International Journal of Biotechnology and Recent Advances

ISSN: 2639-4529

International Biotechnology and Research Conference

April 25-27, 2018, Rome, Italy
Scientific Session Abstracts
DOI: 10.18689/2639-4529.a1.002

New Opportunities of Biocatalysis in Industrial Biotechnology

Luis P. Fonseca

Institute Superior Técnico, University of Lisbon, Portugal

Chemical products continue to have vital importance to the world economy and are essential to the health, food and consumer product industries. However, they led to environmental damage and a low public perception of chemical industry. For that reason, there is increasing pressure from both society and Governments for development of new industrial processes to become more sustainable, reducing waste and preventing the use of toxic substances. Consequently, there is a huge opportunity for research and implementation of new cleaner and green processes in the field of industrial biotechnology namely Biocatalysis and Biotransformation.

Within this scope, Luis P. Fonseca has focus his research interests on application of biocatalysis in green processes, and development and implementation of reaction media using basically water due to higher enzyme stability and also increasing environment concerns. The design of oil-in-water emulsions (also named miniemulsions) have allowed the production of a high range of high-value products from flavors and fragrances, emollients, polymers, nutraceuticals, and specialty and fine chemicals that led to the development of Aromase technology. This technology is characterized by the enzymatic catalysis, utilization of renewable material resources, high energetic efficiency, no toxicity by using reactions based in water, prevention of waste and sub-products,, among many others advantages.

Later modification of the miniemulsion composition evolved to NanoLipCar technology with main goal to design and processing of new drug delivery carriers for encapsulation of cosmeceuticals, nutraceuticals and pharmaceuticals at form of Lipid Nano-Emulsions (LNEs) and Lipid Nano-Particles (LNPs) in this last using long fatty acids that solidify at higher than 37°C.

Luis P. Fonsecais, Associate Professor with Habilitation and Ph.D. of Department of Bioengineering at Instituto Superior Tecnico (I.S.T.) of University of Lisbon (U.L.), Portugal and Senior Research Scientist (PI) at the Institute for Bioengineering and Bioscience (iBB) at I.S.T. His prior positions included Chemical Engineer at Hovione, Lda (1985-1986), a Visiting Researcher at CIPAN, Lda (1987-1988), Post-Doc at The School of Biochemistry and Molecular Biology, University of Leeds (1997-1998), Visiting Scholar at Chemical Engineering Department, University of California, Berkeley (2004-2005), and at Institute of Chemistry at University Federal of Rio Grande do Sul, Brazil (2011-2012) and Sao Carlos at University of Sao Paulo, Brazil (2015 and 2017).

Immobilization of ARM Lipase for Industrial Use

Raja Noor Zaliha Raja Abd Rahman1,2,3*, Nur Syazwani Mohtar2,3, Shuhaimi Mustafa1,2 and Mohd Basyaruddin Abdul Rahman2,4

1Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia, Malaysia
2Enzyme and Microbial Technology Research Centre, Universiti Putra Malaysia, Malaysia
3Halal Products Research Institute, Universiti Putra Malaysia, Malaysia
4Faculty of Science, Universiti Putra Malaysia, Malaysia

ARM lipase was isolated from Geobacillus sp. strain ARM, holds many potentials for industrial applications as it is thermostable, organic solvent tolerant, 1,3-regioselective. It prefers medium and long chain fatty acids as substrate. In this study, the enzyme was overexpressed in Escherichia coli system and then purified using affinity chromatography. The purified enzyme was immobilized in gelatinized sago solution and spray-dried by entrapment technique in order to enhance the enzyme operational stability for handling at high temperature and also for storage. The physical characteristic of the immobilized enzyme was studied by scanning electron microscopy, surface area and porosity. The immobilized ARM lipase showed good performance at high temperature. The immobilized enzyme could be stored at for few months without loss of activity. Collectively, the immobilized lipase shows promising capability for industrial use.

Raja Noor Zaliha Raja Abd. Rahman obtained her Doctor of Engineering in Molecular Biology from Kyoto University, Kyoto, Japan in 1998. She is currently the Professor and Head of Enzyme and Microbial Technology Research Centre, Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia. Raja Noor Zaliha is without any doubt a dedicated researcher, project leader and graduate advisor. This is reflected in the number of Top Down research projects that she has contributed to either as project leader or principal investigator as well as the number of international publications of high impact which are often cited by international investigators. In recognition of her scientific findings, she has been awarded patents, prizes and commendations and has derived personal satisfaction from the success of her graduate students, her participation on national and international governmental projects and not least, having a bacterial species named after her.

Towards the Development of a Multidimensional Multisensor Spatiotemporal Model for Disease Detection and Spread

Maria Regina Justina E. Estuar*, Hadrian Paulo M. Lim and Raphael B. Alampay

Information Systems and Computer Science, Ateneo de Manila University, Philippines

Conductivity, amount of light, and air temperature. Weather data include air temperature, relative humidity, rainfall, and air pressure. Mobile phone data include leaf images captured from infected and non-infected sites. Weather data is used to impute with the increasing availability of low cost devices in monitoring crops, small scale farmers, especially in developing countries, now have the opportunity to access Smart farming technologies in monitoring crops. The Philippines is the 3rd largest export producer of Cavendish banana. There are also over 180 small to medium scale banana plantations, nationwide. This study attempts to develop a predictive model for the early detection of Fusarium oxysporum f. sp. cubense Tropical Race 4 or more commonly known as Panama Wilt, a soil born fungi based disease that greatly affects the production of bananas in the country. The study attempts to test whether a multidimensional approach, using multi-sensor data can be used to develop a geospatial predictive model for early detection of disease. Data is extracted from three sources including soil sensors, weather station, and mobile phone. Soil data include data coming from home grown pH sensors calibrated to work with an off the shelf soil sensor that captures soil temperature, soil moisture, soil data that was corrupted or not captured for some time period. A resulting ARIMAX model has been created that had an RMSE of roughly 16 epidemic incidences for short-term forecasting. Image data has also been explored to generate structural features of possibly infected banana leaves through an autoencoder, which allows the feature extraction for a single class. This results to latent variables that efficiently encode significant information in the images. The success rate of the autoencoder is at 91% for healthy specimens, while only at 78% for infected samples. Integrating the various data extracted from both time-series and image-based models, a linear model that incorporates both spatial and temporal factors can be created to quantify and predict disease spread.

Maria Regina is a full-time Professor at the Department of Information Systems and Computer Science, Ateneo de Manila University. She currently manages two laboratories: Ateneo Java Wireless Competency Center and the Ateneo Social Computing Science Laboratory where research focus is on in the design and development of social, mobile and wireless systems to understand and model collective behavior and capacity. She teaches Social Computing, Information Systems for Disaster and Health.

A Framework for Real Time Detection and Monitoring towards of Plant Diseases: A Case Study on Use of Web and Mobile Applications on Foc TR4

Marlene M. De Leon* and Maria Regina E. Estuar

Ateneo de Manila University, Philippines

The proliferation of Internet of Things (IoT) devices and cyberphysical structures allowed mobile and web applications to be used for early detection and monitoring of plant diseases. The Cloud-based Intelligent Total Analysis System (CITAS) is an intelligent farming system composed of mobile applications, web applications, and wireless sensor networks (WSNs). It aims to detect early and monitor the presence of Fusarium oxysporum cubense TR4 (Foc TR4) on Cavendish bananas cultivated in the Philippines using modeling and analysis.

CITAS Mobile was developed as a data collection tool for farmers. Data collected include: 1) plant leaf images; 2) qualitative plant characteristics; 3) soil WSN parameter readings (e.g., humidity, pH); and 4) GPS coordinates. CITAS Mobile analyzes plant leaf images in real-time to determine the presence or absence of the disease. Data collected by the mobile application are sent to the CITAS Server through its Application Program Interface (API) end-point via mobile internet connection for multidimensional modeling and analysis. CITAS Web provides farm owners and researchers access to historical data collected by CITAS mobile. CITAS Web has the following features: 1) data-entry of soil sample characteristics; 2) database display of farms and plants; 3) historical trend visualization of soil WSN parameter readings; and 4) visualization of multidimensional analysis results involving plant leaf images, microscopic soil images, soil WSN parameter readings, and soil physico-chemical and nutrient characteristics.

CITAS provides a framework for collection, management, analysis, and visualization of plant disease spread in a geospatial interface. Currently, total analysis allows for early detection and monitoring of Foc TR4. Farmers, farm owners, and researchers may use CITAS for early detection and monitoring of other plant diseases through extension of its analysis parameters. Through this study, the framework can be modified and used to monitor other plant diseases.

