CARBOHYDRATE COMPOSITION AND ANTIOXIDANT ACTIVITY OF ROOT EXTRACTS OF INULA HELENIUM L.

СУ “СВ. КЛИМЕНТ ОХРИДСКИ”

Б И О Л О Г И Ч Е С К И Ф А К У Л Т Е Т

МЛАДЕЖКА НАУЧНА КОНФЕРЕНЦИЯ

“КЛИМЕНТОВИ ДНИ”

22-23 НОЕМВРИ, 2012 г.

СОФИЯ

Софийски университет “Св. Климент Охридски”

БИОЛОГИЧЕСКИ ФАКУЛТЕТ

Фонд „Научни изследвания” на СУ Научноизследователски сектор на СУ

Съюз на учените в България

МЛАДЕЖКА НАУЧНА КОНФЕРЕНЦИЯ

“КЛИМЕНТОВИ ДНИ”

22-23 НОЕМВРИ, 2012 г.

СБОРНИК РЕЗЮМЕТА

Книжка трета

СОФИЯ, 2012

Организационен комитет: Научен комитет:

Председател: доц. д-р Божидар Галуцов

Председател: доц. д-р Мариела Оджакова

Членове: проф. д-р Василий Голцев проф. д-р Венета Капчина гл. ас. д-р Яна Евстатиева гл. ас. д-р Кръстьо Димитров ас. д-р Любен Загорчев

Членове: проф. дбн Диана Петкова - СУБ чл. кор.проф. Здравко Лалчев, дбн чл. кор.проф. Румен Панков, дбн проф. дбн Яна Топалова проф. дбн Спасимир Тонков проф. д-р Пенка Мончева проф. д-р Христо Гагов

Секретари: гл. ас. д-р Диляна Николова Мариана Модрева

Секретари: гл. ас. д-р Иван Трайков гл. ас. д-р Анита Тошева

Уебсайт: гл. ас. д-р Любомир Кендеров

www.uni-sofia.bg http://klimentovi-dni.biofac.info

www.usb-bg.org

Младежката научна конференция “Климентови дни” се финансира от: Проект № 020/2012 г. “Фонд научни изследвания” на СУ Договор № 3022/2012 г. НИС на СУ ©Младежка научна конференция “Климентови дни”, 2012 г. СБОРНИК РЕЗЮМЕТА, книжка трета, българска първо издание ISSN: 1314–4960

МЛАДЕЖКА НАУЧНА КОНФЕРЕНЦИЯ “КЛИМЕНТОВИ ДНИ” 22-23, НОЕМВРИ 2012 3

ПРОГРАМА

22 НОЕМВРИ, ЧЕТВЪРТЪК

Час Сесия Аудитория

08.00 – 10.00 Регистрация фоайе, ІІ етаж

08.00 – 10.00 Монтиране на постерните табла ІІ етаж

09.00 – 9.30 Откриване на конференцията доц. д-р Мариела Оджакова, Декан на Биологическия факултет

239

Час Доклади на преподаватели от БФ

239 Председател – проф. дбн Диана Петкова Секретар – ас. д-р Любен Загорчев

09.30 – 10.00 Проф. д-р Пенка Мончева – “Бактерии – патогени по растенията” 239

10.00 – 10.30

Проф. д-р Гинка Генова – “Неврогенетика. Drosophila като модел за откриване на нови гени, свързани със синдрома на чупливата Х-хромозома”

239

10.30 – 11.00 КАФЕ ПАУЗА


239 Председател – проф. дбн Диана Петкова Секретар – ас. д-р Любен Загорчев

11.00 – 11.30

доц. д-р Георги Ас. Георгиев “Биофизикохимичният подход в изучаването на повърхностните свойства на слъзният филм: нов поглед към молекулните механизми на Синдромът на Сухото Око и към възможностите за терапия”

239

11.30 – 12.00

доц. д-р Иванка Цачева “Комплементният белтък C1q – функциониране на границата норма-патология”

239

12.00 – 12.30

доц. д-р Вяра Маматаркова “Уникалните свойства на биофилмите – основа за създаване и усъвършенстване на биопроцесни системи”

239

12.30 – 13.30 ОБЯД



239 Председател – проф. д-р Христо Гагов Секретар – гл. ас. Илияна Саздова

13.30 – 14.00 доц. д-р Петя Христова "Млечнокисели бактерии – за и против" 239

14.00 – 14.30 доц. д-р Ивелин Панчев “Биологични подходи за анализ на макромолекулни взаимодействия”

239

14.30 – 15.00 доц. д-р Светослав Димов „ГМО” – мръсна дума и/или спасение 239

15.00 – 15.30 КАФЕ ПАУЗА

15.30 – 17.30 ПОСТЕРНА СЕСИЯ фоайе, ІІ етаж

23 НОЕМВРИ, ПЕТЪК

Час Сесия Аудитория

09.00 – 10.30 ПОСТЕРНА СЕСИЯ фоайе, ІІ етаж

10.30 – 11.00 КАФЕ ПАУЗА

Час КРЪГЛА МАСА

„Кариерно развитие” 239 Председател – гл. ас. д-р Иван Трайков

11.00 – 12.30 Участват представители на институции и фирми, потребители на кадри, завършили Биологическия

факултет

12.30

Закриване на конференцията доц. д-р Мариела Оджакова, Декан на Биологическия факултет Награждаване „Най-добър постер”

239

13.00 ИГРАТА – ежегодна игра-викторина,

организирана от ФСС на БФ 239

14.30 Демонтиране на постерните табла


P1 MICROHABITAT SELECTION BETWEEN SEX AND AGE CLASSES IN LACERTA AGILIS CHERSONENSIS ANDREJOWSKI, 1832 (SQUAMATA: SAURIA: LACERTIDAE) ATANAS GROZDANOV1*, NIKOLAY TSANKOV2, STELIYANA POPOVA1 1 Department of Anthropology and Zoology, Faculty of Biology – Sofia University ”St. Kliment Ohridski”, blvd. Dragan Tsankov 8, 1164 Sofia; 2 National Museum of Natural History – BAS, blvd. Tsar Osvoboditel 1, 1000 Sofia * Corresponding author: [email protected]

Keywords: Lacerta agilis, microhabitats, ontogenetic shift

Habitat selection is an important tool for spatial segregation among sexes and age classes in lizards. The purpose of this study was to explore the microhabitat selection in both sexes of Lacerta agilis chersonensis at different ontogenetic stages in order to test the generalist/specialist hypothesis already stated to be valid for this species. Both sexes were separated in to groups according to their age class. A total 69 specimen, 18 adult and 16 juvenile females, 20 adult and 14 juvenile males have been studied. For every specimen common microhabitat conditions at 5 meter radius around the point location were stated. The next parameters in every polygon were counted: grass density in 4 classes, grass height in 3 classes, presence of country road, -trees, -shrubs, -stones or - artificial shelters. Multivariate analyses by mean of correspondent analyses well separate the four designated groups. According to data analyzed males were attributed to the microhabitats with shrubs and high/dense grasses. Females were affiliated to a lower high/dense grasses class and artificial shelters. A clear segregation among sexes at their juvenile stage was also demonstrated. Data are discussed in the light of the results obtained.

P2 XYLOGENESIS OF PINUS HELDREICHII AND PINUS PEUCE IN PIRIN MTS

ALBENA IVANOVA*, YANITSA TODOROVA, PEPA RANGELOVA AND MOMCHIL PANAYOTOV

University of Forestry, Dendrology Dept. Kliment Ohridski 10 Blvd., Sofia 1756, Bulgaria; * Corresponding author: [email protected] Keywords: tree rings, xylogenesis, Pinus heldreichii, Pirin

Pinus heldreichii and Pinus peuce are tree species characterized by

longevity. Previous studies have outlined the potential for constructing long tree ring chronologies which may be used for studying the climatic variation in the past. A


difficulty is the mixed climate signal with negative effect of summer droughts, but also of extremely cold summers and the positive effect of warm winters. Therefore successful dendroclimatic analysis requires better understanding of the processes of tree ring production.

Our aim within this project was to study the cambial activity, production of tracheids and their differentiation during the growth period. We chose 100-250 yrs. old Pinus heldreichii and Pinus peuce trees from natural forests in Pirin Mts. Beginning from 2010 we took microcores every 10 to 14 days, prepared samples with sliding microtom GSL-1, recorded them with digital camera and then analyzed the images to obtain information for the onset of cambial activity, the period of production of first cells, period until cells were produced, the moment of highest division rate and the differentiation of the cells. We found that in 2010 and 2011 the onset of cambial activity was delayed until the end of June - beginning of July. In 2012 it started at the beginning of June. The late start in 2010 and 2011 was probably due to cold periods in April and May. Despite this the trees produced tree rings with more rows of cells and higher width than all the rings in the period 2000-2009. This confirms our initial hypothesis that warmer than usual winters promote the production of wider tree rings. The production of new cells continued until the end of September, while the differentiation until the middle of October. In both years Pinus heldreichii trees produces more cells than Pinus peuce trees.

P3 CONTRIBUTION TO THE CHOROLOGY OF SOME AQUATIC PLANTS IN BULGARIA

TOSHEVA A. 1*, TRAYKOV I. 2

1 Sofia University "St. Klmient Ohridski", Faculty of Biology, Department of Botany, 2 Sofia University "St. Klmient Ohridski", Faculty of Biology, Department of Ecology and Environmental Protection *Corresponding author: [email protected]

Keywords: macrophytes, chorology, reservoirs, Bulgaria.

Despite its small territory Bulgaria has quite rich and diverse flora, which is due to the medial geographic location of the country, its varied landscape and its geological history. As a result, total of 4030 vascular plant species (mosses excluded) occur in the country, of which the aquatic plants account for up to 9%.

The aim of this study is to collect and analyze the data about the biodiversity of vascular plants in the water bodies. Up to now more than 100 standing water bodies (lakes and reservoirs) situated at different altitudes, from sea level to 2535m a.s.l. have been investigated. Sampling of macrophytes was carried out in the lakes between June and September 2009-2012.

In the course of the survey of macrophytes according to the aim of the project “Trophic state assessment and distribution of main monitoring groups –


phytoplankton and aquatic macrophytes in different types of limnetic water bodies from the ecoregions of Bulgaria” (MU-0114/2008), new chorological data for 9 species were found: Ceratophyllum demersum, Lemna gibba, Myriophyllum spicatum, Myriophyllum verticilatum, Najas marina, Potamogeton gramineus, Potamogeton pectinatus, Potamogeton perfoliatus and Typha laxmanii. Acknowledgment: The financial support through Project № DO 02-341/2008 provided by National Science Fund at the Ministry of Education, Youth and Science is gratefully acknowledged.

P4

DIURNAL ACTIVITY, FREQUENCY OF OCCURRENCE AND FEEDING CAPACITY IN DIFFERENT HABITATS OF THE COMMON PIPISTRELLE BAT PIPISTRELLUS PIPISTRELLUS (CHERBER, 1774)

DANIELA DIMITROVA, VENCESLAV DELOV*

Sofia University, Faculty of Biology, Dept. of Zoology and Anthropology, 1164 Sofia

Keywords: diurnal activity, habitats, Pipistrellus pipistrellus

Тhe common pipistrelle bat Pipistrellus pipistrellus is one of the most common and wide spread bat species in Bulgaria, especially in settlements. The present study was carried out in different habitats in Bulgaria during 2010-1012 period. The habitats included are urban habitats in city of Sofia, semi-natural habitats in some large city’s parks, villages and natural habitats around the upper stream of the river Maritza, costal forest habitats, steppe grassland habitats in Dobrudja plateau and wetlands. The specific equipment for field studies includes bat detector Peterson 240 X and recording devices Transcend MP860, stereo recorder Olympus with voice activation and voice detection functions, and laptop. The recording devices were used in different way, according to the specific purpose. The feeding potential of the specific habitat was estimated on the basis of the number of specific feeding calls per unit of time. The highest feeding potential has been estimated for wetlands. Diurnal activity show it has highest peak during the first half of the night between sunset and midnight. The frequency of occurrence in urban and wetland habitats reaches a few records per minute during this period. Urban habitats in Sofia and settlements along the upper stream of the river Maritza show the highest frequency of occurrence. In costal forest habitats the it is also comparatively high. Its lowest value was found in steppe habitats.


P5 HABITAT VARIABLES ASSOCIATED WITH WOLF (CANIS LUPUS L.) DISTRIBUTION AND ABUNDANCE IN BULGARIA

DIANA ZLATANOVA*, ELITZA POPOVA Faculty of Biology, Sofia University “St. Kliment Ohridski”, Dragan Tzankov 8, Sofia 1164, *Corresponding author: [email protected]

Keywords: habitat variables, wolf

The wolf is highly adaptable and at the same time a conflict species which demands for a better understanding of the variables associated with its presence. Based on data for 1323 wolf locations collected during the last 3 years and updated distribution in Bulgaria, we analyzed the main habitat factors influencing the distribution and abundance of the wolf in the country. The outcome of these analyses shows that although being mainly a forest species, the wolf does not avoid pastures and meadows and is highly tolerable to urban areas, although prefer areas with smaller settlements. The wolves also avoid altitude below 500 m (correlated with densely populated areas and bigger settlements) and above 2000 (above forest level) and show no significant preference to certain inclinations of the slope. The road density is also of no significant importance for the wolf distribution but there is a clear avoidance of the busy first and second class roads. The density of water areas is also of no significant influence for the wolf presence. The preybase (ungulate) density is one of the most important factors defining the wolf distribution and abundance as most of wolf the locations are collected in areas with ungulate biomass of 100-150 кг/км2. Indirectly the wolf is affected by the number of hunters per area mainly through the competition for the prey. Directly the species is affected only when the hunters density is above 3 hunters/km2. Highest wolf density is observed in areas with 1-1,5 hunters/км2 which is also correlated with high numbers of undulates.


P6 PRESENT DISTRIBUTION AND CONSERVATION SPECIFICS OF THE EGYPTIAN VULTURE (NEOPHRON PERCNOPTERUS LINNAEUS, 1758) IN SOUTHWEST BULGARIA E. STOYNOV1, GROZDANOV A.2, PESHEV H.1, PESHEV D.2

1 Fund for Wild Flora and Fauna, 49 “Ivan Mihaylov” Str, room 327, P.O.Box 78, 2700 Blagoevgrad 2 Faculty of Biology, Sofia University, 8 Dragan Tzankov blvd., 1164 Sofia, Bulgaria *Corresponding author: [email protected] Keywords: Egyptian vulture, Neophron percnopterus, distribution, conservation

The present research concentrates on the actual status of the Egyptian vulture (Neophron percnopterus) in Southwest Bulgaria and the factors for the species critical decrease in the last decade. Information for the extinction of the last breeding pair in the region was included. The last succesful breeding was documented in 2003. The research covered also observations during migration period and observations of non-breeding individuals. Conservation specifics for the species were presented in relation with the Griffon vulture reintroduction activities in the area. As a result of the regular artificial feeding, one immature individual was attracted and observed in Kresna gorge for a period of two months. The regular observations of the bird present the first success of species recent conservation, after the loss of all breeding pairs in the region. Supplemental feeding was also identified as a positive measure applied for the breeding pair in 2003. Two chicks were fledged succesfuly in the same year. Recommendations for future conservation activities were made.

P7 DISTRIBUTION OF CRAYFISH AND CRABS IN THE UPPER REACHES OF THE KAMCHIYA RIVER

E. STOYNOV1, PARVANOV D. 2, GROZDANOV A.2 1 Fund for Wild Flora and Fauna, 49 “Ivan Mihaylov” Str, room 327, P.O.Box 78, 2700 Blagoevgrad 2 Faculty of Biology, Sofia University, 1164 Sofia, Bulgaria *Corresponding author: [email protected] Keywords: Astacus astacus, Austropotamobius torrentium, Astacus leptodactylus, Potamon ibericum, mixed populations, Kamchiya River.

Twelve localities on the upper reaches of the Kamchiya River were sampled between 2004 and 2011 to determine the distribution and presence of the freshwater crayfish and crabs. In addition to the standard trapping techniques a new, highly effective crayfishing method developed by local people was applied.


Four indigenous species (noble crayfish - Astacus astacus, narrow-clawed crayfish - Astacus leptodactylus, stone crayfish - Austropotamobius torrentium and Potamon ibericum) were found in a relatively small area in Kotel Mountain and adjacent territories (Eastern Stara Planina Mountain, Bulgaria). Extremely rare in Europe mixed populations of A. torrentium and A. astacus were detected in Golyama Kamchiya and Luda Kamchiya Rivers. The ratio of A. torrentium to A. astacus in the total catch decreased considerably from habitats with large stones and high water velocity (2.31 ± 0.43) to sites with small stones and relatively slow velocity (0.96 ± 0.12) for both locations with observed mixed populations.

The obtained results are fundamental for the development of management strategies with the aim of better crayfish and crab habitat protection in the circumstances of increasing anthropogenic impact.

P8 DACTYLOGYRUS VOLFI LUCKY,1970 FOUND ON BARBUS TETRAZONA (BLEEKER,1855) IN BULGARIA

EMIL V. BORISOV e-mail: [email protected] Keywords: ornamental fishes, Monogenea, Dactylogyrus sp.

A new species for the helmithofauna of the ornamental fishes was found in Bulgaria. Ten specimens were examined. The parameters of invasion are given. Measurements are compared with those of the holotype, described by Lucky (1970).

P9 REPRESENTATIVES OF DACTYLOGYRIDAE FAMILY OF THE MONOGENEA CLASS IN GOLD FISH (CARASSIUS AURATUS AURATUS) IMPORTED IN BULGARIA FROM SINGAPORE

EMIL V. BORISOV e-mail: [email protected] Keywords: ornamental fishes, Monogenea, Dactylogyrus sp.

The introducing of new species in different parts of the world is the main reason for changing biodiversity.

A lot of ornamental fishes are bearers of exotic parasites, which spread in environmental waters in different ways (releasing, throwing out death fish in sewage). This gives possibility of their parasites to go in environmental waters of the country where they invade new hosts (except parasites there are viruses and bacterial agents spreading in this way). In this study we find 8 species of the genus


Dactylogyrus, which are typical for goldfish Carassius auratus auratus (Linnaeus, 1758), which is similar to the native for Bulgaria Carassius auratus gibelio (Linnaeus, 1758). Two of the dactylogyrids Dactylogyrus vastator (Nybelin, 1924), D. intermedius (Weger, 1910) are extremely pathogenic. Three of the others: D. dulkeiti (Bychwsky, 1936), D. inexpectatus (Gussev 1955), D. baueri (Gussev,1955) are novelty to the helminthofauna of the country.

P10 THE ECOLOGICAL AND FLORISTIC CHARACTERISTICS OF POPULATIONS OF SIDERITIS SCARDICA GRISEB. IN SLAVYANKA MOUNTAIN

INA ANEVA1*, PETER ZHELEV2, LJUBA EVSTATIEVA1, DIMITAR DIMITROV3

1Institute of Biodiversity and Ecosystem Research, BAS, 23, Acad. G. Bonchev Str., 1113 Sofia 2University of Forestry, 10 Kliment Ohridsky blvd., 1756 Sofia, Bulgaria 3National Museum of Natural History, 1 Tsar Osvoboditel Blvd *Corresponding author: [email protected] Keywords: flora, population, Sideritis scardica

Sideritis scardica Griseb. is an endangered species with threatened and extremely vulnerable populations. The current study provides information about the population state and structure, vegetation dynamics and the place of the species in habitats in all locations of Slavyanka Mountain. Three localities of the species are investigated – at the base of the peaks Golyam Tsarev Vrah, Shabran and Gotsev Vrah. A list of diversity of vascular plants is made for each locality. The taxonomic structure of the flora and the ecological and biological characteristics are defined.

P11 NOTES ON THESIUM BAVARUM AND T. LINOPHYLLON IN BULGARIA KIRIL VASSILEV1*, ANITA TOSHEVA2, TSVETA GANEVA2, HRISTO PEDASHENKO1, ANTOANETA PETROVA3, BORIS ASSYOV1 1Institute of Biodiversity and Ecosystem Research, Bulgarian Academy of Sciences 2Faculty of Biology, Sofia University “St. Kliment Ohridski” 3Botanical Garden, Bulgarian Academy of Sciences *Corresponding author: [email protected] Keywords: pollen, phytocoenology, Santalaceae, taxonomy

Thesium bavarum and T. linophyllon are two morphologically similar entities and their taxonomic treatment is still controversial, the decisions ranging from separation as two self-standing species, through recognition as infraspecific taxa and even synonymization of T. linophyllon to T. bavarum. In an attempt to


solve this problem, we have explored macromorphological characters, scanning electron microscopic study of pollen and peculiarities of the leaf cuticle. Phytocoenological data for the taxa are also presented.

P12 OVERVIEW ON DISTRIBUTION AND COMMUNITIES OF EDRAIANTHUS SERBICUS IN BULGARIA KIRIL VASSILEV1*, ASEN ASENOV2, IVA APOSTOLOVA1

1 Institute of Biodiversity and Ecosystem Research, Bulgarian Academy of Sciences, Acad G. Bonchev str., bl. 2,1113, Sofia, Bulgaria, 2 University of Sofia, Faculty of Biology, Department of Botany, 8 Dragan Tzankov blvd., Sofia 1164 *Corresponding author: [email protected] Keywords: Balkan endemic, dry grasslands, Festuco-Brometea, rare plant species

Edraianthus serbicus (A. Kern.) Petrovič is a Balkan endemic species located in western part of Bulgaria. The study provides new data and overview on the current knowledge about its distribution. The species is rare and grows in isolated populations exceptionally on calcareous terrains. Edraianthus serbicus takes part in xerophytic herbaceous vegetation of Saturejion montanae alliance. High abundance of Euro-Asiatic and sub-Mediterranean species is characteristic for the plant communities. Also many rare and endemic species take part in their species composition which determines their high conservation value.

P13 ABUNDANCE AND ACTIVITY OF SOIL ACTINOMYCETES FROM LIVINGSTON ISLAND, ANTARCTICA

MARTA ENCHEVA1, NADEZHDA ZAHARIEVA1, ANELIA KENAROVA1, NESHO CHIPEV2, VALENTINA CHIPEVA1, PETYA HRISTOVA1, ISKRA IVANOVA1, PENKA MONCHEVA1*

1Sofia University, Biological Faculty, 8 Dragan Tsankov Blvd., 1164 Sofia, Bulgaria 2Institute of Biodiversity and Ecosystem Research, 2 Yuri Gagarin Str., 1113 Sofia, Bulgaria *Corresponding author: [email protected] Keywords: Actinomycetes, antimicrobial activity, Biolog EcoPlates, PCR-amplification

The soils of Antarctica are severe environments inhabited by well adapted

microorganisms, the knowledge of which is scarce. The study on their abundance, diversity and physiology will provide the investigators with new data on the mechanisms of their adaptation and gives new opportunities to isolate microorganisms with unique properties for practical use.


