Biotechnology & Biotechnological Equipment

Leuconostoc mesenteroides NRRL B-1149 produces extracellular dextransucrase which in this study was purified using different concentrations of polyethylene glycol (PEG). The dextran produced by this enzyme is unique in that it contains α - (1→6) and α-(1→3) linkages which have clinical applications. The cell free supernatant with 0.9 U/mg enzyme specific activity was subjected to fractionation by PEG-400 and PEG-1500. The 33% PEG-400 gave dextransucrase with specific activity of 9.2 U/mg and 10 fold purification and the 15% PEG-1500 gave dextransucrase with maximum specific activity of 15 U/mg and 17 fold purification in a single step. The purified enzyme showed multiple molecular forms on denaturing SDS-PAGE with three prominent bands. The purified dextransucrase confirmed the presence of glucan, after in-situ activity detection by Periodic acid Schiff's staining after running under denaturing SDS-PAGE. The three bands that appeared on denaturing SDS-PAGE stained with silver nitrate solution, corresponded to the three activity bands.

The influence of γ- rays on genetic variability is in direct correlation with the condition of irradiated material. Here we tried to increase the variability of Vicia faba L. var. major karyotype, after contamination with 131I (a kind of radioactive rain) in elongation phase in ten plant groups. The effects of irradiation were observed not only by controlling mitotic activity and calculating mitotic index in control and treated groups, but also by examination of every phase of mitotic and meiotic division. The values for normal and defect dividing cells are given in percentages. The most common types of aberrations were recorded during M1 generation.

The researches on the skeletal elements astragalus and calcaneus have been made in order to determine differences among deer (Cervus elaphus L) populations. Specimens were taken from several Neolithic localities on the territory of Balkan Peninsula. Possible differences among the populations are possible due to different geographic and ecologic factors at the research localities, and due to influence of artificial selection. The artificial selection is evidenced by the choice of game, taking into consideration usability of deer as a hunting game for Neolithic people. Statistic analysis has shown that there is variability among individuals of the species Cervus elaphus L. at the localities. However, there are no statistically significant differences among the populations. The individual variations among wild populations have been confirmed, while the most significant anthropogenic influence is evidenced by prominent selection of hunting game. Scatter analysis has shown that there is significantly larger number of male individuals, in comparison to female ones, in the specimen. This data is in correlation with the already mention fact that usability of male individuals is greater.

During the parasitological investigations on the gills of Leuciscus cephalus albus from Lake Ohrid (Macedonia), are found 4 monogenean species: Dactylogyrus sphyrna, Dactylogyrus folkmanovae, Dactylogyrus vistulae and Paradiplozoon ergensi. The total prevalence of infestation is 58,0% and the highest prevalence is of Dactylogyrus sphyrna (found in 26,0% of chubs). The average intensity of infestation is 5,50, and the highest level is that of Paradiplozoon ergensi. All monogenean species mentioned in this study represents a first record for the fishes from natural lakes in Macedonia, with the exception of Paradiplozoon ergensi, previously found in the Lake Ohrid. Among the monogenean species, found out in Lake Ohrid, the greatest pathological influence is associated with Dactylogyrus vistulae and Paradiplozoon ergensi.

The extracellular enzyme cyclodextrin glucanotransferase produced by Paenibacillus macerans ATCC 8244 cells was immobilized by covalent linking via glutaraldehyde to silanized magnetic nanoparticles treated with PEI and on cellulose- coated magnetite microparticles in order to study the immobilization capacities of the carriers, the activity recovery and some biochemical characteristics of the enzyme after immobilization. These biocatalysts were used for an efficient production of cyclodextrins from starch in repeated-batch runs, retained more than 52–83% of their initial activity and were stable after 60 day storage at 4°C. No significant increase of pH stability profile was observed, whereas the thermal stability of immobilized GTase was superior to that of free CGTase.

The recent advancement in the field of biomedical technologies has opened up new possibilities in the treatment of autoimmune disorders such as multiple sclerosis (MS). Here, we report the production of soluble complexes between the myelin oligodendrocytic glycoprotein (MOG) 79–90 peptide and a genetically engineered murine MHC class II molecule Aq. Using mouse model of MS, we demonstrate that the generated complexes are functional and able to ameliorate the clinical signs and reduce the incidence of experimental autoimmune encephalomyelitis (EAE). Our findings offer a new possibility for the treatment of chronically active autoimmune inflammation such as multiple sclerosis.