Marlene De Leon earned her Ph.D. in Computer Science degree from the Ateneo de Manila University, Philippines. She currently works as a full-time Associate Professor at the Department of Information Systems and Computer Science (DISCS), where she teaches Software Engineering, Systems Analysis and Design, and Database Management. She serves as one of the mentors in the Ateneo Social Computing Science Laboratory and the Ateneo Java Wireless Competency Center where disaster management systems and healthcare management systems are developed.

The Automated Detection of Fusarium Oxysporum Sp. in Soil Samples through Mapped Methods of Image Analysis and Machine Learning

Andrei D. Coronel* and Maria Regina E. Estuar

Ateneo de Manila University, Philippines

The export of bananas is a major area of trade in the Philippines, with Cavendish cultivars making up 50% of the export volume of this market. This type of banana, however, is susceptible to the infection of Fusarium oxysporum sp. Cubense, Tropical Race 4 (Foc TR4), the fungus that causes Fusarium Wilt. Capable of rendering wide areas of banana farms with infected plantations, Foc is a significant threat to this commerce. There is therefore a need for the early detection of Foc, especially in juvenile and asymptomatic bananas. This study has implemented an image analysis and machine learning approach in detecting Foc in magnified soil samples. Several image analysis methods were tested alongside a number of machine learning techniques towards the goal of identifying an ideal mapping of these procedures. This study not only served to improve the classification accuracy of Foc identification, but the results also acted as a guide to the development of both the soil preparation protocol and mobile microscope design. The results show that CNN, ANN, and SVM machine learning methods yield classification accuracies of 72.6%, 82.23%, and 81.12%, respectively, where SVM performed with the fastest processing time. It is conclusive that a fluorescence-based soil preparation protocol paired with at least 100x magnification and a phase contrast modality produces a viable input for a shape-recognition based image analysis technique. The study is an important step towards a multi-parameter approach in the early detection of Foc TR4 infection, thus potentially changing farmer behavior from reactive practices to preventive measures in the context of Fusarium Wilt. Similar methods can be applied in other plant diseases that make use of image analysis for early detection of diseases.

Dr. Andrei Coronel graduated with Bachelor of Science degree in Biology in 1999, a Master of Science degree in 2005, and a Ph.D. degree in Computer Science in 2014. He was a Research Engineer for the IIR (A-Star)/NUS in Singapore last 2008, where he contributed in the development of an ontology-based cancer database system. He involves himself with projects that intersect CS with health or environmental science, such as this recent project in UC Berkeley. He is currently the head of the Ateneo Computational Sound and Music Lab in his University, furthering his research oriented in data analytics.

QCM-based Biosensors for the Detection of Tumor Released Exosomes

Agnese Magnani1*, Gemma Leone1, Marco Consumi1, Ada Fort1, Valerio Vignoli1, Giorgia Radano2, Natasa Zarovni2 and Antonio Chiesi2

1Siena University and INSTM Siena Research Unit, Italy
2Exosomics Siena S.p.A., Italy

Biosensors can satisfy the rapidity and accuracy diagnosis requirements in cancer biomarkers (tumor associated antigens) detection during early stages of the disease, thus overcoming many of the problems related to the classical diagnostic methods more expensive and often time consuming. Modern bioaffinitysensors, such as DNA- or immunosensors, have recently demonstrated great potential for monitoring cancer-related protein markers and DNA mutations.

Mass-sensitive devices like Quartz Crystal Microbalances (QCMs) are commonly used to develop bio-affinity sensors: this kind of sensor is usually made of an AT-cut quartz disk with electrodes on both sides, one of which is functionalized with a receptor selective to the target analyte. If the quartz is used as a feedback element in an electronic sinusoidal oscillator (setting in this way the oscillation frequency), in case of a bio-recognition event, the change of the mass of the quartz is proportionally converted to a frequency shift of the oscillator frequency, providing an indirect measure of the adsorbed mass with a good sensitivity (1Hz/ng for 10 MHz AT-cut quartzes). The development and the characterization of QCM biosensors for the detection of exosomes are presented. Exosomes are cell-derived vesicles that are present in many biological fluids, and that possess diagnostic potential in the oncologic field. From tests with physiological solutions and human plasma, the developed biosensors have proved to give a rapid response (within minutes) with high sensitivity and specificity against the PSMA (prostate-specific membrane antigen).

Agnese Magnani is a Professor of Inorganic Chemistry- University of Siena, Department of Biotechnology, Chemistry and Pharmacy.
Her research topics are:
- Functional polymers, materials and coatings for biomedical and agriculture applications
- Biosensors for biomarkers identification and determination in body fluids
- Nano carriers for drug target delivery
- Study of molecular recognition processes: protein-ligand interactions; protein and cell interaction with solid surfaces; biofilm formation
- Application of IR and ToF-SIMS to materials and biological systems: thin films, SAMs and Nano materials characterization; geographical characterization of agrifood products; development of micro-imaging methods for biological processes in tissues and cells.

Removal of Lead and Mercury from Contaminated Water

Hossein Rostami1 and Mozhgan Bahadory2

1Philadelphia University, USA
2Community College of Philadelphia, USA

Over the past 15 years, about one half million sites with potential contamination have been reported to federal or state authorities. Of these, about 217,000 sites still need remediation and new contaminated sites continue to appear each year. The most common type of contaminants is metals, solvents and petroleum products. Heavy metals are present in two thirds of Department of Defense (DOD) and superfund sites and about 50% of Department of Energy (DOE) and Resource Conservation and Recovery Act (RCRA) sites. Heavy metals are the largest class of contaminates and also the most difficult to treat. Heavy metals most frequently found in the waste stream are lead, mercury, chromium, cadmium, arsenic and zinc. This work focuses on the removal of lead from aqueous solution.

Lead and Mercury has long been acknowledged as a harmful environmental contaminant. Lead pollution impacts all of the bodyʼs systems. Lead exposure can adversely influence the brain, central nervous system, blood cells, and kidneys. Mercury exposure can adversely influence entire body including the brain, central nervous system, lung, skin, and kidneys. To remove lead and mercury from wastewater, several methods are utilized. Some of these methods are chemical precipitation, ion exchange adsorption, reverse osmosis and electrodialysis. All of these techniques have shortcomings.

Alkali Fly Ash Permeable Reactive Barrier (AFA-PRB) is a newly developed novel material made with fly ash alkali activating solution and filler material (sand and coarse aggregates). AFA-PRB was used to remove leadand mercury from contamination water. AFA-PRB materials with high permeability were created. For reactive barrier material permeability, must be rapid, in the range of 10-2 cm/sec to 10-1 cm/sec. AFA - PRB from three ash sources with permeability of 10-1 cm/sec were produced and crushed into pelletized form. Effectiveness of the various barriers was determined by batch and column tests. Laboratory experiment indicates lead ion reduces from 1000 ppm to less than 2 ppm with 10 liters of solution and Lead ion from 10 ppm to less than 0.01 ppm. Laboratory experiment also indicates mercury ion reduces from 1000 ppm to less than 0.5 ppm with 10 liters of solution and mercury ion from 10 ppm to less than 0.01 ppm.

Diversity of Microorganisms in Heavy Oil Reservoirs (Russia) and their Possible Application in Meor

Diyana Sokolova1*, Tamara Babich1, Ekaterina Semenova1, Alexey Ershov1, Salimat Bidzhieva1, Igor Borzenkov1, Marat Khisametdinov2, Tatiana Tourova1 and TamaraNazina1

1Winogradsky Institute of Microbiology, Research Center of Biotechnology of the Russian Academy of Sciences, Russia
2Tatar Oil Research and Design Institute, Russia

Geological reserves of heavy and high-viscosity oil in Russia reach 6–7 billion tons and exceed residual reserves of light conventional oil. The goal of the present work was to study the diversity and activity of microorganisms from low-temperature heavy oil reservoirs (Russia) and their biotechnological potential for development of the microbiological method for enhanced recovery of heavy oil. Microorganisms are able to produce diverse oil-displacing agents (organic and mineral acids, biosurfactants, solvents, biopolymers, gases), which can be in technologies for microbial enhancement of oil recovery (MEOR).

The functional and phylogenetic diversity of microorganisms at five heavy oil deposits (Russia) was studied. The oilfield exploited without water-flooding, were shown to harbor scant microbial communities, while microbial numbers in the water-flooded strata was high. Methanogens of the genera Methanothrix, Methanobacterium and Methanoregula were revealed by high-throughput sequencing of 16S rRNA genes and DGGE analysis of mcrAgenes. Aerobic bacteria of the genera Rhodococcus, Pseudomonas, Gordonia, Cellulomonas, etc., capable of biosurfactant production, were isolated. Fermentative enrichments producing volatile organic acids (acetic, propionic, and butyric) from sugar-containing substrates were obtained. These acids dissolved the carbonates of oil-bearing rock efficiently. The major microbial groups of interest for development of MEOR technology are aerobic oil-oxidizing bacteria, anaerobic fermenting bacteria, and methanogenic prokaryotes. Core-flooding experiments with injection culture liquid of aerobic bacteria Gordoniaamicalis 6-1 and Rhodococcuserythropolis KS22 resulted in recovery of 17.6% of additional oil from the models because of biosurfactants and biomass production. Application of starch and culture liquid of fermenting enrichment capable of producing low fatty acids resulted in recovery of 13.4% of additional oil from the models. Selection of the most efficient microbial technology for enhanced recovery of heavy oil from terrigenous and carbonate strata is discussed.