The aim of the present work was to study bacterial abundance including actinomycetes and physiological diversity of Antarctic soils, by agar plate method and BIOLOG EcoPlates technique, respectively, and based on this actinomycete strains to be isolated, identified and screened for antimicrobial activity against different test bacteria.

The objectives of our investigation were seven Antarctic soils taken from moss, Deschampsia vegetated and fell field habitats of Livingston Island. Some of the soil physicochemical parameters, like pH, moisture and humus, were determined. Bacterial physiological activity and diversity were assessed based on the average color development of EcoPlate wells, and community capacity to utilize different chemical categories of carbon sources, respectively.

The results showed that soil bacteria from habitats with cryptogam (moss) coverage were characterized by highest physiological diversity and more intensive utilization of polyols, amino acids and phenolic compounds, and high abundance of actinomycetes. Most morphologically different actinomycete strains were isolated from the moss habitat denoted in the study as S6. The affiliation of the strains to genus Streptomyces was proved by the PCR amplification of 16S rDNA, using genus-specific primers. The screening of antimicrobial activity of the isolates by diffusion bioassay, using agar plugs showed that the strains synthesized antibacterial substances, active against both Gram-negative and Gram-positive bacteria. We conclude that the soils of moss habitats are favorable environments supporting high bacterial physiological activity, and high number and diversity of actinomycetes most of which are active producers of antibacterial metabolites.

P14

MACTOPHYTES MAPPING AND PHYSICOCHEMICAL PARAMETERS IN OGNYANOVO RESERVOIR

SAVCHOVSKA M.1, TOSHEVA A.2*, TRAYKOV I.3

1MSc. Student: Dep. of Botany, Faculty of Biology, Sofia University St.”Kl.Ohridski” 2Dep. of Botany, Faculty of Biology, Sofia University St.”Kl.Ohridski” 3Dep. of Ecology and EP, Faculty of Biology, Sofia University St.”Kl.Ohridski” * Corresponding author: [email protected]

Keywords: invasive species, macrophytes, reservoirs, spatial heterogeneity, WFD.

Longitudinal differences in physical and chemical parameters lead to a trophic gradient in the reservoirs. Marked differences in abiotic parameters and trophic response were observed between the riverine and the lacustrine part of Ognyanovo reservoir in relation to nutrients loading. Abundance and species composition of aquatic macrophytes were studied between 2009 and 2012. The observations at each sampling station are based on transect monitoring data. Strong influence of water level fluctuations and seasonal dynamics were observed, which emphasizes the problem of what the proper time is for macrophytes sampling in


reservoirs to provide an adequate definition of the ecological state in the future. The gathered data are used to produce a map of the macrophytes distribution in the reservoir. Rapid spread of the invasive species Elodea nuttallii is described after it appeared in the reservoir. A multiple point of spreading of E. nuttallii is observed in relation to fishermen and recreational activities. The E. nuttallii range expansion is assessed by its maximum linear rate of spread, with estimates of approximately 150 – 200 m yr−1, suggesting that invasion started around 7 to 10 years ago. Acknowledgment:The financial support through Project № DO 02-341/2008 provided by National Science Fund at the Ministry of Education, Youth and Science is gratefully acknowledged.

P15

STRUCTURE OF THE ZOOPLANKTON COMMUNITIES FROM SEVERAL WATER BODIES IN NATURA 2000 ZONES MIRA TISHEVA, DIMITAR KOZUHAROV*

Biological Faculty, Sofia University, Sofia, Bulgaria, 8 Dragan Tsankov Blvd., Sofia 1164, Bulgaria *Corresponding author: [email protected] Keywords: community structure, Rotifera, swamps, zooplankton

Zooplankton community structure in the standing water bodies can be affected by various ecological factors, including water chemistry, lake typology (such as depth and type of water inflowing) and human activity in the watersheds.

Examination of the zooplankton assemblages of vulnerable to the human impacts freshwater bodies was conducted during the period June 2009 - June 2012. All the studied water bodies are situated in the ecological frame Natura 2000 protected zones. The research was carried out on micro reservoir and four swamps. The swamps were strongly overgrown by macrophyte vegetation – over 70%. The studied objects are located in three zones of karstic area in western part of Bulgaria. The qualitative structure of the zooplanktonic communities was determined. As a whole 150 plankton components, that belong to different taxa ranks were identified such as genus, species and subspecies. There were found 5 new records for Bulgarian Rotifera fauna.

The most numerous were: Protozoa – 15 taxa, Rotifera – 103 taxa, Cladocera – 15 taxa and Copepoda – 9 taxa. There were also found in samples small number of Gastrotricha, Ostracoda, Harpacticoida organisms and Chironomid larvae. The zooplankton communities were characterized by a distinct richness of littoral - phytophilous species and a paucity of typical pelagic elements. Some bentho-plankton forms were found as well. Most of the registered zooplankton elements indacate the advanced eutrophication process in the studied water bodies.


P16 SCI “ZAPADNA STARA PLANINA I PREDBALKAN” - FLORISTIC STUDIES ON XEROTHERMIC OAK FORESTS

NADEZHDA GEORGIEVA*, KALINA PACHEDJIEVA, MARIYANA LYUBENOVA

Sofia University “St. Kliment Ohridski” – Faculty of Biology, Department of Ecology and Environmental Protection, 8 Dragan Tzankov Blvd, 1164 Sofia, Bulgaria *Corresponding author: [email protected] Keywords: flora, SCI, xerothermic oak forests

The main aim in the present study is to identify the floristic diversity of xerothermic oak forests with predomination of Quercus cerris L. and Quercus frainetto Ten. in SCI “Zapadna Stara planina i Predbalkan”. The inventory of the floristic complex resulted in more than 100 species of higher plants that belong to almost 80 genera and 40 families represented in the Bulgarian vascular flora. The biological specter of the xerothermic oak forests in the investigated region is made on the basis of Raunkiaer’s life forms. The specter shows the predominance of hemicryptophytes which takes an intermediate position between the specters of the forests in the temperately warm and temperately cold zone. The phytogeographical specter shows the predominance of European-Asian and sub-Mediterranean floral elements which are represented equally (about 20% each). The European, European-Mediterranean and sub-Boreal floral elements are also well represented (about 11% each), the anthropophytes is also discussed. The results of the present study will be a fundament of a further complex investigations of the xerothermic oak forests in the SCI and also will be useful for the defining of succession processes in the discussed forests.

P17 RELATIONSHIP BETWEEN TEMPERATURES AND PRECIPITATION AND PINUS NIGRA TREE RING WIDTH CHRONOLOGY FROM RHODOPA MTS VELISLAVA SHISHKOVA* AND MOMCHIL PANAYOTOV

University of Forestry, Dendrology Dept., Kliment Ohridski 10 Blvd., Sofia 1756, Bulgaria *Corresponding author: [email protected] Keywords: tree rings, dendroclimatology, Pinus nigra, Rhodopa Mts.

Previous studies of Austrian pine (Pinus nigra Arnold) from different parts of Europe outline the species as potentially valuable for dendroclimatic analysis. It is usually suitable for high quality precipitation reconstruction for periods without instrumental data. Despite this and the fact that in Bulgaria there are centuries-old Pinus nigra forests, local dendroclimatic studies are scarce.


We aim at studying the climate-growth relationship in tree-ring width chronology constructed from Pinus nigra trees from the region of Dobrostan in the Rhodopa Mts., Bulgaria. Using standard dendrochronological methods we constructed a chronology from 32 cores from 17 trees for the period 1741-2007. The statistical analysis shows high inter-series correlation (r = 0.645) and population signal (EPS) above 0.85 after 1850. We performed correlation analysis of the constructed chronology with the climate series from Sadovo city, which is close to the study site and one of the longest reliable series in Bulgaria with continuous measurements after 1893. Our results show high statistically significant negative correlation with the temperatures from the summer of growth and the summer prior to growth (R > 0.4). The relationship with precipitation amounts is high and positive (r > 0.5). Therefore we consider that there is clear and temporally stable drought signal in the constructed chronology and it may successfully be used to reconstruct precipitation for the period without instrumental data.

P18 INFLUENCE OF ENVIRONMENTAL FACTORS OVER MORPHOLOGY OF SNAKE EYED SKINK ABLEPHARUS KITAIBELII (BIBRON & BORY DE SAINT-VINCENT, 1833) (SQUAMATA: REPTILIA: SCINCIDAE)

VLADISLAV VERGILOV1*, NIKOLAY TZANKOV2

1 Department of Anthropology and Zoology, Faculty of Biology – Sofia University ”St. Kliment Ohridski”, blvd. Dragan Tsankov 8, 1164 Sofia; 2 National Museum of Natural History – BAS, blvd. Tsar Osvoboditel 1, 1000 Sofia *Corresponding author: [email protected] Keywords: Ablepharus kitaibelii, environmental factors, morphology

The snake-eyed skink (Ablepharus kitaibelii) is the only species of the Scincidae family in Bulgaria and also the northernmost species of this family on the continent. The relations between climatic and geographic parameters and morphological features, as a whole, have not been studied within the family. In this study material from the entire country (n=167) was included. In regards to the age and sexual-size dimorphism, only adult specimens were studied, and the dataset was separated by sex and includes 89 males and 78 females. Four meristic and eight metric traits were taken, and the metric ones were converted into 17 indices. Five climatic and geographic factors were also taken into account for each specimen. The data was tested for correlation between variables. Principal Component Analyses (PCA) was used for assessing the effect of environmental parameters over the species morphology. Strongest relation was found in regards to geographic latitude. The latitude correlates with metric and meristic signs including relative proportions, as well as with countable features of the body and head. The climatic zones also have an influence, yet to a smaller extent. According to the fact that these zones have a northwest-southeast orientation this factor partially overlaps and partly


corresponds to the geographic latitude. The main conclusion is that there is geographic intraspecific variability with expressed zonal character, and in contrast to the schemes suggested until now, which point out variability in north-south direction this study results suggest that the east-west directional variability seems to be more credible.

P19 BULGARIAN BLACK SEA GOBIES AS IMPORTANT OBJECT IN THE COMMERCIAL FISHING.

VELISLAV ZAREV*, APOSTOLOS APOSTOLU, BORIS VELKOV, MILEN VASSILEV

Institute of biodiversity and ecosystem research, Bulgarian Academy of Sciences, 1 Tsar Osvoboditel blvd., Sofia, Bulgaria, *Corresponding author: [email protected]

Keywords: Black Sea, family Gobiidae, gobies, industrial fishing

According to the official sources (National Agency of Fisheries and Aquaculture) gobies in the industrial fish catches take second place only after the sprat. In fact these catches are much higher, because the registered in the NAFA landings are less than 10% (unofficial data). The current survey covered most of the Bulgarian Black Sea coastline for the period 2009-2012. The study sites, equipment and techniques were chosen to be as close as possible to these of the local fishermen, observations on the local markets and interviews with fishermen were also carried out. 8 of the 24 native to Bulgaria goby species were presented in the sampling. The largest percentage has taken the round goby (Neogobius melanostomus), followed by the black goby (Gobius niger), the giant goby (Gobius cobitis) and the mushroom goby (Neogobius eurycephalus); smaller part was taken by the knout goby (Mesogobius batrachocephalus). The smaller quantities were established for the grass goby (Zosterisessor ophiocephalus), broad-headed (pinchuk’s) goby (Neogobius cephalargoides) and rock goby (Gobius paganellus).


P20 HABITAT PREFERENCES OF THE COMMON PIPISRELLE BAT PIPISTRELLUS PIPISTRELLUS (CHERBER, 1774) AND THE NATHUSIUS PIPISTRELLE BAT PIPISTRELLUS NATHUSII (KEYSERLING & BLASIUS, 1839) IN SOME BLACK SEA COSTAL AREAS DANIELA DIMITROVA, VENCESLAV DELOV* Sofia University, Faculty of Biology, Dept. of Zoology and Anthropology, 1164 Sofia Keywords: habitat, Pipistrellus nathusii, Pipistrellus pipistrellus.

The common pipisrelle bat Pipistrellus pipistrellus and the Nathusius pipistrelle bat Pipistrellus nathusii are the most numerous small bat species along Bulgarian Black Sea Coast. The aim of this study is to clarify habitat preferences and seasonal differences of occurrence in both species. This investigation was carried out during 2010-2012 period in the region of the northern Black Sea Coast of Bulgaria from Varna bay to Durankulak Lake. The research equipment includes bat detector Peterson 240 X and recording devices Transcend MP860, sound recorder Olympus with voice activation and voice detection functions, and laptop. The recording devices were used in different way, according to the specific purpose. The main habitats in the area are freshwater and salt-water wetlands, beach, costal forest habitats, agricultural lands, steppe grassland habitats, and settlements. The common pipisrelle bat and the Nathusius pipistrelle bat were found to be a dominating species in bat communities of costal forest habitats and settlements. In settlements and wetlands the first species prevails over the second one. The common pipisrelle bat forms remarkable concentrations around powerful artificial light sources. In costal forest habitats, the distribution of these species depends of the microhabitat specificity. The Nathusius pipistrelle bat predominates in costal forest near to the beach, up to 500 m from the sea, whereas the Common pipisrelle bat is most numerous in similar inland forest habitats. The both species showed clear seasonal differences in predomination. The Common pipisrelle bat predominates during spring and early summer whereas the Nathusius pipistrelle bat predominates during autumn.


P21 APPLICATION OF TWO TROPHIC INDICES BASED ON MACROZOOBENTHOS IN ECOLOGICAL ASSESSMENT OF POTENTIALLY REFERENT SITES OF BULGARIAN RIVERS MARIA KERAKOVA, MILA IHTIMANSKA, EMILIA VARADINOVA* Institute of Biodiversity and Ecosystem Research, Bulgarian Academy of Science Bulgaria, 1113 Sofia *Corresponding author: [email protected] Keywords: ecological state, macrozoobenthos, referent sites, trophic groups, trophic index,

Benhtosological data from 39 potentially referent sites situated on 33 Bulgarian rivers are used for this study. Functional feeding group distribution along the studied sites and the main indicative macroinvertebrate groups are analyzed. Similarities between species composition of the macrozoobenthos, as well functional feeding groups of investigated rivers are discussed. Current ecological state, based on German Feeding Type index (RETI/PETI) and Index of Trophic Completeness (ITC), of the studied sites was determined. Assessment was compared with the one made in the adapted version of Irish Biotic Index (BI), widely applied in Bulgarian benthosological practice.

Percentage distribution of the species in the main indicative groups demonstrates that species which belong to functional group shredders are most sensitive, while scrapers, predators, collectors and filtrators are relatively tolerant and deposit feeders are characterized as most tolerant. Cluster analysis results demonstrate higher similarity in species composition between different studied sites rather than similarity between different functional groups. Contrariwise the results of the cluster analysis based on trophic structure do not show clearly pronounced grouping of the river types, elevation or catchment areas. That result is due to the fact that insofar the trophic structure is more sensitive and characterizes the functioning of benthic community, at studied referent sites it is probably influenced mainly by hydromorphological and hydrological changes in water ecosystems.

According the values of RETI/PETI 19 sites are assessed in very good ecological state, 17 – in good and 3 – in moderate. In comparison, ITC evaluation shows worse results. The highest coincidence is observed between the BI and RETI/PETI results – at 19 sites. At 10 the assessment based on the two trophic indices coincide. Considerable discrepancy is observed between the results of BI and ITC, where at only 6 sites the categories (ecological quality ratio) are the same.


P22 SPECIES DIVERSITY IN RELATION TO HABITATS DIVERSITY

MIROSLAV SLAVCHEV1*, NIKOLAY TZANKOV2, GEORGI POPGEORGIEV3

1 Department of Anthropology and Zoology, Faculty of Biology – Sofia University 2 National Museum of Natural History – BAS, blvd. Tsar Osvoboditel 1, 1000 Sofia 3 Regional Natural History Museum, Hristo G. Danov 34 Str., 4000 Plovdiv *Corresponding author: [email protected] Keywords: amphibians, diversity, NATURA 2000, reptiles

NATURA 2000 is the biggest network of protected areas in European Union. Management Plan for every each zone is one of the most important documents. For it proper preparation detailed study of the subject plan territory is necessary. For a model object of this research a medium size NATURA 2000 protected area have been chosen – Pastrina BG0001037. Field data for five years period (2008-2012) have been analyzed. Data set includes 467 point locations of 24 amphibians and reptiles species. The study territory has been divided into 1x1 km grid. For every single cell of the grid we count the number of the species and the number of the habitats. For open and agricultural habitats data has been taken from the physical blocks. Forest data base have been used for the forest lands. Polygons with missing data have been filled with CORINE Land Cover. To estimate the habitat and species diversity Shannon Diversity Index has been used. Habitat diversity is slightly higher then species diversity, i.e. 1.451 (from 1.332 to 1.570) vs. 0.629 (from 0.420 to 0.838). It is noteworthy that habitat diversity does not correlate with species diversity. Species diversity is rather determined by presence of certain habitats that maintain high species diversity as a rule. Habitats as thermophiles delicious forests and nearby territories are the most important habitats with higher species diversity and with higher percent of species of conservation priority as Triturus cristatus, Bombina bombina, B. variegata, Eurotestudo hermanni. They cover only 11.7% of the territory of the protected area that make those habitats and affiliated species vulnerable and with uncertain future.


P23 CHANGES IN BIOLOGICAL AND PHYSICO-CHEMICAL PARAMETERS OF RIVER WATER IN A SMALL HYDROPOWER RESERVOIR CASCADE

MIHAILOVA P.1, TRAYKOV I.2*, TOSHEVA A.3, NACHEV M.4

1MSc. Student: Dep. of Ecology and EP, Faculty of Biology, Sofia University St.”Kl.Ohridski” 2Dep. of Ecology and EP, Faculty of Biology, Sofia University St.”Kl.Ohridski” 3Dep. of Botany, Faculty of Biology, Sofia University St.”Kl.Ohridski” 4Dep. of Aquatic Ecology, Faculty of Biology, University of Duisburg-essen *Corresponding author: [email protected] Keywords: chlorophyll-a, HPP, macrophytes, reservoir cascade, river water quality, run-of-the-river reservoirs.

The impact of single small hydropower plants on the environment is thought to be negligible. Their effect progressively increases with the cascade development of the river basins, thus the cumulative environmental effects of small HPP can have a significant impact on the ecological state of the rivers. We have studied the effect of two small HPP reservoirs on some parameters of river water quality. The Lakatnik Reservoir (run-of-the-river, mean depth 2.7 m, area 11.1 ha, mean residence time 0.2 days) and the Svrazhen Reservoir (run-of-the-river, mean depth 3.1 m, area 22.57 ha, mean residence time 0.44 days) are situated in the middle reaches of the Iskar river. The river water is rich in nutrients and organic substances and carries high amounts of suspended sediments. As it flows through the cascade, the river temperature and electrical conductivity increase, while oxygen concentration, pH and suspended sediments decrease, mostly due to the enhanced degradation and transformation of the organic substances in the water. The amount of the phytoplankton depends on the residence time and the amount of the abioseston in the river water. Despite their recent construction, the reservoirs support rich assembly of macrophytes, mostly indicators of eutrophic conditions. The observed changes in biological and physico-chemical parameters of river water are a direct result of increased retention of water in the reservoirs. The study shows the potential of small hydropower cascades to influence the river water quality. Acknowledgment: The financial support through Project № DO 02-341/2008 provided by National Science Fund at the Ministry of Education, Youth and Science is gratefully acknowledged.


P24 STUDY ON THE IMPACT OF GOOSE GRAZING ON CROP YIELDS IN THE AREA AROUND SHABLA AND DURANKULAK LAKES

PENCHO PANDYKOV1, ANTON STAMENOV2 AND NICKY PETKOV3* 1 Faculty of Biology of Sofia University “St. Kl. Ochridski”; 2 University of Forestry; ***Bulgarian Society for the Protection of Birds 3 c/o BSPB/BirdLife Bulgaria, PO Box 50, BG-1111 Sofia, Bulgaria, *Corresponding author: [email protected] Keywords: wintering geese, grazing impact, Shabla, Durankulak, Branta ruficollis, Anser albifrons, crop yield

We present results of one year study (2011/2012) on the impact of grazing

by wintering geese on the crop yield of wheat near Shabla and Durankulak lakes. The main wintering species are Anser albifrons and Branta ruficollis. The study design is based on open for grazing experimental plots and exclosure areas/control plots using a metal cage. The study sites were located between the lakes of Durankulak and Shabla. The exclosure were installed at the beginning of November 2011 -cages of 1x1m. Open experimental grazing plots of 1x1m were marked with 20cm wooden sticks, around each was marked 3x3m square where faeces were counted. Altogether six faeces counts were done between December and March. During harvest time the whole plants with stems were collected and removed from all plots. We collected the following measurements for each plot in the study: Number of wheat plants per unit area; Length of the ears; Length of the plants; Number of grains/seeds per ear; Weight of grain/seed.

From each plot randomly selected 100 plants were taken. The length of the ears and the stem of each of them were measured. Then from 60 of these plants we took the seeds/grains and counted them. From those seeds on a random base were selected 50 and measured their weight.

Result shower no significant differences between the crop yields in experimental and control plots. The goose grazing pressure showed positive correlation with number of plants and negative correlation with the grain weight. The grazing pressure did not had any significant correlation with the crop yield. Thus the sprout of more plants from one seed compensated for the lighter and fewer grains found in the grazed plots. We found statistically no significant results to support any impact of grazing by wintering geese on the overall wheat crop yield in the studied area.


P25 STRUCTURE OF OLD-GROWTH PINUS HELDREICHII FORESTS IN PIRIN MTS

PEPA TSVETANOVA AND MOMCHIL PANAYOTOV University of Forestry, Dendrology Dept. Kliment Ohridski 10 Blvd., Sofia 1756, Bulgaria *Corresponding author: [email protected] Keywords: old-growth forests, Pinus heldreichii, structure, Pirin

Bosnian pine (Pinus heldreichii Christ) is a tertiary relict and Balkan sub-

endemic species. Its communities are of high conservational value because they are extremely rich in rare and endemic plant and mushroom species. In Bulgaria, Pinus heldreichii forests are found only in the Pirin and Slavyanka Montains. Despite the necessity of better knowledge of the species and the forests, many questions are still poorly studied.

We aim at studying the structure and natural dynamics of old-growth Pinus heldreichii forests in Pirin. For the purpose we selected 4 regions in which we set 4 sample plots. Within them we measured the position in local coordinate system, DBH, height and crown dimensions of each tree. We extracted tree ring cores to estimate tree ages, study growth dynamics and the influence of external abiotic factors. We compared our results with the results from a previous study of younger forests in Pirin.