HPV infection is a major pathogenetic factor in cervical carcinoma as well as in many of the squamous cancers of head and neck and other epithelial cancers. Persistence of HPV DNA detectable by routine methods is considered to be a risk factor for advanced CIN and, in patients treated by surgery or non-surgical treatment modalities (radiotherapy, chemotherapy), HPV persistence is believed to be associated with increased risk for local recurrence. In terms of survival, however, it has been repeatedly proven that patients with cervical cancer and other HPV-associated cancers with detectable HPV DNA tend to have better outcomes than patients with HPV-negative tumours. The P72R polymorphism in the human TP53 gene has been contemplated as an independent phenotype modifier in cancers, especially the R allele which has been shown to confer higher pro-apoptotic properties to the resultant p53 protein. It has been demonstrated, however, that RR homozygotes were much more common in study groups with HPV-associated tumours than the other two genotypes and that the P allele in P/R heterozygotes was preferentially lost while the R allele was preferentially retained and mutated. It is possible that HPV-dependent carcinogenesis strictly relies on the presence of HPV and the expression of the E6 and E7 oncoproteins only in the initial phases of transformation of infected cells (e.g. CIN). It may be associated with activation of latent HPV that would create a background of decreased control over the integrity of the genome of the host cell. The process can develop further by mechanisms independent of the presence of HPV and if the virus clears at some later point, that would not halt the already ongoing neoplastic transformation. Absence of HPV DNA in cervical tumours, whether before or after treatment, is not a reason to decrease vigilant monitoring and rule out the need for further treatment, as it may be quite possible that the TP53 gene of the infected cells has already been modified in the course of cancer progression by HPV-independent mutagenesis. Cervical tumours that are HPV-negative ought to alert attending oncologists for the possibility for increased growth potential and invasiveness of the tumour so as to contemplate more aggressive anticancer therapies, especially in carriers of the R allele of the P53R polymorphism.

The effect of landscape structure on distribution of the Günthers vole in typical and mostly widespread habitats in the region of Strandja Mountain was studied. In the studied region, which represents the most northeastern part of the range of voles of guentheri group in Europe, general trends in distribution and biotopic adherence of M. g. strandzensisin have been outlined. The established distribution of Microtus guentheri in the studied biotopes revealed the presence of Günther's vole only in wet habitats with hydrophilic vegetation. The established relative numbers of the Günther's vole in natural and semi-natural habitats and agricultural areas under autumn crops determined this species as rare in Strandja region, while in the preferred habitat in natural wet zones along Veleka River it was presented as common species. It was recommended to carry out regular examinations of M. g. strandzensis in the region, focused particularly in clover and wheat fields and their neighbouring roadsides; species numbers in these habitats should be used as indicator of its population state.

The former uranium mine Senocos (Blagoevgrad district, Bulgaria) has been exploited until 1991 when it was closed and later (1994–1997) reclaimed. Recently, the reclamation is compromised due to the erosion of protective layers and the mine wastes continue to affect the area resulting in increased radiation and uranium concentration in mine area. Soil bacterial abundance in more radioactive environment of mine remains unchanged (on average ((8.52±5.1) x 108 cell g−1) compared to the control ((8.76±3.5) x 108 cell g−1) in contrast to their dehydrogenase activity which decreases more than two times. Physiological and species diversity of bacterial mine communities are also affected by the pollution as it is expressed by lower values of Shannon indices compared to the control communities. All carbon sources in BIOLOG assay except 2-hydroxy benzoic acid are available for control bacterial communities and the number of unavailable carbon sources increases linearly with increasing the concentration of uranium in soil. The high levels of radionuclide pollution decrease the biodiversity to 3–4 species (66% of total) in the most polluted point of the mine territory (SPS(5)).

Forty-two Lactobacillus strains isolated from „Lukanka” sausage were examined on the level of growth monitored by optical density within 72 hours spread on twelve time classes. The range of the OD comprises recorded values within the range of minimum (0) and maximum (2.31). Standard deviation of the observed OD varies between. The pH level of the cultures with recorded OD600 comprises values between minimum and maximum (3.60 to 6.00) with standard deviation (0.84). We applied a linear regression models in the study for statistical analysis of the variation of the recorded traits considered with the categories of the measured fixed effects of strain and hour. The GLM estimates of the recorded ODs and the induced acidity have high significance of the variances (F-statistics, P <0.001). The differences between the averages of the categories within the strains and hours give us information about the possible variation dependences at following significance.