The research was supported by the Russian Foundation for Basic Research, project no. 16-14-00028.

Dr. Diyana Sokolova graduated from the Mendeleyev University of Chemical Technology of Russia (MUCTR) in 2001. From 2001 she has been working as a researcher at the Laboratory of Petroleum Microbiology, Winogradsky Institute of Microbiology, Research Center of Biotechnology of the Russian Academy of Sciences,Moscow, Russia.
She did her Ph.D., on “Formation of biosurfactants by aerobic organotrophic bacteria from oil reservoir”, under the supervisor Dr. T.N. Nazina.Diyana Sokolova worked at petroleum reservoirs of Russia, China and USA.
She has an area of specialization in petroleum microbiology, biosurfactants, physiology of microorganisms, heavy oil, biodegradation, biotechnology, microbial enhancement for oil recovery (MEOR).

Assessing the Microbiome Dynamics in Three Photo-Bioreactors Established for Coking Wastewater Treatment: An Orchestration between Microalgae and Bacterial Communities

Mariam Hassan1*, Tamer Essam1 and Salwa Megahed1,2

1Department of Microbiology and Immunology, Cairo University, Egypt
2Department of Microbiology and Immunology, October University for Modern Sciences and Arts, Egypt

The investigation of microbial community structures is a significant way to understand biodegradation capacities in biological wastewater treatment processes. Photo-bioreactors A, B and C received real coking-wastewater as influent with COD 776 ± 56, 1229 ± 85 and 2033 ± 27 mg/l, respectively. In phase-1 phenol was added to the influent, while dichlorophenol was added in combination with phenol in phase-2. Treatment efficiency of algal-bacterial systems was biomonitored using different bioassays (phytotoxicity, Artemia toxicity, cytotoxicity, algal-bacterial ratio and settleability). COD removal %, phenol and dichlorophenol concentrations were also monitored. All systems efficiently detoxified the influents in phase-1. In phase-2, Systems B and C failed to detoxify the influents. Illumina-sequencing generated 2119749 effective sequences of 16S-rRNA gene from 21 samples collected from different influents and effluents. The number of observed species was significantly lower in effluent samples than influent samples, as some taxa dominated in photo-bioreactors and contributed to the systems performance. Significant difference in the microbial diversity between influent and effluent samples was detected. Proteobacteria (78%), Firmicutes (12%), Bacteroidetes (5%) and Deferribacteres (2%) were the dominant phyla in influent samples. While in effluent samples Proteobacteria (68%) and Bacteroidetes (25%) dominated. Failure in treatment process in systems B and C at phase-2 was accompanied with significant difference in the microbial diversity. Significant relative abundance of anaerobic bacteria from Deferribacteraceae and Peptococcaceae families in influent samples conformed to the nature of coking-wastewater. The co-culture of microalgae shifted the microbiome and promoted the activity of genera affiliated to Chitinophagaceae, Pseudomonadaceae and Xanthomonadaceae families, which dominated in effluent samples. These bacteria are known for their catabolic diversity that enables xenobiotic degradation. The superiority of algal-bacterial systems for coking-wastewater treatment was confirmed as co-culture of microalgae eradicated pathogenic bacteria such as Arcobacter and Legionella genera in the treated effluent.

Mariam Hassan is an Assistant Lecturer in Department of Microbiology and Immunology, Faculty of Pharmacy, Cairo University. She teaches the practical courses to the undergraduate and graduate students. She is working on her PhD project on biomonitoring water microbiome(s) involved in biological wastewater treatment processes. She had a experience on experience: microbiology, biotechnology, biodiversity, bioinformatics and high through put data analysis.

Increased Concentration of Diesel in Soil has Varying Impacts on Different Soil Biological Activities

Chukwudi O. Onwosi1,2*, Joyce N. Odimba1,2, Victor C. Igbokwe1,2 and Chinwe J. Aneke3

1Department of Microbiology, Faculty of Biological Sciences, University of Nigeria, Nigeria
2Bioconversion and Renewable Energy Research Unit, University of Nigeria, Nigeria
3Department of Biochemistry, Faculty of Applied and Natural Sciences, Enugu State University of Science and Technology, Nigeria

The resultant effects of the persistent contamination of the soil environment by organic pollutants are known to be deleterious to soil components and by extension, humans. Therefore, bioremediation monitoring is pertinent in ensuring the effective and efficient restoration of soil activities. In this study, soils polluted with varying concentrations of diesel (IC 1, IC 2, IC 3) at initial TPH concentrations of 14785.84 mg/kg, 23859.52 mg/kg and 42134.96 mg/kg, respectively, were bioremediated using rice husk as biostimulant. Different soil biological parameters namely soil enzyme activities (catalase and b-glucosidase), soil microbial biomass carbon (MBC), nitrogen (MBN) and phosphorus (MBP), soil microbial respiration as well as the soil phytotoxicity were used to monitor the bioremediation process. At the end of 56-day study, the degradation rate for IC 1, IC 2 and IC 3 were recorded at 99.1%, 98% and 97.6% respectively. The 1st, 2nd and nth-order kinetic equations were used in determining the efficiency of the treatment for the increasing concentrations of diesel polluted soils. The results of 1st order kinetics for IC 1 (k= 0.6745 d-1, R2= 0.9388); IC 2 (k= 0.5738 d-1, R2= 0.9287); IC 3 (k= 0.6058 d-1, R2= 0.9626); 2nd order kinetics for IC 1 (k= 8.748×10-5 d-1, R2= 0.8594); IC 2 (k= 4.301×10-5 d-1, R2= 0.8457); IC 3 (k= 3.046×10-5 d-1, R2= 0.9839); nth order kinetics for IC 1 (R2= 0.9492, k= 4.472 d-1, n= 0.7919); IC 2 (R2= 0.9394, k= 6.239 d-1, n= 0.7517); IC 3 (R2= 0.9882, k= 0.0028 d-1, n= 0.3082). From the results obtained, all biological activities for IC 1 except MBN were most responsive to the rice husk treatment than those of the IC 2 and IC 3. Improved plant growth was also observed in IC 1 and IC 2, as compared to IC 3, towards the end of the bioremediation study. These outcomes showed that the use of biological parameters is indispensible in monitoring the efficacy of a bioremediation process on contaminated soil.

Potential of Indigenous Lactococci as Starter Cultures in Dairy Industry

Mirjana Bojanic Rasovic

Biotechnical faculty, University of Montenegro, Montenegro

The diversity of lactic acid bacteria in traditional dairy products represents great potential in biotechnology. In order to standardize indigenous products, the basic requirement is the application of the determined indigenous lactic acid bacteria as starter cultures affecting their specific characteristics by performing fermentation and influencing the ripening process. Starter microorganisms produce lactic acid, which is very important during the coagulation and texturizing of the curd cheese. Production of volatile compounds (e.g. diacetyl and acetaldehyde) contribute to the flavor of these dairy products. Starter culture may posses a proteolytic and lipolytic activity that may be desirable, especially during the maturation of some types of cheese. They also produce bacteriocins, which prevents the growth of pathogens and many spoilage microorganisms. In the process od fermentation of cheese usually participate bacteria of the genus Lactococcus – Lc. lactis ssp. lactis, Lc. lactis ssp. cremoris and homofermentative lactobacilli. Lactococcus lactis species is one of the most important of lactic acid bacteria that are used in the dairy industry. The major functions of this species in dairy fermentation are the production of lactic acid from lactose, hydrolysis of casein and citric acid fermentation. Their metabolic end products and enzymes directly or indirectly have significant influence in determining the texture and flavour of the final products. The utilization of lactococci isolated from indigenous fermented milk products would lead to potential starter cultures with the necessary properties for typical local products that are well accepted by the local population. Also, the use of such starter cultures would allow the production of cheese with designated geographical origin.

