program for Excellence in SW (20170001000011001) supervised by the IITP(Institute for Information and Communications Technology Promotion).

Keywords: auto-encoder neural network, CT image artefact, deep learning, intrinsic image representation, noise reduction, total variation

Procedia PDF Downloads 189

Authors: Gadanya, A.M., Ponfa, S., Jibril, M.M., Abubakar, S. M.

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Background: Edible insects such as grasshopper are important sources of food for humans, and have been consumed as traditional foods by many indigenous communities especially in Africa, Asia, and Latin America. These communities have developed their skills and techniques in harvesting, preparing, consuming, and preserving edible insects, widely contributing to the role played by the use of insects in human nutrition. Aim/ objective: This study was aimed at determining the effect of insect enriched sprouted soyabean /millet based food on some gut health markers in albino rats. Methods. Four different formulations of Complementary foods (i.e Complementary Food B (CFB): sprouted millet (SM), Complementary Food C (CFC): sprouted soyabean (SSB), Complementary Food D (CFD): sprouted soybean and millet (SSBM) in a ratio of (50:50) and Complementary Food E (CFE): insect (grasshopper) enriched sprouted soybean and millet (SSBMI) in a ratio of (50:25:25)) were prepared. Proximate composition and short chain fatty acid contents were determined. Thirty albino rats were divided into5 groups of six rats each. Group 1(CDA) were fed with basal diet and served as a control group, while groups 2,3,4 and 5 were fed with the corresponding complimentary foods CFB, CFC, CFD and CFE respectively daily for four weeks. Concentrations of fecal protein, serum total carotenoids and nitric oxide were determined. DNA extraction for molecular isolation and characterization were carried out followed by PCR, the use of mega 11 software and NCBI blast for construction of the phylogenetic tree and organism identification respectively. Results: Significant increase (P<0.05) in percentage ash, fat, protein and moisture contents, as well as short chain fatty acid (acetate, butyrate and propionate) concentrations were recorded in the insect enriched sprouted composite food (CFE) when compared with the CFA, CFB, CFC and CFD composite food. Faecal protein, carotenoid and nitric oxide concentrations were significantly lower (P>0.05) in group 5 in comparison to groups 1to 4. Ruminococcus bromii and Bacteroidetes were molecularly isolated and characterized by 16s rRNA from the sprouted millet/sprouted soybean and the insect enriched sprouted soybean/sprouted millet based food respectively. The presence of these bacterial strains in the feaces of the treated rats is an indication that the gut of the treated rats is colonized by good gut bacteria, hence, an improved gut health. Conclusion: Insect enriched sprouted soya bean/sprouted millet based complementary diet showed a high composition of ash, fat, protein and fiber. Thus, could increase the availability of short chain fatty acids whose role to the host organism cannot be overemphasized. It was also found to have decrease the level of faecal protein, carotenoid and nitric oxide in the serum which is an indication of an improvement in the immune system function.

Keywords: gut-health, insect, millet, soybean, sprouted

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Authors: Nada Verdel, Tomaz Rijavec, Albin Pintar, Ales Lapanje

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Objectives: White water circuits of woodfree paper mills contain suspended, dissolved, and colloidal particles, such as cellulose, starch, paper sizings, and dyes. By closing the white water circuits, these particles start to accumulate and affect the production. Due to high amount of organic matter that scavenge radicals and adsorbs onto catalyst surfaces, treatment of white water with photocatalysis is inappropriate. The most suitable approach should be bioaugmentation-assisted bioremediation. Accordingly, objectives were: - to isolate bacteria capable of degrading organic compounds used for the papermaking process - to select the most active bacteria for bioaugmentation. Status: The state-of-the-art of bioaugmentation of pulp and paper mill effluents is mostly based on biodegradation of lignin. Whereas in white water circuits of woodfree paper mills only papermaking compounds are present. As far as one can tell from the literature, the study on degradation activities of bacteria for all possible compounds of the papermaking process is a novelty. Methodology: The main parameters of the selected white water were systematically analyzed during a period of two months. Bacteria were isolated on selective media with particular carbon source. Organic substances used as carbon source either enter white water circuits as base paper or as recycled broke. The screening of bacterial activities for starch, cellulose, latex, polyvinyl alcohol, alkyl ketene dimers, and resin acids was followed by addition of lugol. Degraders of polycyclic aromatic dyes were selected by cometabolism tests; cometabolism is simultaneous biodegradation of two compounds, in which the degradation of the second compound depends on the presence of the first. The obtained strains were identified by 16S rRNA sequencing. Findings: 335 autochthonous strains were isolated on plates with selected carbon source. The isolated strains were selected according to degradation of the particular carbon source. The ultimate degraders of cationic starch, cellulose, and sizings are Pseudomonas sp. NV-CE12-CF and Aeromonas sp. NV-RES19-BTP. The most active strains capable of degrading azo dyes are Aeromonas sp. NV-RES19-BTP and Sphingomonas sp. NV-B14-CF. Klebsiella sp. NV-Y14A-BTP degrade polycyclic aromatic direct blue 15 and also yellow dye, Agromyces sp. NV-RED15A-BF and Cellulosimicrobium sp. NV-A4-BF are specialists for whitener and Aeromonas sp. NV-RES19-BTP is general degrader of all compounds. To the white water adapted bacteria were isolated and selected according to their degradation activities for particular organic substances. Mostly isolated bacteria are specialized to lower the competition in the microbial community. Degraders of readily-biodegradable compounds do not degrade recalcitrant polycyclic aromatic dyes and vice versa. General degraders are rare.

Keywords: bioaugmentation, biodegradation of azo dyes, cometabolism, smart wastewater treatment technologies

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Authors: I. C. Orabueze, A. A. Amudalat, S. A. Adesegun, A. A. Usman

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Dental and associated oral diseases are increasingly affecting a considerable portion of the population and are considered some of the major causes of tooth loss, discomfort, mouth odor and loss of confidence. This study focused on the ethnobotanical survey of medicinal plants used in oral therapy and evaluation of the antimicrobial activities of methanolic extracts of two selected plants from the survey for their efficacy against dental microorganisms. The ethnobotanical survey was carried out in six herbal markets in Lagos State, Nigeria by oral interviewing and information obtained from an old family manually complied herbal medication book. Methanolic extracts of Olax subscorpioidea (stem bark) and Bridelia ferruginea (stem bark) were assayed for their antimicrobial activities against clinical oral isolates (Aspergillus fumigatus, Candida albicans, Streptococcus spp, Staphylococcus aureus, Lactobacillus acidophilus and Pseudomonas aeruginosa). In vitro microbial technique (agar well diffusion method and minimum inhibitory concentration (MIC) assay) were employed for the assay. Chlorhexidine gluconate was used as the reference drug for comparison with the extract results. And the preliminary phytochemical screening of the constituents of the plants were done. The ethnobotanical survey produced plants (28) of diverse family. Different parts of plants (seed, fruit, leaf, root, bark) were mentioned but 60% mentioned were either the stem or the bark. O. subscorpioidea showed considerable antifungal activity with zone of inhibition ranging from 2.650 – 2.000 cm against Aspergillus fumigatus but no such encouraging inhibitory activity was observed in the other assayed organisms. B. ferruginea showed antibacterial sensitivity against Streptococcus spp, Staphylococcus aureus, Lactobacillus acidophilus and Pseudomonas aeruginosa with zone of inhibitions ranging from 3.400 - 2.500, 2.250 - 1.600, 2.700 - 1.950, 2.225 – 1.525 cm respectively. The minimum inhibitory concentration of O. subscorpioidea against Aspergillus fumigatus was 51.2 mg ml-1 while that of B. ferruginea against Streptococcus spp was 0.1mg ml-1 and for Staphylococcus aureus, Lactobacillus acidophilus and Pseudomonas aeruginosa were 25.6 mg ml-1. A phytochemical analysis reveals the presence of alkaloids, saponins, cardiac glycoside, tannins, phenols and terpenoids in both plants, with steroids only in B. ferruginea. No toxicity was observed among mice given the two methanolic extracts (1000 mg Kg-1) after 21 days. The barks of both plants exhibited antimicrobial properties against periodontal diseases causing organisms assayed, thus up-holding their folkloric use in oral disorder management. Further research could be done viewing these extracts as combination therapy, checking for possible synergistic value in toothpaste and oral rinse formulations for reducing oral bacterial flora and fungi load.