The studied old-growth forests are characterized by much smaller number of trees per 1 ha (42 to 304) than in younger forests (320 to 1777). There are trees with diameter above 100 cm. The percentage of trees with DbH>50 cm varies between 18 and 51%. In most of the plots there are clear signs of at least several fires. Regeneration is isolated mostly on dead wood in high classes of decomposition. Age of dominant trees is more than 250 years. Our data might be valuable in other ecological studies, that need to have better understanding of forest structure and history and might serve as a basis for definition of criteria for old-growth Pinus heldreichii forest.


P26 SHORT-TIME EFFECT OF HEAVY METAL STRESS ON KEY ENZYME INDICATORS IN RIVER SEDIMENTS

YOVANA TODOROVA, YANA TOPALOVA

Sofia University, Faculty of Biology, Department of General and Applied Hydrobiology, *Corresponding author: [email protected] Keywords: heavy metal stress, Phosphatase Activity Index, river sediments, Total Dehydrogenase Activity

The pollution of aquatic ecosystems made the necessity of new technologies for environmental decontamination more urgent. Bioremediation approaches involving microbial degradative capacities are regarded as effective and environmental friendly alternatives. As mediators in key transformation processes, the complex enzyme activities (measured as a total of extracellular and intracellular activity on organism and sub-organism level) have the potential to be used as bioremediation indicators for response of whole biological system and for the mechanisms and rate of detoxification processes. But the presence of some toxic and xenobiotic substrates can modulate (activate or inhibit) the enzyme activities and affects the degree of pollutants transformation. The typical examples for study of these complex effects in Bulgaria are the sediments in the middle part of Iskar River. The environmental state of river sector has been strongly influenced by the organics, nutrients, xenobiotics pollutants and by the intensive hydrotechnical activity for construction of 9 micro-hydro power plants. The change of hydrological regime is a prerequisite for intensive sediment concentration in areas of water inundation and uncontrollable deaccumulation/accumulation of hazard pollutants.

The main objective of this study was to assess the effect of heavy metals (Cu, Cd, Zn and Pb) on key enzyme indicators (total dehydrogenase activity – DHA and phosphatase activity index – PAI) in sediments of Iskar River middle part. The inoculation with moderate concentrations of cadmium stimulated the DHA but the further increasing inhibited the activity with 80%. The Cu2+ had a specific effect by the location of sediments. The Pb2+ and Zn2+ in all used concentrations inhibit the DHA with 35-95% and the registered effect was proportional to the ion concentration. The PAI was inhibited by all tested heavy metals but the Zn2+-ions have the higher impact (over 75% inhibition). The obtained results show some of the ways to assess the effect of heavy metal stress on enzyme indicators in aquatic environments and give a possibility to make accurate predictions of ecosystem response to future application of bioremediation technologies.


P27 EFFECT OF NANODIAMONDS ON TECHNOLOGICAL PARAMETERS OF MODEL BIPHASIC AZO-DETOXIFICATION PROCESS

MIHAELA V. BELOUHOVA*, IRINA D. SCHNEIDER, YANA I. TOPALOVA

Department of General and Applied Hydrobiology, Faculty of Biology, Sofia University *Corresponding author: [email protected]

Keywords: nanodiamonds, azo-detoxification process, sand biofilters.

Nanotechnologies are developing extremely fast by entering in diverse fields

of science, including medicine and biology. These innovative technologies are high technologies and reveal new potential of their integration in different areas of practice and especially in the working environmental technologies.

The effect of nanodiamonds (ND), which are products of the nanotechnologies, is almost unknown in the scientific studies of the biological systems, perspective for application in the environmental technologies – the functioning biofilms. In the present work the effect of ND on the technological parameters of azo-detoxification process has been studied. The model wastewater treatment process in two sand lab scale biofilters in semi continuous regime of functioning and model wastewater with increasing concentration of amaranth has been realized: biofilter with ND added in early phase – 266 h - (Biofilter EPh) and biofilter with ND added in late phase of the process – 485 h (Biofilter LPh). The general parameters for monitor of the process were: efficiency and rate of amaranth removal, COD /Chemical Oxygen Demand/ and TOC /Total Organic Carbon/. The obtained results showed that ND added in early phase stronger influenced the kinetic parameters of amaranth removal – increased with 4% efficiency of the detoxification, with 90% the rate of transformation and with 15% decreased of COD of the effluent. In the same time the adding of ND at the late phase increased weaker the key parameters of the amaranth detoxification - efficiency with 2% and rate of removal with 45 %. Summarizing these results we can conclude that the biofilms, functioning in early phase of detoxification are more reactive to the positive modulation effect of ND. The future investigations will be directed to the elucidation of the probable mechanisms of ND effect of the amaranth removing biofilms.


P28 INTERACTION OF CELLS WITH MODIFIED POLYETHYLENGLYCOL SURFACES

A. KOSTADINOVA1, N. ZAEKOV1, I. KERANOV2

1IBBME Acad.G.Bonchev 21,1113,Sofia,Bulgaria 2Centre de Recherche Public Henri Tudor, Department of Advanced Materials and Structures, Esch-sur-Alzette, Luxembourg Keywords: Tissue Engineering, cell attachment, biomaterials, PEG

The use of a polymer as a biomaterial, as material touching the tissue, blood,

proteins and other biological fluids inside or outside the human body requires solving serious problems biocompatibility. There are numerous opportunities for improving, of polymer surfaces, enabling the creation of uncharged, positively or negatively charged and contain various surface functional groups. The aim of this work is to fined opportunities to improve bio-compatibility of silicone rubber which pre grafted acrylic acid with different density and plasma treated surface with subsequent grafting PEG with different density and chain length. The maximum of cell adhesion and survival received on a surface with an average density of PEG graft.

P29 NITRIC OXIDE (NO·)-SCAVENGING ACTIVITY OF THE METHANOLIC EXTRACTS OF IN VITRO CULTURES OF SOME MEDICINAL PLANTS CHARACTERISTIC FOR THE BALKAN FLORA ALEKSANDAR MEHANDZHIYSKI1, DANIELA BATOVSKA1, DIMITAR DIMITROV2, LJUBA EVSTATIEVA3, KALINA DANOVA1

1 Department of Chemistry of Natural Compounds, Institute of Organic Chemistry with Centre of Phytochemistry, Bulgarian Academy of Sciences, Acad G. Bonchev bl. 9, Sofia 1113, Bulgaria 2 Botany Department, National Museum of Natural History, Bulgarian Academy of Sciences, 3 Institute for Biodiversity and Ecosystem Research, Bulgarian Academy of Sciences *Corresponding author: [email protected], [email protected], [email protected] Keywords: Clinopodium vulgare, Artemisia alba, Hypericum species, medicinal and aromatic plants biotechnology, NO• - scavenging activity

Here, we report a part of our continuing study on the biotechnological development of Balkan medicinal and aromatic plants some of which are poorly investigated. In this work, we obtained methanolic extracts from the aerial parts of in vitro cultured Clinopodium vulgare, Hypericum tetrapterum, two endemic representatives of the Hypericum genus – H. rumeliacum and H. richeri and Artemisia alba (aerial and root parts) and evaluated their NO• scavenging capacity. The extracts from the not well-studied H. rumeliacum, H. richeri and H. tetrapterum exhibited remarkably higher activity (0.18, 0.17 and 0.97 mg/ml, respectively)


compared to C. vulgare and A. alba (aerials and roots) (SC50 = 3.45, 2.93 and 2.62 mg/ml, respectively). Vitamin C was used as a reference compound, exhibiting NO•-scavenging activity of 0.26 mg/ml.

The presented results are indicative of the high therapeutic potential of the extracts derived from in vitro systems of the studied plants and are a good basis for further more detailed research.

P30 AMMONIUM VANADATE DECREASES VIABILITY AND PROLIFERATION OF CULTURED RETROVIRUS-TRANSFORMED CHICKEN HEPATOMA CELLS ABDULKADIR MAHDI ABUDALLEH1,2, TANYA ZHIVKOVA2, LORA DYAKOVA3, BOYKA ANDONOVA-LILOVA2, RADOSTINA ALEXANDROVA2*

1Faculty of Biology, Sofia University “St. Kliment Ohridski”, 8 Dragan Tsankov Blvd, Sofia, Bulgaria;

2Institute of Experimental Morphology, Pathology and Anthropology with Museum, Bulgarian Academy of Sciences, Acad. G. Bonchev Str., Bl. 25, Sofia 1113, Bulgaria; 3Institute of Neurobiology, Bulgarian Academy of Sciences, Acad. G. Bonchev Str., Bl. 23, Sofia 1113 *Corresponding author:[email protected] Keywords: ammonium vanadate, cell culture, chicken hepatoma, cytostatic/cytotoxic activity, myelocytomatosis virus.

Vanadium compounds have been reported to possess anti-neoplastic potential especially against chemically-induced liver, mammary, and colon carcinogenesis in rodents. According to the literature available, the anticancer activity of vanadium compounds in virus-transformed tumor cells has not been clarified yet. The aim of the study presented was to evaluate the effect of ammonium vanadate (NH4VO3) on viability and proliferation of cultured LSCC-SF-Mc29 cell line established from a transplantable chicken hepatoma, induced by the myelocytomatosis virus Mc29. The cells carry specific viral oncogene – v myc, and it is well known that myc (c-myc, L-myc, N-myc) proto oncogenes are involved in pathogenesis of > 80% of human malignancies. The investigations were performed by thiazolyl blue tetrazolium bromide (MTT) test, neutral red uptake cytotoxicity assay and double staining with acridine orange and propidium iodide. The results obtained revealed that applied at a concentration range of 0.1–20 µg/ml for 24-72h NH4VO3 expresses significant cytotoxic and/or cytostatic effects that are time- and concentration dependent. Acknowledgement: This study was supported by Grant № 24/05.04.2012, Sofia University “St. Kliment Ohridski” and European Social Fund and Republic of Bulgaria, Operational Programme “Development of Human Resources” 2007-2013, Grant № BG051PO001-3.3.06-0048 from 04.10.2012.


P31 SOLID PHASE EXTRACTION AND HPLC DETERMINATION OF PHLORIDZIN IN NATURAL PRODUCTS ANDREY MARCHEV1*, IVAN IVANOV2, RADKA VRANCHEVA3, ATANAS PAVLOV1,2

1Laboratory of Applied Biotechnologies Plovdiv, The Stephan Angeloff Institute of Microbiology, Bulgarian Academy of Science, 139 Ruski Blvd, 4000 Plovdiv, Bulgaria 2Department of Organic Chemistry, University of Food Technologies, 4002 Plovdiv, Bulgaria 3Department of Analytical Chemistry, University of Food Technologies, 4000 Plovdiv, Bulgaria *Corresponding author:[email protected] Keywords: amberlite XAD4, HPLC, phloridzin, solid phase extraction

The dihydrochalcone phloridzin is a natural product and presences as a dietary constituent in many fruit trees. For over 150 years this flavonoid attracts interests because of its pharmaceutical activities to decrease blood glucose levels and support diabetes treatment. This compound exhibits also antioxidant, anti-inflammatory, immunosuppressive, antimutagenic and antiobesity effects.

A reliable technique for routine analysis of phloridzin from natural products (apples and plums) combining solid phase extraction with Amberlite XAD4 and subsequent HPLC analysis was developed. The proposed methodology is less complex than other methods for flavonoid analysis from natural samples. The samples were extracted with acidified water (pH2) and than adsorption on resin XAD4 was carried out. To estimate the most selective eluent of phloridzin, organic solvent with different polarity and acidity were used. The highest recovery value of 90% was achieved when methanol was applied as eluent. With the developed HPLC method the analysis were performed for 20 minutes using Waters HPLC system with UV detection at 285 nm and Supelco Discovery HS C18 column (5 μm, 25 cm × 4.6 mm) operated at 26 °C. The mobile phase consisted of solvent A: 2% acetic acid and solvent B: acetonitrile under gradient flow rate. Under these operating conditions the method was precise and linear relationship was established in the range of 10-100 μg/mL and the coefficient of correlation (r2) was > 0.99. This technique was applied for the analysis of phloridzin in different samples of Malus “Golden delicious” and Prunus “Troyan blue”. The content of phloridzin in the different samples ranged between 14.0 and 24.0 μg/g of fruits.

The developed procedure for extraction and analysis of phloridzin is convenient and can be applied successfully to estimate this flavonoid in fruits and as a quality control during the production of fruit jams as well.


P32 VIABILITY AND PROLIFERATION OF MURINE BONE MARROW CELLS CULTURED IN THE PRESENCE OF NEWLY SYNTHESIZED COMPOSITE MATERIALS FOR BONE IMPLANTS BOYKA ANDONOVA-LILOVA1, TANYA ZHIVKOVA1, LORA DYAKOVA2, DIANA RABADJIEVA4, STEFKA TEPAVITCHAROVA4, RADOSTINA ALEXANDROVA1

1Institute of Experimental Morphology, Pathology and Anthropology with Museum, Bulgarian Academy of Sciences, 2Institute of Neurobiology, Bulgarian Academy of Sciences; 3Faculty of Biology, Sofia University “St. Kliment Ohridski”, 4Institute of General and Inorganic Chemistry, Bulgarian Academy of Sciences *Corresponding author:[email protected] Keywords: bone marrow cells, trypan blue dye, composite matrials, bone implants, biocompatibility

Calcium phosphate bioactive materials have been used in the medicine and dentistry due to their chemical and structural similarity with bone tissue mineral composition. The aim of our study was to evaluate the influence of three newly synthesized composite materials for bone implants on viability and proliferation of cultured murine bone marrow cells (BMC). BMC were obtained according to the standard procedure from femurs of ICR mice (2 months old) from both sexes. The cells were grown in 24 well/plates in the presence of the composite materials. The BMC cultured in non-modified medium served as a control. The investigation was performed after 72 h of treatment by trypan blue dye exclusion test using a Cell counter (Invitrogen). The percent of viable cells was found to vary between 46.67% and 67.16% (after 3 day treatment period) and to be 14.58% - 79.4% (on the 6th day) as compared to the control. Additional experiments are underway to clarify better the biocompatibility of the examined composite material as well as its osteoconductivity and osteoinductivity. Acknowledgement: Supported by Grant DTK-02-70/2009, National Science Fund, Bulgaria; and European Social Fund and Republic of Bulgaria, Operational Programme “Development of Human Resources” 2007-2013, Grant № BG051PO001-3.3.06-0048 from 04.10.2012.


P33 IDENTIFICATION OF SNP MUTATIONS IN MYBЕ-1 GENE INVOLVED IN DROUGHT STRESS TOLERANCE IN MAIZE BORISLAV ASSENOV1, VIOLETA ANDJELKOVIC2, DRAGANA IGNJATOVIC-MICIC2, JELENA VANCETOVIC2, ANA NIKOLIC2, NIKOLAI K. CHRISTOV1, STEFAN TSONEV1, NABIL ABU-MHADI1, DIMITAR VASSILEV1, YORDAN MUHOVSKI3, MIMA ILCHOVSKA4, ELENA TODOROVSKA1*

1AgroBioInstitute, 8 Dragan Tsankov blvd., 1164 Sofia, Bulgaria 2Maize Research Institute “Zemun Polje”, Slobodana Bajica 1, 11185 Zemun Polje, Belgrade, Serbia 3 Département Sciences du vivant Centre wallon de Recherches Agronomiques, Gembloux, Belgium 4Maize Research Institute, Kneja, Bulgaria *Corresponding author: [email protected] Keywords: maize, drought, MYBE-1 gene, SNPs

As one of the most important agricultural crops, maize is a staple diet for a large portion of the continuously growing world’s human population. Unfortunately, its production is severely affected by adverse environmental stresses such as drought, salt, low and high temperatures.The mechanisms of drought stress tolerance in maize are quite complex and involves a signaling network of genes that controls its adaptation to the stress. Recently, many transcription factors (TFs) for tolerance to drought stress have been identified in maize. In this study, specifically designed primers were used to identify functional single nucleotide polymorphisms (SNPs) in the MYB-like protein E1 (MYBE-1) gene of 13 tolerant and sensitive maize inbred lines from the gene banks of Maize Research Institute “Zemun Polje”, Serbia and Maize Research Institute, Kneja, Bulgaria by PCR amplification and direct sequencing. Multiple alignments of the sequenced fragments to the drought sensitive US maize inbred B73 and other inbred lines, representing the functional diversity of maize, from the Panzea database (http://www.panzea.org/) was performed. The sequence alignment revealed several SNPs in inbred lines object of this study, one of which unique to the drought tolerant Serbian inbred T3. In this line, the A→G transition, located in the third exon of the gene, leads to amino acid change from Tryptophan to Alanine at the Corresponding position of the protein. This replacement doesn’t affect the binding site of MYBE-1 transcription factor, but probably reflects on the spatial conformation trough inhibition of it’s functional activity.

This study will contribute the identification of functional markers in genes implicated in the development of drought stress tolerance and their further use for genomic improvement in the national maize selection programs.


P34 LACTIC ACID FERMENTATION WITH ENCAPSULATED LACTOBACILLUS CASEI SSP. RHAMNOSUS ATSS 11979 (NBIMCC 1013) IN ALGINATE/CHITOSAN MATRICES

BOGDAN GORANOV*, DENITZA BLAZHEVA, GEORGI KOSTOV, YODANKA GERMANOVA, ZAPRIANA DENKOVA

Department “Microbiology”, University of Food Technologies, Plovdiv, boul. "Maritza" № 26, *Corresponding author: [email protected]

Keywords: lactic acid bacteria immobilization, microcapsules, lactic acid

Microbial lactic acid production is one of the most studied processes. The

interest in this process is ascribed to the wide application of lactic acid as well as the lactobacilli producing it. The efficiency of the batch fermentation processes can be improved within limits by varying culture conditions and medium composition. The immobilized biocatalysts represent an attractive alternative for various fermentation processes. They have several advantages: high concentration of cells in the working volume of the bioreactor and hence high specific productivity, reducing the possibility of substrate and product inhibition. At the same time, they allow the transfer of the process into continuous mode with stable and reproducible control. An important requirement for implementing the technology with immobilized microbial cells is its complete characterization.

In the present work was investigated the lactic acid production with encapsulated lactic acid bacteria in alginate/chitosan complexes with solid and liquid core was investigated. It was studied two different alginate form – Ca and Ba, providing different mechanical stability and survival of microorganisms. The lactic acid production with immobilized cell was compared with the process with free lactic acid bacteria. pH and temperature optimum of the processes with free and immobilized cells were determined and a shifted temperature optimum of process with immobilized biocatalysts was observed. After having studied the compressibility of microcapsules, a biocatalyst with suitable mechanical profile for fermentation was chosen.

Batch fermentation carried out in a bioreactor with mechanical stirring. The dynamics of substrate uptake, accumulation of lactic acid, pH and reductive-oxidative potential of the fermenting medium were determined. The results will be used for the construction of fermentation system for continuous production of lactic acid with immobilized biocatalysts.


P35 CHANGES IN GLUTAMATE-CYSTEINE LIGASE (GCL) AND GLUTATHIONE PEROXIDASE (GPX4) EXPRESSION LEVELS IN 3T3-L1 CELLS AFTER EXPOSURE TO AGRIMONIA EUPATORIA L. EXTRACT

DIANA IVANOVA*, OSKAN TASINOV, DEYANA VANKOVA, YOANA KISELOVA-KANEVA Department of Biochemistry, Molecular Medicine and Nutrigenomics with Laboratory of Nutrigenomics, Functional Foods and Nutraceuticals, Medical University “Prof. Dr. P. Stoyanov”, 55 Marin Drinov Str., 9002 Varna, Bulgaria; *Corresponding author: [email protected] Keywords: Agrimonia eupatoria, gene expression, glutamate-cysteine ligase, glutathione peroxidase, 3T3-L1

Oxidative stress is implicated in a number of diseases and pathologic conditions and can result from excessive free radical production and/or insufficient cellular free radical antioxidant defense. Expression of antioxidant enzyme may be influenced by variety of substances, including antioxidants and plant polyphenols. Glutamate-cysteine ligase is the first and rate-limiting enzyme in glutathione biosynthesis. As an antioxidant enzyme glutathione peroxidase 4 is involved in reduction of organic hydroperoxides. The aim of the present study was to assess the expression levels of abovementioned enzymes in cell culture model upon treatment with Agrimonia eupatoria extract, previously established to be rich in polyphenols and to exert high antioxidant activity in vitro. As a cell culture model 3T3-L1 preadipocyte cell line was used. In order to determine the effect of the extract in standard conditions cells were incubated for 24h in phenol red free medium, containing Agrimonia eupatoria extract. To assess the effect of the extract in condition of oxidative stimulation it was applied as pretreatment for 24h, followed with 24h incubation with 100 µM tert-butyl-hydroperoxide. Gene expression level of the respective genes was determined with two-step Real-Time PCR. Relative quantification analysis was performed using 2-ΔΔCt method. Incubation of 3T3-L1 cells with Agrimonia eupatoria extract resulted in significant increase in mRNA levels of glutamate-cysteine ligase up to 240% (p<0.05). No no significant change in transcription level of glutathione peroxidase 4 was detected. tert-butyl-hydroperoxide significantly upregulated expression of both enzymes. Pretreatment of cells with Agrimonia eupatoria extract significantly reduced the stimulatory effect of the oxidizing agent on glutathione peroxidase 4 expression with 35% (p<0.001). Data from the current study show that Agrimonia eupatoria extract may influence cell behavior modulating expression of genes involved in the antioxidant cell defense and may exert protective activity against oxidative stimulation.


P36 EFFECT OF ABA PRETREATMENT ON CHLOROPLASTS STRUCTURE IN HYPERICUM RUMELIACUM BOISS. PLANTS SUBJECTED TO CRYOPRESERVATION DIMITRINA KOLEVA*, TSVETA GANEVA, MIROSLAVA STEFANOVA

Department of Botany, Faculty of Biology, Sofia University “St. Kliment Ohridski”, 8 Dragan Tsankov blvd., 1164 Sofia, Bulgaria Keywords: abscisic acid, chloroplasts, cryopreservation, Hypericum rumeliacum

Chloroplast ultrastructure in mature leaves’ cells of in vitro cultured Hypericum rumeliacum Boiss. plants regenerated from cryopreserved shoot tips pretreated with abscisic acid (ABA) for 3, 7 and 10 days periods respectively was analyzed by transmission electron microscopy (TEM). The chloroplasts in the control unfrozen plants were lens shaped with well developed peristromium and thylakoids. The chloroplasts in regenerants passed 3-days period of pretreatment with ABA were structured almost the same to the control ones while in regenerated after 7 days pretreatment plants, reduced volume of the thylakoids was observed. In 10 days pretreated regenerants TEM revealed round shaped chloroplasts and both granna and stromal thylakoids were destructed. The structural analysis of the plastid apparatus in regrown H. rumeliacum plants pointed out the 3-days ABA pretreatment as the most efficient for cryopreservation on subcellular level of organization. Acknowledgement: This work was supported by grant of the Bulgarian Ministry of Education Youth and Science (DNTC/BG-SK-01/2012) and from Slovak Research and Development Agency (APVV SK-BG 0012-10).