The antioxidant potential of methanol extracts and ethyl acetate fractions of 10 Artemisia species was investigated using the 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radical scavenging assay. The screening of all extracts showed that the ethyl acetate fractions possessed the highest antioxidant activity. The fractions of A. vulgaris, A. campestris, A. annua, A. alba, A. austriaca were the most active and their scavenging potencies as indicated in IC50 values, being 11.96, 12.50, 15.85, 30.79, 37. 35 μg/ml, respectively. Among the methanol extracts these of A. pontica and A. annua (Bulgarian origin) exhibited the strongest antioxidant capacity—61.27 and 67.45 μg/ml. Five flavonoidglycosides were identified by TLC analysis.

The legal requirements stated in the EU Water Framework Directive (WFD) have led to increased activity concerning macrophyte indicator metrics in Europe. Nevertheless in Bulgaria such indexes are still under consideration because of the lack of official methodologies. The macrophyte surveys undertaken as a part of two EU-funded projects are a unique resource allowing aquatic plant communities to be studied. The Reference Index (RI) was chosen as macrophyte assessment method since classifies rivers by using regional approach and reflects different kinds of environmental pressures. RI was implemented on 73 sampling sites in Bulgaria. On the basis of this dataset, our attempt to redesign the RI by the addition of further species, and the re-grouping of existing species, resulted in a considerable improvement in relation with regional conditions. The chosen macrophyte method can be applied at Bulgarian rivers in order to assess their ecological status.

The genome response of model invertebrate and vertebrate species to stress agent in the environment was studied. The both group of species were collected from contaminated water stations along Marisa (Kemera) and Chaya (Asenovgrad) Rivers and terrestrial areas near Asenovgrad. The structural and functional alterations of the salivary gland chromosomes of Chironomus riparius as well as the aberrations of mitotic chromosomes of Apodemus flavicollis and Microtus arvalis were analyzed. In the polytene chromosomes of C. riparius a high spectrum of somatic aberrations were detected, which appeared in significantly higher frequency in comparison of the control (Kemera: G=37.99, df=1, P<0.001; Asenovgrad, G = 42.82, df = 1, P<0.001). Also, the key structure of the polytene chromosomes: Balbiani rings and Nucleolar Organizer decreased their normal function activity which indicates that they are direct target for the stress of contaminants in the environment. The frequency of aberrant cells of small rodents (Apodemus flavicollis and Microtus arvalis) was significantly higher than the control, well expressed in M. arvalis (G = 4.54, df =1, P <0.05). In both groups of studied species genome instability were found, realized by many structure chromosome rearrangements which can be used as a cost-effective indicator of genotoxicity, and hence suitable markers of potential environmental stress. However, the studied species show differences in their response which might be depended on their biology.

Airpollution investigations have not been done in Knjaževac until now. In this work, method of bioindication by lichens has been used. On 18 investigated points it has been found 22 lichen taxa from 12 genera. By calculating Index of Atmospheric Purity (IAP) 2 zones with different air pollution (”lichen desert” and “transitional” zone) has been found. The most sensitive species in Knjaževac are Buelia punctata, Melanelia acetabulum, M. exasperata, and Parmelia tiliacea and most tolerant are Lecanora alophana, L. intumescens, L. pulicaris, Lecidella elaeochroma and Phaeophyscia orbicularis.

Antimicrobial activity of two trade lots of essential oils from Rosa alba L. against Gram-positive bacteria, belonging to genera Staphylococcus and Bacillus, Gram-negative bacteria, belonging to genera Escherichia, Pseudomonas, Salmonella and Citrobacter and yeasts, belonging to genera Candida was investigated. It was determined that Gram-positive bacteria were more sensitive and Gram-negative bacteria, especially Pseudomonas spp., were more resistible to essential oils and pure components. The major pure constituents citronellol, geraniol and nerol demonstrated higher antimicrobial activity in comparison with essential oil samples.