Keywords: antimicrobial activities, Bridelia ferruginea, dental disinfection, methanolic extract, Olax subscorpioidea, ethnobotanical survey

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Authors: Tomasz Borowski, Dawid Sołoducha, Agata Markowska-Szczupak, Aneta Wesołowska, Marian Kordas, Rafał Rakoczy

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Essential oils (EOs) are fragrant volatile oils obtained from plants. These are used for cooking (for flavor and aroma), cleaning, beauty (e.g., rosemary essential oil is used to promote hair growth), health (e.g. thyme essential oil cures arthritis, normalizes blood pressure, reduces stress on the heart, cures chest infection and cough) and in the food industry as preservatives and antioxidants. Rosemary and thyme essential oils are considered the most eminent herbs based on their history and medicinal properties. They possess a wide range of activity against different types of bacteria and fungi compared with the other oils in both in vitro and in vivo studies. However, traditional uses of EOs are limited due to rosemary and thyme oils in high concentrations can be toxic. In light of the accessible data, the following hypothesis was put forward: Low frequency rotating magnetic field (RMF) increases the antimicrobial potential of EOs. The aim of this work was to investigate the antimicrobial activity of commercial Salvia Rosmarinus L. and Thymus vulgaris L. essential oil from Polish company Avicenna-Oil under Rotating Magnetic Field (RMF) at f = 25 Hz. The self-constructed reactor (MAP) was applied for this study. The chemical composition of oils was determined by gas chromatography coupled with mass spectrometry (GC-MS). Model bacteria Escherichia coli K12 (ATCC 25922) was used. Minimum inhibitory concentrations (MIC) against E. coli were determined for the essential oils. Tested oils in very small concentrations were prepared (from 1 to 3 drops of essential oils per 3 mL working suspensions). From the results of disc diffusion assay and MIC tests, it can be concluded that thyme oil had the highest antibacterial activity against E. coli. Moreover, the study indicates the exposition to the RMF, as compared to the unexposed controls causing an increase in the efficacy of antibacterial properties of tested oils. The extended radiation exposure to RMF at the frequency f= 25 Hz beyond 160 minutes resulted in a significant increase in antibacterial potential against E. coli. Bacteria were killed within 40 minutes in thyme oil in lower tested concentration (1 drop of essential oils per 3 mL working suspension). Rapid decrease (>3 log) of bacteria number was observed with rosemary oil within 100 minutes (in concentration 3 drops of essential oils per 3 mL working suspension). Thus, a method for improving the antimicrobial performance of essential oil in low concentrations was developed. However, it still remains to be investigated how bacteria get killed by the EOs treated by an electromagnetic field. The possible mechanisms relies on alteration in the permeability of ionic channels in ionic channels in the bacterial cell walls that transport in the cells was proposed. For further studies, it is proposed to examine other types of essential oils and other antibiotic-resistant bacteria (ARB), which are causing a serious concern throughout the world.

Keywords: rotating magnetic field, rosemary, thyme, essential oils, Escherichia coli

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Authors: Rossana Roila, Raffaella Branciari, Lorenzo Cardinali, David Ranucci

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The wild boar (Sus scrofa) population has strongly increased across Europe in the last decades, also causing severe fauna management issues. In central Italy, wild boar is the main hunted wild game species, with approximately 40,000 animals killed per year only in the Umbria region. The meat of the game is characterized by high-quality nutritional value as well as peculiar taste and aroma, largely appreciated by consumers. This type of meat and products thereof can meet the current consumers’ demand for higher quality foodstuff, not only from a nutritional and sensory point of view but also in relation to environmental sustainability, the non-use of chemicals, and animal welfare. The game meat production chain is characterized by some gaps from a hygienic point of view: the harvest process is usually conducted in a wild environment where animals can be more easily contaminated during hunting and subsequent practices. The definition and implementation of a certified and controlled supply chain could ensure quality, traceability and safety for the final consumer and therefore promote game meat products. According to European legislation in some animal species, such as bovine, the use of weak acid solutions for carcass decontamination is envisaged in order to ensure the maintenance of optimal hygienic characteristics. A preliminary study was carried out to evaluate the applicability of similar strategies to control the hygienic level of wild boar carcasses. The carcasses, harvested according to the selective method and processed into the game-handling establishment, were treated by nebulization with two different solutions: a 2% food-grade lactic acid solution and aromatic vinegar. Swab samples were performed before treatment and in different moments after-treatment of the carcasses surfaces and subsequently tested for Total Aerobic Mesophilic Load, Total Aerobic Psychrophilic Load, Enterobacteriaceae, Staphylococcus spp. and lactic acid bacteria. The results obtained for the targeted microbial populations showed a positive effect of the application of the lactic acid solution on all the populations investigated, while aromatic vinegar showed a lower effect on bacterial growth. This study could lay the foundations for the optimization of the use of a lactic acid solution to treat wild boar carcasses aiming to guarantee good hygienic level and safety of meat.

Keywords: game meat, food safety, process hygiene criteria, microbial population, microbial growth, food control

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Authors: Jia H. Wong, Jingli Zhang, Habsah Mohamad, Iswatun H. Abdullah Ripain, Muhammad Bilal, Amelia J. Lloyd, Abdul A. Mohamed Yusoff, Jafri M. Abdullah

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Epilepsy is one of the most common neurological diseases affecting more than 50 million of people worldwide. Epilepsy is a state of recurrent, spontaneous seizures with multiple syndromes and symptoms of different causes of brain dysfunction, prognosis, and treatments; characterized by transient, occasional and stereotyped interruptions of behavior whereby the excitatory-inhibitory activities within the central nervous system (CNS) are thrown out of balance due to various kinds of interferences. The goal of antiepileptic treatment is to enable patients to be free from seizures or to achieve control of seizures through surgical treatment and/or pharmacotherapy. Pharmacotherapy through AED plays an important role especially in countries with epilepsy treatment gap due to costs and availability of health facilities, skills and resources, yet there are about one-third of the people with epilepsy have drug-resistant seizures. Hence, this poses considerable challenges to the healthcare system and the effort in providing cost-effective treatment as well as the search for alternatives to treatment and management of epilepsy. Enhancement of γ-aminobutyric acid (GABA)-mediated inhibitory neurotransmission is one of the key mechanisms of actions of antiepileptic drugs. GABA type > a receptors (GABAAR) are ligand-gated ion channels that mediate rapid inhibitory neurotransmission upon the binding of GABA with a heteropentameric structure forming a central pore that is permeable to the influx of chloride ions in its activated state. The major isoform of GABAA receptors consists of two α1, two β2, and one γ2 subunit. It is the most abundantly expressed combinations in the brain and the most commonly researched through Xenopus laevis oocytes. With the advancing studies on ethnomedicine and traditional treatments using medicinal plants, increasing evidence reveal that spice and herb plants with medicinal properties play an important role in the treatment of ailments within communities across different cultures. Capsaicin is the primary natural capsaicinoid in hot peppers of plant genus Capsicum, consist of an aromatic ring, an amide linkage and a hydrophobic side chain. The study showed that capsaicins conferred neuroprotection in status epilepticus mouse models through anti-ictogenic, hypothermic, antioxidative, anti-inflammatory, and anti-apoptotic actions in a dose-dependent manner. In this study, five capsaicin derivatives were tested for their ability to increase the GABA-induced chloride current on α1β2γ2S of GABAAR expressed on Xenopus laevis oocytes using the method of two-microelectrode voltage clamp. Two of the capsaicin derivatives, IS5 (N-(4-hydroxy-3-methoxybenzyl)-3-methylbutyramide) and IS10 (N-(4-hydroxy-3-methoxybenzyl)-decanamide) at a concentration of 30µM were able to significantly increase the GABA-induced chloride current with p=0.002 and p=0.026 respectively. This study were able to show the enhancement effect of two capsaicin derivatives with moderate length of hydrocarbon chain on this receptor subtype, revealing the promising inhibitory activity of capsaicin derivatives through enhancement of GABA-induced chloride current and further investigations should be carried out to verify its antiepileptic effects in animal models.

