Antifungal and antigenotoxic effects of oleuropein

Učinak sekundarnog metabolita masline (Olea europaea L.) oleuropeina, njegovog razgradnog produkta hidroksitirosola i vodenog ekstrakta maslinova lista na staničnu vijabilnost medicinski značajne gljivične vrste Candida albicans ispitan je in vitro testovima. Metodom mikrodilucije određene su minimalne inhibitorne koncentracije (MIK) koje uzrokuju 80 % smanjenja stanične vijabilnosti i koje su iznosile 12,5 mg/ml za oleuropein, 6,25 mg/ml za hidroksitirosol i 25 mg/ml za vodeni ekstrakt maslinova lista. Kvantitativna analiza s fluorescentnim bojanjem upućuje na zaključak da je apoptoza primarni način stanične smrti u uzorcima tretiranim sa sub-MIK koncentracijama ovih tvari. Budući da je patogenost vrste određena virulentnim čimbenicima, ispitani su učinci oleuropeina i hidroksitirosola na najznačajnije virulentne čimbenike vrste C. albicans. Utvrđen je inhibitorni učinak ovih fenolnih tvari na promjenu u hifalni oblik rasta kod vrste C. albicans induciranu u uvjetima staničnog gladovanja. Pri sub-MIK koncentracijama, oleuropein i hidroksitirosol nisu pokazali inhibitorno djelovanje na stvaranje biofilma ove vrste. Pri subinhibitornim koncentracijama nakon 24 sata inkubacije, oleuropein i hidroksitirosol su uzrokovali modulaciju stanične površinske hidrofobnosti koja se dovodi u svezu s adherencijom vrste C. albicans na biomaterijale. Uzgoj vrste C. albicans u prisutnosti oleuropeina i hidroksitirosola je uzrokovao inhibiciju aktivnosti kandidinih hidrolitičkih enzima, aspartil-proteaza (Sap) i α-glukozidaza. Tretiranjem vrste C. albicans s oleuropeinom i hidroksitirosolom utvrđeno je višestruko djelovanje ovih tvari uključujući oštećenje stanične stijenke, stanične membrane i otpuštanje citoplazmatskog sadržaja (DNA i proteina). S obzirom da neki antimikotici djeluju kao inhibitori biosinteze ergosterola, provedeno je ispitivanje učinka oleuropeina i hidroksitirosola na modulaciju ergosterola kojima je utvrđeno smanjenje ovog sterola u ovisnosti o primijenjenoj koncentraciji. Uz testove određivanja djelovanja na gljivičnu vrstu, ispitani su mogući citotoksični i genotoksični učinci na stanice nositelja. Navedene tvari nisu pokazale hemolitičku aktivnost na humanim eritrocitima pri testiranim koncentracijama u rasponu 3-60 μM. Ispitivanje učinka na humanim limfocitima periferne krvi provedeno kometnim testom pokazalo je protektivno djelovanje ovih tvari na oštećenja DNA inducirana s mutagenom H2O2, pri čemu je hidroksitirosol imao bolji protektivni učinak u usporedbi s oleuropeinom nakon 120 min predtretmana s ispitivanim tvarima u koncentracijama 1, 5 i 10 μM. Navedeni genoprotektivni učinci su posljedica snažnog antioksidativnog djelovanja ovih fenolnih spojeva utvrđenih ABTS, FRAP i CUPRAC metodom.Activity of oleuropein, a phenol secoiridoid present in olive tree products, its derivative hydroxytyrosol and water extract of olive leaf were investigated using in vitro tests against opportunistic fungal pathogen Candida albicans. Antifungal susceptibility testing was used to estimate the drop of viability up to 80 % in comparison to the control (untreated cells). Minimal inhibitory concentration (MIC) against C. albicans for oleuropein was 12,5 mg/ml, for hydroxytyrosol was 6,25 mg/ml and for water extract of olive leaf was 25 mg/ml. Morphological changes in the nuclei after staining with fluorescent DNA-binding dyes revealed that apoptosis was a primary mode of cell death in analyzed samples treated with sub-MIC concentrations of tested compounds. In order to understand mode of action of oleuropein and hydroxytyrosol, their effect on C. albicans virulence factors essential for development of infection in the host have been investigated. Both oleuropein and hydroxytyrosol modulate morphogenetic coversion and inhibit filamentation of C. albicans induced in the conditions of reduced nutrient availability. On the other hand, treatment with sub-MIC concentractions of these phenolic compounds did not affect C. albicans biofilm formation. The hydrophobicity assay showed that both compounds, in sub-MIC values, have decreased the cellular surface hydrophobicity, a factor associated with adhesion to epithelial cells. Cultivation of C. albicans in the presence of oleuropein and hydroxytyrosol also inhibits, in the dose-dependent manner, acitivity of aspartyl-proteases (Sap), hydrolytic enzymes important in adherence, tissue penetration, invasion and destruction of host tissue. Most therapies for fungal infections target the ergosterol biosynthesis pathway or its end product ergosterol which is necessary for growth and normal function of fungal cells. At subinhibitory concentrations tested compounds have altered sterol content and subsequently affected the cell membrane of C. albicans. Both compounds induced dose-dependent loss of intracellular material to outer space (DNA, protein leakage) and caused membrane depolarisation indicating their direct effect on the membrane. In addition, cytotoxicity and genotoxicity assays were performed to elucidate the effect of oleuropein and hydroxytyrosol on the host cells. Oleuropein and hydroxytyrosol exhibited no hemolytic activity on human erythrocytes at tested concentrations (3-60 μM). Antigenotoxic effects of oleuropein and hydroxytyrosol against H2O2-induced damage in human lymphocytes were evaluated in vitro by alkaline comet assay. Pretreatment of human lymphocytes with each of the substance for 120 min produced dose-dependent reduction of primary DNA damage in the tested cell type. At tested concentrations (1, 5, 10 μM), hydroxytyrosol showed better protective effect than oleuropein against H2O2-induced DNA breaks. Our results suggest that genoprotective effects of these phenolic compounds could be attributed to their radical scavenging and metal ions chelating activity which were determined using ABTS, FRAP and CUPRAC assays

