46^th World Congress on Microbiology September 18-19, 2017 Dublin, Ireland

Alisa Gricajeva is currently a PhD student at the Department of Microbiology and Biotechnology, Institute of Biosciences, Life Sciences Center, Vilnius University. Her final thesis work is associated with the study of microbial lipolytic enzymes: search of unstudied enzymes and their activity analysis. Alisa Gricajeva is an author of publications and conference theses dealing with characterization of lipolytic enzymes, improvement of their properties and assumption of their use in biotechnology. Alisa Gricajeva is also a junior researcher and lecturer of Food Microbiology and Biotechnologies and Enzymology lectures at Vilnius University.

Today microbial lipases find vast applications and have firmly established themselves in diverse areas of industrial microbiology and biotechnology. Among hydrolytic enzymes which are currently published to be the most widely used in industrial processes, lipases represent the third most commercialized group of enzymes and their production need is constantly increasing. For that reason, there is a perpetual interest in obtainment of low-cost, highly active and stable lipases which could be applied in different biotechnology branches. Furthermore, usually, immobilized enzymes are preferred over their free forms since immobilization not only improves enzyme features but also makes it possible to reuse the enzyme and increases its turnover. Whereas easily cultivable bacteria are one of the cheapest sources of lipases with different suitable features, in the present study, potential of Staphylococcus saprophyticus as a lipase producer was assayed. S. saprophyticus GTC1 isolate which was found and identified previously in our work showed remarkable lipolytic activity. As a species it is published to be a part of natural human flora sometimes causing uncomplicated urinary tract infections and is also found in soils. Secretion of the enzyme (LipGTC) was obtained in TSB liquid medium without addition of any inducers. After (NH4)2SO4 precipitation, using SDS-PAGE and tributyrin overlay zymography, LipGTC was determined to be ~50 kDa. LipGTC was eluted from the gel and was further analyzed. Enzyme had optimal activity at 35 – 50oC, pH 9 and hydrolyzed synthetic p-NP substrates and different natural lipids releasing highly valuable fatty acids. Some tested chemicals (detergents, different metal ions), as well as immobilization of the enzyme onto octyl-sepharose, hyperactivated LipGTC (relative activity in methanol and Ca2+ were 113% and 122%, respectively). LipGTC showed to be highly stable in the range of 30 – 50oC up to 6 h without remarkable loss of activity. To conclude, until now there were works published dealing with only one S. saprophyticus membrane associated Ssp lipase, therefore, present study widens knowledge about secreted S. saprophyticus lipolytic enzymes and emphasizes its potential to be applied industrially.

Gemma is currently a fourth year medical student of the international “Medicine and Surgery” English-taught program activated at Università Cattolica del Sacro Cuore, Rome, Italy. She is interested in child neuropsychiatric disorders. Gemma had the opportunity to join GNOSIS, a non profit research organiza-tion devoted to the study of movement disorders and behavioural child and is grateful for the privilege of being involved in a research project on the psychiatric aspects of infectious diseases.

A girl with Tourette syndrome (TS) and increased an-tibody levels against Streptococcus pyogenes was monitored longitudinally for the presence of nasopha-ryngeal bacteria, specific antibody titres, and autoim-munity directed against brain antigens (Fig.1). This case presented overlapping similarities with PAN-DAS patients (Paediatric Autoimmune Neuropsychi-atric Disorders Associated with Group A Streptococ-cal Infections) where tics and obsessive compulsive disorders follow acute Streptococcus pyogenes infec-tions. Microbiological monitoring indicated that the child was an intermittent Staphylococcus aureus naso-pharyngeal carrier. Clinical improvements in motor tic frequency and severity were observed during the S. aureus colonization phase and were temporally corre-lated with the downregulation of anti-streptococcal and anti-D1/D2 dopamine receptor antibody produc-tion. S. aureus is known to be very effective in the downregulation of the host immune response to pro-mote immune evasion. Dopamine is a crucial neuro-transmitter in motor control, and autoimmunity against its neuronal receptors may alter central dopa-mine pathways leading to movement and neuropsy-chiatric disorders, especially in childhood. After de-colonization, clinical conditions reverted to the poor scores previously observed, suggesting a possible role of the immune response in bacterial clearance as a trigger of symptom recrudescence. This hypothesis is consistent with the data from animal models showing that a pro-inflammatory, Th17 cell-associated immune response, is required for S. aureus nasal decoloniza-tion. These findings imply that a cause– effect rela-tionship exists between S. aureus colonization and tic improvement, as well as between bacterial decoloni-zation and tic exacerbation. Conclusion & Signifi-cance: This case is the first demonstration of the mod-ulation of tic manifestation in a S. aureus intermittent carrier with TS. A shift occurred from an anti-inflammatory modulatory response during the coloni-zation phase to a pro-inflammatory state during the clearing process. Thus, S. aureus nasal carriage possi-bly provides a new human model for the in vivo study of the interplay between infections, immunity, auto-immunity, and tic disorders.