Keywords: α1β2γ2 GABAA receptors, α1β2γ2S, antiepileptic, capsaicin derivatives, two-microelectrode voltage clamp, Xenopus laevis oocytes

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Authors: Norihiro Kato, Yuriko Takayama

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Some gram-negative bacteria enable the simultaneous activation of gene expression involved in N-acylhomoserine lactone (AHL) dependent cell-to-cell communication system. Such regulatory system for the bacterial group behavior is termed as quorum sensing (QS) because a diffusible AHL signal can accumulate around the cell during the increase of the cell density and trigger activation of the sequential QS process. By blocking the QS, the expression of diverse genes related to infection, antibiotic production, and biofilm formation is inhibited. Conditioning of QS by regulation of the DNA-receptor-AHL interaction is a potential target for enhancing host defenses against pathogenicity. We focused on engineered application of transcriptional regulator SpnR produced in opportunistic human pathogen Serratia marcescens. The SpnR can interact with AHL signals at an N-terminal domain and also with a promoter region of a QS target gene at a C-terminal domain. As the initial process of the QS activation, the SpnR forms a complex with the AHL to enhance the expression of pig cluster; the SpnR normally acts as an activator for the expression of the QS-dependent gene. In this research, we attempt to artificially control QS by changing the role of SpnR. The QS-dependent prodigiosin production is expected to inhibit by externally added SpnR in the culture broth of AS-1 strain because the AHL concentration was kept below the threshold by AHL-SpnR complex formation. Maltose-binding protein (MBP)-tagged SpnR (MBP-SpnR) was overexpressed in Escherichia coli and purified using an affinity chromatography equipped with an amylose resin column. The specific interaction between AHL and MBP-SpnR was demonstrated by quartz crystal microbalance (QCM) sensor. AHL with amino end-group was coupled with COOH-terminated self-assembled monolayer prepared on a gold electrode of 27-MHz quartz crystal sensor using water-soluble carbodiimide. After the injection of MBP-SpnR into a cup-type sensor cell filled with the buffer solution, time course of resonant frequency change (ΔFs) was determined. A decrease of ΔFs clearly showed the uptake of MBP-SpnR onto the AHL-immobilized electrode. Furthermore, no binding affinity was observed after the heat-inactivation of MBP-SpnR at 80ºC. These results suggest that MBP-SpnR possesses a specific affinity for AHL. MBP-SpnR was added to the culture medium as an AHL trap to study inhibitory effects on intracellularly accumulated prodigiosin. With approximately 2 µM MBP-SpnR, the amount of prodigiosin induced was half that of the control without any additives. In conclusion, the function of SpnR could be switched by adding it to the cell culture. Exogenously added MBP-SpnR possesses high affinity for AHL derived from cells and acts as an inhibitor of AHL-mediated QS.

Keywords: intracellular signaling, microbial biotechnology, quorum sensing, transcriptional regulator

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Authors: Claudia Cocozza, Niccolò Frassinelli, Enrico Marchi, Cristiano Foderi, Alessandro Bizzarri, Margherita Paladini, Maria Laura Traversi, Eleftherious Touloupakis, Alessio Giovannelli

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The development of tools to quickly identify the fate of injured trees after stress is highly relevant when biodiversity restoration of damaged sites is based on nature-based solutions. In this context, an approach to assess irreversible physiological damages within trees could help to support planning management decisions of perturbed sites to restore biodiversity, for the safety of the environment and understanding functionality adjustments of the ecosystems. Tree vitality can be estimated by a series of physiological proxies like cambium activity, starch, and soluble sugars amount in C-sinks whilst the accumulation of ethanol within the cambial cells and phloem is considered an alert of cell death. However, their determination requires time-consuming laboratory protocols, which makes the approach unfeasible as a practical option in the field. The project aims to develop biosensors to assess the concentration of soluble sugars and ethanol in stem tissues. Soluble sugars and ethanol concentrations will be used to define injured trees to discriminate compromised and recovering trees in the forest directly. To reach this goal, we select study sites subjected to prescribed fires or recent wildfires as experimental set-ups. Indeed, in Mediterranean countries, forest fire is a recurrent event that must be considered as a central component of regional and global strategies in forest management and biodiversity restoration programs. A biosensor will be developed through a multistep process related to target analytes characterization, bioreceptor selection, and, finally, calibration/testing of the sensor. To validate biosensor signals, soluble sugars and ethanol will be quantified by HPLC and GC using synthetic media (in lab) and phloem sap (in field) whilst cambium vitality will be assessed by anatomical observations. On burnt trees, the stem growth will be monitored by dendrometers and/or estimated by tree ring analyses, whilst the tree response to past fire events will be assessed by isotopic discrimination. Moreover, the fire characterization and the visual assessment procedure will be used to assign burnt trees to a vitality class. At the end of the project, a well-defined procedure combining biosensor signal and visual assessment will be produced and applied to a study case. The project outcomes and the results obtained will be properly packaged to reach, engage and address the needs of the final users and widely shared with relevant stakeholders involved in the optimal use of biosensors and in the management of post-fire areas. This project was funded by National Recovery and Resilience Plan (NRRP), Mission 4, Component C2, Investment 1.1 - Call for tender No. 1409 of 14 September 2022 – ‘Progetti di Ricerca di Rilevante interesse Nazionale – PRIN’ of Italian Ministry of University and Research funded by the European Union – NextGenerationEU; Grant N° P2022Z5742, CUP B53D23023780001.

Keywords: phloem, scorched crown, conifers, prescribed burning, biosensors

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Authors: Yuriko Takayama, Norihiro Kato