Dr. Mirjana Bojanic Rasovic began to work in 1994 at Biotechnical Institute, now Biotecnical faculty in Podgorica. From 2012, she is a senior research associate in the field of Animal hygiene and Preventive Diseases and Microbiology.
For two terms (2003 to 2007), she was president of the Society of Microbiologists of Montenegro for which time she organized several scientific meetings and lectures. She has participated in the work of a number of domestic and foreign scientific and professional conferences. As author or co-author, about 80 scientific and professional papers have been published. She was the head of national projects “Influence of zoohygienic conditions on the appearance of mastitis in high dairy cows”, 2001-2003., “Examination of the spread of bovine infections with Mycoplasma bovis in the Montenegro”,Isolation and characterization of indigenous lactic acid bacteria for the production of specific cheeses in Montenegro”, (2009-2012).
Dr. Mirjana Bojanic Rasovic is Member of following committees: The Veterinary Chamber of Montenegro, Commission for Accreditation of the Accreditation Body of Montenegro, Technical Committee for Laboratories of the Accreditation Body of Montenegro, Sectoral Commission for Agriculture, Food Processing, Committee for Agriculture and Forestry of the Montenegrin Academy of Science and Arts, Commission of the Ministry of Agriculture and Rural Development for professional exam for graduate veterinarians.

Effect of Clarifying Agents on the Volatile Composition of Mead

Leticia M. Estevinho1,2*, Ananias Pascoal1,2, J.M. Oliveira3, A.P. Pereira1,2, Xesus Féas4 and Ofelia Anjos5,6

1Escola Superior Agraria, Instituto Politécnico de Bragança, Portugal
2Centro de Investigacao de Montanha, Escola Superior Agraria, Instituto Politecnico de Braganca, Portugal
3Centro de Engenharia Biologica, Universidade do Minho, Portugal
4Academia das Ciencias Veterinarias da Galiza, Edificio EGAP, Espanha
5Instituto Politecnico de Castelo Branco, Portugal
6Centro de Estudos Florestais, Instituto de Agronomia, Universidade Lisboa, Portugal

Mfead is an ancient alcoholic beverage containing between 8% and 18% alcohol by volume. It is obtained by fermentation of honey-wort though a complex process demanding both long-term fermentation and maturation. In wine production different procedures are applied for organoleptic propertiesʼ stabilization and improvement, among which clarification. However, studies regarding those procedures in the context of mead fermentation are practically non-existent.

This study aimed to assess the effect of several clarifying agents (i. casein, ii. gelatin, iii. silica, iv. bentonite, v. tannins and vi. bentonite + gelatine + egg yolk) on the volatile composition of mead. The volatile compounds were determined by gas chromatography fitted with a flame ionisation detector (FID) and by gas chromatography-mass spectrometry (GC-MS).

Thirty-six volatile compounds were identified, among which 42.50% belong to the group of alcohols, 40.40% were carbonyl compounds, 14.40% were acetates and 1.8% were esters. Volatile compoundsʼ concentration differed according to the concentration and type of fining agent used. Significant differences (p < 0.05) were found in ten volatile compounds independently of the type of treatment used. Highest volatile compoundsʼ concentration, mainly esters, lactones, terpenes and norisopenoids, were observed in meads clarified with silica; the lowest values were obtained for samples in which tanines were used. From the identified compounds, eleven had major impact on meadsʼ aroma, with OAV>1.

These results contribute to select the fining agent to be used in mead fermentation and, to a later extent, to improve the sensorial characteristics of this beverage.

Leticia M. Estevinho received the Zootechnical Engineering degree in 1985, the MS Degree in Biotechnology in 1989 and the PhD Degree in Science in 1995. In 2009 passed with merit the proofs of “Agregacao”. Leticia M. Estevinho published more than 100 articles in indexed international journals, wrote several book chapters and presented more 160 oral and written communications. Leticia Estevinho is the Head of the Microbiology Laboratory of Escola Superior Agraria de Braganca since 1986 and has been the principal investigator of more than 30 financed research projects.

Production and Evaluation of Protease Enzyme (B.subtilis strain) for Eco-friendly Leather Processing Empowerment in Ethiopian Tanneries

Biruk Abate

Ku Lueven Engineering College, Belgium

Across the world the tanning industries are facing a high challenge pressure from strict legislation articulated for the purpose of protecting and preserving the environment. In Ethiopia leather processing industry is considered from the directions of economic activity and environment polluting concern. In order to operate it within the environmentally compatible situations and sustain the agriculture development led industrialization policy and the ongoing development strategy of the tanning industry in Ethiopia should come out that can mitigate the adverse effect of the conventional leather processing methods particularly that of lime-sulphide dehairing in beam house operation. This study was focused on dehairing enzyme production as green technology alternative for the existed conventional unhairing practice. The B.subtilis strain was produced from microorganism from beam house waste from the local leather processing areas, sub-cultured and characterized for its growth and dehairing protease production in terms of pH, temperature, incubation time and growth and production media composition. The combined effects of pH and temperature on protease production also investigated and they were found to have high interactive effect. After the culture conditions for production were clearly studied the alkaline protease was produced in a laboratory scale fermenter containing ingredients with required concentration of glucose, peptone, MgSO4.7 H2O, KH2PO4 and FeSO4.7H2O and it was maintained at a temperature about 37°C for 24 to 48 hoursand trembled uncontrollably in the incubator operated close to 140rpm. After completion of fermentation time all fermented sauce base of the broth was separated by centrifuge rotated at 10,000 rpm at 4°C for 15-18 min and the clear liquid part was taken for crude enzyme preparation. From this clear supernatant liquid part the concentration of protease enzyme produced in the production medium was analyzed at best conditions near to PH 8.5, temperature 37°C and incubation time of 24 hours and closer to the 96 hrs operations maximum amunt of enzyme was yielded. The final protease product was recovered, partially purified and stabilized by primary downstream processing such as crude enzyme formulation, ammonium sulphate precipitation. The use of the protease product of this technique on sheep skins resulted in a highly promising hair removal efficiency that can really compete with lime-sulphide chemical unhairing process.

Biruk Abate Fenta is Lecturer and researcher at Bahir Dar Institute of Technology, Bahir Dar University in the Faculty of Chemical and Food Engineering and Biotechnology Research Institute. He received a B.S. degree in Chemical Engineering from Bahir Dar Institute of Technology, Bahir Dar University, Ethiopia and the Master of Technology in Chemical Engineering from the Addis Ababa Institute of Technology, Addis Ababa University, Ethiopia. He has been active in the area of Postharvest and Energy technologies, water quality and environment, chemicals and Pharmaceuticals researches in addition to his regular work of lecturing undergraduate students and mentoring postgraduate students during their thesis work. His national experience includes consulting feasibility project works of household/small scale, medium and large scale industries across his country. Currently he is an international PhD student in Ku Leuven Engineering Technology, Leuven, Belgium

The Casein Kinase 2 (CK2) and PIm Kinase as a Target for Anticancer Therapy

Maria Bretner*, Monika Wielechowska, Konrad Chojnacki and Patrycja Wińska

Warsaw University of Technology, Faculty of Chemistry, Poland

Protein kinases CK2 and PIM belong to serine/threonine kinases that are involved in the regulation of number of signalling pathways. They are upregulated in multiple cancers, including lymphoma, leukemia, breast, prostate, head and neck cancers and act as repressors of apoptosis, contributing to chemoresistance. Over 30 drugs, that target kinases, have been approved for clinical use over the past decade, and hundreds more are undergoing clinical trials, among them are: specific inhibitor of CK2–CX4945 and few specific inhibitors of PIM family e.g. AZD1208, CX-4595, SGI-9481 or LGB321. But their efficiency is not satisfactory so the increasing attention has been recently focused on the significant role of dual kinase inhibitors. Until now a few examples of the benzimidazole derivatives, which affect the activity of both kinases, CK2 and PIM1 have been known. We designed and synthesized novel 4,5,6,7- tetrabromo-1H-benzimidazole (TBBi) and 4,5,6,7- tetrabromo-1H-benzotriazole (TBBt) derivatives with alkylamine substituents. To test the inhibitory properties recombinant human kinase CK2α, CK2 holoenzyme, and kinase PIM1 was obtained in E. coli bacterial expression system. The most promising compound, the 3- (4,5,6,7-tetrabromo-2 methyl-1H-benzimidazol-1-yl) propan-1-amine (14b) inhibited the activity of CK2 with an IC50 of 0.35 M and PIM1 0.15 M. Furthermore, the influence of new active dual inhibitors on the cell viability was evaluated and EC50 determined for 14b with the use of CCRF-CEM, MCF-7 and PC3 cell lines were in the range of 2-4 M.

This work was supported by an NSC Poland grant 2014/13/B/NZ7/02273 and by WUT.

Maria Bretner received M.S. degree in chemistry from the Faculty of Chemistry, University of Warsaw in 1974 and Ph.D in biological sciences from the Institute of Biochemistry and Biophysics in 1997. She completed postdoctoral studies at the University of Maryland Baltimore County, USA. (1997-1999). She worked at the IBB PAS developing the methods of new inhibitors synthesis of Thymidylate Synthase, HIV RT and HCV helicase. Since 2007 she is working at the Warsaw University of Technology, Faculty of Chemistry. Research areas include chemistry andenzymology, biocatalysis, biochemistry of the cancer processes. She is a Co-author of 69 scientific papers.