Major Histocompatibility complex (MHC) plays a key role in immune response. We intend to study the polymorphism in two microsatellite loci of MHC complex, for two populations of Shkodrane sheep breed, whose location is North of Albania. The study was carried out in 113 unrelated individuals. Both markers were highly polymorphic. The analysis revealed a great number of alleles 14 and 40 per OMHC1 and OLADRB respectively. The level of heterozygosity was very high: 80% for OMHC1 and 60% OLADRB. This can be explained with the participation of the molecules encoded by MHC complex in the immune response, and with the balanced selection which act on antigen recognition site. PIC values were higher than 0.5. Therefore, both loci can be used as genetic markers.

Recka is an autochtonous sheep breed in Albania. We intend to analyze the genetic diversity of Recka sheep breed, by the use of 15 microsatellite markers. Allelic and genotypic frequencies, heterozygosities and gene diversity were estimated. A total of 143 alleles were distinguished by the 15 microsatellite markers used. All the microsatellites were highly polymorphic, with mean allelic number of 9.53, ranging 5–15 per locus. The observed heterozygosity ranged between 0.625 to 0.968, with mean of 0.785, indicating high genetic variation in this breed. PIC values were higher than 0.5, indicating that the set of used markers was highly informative. It was noticed a low rate of inbreeding within breed (FIS = 0.013). The results suggest that this set of loci was very effective for testing genetic variation. This is the first report of microsatellite variation in Recka sheep breed in Albania.

This research tested the rearing method of Queen Bees according to Jenter's. Two consecutive planting were realized, respectively on June 24 and July 12, each of these time-periods representing one stage of the experiment. To implement this research, the strongest bee-colonies of the apiary were used. During each stage of experiment 10 Queen Bees were produced. Their quality was judged and decided based on their growth parameters and performance. During each stage, the percentage of larvae's sealing, young Queen-Bees emergence, their time of copulation, their body size, the number of laid eggs were surveyed and measured. At the end of the research, it was concluded that Queen-Bees rearing parameters were better on the first stage of the study, all this due to the sufficient food availability and favourable climate conditions during this period of time. No significant differences were observed for the copulation time between two stages. During the second stage, the egg production is 27.7% lower than the first stage. The differences in value for the number of laid eggs in each stage are significant. The biggest size of the body-length and the maximum body weight of the Queen?Bees were reached during the June-July period (respectively 271.5 mg and 2.01mm). However, these parameters have been within the limits during the period of study.

The genus Lamium L. (Lamiacea) comprises about 40 species of annuals and perennials distributed in Europe, Asia and Africa. Lamium album L. possesses astringent, antispasmodic, anti-inflammatory, antibiotic and bacteriostatic properties. The effect of auxin IBA (Indole-3-butyric acid) on micropropagation of Lamium album L. has been investigated. On IBA-supplemented MS medium all concentrations stimulated the length of shoots, average number of leaves and roots. IBA did not influence number of shoots, callusogenesis and dry weight of in vitro propagated plants. To the best of our knowledge, this is the first report on in vitro multiplication of Lamium album through direct plant regeneration technique offers an effective alternative method of propagation for this important multipurpose medicinal plant.

Extracellular and membrane-bound alkaline phosphatases were produced at the middle stationary phase of growth by a strain Bacillus cereus. Twenty two percent of the enzyme activity was secreted into the culture media. An extracellular alkaline phosphatase (AP I) and a membrane-bound alkaline phosphatase (AP II) were purified 282-fold and 70-fold, respectively by a combination of chromatographic methods. Enzyme activity of alkaline phosphatase preparations was maximal at pH 9.5. Both enzymes were inhibited by EDTA and were reactivated by addition of Ca2+. The molecular weight of AP I was estimated to be 43 ± 1 kDa, and that of AP II was estimated to be 44 ± 1 kDa. Alkaline phosphatase activity of both enzyme preparation s was completely lost by heating at 80°C.