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Quorum sensing (QS) is a cell-to-cell communication system in bacteria to regulate expression of target genes. In gram-negative bacteria, activation on QS is controlled by a concentration increase of N-acylhomoserine lactone (AHL), which can diffuse in and out of the cell. Effective control of QS is expected to avoid virulence factor production in infectious pathogens, biofilm formation, and antibiotic production because various cell functions in gram-negative bacteria are controlled by AHL-mediated QS. In this research, we applied cyclodextrins (CDs) as artificial hosts for the AHL signal to reduce the AHL concentration in the culture broth below its threshold for QS activation. The AHL-receptor complex induced under the high AHL concentration activates transcription of the QS-target gene. Accordingly, artificial reduction of the AHL concentration is one of the effective strategies to inhibit the QS. A hydrophobic cavity of the CD can interact with the acyl-chain of the AHL due to hydrophobic interaction in aqueous media. We studied N-hexanoylhomoserine lactone (C6HSL)-mediated QS in Serratia marcescens; accumulation of C6HSL is responsible for regulation of the expression of pig cluster. Inhibitory effects of added CDs on QS were demonstrated by determination of prodigiosin amount inside cells after reaching stationary phase, because production of prodigiosin depends on the C6HSL-mediated QS. By adding approximately 6 wt% hydroxypropyl-β-CD (HP-β-CD) in Luria-Bertani (LB) medium prior to inoculation of S. maecescens AS-1, the intracellularly accumulated prodigiosin was drastically reduced to 7-10%, which was determined after the extraction of prodigiosin in acidified ethanol. The AHL retention ability of HP-β-CD was also demonstrated by Chromobacterium violacuem CV026 bioassay. The CV026 strain is an AHL-synthase defective mutant that activates QS solely by adding AHLs from outside of cells. A purple pigment violacein is induced by activation of the AHL-mediated QS. We demonstrated that the violacein production was effectively suppressed when the C6HSL standard solution was spotted on a LB agar plate dispersing CV026 cells and HP-β-CD. Physico-chemical analysis was performed to study the affinity between the immobilized CD and added C6HSL using a quartz crystal microbalance (QCM) sensor. The COOH-terminated self-assembled monolayer was prepared on a gold electrode of 27-MHz AT-cut quartz crystal. Mono(6-deoxy-6-N, N-diethylamino)-β-CD was immobilized on the electrode using water-soluble carbodiimide. The C6HSL interaction with the β-CD cavity was studied by injecting the C6HSL solution to a cup-type sensor cell filled with buffer solution. A decrement of resonant frequency (ΔFs) clearly showed the effective C6HSL complexation with immobilized β-CD and its stability constant for MBP-SpnR-C6HSL complex was on the order of 102 M-1. The CD has high potential for engineered control of QS because it is safe for human use.

Keywords: acylhomoserine lactone, cyclodextrin, intracellular signaling, quorum sensing

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Authors: Befekadu Urga Wakayo

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Introduction: Bacterial infections represent major human and animal health problems in Ethiopia. In the face of poor antibiotic regulatory mechanisms, development of antimicrobial resistance (AMR) to commonly used drugs has become a growing health and livelihood threat in the country. Monitoring and control of AMR demand close coloration between human and veterinary services as well as other relevant stakeholders. However, risk of AMR transfer from animal to human population’s remains poorly explored in Ethiopia. This systematic research literature review attempted to give an overview on AMR challenges of bovine mastitis bacteria in Ethiopia. Methodology: A web based research literature search and analysis strategy was used. Databases are considered including; PubMed, Google Scholar, Ethiopian Veterinary Association (EVA) and Ethiopian Society of Animal Production (ESAP). The key search terms and phrases were; Ethiopia, dairy, cattle, mastitis, bacteria isolation, antibiotic sensitivity and antimicrobial resistance. Ultimately, 15 research reports were used for the current analysis. Data extraction was performed using a structured Microsoft Excel format. Frequency AMR prevalence (%) was registered directly or calculated from reported values. Statistical analysis was performed on SPSS – 16. Variables were summarized by giving frequencies (n or %), Mean ± SE and demonstrative box plots. One way ANOVA and independent t test were used to evaluate variations in AMR prevalence estimates (Ln transformed). Statistical significance was determined at p < 0.050). Results: AMR in bovine mastitis bacteria was investigated in a total of 592 in vitro antibiotic sensitivity trials involving 12 different mastitis bacteria (including 1126 Gram positive and 77 Gram negative isolates) and 14 antibiotics. Bovine mastitis bacteria exhibited AMR to most of the antibiotics tested. Gentamycin had the lowest average AMR in both Gram positive (2%) and negative (1.8%) bacteria. Gram negative mastitis bacteria showed higher mean in vitro resistance levels to; Erythromycin (72.6%), Tetracycline (56.65%), Amoxicillin (49.6%), Ampicillin (47.6%), Clindamycin (47.2%) and Penicillin (40.6%). Among Gram positive mastitis bacteria higher mean in vitro resistance was observed in; Ampicillin (32.8%), Amoxicillin (32.6%), Penicillin (24.9%), Streptomycin (20.2%), Penicillinase Resistant Penicillin’s (15.4%) and Tetracycline (14.9%). More specifically, S. aurues exhibited high mean AMR against Penicillin (76.3%) and Ampicillin (70.3%) followed by Amoxicillin (45%), Streptomycin (40.6%), Tetracycline (24.5%) and Clindamycin (23.5%). E. coli showed high mean AMR to Erythromycin (78.7%), Tetracycline (51.5%), Ampicillin (49.25%), Amoxicillin (43.3%), Clindamycin (38.4%) and Penicillin (33.8%). Streptococcus spp. demonstrated higher (p =0.005) mean AMR against Kanamycin (> 20%) and full sensitivity (100%) to Clindamycin. Overall, mean Tetracycline (p = 0.013), Gentamycin (p = 0.001), Polymixin (p = 0.034), Erythromycin (p = 0.011) and Ampicillin (p = 0.009) resistance increased from the 2010’s than the 2000’s. Conclusion; the review indicated a rising AMR challenge among bovine mastitis bacteria in Ethiopia. Corresponding, public health implications demand a deeper, integrated investigation.

Keywords: antimicrobial resistance, dairy cattle, Ethiopia, Mastitis bacteria

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Authors: Fawzyah Albaldi

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Urinary tract infections (UTIs) are the most prevalent of infectious diseases and > 80% are caused by uropathogenic E. coli (UPEC). Infection occurs following adhesion to urothelial plaques on bladder epithelial cells, whose major protein constituent are the uroplakins (UPs). Two of the four uroplakins (UPIa and UPIb) are members of the tetraspanin superfamily. The UPEC adhesin FimH is known to interact directly with UPIa. Tetraspanins are a diverse family of transmembrane proteins that generally act as “molecular organizers” by binding different proteins and lipids to form tetraspanin enriched microdomains (TEMs). Previous work by our group has shown that TEMs are involved in the adhesion of many pathogenic bacteria to human cells. Adhesion can be blocked by tetraspanin-derived synthetic peptides, suggesting that tetraspanins may be valuable drug targets. In this study, we investigate the role of tetraspanins in UPEC adherence to bladder epithelial cells. Human bladder cancer cell lines (T24, 5637, RT4), commonly used as in-vitro models to investigate UPEC infection, along with primary human bladder cells, were used in this project. The aim was to establish a model for UPEC adhesion/infection with the objective of evaluating the impact of tetraspanin-derived reagents on this process. Such reagents could reduce the progression of UTI, particularly in patients with indwelling catheters. Tetraspanin expression on the bladder cells was investigated by q-PCR and flow cytometry, with CD9 and CD81 generally highly expressed. Interestingly, despite these cell lines being used by other groups to investigate FimH antagonists, uroplakin proteins (UPIa, UPIb and UPIII) were poorly expressed at the cell surface, although some were present intracellularly. Attempts were made to differentiate the cell lines, to induce cell surface expression of these UPs, but these were largely unsuccessful. Pre-treatment of bladder epithelial cells with anti-CD9 monoclonal antibody significantly decreased UPEC infection, whilst anti-CD81 had no effects. A short (15aa) synthetic peptide corresponding to the large extracellular region (EC2) of CD9 also significantly reduced UPEC adherence. Furthermore, we demonstrated specific binding of that fluorescently tagged peptide to the cells. CD9 is known to associate with a number of heparan sulphate proteoglycans (HSPGs) that have also been implicated in bacterial adhesion. Here, we demonstrated that unfractionated heparin (UFH)and heparin analogs significantly inhibited UPEC adhesion to RT4 cells, as did pre-treatment of the cells with heparinases. Pre-treatment with chondroitin sulphate (CS) and chondroitinase also significantly decreased UPEC adherence to RT4 cells. This study may shed light on a common pathogenicity mechanism involving the organisation of HSPGs by tetraspanins. In summary, although we determined that the bladder cell lines were not suitable to investigate the role of uroplakins in UPEC adhesion, we demonstrated roles for CD9 and cell surface proteoglycans in this interaction. Agents that target these may be useful in treating/preventing UTIs.