Uloga i aktivnosti Sanitarne inspekcije

U obavljanju nadzora na poplavljenom području i području koje graniči sa poplavljenim područjem na području Vukovarsko-srijemske,Brodsko-posavske i dijelovima Požeško-slavonske županije bilo je uključeno 13 (trinaest) sanitarnih inspektora, od toga 10 (deset) sanitarnih inspektora iz PJ-Odjel za istočnu Hrvatsku, a 3 (tri) sanitarna inspektora iz drugih Odjela bila su u ispomoći. Po potrebi je bio organiziran rad u dvije smjene (preraspodjela radnog vremena). Sanitarna inspekcija obavila je 172 nadzor

Free planar vibration of structures composed of rigid bodies and elastic beam segments

Ovaj rad predstavlja analizu slobodnih vibracija struktura sastavljenih od krutih tela međusobno spojenih sa elastičnim gredama. Pretpostavlja se da se centri masa krutih tela ne nalaze na neutralnoj osi nedeformisanog elastičnog grednog segmenta kao i da kruta tela vrše ravno kretanje u istoj ravni i da se njihovi centri masa nalaze u toj istoj ravni. Za određivanje frekvencija sistema, modifikacija klasične 'CTMM' metode je upotrebljena. Elastični gredni segmenti se tretiraju kao Ojler-Bernulijeve grede. Prikazan je numerički primer.This article presents free vibration analysis of structures composed of rigid bodies connected with elastic beam segments. It is assumed that the mass centers of rigid bodies are not located on the neutral axes of undeformed elastic beam segments as well as rigid bodies perform planar motion in the same plane and their mass centers are located in that plane. For determination of natural frequencies of the system, modification of the conventional continuous-mass transfer matrix method has been performed. The elastic beam segments are treated as Euler-Bernoulli beams. Numerical example is presented

Protective effects of olive oil phenolics oleuropein and hydroxytyrosol against hydrogen peroxide-induced DNA damage in human peripheral lymphocytes

This study investigates antioxidant capacity and protective effects of phenolic compounds oleuropein (OLP) and hydroxytyrosol (HT), present in olive oil and olive leaves, against H2O2-induced DNA damage in human peripheral lymphocytes. Antioxidant potency was determined using the measurement of radical-scavenging activity (ABTS∙+ assay), ferric reducing power (FRAP assay) and cupric reducing antioxidant capacity (CUPRAC assay). Both substances were found to be potent antioxidant agents due to their free radical-scavenging activities. Antigenotoxic effects of oleuropein and hydroxytyrosol against H2O2-induced damage in human lymphocytes were evaluated in vitro by alkaline comet assay. At tested concentrations (1, 5, 10 µmol L–1), oleuropein and hydroxytyrosol did not induce a significant increase of primary DNA damage in comparison with the negative control. Pretreatment of human lymphocytes with each of the substances for 120 min produced a dose-dependent reduction of primary DNA damage in the tested cell type. Hydroxytyrosol showed a better protective effect against H2O2-induced DNA breaks than oleuropein which could be associated with their free radical-scavenging efficacy

Ispitivanje uticaja različitih adjuvanasa na imunogenost vakcine protiv virusa parainfluence 3 kod goveda