P37 INFLUENCE OF THE EXTRACTION CONDITIONS OVER THE ANTIOXIDANT ACTIVITYOF CARDUUS THOERMERI WEINM. WATER EXTRACTS

DASHA MIHAYLOVA1*, LIDIYA GEORGIEVA1, ILIYA ZHELEV2, IVANKA DIMITROVA-DYULGEROVA3

1Department of Biotechnology, University of Food Technologies, 26 Maritza Blvd., Plovdiv, Bulgaria 2Department of Pharmacognosy, Faculty of Pharmacy, Medical University of Varna, Varna, Bulgaria 3Department of Botany, Faculty of Biology, Plovdiv University”Paisii Hilendarski”, Plovdiv, Bulgaria

*Corresponding author: [email protected] Keywords: Carduus thoermeri Weinm., flower heads, antioxidant activity, water extracts

The influence of the extraction technique over the antioxidant properties of Carduus thoermeri Weinm. water extracts, expressed as Trolox equivalent antioxidant capacity (TEAC), was studied using DPPH (2,2-diphenil-1-picrylhydrazyl) scavenging, ABTS (2,2′-azino-bis-3- ethylbenzothiazoline-6-sulfonic acid) scavenging and reducing power assays. The phenolic concentration in the examined extracts, calculated as mg gallic acid equivalent (GAE)/g dry weight (DW), ranged from 7.65 ± 0.91 to 19.57 ± 1.06 mg GAE/g DW. The results from the total phenolics assay and the antioxidant activity tests were significantly correlated. Among the different C. thoermeri water extracts the decoct shows higher antioxidant activity and content of total phenolic compounds, which suggested that polyphenols were responsible for the antioxidant abilities.

P38 CHANGES IN RAT TESTIS AND SPERM COUNT AFTER ACUTE TREATMENT WITH SODIUM NITRITE

EKATERINA H. PAVLOVA*, DONIKA I. DIMOVA, EMILIA B. PETROVA, YORDANKA G. GLUHCHEVA, NINA N. ATANASSOVA

Institute of Experimental Morphology, Pathology and Anthropology with Museum, Bulgarian Academy of Sciences Acad. G. Bonchev Str., Bl. 25, 1113 Sofia, Bulgaria, *Corresponding author: [email protected]

Keywords: rat testis, sodium nitrite, sperm count

Background: Sodium nitrite (NaNO2) is an inorganic salt with various

industrial applications. The adverse health effects of NaNO2 in animals and humans are typically due to the formation of methemoglobin in the blood. This can lead to cyanosis and, at very high levels, death. Humans are constantly exposed to sodium nitrite through food and drinking water. The aim of the present study is to follow up the changes in rat testis and sperm count after acute treatment with sodium nitrite.


Methods: Four-month-old male Wistar rats were intraperitoneally injected with NaNO2 at dose of 50 mg/kg body weight (distilled water for controls). Treated animals were sacrificed at different time intervals (1h, 5h, 24h and 48h) following the administration. Testes and epididymides were sampled, weighed and embedded in paraffin using routine histological practice. Spermatozoa were isolated from both vasa deferentia and counted.

Results: Histological observations of the testis in all investigated groups demonstrated disorganization of seminiferous epithelium and assemblance of undifferentiated germ cells in the luminal area of the tubules. Testis weight/body weight index was increased in the first hours after administration which is probably due to higher seminal fluid volume. Statistically significant reduction in sperm count ranging between 48% (5h) and 28% (48h) was also observed. These results may be associated with impaired vascular permeability and water-soluble balance.

In conclusion, acute treatment with NaNO2 affects testis morphology, some weight indices and sperm count in mature rats.

Acknowledgements: This work is supported by a grant No DMU 03/18 for Young scientists from the Bulgarian National Science Fund.

P39 COMPARATIVE ANTI-HERPES VIRAL EFFECTS OF IN VITRO AND IN VIVO EXTRACTS DERIVED FROM LAMIUM ALBUM L. D. TODOROV1*, M. DIMITROVA2, K. KOSTOVA1, J. YORDANOVA2, V.KAPCHINA-TOTEVA2, S. SHISHKOV1

1Laboratory of Virology, Faculty of Biology, St. Kl. Ohridski Sofia University

2Department of Plant Physiology, Faculty of Biology, St. Kl. Ohridski Sofia University, *Corresponding author: [email protected]

Keywords: Antiherpes activity, Herpes Simplex Virus, Lamium album L.

Lamium album L. is valuable medicinal plant which possesses astringent, antispasmodic, anti-inflammatory, antibiotic and bacteriostatic properties. In our in vitro study, were tested for antiviral activity chloroform and methanol extracts, receiveded by Soxhlet extraction from in vivo and in vitro propagated plants. The extracts inhibited the replication of herpes simplex virus type 1 (HSV-1) and type 2 (HSV-2) in MDBK cells significantly without apparent cytotoxicity. The chloroform extracts shown strong antiviral effect. The 50% effective doses (EC50) of the chlorophorm in vitro extract was 552 μg/ml for HSV-1 and 487μg/ml for HSV-2. The EC50 of the chloroform in vivo extract was 668μg/ml and 780μg/ml respectively. The viral replications ware suppressed at 90% from the both chloroform extracts applied in maximal nontoxic concentrations. The test methanol extracts inhibited viral replications same mode, but EC50 values was higher then the chloroform extracts. The results suggested better inhibitory effects of the extracts,


obtained from wild plant then in vitro extracts. In addition, the chloroform in vivo extract shown strong virucidal effect. The extract applied in MNC inactivated the extracellular HSV-1 was above 90% (∆log1,5). This research is funded by contract №DTK-02-29/2009 of МEYS.

P40 ANTIBIOTIC COMPLEX, PRODUCED BY AN ANTARCTIC ACTINOMYCETE STRAIN S. ANULATUS 39 LBG09

DARINA DIMITROVA1, PETAR DORKOV2, BLAGOVESTA GOCHEVA3*

1Medical College “Y. Filaretova”, Medical University - Sofia, “Y. Filaretova” Str. No.3, Sofia, 2Biovet Ltd, R & D Directorate,"Peter Rakov” Str. 39, Peshtera, Bulgaria 3Faculty of Biology, Sofia University, 8 "Dragan Tzankov” Blvd, 1164 Sofia, Bulgaria; *Corresponding author: [email protected] Keywords: activity spectrum, antibiotic complex, component composition, screening

The strain Streptomyces anulatus 39 LBG09 is selected after screening of 17 actinomycete strains, isolated from soil examples of the isle Livingston – Antarctida, for producing antimicrobial substances. The strain synthesizes extracellular (86%) wide spectrum antibiotic complex both against Grampositive and Gramnegative bacteria, including phytopathogenic bacteria. For the purpose of isolation of the active substance, the mycelium is filtered and discarded. The obtained filtrates are extracted with different polarity organic solvents which aimed to determine the polarity of the active substance. The best for this purpose is etylacetate, followed by acetone and diethyl ether. This determines a complex mixture of several different polarity antibiotic substances. The obtained active extracts are subjected to Thin-Layer Chromatography analyse (TLC) to determine the polarity of the mobile phase for chromatographic separation column. Additional chromatography purification in Silica gel 60 column is made and afterwards a HPLC analyse is performed for determination of the component composition. It is proved that the antiobiotic complex is composed of three components: C1 (32.21%), C2 (13.21%) and C3 (18.33%). Preliminary quantitative analyse for functional groups identification shows that there are no polyene structures, spironolactones and macrolactone cycles in the composition of the three components. The antibiotic complex, synthesized from the strain S. anulatus 39 LBG09, is heat- and alkaline stable.


P41 SUBCHRONIC TOXICITY STUDY OF ETHANOL INFUSION FROM COTINUS COGGYGRIA WOOD IN RATS DIANA IVANOVA1, DANAIL PAVLOV1*, MIROSLAV EFTIMOV2, KALIN KALCHEV3, MILKA NASHAR1, MARIA TZANEVA4, STEFKA VALCHEVA-KUZMANOVA2 1Department of Biochemistry, Molecular Medicine and Nutrigenomics 2Department of Preclinical and Clinical Pharmacology and Toxicology 3Department of General and Clinical Pathology 4Department of Preclinical and Clinical Sciences Medical University “Prof. Dr. Paraskev Stoyanov” – Varna, Bulgaria Address: 55 Marin Drinov Str., 9002 Varna, Bulgaria; *Corresponding author:[email protected] Keywords: Biochemistry, European smoke tree, Histopathology, Phytopharmacology

The Eurasian smoke tree (Cotinus coggygria Scop., Anacardiaceae) is used by the Balkan folk medicine for its antiseptic and antimicrobial properties as well as for treatment of gingival and throat inflammations. Although that aqueous infusion from C. coggygria leaves has been applied mainly externally because of the large gallotannins content, there are few reports for an internal use of ethanol infusions from the wood against gastric ulcer and diarrhea.

The aim of this study was to examine the subchronic toxicity of ethanol infusion from C. coggygria wood in experimental animals. Three groups of 10 male Wistar rats were treated by stomach gavage as follows: (1) 20% ethanol infusion from wood, (2) 20% ethanol, (3) distilled water at doses of 10 ml/kg b.w. After 30 days of treatment the animals were sacrificed and the blood and organs were collected for biochemical and histopathological analyses.

Results showed that the treatment with ethanol infusion from C. coggygria wood did not cause subchronic toxicity on liver, kidney and brain. No treatment-related effects were identified for the general condition of rats and their body or liver weight. Histological investigation did not detect pathological deviations in organs of the treated group compared to both controls. No significant changes were observed in plasma levels of biochemical markers: hepatic enzymes (aspartate aminotransferase, alanine aminotransferase, and alkaline phosphatase), urea, uric acid, creatinine, triacylglycerols, malondialdehyde and thyols. No significant changes were observed in MDA and thyols in the liver, kidney and brain homogenates. Significant increase in uric acid levels in liver homogenates was registered and could be due to specific effects of bioflavonoids.

We concluded that the subchronic administration of ethanol infusion from C. coggygria wood is non-toxic in the applied concentration and therefore it can be used for further investigation of possible effects in animal models.


P42 ELECTROKINETIC POTENTIAL OF PEA THYLAKOID MEMBRANES UNDER STATIC MAGNETIC FIELD (2T) TREATMENT: ROLE OF IRON IONS AND LIGHT EXPOSURE ELITSA IVANOVA 1,4*, VIRJINIA DOLTCHINKOVA2, CHAVDAR ROUMENIN3

1 Institute for Molecular Biology “Acad Roumen Tsanev”, Department of Gene Regulations, Bulgarian Academy of Sciences, Acad. G. Bonchev Str., Block 21, 1113 Sofia, Bulgaria 2Department of Biophysics and Radiobiology; Faculty of Biology, Sofia University “ 3 Institute of Systems Engineering and Robotics (ISER), Bulgarian Academy of Sciences, Acad. 4 Katholieke Universiteit Leuven, Laboratory of Protein Phosphorylation, 3000 Leuven, Belgium * Corresponding author - [email protected] Keywords: Surface charge density; Static magnetic field; Light scattering; Thylakoid membranes; Lipid peroxidation

The effects of static magnetic field (SMF, 2T) on pea thylakoid membranes were investigated under photosynthetic and actinic light exposure. The pea plants were subjected to light intensity of 2000 lx (HL, high light) and to 650 lx (IL, intermediate light), respectively in order to specify the effect of SMF (2T) on the electrokinetic potentials at the membrane surfaces. We determined the electrophoretic mobility, zeta potential and surface charge density of thylakoids in dependence on the iron content of the ‘Fe-sufficient’, ‘Fe-starved’ and ‘2 x Fe’ variants. The generation of light-induced increase in negative zeta potential of ‘Fe-sufficient’ SMF (2T) pre-exposed thylakoids (HL, IL) was due to an enhancement of net negative surface electric charge on the outer surface of the membranes.

The primary electron-transport processes across the thylakoid membrane was increased due to actinic light treatment to the ‘Fe-starved’ SMF pre-exposed thylakoids (HL) because of most densely packed thylakoids.

A decreased lipid peroxidation by SMF (2T) exposure before or after actinic illumination of thylakoids was observed. It could be due to a recombination of radical products as a consequence of an indirect action of magnetic field on thylakoids from ‘Fe-sufficient’ and‘2 x Fe’ plants.

The present results provide information by which we could understand the nature of magnetic field energy storage and transfer and photochemical conversion of solar energy due to the electrokinetic approach. In the case of photosynthetic membrane system, the input is light and the response can be either a change in electron transport processes (∆LS) or change in the physico-chemical properties (electrokinetic potential).

The electrokinetic potential changes in pea thylakoids upon SMF (2T) exposure depend on Fe content in growing medium as well as light intensity (HL, high light; IL, intermediate light) during pea plant growing.


P43 ESTABLISHMENT AND CHARACTERIZATION OF STABLY TRANSFECTED MDCK CELL LINE, EXPRESSING HBEST1 PROTEIN

KIRILKA MLADENOVA1, VESELINA MOSKOVA-DOUMANOVA1, IRENA TABASHKA2, SVETLA PETROVA2, ZDRAVKO LALCHEV2, JORDAN DOUMANOV2*

1Department of Cytology, histology and embryology, Faculty of Biology, SU “St. Kl. Ohridski” 2Department of Biochemistry, Faculty of Biology, SU “St. Kl. Ohridski”, Sofia, 1164 *Corresponding author: [email protected] Keywords: Best1 protein, BVMD, MDCK II cells

Bestrophin-1(Best1) is a transmembrane protein specifically expressed in the retinal pigment epithelial cells. It is localized on their basolateral plasma membrane. Malfunctions of the protein lead to retinal pathologies, named Bestrophinophaties, including the Best Vitelliform Macular Dystrophy (BVMD). Best1 is thought to be a Ca2+- activated Cl- channel or a regulator of ion transport, or both. Some other functions in the maintaining of cellular homeostasis are also attributed to this protein.

In order to obtain additional information about protein structure and functions we established a stably transfected MDCK II cell line expressing human bestrophin-1, conjugated with Myc and His- tags at C terminus.

The presence of stably expressed Best1 protein was demonstrated by Western blot analyses and immunofluoresce studies. Confocal microscopy images of transfected MDCK cells confirmed its basolateral localization.

Additionally, no differences in the cell growth ratio and mitotic index of established cell line, in comparison with conventional MDCK cells were observed. Our studies suggest that Best1 does not influence cell growth and cell polarity in transfected cells, making our stably transected cell line an appropriate model for investigations of Best1 functions.

Acknowledgments: These studies were supported by Bulgarian Science Fund, grant № DDVU 02/10 and DO02/08.


P44 DROUGHT INDUCED DAMAGES OF PHOTOSYNTHETIC MACHINERY IN PHASEOLUS VULGARIS AND PLANTAGO MAJOR L. PLANTS ANALYSED BY SIMULTANEOUSLY RECORDED PROMPT AND DELAYED CHLOROPHYLL A FLUORESCENCE INDUCTIONS

VASSILENA KRASTEVA, VLADIMIR ALEXANDROV, MARIYA CHEPISHEVA, SHTERJAN DAMBOV, DETELIN STEFANOV, IVAN YORDANOV, VASILIJ GOLTSEV* St Kliment Ohridsky Sofia University, Department Biophysics and Radiobiology, Dragan Tsankov 8 Blvd. 1164 Sofia, Bulgaria *Corresponding author: [email protected]

Plant uses complex responses to protect against drought, including changes in gene expression, cell metabolism and whole plant physiology. Drought affects photosynthetic apparatus at different levels of organization. Using of new developed techniques for simultaneous registration of prompt and delayed fluorescence, give opportunity to achieve substantial progress in investigation of the drought effects on photosynthesis in intact leaves. The stress reaction of photosynthetic machinery is monitored in leaves of whole plants of Phaseolus vulgaris and Plantago major during 12-days cessation of irrigation. In opposite to drought sensitive bean plants, photosynthetic apparatus of plantain reveals earlier response to increasing water deficit. It was expressed by a visible decrease in fluorescence yield and suppression of photosynthetic efficiency. The shape of delayed fluorescence (DF) induction is found to be the most sensitive indicator for changes in photosynthetic electron transport in plants subjected to drought stress. As a parameter for estimation of drought stress effects could be considered the second maximum of sub-millisecond DF induction curve, while both microsecond and millisecond DF components did not change significantly. The changes of the second DF induction maximum could be interpreted as decreased rate of intersystem electron flow in tolerant Plantago plants. This suggestion is also supported by significant decrease in the rate of P700+ reduction that was monitored by light reflection at 820 nm. We proposed that intersystem electron transport suppression could be related to adjustment of electron transport chain for following switch to cyclic electron flow around PSI to secure photosynthetic electron transport with protection against drought induced damages.

Acknowledgments: Authors thank to St. Kliment Ohridski Sofia University Science Fund, Project № 021/05.04.2012 for the financial support.


P45 STUDY OF PROTEASES AND PROTEASE INHIBITORS FROM STREPTOMYCES STRAINS

KARAMFIL KALCHEV1, YAVOR RABADJIEV3, DIMITAR GANCHEV3, MARGARITA TSENOVA3, ILIYA ILIEV2, ISKRA IVANOVA3* 1 - Shoumen University ; 2 - Plovdiv University ; 3 - Sofia University Department of MIcrobiology, , 8, Dragan Tsankov, bld *Corresponding author: [email protected]

Keywords: Streptomyces; protease; protease inhibitors

Proteases and protease inhibitors are important enzymes in Streptomyces

physiology and differentiation. The objective of this paper was to investigate the optimal conditions for production of proteases and protease inhibitors secreted by three newly isolated Streptomyces strains. The optimal growth and inhibition activities were detected after cultivation on peptone media. The maximum of protease and protease inhibition activities were specific for the examined strains. The extracellular proteins were purified by ammonium sulfate precipitation, HPLC analysis and SDS-PAGE. One of the secreted proteases showed specific activity against beta-lactoglobulin.

P46 INFLUENCE OF COMPLEMENT ACTIVATION ON ESTABLISHED ZYMOSAN-INDUCED ARTHRITIS LYUDMILA BELENSKA1*, PETYA DIMITROVA2, NINA IVANOVSKA2

1Department of Biology, Medical Genetics and Microbiology, Faculty of Medicine, Sofia University 2 Department of Immunology, Institute of Microbiology, Sofia, Bulgaria *Corresponding author: [email protected] Keywords: complement, rheumatoid arthritis, zymosan-induced arthritis

Zymosan-induced arthritis is an experimental model of rheumatoid arthritis used for analyzing cells and molecules that mediate the pathogenesis of inflammatory joint diseases. Although, the alternative pathwayof complement is involved in the pathogenesis of arthritis much issues are not well elucidated. The aim of the present study was to evaluate the effect of complement depletion on cell populations that participate in the course of zymosan-induced arthritis. The decomplementation was performed using cobra venom factor, a peptide fragment of cobra C3 component which is capable of activating the alternative complement pathway. We established that CVF treatment affected proteoglycan loss and collagen synthesis dependent on time of injection. Absence of complement activity


differentially affected CD49+ and CD49/CD94+ cell populations in the synovium and blood. Also, the influx of synovial CD4+ T cells is ameliorated in CVF-treated mice, compared to untreated arthritis mice. Success of complement inhibition in the experimental models encourages novel therapeutic approaches to the treatment of human rheumatoid arthritis.

P47 MODULATION OF GCL AND GPX4 GENE EXPRESSION BY SAMBUCUS EBULUS L. FRUIT AQUEOUS INFUSION TASINOV O.*, KISELOVA-KANEVA Y., IVANOVA D. Department of Biochemistry, Molecular Medicine and Nutrigenomics, Medical University “Prof. Dr. P. Stoyanov”, 55 Marin Drinov Str., 9002 Varna, Bulgaria; *Corresponding author: [email protected] Keywords: antioxidant, GCL, GPx4, Sambucus ebulus, 3T3-L1 cells

Glutamate-cysteine ligase (GCL) and glutathione peroxidase 4 (GPx4) are

enzymes involved in cellular antioxidant defense. Their gene expression may be modulated by biological active compounds, such as polyphenols. Sambucus ebulus L. (SE) is a plant which fruit extract exhibits high antioxidant activity (AOA), due to its high polyphenol content. We analyzed the effect of SE fruit extract on GCL and GPx4 gene expression levels in the presence and in the absence of oxidizing agent tert-butyl hydroperoxide (tButOOH) in a 3T3-L1 cell culture model. Gene expression was analyzed performing Real Time quantitative polymerase chain reaction (RT-qPCR), and relative gene expression levels were calculated using 2-ΔΔ Ct method. Viability of SE extract treated preadipocytes was evaluated using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazoliumbromid (MTT) reduction assay. In the range of 1.25% to 12.5% content of the extract in medium, cell viability was not affected significantly. GCL mRNA levels were increased significantly (p<0.05) after treatment with SE fruit extract. Pretreatment with SE fruit aqueous extract decreased tButOOH induced gene expression of GCL (p<0.05) and of GPx4 (p<0.05). These data suggest that SE fruit extract exhibits strong AOA by modulating GCL and GPx4 gene expression. This study provides first scientific data about the AOA of SE fruit extract in a cell culture model.


P48 CHANGES IN THE ACTIN CYTOSKELETON AND ACTIN-ASSOCIATED PROTEINS IN HELA CELLS CO-CULTIVATED WITH E. COLI O157:Н7 T. TOPOUZOVA-HRISTOVA1, S. STOITSOVA2, T. PAUNOVA-KRUSTEVA2, B. ATANASOVA1, J.DOUMANOV3, E. STEPHANOVA1

1Department of Cytology, Histology and Embryology, Faculty of Biology, Sofia University, 2Department of Morphology of Microorganisms and Electron Microscopy, Institute of Microbiology, Bulgarian Academy of Sciences 3 Department of Biochemistry, Faculty of Biology, Sofia University Keywords: E. coli O157:Н7, HeLa, actin

Escherichia coli strains with the O157 LPS serotype may cause health disorders with variable severity. These bacteria are extracellular pathogens which have developed a specific mode to manipulate host cell cytoskeleton by means of a Type III secretion system. Most typically in polarized epithelial cells, the bacteria inject a translocated intimin receptor (Tir) into the host cells and thus initiate a signaling cascade that results in host cell-actin rearrangements. On the background of the very well analyzed F-actin rearrangements, little is known about the possible interference of E. coli O157 adherence with other cytoskeleton-related proteins and events in non-polarized host cells.