Safoura Salahi has her expertise in food microbiology, probiotics and industrial microbiology. Her studies is on isolation and identification of applied bacteria. Which is very important for the industrials of food and necessary for the healthcare.

Lactic acid bacteria are gram-positive, non-spore forming, rod-shaped and cocci and have anaerobic respiration, and produce lactic acid as a final product during the fermentation of carbohydrates. These bacteria are probably abundant bacteria associated with human beings, which normally are associated with the digestive system and are also are present in habitats associated with food, including cereals.rnThe aim of this study is to isolate and identify different species of lactic acid bacteria in traditional sourdough in Chaharmahal and Bakhtiari Province, and introduce them to the industry as probiotics.rnAfter characterization of isolated bacteria, their probiotic properties were evaluated, such as resistance to acid, resistance to bile salts, Resistance to gastric juice enzymes such as pepsin and trypsin, the ability to produce antibacterial compounds, gas production and Exopolysaccharide production, then, the safety assessment of selected isolates was performed according to national standards and ultimately molecular detection was performed using 16S rDNA sequencing method.rnThe results showed that 3 strains were selected with codes A0302, A0402, A0501, which have no pathologic activity, they had the characteristics of resistance to acid over 106 cfu / ml and resistance to bile salts with deterrence factor less than 0.4, respectively, the characteristics of anti-bacterial compounds production against Escherichia coli and Staphylococcus aureus bacteria, the ability of resistance to gastric juice enzymes and A0402 had the ability to produce gas, which according to 16S rDNA sequencing, it was found that A0501 and A0302 strains had the highest affinity or Pediococcus lolii and A0402 strain had the closest with Lactobacillus fermentum which could improve the quality and quantity of sourdough and subsequently breads were produced with better quality.rn

Vilius Malunavicius is currently a Microbiology and biotechnology MSc student at the Institute of Biosciences in Vilnius University. Vilius Malunavicius has 4 years of experience working in protein engineering field for lipolytic enzymes improvement. He is as co-author of a publication and several conference theses, associated with lipolytic enzymes from Geobacillus sp. 95. He has also received prof. K. Jankevicius scholarship (2016) and worked at Greifswald University prof. U.T. Bornscheuer group under the Erasmus program for 3 months (2017.02-2017.05).

Geobacillus species show high potential as biocatalysts suitable for industrial biotechnology applications. The ability of these bacteria to produce a variety of extracellular enzymes, such as amylases, xylanases, proteases, lipases, esterases and ureases has ranked them among the most important enzyme producers. Thermostable and thermoactive lipolytic enzymes have gained remarkable importance over other industrially used biocatalysts due to their versatility regarding catalytic behavior. The main advantages of performing industrial processes at higher temperatures are reduced risk of microbial contamination and lower viscosity. It was showed that Geobacillus sp. strain 95 produces two types of thermostable, thermoactive and organic solvent-tolerant lipolytic enzymes: lipases (named GD-95) and carboxylesterases (named GDEst-95). The main goal of this research was to investigate the potential of recombinant variant of GD-95 lipase and GDEst-95 esterase for the application in industry. It was shown that both enzymes displayed an ability to perform catalysis at temperatures ranging from 5 oC to 75 °C while retaining more than 50 % of lipolytic activity after incubation at temperature range of 30-65 oC. Both biocatalysts also possessed long-term (216 h) stability in isopropanol, methanol and hexane (25 % and 50 %). Our results also showed that new esters can be obtained using a mixture composed of coconut, peach, macadamia or canola oils, 1/10 (w/w) ethanol or methanol and GD-95 lipase as biocatalyst. The major ingredient of coconut oil is lauric acid which can be turned into ethyl laureate (flavour ingredient) via esterification reaction with ethanol. The results also suggested that GD-95 lipase can be a powerful tool for the production of emulsifiers (mono- or diacylglycerols). In conclusion, the present study demonstrated that lipolytic enzymes (lipase and carboxylesterase) produced by Geobacillus sp. strain 95 have suitable properties for industrial applications and can offer an eco-friendly alternative for chemical synthesis.