The objective of our research was to examine the influence of different adjuvants on immunogenicity of vaccine against parainfluenza 3 virus in cattle. In the research there was used the commercial vaccine Respi-ol against PI3 virus in cattle, produced by VZS from Subotica as well as the experimental vaccine against the same virus prepared with completed adjuvant Emulsigen. The research was performed on two experimental groups of 12 calves each. The animals were immunized twice, in the interval of 21 days. The third group of 6 calves was a control. The results of the research showed that a smaller amount of adjuvent Emulsigen per vaccine dose provided a satisfactory height of titre of virus neu­tralizing antibodies in the blood serum samples of vaccinated calves in regard to prescribed greater amount of standard oil adjuvant contained in commercial Respi-ol vaccine.Cilj naših istraživanja je bilo ispitivanje uticaja različitih adjuvanasa na imunogenost vakcine protiv parainfluence virusa 3 kod goveda. U ispitivanjima je korišćena komercijalna vakcina Respi-ol protiv virusa PI3 goveda, proizvođača VZS iz Subotice i eksperimentalna vakcina protiv navedenog virusa pripremljena sa gotovim adjuvansom Emulsigen-om. Ispitivanja su vršena na dve ogledne grupe teladi od po 12 životinja koje su dvokratno imunizovane u razmaku od 21 dan. Treća grupa teladi od ukupno 6 životinja je služila kao kontrola u ispitivanjima. Rezultati izvršenih ispitivanja su pokazali da je manja količina adjuvansa Emulsigen-a po dozi vakcine omogućavala postizanje zadovoljavajuće visine titra virus neutralizujućih antitela u uzorcima krvnog seruma vakcinisanih teladi u poređenju sa propisanom, većom količinom standardnog uljnog adjuvansa sadržanog u komercijalnoj vakcini Respi-ol

First report on the non-indigenous triclad girardia tigrina (Girard, 1850) (Tricladida, Dugesiidae) in Serbia, with Notes on its Ecology and Distribution

Girardia tigrina (Girard, 1850) (Turbellaria, Tricladida, Dugesiidae), a native freshwater triclad of North America, is a widespread species, which has been introduced by human activities into various parts of the world, including Europe. The aim of this paper is to contribute to the knowledge of this species by presenting its frst record and recent distribution in Serbia. Girardia tigrina was recorded in Serbia in 2008 in the lower stretch of the Kolubara River. After this initial fnding, the species was identifed only a few more times, but with low abundance. Its presence only in the northern part of Serbia, in the Danube River and in its proximity, points to the Danube River and its larger tributaries (the South invasive corridor) as the main route of its spread in the country. Our data suggest that G. tigrina should not be considered an invasive species but as an alien species with a minor impact on native communities

The Bologna reform of subject teacher education in the newly founded states in the territory of the former Yugoslavia

The article provides an overview of carrying out the principles of the Bologna reform in the education of subject teachers in the newly founded states in the territory of the former Yugoslavia. Drawing upon official documents, particularly laws and by-laws, study programmes and constituent documents of individual universities, the comparative analysis of the reform processes between 2004 and 2013 is made within a relatively homogeneous area in teacher education that existed before the break-up of the former joint state. Positive effects and weak points of the reform activities are observed and detected. The analysis has shown that by implementing the Bologna process the differences in the training of subject teachers among the states and universities, and even among individual universities, increased significantly compared to the previous state of education. This is evident not only in the simultaneous implementation of different models (i.e., the duration of studies (3+2, 4+1, 5+0), but also in concurrent application of simultaneous and successive forms of acquiring teacher competences, different academic titles, and particularly in the greatest issue - different levels of education at which teachers acquire teaching competences for the same teacher profile

The bologna reform of subject teacher education in the newly founded states in the territory of the former Yugoslavia

The article provides an overview of carrying out the principles of the Bologna reform in the education of subject teachers in the newly founded states in the territory of the former Yugoslavia. Drawing upon official documents, particularly laws and by-laws, study programmes and constituent documents of individual universities, the comparative analysis of the reform processes between 2004 and 2013 is made within a relatively homogeneous area in teacher education that existed before the break-up of the former joint state. Positive effects and weak points of the reform activities are observed and detected. The analysis has shown that by implementing the Bologna process the differences in the training of subject teachers among the states and universities, and even among individual universities, increased significantly compared to the previous state of education. This is evident not only in the simultaneous implementation of different models (i.e., the duration of studies (3+2, 4+1, 5+0), but also in concurrent application of simultaneous and successive forms of acquiring teacher competences, different academic titles, and particularly in the greatest issue different levels of education at which teachers acquire teaching competences for the same teacher profile

Olive leaf extract activity against Candida albicans and C. dubliniensis – the in vitro viability study

Olive leaf extract is characterized by a high content of polyphenols (oleuropein, hydroxytyrosol and their derivatives), which is associated with its therapeutic properties. The objective of the present research was to evaluate the antifungal activity of olive leaf aqueous extract against Candida albicans ATCC 10231 and C. dubliniensis CBS 7987 strains. Minimum inhibitory concentration (MIC) of the extract was determined by several in vitro assays. The extract showed a concentration dependent effects on the viability of C. albicans with MIC value of 46.875 mg mL–1 and C. dubliniensis with MIC value 62.5 mg mL–1. Most sensitive methods for testing the antifungal effect of the extract were the trypan blue exclusion method and fluorescent dye exclusion method while MIC could not be determined by the method according to the EUCAST recommendation suggesting that herbal preparations contain compounds that may interfere with this susceptibility testing. The fluorescent dye exclusion method was also used for the assessment of morphological changes in the nuclei of the treated cells. According to the obtained results, olive leaf extract is less effective against the tested strains than hydroxytyrosol, an olive plant constituent tested in our previous study