Here we describe data on changes of the actin-associated proteins ZO1 and villin during co-incubation of HeLa cells with a strain of E. coli O157:H7. Using laser scanning confocal microscopy our data showed unusual cytoskeletal actin rearrangement in HeLa cell line comparing to polarized epithelial cells. Additionally, behavior of actin-associated proteins ZO1 and villin, as well as lack of F-actin pedestals, suggested a different manner of bacterial pathogenesis in non-polarized epithelial cells. Acknowledgements. This study was supported by the National Research Fund, Republic of Bulgaria, Contract DO02-301/08.


P49 PUTATIVE DEGLYCATING ENZYME FROM EMBRYOGENIC SUSPENSION CULTURES OF D. GLOMERATA L.

EVA POPOVA1, ROUMYANA MIRONOVA2, MARIELA ODJAKOVA1*

1Department of Biochemistry, Faculty of Biology, Sofia University, 1164, Sofia, Bulgaria 2Insitute of Molecular Biology, Bulgarian Academic of Science *Corresponding author: [email protected] Keywords: amadoriase, glycation, plants

Glycation is a major cause of spontaneous damage to proteins and other biological amines in physiological systems. This process includes a series of reactions leading to formation of early, intermediate and advanced glycation end products. To date, three groups of enzymes capable of catalyzing the degradation of early glycation products have been discovered and their presence was demonstrated in animals, many bacterial species, and plants (Arabidopsis thaliana and Spinacia oleracea).

The aim of the present study was to investigate the monocot plant, Dactylis glomerata for the presence of putative deglycating enzymes. We used human single fold scFv libraries Tomlinson I&J to produce monoclonal antibodies against amadoriase from Escherichia coli. 80 clones were selected against the target antigen and tested for their ability to recognize it by ELISA and Western blot analysis. To detect the presence of a putative deglycating enzyme in our plant model system, different protein fractions from embryogenic suspension cultures of D. glomerata were isolated and separated by SDS PAGE. The Western blotting was performed with a monoclonal antibody that recognize the E. coli amadoriase with high affinity confirmed by ELISA. The soluble antibody was shown to recognize a single protein band from a total cell wall protein with apparent Mw of 37 kDa which might be a deglycating enzyme from D. glomerata. The protein fraction was purified against scFv from the high affinity clone and will be send for custom sequencing by mass spectrometry.


P50 ELECTROPHORETIC CHARACTERIZATION OF MILK PROTEINS FROM BULGARIAN RHODOPEAN CATTLE – APPLICATION IN SELECTION AND BIODIVERSITY PRESERVATION LYUBEN ZAGORCHEV1, MARIA DIMITROVA1, MARIELA ODJAKOVA1, DENITSA TEOFANOVA2, PETER HRISTOV2

1Department of Biochemistry, Faculty of Biology, Sofia University1164, Sofia, Bulgaria 2Institute of Biodiversity and Ecosystem Research at the Bulgarian Academy of Sciences, *Corresponding author: [email protected] Keywords: 2D electrophoresis, β-lactoglobulin, milk proteins polymorphism

αs1-casein, κ-casein and β-lactoglobulin show comparatively the highest

polymorphism among the milk cattle proteins. Therefore they have been extensively investigated in respect to identify the correlation between this polymorphism and the qualitative and quantitative milk traits. It is possible, based on the variability of milk proteins, to clarify the phylogenetic relationships between different cattle breeds and to gather the necessary data to preserve the genetic fund and biodiversity of the local breeds. In the present study an optimized 2D electrophoretic method for milk protein profiling is used to determine the variation in the isoforms of αs1-casein, κ-casein and β-lactoglobulin in the Bulgarian Rhodopean cattle. Also the possible relation between this polymorphism and the qualitative features of milk is discussed. Several different genotypes for each of the proteins were established in the investigated population. The method proposed here is a highly sensitive system for milk protein investigation. It could be used to determine the phylogenetic origin of a given cattle breed but also to maintain a highly-productive selection and preservation strategy. In addition the electrophoretic method could be successfully applied to industry in respect of quality control of commercial milk products. Acknowledgments: This study was supported by grant YRG 02/23 28.12.2009 from the National Science Fund of the Bulgarian Ministry of Education, Youth and Science, Sofia, Bulgaria.


P51 GLUTATHIONE REDUCTASE ACTIVITY DURING SOMATIC EMBRYOS FORMATION IN SUSPENSION CULTURES OF DACTYLIS GLOMERATA, TREATED WITH DIFFERENT NaCl CONCENTRATIONS

LYUBEN ZAGORCHEV, MIROSLAVA TERZIEVA, DIMITUR DIMITROV, MARIELA ODJAKOVA*

Department of Biochemistry, Faculty of Biology, Sofia University,. 1164, Sofia, Bulgaria *Corresponding author: [email protected] Keywords: glutathione, glutathione reductase, salt stress, somatic embryogenesis

The concentration of the tripeptide glutathione (GSH; γ-L-glutamyl-L-cysteinyl-glycine) and its Corresponding disulphide, GSSG is of great importance to the ability of plant cells to overcome abiotic stress. GSH plays a central role in ROS scavenging and the GSH/GSSG ratio is maintained by glutathione reductase (GR, EC 1.6.4.2). The activity of GR was investigated in suspension cultures of orchardgrass (Dactylis glomerata), subjected to different salt stress levels. It was found that high GSH/GSSG ratio is associated with accumulation of fresh mass in the initial days of the cultivation period while lower GSH/GSSG ratio is associated with the formation of somatic embryos both in controls and cultures, subjected to low salt concentration (85 mM NaCl). Correspondingly the GR activity was higher during the initial stages and decreased during the formation of somatic embryos thus providing a favorable intracellular environment for the different developmental stages. In cultures, treated with higher salt concentrations (170 mM NaCl) the GR activity remains comparatively lower during the whole cultivation period that results in unfavorably low GSH/GSSG ratio. At this salt concentration both fresh mass accumulation and somatic embryogenesis rate are significantly lower than in controls. A single GR band with apparent molecular weight of 60 kDa was found in all cultures by immunoblot analyses but it was presented in lower amounts in cultures, treated with 170 mM NaCl. Our results are in good agreement with the general understanding that more reducing conditions (high GSH/GSSG ratio) are required for cell proliferation and more oxidizing conditions (low GSH/GSSG ratio) are required for cell differentiation. Besides the regulation of the GSH synthetic pathway GR is also an important regulator of the GSH turn-over and the inability of the high stressed-cultures to maintain its activity leads to inhibition of both growth and somatic embryogenesis.


P52 CHANGES IN ARABINOGALACTAN PROTEINS DURING SOMATIC EMBRYOGENESIS IN SUSPENSION IN VITRO CULTURES OF DACTYLIS GLOMERATA L.

LYUBEN ZAGORCHEV, RADOST STOINEVA, MARIELA ODJAKOVA*

Department of Biochemistry, Faculty of Biology, Sofia UniversitySofia 1164, Bulgaria *Corresponding author: [email protected] Keywords:arabinogalactan proteins, salt stress, somatic embryogenesis, Yariv-reagent

Somatic embryogenesis is a remarkable process in plant cells in which somatic embryos are formed from a single somatic cell that could further regenerate into a whole plant. Arabinogalactan proteins (AGPs), a family of heavily glycosylated cell surface glycoproteins, are among the key regulators of somatic embryogenesis. The aim of the present study is to investigate the expression and localization of AGPs in embryogenic suspension cultures of Dactylis glomerata L., treated with NaCl. We have used two concentrations of NaCl - 85 mM, which stimulates somatic embryogenesis and 170 mM, which inhibits the process. The addition of (β-D-glucosyl)3 Yariv reagent to the growth medium, a synthetic phenylglycoside that specifically reacts with AGPs, inhibited both fresh mass accumulation and somatic embryogenesis. A lot of cells with abnormal shape were observed in cultures with Yariv reagent compared to controls showing the importance of AGPs for the development of plant cells. Salt stress showed differential effect on distribution of AGPs, recognized by LM2 and MAC207 antibodies as shown by immunoblot and immunofluorescence analyses. While LM2 antibody recognizes a glucuronic acid – containing epitop, MAC207 is directed against L-arabinose containing epitop. MAC207-recognized AGPs are found predominantly in the intracellular fraction in controls and migrates to the cell wall in salt-treated cultures. LM2-recognized AGPs almost disappeared from the cell wall at 170 mM NaCl. The pattern of AGPs is differing according to the applied salt stress and at least some of them could be associated with the process of somatic embryogenesis.


P53 GROWTH CHARACTERISTIC OF LACTOBACILLUS DELBRUECKII SPP. BULGARICUS 286 IN CORRELATION WITH LACTIC ACID PRODUCTION V. KALICHUK, G. VELIKOVA, S. G. DIMOV Sofia University "St. Kliment Ohridski", Faculty of Biology, 1164 – Sofia, Bulgaria *Corresponding author: [email protected]

Homofermentative lactic acid bacteria from genus Lactobacillus are well-

recognized producers of lactic acid, a primary product which takes part in multifarious manufactories. Also, the quantity of the synthesized lactic acid plays an essential part in the product properties. For the purpose of gaining an understanding for the industrial perspective of the strain Lactobacillus delbrueckii spp. bulgaricus 286, its production of lactic acid and growth characteristics were evaluated by conducting cultivation experiments. It was cultivated in MRS broth with different acidity, and its growth and the lactic acid production were monitored. In the light of the obtained data, the analysis represented a correlation between pH of the growth medium and the behavior of the bacterial population (in both the amount synthesized lactic acid and the measured biomass) at the time of observation.

P54 IN VIVO ANTIOXIDANT ACTIVITY EVALUATION OF PEPTIDES PRODUCED DURING THE FERMENTATION OF YOGHOURT-LIKE DAIRY PRODUCTS G. CHIKOV, V. ALEKSANDROVA, M. DIMITROV AND S. G. DIMOV

Sofia University "St. Kliment Ohridski", Faculty of Biology, 1164 – Sofia, Bulgaria *Corresponding author: [email protected]

A collection of few dozens Lactobacillus strains belonging to different species was previously screened for proteolytic activity by polyacrylamide gel electrophoresis during cultivation in skim milk (data not present). Four Lb. delbrueckii strains showing good proteolytic activity and belonging to the subspecies bulgaricus and lactis were selected, and used in combination with the commercial yoghourt starter LBB BY D4 (“LB Bulgaricum” PLC, Sofia, Bulgaria) for the production of yoghourt-like dairy products which were further analyzed for in vivo antioxidant activity caused by the peptides obtained during the fermentation of the milk proteins. It was found that the concentration of the reactive oxygen species (ROS) within living yeast cells (used as indicator for the test) was lower when treated with samples obtained by the combined starters in comparison with the commercial starter only. Differences between the strains were also observed, thus


suggesting that different strains possess different levels of probiotic antioxidant properties in vivo.

P55 ENCAPSULATION OF BREWING YEAST IN ALGINATE/CHITOSAN MATRIX: COMPARATIVE STUDY OF BEER FERMENTATION WITH IMMOBILIZED AND FREE CELLS

VESELA NAYDENOVA*, STOYAN VASSILEV, MARIA KANEVA, GEORGI KOSTOV

University of Food Technology, PlovdivDepartment “Technology of wine and beer”, University of Food Technology, Plovdiv, 26 Maritza blvd., *Corresponding author: [email protected]

Keywords: brewing, flavor profile, immobilized yeast

Beer production with immobilized cells has been extensively investigated over the last 30 years. The main advantages of immobilized cell technology (ICT) are: enhanced fermentation productivity due to higher biomass densities, possibility for using in continuous processes and improved cell stability. Nowadays, the application of ICT in main fermentation is on laboratory and pilot scale. On the other hand, immobilized cells are used on industrial scale for production of alcohol-free and low-alcohol beer or maturation of green beer.

In the present study, possibilities for beer production with lager brewing yeast, encapsulated in alginate/chitosan matrix with an internal liquid core were investigated. Batch fermentations with immobilized and free cells were carried out at 15°C. The dynamics of extract, alcohol and color of beer were studied at different initial wort extracts. Simultaneously, the dynamics of substances, forming the beer flavor profile (esters, aldehydes and higher alcohols) were investigated. As a result, significant differences between fermentations with free and immobilized cells were observed.

Kinetic parameters of development of the microbial population were determined on the base of the data for the dynamics of fermentations. Immobilized cells showed lower specific growth rates but they indicated higher specific rates of accumulation of ethanol.

The obtained data on the dynamics of fermentation processes and the accumulation of metabolites will be used to develop a continuous system for beer production with immobilized cells, because microcapsules showed high mechanical stability and possibility for repeated use in alcoholic fermentations.


P56 IN VITRO ASSESSMENT OF PREBIOTIC UTILIZATION BY DAIRY AND VAGINAL LACTOBACILLI ROSITSA TROPCHEVA1*, SILVIA DIMITONOVA1, RALITSA GEORGIEVA1, HRISTOVA JANA1, AURORA SALAGEANU2, DIONYSSIOS N. SGOURAS3 AND SVETLA DANOVA1 1-Department of Microbial Genetics, The Stephan Angeloff Institute of Microbiology, Bulgarian Academy of Sciences, 26, Acad. Georgi Bontchev str., 1113- Sofia, Bulgaria; 2- Infection and Immunity Laboratory, Cantacuzino Institute 103, Spl. Independentei, Sector 5050096, Bucharest, Romania; 3- Laboratory of Medical Microbiology, Hellenic Pasteur Institute, Athens, Greece. *Corresponding author: [email protected] Keywords: “katаk”, prebiotics, vaginal and dairy lactobacilli

The new consumers’ demand for healthy products on the market enhances intensive researches on lactic acid bacteria (LAB) and prebiotics. Individually and in combinations (in a synbiotic form), they can be effective microbial modulators, with a positive impact on human health and wellbeing. Thus, the selection of successful combinations LAB strain-prebiotic is a milestone in the development of new functional foods and products. With this aim, the potential of 54 Bulgarian LAB strains to growth in the presence of galactoologosacharide (GOS), glucooligosacharide (GLOS) and fructooligosacharide (FOS) were estimated. Twenty one vaginal lactobacilli and 33 original LAB strains from traditional fermented dairy products (yogurt, cheese and “katak”) have been identified and pre-selected on the base of widely accepted in vitro criteria for probiotics. The in vitro assessment of prebiotic’s utilization was evaluated in a mini-plate model system with a modified MRS broth. The major part of tested LAB was able to growth in medium containing 10% w/v GLOS (80-93% from the tested strain) and 10% w/v GOS (75-91%) as a single carbon source. However, both – vaginal and dairy lactobacilli do not metabolize the FOS. A higher metabolic activity has been proved for LAB strains belonging to the species Lactobacillus plantarum, Lactobacillus brevis and Lactobacillus fermentum. A strict correlation has been reported between the ability of different L. plantarum to ferment GOS and the β-galactosidase activity. The results revealed that the oligosaharide utilization patterns are strain specific and they do not dependent from the origin of the tested LAB. Two strains - a vaginal L. fermentum and a dairy L. plantarum, were selected and additionally characterized. Obtained results are promising for further development of new synbiotic formulas.


P57 A BIOSENSOR FOR ASSESSING COMPATIBILITY OF NEW MATERIALS IN REGENERATIVE AND REPARATIVE MEDICINE SVETLANA IVANOVA1*, STOYAN CHAKAROV2, ROUMEN PANKOV3

1Scientific Technological Service Ltd., Sofia, Bulgaria 2Sofia University "St. Kliment Ohridski", Faculty of Biology, Department of Biochemistry, Sofia 3Sofia University "St. Kliment Ohridski", Faculty of Biology, Department of Cytology, Histology and Embryology, Sofia, Bulgaria *Corresponding author: [email protected] Keywords: biocompatibility, biomimetic materials, biosensor, regenerative and reparative medicine

The concept for invisibility and inertness of tissue implants has recently been shifted towards the perception that prostheses must mimic to a maximum extent the natural environment to effectively secure implantation. Newly developed biomimetic materials have to provide a three-dimensional scaffold and the required signals for cells adhesion, proliferation, differentiation and formation of new tissue with deposition of own extracellular matrix. With the advances in material science, polymer chemistry and molecular biology innovative biomaterials are constantly evolving and the necessity of rapid and unambiguous assessment of their biocompatibility arises.

The initial interaction of a cell with a substrate commences with its spreading and formation of primary adhesion contacts which develop into stable and organized structures. Efficient approach to evaluate surface biocompatibility is the characterization of these cell-substrate adhesions.

Here we present a living cell-based biosensor to estimate the type, area and morphological characteristics of adhesion contacts as an indication of the functional status of cells and hence the biocompatibility of the materials studied. Unlike molecular biosensors which detect the effect of a single target substance, cell biosensors are complex systems and offer the advantage to simultaneously receive, process and react to numerous and diverse signals such as adhesivity, elasticity, chemical complexity and topology through continuous expression of genes coding tagged proteins.

We have transfected the fibroblast cell line NIH/3T3 with two expression plasmid constructs and cloned cells after antibiotic selection to generate biosensor cells that are stably expressing two chimeric fluorescent tagged proteins, namely GFP-tensin and mCherry-vinculin/paxillin (components of the focal and fibrillar adhesion contacts). Further fluorescent microscopic analysis and specific image processing allows us to quantitatively assess the surface properties of novel biomaterials for regenerative medicine.


P58 REPAIR PLAN VS. PLAN REPAIR - A PANEL FOR ASSESSMENT OF INDIVIDUAL REPAIR CAPACITY UNDER PHYSIOLOGICAL AND PATHOLOGICAL CONDITIONS CHICHEVA Z.1, CHELENKOVA P.1, *PETKOVA R.1, CHAMOVA T.2, JELYAZKOVA-GLAVEEVA S.2, TOURNEV I.2, CHAKAROV S.3* 1 Scientific Technological Service (STS), Ltd., Sofia, Bulgaria 2 Clinic of Neurology, University Hospital “Alexandrovska”, Sofia, Bulgaria 3 Sofia University "St. Kliment Ohridsky", Sofia, Bulgaria Corresponding authors: Rumena Petkova, Stoyan Chakarov

Individual capacity for repair of everyday DNA damage or of specific types of lesions (e.g. environmental genotoxicity or iatrogenic interference, e.g. chemo- or radiotherapy) can be used as a valid marker for the ability to maintain the integrity of the genome and to assess the propensity for induction of apoptosis. Virtually in all genes coding for products acting in DNA repair and damage-associated apoptosis have been identified polymorphisms that modulate the efficiency of recognition of lesions in DNA or the effectiveness of repair. These polymorphisms alter the repair capacity subtly but distinctly enough to account for differential patterns in succeptibility to certain diseases and conditions, longevity, eligibility for different therapies and outcomes after therapies. We have developed a panel for assessment of individual DNA repair capacity (XPCins83PAT, P53P72R, ЕRCC1C8092A, rate of telomere attrition) which has been used to assess the limits of variance in efficiency of DNA repair and cell and tissue renewal in healthy Bulgarian individuals. The resultant findings reveal features in the population structure which are likely to reflect ancient selection patterns. Also, a preliminary study of marker variance has been carried out on a study group of patients with cardiovascular disease in order to assess the ability to repair oxidative damage and propensity for apoptosis in damaged tissues. Presumably, the panel for assessment of individual repair capacity may be modified further for use in assessment of risk for developing disease or, when the associated diseases or conditions has already developed, for risk of complications.

Acknowledgements: This research has been supported by grants No. DO02-69 and DMU03/112 at the Ministry of Education, Youth and Science of Republic of Bulgaria.


P59 HALF TROUT, HALF SALMON, ALL WRONG. EVIDENCE OF INBREEDING AND GENETIC ADMIXTURE OF RELATED SPECIES IN BULGARIAN NATURAL BROWN TROUT POPULATIONS CHELENKOVA P.1, PETKOVA R.1, YOCHEV S.2, *CHAKAROV S.3

1 Scientific Technological Service (STS), Ltd., Sofia, Bulgaria 2 Bulgarian Ichtyological Society, Sofia, Bulgaria 3 Sofia University "St. Kliment Ohridsky", Sofia, Bulgaria Corresponding author: Stoyan Chakarov

Brown trout (Salmo trutta fario) is a native species to the coldwater rivers of

Europe. It also commonly bred in European and Bulgarian fish farms using breeding material produced locally or imported from abroad. The foreign breeding material often originates from Northern Europe, where related salmonid species such as Atlantic salmon (S. salar) are often bred as well. The occurrence of salmon-trout hybrids has been previously reported to amount to above 10 % in some trout populations from Northern and Western Europe and the issues related to risk of introgression have been brought to the attention of the European environmental control bodies. We found an unusually high level of trout-salmon hybrids in a study group representative of Bulgarian native populations of S. trutta fario. The hybrids are likely to originate from escaped or inadvertently or deliberately released hybrid fish and/or release of breeding stock contaminated with fertile hybrids in the natural water basins. The level of genetic diversity in Bulgarian native populations of brown trout also presents a cause for concern, as we have found apparent evidence of inbreeding which might have resulted from releases or escapes of hatchery material in natural waters. It is advisable that measures be taken in order to prevent further disturbance of the genetic pool of native brown trout populations in Bulgaria.

Acknowledgements: This research was partially supported by grant No. DO02-69 at the Ministry of Education, Youth and Science of the Republic of Bulgaria.


P60 BMP INDUCED DIFFERENTIATION OF HESC BORISLAV ARABADJIEV1,2, STOYAN CHAKAROV1, ALBENA MOMCHILOVA3, ROUMEN PANKOV1*

1Sofia University “St. Kliment Ohridski”, Sofia, Bulgaria 2 Scientific Technological Service (STS) Ltd., Sofia, Bulgaria 3 Institute of Biophysics and Biomedical Engineering, Bulgarian Academy of Sciences, Sofia, Bulgaria *Corresponding author: Roumen Pankov

Keywords: human embryonic stem cells, bone morphogenetic proteins, differentiation

Human embryonic stem cells (hESC) are pluripotent cells, derived from the

Inner Cell Mass (ICM) of in-vitro fertilized pre-implantation stage blastocysts. Embryonic stem cells are localized to the inner cell mass of the developing mammalian blastocyst and can give rise to all future cell types of the organism. In addition, a small population of embryonic stem cells is allocated to the germline - they are called primordial germ cells. The fate of these germline stem cells is different from the remaining cells, which undergo gastrulation and give rise to the three germ layers (endoderm, mesoderm and ectoderm) of the developing embryo.

Here we show different cell types, differentiated in vitro from our previously derived hESC lines B1 and B2. Some of the cell types were spontaneously differentiated, where others were derived using molecular cues such as bone morphogenetic proteins (BMPs) to “push” the cells in the desired direction.