Ahmad Nikseresht was affiliated with Payame Noor University as a Ph.D student until May 2013. His thesis was conducted under the supervision of Prof. M. Bakavoli in the Ferdowsi University. February 2011, Ahmad commenced research in the field of Organic Chemistry involving carbohydrates, as a Gust Ph.D-student at the laboratory of Organic Chemistry of Wageningen University. He was worked as a Gust Ph.D-student under the supervision of Prof. H. Zuilhof and Dr. Tom Wennekes for a period of nine month. He is the Director of Department of Research and Entrepreneurship of Payame Noor University of Ilam, Iran. He has published more than 37 papers in reputed journals and international conferences in the field of organic chemistry.

Umbilicus intermedius boiss (nafe venus) is important medicinal plants, especially distributed in southern regions of west Iran Ilam. Nafe venus is from crassulaceae family and thriving in the desert, and has a greenish-pink flowers, which is traditionally used to treat various infecious disease. In recent years, due to drug resistant using of medicinal plants against bacterial infecious has received attention as valuable tool to gain insight into the function of anti bacterial and improvement of their functionality. In the present study, we aim to evaluated of inhibitory effect of alcoholic, aqueous and chloroform extract of Umbilicus intermedius boiss (nafe venus) on Escherichia Coli in vitro. The plant was collected and dried in worm and low humid environment. Alcoholic, aqueous and chloroform extract prepared according to the standard method. Clinical samples of Escherichia Coli was used. Inhibitory effect of alcoholic, aqueous and chloroform extract was considered in Muller Hinton Agar medium by disk diffusion method. Inhibitory zone of alcoholic extract on Escherichia Coli was 13 mm. While zone of growth inhibition of aqueous and chloroform extract were not observed. The alcoholic extract had higher antibacteial effect than the aqueous and chloroform extract.

Veerendra Koppolu is affiliated with both University of Kansas and AstraZeneca/Medimmune, Maryland.

Enterotoxigenic Escherichia coli (ETEC) is a diarrhea causing pathogen which causes ~500,000 deaths/ year worldwide. Rns is a transcriptional activator of the AraC family and is required for virulence gene activation in ETEC. Rns has 2 domains: an amino terminal domain (NTD) and a DNA binding domain (DBD). Current evidence indicates that Rns functions as a monomer and may not require an effector to activate transcription. The main goal of this study is to identify the role of Rns-NTD in transcription activation. In previous work, two individual mutations I14T and N16D, near the amino terminus of Rns and far from known DNA-binding determinants, abolished Rns activation at its own promoter by hindering the binding of Rns to DNA. We have found that deleting the Rns-NTD (residues 1-156) leads to loss of Rns activity in vivo. Sequence analysis of Rns’ closest homologs showed that a region of the NTD (residues 12- 30) shares high identity (74%), much higher than the 26% overall NTD identity. Further, this region aligns with a region in the ToxT structure that contacts the DBD. We hypothesize that residues in this region contact Rns DBD, causing structural rearrangements in the DBD that facilitate DNA binding. One test of our hypothesis is site-directed random mutagenesis to identify mutants in this region that are defective in transcription activation. To date, we have mutagenized residues 12 to16 and identified eight variants (at positions I12, I14, N15 and N16) that decreased Rns activity by > 2 fold, indicting that these residues are important for Rns activity. On the other hand none of the mutants of K13 had decreased Rns activity. We will perform Western blots to eliminate unstable variants from consideration. We have purified Rns DBD to near homogeneity by adding a solubility tag, GB1, at the C-terminus of the DBD. The purified Rns DBD will be used for antibody generation and structural studies. Our electrophoretic mobility shift analysis (EMSA) with purified Rns DBD-GB1 showed that Rns DBD can bind to DNA in vitro at sufficiently high protein concentrations.