Keywords: UTIs, tspan, uroplakins, CD9

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Authors: Elisabetta Ceretti, Silvia Bonizzoni, Alberto Bonetti, Milena Villarini, Marco Verani, Maria Antonella De Donno, Sara Bonetta, Umberto Gelatti

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Introduction: Air pollution is a global problem. In 2013, the International Agency for Research on Cancer (IARC) classified air pollution and particulate matter as carcinogenic to human. The study of the health effects of air pollution in children is very important because they are a high-risk group in terms of the health effects of air pollution and early exposure during childhood can increase the risk of developing chronic diseases in adulthood. The MAPEC_LIFE (Monitoring Air Pollution Effects on Children for supporting public health policy) is a project founded by EU Life+ Programme which intends to evaluate the associations between air pollution and early biological effects in children and to propose a model for estimating the global risk of early biological effects due to air pollutants and other factors in children. Methods: The study was carried out on 6-8-year-old children living in five Italian towns in two different seasons. Two biomarkers of early biological effects, primary DNA damage detected with the comet assay and frequency of micronuclei, were investigated in buccal cells of children. Details of children diseases, socio-economic status, exposures to other pollutants and life-style were collected using a questionnaire administered to children’s parents. Child exposure to urban air pollution was assessed by analysing PM0.5 samples collected in the school areas for PAHs and nitro-PAHs concentration, lung toxicity and in vitro genotoxicity on bacterial and human cells. Data on the chemical features of the urban air during the study period were obtained from the Regional Agency for Environmental Protection. The project created also the opportunity to approach the issue of air pollution with the children, trying to raise their awareness on air quality, its health effects and some healthy behaviors by means of an educational intervention in the schools. Results: 1315 children were recruited for the study and participate in the first sampling campaign in the five towns. The second campaign, on the same children, is still ongoing. The preliminary results of the tests on buccal mucosa cells of children will be presented during the conference as well as the preliminary data about the chemical composition and the toxicity and genotoxicity features of PM0.5 samples. The educational package was tested on 250 children of the primary school and showed to be very useful, improving children knowledge about air pollution and its effects and stimulating their interest. Conclusions: The associations between levels of air pollutants, air mutagenicity and biomarkers of early effects will be investigated. A tentative model to calculate the global absolute risk of having early biological effects for air pollution and other variables together will be proposed and may be useful to support policy-making and community interventions to protect children from possible health effects of air pollutants.

Keywords: air pollution exposure, biomarkers of early effects, children, public health policy

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Authors: Moustafa Osman Mohammed

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This paper introduces Luhmann’s autopoietic social systems starting with the original concept of autopoiesis by biologists and scientists, including the modification of general systems based on socialized medicine. A specific type of autopoietic system is explained in the three existing groups of the ecological phenomena: interaction, social and medical sciences. This hypothesis model, nevertheless, has a nonlinear interaction with its natural environment ‘interactional cycle’ for the exchange of photon energy with molecular without any changes in topology. The external forces in the systems environment might be concomitant with the natural fluctuations’ influence (e.g. radioactive radiation, electromagnetic waves). The cantilever sensor deploys insights to the future chip processor for prevention of social metabolic systems. Thus, the circuits with resonant electric and optical properties are prototyped on board as an intra–chip inter–chip transmission for producing electromagnetic energy approximately ranges from 1.7 mA at 3.3 V to service the detection in locomotion with the least significant power losses. Nowadays, therapeutic systems are assimilated materials from embryonic stem cells to aggregate multiple functions of the vessels nature de-cellular structure for replenishment. While, the interior actuators deploy base-pair complementarity of nucleotides for the symmetric arrangement in particular bacterial nanonetworks of the sequence cycle creating double-stranded DNA strings. The DNA strands must be sequenced, assembled, and decoded in order to reconstruct the original source reliably. The design of exterior actuators have the ability in sensing different variations in the corresponding patterns regarding beat-to-beat heart rate variability (HRV) for spatial autocorrelation of molecular communication, which consists of human electromagnetic, piezoelectric, electrostatic and electrothermal energy to monitor and transfer the dynamic changes of all the cantilevers simultaneously in real-time workspace with high precision. A prototype-enabled dynamic energy sensor has been investigated in the laboratory for inclusion of nanoscale devices in the architecture with a fuzzy logic control for detection of thermal and electrostatic changes with optoelectronic devices to interpret uncertainty associated with signal interference. Ultimately, the controversial aspect of molecular frictional properties is adjusted to each other and forms its unique spatial structure modules for providing the environment mutual contribution in the investigation of mass temperature changes due to pathogenic archival architecture of clusters.

Keywords: autopoiesis, nanoparticles, quantum photonics, portable energy, photonic structure, photodynamic therapeutic system

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Authors: Mohamed M. Zaki, Maha M. Hady

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Salmonella spp. have been categorized as the world’s biggest threats to human health and poultry products are mostly incriminated sources. In Egypt, it was found that S. enteritidis and S. typhimurium are the most prevalent ones in poultry farms. It is recommended to eliminate salmonella from living bird by competing for salmonella contamination in feed in order to establish a healthy gut. The Feed acidifiers are the group of feed additives containing low-molecular-weight organic acids and/ or their salts which act as performance promoters by lowering the pH in the gut, optimizes digestion and inhibit bacterial growth. The inclusion of organic acid in pure form nonetheless effective in feed, yet, it is difficult to handle in feed mills as it is corrosive and produce more losses during pelleting process. The current study aimed at to evaluate the impact of incorporation of sodium diformate (SDF) and a commercial acidifier, CA (a mixture of butyric and propionic acids and their ammonium salts) at 0.4% dietary levels on broilers performance and the control S. enteritidis infection. Two hundreds and seventy unsexed cobb chickens were allotted in one of three treatments (90/ group) which were, the control (no acidifier, C- &C+), the 0.4% SDF (SDF- & SDF +) and the 0.4% CA (CA- & CA +) dietary levels for 35 days. Before the allocation of the groups, ten extra birds and a diet sample were bacteriologically examined to ensure negative contamination with salmonella. The birds were raised on deep-litter separated pens and had free access to feed and water all the time. The experimentally formulated diets were kept at 40C. After 24h access to the different dietary treatments, all the birds in the positive groups (n=15/ replicate) were inoculated intra-crop with 0.2 ml of 24 h broth culture of S. entertidis containing 1X 107 organisms while the negative-treated groups were inoculated with the same amount of the negative broth and second inoculation was done at 22 d of age. Colocal swabs were collected individually from all birds 2 h pre-inoculation to assure the absence of salmonella, then 1, 3, 5, 7, 21 days post-inoculation to recover salmonella. Performance parameter (body weight gain and feed efficiency) were calculated. Mortalities were recorded and reisolation of the salmonella was adopted to ensure it was the inoculated ones. The results revealed that the dietary acidification with sodium diformate significantly improved broilers performance and tends to produce heavier birds as compared to the negative control and CA groups. Moreover, the dietary inclusion of both acidifiers at level of 0.4% was able to eliminate mortalities completely at the relevant inoculation time. Regarding the shedding of S. enteritidius in positive groups, the SDF treatment resulted in significant (p<0.05) cessation of the shedding at 3 days post-inoculation compared to 7 days post-inoculation for the CA-group. In conclusion, sodium diformate at 0.4% dietary level in broiler diets has a valuable effect not only on broilers performance but also by eliminating S. enteritidis the main source of salmonella contamination in poultry farms which is feed.