Native Cell Membrane Nanoparticles System for Membrane Proteins

Youzhong Guo1,2*, Weihua Qiu1,2, Ziao Fu3, Guoyan G. Xu1, Robert A. Grassucci4, Yan Zhang1, Joachim Frank4,5 and Wayne A. Hendrickson4,6,7

1Department of Medicinal Chemistry, Virginia Commonwealth University, USA
2Institute for Structural Biology, Virginia Commonwealth University, USA
3Integrated Program in Cellular, Molecular, and Biomedical Studies, Columbia University, USA
4Department of Biochemistry and Molecular Biophysics, Columbia University, USA
5Department of Biological Sciences, Columbia University, USA
6Department of Physiology and Cellular Biophysics, Columbia University, USA
7New York Structural Biology Center, USA

We devised a native cell membrane nanoparticles system, which we applied in a single-particle cryo-EM study of the multidrug exporter AcrB. Lipid-AcrB nanoparticles were prepared directly from membranes without any use of detergents. A 3D reconstruction in C1 symmetry achieved a final density map at 3.2 Å resolution, an atomic model of quasi-C3-symmetric AcrBwas fitted to this map, and the residual density revealed many ordered lipid molecules. Most remarkably, a central cavity between the three transmembrane domains contains a 24-lipid patch of well-ordered bilayer structure. Inner leaflet lipid chains pack in a hexagonal array like that in phosphatidylethanolamine crystal structures, whereas the outer leaflet has highly irregular packing. Protein side chains interact with both leaflets and participate in the hexagonal pattern. The AcrB export mechanism requires reorganization of the lipid bilayer structure. This system should be broadly applicable for membrane protein structural biology and structure-based drug discovery and development.

Youzhong Guo was born in Xiangcheng, Henan, China, in 1974. He received the B.S. degree in biology from Henan Normal University, Xinxiang, Henan, China, in 1997. He received the Ph.D. degree in pharmacy/structural biology from the University of Texas at Austin, Austin, TX, U.S.A., in 2010. From 2010 to 2016, he worked with Dr. Wayne A. Hendrickson as a postdoc in Columbia University, New York, NY, U.S.A. In 2016, he joined the Department of Medicinal Chemistry, Virginia Commonwealth University, Richmond, VA, U.S.A. as an Assistant Professor. His current research interests include membrane protein structural biology and structure-based drug discovery.

Small Molecular Weight Peptides as Potential Therapeutics: Anticancer and Antimicrobial Effects of an Analogue to a Viral Protein

Taghrid Istivan1*, Elena Pirogova1, Terrence Piva1 and Victoria Korolik2

1Science Engineering and Health College, RMIT University, Australia
2Institute for Glycomics, Griffith University, Australia

The resonant recognition model (RRM) can be employed to de novo design small molecular weight peptide analogues to known anticancer proteins, like viral proteins, interleukins, and tumor necrosis factor (TNFα). The biological effects of RRM-MV, an 18 aabioactive peptide analogue to myxoma virus NM-T5protein (AAC55050, 483 aa), were investigated on mammalian cell cultures and selected bacterial pathogens using qualitative and quantitative cell survival methods. Human apoptosis protein arrays were used to detect the levels of pro-apoptotic and anti-apoptotic proteins in treated cancer cells versus non treated cells. The small molecular weight peptide (2.3 kDa) produced cytotoxic effects on cancer cell lines including mouse melanoma (B16), human melanoma (MM96L), squamous cell carcinoma (COLO16), prostate cancer (PC3) and breast cancer (MCF7). Yet, no cytotoxic effects were detected on human red blood cells, skin fibroblasts and other normal mammalian cell lines. Furthermore, RRM-MV exhibited a bacteriostatic effect on Gram positive bacteria such as Staphylococcus spp. Furthermore, in treated cancer cells and prior to necrosis stage, the peptide was located inside cytoplasmic components predicted to be glycoproteins, and it also expressed different levels of binding specificity to glycans like Sialy Lewis X, gangliosides and mannoses when glycan arrays were performed to evaluate the binding affinity of RRM-MV. The data from this study indicated that RRM designed peptides such as RRM-MV have a potential to be developed as effective therapeutics, mostly for their ability to penetrate cellular membranes, and to interfere with specific cellular biological functions.

Dr. Istivan is a senior academic at RMIT University, Melbourne, Australia within the Biotechnology Discipline. Her research interests are in bacterial virulence, therapies and drug development. She was awarded a PhD in molecular microbiology in 2005 and has been affiliated with RMIT University as a researcher and a lecturer for 15 years. She has also supervised several PhD and Masters research projects within her research team and authored and coauthored research articles and book chapters in the fields of microbiology and novel therapeutics. She is currently the Biosciences senior program manager (Teaching and Learning) within the School of Science.

The Effect of Etoposide Drug on Chromosomal Changes and Mitotic Index in Mouse Bone Marrow Cells

Abdul Rahman A. I. Alyahya

College of Applied Medical Science, Shaqra University, Saudi Arabia

Etoposide is one of the important chemical drugs commonly used worldwide in the treatment of many cancers and it is classified as an anti metabolic drug as it disrupt the process of multiplication of DNA in cancer cells by inhibiting DNA topoisomerase II causing the death of cancer cells. We select short-term tests in the current study, because it is a quick and accurate indicator to study the effect of chemicals and drugs on biological systems. As there are changes can determined accurately by cytological examination. Laboratory male mice of pure SWR / J were exposed to three types of concentrations (20, 40, 60 mg / kg) of Etoposide at different times to study its effect on the index rate of indirect cell division, and the extent of chromosomal abnormalities that can be caused. Treatment with Etoposide drug led to significant decrease in the index rate of cell division and significant increase of chromosomal abnormalities, and it correlates directly with concentration and inversely with the passage of time after treatment.

Dr. Abdul Rahman A. I. Alyahya is working in the Department of Clinical Laboratory Sciences, College of Applied Medical Sciences, Shaqra University, KSA. He is a cell Biologist scientist since 2005. He has strong research and teaching experience in cell Biology, toxicology, animal cell culture, Anticancer Drugs and pharmaceutical biotechnology. He obtained his PhD from King Saud University in Biology. He joined Shaqra University in 2009 as assistant professor and promoted to associate professor. Since 2009, he has secured many national grants for many research projects. He has published 31 refereed Journal papers, 12 conference presentations, 3 industrial reports, two conference proceedings. Formerly he was the dean College of science and currently he is a Vice Rector of Shaqra University for services and consultancy affairs. He has research interest in Cell biology, Toxicology, Animal Cell Culture, Pharmaceutical Biotechnology and Environmental Biotechnology.

Effect of Porphyrinoids on the Infectivity of Nosema spp. Microsporidia

Mariusz Trytek1*, Katarzyna Buczek1, Grzegorz Borsuk2, Aneta Ptaszyńska3, Anna Gromada1, Katarzyna Rybicka-Jasińska4 and Dorota Gryko4

1Department of Industrial Microbiology, Faculty of Biology and Biotechnology, Maria Curie-Skłodowska University, Poland
2Institute of Biological Basis of Animal Production, Faculty of Biology, Animal Sciences and Bioeconomy, University of Life Sciences in Lublin, Poland
3Department of Botany and Mycology, Faculty of Biology and Biotechnology, Maria Curie-Skłodowska University, Poland
4Institute of Organic Chemistry, Polish Academy of Sciences, Poland

Pathogenic fungi from the phylum Microsporidia are intracellular parasites found in both invertebrates and vertebrates. Nosema apis and N.ceranae are particularly devastating to honeybees (Apis mellifera carnica) as they complete their life cycle in the insectsʼ intestinal tract, causing total colony collapse. In the present work, protoporphyrin amide derivatives [PPIX (Lys)2, PPIX(Asp)2, PPIX(Lys-Lys)2] and porphyrins H2TTMePP and H2TmePyP were studied for their bioactivity against microsporidia isolated from dead honeybees. The microsporidia were treated in vitro with aporphyrin before they were usedto infect the honeybees. The Nosema spp. spores were isolated from winter beehive debris, purified, and incubated in a 0.5% sucrose solution (2 × 10-7 spores/mL) with one of the five porphyrins (100 µM)in the dark on a shaker at 30°Cfor 24 h. A control experiment without aporphyrin was performed simultaneously. Next, the spores were centrifuged, washed with sterile solutions of 0.9% NaCl and 0.5% sucrose to remove porphyrins, and used to infect honeybees. Differences in the infectivity of pretreated and untreated spores were determined in vivo by measuring the number of spores developed in living honeybees on the 7th, 12th and 20th day of the cage test experiment. Microsporidia preincubated with the porphyrins showed a lower infectivity. The largest differences were observed on days 12 and 20. The level of infection in bees infected with porphyrin-treated spores was 2-fold lower than in the control, and in the case of PP (Lys-Lys)2, even 3.3-fold lower. At the same time, lower bee mortality (up to 50 %) was observed compared to the control group. Morphological changes and deformations of the cell wall of the microsporidia treated with porphyrins were observed by light and scanning electron microscopy (SEM). Inactivation of Nosema spp. spores with porphyrins reduces their ability to infect honeybees and develop in their intestines, thus diminishing bee mortality. This work was financially supported by the National Science Centre, Poland (2015/17/B/NZ9/03607).