Bone morphogenetic proteins (BMPs) are multifunctional cytokines belonging to the transforming growth factor-β superfamily. We used a combination of BMP-4, BMP-7 and BMP8b in the presence of FBS to differentiate VASA positive cell resembling germ cells, from adherent cultures of B1 hESC. Along the way to differentiating germ cells we observed that addition of BMP-4 to the culture medium of developing embryoid bodies (EBs) in suspension culture increased the rate of incidence of beating cardiomyocytes within them. The occurrence of functional cardiomyocytes under those conditions is not surprising, since BMP signals play a critical role in vertebral mesodermal induction, as well as in heart development. In addition to induced differentiation we also observed spontaneous differentiation of neuron like cells during routine culture of B1 and B2 hESC lines, demonstrating their pluripotency. Acknowledgements: This study was supported by grant No DO02-180 of the Ministry of the Education, Youth and Science of Republic of Bulgaria.


P61 IDENTIFICATION OF XANTHOMONAS STRAINS FROM TOMATO AND PEPPER AND THEIR SENSITIVITY TO ANTIBIOTICS AND COPPER YOANA KIZHEVA1, TACA VANCHEVA1, PETYA HRISTOVA1, MARIYA STOYANOVA2, MONIKA STOJANOVSKA1, NEVENA BOGATZEVSKA2 AND PENKA MONCHEVA1*

1Sofia University, Sofia, Bulgaria, 2 Institute of Soil Science, Agrotechnologies and Plant Protection, Sofia, Bulgaria *Corresponding author: [email protected] Keywords: pepper, tomato, Xanthomonas euvesicatoria, Xanthomonas gardneri, Xanthomonas vesicatoria

Bacterial spot is a serious disease on tomato and pepper in the vegetable-producing fields of Bulgaria and Macedonia. This study aims to characterize 136 strains (62 isolates from tomato, 27 from weeds-resident phase, and 47 – from pepper) from different regions in these two countries. Three bacterial species were identified by the means of PCR with species-specific primers. The analysis proves that Xanthomonas vesicatoria and Xanthomonas euvesicatoria have been present in Bulgaria and Macedonia for the last years. X. vesicatoria affects tomato and pepper and is the dominant species in Bulgaria on tomato and weeds while X. euvesicatoria affects only pepper. Xanthomonas gardneri emerges as a pathogen of tomato in Bulgaria in the last two years. This study reports for the first time in Bulgaria the species X. euvesicatoria and X. gardneri as pathogens of pepper and tomato, respectively.

The strains were compared for amylase activity and ability to utilize cis-aconitate. The analysis shows that these two characteristics vary and cannot be used for species differentiation on phenotypic level. Twelve antibiotics and copper ions in five concentrations were tested for activity against the isolated strains. Tetracycline, kanamycin, and vancomycin were effective against these leaf pathogens. Streptomycin was effective only against isolates from pepper. Copper ions in concentrations over 0,4 % fully suppressed the growth of bacterial isolates from pepper. Isolates from tomato and weeds were more resistant and only a concentration of 0,5% was needed to achieve the same effect. Most of the strains developed unconstrained in the presence 0,1% copper ions. The analysis of data shows that isolates from tomato and weeds are more resistant to copper ions than the isolates from pepper which can be explained by the use of copper bactericides for disease control of tomato. Acknowledgments: This study was supported by the Scientific Foundation of Sofia University, Project No 059/05.04.2012.


P62 IN VITRO REGENERATION AND AGROBACTERIUM - MEDIATEDGENETIC TRANSFORMATION OF THE RESURRECTION PLANT HABERLEA RHODOPENSIS FRIV.

GALYA PETROVA* AND DIMITAR DJILIANOV

Abiotic stress, AgroBioInstitute, 8 Dragan Tzankov Blvd., 1164 Sofia, BULGARIA *Corresponding author: [email protected] Keywords: Agrobacterium, Haberlea rhodopensis, Regeneration, Transformation

Resurrection plants are widely used models for desiccation tolerance studies. Several gene products have been successfully isolated from these species. In an attempt to study their role, these products have been successfully transferred in other model plants. Since resurrection plants are as a rule, polyploids, they are pure targets for mutational studies. In this respect, the establishment of efficient and repeatable transformation system will contribute significantly for the elucidation of stress tolerance.

In this study we describe for the first time, a procedure for Agrobacterium tumefaciens -mediated genetic transformation of the resurrection plant Haberlea rhodopensis Friv. We developed a new system for in vitro regeneration of Haberlea in liquid media. It enables us to achieve direct regeneration and transformation system, which is an alternative to the callus-based transformation, used in other resurrection plants.

The A. tumefaciens strain LBA4404 harbouring the plasmid pCAMBIA 1305.1 which contains the gus gene as a reporter gene and hpt gene as a selectable marker gene was used. The initial experiments were conducted in order to establish the suitable concentration of cefotaxime for the elimination of Agrobacterium from cultures, as well as the optimal concentration of hygromycin for the selection of transformed plants. It was found that the highest concentration of cefotaxime that protocorms of H. rhodopensis could tolerate is 500 mg/l and they are completely inhibited at 0.75 mg/l hygromycin.

Histochemical GUS assay and PCR analysis confirmed the presence of the gus and hpt genes, respectively. The percentage of GUS activity was 3% and the optimal co-cultivation time was 60 minutes.


P63 EFFECT OF CHILLING AND FREEZING STRESSES ON JASMONATE CONTENT IN ARABIS ALPINA MARKO KOLAKSAZOV1, FRÉDÉRIC LAPORTE2, KALINA ANANIEVA3, PETRE DOBREV4, MICHEL HERZOG5, EVGUÉNI ANANIEV1* 1 University of Sofia, Faculty of Biology, Dept. Plant Physiology 2 University of Grenoble Joseph Fourier, Laboratoire d’Ecologie Alpine – LECA 3 Institutes of Plant Physiology and Genetics, Bulgarian Academy of Sciences – Sofia 4 Institute of Experimental Botany, Prague, Czech Republic *Corresponding author: [email protected] Keywords: Arabis alpina, cold stress, freezing and chilling stress, jasmonates, JA and JA-ile

Jasmonic acid (JA) and its derivatives MeJA and JA-ile are known as active mediators of plant cell response to various environmental stresses, including wounding, salinity and drought. For studying the role of jasmonates in the acquisition of cold stress tolerance in plants, we used the model system of the perennial mountain plant Arabis alpina (Brassicaceae). Three A. alpina populations were selected based on their resistance to frost at –6, –7ºC. These populations included T-plants (tolerant to frost, collected in the higher French Alps, Col du Galibier), NT-pants (non-tolerant, growing in the mountain of Vercors) and SH-plants (short hypocotyl phenotype), the latter considered as an adaptive feature to stress. The experimental scheme included plant development at 22ºC (control temperature), transfer of plants to 4ºC for 4 days (acclimation phase during chilling stress) and exposure to frost at –7ºC for 12h (freezing stress). The endogenous jasmonates content was measured by means of HPLC-MS-MS analyses. Our results showed very high content of JA (4-6 nmol g-1DW) in the three studied populations at control temperature. After chilling stress, the content of JA in T-plants did not decrease in contrast to the 10-fold reduction of JA in NT and SH populations (mean value of 0,5 nmol g-1DW). Thecontent of the biologically active compound jasmonate-isoleucine (JA-ile) was considerably higher in control conditions in T- (1,5 nmol g-1DW) compared with NT- and SH-plants (0,26 and 0,6 nmol g-1DW, respectively). JA-ile decreased after chilling stress in the three populations, remaining considerably higher in T-plants (0,67 nmol g-1DW) as compared with NT (0.07 nmol g-1DW). Plant exposure to freezing temperature led to a very sharp decrease of JA and JA-ile in all three populations. The obtained results indicate that JA and its amino acid conjugate JA-ile may play an important role in the acclimation of Arabis alpina to cold stress. The investigation of JA synthesis genes (LOX and AOS) at transcriptional level is in progress.


P64 BIOFILM FORMING BY LACTOBACILLUS STRAINS ON METAL SURFACES

IGNATOVA-IVANOVA TS.1*, R. IVANOV1

1 Department of Biology, Shumen University, Shumen, Bulgaria *Corresponding author: [email protected] Keywords : biofilm, corrosion, inhibitor, Lactobacillus sp.

Corrosion of metals is one of the most serious and challenging problems faced by industries worldwide. Biofilms composed of a secreted polymeric substance containing microbial population have shown to inhibit corrosion in metals.

It was proved that Lactobacillus strains constituted biofilms in the presence of different amounts carbohydrates (10 and 15% lactose, 4% sucrose and mixture 2% sucrose and 4% maltose). The obtained information was used in a study treating the anticorrosive properties of microbial biofilms synthesized by the latter strain. The study of the corrosive stability of steel samples was conducted on the gravimetrique method. The rate of corrosion, the degree of protection, and coefficient of protection has been calculated. A microscope pictures of the treated steel samples confirmed the corrosive activity.

P65 SUB-CELLULAR FRACTIONATION AND GEL BASED PROTEOMICS OF HABERLEA RHODOPENSIS: A PROMISING APPROACH TO BREAK THE BLACK BOX OF RESURRECTION PLANTS PETKO MLADENOV 1*, DIANA ZASHEVA 2, NIKOLAI K. CHRISTOV 1, MAGDALENA TCHORBADJIEVA 3, DIMITAR DJILIANOV 1 1-Agrobioinstitute 8 Dragan Tzankov Bvld. 1164 Sofia Bulgaria 2-Institute of Biology and Immunology of Reproduction "Acad. Kiril Bratanov “ 73 Tzarigradsko shose Bvld. 1113 Sofia, Bulgaria 3- Sofia University, Faculty of Biology 8 Dragan Tsankov Blvd. 1164 Sofia, Bulgaria *Corresponding author: [email protected] Keywords: BN/PAGE, 2D IEF/SDS PAGE, Haberlea rhodopensis

Haberlea rhodopensis (Gesneriacea) is a Balkan endemite and a glacial relict belonging to the group of the resurrection plants. They are unique among angiosperms with their ability to survive the loss of almost all water content. After rewatering these plants rapidly restore their former state and thus provide a very suitable model to study plant reaction to dehydration. In unicellular organisms a large number of comprehensive flux maps and control mechanisms have been characterized. The complexity of plant metabolic networks, due to the presence of


multiple cell types and the high degree of sub-cellular compartmentation is very challenging for modern biology. Thus the assessment of the specific functions of sub-cellular compartments could add a new dimension of proteome analysis. In this respect, we developed a protocol for simultaneous isolation of pure and intact chloroplasts and mitochondria from plants of Haberlea rhodopensis with subsequent gel-based proteomics (BN/PAGE and 2D IEF/SDS PAGE). The resulting 2D sub-cellular catalogues of proteins from H. rhodopensis will help to elucidate their role in the cell as molecules, spatially organized according to their function. The attribution of certain activities to specific organelles will contribute for drastic reduction in overall sample complexity and to improve the resolution and sensitivity of the analyses.

P66 EFFECT OF SALICYLIC ACID ON PRIMARY PHOTOSYNTHETIC REACTIONS IN WINTER WHEAT CULTIVARS AFTER COLD HARDENING AND FREEZING

PAVLINA SASHEVA 1, RUSINA YORDANOVA 2*, TIBOR JANDA3, LILIANA MASLENKOVA2

1Medical University of Sofia, Faculty of Pharmacy, Dunav str. 2, Sofia 1000, Bulgaria 2Institute of Plant Physiology and Genetics, BAS, 1113 Acad. G. Bonchev 21, Sofia 3Agricultural Institute, Centre for Agricultural Research, HAS, H-2462, Martonvasar, POB 19, Hungary *Corresponding author: [email protected] Keywords: cold hardening, freezing, salicylic acid, thermoluminescence, winter wheat.

Thermoluminescence emission in winter wheat cultivar (Triticum aestivum

L. var. “Sadovo-1”), subjected to short period of low positive temperatures (cold hardening) and subzero temperatures (freezing) followed by period of recovery was measured in order to determinate changes in the redox cycling of Photosystem II centers. Main goal of this study is to investigate weather exogenously applied salicylic acid modulated plant resistance to low temperature. Some parameters, considered like stress markers – the levels of hydrogen peroxide, lipid peroxidation, free proline, rate of electrolyte leakage and the activity of some antioxidative enzymes and were also measured. Data for free and bound salicylic acid accumulation are presented.


P67 DROSOPHILA dmfr1 (DROSOPHILA FRAGILE X MENTAL RETARDATION1) INTERACTS WITH GENES INVOLVED IN SYNAPTIC FORMATION MARIA H. PETROVA, DIMITRINA G. GEORGIEVA, MEGLENA L. KITANOVA, GINKA K. GENOVA*

Department of Genetics, SU”St.Kl.Ochridski”, Sofia, Bulgaria *Corresponding author: [email protected]

Keywords: Drosophila melanogaster; Fragile X syndrome; cell polarity and contacts;synapses.

Fragile X mental retardation is the most common form of human mental

retardation, which is caused by the functional loss of the fragile X mental retardation protein (FMRP). It is not well understood how the absence of this protein is related to this disease pathology.

Drosophila FMRP1, encoded by the gene dfmr1 has a global participation in different processes in the central nervous system of Drosophila. It negatively regulates translation of different neuronal transcripts, takes parts in the mRNA transport in the neurons and in the regulation of the mRNA stability.

We used the GAL4/UAS system to ectopically express dfmr1 in the wing imaginal discs to obtain an abnormal “wing” phenotype”. On the background of this mutant phenotype we investigated the genetic interactions in the adult wings of dfmr1 with genes involved in cell polarity, cell contacts and synaptic formation. Mutations in these genes modified the mutant wing phenotype. Mutant scribbled (scrib) made this phenotype worse and was classifies as an enhancer, and mutant discs large 1 (dlg1) rescued the same phenotype and was classifies as a suppressor.

We accomplished a semi-quantitative RT-PCR to analyze the relative expression levels of mRNAs, encoded by these two genes in wild type flies and in the mutant stock, in which dFMRP was overexpressed. We observed increased dlg1-and scrib mRNA levels in the stock overexpressing dFMRP, as compared to the wild type. We speculate that dFMRP exerts a positive control at the level of transcription of both genes.

Acknowledgments: The authors express sincere gratitude for the funds of the budget of the SU “St.Kl.Ochridski” supporting research in our work (023/05.04.2012).

The authors express grateful acknowledgements to the National Science Fund of the Ministry of Education and Science (DID-02/35/2009) for support and funding.

The authors express sincere gratitude to Bloomington Drosophila Stock Center at Indiana University, USA for supporting research in our work.


P68 CYSTATHIONINE GAMMA–LYASE AS A REGULATOR OF RESISTANCE ARTERY CONTRACTION UNDER NORMAL AND HYPERGLYCEMIC CONDITIONS RADOSLAVA EMILOVA1*, DANIELA DIMITROVA 2, VENKO GEORGIEV1, TEODORA DANEVA3 ,HRISTO GAGOV 1

1 Dept. of Animal and Human Physiology, Faculty of Biology, Sofia University 2 Dept. of Excitable Structures, Institute of Biophysics, Bulgarian Academy of Sciences (BAS) 3Dept. of Immunobiology of Reproduction, Institute of Biology and Immunology of Reproduction, BAS *Corresponding author: [email protected] Keywords: artery, contraction, cystathionine gamma–lyase, hyperglycemia

The aim of our study is to clarify the participation of cystathionine gamma–lyase (CSE) that produces H2S in smooth muscle and perivascular adipose tissues but not in vascular endothelium in the regulation of resistance size rat artery contraction.

Study design and methods: In this research the isometric contraction of de-endothelised rat gracilis arteries (a. gracilis) was measured with Small Vessel Myograph (DMT 410M, Denmark). Isometric wire myography is an in vitro technique that examines the functional responses and vascular reactivity of isolated rings of vessels with small diameters. In our study rat a. gracilis were dissected of approximately 2mm ring preparations in chilled physiological saline solution (PSS). Half of the isolated rings were incubated in PSS with 20 mmol/l D-glucose and the others in normal glucose concentration (5.5 mmol/l). Increasing concentrations of serotonin (5-hydroxytryptamine, - from 10-10 to 10-5 mmol/l) were applied to induce gradual constriction of circular artery segments. The absence of endothelium was confirmed by the lack of the relaxation to acetylcholine of 60mM KCl - contracted arteries. The relaxing effect of CSE was inhibited by DL-propargyl glycine. Statistical analyses were performed by SPSS. All results are given as means ± S.E.M of six separate experiments. Statistical significance was determinate using Student t-test.

Results and discussion: The extracellular glucose concentration of 20 mmol/l do not influence the contractile effect of serotonin. The inhibition of CSE significantly enhanced the maximal force of serotonin-induced contraction under normal and hyperglycemic conditions. It is concluded that CSE is an important regulator of endothelium denuded rat resistance arteries. It is suggested that CSE in smooth muscle cells of artery wall released mediator that antagonised the constricting action of serotonin. This regulation is insensitive to extracellular glucose concentration. Acknowledgments:Supported by Grant № 57/2012 from Sofia University St. Kliment Ohridski, Sofia, Bulgaria: Role of cystathionine gamma-lyase as a regulator of arterial tension in hyperglycemia and diabetes.


P69 CARBOHYDRATE COMPOSITION AND ANTIOXIDANT ACTIVITY OF ROOT EXTRACTS OF INULA HELENIUM L. R. Z. VRANCHEVA2, N. TR. PETKOVA1*, I. G. IVANOV1, P. P. DENEV1*, A. I. PAVLOV1,4, J. N. ALEKSIEVA3

1Department of Organic Chemistry, 2Department of Analytical Chemistry, 3Departament of Catering and Tourism University of Food Technology, 26, Maritza Blvd., Plovdiv, 4002 4Institute of Microbiology Stephan Angelov, BAS Sofia *Corresponding authors: [email protected], [email protected] Keywords: elecampane, inulin, antioxidant activity

Ultrasonic and microwave-assisted extractions of biologically active substances from the roots of elecampane (Inula helenium L.) with three different polar solvents - water, methanol and 70 % (v/v) ethanol were investigated. The carbohydrate composition in all extracts was qualitatively determinated by TLC analysis. The fructan content was analyzed by spectrophotomeric method based on the resorcinol reagent. The highest yields of inulin and FOS (42 g/100g dry weight) were obtained by the performed water extraction combined with ultrasonic irradiation. The total phenolic and flavonoids quantities were analyzed by using a Folin–Ciocalteu’s and Al(NO3)3 reagents, respectively. The total antioxidant activity of different extracts from elecampane roots was estimated by: DPPH (1,1-diphenyl-2-picrylhydrazyl radical), ABTS (2,2`azinobis-(3-ethylbenzthiazoline-6-sulfonic acid)), FRAP (ferric reducing antioxidant power) and CUPRAC (cupric reducing antioxidant capacity ). The 70 % (v/v) ethanol extracts obtained by the ultrasound-assisted extraction showed the highest antioxidant activity determinated by the following methods DPPH -107,20 mM TE/g dw; ABTS -86,05 mM TE/g dw, FRAP - 67,05 mM TE/g dw and CUPRAC - 173,02 mM TE/g dw. The reason for these was probably depend on the highest content of total phenols and flavonoids (7,87 mg /g GAE dw and 23,87 mM QE/g dw, respectively).


P70 STUDY ON TRIPEPTIDYL PEPTIDASE I ACTIVITY IN THE RAT BRAIN AFTER SODIUM NITRITE-INDUCED ACUTE HYPOXIA

STELLA DIMITROVA, EMILIA PETROVA*, LILIA GEORGIEVA, MASHENKA DIMITROVA, DIMITAR KADIYSKY

Institute of Experimental Morphology, Pathology and Anthropology with Museum, Bulgarian Academy of Sciences, Acad. G. Bonchev Str., Bl. 25, 1113 Sofia, Bulgaria, *Corresponding author: [email protected] Keywords: acute hypoxia, rat model, sodium nitrite, tripeptidyl peptidase I

Background: Acute hypoxia could appear during different conditions – shock, severe intoxications, multiple injuries, acute respiratory or heart failure, etc. It might cause a substantial brain damage and lasting neurological sequelae. Tripeptidyl peptidase I (TPPI) activity is crucial for the neuronal functions. Little is known about the enzyme pattern after a hypoxic shock. The aim of the present study is to follow up TPPI activity changes in the rat brain after NaNO2-induced acute hypoxia.

Methods: Four-month-old male Wistar rats were injected intraperitoneally with 50 mg/kg body weight NaNO2 (distilled water for controls) and sacrificed at different time intervals (1h, 5h, 24h, and days 2, 10, 20) following the administration. TPPI activity was visualized in the cerebral cortex and cerebellar cortex using enzyme histochemical method with the substrate Gly-Pro-Met-1-anthraquinonyl hydrazide. Silver-copper staining for neurodegeneration was also carried out.

Results: TPPI activity was visibly lower in the studied brain regions from the 1st hour up to the 10th day after intoxication as compared to controls. Signs of enzyme activity recovery were observed on the 10th day and on the 20th day the enzyme pattern was similar to the one in controls. These changes corresponded to the damaged nerve fibers and profound neuronal loss from the 5th hour after treatment as demonstrated by the silver-copper impregnation technique.

Conclusion: TPPI activity is down-regulated after acute hypoxic treatment in the rat cerebral cortex and cerebellar cortex. The enzyme might participate in the tissue response to the hypoxic shock or, otherwise, its lowered activity could be one of the reasons for the neuronal loss. Thus, the enzyme pattern in the rat model of acute hypoxia deserves to be studied in more details in order to elucidate the intrinsic mechanisms of the brain response after hypoxia.

Acknowledgements: This work is supported by a grant No DMU 03/18 for Young scientists from the Bulgarian National Science Fund.


P71 EXOGENOUS TREATMENT BY PLANT GROWTH REGULATORS AFFECTS ARTEMISIA ALBA TERPENOID PROFILE BY ALTERING MORPHOGENESIS, PHYSIOLOGICAL STATUS AND ENDOGENOUS CYTOKININ LEVELS IN VITRO SASHKA KRUMOVA1, VACLAV MOTYKA2, PETRE DOBREV2, MILKA TODOROVA3, ANTOANETA TRENDAFILOVA3, LJUBA EVSTATIEVA4, KALINA DANOVA3 1 Institute of Biophysics and Biomedical Engineering, BAS, 1113 Sofia, Bulgaria 2 Institute of Experimental Botany AS CR, Prague, Czech Republic 3 Institute of Organic Chemistry with Centre of Phytochemistry, BAS, 1113 Sofia, Bulgaria 4 Institute for Biodiversity and Ecosystem Research, BAS, J. Gagarin 2, Sofia 1113, Bulgaria *Corresponding authors: [email protected] ; [email protected]; [email protected]; [email protected], [email protected]

Keywords: Artemisia alba in vitro, endogenous cytokinins, Photosystem II, plant growth regulators, terpenoid profile

Modifications of plant growth regulators (PGRs) supplied to Artemisia alba in vitro cultures have been previously shown to affect plant morphogenesis and terpenoid profile of the essential oil of aerial parts. In continuation of this research we report here on the effects of PGRs on the terpenoid profile, structure of photosystem II in vitro as well as on the endogenous cytokinin (CK) levels of both above- and underground parts. The contents of CK bioactive forms (free bases and ribosides) were followed. It was revealed that the PGR-modified growth and development as well as the alterations of photosystem II structural organization were related to the CK levels of Artemisia alba. Thus, elevated monoterpenoid levels were associated with a higher peripheral antennae aggregation and elevation of trans-zeatin riboside, dihydrozeatin and dihydrozeatin riboside as well as N6-(2-isopentenyl)adenine in the aerials of the respective PGR treatments. These results imply of the role of exogenous factors such as PGR supplementation in affecting the terpenoid biogenesis in vitro by altering the levels of endogenous CKs and physiological status of the plant organism.