Keywords: acidifier, broilers, Salmonalla spp, sodium diformate

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Authors: Nourhan Hussein Fanaki, Hoda Mohamed Gamal El-Din Omar, Nihal Kadry Moussa, Eva Adel Edward Farid

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The resistance of staphylococci to various antibiotics has become a major concern for health care professionals. The efficacy of the combinations of selected glycopeptides (vancomycin and teicoplanin) with gentamicin or rifampicin, as well as that of gentamicin/rifampicin combination, was studied against selected pathogenic staphylococcus isolated from Egypt. The molecular distribution of genes conferring resistance to these four antibiotics was detected among tested clinical isolates. Antibiotic combinations were studied using the checkerboard technique and the time-kill assay (in both the stationary and log phases). Induction of resistance to glycopeptides in staphylococci was tried in the absence and presence of diclofenac sodium as inducer. Transmission electron microscopy was used to study the effect of glycopeptides on the ultrastructure of the cell wall of staphylococci. Attempts were made to cure gentamicin resistance plasmids and to study the transfer of these plasmids by conjugation. Trials for the transformation of the successfully isolated gentamicin resistance plasmid to competent cells were carried out. The detection of genes conferring resistance to the tested antibiotics was performed using the polymerase chain reaction. The studied antibiotic combinations proved their efficacy, especially when tested during the log phase. Induction of resistance to glycopeptides in staphylococci was more promising in presence of diclofenac sodium, compared to its absence. Transmission electron microscopy revealed the thickening of bacterial cell wall in staphylococcus clinical isolates due to the presence of tested glycopeptides. Curing of gentamicin resistance plasmids was only successful in 2 out of 9 tested isolates, with a curing rate of 1 percent for each. Both isolates, when used as donors in conjugation experiments, yielded promising conjugation frequencies ranging between 5.4 X 10-2 and 7.48 X 10-2 colony forming unit/donor cells. Plasmid isolation was only successful in one out of the two tested isolates. However, low transformation efficiency (59.7 transformants/microgram plasmid DNA) of such plasmids was obtained. Negative regulators of autolysis, such as arlR, lytR and lrgB, as well as cell-wall associated genes, such as pbp4 and/or pbp2, were detected in staphylococcus isolates with reduced susceptibility to the tested glycopeptides. Concerning rifampicin resistance genes, rpoBstaph was detected in 75 percent of the tested staphylococcus isolates. It could be concluded that in vitro studies emphasized the usefulness of the combination of vancomycin or teicoplanin with gentamicin or rifampicin, as well as that of gentamicin with rifampicin, against staphylococci showing varying resistance patterns. However, further in vivo studies are required to ensure the safety and efficacy of such combinations. Diclofenac sodium can act as an inducer of resistance to glycopeptides in staphylococci. Cell-wall thickness is a major contributor to such resistance among them. Gentamicin resistance in these strains could be chromosomally or plasmid mediated. Multiple mutations in the rpoB gene could mediate staphylococcus resistance to rifampicin.

Keywords: glycopeptides, combinations, induction, diclofenac, transmission electron microscopy, polymerase chain reaction

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Authors: Celine Shi Ying Lee, Jong Jian Lee

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Background: Infectious keratitis is one of the leading causes of blindness worldwide. Prompt treatment with effective medication will control the infection early, preventing corneal scarring and visual loss. fluoroquinolones ophthalmic medication is used because of its broad-spectrum properties, potency, good intraocular penetration, and low toxicity. The study aims to evaluate the treatment outcome of corneal ulcers using Levofloxacin 1.5% ophthalmic solution (LVFX) with adjuvant oral ciprofloxacin when indicated and apply this treatment strategy in primary health care as first-line treatment. Methods: Patients with infective corneal ulcer treated in an eye center were recruited. Inclusion criteria includes Corneal infection consistent with bacterial keratitis, single or multiple small corneal ulcers. Treatment regime: LVFX hourly for the first 2 days, 2 hourly from the 3rd day, and 3 hourly on the 5th day of review. Adjuvant oral ciprofloxacin 500mg BD was administered for 5 days if there were multiple corneal ulcers or when the location of the cornea ulcer was central or paracentral. Results: 47 subjects were recruited. There were 16 (34%) males and 31 (66%) females. 40 subjects (85%) were contact lens (CL) related to corneal ulcer, and 7 subjects (15%) were non-contact lens related. 42 subjects (89%) presented with one ulcer, of which 20 of them (48%) needed adjuvant therapy. 5 subjects presented with 2 or 3 ulcers, of which 3 needed adjuvant therapy. A total of 23 subjects (49%) was given adjuvant therapy (oral ciprofloxacin 500mg BD for 5 days).21 of them (91%) were CL related. All subjects recovered fully, and the average duration of treatment was 3.7 days, with 49% of the subjects resolved on the 3rd day, 38% on the 5thday of and 13% on the 7thday. All subjects showed symptoms of relief of pain, light-sensitivity, and redness on the 3rd day with full visual recovery post-treatment. No adverse drug reactions were recorded. Conclusion: Our treatment regime demonstrated good clinical outcome as first-line treatment for corneal ulcers. A corneal ulcer is a common eye condition in Singapore, mainly due to CL wear. Pseudomonas aeruginosa is the most frequent and potentially sight-threatening pathogen involved in CL related corneal ulcer. Coagulase-negative Staphylococci, Staphylococcus aureus, and Streptococcus Pneumoniae were seen in non-CL users. All these bacteria exhibit good sensitivity rates to ciprofloxacin and levofloxacin. It is therefore logical in our study to use LVFX Eyedrops and adjuvant ciprofloxacin oral antibiotics when indicated as first line treatment for most corneal ulcers. Our study of patients, both CL related and non-CL related, have shown good clinical response and full recovery using the above treatment strategy. There was also a full restoration of visual acuity in all the patients. Eye-trained primary Healthcare practitioners can consider adopting this treatment strategy as first line treatment in patients with corneal ulcers. This is relevant during the COVID pandemic, where hospitals are overwhelmed with patients and in regions with limited access to specialist eye care. This strategy would enable early treatment with better clinical outcome.

Keywords: corneal ulcer, levofloxacin hydrate, treatment strategy, ciprofloxacin

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Authors: Yuvraj Singh Dangi, Brajesh Kumar Tiwari, Ashok Kumar Jain, Kamta Prasad Namdeo

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The potential use of liposomes as drug carriers by i.v. injection is limited by their low stability in blood stream. Firstly, phospholipid exchange and transfer to lipoproteins, mainly HDL destabilizes and disintegrates liposomes with subsequent loss of content. To avoid the pain associated with injection and to obtain better patient compliance studies concerning various dosage forms, have been developed. Conventional liposomes (unilamellar and multilamellar) have certain drawbacks like low entrapment efficiency, stability and release of drug after single breach in external membrane, have led to the new type of liposomal systems. The challenge has been successfully met in the form of Double Liposomes (DL). DL is a recently developed type of liposome, consisting of smaller liposomes enveloped in lipid bilayers. The outer lipid layer of DL can protect inner liposomes against various enzymes, therefore DL was thought to be more effective than ordinary liposomes. This concept was also supported by in vitro release characteristics i.e. DL formation inhibited the release of drugs encapsulated in inner liposomes. DL consists of several small liposomes encapsulated in large liposomes, i.e., multivesicular vesicles (MVV), therefore, DL should be discriminated from ordinary classification of multilamellar vesicles (MLV), large unilamellar vesicles (LUV), small unilamellar vesicles (SUV). However, for these liposomes, the volume of inner phase is small and loading volume of water-soluble drugs is low. In the present study, the potential of phosphatidylethanolamine (PE) lipid anchored double liposomes (DL) to incorporate two drugs in a single system is exploited as a tool to augment the H. pylori eradication rate. Preparation of DL involves two steps, first formation of primary (inner) liposomes by thin film hydration method containing one drug, then addition of suspension of inner liposomes on thin film of lipid containing the other drug. The success of formation of DL was characterized by optical and transmission electron microscopy. Quantitation of DL-bacterial interaction was evaluated in terms of percent growth inhibition (%GI) on reference strain of H. pylori ATCC 26695. To confirm specific binding efficacy of DL to H. pylori PE surface receptor we performed an agglutination assay. Agglutination in DL treated H. pylori suspension suggested selectivity of DL towards the PE surface receptor of H. pylori. Monotherapy is generally not recommended for treatment of a H. pylori infection due to the danger of development of resistance and unacceptably low eradication rates. Therefore, combination therapy with amoxicillin trihydrate (AMOX) as anti-H. pylori agent and ranitidine bismuth citrate (RBC) as antisecretory agent were selected for the study with an expectation that this dual-drug delivery approach will exert acceptable anti-H. pylori activity.