Dr. Mariusz Trytek is Assistant Professor at the Department of Industrial Microbiology, the Faculty of Biology and Biotechnology of Maria Curie-Skłodowska University in Lublin, Poland. He received his M.S. degree in organic chemistry in 2000 and earneda PhD in biological sciences in 2007 from MCSU, with which he has been affiliated as a researcher and a lecturer for 16 years now. Dr. Trytek has supervised several bachelor and master theses. His current research interests include biomimetic catalysis with porphyrins, biotransformation of organic compounds, and biomedical applications of porphyrinoid compounds. He has co-authored over30 publications (including research papers, reviews, and book chapters), and four patents. He is currently the principal investigatorin the national project “The biological activity of porphyrinoids and mechanisms of their action against intracellular bee pathogens of the genus Nosema”, 2016-2019.

Curcumin Reduces Antioxidant Activity Following Spinal Cord Injury in Rat Model

Ahmed Abdellatif1,2*, Sara K. Abu Hussein1, Hana H. Heiba2, Yomn Abdullah2, Amged Ouf2 and Hamdino Attia3

1Biotechnology Program, The American University in Cairo, Egypt
2Department of Biology, School of Sciences and Engineering, The American University in Cairo, Egypt
3Anatomy Department, Faculty of Medicine, Alazhar University, Egypt

Spinal cord injury is a debilitating disability. Oxidative damage and inflammation are two hallmarks of the secondary spinal cord injury. The objective of this study was to evaluate the potential of Curcumin (a polyphenolic compound extracted from the rhizome of Curcuma longa that has been known to possess antioxidant and anti-inflammatory properties) as an antioxidant and anti-inflammatory agent following spinal cord injury in rats, and to compare its therapeutic effects following local application directly to the injury versus its oral dietary supplementation in a spinal cord hemisection model at T9-T10. Female Sprague Dawley rats were randomized into a control, injury, and treatment groups of local single dose of Curcuma longa extract immediately on the injury site and a Dietary supplement group. Crude Curcumin was added to the animalsʼ feed (10% of daily feed) one week before and week after injury. Oxidative stress parameters were Malondialdehyde (MDA) and total antioxidant capacity (TAC). Expression of tumor necrosis factor alpha (TNF α) and interleukin 6 (IL-6) was detected using Enzyme Linked Immunosorbent Assay (ELISA). Our results show that at 7 days, although Dietary supplement was effective in increasing TAC levels and lowering TNF α expression levels, it did not affect MDA levels. Local treatment regimen has shown to be more effective on all four parameters. Our results demonstrate that local Curcumin application directly on the injury site might be more efficacious in alleviating oxidative damage and reducing inflammation following spinal cord injury.

Ahmed Abdellatif is an assistant professor of biology at the School of Sciences and Engineering, The American University in Cairo, Egypt. He received an MSc and a PhD from the University of Louisville, School of Medicine in Anatomical Sciences and Neurobiology. He also earned an MBBCH and a Master of Science in anatomy and embryology from Alexandria University School of Medicine. Before joining AUC, he was involved in curriculum design for medical education and taught medical and graduate level courses in human gross anatomy, neuroscience. Dr. Abdellatif has been interested in drug delivery and the development of cost effective treatments using natural herbal extracts for various applications.

Genomic Modification by Ocimum canum against Lead-Induced Chromosome Aberration and itʼs Effect on Antioxidant Enzymes

Tugbobo O Samuel1*, Akinyede K Adedamola1 and Oloyede O Ibidun2

1Department of Science Technology, Federal Polytechnic, Nigeria
2Department of Biochemistry, Ekiti State University, Nigeria

Anticlastogenic potential of Ocimum canum (Black leaf) extract was studied in bone marrow cells of mice using micronucleus assay. 200mg/kg of Ocimum canum aqueous extract was administered as dietary supplement for 30-days. The mice were divided into three groups A, B and C. Animals in group A were fed with distilled water, B were treated with 2.5mg/kg lead acetate while group C were fed with 200mg/kg Ocimum canum aqueous extract and 2.5mg/kg lead acetate simultaneously. After 30-days, mice were sacrificed and chromosome preparations were made from bone marrow according to colchicines hypotonic-fixation air drying Giemsa schedule. The cytogenic end-point observed was chromosomal aberration which increased significantly (P<0.05) in group B animals treated with lead acetate only. However, the chromosomal aberration was significantly (P<0.05) reduced by the extract fed to animals in group C. In addition, the effect of the extract on the defensive antioxidant enzymes of the test animals was also assessed. The results indicate synergistic effect of the extract on the antioxidant enzymes in the liver tissues. Hence, the results of this study suggest viable anticlastogenic and antioxidant potentials of Ocimum canum extract which could protect against leadinduced chromosomal aberration and as well enhance activities of antioxidant enzymes.

Molecular Characterization, Genetic Diversity and Similarities of Cladosporium species as Revealed by: Internal Transcribed Spacer – Polymerase Chain Reaction (ITS-PCR)

Mohammed S. Alhussaini

Department of Clinical Laboratory Sciences, Shaqra University, Saudi Arabia

The current investigation compared genetic diversities, and genetic similarities within and among Cladosporium species populations using PCR-based markers. Nuclear ribosomal DNA internal transcribed spacers have been used successfully to analyze intraspecific and interspecific relationships in various fungi. In the current study, we have used the internal transcribed spacer (ITS) to aid compare the ITS in length and restriction patterns. The internal transcribed spacer (ITS) was amplified using polymerase chain reaction combining primers ITS4 and ITS5. PCR products were digested with three restriction enzymes and separated by agarose electrophoresis. Restriction patterns generated by CfoI and MspI and RsaI were unique for most species assayed. Clear results were obtained by using ITS-PCR in the present study. The results were consistent with those based on biological characteristics and morphological features. The ITS-PCR fingerprinting methods presented here led to clear differentiation of the isolates at the species level. Fingerprinting profiles generated readily discriminated between each of the 6 species. Cluster analysis further supported this observation and clusters corresponding to each species are readily identified in the dendrograms.

Fine Tuning of the Silver Bullet; Epigenetic Manipulation of Immune Checkpoints in Breast Cancer

Hend M. El Tayebi

Genetic Pharmacology Research Group, German University in Cairo, Egypt

Programmed cell death protein-1 (PD-1) is an immune checkpoint receptor that prevents overstimulation of immune responses with an essential role in cancer immunity. Blockade of immune checkpoints especially PD-1/PD-L1 became a principle approach in cancer therapy. Different subtypes of Breast Cancer (BC) have different biological behaviors and distinct gene expression profiles. Moreover, microRNAs and Long-noncoding RNAs (LncRNAs) have gained wide spread attention recently as a critical key players in carcinogenesis but their underlying mechanisms remain limited. Our previous data revealed that overexpression of LncRNA; X inactive –specific transcript (XIST); negatively regulates PDL-1 in BC. Our work aims to investigate the impact of microRNAs and LncRNAs on PD-L1 in MDA-MB-231 cells, in addition to analyzing the differential expression of PD-L1 and non coding RNAs in different BC subtypes. Our data sheds the light on the differential expression of miR-182, MALAT-1 and XIST with a significant correlation to PD-L1 in TNBC and IDC. The data suggests a crucial role for these genes in the extent of evasion of cancer immunity in different BC subtypes.

Dr. EL Tayebi is an assistant professor of Genetics and Genetic Engineering and she is heading the “Genetic Pharmacology Research Group” in the faculty of Pharmacy and Biotechnology, German University in Cairo (GUC), Egypt. El Tayebi received her bachelor degree with highly honored excellence from faculty of Pharmacy and Biotechnology, GUC, in 2008. In 2009, she has been appointed as assistant lecturer of molecular pathology and pathophysiology at the GUC. And in 2014, she started teaching Cell Biology and Genetic Engineering as an assistant professor.
She pursued her M.Sc. and PhD studies in the field of Molecular Pathology in the period from 2008-2012. She received her PhD degree with “Summa cum laude” from the GUC in 2012. After PhD, El Tayebi worked as a senior scientist and as a team leader of hepatocellular carcinoma (HCC) group under the umbrella of the MPRG. During her postdoctoral period (2012-2015), she focused on developing the research skills of her HCC team as well as her by developing new lab techniques, writing proposals and earning research funds. She has published more than 15 publications in peer-reviewed international journals. El Tayebi has supervised 10 M.Sc. theses and currently she is supervising another 8 M.Sc. thesis.