P72 THE IMPORTANCE OF ENZYMES IN FERMENTATION OF RAW DRIED TRADITIONAL SAUSAGES BULGARIAN TYPE LUKANKA

SASHO STOYANOVSKI1*, ZIVKO GACOVSKI1, STEFKA ANTONOVA-NIKOLOVA2, NIKOLAY KIRILOV2, ISKRA IVANOVA2, TENYO TENEV3, VALENTINA HADJINESHEVA3

1Bitola University “St.Kliment Ohridski”, Faculty of Veterinary 2Sofia University “St.Kliment Ohridski”, Faculty of Biology, Department of General and Industrial Microbiology 3Sofia University “St.Kliment Ohridski”, Faculty of Biology,Departemnet of Cytology,Histology and Embriology *Corresponding author: [email protected] Keywords: sausage, taste, fermentation, variety

The first recorded evidence of the meat processing industry dates back to Roman times (Lucke, 1974). During fermentation, the result of complex biochemical and physico-chemical reactions occurring changes in the composition and structure of meat in aromatic and taste the finished product, which are different from those of raw material. The typical taste of dry sausages is result of microbial and enzymatic metabolism of carbohydrates, proteins and lipids. In combination with the spices, the complex metabolic products affect the sensory profile of the sausages.Fermentation of meat is a process used away back to extend the shelf life of otherwise perishable commodity. Production of naturally fermented sausages is traditional in countries of southern Europe, but also occurs in some Scandinavian countries and Latin America. Specific taste, odor, color and structure of the sausages is due to the characteristics of the input in their raw meat and spices and applied technology. Of great importance for the final product is the composition of natural microflora of meat and fermentation process, which it operates. Therefore, the variety of ranges in different geographical areas is huge. Study of enzymes in the fermentation of raw dried sausages traditional Bulgarian type of sausage for obtaining of taste is of great importance the finished product. More then 39 strains of genera Lactobacillus were isolated during the different stages of the fermentation process of the naturally fermented sausages.The strains were characterized in terms of some enzymatic activities using the system API ZYM. Evaluation of the activity was carried out on 5-point scale, composed according to the intensity of discolored.Enzymatic potential of bacteria is an important factor in shaping the characteristic flavor of meat products, but also for its diversity in different products. This starter cultures reflect on the taste and aroma of fermented meat products.


P73 DEVELOPMENT OF MULTIPLEX PRIMER SETS FOR COST EFFICIENT SSR GENOTYPING OF MAIZE (ZEA MAYS) MAPPING POPULATIONS ON A CAPILLARY SEQUENCER STEFAN TSONEV*, MICHAELA VELICHKOVA, ELENA TODOROVSKA, VIKTORIA AVRAMOVA, NIKOLAI K. CHRISTOV

AgroBioInstitute, 8 Dragan Tsankov blvd., 1164 Sofia, Bulgaria *Corresponding author: [email protected] Keywords: maize, mapping, multiplex PCR, SSRs

Because of their accessibility, genomic abundance and high level polymorphism SSRs are often markers of choice in many QTL mapping studies. To obtain a good mapping resolution one have to genotype a large number of individuals in a large number of marker loci, which is both labour-intensive and expensive task. The multiplex-ready PCR technique (MRTTM) provides possibility for fast optimisation with simultaneous amplification and flourescent labeling of several amplicons in one reaction tube. Thus, avoiding the complex optimisation work for a standart multiplex PCR applications. In this study we propose a low-cost protocol for optimizing high-throughput genotyping of maize mapping populations using SSR markers and capillary electrophoresis. The method is based on MRTTM

and consists of three stages. In the first stage the parental lines and F1 were screened in 176 public SSR

loci. Fifty four of the analyzed loci were found to be polymorphic between parental lines. In the second stage, primer concentrations were optimized in uniplex reactions. In the third stage the polymorphic markers were grouped in 19 sets containing 2 to 6 primer pairs and tested for successful amplification in multiplex PCR reactions. Using the optimised protocol, a maize DH population consisting of 192 individuals was genotyped. The cost and effort benefits of the proposed method will be discussed.


P74 CALLUS CULTURES AND INDIRECT REGENERATION OF RUSCUS HYPOGLOSSUM IN VITRO

TEODORA IVANOVA*, DESSISLAVA DIMITROVA, GEORGI ANGELOV, CHAVDAR GUSSEV, YULIA BOSSEVA, TATYANA STOEVA

Department of Plant and Fungal Diversity and Resources, Institute of Biodiversity and Ecosystem Research, Bulgarian Academy of Sciences, 23, Acad. G. Bonchev Str., 1113, Sofia, Bulgaria *Corresponding author: [email protected]

Keywords: genome size, isoenzymes, micropropagation, Ruscaceae

Ruscus hypoglossum L. (Ruscaceae) is highly praised ornamental plant collected predominantly from the wild and threatened in several countries due to overcollection. Pest and disease sensitivity and low germination rate hamper it’s cultivation. Thus micropropagation has been considered to be advantageous for rapid production of planting material and conservation of genetic resources. Callus cultures of R. hypoglossum were induced on thidiazuron (TDZ) containing medium and indirect regeneration rates were tested on media with different growth regulators and sucrose concentrations. Results showed variability in regeneration rates and quality of obtained plants depending on the clonal identity of the initial cultures. TDZ ensured higher regeneration rates and caused marked cladode alteration in regenerants that were persistent through whole culture process. Callus induction was optimal at 30 g.l-1 sucrose and dropped down with the increase of sucrose concentration. Shoot proliferation was positively influenced by the increase of the sucrose 15-60 g.l-1. Isozyme profiles (esterase, peroxidase and acid phosphatase) and DNA content were assessed and possible causes for the observed variation on molecular level was discussed.

P75 THE SALINITY EFFECT ON THE MORPHOLOGY AND THE PIGMENTS CONTENT IN THREE PAULOWNIA CLONES GROWN EX VITRO

K. MILADINOVA1, K. IVANOVA2, T. GEORGIEVA1, M. GENEVA3, Y. MARKOVSKA2*

1 Biotree, 8 Iliensko shoes str., 1220 Sofia, Bulgaria; 2 Faculty of Biology, University of Sofia, 8 Dragan Tsankov Blvd., Sofia 1164, Bulgaria 3 Institute of Plant Physiology and Genetics, BAS, Acad. Georgi Bonchev str., bl. 21, Sofia *Corresponding author: [email protected] Keywords: hydroponic, morphology, Paulownia, pigments, salinity.

Paulownia is an important woody species using in reforestation programs and will be an intercropping species too. We evaluated the effect of salt stress on growth


parameters, chlorophyll a, chlorophyll b and carotenoids in Paulownia clones (P. elongata x fortunei x elongata - T2, P. elongata x elongata - T4, P. elongata x kawakarnii - EK) grown in hydroponic at three levels of salinity, 50 mmol/l, 100 mmol/l, 200 mmol/l sodium chloride (NaCl) solution. When in vitro propagated plants reached a determined size, they were transferred to nutrient solution for 48 days and than were treated with NaCl solutions for 10 days. We examined the physiological indicators in order to develop practicable strategie for selecting salt tolerant clones of Paulownia, which were produced by BioTree Ltd., Bulgaria. The root and stem length, as well as leaf number and total leaf area of T2 clone were reduced insignificantly in comparison with these of T4 and EK clones during NaCl treatment. Control of T2 clone was characterized with approximately twice higher total dry mass per plant than EK and T4 clones. The root dry mass to shoot dry mass ratios of three clones changed in a different manner with increasing salinity levels and was highest in EK clone at 200 mmol/l NaCl, followed by T4 and T2. Total leaf area showed the capability of the plants in forming of photosynthetic surface and was reduced more in T4 and EK clones under salt stress. The chlorophyll a to chlorophyll b ratio as well as the green to yellow pigments ratio increased only in the leaves of T4 clone. Our results suggest that T2 clone was more tolerant to salt stress than EK and T4 clones.

P76 EFFECT OF 6-BENZYLAMINOPURINE ON MICROPROPAGATION OF ARTEMISIA CHAMAEMELIFOLIA VILL. (ASTERACEAE) LYUBOV HRISTOVA*, EVELINA DAMYANOVA, ZORNICA DOICHINOVA AND VENETA KAPCHINA-TOTEVA Department of Plant Physiology, Faculty of Biology, Sofia University “St. Kliment Ohridski”, 8 Dragan Tsankov Blvd., 1164 Sofia, Bulgaria, *Corresponding author: [email protected] Keywords: in vitro propagation, endangered species, cytokinin

The influence of different concentrations (0.1 – 1.0 mg.L-1) of the cytokinin 6-benzylaminopurine (BA) on the in vitro propagation of the endangered aromatic plant Artemisia chamaemelifolia Vill. (Asteraceae) was examined. The increase of concentration stimulated the formation of shoots but shortened their length. The middle concentrations of 0.5, 0.6 and 0.7 mg.L-1 BA gave the highest number of shoots but they also appeared to promote negative effects like vitrification and necrosis more than the other variants. Although there is no clear trend, all the variants have higher fresh weight and form more primary callus than the control explants grown on Murashige & Skoog medium with Gambourg vitamins. Over 50 % of the explants gave rise to more than 10 new shoots per explant at concentrations of 0.2, 0.3 and 0.9 mg.L-1 BA. All concentrations of BA suppressed root formation and manifested different extent of vitrification. This is the first report to indicate a


stimulation activity of BA on the micropropagation and growth of Artemisia chamaemelifolia Vill. and the results are perspective for a biotechnological application in future experiments for biomass and useful metabolite production.

P77 THE ECOLOGICAL AND FLORISTIC CHARACTERISTICS OF POPULATIONS OF SIDERITIS SCARDICA GRISEB. IN SLAVYANKA MOUNTAIN LJUBA EVSTATIEVA1, INA ANEVA1*, PETER ZHELEV2, DIMITAR DIMITROV3 1Institute of Biodiversity and Ecosystem Research, BAS, 23, Acad. G. Bonchev Str., 1113 Sofia 2University of Forestry, 10 Kliment Ohridsky blvd., 1756 Sofia, Bulgaria 3National Museum of Natural History, 1 Tsar Osvoboditel Blvd *Corresponding author: [email protected] Key words: flora, population, Sideritis scardica

Sideritis scardica Griseb. is an endangered species with threatened and extremely vulnerable populations. The current study provides information about the population state and structure, vegetation dynamics and the place of the species in habitats in all locations of Slavyanka Mountain. Three localities of the species are investigated – at the base of the peaks Golyam Tsarev Vrah, Shabran and Gotsev Vrah. A list of diversity of vascular plants is made for each locality. The taxonomic structure of the flora and the ecological and biological characteristics are defined.

P78 EVALUATION OF THE SELF-PURIFICATION IN THE WATERS OF THE MICRO-DAMS IN THE SMALL HYDRO ELECTRIC POWER PLANTS (HEPPS) LAKATNIK AND SVRAZHEN: POTENTIAL OF THE BIOALGORITHMS

I. YOTINOV*, ST. LINCHEVA, L. KENDEROV, I. SCHNEIDER, Y. TOPALOVA*

Department “General and Applied Hydrobiology”, Faculty of Biology, Sofia University “ *Corresponding authors: [email protected]; [email protected]

Keywords: bioalgorithms, chemical and microbiological parameters, waters, self-purification

Environmental management requires the solution of problems through a comprehensive and thorough examination of all levels - from biochemical analysis to bio- managerial approaches. The chemical and microbiological parameters of water in depth in micro-dams Lakatnik and Svrazhen have been investigated in June and in September, 2011. Linear correlations between microbiological and hydrochemical indicators have been found. They give key information about the


biochemical processes underlying the self-purification. Moreover, these correlations create a highly effective indicator system. The information from this indicator device will enable the preparation of forecast bioalgorithms for the express diagnostics of risk events in the waters of the studied micro-dams in the small HEPPs Lakatnik and Svrazhen. These algorithms will have a high potential for control and indication of self-purification processes.

Our results show that these small dams provide excellent conditions for the implementation of self-purification processes in the water because the speed of the water in these reservoirs is slowing, which creates favorable conditions for the growth of microorganisms in the water. The dams can be considered as natural treatment reactors where the microbial communities, operating in different layers of water, play the key role. As a result, the quality of the aquatic ecosystem is improved, in particular, as well as the quality of the environment, in general.

P79 DYNAMICS OF THE FUNCTIONAL STRUCTURE OF THE SEDIMENT MICROFLORA DURING THE MODEL PROCESS OF PHENOL BIODEGRADATION

S. LINCHEVA*, I. SCHNEIDER, Y. TOPALOVA

Sofia University, Faculty of Biology, Department of General and Applied Hydrobiology, 8 Dragan Tzankov blvd., 1164, Sofia, Bulgaria *Corresponding author: [email protected] Keywords: functional microbial diversity, phenol biodegradation, sediments

The microbial communities are main biological factor realizing biodegradation of toxic compounds. Microorganisms from g.Pseudomonas and g.Acinetobacter have a key position in the detoxification segment. They have options for alternative ways of energy supply and powerful potential for rapid inclusion of metabolic pathways for degradation of xenobiotics. Oil products and phenols are often at concentrations above exposure limit in the sediments of the Iskar River and created reservoirs in the region of Cascade "Middle Iskar". Transformation of toxic pollutants is essential for self-purification potential of the river and reservoirs because of their inhibitory effect on transformation processes. Degradation of xenobiotics is a complex process, which mainly determined by concentration of toxic pollutant and the adaptive capacity of the sediments communities.

The biodegradation of the toxic compound phenol in the sediments of reservoir Lakatnik situated in the Iskar River was simulated in the laboratory model process with duration of 336 hours. Phenol at a concentration of 200 mg /l was the investigated xenobiotic. The process was characterized by the following microbial and chemical indicators: the amount of the key functional groups of microorganisms - aerobic heterotrophic bacteria, anaerobic heterotrophic bacteria, g.Pseudomonas,


g.Acinetobacter; residual phenol; effectiveness of phenol removal. The dynamics of microorganisms showed greater changes after 72 hours of the process. All microbial groups quickly adapted to the amount of 200 mg/l phenol. In the initial phase of the experiment (0 h -72 h) the phenol transformation was negligible. First peak of g.Pseudomonas and g.Acinetobacter was found in the initial phase, related to adaptation to the modulator. The results confirmed that at the end of the process the phenol concentration decreased by 69%. In same time the amount of the key microorganisms for phenol biodegradation increased with 13% for g. Acinetobacter and with 6% for g.Pseudomonas.

P80 ENZYME ACTIVITIES AS A TOOL FOR BIOLOGICAL CONTROL IN DAIRY WASTEWATER TREATMENT

IRINA SCHNEIDER* AND YANA TOPALOVA

Department “General and Applied Hydrobiology”, Faculty of Biology, Sofia University “St. Kliment Ohridski”, 8 Dragan Tzankov blvd., 1164 Sofia, Bulgaria *Corresponding author: [email protected] Keywords: biofilm, β-galactosidase activity, nitrate reductase activity, phosphatase activity, total dehydrogenaze activity

The biological control, especially the selection of appropriate biological indicators, is important for receipt of more adequate assessment and for better management of wastewater treatment processes. Increasing application of biofilm processes in wastewater treatment requires the development of a new analytical approach for fixed biomass activity characterization. The evaluation of opportunity for application of enzyme activities of the total dehydrogenase, phosphatase, nitrate reductase and ß-galactosidase as biological indicators of well functioning biofilms in dairy wastewater treatment is the aim of this article.

The lab scale wastewater treatment process was modelled in biofilters (with carrier gravel) in semi-continuous regime for 282 days. A model wastewater was used at cycle of feeding 19±2d. It contained mineral media and whey as a bi-product from milk processing. Three biological systems with different inocula for initial biofilter inoculation: i/ acclimated activated sludge (AAS); ii/ AAS with addition of microbial preparation HydroPacks and iii/ AAS with addition of microbial preparation Laktazym, were investigated. The fixed biomass activity was studied in parallel with the activity of the suspended biomass. The enzymological parameters were discussed with removal effectiveness for key pollutants (COD, proteins and lactose) at the 216th hour which corresponded to the phase of stabilization of sequencing batch process (from 105 to 282 day) and the stable biofilm development. The obtained data showed two important tendencies for biofilm activity assessment: i/ the total dehydrogenase activity was appropriate indicator for assessment of total organic matter (measured as COD) mineralization and ii/ the phospatase and nitrate


reductase activities were indirectly related to protein biotransformation. The ß-galactosidase activity of suspended biomass was more appropriate indicator for assessment of lactose biodegradation in comparison with the ß-galactosidase activity of biofilm. The investigated enzyme activities are quick, accurate and easy applicable tool for control and management of wastewater treatment in the dairy industry.

P81 EFFECT OF CHOLESTEROL ON THE PHASE SEPARATION OF AMINOPHOSPHOLIPIDS: ELUCIDATION OF THE ROLE OF AMINOPHOSPHOLIPIDS IN THE FORMATION OF LIPID RAFTS IN MODEL MEMBRANES RUSINA HAZAROSOVA*, GALYA STANEVA, DIANA PETKOVA, ALBENA MOMCHILOVA Institute of Biophysics and Biomedical Engineering, Bulgarian Academy of Sciences *Corresponding author: [email protected]

Plasma membranes of cells exhibit asymmetrical distribution of the lipid molecules between the two membrane monolayers. The aminophospholipids phosphatidylserine and phosphatidylethanolamine are located in the inner leaflet of the plasma membrane whereas phosphatidylcholine and sphingomyelin – in the outer one. The existence of liquid-ordered domains (Lo phase) in glycerophospholipid matrix (liquid-disordered (Ld) phase) mimics the properties of membrane domains, called “rafts” at cell level. Rafts are enriched in sphingomyelin (SM) and cholesterol (CHOL). Nowadays, little is known about the ability of the aminophospholipids to form Lo domains. That is why we focused our attention on the phase separation of aminophospholipids in glycerophospholipid matrix as well as the effect of CHOL on it. All studies were performed by means of fluorescence microscopy of giant unilamellar vesicles (GUV). GUVs were composed of egg yolk phosphatidylcholine/egg yolk phosphatidylethanolamine (EYPC/EYPE), egg yolk phosphatidylcholine/brain phosphatidylserine (EYPC/BPS) and in the ternary mixtures, where the two aminophospholipids in question are combined with each other (EYPC/EYPE/BPS). Gel/liquid-disordered phase coexistence was detected in these mixtures, where aminophospholipids segregate in gel leaf-like domains. When cholesterol was added to each one above-mentioned mixture, the phase separation was shifted at lower temperatures, namely it increase miscibility of the inner leaflet lipids. Therefore, aminophospholipids and cholesterol did not form micron-scale Lo domains. Addition of aminophospholipids to the control mixture (EYPC/EYSM/CHOL, where Lo/Ld immiscibility is observed) induced raft formation at higher temperature. Based on these results, one can conclude that aminophospholipids promote the formation of Lo domains.


Financial assistance was provided from Bulgarian Fund for Scientific Research (D002-212/2008, DTK 02/5-2009) and Operational Program ‘‘Human Resources Development” (BG 051PO001-3.3.04/42) co-financed by the European Social Fund of the European Union.

P82 INFLUENCE OF SODIUM NITROPRUSSIDE AS A DONOR OF NITRIC OXIDE ON ATP-ASE ACTIVITY OF RAT LIVER MITOCHONDRIA

MILENA SHKODROVA*, MARIELA CHICHOVA, DIMITAR GANCHEV, ILIYANA SAZDOVA, IGNAT MINKOV

Department of Animal and Human Physiology, Faculty of Biology, University of Sofia, *Corresponding author: [email protected] Keywords: mitochondria, mitochondrial ATPase, nitric oxide, sodium nitroprusside

Nitric oxide (NO) is a signal molecule, which is unstable in aqueous solution and rapidly converts to peroxynitrite and nitrate. Regardless of the strong evidences that NO and its derivatives inhibit the mitochondrial function as a result of interactions with most of the respiratory chain transporters, the data concerning the overall process of the oxidative phosphorylation are contradictory.

The present work was undertaken to investigate the influence of sodium nitroprusside (SNP) as a donor of NO on ATPase activity of rat liver mitochondria. ATPase activity was determined by measurement of inorganic phosphate increase in the reaction medium. The results obtained when SNP was added directly to the reaction medium were rather contradictory. We observed an inhibition of the ATPase activity of rat liver mitochondria uncoupled by freezing/thawing, whereas preliminary deactivation of the compound provoked the opposite effect. SNP added directly to the reaction medium led to an increase of the ATPase activity of submitochondrial particles (SMP). The discrepancies in the observed effects may result from the influence of NO itself, donor molecule, products of its degradation and/or from their combined effect. Therefore, a specially constructed experimental system allowing separation of NO from the molecule of its donor was used. In these experiments a thin polytetrafluoroethylene membrane separated the cuvette where SNP was added from that one where ATPase reaction was performed, thus isolating solutions in the cuvettes, but allowing gas diffusion. Using this system we found an inhibition of ATPase activities of rat liver uncoupled mitochondria and of SMP. This effect was abolished after preliminary deactivation of SNP. We concluded that the ATPase activity changes were the result of influence of NO diffusing through the membrane. The present experimental system can be applicable to studies of different enzyme systems sensitive to NO.


P83 SURVEY AND HEALTH ASSESSMENT OF NEW COSMETIC PRODUCT BASED ON NATURAL INGREDIENTS NATALIYA GERGINOVA1*, ELENA IVANOVA1, KALOYAN BOZHANCHEV1 , SILVIA GENEVA1, ANTON TACHEV 2

1Student: Sofia University "St. Klmient Ohridski", Faculty of Biology, 1164 Sofia, Bulgaria , 2National center of Public Health and Analysis *Corresponding author: [email protected] Keywords: cosmetics, Vaseline, natural products

The modern trends in cosmetics are orientated towards searching for the use of more natural products which are a part of a complex approach to a more natural way of life. In answer for this search the purpose of this research was to create a new natural cosmetic product (hand-made cosmetics), which is a mixture of compounds with natural and animal origins which have a revitalizing effect on the skin.

For the creation of the product a set of classical technologies was used: gradually adding the raw material, mixing and dissolving until a stable and homogenized product is created. The order in which the compounds were added is based on their physical chemistry properties. The product has been subdued to standard cosmetics quality tests, which it has passed: pH of 10% water solution, thermostability, homogeneity and a microbiological analysis.