Formaldehyde (F) is a ubiquitous chemical agent, part of our working and residential environment. It is a well-known mucous membrane irritant and a skin sensitizing agent associated with contact dermatitis. The concomitant exposure to formaldehyde and glutaraldehyde is common in dental practice. The aim of the present study was to perform a comparative overall evaluation of the health status of subjects with formaldehyde and glutaraldehyde-specific allergic contact dermatitis, in occupational and nonoccupational exposure. A detailed and intentionally conducted questionnaire survey was performed among a total of 78 participants: 30 dental professionals occupationally exposed to formaldehyde, and 48 non-exposed healthy subjects (referents). All participants were clinically assessed, including a review of medical summary of history and physical examination. Skin patch testing with formaldehyde and glutaraldehyde was performed and a possible co-reactivity was assessed. The general incidence of formaldehyde-specific allergic contact dermatitis was 28.2%, and of glutaraldehyde-specific allergic contact dermatitis, 24.4%. The results demonstrated a stronger irritant effects of formaldehyde compared to glutaraldehyde, expressed in significantly higher incidence and relative risk of subjective upper respiratory tract (p = 0,029, OR=3,049 {1,102-8,442}), respiratory system (p = 0,002, OR=4,916 {1,699-14,228}) and skin (p = 0,019, OR=3,314 {1,191-9,218}) symptoms among subjects with formaldehyde-specific contact dermatitis. The gender analysis showed a significantly higher frequency and relative risk for sensitization to formaldehyde among women (x2 = 4,371, p = 0.037, OR = 3,219 (1,053-9,838). Women with formaldehyde-specific allergic contact dermatitis seem to be a vulnerable group for clinical upper respiratory tract alterations. No increased incidence of upper respiratory tract infectious pathology among subjects with allergic contact dermatitis was established. Tobacco smoking seems to be a risk factor for sensitization to formaldehyde, but not to glutaraldehyde. Allergic predisposition is a risk factor for sensitization to glutaraldehyde, but not to formaldehyde. On other hand, its role for co-reactivity was categorically demonstrated. Studies of this kind, including patch-testing for more dental materials compounds, would be beneficial for early diagnosis of contact allergy, as well as risk assessment and management.

Genetic variation of honey bee populations from six different locations corresponding to tree geographic regions: Northwestern, North-central and North-eastern of Bulgaria was studied on 6 enzymic systems (MDH, ME, EST, ALP, PGM and HK) corresponding to 6 genetic loci. Allozyme analysis revealed that all loci studied were polymorphic in almost all populations studied. The mean number of alleles per locus varied from 1.8 to 2.5. The estimated percentage of polymorphic loci was between 50% and 100%. The observed and expected heterozygosities (Ho and He) ranged from 0.17 to 0.221 and 0.250 to 0.315, respectively. There are not significant deviations of genotype frequencies from Hardy-Weinberg expectations at most of the loci in most populations (0.99 > P > 0.1). The estimated mean FST value from allozyme data was 0.0443 which shows that 4.43% of the overall genetic diversity observed was among populations, as opposed to 95.57% within populations. The values of genetic distance range from 0.002 to 0.036. UPGMA dendrograms were constructed.

The leaf epidermis structure of Amelanchier ovalis Medic. was studied by light and scanning electron microscopy. Rodlets of crystalloid epicuticular waxes and papillae were observed. Simple single trichomes and cyclocytic stomatal type were described. An attempt was made to estimate all features of taxonomic value which are relevant for further clarification of the relationships within subfamily Maloideae. The epidermal structure shows adaptation to various environmental conditions which explains the broad geographical distribution of the specie.

A study on the viability of pollen grains and embryos in Gentiana lutea distributed in Bulgarian flora has been carried out. Degenerative processes predominantly of the mature pollen were observed. As a result of that, in the studied populations, the effective mature pollen grains in the anthers reduce in different amount. The used tetrazolium test shows that in the mature seeds the viability of embryos reduced progressively during the three consecutive years of the present study.

Continuous intoxication with heavy metals seems to be the major problem of the modern societies, which results most of the morphologic-physiological, as well as genetic anomaly in plants. The aim of this study is to analyze intoxicated tobacco material (Nicotiana tabacum L.) with heavy metals (copper, cadmium, lead) and overdoses of fungicide (Ridomil gold) for enzymatic (catalase activity) and biopigment activity (chloroplast pigments) and to make morpho-anatomic examinations (lateral cut of stem and leaf). The plants were from the crop 2008 of two oriental (half oriental) types: PRILEP (P-156) and JAKA (Jv-125/3). They were cultivated in controlled experimental conditions in the Botanical Garden by the Department of Botany, Institute of Biology, Faculty of Natural Sciences and Mathematics-Skopje. Analysis material was taken from leaves in the first insertion or lower leaves (I phase).