A Comprehensive Dynamic Model of Gut Microbiome

Jun Geng1*, Boyang Ji1, Gang Li1, Felipe lopez Isunza4 and Jens Nielsen1,2,3

1Department of Biology and Biological Engineering, Chalmers University of Technology, Sweden
2Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, Denmark
3Wallenberg Center for Protein Research, Chalmers University of Technology, Sweden
4Departamento de Ingeniería de Procesos e Hidráulica, Metropolitan autonomous University, Mexico

Human microbiome, predominately residing in the large intestine constitutes a vastly complex ecosystem encompassing trillions of bacterial cells belonging to hundreds of species/taxa, have recently been revitalized as its extraordinary symbiosis relationship with host during their coevolution history. Long period ecological stability of this complex system with balanced composition is extremely critical to maintain host well-beings by exerting beneficial influence on host immune system development, energy harvest and reproductive physiology mediation. Advances in metagenomics and 16S pyrosequencing promote enormous efforts in uncovering the association between distinct microbial composition and specific host disease progress. Nevertheless, phenomenological prediction of correlation pattern through using taxonomic and genetic repertoire as a proxy of gut content cannot reveal the underlying deterministic processes. Knowledge that demonstrates the non-neutral assembly process of this complex ecosystem is required to develop ecosystem-wide incorporation approach that reveal ecological principles underlying the community assembly, however is still scarce. The challenges impeding theoretical development of decoding gut microbial ecosystem complexity are rooted in three indiscerptible aspects: i) endogenously, the inherently emergent complexity arising not simply due to the inextricable number of components the system is composed of, but rather of time-dependent and non-linear way that these components interact. ii) exdogenously, multi-dimensional host-microbe metabolic axes which contain a series of environmental factors, such as physical force, water absorption, hydrodynamics of colon microenvironment, initial colonization from maternal hand-over and subsequent diet regime that microbiota are exposed to have driven the complex and dynamic development trajectory is crucial while challenging to reveal the hidden ecological forces. iii) Spatial heterogeneity of microbial localization which play a significant impact on microbiota stable colonization and disease development should be incorporated into the model. Therefore, we develop a multidimension modeling framework that comprehensively address aforementioned bottlenecks hereby could be used to predict the microbiome shaping landscape and development process from newborn to first year life. Our approach, incorporates both dynamic behavior of individual species and community assembly rules which is supervised by resource allocation strategy in an intelligent way that is updated in context-dependent environment, hereby begetting higher-order community fitness function and elucidating the microbial-microbial interactions from the ecological perspective. The complex interaction network among host, microbiome and diet are implemented through the transport phenomena involving colon longitudinal direction and cross sectional direction that link differentiated phases which would shape the corresponding metabolic dynamics of spatially-specific microbiome. Our approach is applied to predict the infant microbiome evolution process. Both the microbial development process and microbial composition profile at known stages are reproduced by our model, through comparing our prediction of fecal composition with the measured 16S-rRNA data. Besides, our model predict the time-resolved metabolic and microbial profiles inside colon that is not accessible in vivo. Our in silico predictions are consistent with reported phenomena very well. Therefore, the integrative models proposed in our work would be of paramount importance to deconvolvethe embedded complexity and achieve the final controllability by simultaneously encompassing the most challenging issues in gut ecosystem across temporal and organizational scales, contextualized with human physiology.

She is a postdoc in Chalmers University of Technology, the major project she working is to investigate dynamic modeling of gut microbiota and how its interaction relationship with host, contributing to host health. Before coming to Chalmers, she graduated from Shanghai Jiao Tong University and got her PhD degree there. During her PhD study, she have been in Purdue University for two years as a visiting Scholar.

Evaluation of the Pharmacological Profile of Virgin Coconut Oil (VCO) in Indomethacin-Induced Gastric Ulcer Models

Joshua Parker E* and Okafor Joshua O

Department of Biochemistry, University of Nigeria, Nigeria

Several studies have reported various health benefits of virgin coconut oil (VCO) including its use for weight management, treatment of burns, various infections, and even HIV/AIDS. The present study was conducted to evaluate the pharmacological profile of VCO in indomethacin-induced gastric ulcer models. Twenty-four (24) Wistar albino rats were used for the study and were divided into 6 groups of 4 rats each. Group 1 rats served as the normal control; Group 2 (positive control) rats were administered with indomethacin only, at a dose of 100 mg/kg b.w. Group 3 rats were treated with a standard drug (cimetidine) at a dose of 100 mg/kg b.w. Groups 4 (3 ml/kg b.w. VCO), 5 (6 ml/kg b.w. VCO) and 6 (9 ml/kg b.w. VCO) rats were treated as stated. The extract (VCO) and the standard drug were dissolved in distilled water, and were administered orally to the rats. Treatment with the standard drug (group 3) and VCO (groups 4, 5 and 6) lasted for four days, followed by indomethacin induction (groups 2 to 6) after an overnight fasting. Four (4) hours after induction, the rats were sacrificed and the blood samples collected for biochemical analyses. Stomach tissues were also harvested for histological examination. The gastric ulcer index of rats in groups 4 and 6 was found to be significantly (p < 0.05) higher compared to that of group 1, while group 5 rats treated with 6 ml/kg b.w. VCO, showed a nonsignificant (p > 0.05) increase in the gastric ulcer index compared to that of group 1. The ulcer protective indices of low, mid and high doses of the oil were found to be 38.71, 54.84 and 48.39 respectively, which were found to be higher compared to the standard control (6.45). Treatment with VCO resulted in a non-significant (p > 0.05) increase in the gastric juice volume of rats in groups 4, 5 and 6 compared to that of group 1. There was no significant (p > 0.05) decrease in the gastric pH of groups 4 and 5 rats when compared to the normal control. However, a significant (p < 0.05) decrease in the gastric pH was observed when a high dose of the extract was administered. Administration of the extract resulted in no significant (p > 0.05) differences in the lipid profile, lipid peroxidation and antioxidant parameters, compared to the normal control. Histological findings revealed that stomach sections of rats in groups 4 and 5 showed moderate widespread mucosal necrosis and ulceration, while that of group 6 rats showed focal area of mucosal ulceration with evidence of healing by fibrosis when compared to group 1. The findings of this research revealed that virgin coconut oil (VCO) possesses ulcer ameliorative properties and could therefore be used for the treatment of gastric ulcers. The pharmacological properties of virgin coconut oil could be anchored on mechanisms such as the ability to scavenge free radicals, stabilize cell membranes, stimulate mucous and prostaglandin secretion, inhibit Helicobacter pylori and modulate lecithin cholesterol acyltransferase (LCAT) activity.

Dr. Parker Elijha Joshua is currently working as a faculty in department of Biochemistry at University of Nigeria. He got his Phd in Medical Biochemistry from University of Nigeria in 2009. He completed his master degree from University of Nigeria in 2005.

Determination of Haemolytic Effect and Spectral Analysis Using Gas Chromatography Mass Spectrometry (GC-MS), Fourier Transform Infrared (FTIR) and Ultraviolet Visible (UV-Vis) Spectroscopy of Different Extracts of Cucumis melo L. var. inodorus (Sweet Melon) Fruit

Joshua Parker Elijah*, Boyi Richard-Harris Nsenreuti and Onwubiko Henry A

Department of Biochemistry, University of Nigeria, Nigeria

Introduction: The concept that fruits and vegetables contribute to a personʼs wellbeing is as old as Hippocrates, the father of medicine, who more than 2000 years ago told his patients “let your food be your medicine and your medicine be your food”. Today, this same philosophy with regard to fruits and vegetables being more than just nutrition but medicine as well is experiencing rejuvenation. Indeed, a positive correlation has been reported between fruit consumption and the decreased risk of several chronic diseases including obesity, cardiovascular disease, and certain types of cancer etc. (Boeing et al., 2012; Jansen et al., 2011). The study is aimed at determining the haemolytic effect of Cucumis melo L. var. inodorus (sweet melon or honeydew melon) on human erythrocytes and to study this effect by different spectral analyses.

Experimental Design: In this study, haemolytic activity of the aqueous extract and ethanol extract of C. melo L. var. inodorus mesocarp, pericarp and ethanol extract of the seeds were screened individually against normal human erythrocytes.