Keywords: Helicobacter pylorI, amoxicillin trihydrate, Ranitidine Bismuth citrate, phosphatidylethanolamine, multi vesicular systems

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Authors: Vita Krungleviciute, Rasa Zelvyte, Ingrida Monkeviciene, Jone Kantautaite, Rolandas Stankevicius, Modestas Ruzauskas, Elena Bartkiene

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The environmental impact of agricultural bioproducts from the processing of food crops is an increasing concern worldwide. Currently, cereal bran has been used as a low-value ingredient for both human consumption and animal feed. The most popular bioprocessing technologies for cereal bran nutritional and technological functionality increasing are enzymatic processing and fermentation, and the most popular starters in fermented feed production are lactic acid bacteria (LAB) including pediococci. However, the ruminant digestive system is unique, there are billions of microorganisms which help the cow to digest and utilize nutrients in the feed. To achieve efficient feed utilization and high milk yield, the microorganisms must have optimal conditions, and the disbalance of this system is highly undesirable. Pediococcus strains Pediococcus acidilactici BaltBio01 and Pediococcus pentosaceus BaltBio02 from spontaneous fermented rye were isolated (by rep – PCR method), identified, and characterized by their growth (by Thermo Bioscreen C automatic turbidometer), acidification rate (2 hours in 2.5 pH), gas production (Durham method), and carbohydrate metabolism (by API 50 CH test ). Antimicrobial activities of isolated pediococcus against variety of pathogenic and opportunistic bacterial strains previously isolated from diseased cattle, and their resistance to antibiotics were evaluated (EFSA-FEEDAP method). The isolated pediococcus strains were cultivated in barley/wheat bran (90 / 10, m / m) substrate, and developed supplements, with high content of valuable pediococcus, were used for Lithuanian black and white dairy cows feeding. In addition, the influence of supplements on milk production and composition was determined. Milk composition was evaluated by the LactoScope FTIR” FT1.0. 2001 (Delta Instruments, Holland). P. acidilactici BaltBio01 and P. pentosaceus BaltBio02 demonstrated versatile carbohydrate metabolism, grown at 30°C and 37°C temperatures, and acidic tolerance. Isolated pediococcus strains showed to be non resistant to antibiotics, and having antimicrobial activity against undesirable microorganisms. By barley/wheat bran utilisation using fermentation with selected pediococcus strains, it is possible to produce safer (reduced Enterobacteriaceae, total aerobic bacteria, yeast and mold count) feed stock with high content of pediococcus. Significantly higher milk yield (after 33 days) by using pediococcus supplements mix for dairy cows feeding could be obtained, while similar effect by using separate strains after 66 days of feeding could be achieved. It can be stated that barley/wheat bran could be used for higher value feed production in order to increase milk production. Therefore, further research is needed to identify what is the main mechanism of the positive action.

Keywords: barley/wheat bran, dairy cattle, fermented feed, milk, pediococcus

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Authors: V. Schmidt, L. Janovák, N. Wiegand, B. Patczai, K. Turzó

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Large bone defects are not able to heal spontaneously. Bioactive glasses seem to be appropriate (bio)materials for bone reconstruction. Bioactive glasses are osteoconductive and osteoinductive, therefore, play a useful role in bony regeneration and repair. Because of their not optimal mechanical properties (e.g., brittleness, low bending strength, and fracture toughness), their applications are limited. Bioactive glass can be used as a coating material applied on metal surfaces. In this way -when using them as implants- the excellent mechanical properties of metals and the biocompatibility and bioactivity of glasses will be utilized. Furthermore, ion release effects of bioactive glasses regarding osteogenic and angiogenic responses have been shown. Silicate bioactive glasses (45S5 Bioglass) induce the release and exchange of soluble Si, Ca, P, and Na ions on the material surface. This will lead to special cellular responses inducing bone formation, which is favorable in the biointegration of the orthopedic prosthesis. The incorporation of other additional elements in the silicate network such as fluorine, magnesium, iron, silver, potassium, or zinc has been shown, as the local delivery of these ions is able to enhance specific cell functions. Although hip and knee prostheses present a high success rate, bacterial infections -mainly implant associated- are serious and frequent complications. Infection can also develop after implantation of hip prostheses, the elimination of which means more surgeries for the patient and additional costs for the clinic. Prosthesis-related infection is a severe complication of orthopedic surgery, which often causes prolonged illness, pain, and functional loss. While international efforts are made to reduce the risk of these infections, orthopedic surgical infections (SSIs) continue to occur in high numbers. It is currently estimated that up to 2.5% of primary hip and knee surgeries and up to 20% of revision arthroplasties are complicated by periprosthetic joint infection (PJIs). According to some authors, these numbers are underestimated, and they are also increasing. Staphylococcus aureus is the leading cause of both SSIs and PJIs, and the prevalence of methicillin-resistant S. aureus (MRSA) is on the rise, particularly in the United States. These deep infections lead to implant removal and consequently increase morbidity and mortality. The study targets this clinical problem using our experience so far with the Ag-doped polymer coatings on Titanium implants. Non-modified or modified (e.g., doped with antibacterial agents, like Ag) bioactive glasses could play a role in the prevention of infections or the therapy of infected tissues. Bioactive glasses have excellent biocompatibility, proved by in vitro cell culture studies of human osteoblast-like MG-63 cells. Ag-doped bioactive glass-scaffold has a good antibacterial ability against Escherichia coli and other bacteria. It may be concluded that these scaffolds have great potential in the prevention and therapy of implant-associated bone infection.

Keywords: antibacterial agents, bioactive glass, hip and knee prosthesis, medical implants

Procedia PDF Downloads 191

Authors: Oleg Reva, Ilya Korotetskiy, Marina Lankina, Murat Kulmanov, Aleksandr Ilin