The cosmetic product includes in itself ingredients which are traditionally used in our national medicine and some others which are less popular but still have a good perspective. For the base of the mixture Ostrich oil (Oleum Struthio) and Beeswax (Cera flava) were used. Their healthy effect was enforced with Almond oil (Oleum Amygdalae) and Wheat germ (Oleum Tritici germinis). The characteristic aroma was achieved with the use of Cinnamon oil (Oleum Cinnamomi) which also strengthens the bioactive effect. The nature of the used substances does not require the additional use of chemical improvers: emulators and preservatives. The preliminary microbiological test has confirmed the anticipated antiseptic effect.

Due to the non-specific purposefulness of the components the Vaseline is expected to have a wide-array of effect. From hydrating and protecting the skin from atmospheric influences, anti-wrinkling and anti-stretch marks effect, prevents skin irritations, removes scars, nurtures the skin, and also has and antioxidant and aromatherapy effect.


P84 ENHANCEMENT OF XYLANASE PRODUCTION BY SOL-GEL IMMOBILIZATION OF ASPERGILLUS AWAMORI K-1 MARIYA YORDANOVA1*, YANA EVSTATIEVA1, GEORGI CHERNEV2, SVETLA ILIEVA1, ROSITSA DENKOVA1, DILYANA NIKOLOVA1

1 Department of Biotechnology, Faculty of Biology, Sofia University St. Kliment Ohridski, Sofia 2 Department of Silicate Technology, Faculty of Metallurgy and Material Sciences, UCTM, Sofia, *Corresponding author: [email protected] Keywords: Aspergillus, xylanase, immobilization, sol-gel hybrids, alginate

Filamentous fungi from genus Aspergillus have displayed a great capacity to

produce hemicellulases used in pulp and paper industry and also for the production of fuels and chemicals. Technological potential of Aspergillus awamori species is of particular interest in the investigation of new methods for increasing the cell viability and xylanase activity.

The progress in bioencapsulation over the past decades has led to a revolution in the use of immobilized cells for production of extracellular enzymes in biotechnological manufactures. Compared to the other immobilization methods, sol–gel technique has many advantages due to simplicity of preparation of porous matrices with high thermal and chemical stability and flexibility of controlling pore size. The synthesis of solid inorganic materials from alkylsiloxanes such as tetraethylorthosilicate is alternative area being developed to improve the viability of encapsulated microorganisms. The use of organic additives such as calcium alginate for fabrication of hybrid matrices eliminates the cytotoxicity of the silica sol and increases the cell productivity after immobilization.

The purpose of the present investigation was to study the immobilization of Aspergillus awamori K-1 cells for higher xylanase production using sol-gel encapsulation in hybrid matrices composited by tetraethylorthosilicate as an inorganic precursor, 5% xylan and 5, 10 or 15% calcium alginate as organic compounds. The enzyme synthesis of immobilized cells under batch fermentation conditions were compared with xylanase activity of free cell culture used as control. According to the results culture immobilized in hybrid matrix composited by tetraethylorthosilicate and 5% calcium alginate was found to have xylanase activity of 28,88 IU/ml higher than enzyme activity of 22,04 IU/ml of the control. In contrast to free cell culture, immobilized cell retain their viability and biosynthetic capabilities during continuous fermentation processes. Acknowledgment: This work was supported by grant DMU 03/12 of National Science Fund, Bulgaria.


P85 NEW METHOD FOR IMMOBILIZATION OF TRICHODERMA VIRIDE SL-45

YANA EVSTATIEVA1*, MARINA BADALOVA 1, GEORGI CHERNEV2, VALENTIN SAVOV1, DILYANA NIKOLOVA1

1 Department of Biotechnology, Faculty of Biology, Sofia University “St. Kliment Ohridski”, Sofia 2 Department of Silicate Technology, Faculty of Metallurgy and Material Sciences, UCTM, Sofia, *Corresponding author: [email protected] Keywords: immobilization; sol-gel , Trichoderma; cellulase activity

Cellulase production from Trichoderma strains is regarded as the most suitable for hydrolysis of cellulose materials. Trichoderma viride is one of the best known cellulolytic organisms, producing technologically significant quantities of complete set of extracellular cellulases for the degradation of crystalline cellulose.

In this work Trichoderma viride SL-45 having defined cellulose profiles was investigated for specific applications. Vegetative culture of this strain was immobilized in four variations of sol-gel hybrid silica materials composited by tetraethylorthosilicate as an inorganic precursor and 10 % polyethylene oxide and 5, 10% carboxymethylcellulose as an organic compounds and substrate.

The relationship between strain immobilization, cellulase activity and matrices compositions was investigated. It was found that this method of immobilization had positive effect on strain activity. Biosynthetic activity of immobilized cultures was investigated by batch and fed–batch cultivation up to 840 hours. The production of cellulase from immobilized culture in hybrid matrix composited by tetraethylorthosilicate (as precursor) and 5% carboxymethylcellulose (as organic compound and substrate) was found to be about twice higher than activity of free culture of Trichoderma viride SL-45.

Acknowledgment: This work was supported by grant DMU 03/12 of National Science Fund, Bulgaria.


P86 STUDY OF THE ANTIBACTERIAL ACTIVITY OF BACTERIAL STRAINS FROM THE GENUS BACILLUS AGAINST THE PHYTOPATHOGENIC BACTERIUM XANTHOMONAS VESICATORIA

TSVETANA LICHEVA *, MARINA BADALOVA, VALENTIN SAVOV, YANA EVSTATIEVA, DILYANA NIKOLOVA

Sofia University St KlimentOhridski, Faculty of Biology,8”Dragan Tsankov” str., 1164 Sofia,Bulgaria *Corresponding author: [email protected] Kaywords: antibacterial activity, Xanthomonas vesicatoria, phytopathogenic bacteria

The aim of the study was to determine the activity of strains T 1, 2, 3, 4, 5, 10, 11, 12, 14, 15, 16, 17, 18, V 6, 7, 8, 13 and Bacillus pasteurii against the phytopathogenic bacterium Xanthomonas vesicatoria. The antibacterial activity of T 1, 2, 3, 4, 5, 10, 11, 12, 14, 15, 16, 17, 18, V 6, 7, 8, 13 and Bacillus pasteurii was tested by the agar diffusion method. A very good inhibitory effect against the test bacterium was detected after the 24th hour. It can be observed that the Bacillus strains T 2, 10 and 18 have the greatest activity of all tested strains (T 2 - 22mm, T 10 - 22mm and T 18 - 17mm) against the phytopathogenic bacterium. Strains T 4, 5 and 17 (T 4 - 10mm, T 5 - 5mm and T 17- 3mm ) are less active and strains T 1, 3, 11, 12, 14, 15, 16 do not show any antibacterial activity against the phytopatogen Xanthomonas vesicatoria. V8 has the greatest activity among the Bacillus strains V 6, 7, 8, 9, V 13 and Bacillus pasteurii (V 8 - 8mm). Strains T 2, 10 and 18 were chosen as active against the phytopatogen Xanthomonas vesicatoria and were identified. According to the biochemical tests (system BIOLOG) and the molecular characterization they belong to species Bacillus subtilis and Bacillus licheniformis (unpublished data).

The studied strains showed high antibacterial activity against phytopathogens and may have very wide application in plant production and agriculture. Organic farming excludes the use of synthetic chemicals and pesticides; therefore the use of biologically active substances on the basis of microorganisms against pathogenic bacteria is essential.


P87 ANTIMICROBIAL ACTIVITY OF LACTOBACILLUS PLANTARUM X2 AGAINST PATHOGENIC MICROORGANISMS

DENKOVA R.1*, S. ILIEVA1, D. NIKOLOVA1, Y. EVSTATIEVA1, Z. DENKOVA2, M. YORDANOVA1, V. YANAKIEVA2

1Sofia University „St. Kliment Ohridski”, Department of Biotechnology 2University of food technologies, Department of Microbiology *Corresponding author: [email protected] Keywords: antimicrobial, co-cultivation, pathogen, E.coli, Salmonella, Staphylococcus

One of the requirements for a strain to be included in the composition of probiotic preparations is to exhibit antimicrobial activity against pathogenic microorganisms. The antimicrobial action of the strain Lactobacillus plantarum X2, isolated from naturally fermented sourdough, against pathogens is examined through co-cultivation of the Lactobacillus strain and each of the pathogens: Escherichia coli ATCC 25922, Escherichia coli ATCC 8739, Salmonella abony NTCC 6017, Salmonella sp., Staphylococcus aureus ATCC 25093. It is observed that during separate cultivation of Lactobacillus plantarum X2 and each of the pathogens both Lactobacillus plantarum X2 and the pathogen accumulate high concentrations of viable cells, but during co-cultivation Lactobacillus plantarum X2 maintains a high concentration of viable cells, while the number of living cells of the pathogen is reduced. The degree of reduction of the cells of the pathogen is strain specific and is partially due to the changes in the acidity of the medium as a result of the production of acids by Lactobacillus plantarum X2.

P88 CLASSIFICATION OF LACTOBACILLUS DELBRUECKII SSP. BULGARICUS PHAGE GB1 INTO GROUP “B” LACTOBACILLUS DELBRUECKII BACTERIOPHAGES BASED ON ITS PARTIAL GENOME SEQUENCING

VANYA ALEKSANDROVA, DANİELA ISHLİMOVA, ZOLTAN URSHEV *

LB Bulgaricum PLC, 12A Malashevska str, Sofia *Corresponding author: [email protected] Keywords: bacteriophage, Lactobacillus bulgaricus, Lactobacillus delbrueckii, yoghurt

The modular evolution of phage genomes makes their classification extremely difficult. Still in dairy industry the identification of the phage species that disturbs the fermentation process is of crucial importance in the selection of a strategy to avoid or eliminate phage infections. Phages attacking Lactobacillus delbrueckii are currently divided into four groups assigned as “a”, “b”, “c” and “d” which have


substantial differences in their virion morphology and DNA homology. L. delbr. ssp. bulgaricus phage Gb1 was isolated in 2005 from yoghurt production. Restriction fragments of phage Gb1 DNA were cloned into pIL253 in Lactococcus lctis IL1403 and PCR amplified with vector-specific primers. The resulting amplicons were sequenced and identities with existing sequence databases were searched. All of the obtained sequences showed homology only to regions within the genome of phages LL-Ku and c5 with levels of nucleotide identity of 80-99%. No significant sequence matches were found with other L. delbrueckii phages. The sequenced fragments from the genome of phage Gb1 were homologous to key genes in phages LL-Ku and c5 encoding a major head protein, the tape-measure protein, tail proteins, the single strand binding protein and a diesterase. As phages LL-Ku and c5 are typical representatives of group “b” L. delbrueckii bacteriophages that do not share DNA homology with L. delbrueckii phages from other groups we could classify phage Gb1 into group “b” L. delbrueckii phages. This is the first report of a bacteriophage from this group isolated in Bulgaria. Further studies of phage Gb1 will facilitate the work on the selection of phage resistant starters and prevention of phage infections in yoghurt production.

P89 STARTER CULTURES FOR PRODUCTION OF YOGHURT FROM SHEEP’S MILK WITH EXTENDED SHELF LIFE

TEODORA ZVANCHAROVA, KALINKA BALTOVA*, ZOLTAN URSHEV

LB Bulgaricum PLC, 12A Malashevska str, Sofia *Corresponding author: [email protected] Keywords: Lactobacillus bulgaricus, proteolysis, sheep’s milk, shelf life, yoghurt

Originally Bulgarian yoghurt was prepared from sheep’s milk, but the home-made product had only a few days of shelf life. Today under industrial conditions yoghurt with shelf life of 20 days is routinely produced. Nevertheless producers and exporters are still interested in sheep’s milk yoghurt with even longer storage period. In this study we produced yoghurt with three selected starters and followed up its quality during three months of storage at 5oC. All three starters had low postacidification in the products. The freshly produced yoghurt samples had a pH of 4,17-4,39 which decreased gradually, but after 90 days it was still in the range 4,12-4,30 which was within the acceptable range. Viable cells of Streptococcus thermophilus and Lactobacillus delbrueckii ssp. bulgaricus remained above the critical threshold of 108 and 106 cfu/g respectively for the whole period of 90 days. With the selected starters the accumulation of products of proteolysis remained low with a slight increase only during the first month of storage. The limited proteolisys and low postacidification by the starters was achieved by the selection of weakly proteolytic strains of L. bulgaricus and/or a fermentation process with final viable cells of L. bulgaricus not exceeding 107 cfu/g. Organoleptic tests did not reveal any adverse change of product taste and aroma of yoghurt samples at day 90. In


conclusion, production of sheep’s milk yoghurt with selected starters can give a product that will preserve its nutritional and biological value for as long as three months.

P90 FAST ACIDIFYING UREASE-DEFICIENT STREPTOCOCCUS THERMOPHILUS ISOLATE SHOWS SPONTANEOUS DELETION OF ITS COMPLETE UREASE OPERON

NADYA NINOVA-NIKOLOVA*, ZOLTAN URSHEV

LB Bulgaricum PLC, 12A Malashevska str, Sofia *Corresponding author: [email protected] Keywords: Streptococcus thermophilus, urease, urease-deficient mutant, urease operon

Classical starters for fermented milk contain Streptococcus thermophilus strains which are capable of degrading urea present in milk. The ammonium released from urea slows down the decrease of pH during fermentation. This problem may be overcome with the selection of urease-deficient (UD) variants of S. thermophilus. Spontaneous UD-mutants Ft3uD3 and Yt3uD3-1 were previously isolated from the population of S. thermophilus Ft3 and Yt3, respectively, which originate from industrial yoghurt starters. In this study PCR amplification of the genes present in the urease operon and comparison of the macrorestriction profiles with PFGE of the UD-derivatives and the parent strains was performed to explain the loss of urease activity. No amplification of the genes from the urease cluster was observed for the Yt3uD3-1 UD-derivative of Yt3, indicating that the whole urease operon was absent in this mutant strain. The comparison of the SmaI macroresriction profiles of Yt3uD3-1 and its parent strain indicated a 19 kb deletion in the UD-culture. Therefore we concluded that in the UD-mutant Yt3uD3-1 the whole urease operon was deleted. On the contrary, all genes from the urease cluster amplified in Ft3 and its UD-mutant Ft3uD3 with identical size of each amplicon. Also, no differences in the macrorestriction profile of Ft3 and its mutant Ft3uD3 were observed. Consequently the loss of urease activity in Ft3uD3 is most likely result of point mutation within the urease operon. As the stability of the UD-derivatives during serial passage is a key issue in the application of the obtained mutants, we assumed that deletion mutant Yt3uD3-1 is a better candidate as it cannot revert to its active urease phenotype. Nevertheless, both S. thermophilus UD-derivatives Ft3uD3 and Yt3uD3-1 were stable for at least 150 passages. These results confirm the reliability of S. thermophilus UD-strains for industrial application.


P91 APPLICATION OF MODERN INFORMATION TECHNOLOGIES IN BIOLOGY TEACHER TRAINING ASYA ASENOVA

Sofia University “St. Kliment Ohridski”, Faculty of Biology, Department of Biology Education, 8 Dragan Tzankov Blvd, 1164 Sofia, Bulgaria e-mail: [email protected]

Keywords: Interactive whiteboard, tablet, training approaches, virtual practicum SimAula

Today, technological trends in informatics and telecommunications are behind a new change in educational service provision—from e-learning to mobile learning (m-learning). New technology provide users with opportunities to communicate and access information, learning content and services everywhere, at any time. Their use in the process of professional training of students enables universities to offer flexible training and better quality. In this paper main goal is to present several modern technologies and their educational opportunities. This article presents an idea how to organize education process whit new technology. Finally, it considers a platform providing opportunities to students to diversify learning according to their own needs using text, audio, and video educational materials.

P92 THE ESSAY AS A TOOL FOR MOTIVATION AND EVALUATION OF STUDENTS (FUTURE TEACHERS IN BIOLOGY)

KAMELIA YOTOVSKA*, АSYA ASENOVA

Sofia University St. Kliment Ohridski , Methodology of Biology Teaching, Faculty of Biology, 1164 Sofia, 8 Dragan Tsankov Blvd. *Corresponding author: [email protected] Key words: essay, evaluation, motivation

The essay as a literary form is released of the restrictions of genre frames. It gives a possibility for reasoning on the causal relationships, and life and science examples, comparison and / or opposition to the facts, signs, events, thoughts of famous people. The essay express the personal opinion, own idea and perspective on a chosen topic.

Writing the essay presents the student in greatest degree as an author who has a cognitive and affective style.

The application of skills to create an essay in the College and University allows diagnostic of students’ creativity.

In this article we describe the application of the essay at the University as a specific way of thinking and attitudes, as well as a tool for motivation and


evaluation of students from Sofia University St. Kliment Ohridski who are future teachers in biology.

Another topic in the article is a study regarding the student’s attitudes for and against essay as a form of assessment.

As an appendix to the article there are tools – a check list with the evaluation criteria and scale for assessing student’s essays. Clear regulation in the evaluation of the check list allows the students to self-assessment as they compare to the relevant criteria.

МЛАДЕЖКА НАУЧНА КОНФЕРЕНЦИЯ „КЛИМЕНТОВИ ДНИ” 22-23 НОЕМВРИ, 2012 Г. 83

INDEX ПО ПЪРВИ АВТОР

1. ATANAS GROZDANOV 5 стр. P1 2. A. KOSTADINOVA 26 стр. Р28 3. ABDULKADIR MAHDI ABUDALLEH 27 стр. Р30 4. ALBENA IVANOVA 5 стр. P2 5. ALEKSANDAR MEHANDZHIYSKI 26 стр. Р29 6. ANDREY MARCHEV 28 стр. Р31 7. ASYA ASENOVA 81 стр. Р91 8. BOGDAN GORANOV 31 стр. Р34 9. BORISLAV ARABADJIEV 54 стр. Р60 10. BORISLAV ASSENOV 30 стр. Р33 11. BOYKA ANDONOVA-LILOVA 29 стр. Р32 12. CHELENKOVA P. 53 стр. Р59 13. CHICHEVA Z. 52 стр. Р58 14. D. TODOROV 35 стр. Р39 15. DANIELA DIMITROVA 7, 18 стр. P4; Р20 16. DARINA DIMITROVA 36 стр. Р40 17. DASHA MIHAYLOVA 34 стр. Р37 18. DENKOVA R. 78 стр. Р87 19. DIANA IVANOVA 32, 37 стр. Р35; Р41 20. DIANA ZLATANOVA 7 стр. P5 21. DIMITRINA KOLEVA 33 стр. Р36 22. E. STOYNOV 9 стр. P6; Р7 23. EKATERINA H. PAVLOVA 34 стр. Р38 24. ELITSA IVANOVA 38 стр. Р42 25. EMIL V. BORISOV 10 стр. P8; Р9 26. EVA POPOVA 44 стр. Р49 27. G. CHIKOV 48 стр. Р54 28. GALYA PETROVA 56 стр. Р62 29. I. YOTINOV 69 стр. Р78 30. IGNATOVA-IVANOVA TS. 58 стр. Р64 31. INA ANEVA 11 стр. Р10 32. IRINA SCHNEIDER 71 стр. Р80 33. K. MILADINOVA 67 стр. Р75 34. KAMELIA YOTOVSKA 81 стр. Р92 35. KARAMFIL KALCHEV 41 стр. Р45 36. KIRIL VASSILEV 11, 12 стр. Р11; Р12 37. KIRILKA MLADENOVA 39 стр. Р43 38. LJUBA EVSTATIEVA 69 стр. Р77 39. LYUBEN ZAGORCHEV 45, 46, 47 стр. Р50;Р51;Р52 40. LYUBOV HRISTOVA 68 стр. Р76 41. LYUDMILA BELENSKA 41 стр. Р46 42. MARIA H. PETROVA 60 стр. Р67

МЛАДЕЖКА НАУЧНА КОНФЕРЕНЦИЯ „КЛИМЕНТОВИ ДНИ” 22-23 НОЕМВРИ, 2012 Г. 84

43. MARIA KERAKOVA 19 стр. Р21 44. MARIYA YORDANOVA 75 стр. Р84 45. MARKO KOLAKSAZOV 57 стр. Р63 46. MARTA ENCHEVA 12 стр. Р13 47. MIHAELA V. BELOUHOVA 25 стр. Р27 48. MIHAILOVA P. 21 стр. Р23 49. MILENA SHKODROVA 73 стр. Р82 50. MIRA TISHEVA 14 стр. Р15 51. MIROSLAV SLAVCHEV 20 стр. Р22 52. NADEZHDA GEORGIEVA 15 стр. Р16 53. NADYA NINOVA-NIKOLOVA 80 стр. Р90 54. NATALIYA GERGINOVA 74 стр. Р83 55. PAVLINA SASHEVA 59 стр. Р66 56. PENCHO PANDYKOV 22 стр. Р24 57. PEPA TSVETANOVA 23 стр. Р25 58. PETKO MLADENOV 58 стр. Р65 59. R. Z. VRANCHEVA 62 стр. Р69 60. RADOSLAVA EMILOVA 61 стр. Р68 61. ROSITSA TROPCHEVA 50 стр. Р56 62. RUSINA HAZAROSOVA 72 стр. Р81 63. S. LINCHEVA 70 стр. Р79 64. SASHKA KRUMOVA 64 стр. Р71 65. SASHO STOYANOVSKI 65 стр. Р72 66. SAVCHOVSKA M. 13 стр. Р14 67. STEFAN TSONEV 66 стр. Р73 68. STELLA DIMITROVA 63 стр. Р70 69. SVETLANA IVANOVA 51 стр. Р57 70. T. TOPOUZOVA-HRISTOVA 43 стр. Р48 71. TASINOV O. 42 стр. Р47 72. TEODORA IVANOVA 67 стр. Р74 73. TEODORA ZVANCHAROVA 79 стр. Р89 74. TOSHEVA A., 6 стр. P3 75. TSVETANA LICHEVA 77 стр. Р86 76. V. KALICHUK 48 стр. Р53 77. VANYA ALEKSANDROVA 78 стр. Р88 78. VASSILENA KRASTEVA 40 стр. Р44 79. VELISLAV ZAREV 17 стр. Р19 80. VELISLAVA SHISHKOVA 15 стр. Р17 81. VESELA NAYDENOVA 49 стр. Р55 82. VLADISLAV VERGILOV 16 стр. Р18 83. YANA EVSTATIEVA 76 стр. Р85 84. YOANA KIZHEVA 55 стр. Р61 85. YOVANA TODOROVA 24 стр. Р26

CARBOHYDRATE COMPOSITION AND ANTIOXIDANT ACTIVITY OF ROOT EXTRACTS OF INULA HELENIUM L.

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