Test tube 1: 0.5ml erythrocyte (RBC) suspension + 0.5ml SPBS (minimal control)
Test tube 2: 0.5ml erythrocyte (RBC) suspension + 0.5ml distilled water (maximal control)
Test tube 3a: 0.5ml erythrocyte suspension + 0.5ml M AQ at 125µg/ml in SPBS
Test tube 3b: 0.5ml erythrocyte suspension + 0.5ml M AQ at 250µg/ml in SPBS
Test tube 3c: 0.5ml erythrocyte suspension + 0.5ml M AQ at 500µg/ml in SPBS
Test tube 3d: 0.5ml erythrocyte suspension + 0.5ml M AQ at 1000µg/ml in SPBS
Test tube 4a: 0.5ml erythrocyte suspension + 0.5ml M ETH at 125µg/ml in SPBS
Test tube 4b: 0.5ml erythrocyte suspension + 0.5ml M ETH at 250µg/ml in SPBS
Test tube 4c: 0.5ml erythrocyte suspension + 0.5ml M ETH at 500µg/ml in SPBS
Test tube 4d: 0.5ml erythrocyte suspension + 0.5ml M ETH at 1000µg/ml in SPBS
Test tube 5a: 0.5ml erythrocyte suspension + 0.5ml P AQ at 125µg/ml in SPBS
Test tube 5b: 0.5ml erythrocyte suspension + 0.5ml P AQ at 250µg/ml in SPBS
Test tube 5c: 0.5ml erythrocyte suspension + 0.5ml P AQ at 500µg/ml in SPBS
Test tube 5d: 0.5ml erythrocyte suspension + 0.5ml P AQ at 1000µg/ml in SPBS
Test tube 6a: 0.5ml erythrocyte suspension + 0.5ml P ETH at 125µg/ml in SPBS
Test tube 6b: 0.5ml erythrocyte suspension + 0.5ml P ETH at 250µg/ml in SPBS
Test tube 6c: 0.5ml erythrocyte suspension + 0.5ml P ETH at 500µg/ml in SPBS
Test tube 6d: 0.5ml erythrocyte suspension + 0.5ml P ETH at 1000µg/ml in SPBS
Test tube 7a: 0.5ml erythrocyte suspension + 0.5ml S ETH at 125µg/ml in SPBS
Test tube 7b: 0.5ml erythrocyte suspension + 0.5ml S ETH at 250µg/ml in SPBS
Test tube 7c: 0.5ml erythrocyte suspension + 0.5ml S ETH at 500µg/ml in SPBS
Test tube 7d: 0.5ml erythrocyte suspension + 0.5ml S ETH at 1000µg/ml in SPBS

Determination of Haemolytic Activity: In vitro haemolytic activity assay was performed by spectrophotometer method (Yang et al., 2005).

Ultraviolet Visible (UV-Vis) Spectroscopy Analysis: UV-Vis spectrophotometric absorbance spectra of reaction mixtures (Test tubes: 3b, 4b, 5b, 6b and 7b) and the control (Test tube 1) were recorded at 400 – 700nm wavelengths using Agilent Technologies Cary 300 UV-Vis Spectrophotometer. Haemoglobin (Hb) behaviour and possible interaction with the different fruit extracts was also observed.

Fourier Transform Infrared (FTIR) Analysis: The mesocarp, pericarp and seeds of the sample, sweet melon and their respective extracts (M AQ, M ETH, P AQ, P ETH, S ETH); reaction mixtures with the highest extract concentration (Test tubes: 3d, 4d, 5d, 6d and 7d) were analysed with Agilent Technologies Cary 630 Fourier Transform Infrared (FTIR) Spectrometer. FTIR spectra were recorded from the range 650 – 4000 cm-1 wave number. However, the FTIR spectrometer detected only a very broad band of OH group when analysing the reaction mixtures possibly because of the high volume of water in it; but on allowing a small drop on the Diamond sampling window to dry, several interesting peaks were recorded.

Gas Chromatography – Mass Spectrometry (GC-MS) analysis: Chromatographic analysis by GC-MS was carried out on Agilent Technologies GC 7890B Series combined with Agilent Technologies

Identification of the Chemical Constituents: Interpretation of the mass spectrum was conducted using the database of National Institute Standard and Technology (NIST) having more than 62,000 patterns.

Statistical Analysis: All tests were conducted in triplicate. Data are reported as means ± standard deviation (SD). Results were analysed statistically by using SPSS. Significant differences were established by one-way analyses of variance (ANOVA) using Duncan and LSD multiple comparison statistics and the accepted level of significance was p < 0.05 for all the result.

Results: All the samples exhibited no haemolytic effect toward human erythrocytes when concentration was varied from 125 µg/ml to 1000 µg/ml. The percentage haemolysis obtained for all the extracts at the concentrations 125, 250, 500 and 1000 µg/ml respectively were: M ETH 0.011, 0.018, 0.015 and 0.010%; M AQ 0.119, 0.123, 0.064 and 0.087%; P AQ 0.046, 0.143, 0.129 and 0.152%; P ETH 0.135, 0.177, 0.263 and 0.242%; and S ETH 0.062, 0.195, 0.286 and 0.400%. These values were statistically considered not significant when compared to zero value (p > 0.05). Studies of the UV-Vis spectra of all test samples at 250 µg/ml (Test tubes 3b, 4b, 5b, 6b and 7b) and control (Test tube 1) showed similar spectra patterns when scanned from 400 – 700 nm wavelengths, with Soret peaks arising in the 413 – 415 nm region and two other smaller peaks at 540 nm and 577 nm showing the maximum absorption for haemoglobin fully saturated with oxygen. Interestingly, the absorption spectra conform to the spectra obtained from the US National Centre for Biotechnology Information. PubChem Compound Database. The FTIR Spectroscopy results obtained when all the samples (whole fruit parts: pericarp, mesocarp and seeds; extracts: P AQ, P ETH, M AQ, M ETH and S ETH; and the reaction mixtures: Test tubes 3d, 4d, 5d, 6d and 7d) were analysed showed

Conclusion: From the results obtained from this study, Cucumis melo L. var. inodorus (sweet melon) does not have any haemolytic effect on human erythrocytes (RBC) and hence pose no danger to consumers, since different methods used (i.e. Spectrophometric in vitro haemolytic assay method of Yang et al. (2005) and spectral analysis using UV-Vis spectroscopy, FTIR and GC-MS) did not show significant variation in the contents of bioactive compounds detected in the different extracts of sweet melon pericarp, mesocarp and seeds with human erythrocytes. ent of novel drugs.

Dr. Parker Elijha Joshua is currently working as a faculty in department of Biochemistry at University of Nigeria. He got his Phd in Medical Biochemistry from University of Nigeria in 2009. He completed his master degree from University of Nigeria in 2005.

The Influence of Milky-Wax Ripeness Walnuts Extracts on Formation and Properties of Clots

Olga Orlova* and Svetlana Buzhlakova

ITMO University, Institute of Refrigeration and Biotechnologies, Russia

The aim of this work was to study the influence of chemical composition of extracts from the fruits of milky-wax ripeness walnuts on the formation of lactic acid clots and on the duration of their storage. Experimental studies were carried out in the international research centre “Biotechnology of Third Millennium” Institute of Refrigeration and Biotechnology, ITMO University. Modern methods of research using laboratory equipment of companies Shimadzu, Buchi, Zeiss, Kohler and other were used.

Possibility of using of liquid and encapsulated extracts of milky-wax ripeness walnuts in the production of fermented milk drinks was investigated.

In this work, we have identified types, the optimal amount of starter culture and the influence of the extracts on growth of microbial flora of drinks.

The type, optimal dose and stage of introduction of milky-wax ripeness walnuts extract to raw materials were also defined. Changes in physicochemical properties and chemical composition of extracts were studied.

The positive effect of milky-wax ripeness walnuts extract on formation and properties of fermented milk clots was established.

Due to naphthaquinone-juglone, which is a part of walnuts extract, fermented milk drinks had a sustainable microbiological condition throughout the period of storage.

Thus, using of additives based on milky-wax ripeness walnuts enrich products with substances necessary for the daily prevention of organism from disease and environmental hazards. Developed products can be successfully used to supply of different population groups (children, the elderly, athletes, people working in extreme conditions), as well as in the health-promoting purposes.

Olga Orlova is a head of the Committee on Innovation and Technology Implementation, a member of international research centre “ Biotechnologies of the Third Millennium”, Associate Professor of the Applied Biotechnology Department at the ITMO University, Russia. She is also Skolkovo Foundation expert, Food Net adviser in Saint-Petersburg, Leader of the St. Petersburg Project “Nutrition for the Future. She was born on January 31, 1966. Olga Orlova graduated the Institute in 1989. She received her PhD in 2009 from St. Petersburg State University of Refrigeration and Food Engineering. She was the chief technologist at a dairy plant for 12 years. She is married and has 2 children. Scientific interests: food biotechnology, foods for particular nutritional uses, shelf life prolongation.