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Molecular docking approaches are widely used for design of new antibiotics and modeling of antibacterial activities of numerous ligands which bind specifically to active centers of indispensable enzymes and/or key signaling proteins of pathogens. Widespread drug resistance among pathogenic microorganisms calls for development of new antibiotics specifically targeting important metabolic and information pathways. A generally recognized problem is that almost all molecular targets have been identified already and it is getting more and more difficult to design innovative antibacterial compounds to combat the drug resistance. A promising way to overcome the drug resistance problem is an induction of reversion of drug resistance by supplementary medicines to improve the efficacy of the conventional antibiotics. In contrast to well established computer-based drug design, modeling of drug resistance reversion still is in its infancy. In this work, we proposed an approach to identification of compensatory genetic variants reducing the fitness cost associated with the acquisition of drug resistance by pathogenic bacteria. The approach was based on an analysis of the population genetic of Mycobacterium tuberculosis and on results of experimental modeling of the drug resistance reversion induced by a new anti-tuberculosis drug FS-1. The latter drug is an iodine-containing nanomolecular complex that passed clinical trials and was admitted as a new medicine against MDR-TB in Kazakhstan. Isolates of M. tuberculosis obtained on different stages of the clinical trials and also from laboratory animals infected with MDR-TB strain were characterized by antibiotic resistance, and their genomes were sequenced by the paired-end Illumina HiSeq 2000 technology. A steady increase in sensitivity to conventional anti-tuberculosis antibiotics in series of isolated treated with FS-1 was registered despite the fact that the canonical drug resistance mutations identified in the genomes of these isolates remained intact. It was hypothesized that the drug resistance phenotype in M. tuberculosis requires an adjustment of activities of many genes to compensate the fitness cost of the drug resistance mutations. FS-1 cased an aggravation of the fitness cost and removal of the drug-resistant variants of M. tuberculosis from the population. This process caused a significant increase in genetic heterogeneity of the Mtb population that was not observed in the positive and negative controls (infected laboratory animals left untreated and treated solely with the antibiotics). A large-scale search for linkage disequilibrium associations between the drug resistance mutations and genetic variants in other genomic loci allowed identification of target proteins, which could be influenced by supplementary drugs to increase the fitness cost of the drug resistance and deprive the drug-resistant bacterial variants of their competitiveness in the population. The approach will be used to improve the efficacy of FS-1 and also for computer-based design of new drugs to combat drug-resistant infections.

Keywords: complete genome sequencing, computational modeling, drug resistance reversion, Mycobacterium tuberculosis

Procedia PDF Downloads 261

Authors: Vanessa G. P. Severino, Eliangela Cristina Candida Costa, Nubia Alves Mariano Teixeira Pires Gomides, Lucilia Kato, Afif Felix Monteiro, Maria Anita Lemos Vasconcelos Ambrosio, Carlos Henrique Gomes Martins

Abstract:

One of the biomes present in Brazil is known as Cerrado, which is a vast tropical savanna ecoregion, particularly in the states of Goiás, Mato Grosso do Sul, Mato Grosso, Tocantins and Minas Gerais. Many species of plants are characterized as endemic and they have therapeutic value for a large part of the population, especially to the rural communities. Given that, the southeastern region of the state of Goiás contains about 21 rural communities, which present a form of organization based on the use of natural resources available. One of these rural communities is named of Coqueiros, where the knowledge about the medicinal plants was very important to this research. Thus, this study focuses on the ethnobotanical survey of this community on the use of Kielmeyera coriacea to treat diseases. From the 37 members interviewed, 76% indicated this species for the treatment of intestinal infection, leukemia, anemia, gastritis, gum pain, toothache, cavity, arthritis, arthrosis, healing, vermifuge, rheumatism, antibiotic, skin problems, mycoses and all kinds of infections. The medicinal properties attributed during the interviews were framed in the body system (disease categories), adapted from ICD 10; thus, 20 indications of use were obtained, among five body systems. Therefore, the root of this species was select to chemical and biological (antioxidant and antimicrobial) studies. From the liquid-liquid extraction of ethanolic extract of root (EER), the hexane (FH), ethyl acetate (FAE), and hydro alcoholic (FHA) fractions were obtained. The chemical profile study of these fractions was performed by LC-MS, identifying major compounds such as δ-tocotrienol, prenylated acylphoroglucinol, 2-hydroxy-1-methoxyxanthone and quercitrin. EER, FH, FAE and FHA were submitted to biological tests. FHA presented the best antioxidant action (EC50 201.53 μg mL-1). EER inhibited the bacterial growth of Streptococcus pyogenes and Pseudomonas aeruginosa, microorganisms associated with rheumatism, at Minimum Inhibitory Concentration (MIC) of 6.25 μg mL-1. In addition, the FH-10 subfraction, obtained from FH fractionation, presented MIC of 1.56 μg mL-1 against S. pneumoniae; EER also inhibited the fungus Candida glabrata (MIC 7.81 μg mL- 1). The FAE-4.7.3 fraction, from the fractionation of FAE, presented MIC of 200 μg mL-1 against Lactobacillus casei, which is one of the causes of caries and oral infections. By the correlation of the chemical and biological data, it is possible to note that the FAE-4.7.3 and FH-10 are constituted 4-hydroxy-2,3-methylenedioxy xanthone, 3-hydroxy-1,2-dimethoxy xanthone, lupeol, prenylated acylphoroglucinol and quercitrin, which could be associated with the biological potential found. Therefore, this study provides an important basis for further investigations regarding the compounds present in the active fractions of K. coriacea, which will permit the establishment of a correlation between ethnobotanical survey and bioactivity.

Keywords: biological activity, ethnobotanical survey, Kielmeyera coriacea Mart., LC-MS profile

Procedia PDF Downloads 138

Authors: Abirami Baleswaran, Christel Couderc, Loubnah Belahcen, Jean Dayde, Hélène Tormo, Gwénaëlle Jard

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Cheeses are often reported to be spoiled by Pseudomonas spp., responsible for defects in appearance, texture, taste, and smell, leading to their non-marketing and even their destruction. Despite preventive actions, problems linked to Pseudomonas spp. are difficult to control by the lack of knowledge and control of these contaminants during the cheese manufacturing. Lactic goat cheese producers are not spared by this problem and are looking for solutions to decrease the number of spoiled cheeses. To explore different hypotheses, experiments are needed. However, cheese-making experiments at the pilot scale are expensive and time consuming. Thus, there is a real need to develop a miniature cheeses model system under controlled conditions. In a previous study, several miniature cheese models corresponding to different type of commercial cheeses have been developed for different purposes. The models were, for example, used to study the influence of milk, starters cultures, pathogen inhibiting additives, enzymatic reactions, microflora, freezing process on cheese. Nevertheless, no miniature model was described on the lactic goat cheese. The aim of this work was to develop a miniature cheese model system under controlled laboratory conditions which resembles commercial lactic goat cheese to study Pseudomonas spp. spoilage during the manufacturing and ripening process. First, a protocol for the preparation of miniature cheeses (3.5 times smaller than a commercial one) was designed based on the cheese factorymanufacturing process. The process was adapted from “Rocamadour” technology and involves maturation of pasteurized milk, coagulation, removal of whey by centrifugation, moulding, and ripening in a little scale cellar. Microbiological (total bacterial count, yeast, molds) and physicochemical (pH, saltinmoisture, moisture in fat-free)analyses were performed on four key stages of the process (before salting, after salting, 1st day of ripening, and end of ripening). Factory and miniature cheeses volatilomewere also obtained after full scan Sift-MS cheese analysis. Then, Pseudomonas spp. strains isolated from contaminated cheeses were selected on their origin, their ability to produce pigments, and their enzymatic activities (proteolytic, lecithinasic, and lipolytic). Factory and miniature curds were inoculated by spotting selected strains on the cheese surface. The expression of cheese spoilage was evaluated by counting the level of Pseudomonas spp. during the ripening and by visual observation and under UVlamp. The physicochemical and microbiological compositions of miniature cheeses permitted to assess that miniature process resembles factory process. As expected, differences involatilomes were observed, probably due to the fact that miniature cheeses are made usingpasteurized milk to better control the microbiological conditions and also because the little format of cheese induced probably a difference during the ripening even if the humidity and temperature in the cellar were quite similar. The spoilage expression of Pseudomonas spp. was observed in miniature and factory cheeses. It confirms that the proposed model is suitable for the preparation of miniature cheese specimens in the spoilage study of Pseudomonas spp. in lactic cheeses. This kind of model could be deployed for other applications and other type of cheese.

Keywords: cheese, miniature, model, pseudomonas spp, spoilage

Procedia PDF Downloads 132