Honey bee's wings are on dorsal side and are attached to the second segment of the thorax. The system of chitin nerves runs trough them. The wing nervature is variable, but the layout of cells on the wings is not random, however, and is used for the determination of bees. Honey bee is one of the most studied insects with a total of 42 morphometric parameters used for the purpose of racial determination and taxonomy. The aim of this paper is to determine the morphometric parameters of the angles between the nerves of the front wing, which are: A4, B4, D7, E9, G18, J10, J16, K19, L13, N23 and O26, as well as to analyze the morphometric parameters on the rear wing of the honey bee?angles W1, W2 and W3). The bee samples were taken from the area of north part of Kosovo and Metohija on the sites Le?ak, So?anica and Kosovska Mitrovica. The samples were comprised of 10 bees each, and are indigenous material. Also, the variation coefficient and maximum and minimum value of the researched parameters were determined in this study.

Psychrophilic yeast Isolate 100, was selected as a producer of exopolysaccharide. A laboratory scheme was created for obtaining of the biopolymer. Biotechnological and physicochemical factors influencing its biosynthesis were investigated. It was determined that inocolum quantity from 6–10%, cultivated for 48 hours at 22° C, is suitable for maximum synthesis. The exopolysaccharide that was synthessed in the culture liquid was thermostable at 50°C to 70°C and it degrades at 80°C. The quantity of the sedimentary polymer, that was synthesis on environment with arabinose and mannose from Isolate 100, after incubation in ethanol at 4°C for 120 hours staying at the time of 120 hours in ethanol at 4 °C, was increased with 40%. The influence of the temperature over drying of the exopolysaccharide at 65°C and 105 °C was studied. At the low temperature of drying the polymer contained near 20% humidity. The effect of the experimental conditions (temperature, time, concentration of 2%, 3%, 4%, 5%) on the apparent viscosity values of the cultural liquid was studied. It was found to reveal a behaviour of non-Newtonian liquid. The power law equation of Oswald- de Waale describing the change of the apparent viscosity was derived.

The extract was prepared from fresh leaves of Jovibarba heuffelii (Schott.) A Löve & D. Löve and used to evaluate the antimicrobial activity against a panel of Gram-positive and Gram- negative bacteria as well as against fungi. Antimicrobial activity was evaluated using broth microdilution method. For determination of the total phenolic and flavonoid contents methanolic, ethyl acetate, acetone and diethyl ether extracts were prepared. The phenolic content of the extracts was determined spectrophotometrically using Folin—Ciocalteu reagent and the results are expressed as gallic acid equivalents (GAE). Jovibarba heuffelii extract showed moderate antibacterial activity, while its antifungal activity was significantly higher. The high contents of total phenolic compounds and total flavonoids indicated that these compounds contribute to the antimicrobial activity.

Oligosaccharides may alter the antimicrobial activity of lactobacilli strains cultivated in whey media. The utilization of galactooligosaccharides in whey media by Lactobacillus fermentum I-5 was performed. The quantity of the used galactooligosaccharides had a pronounced effect on the growth rate. It was found that concentrations of 15% galactooligosaccharides and 2% of whey proteins contribute to higher growth rate. The obtained results revealed again that the higher percent of oligosaccharides stimulate higher antimicrobial activity.

At current study the antimicrobial activity of Origanum vulgare, Satureja montana, Thymus vulgaris, Pimenta dioica and Syzygium aromaticum against psychrotrophic microorganisms, isolated from spoiled chilled meat products was investigated. MIC, MBC and MFC of the essential oils were determined both at 37° and 4°C. Antimicrobial activity of the essential oils retained unchanged at both temperatures. Among the tested psychrotrophic microorganism Gram-positive bacteria Brochothrix thermosphacta was the most sensitive strain and Gram-negative bacteria Pseudomonas aeruginosa was the most resistible. The results obtained expanded the possibilities for application of studied oils not only as flavour enhancers, but even as natural antimicrobials in chilled meat products.

The paper presents the results of antioxidant activity of water, methanolic, ethyl-acetate, acetone and petroleum ether extract from plant species Teucrium chamaedys L. (Lamiaceae). Antioxidative activity was determined in vitro, using the DPPH reagent by spectrophotometric method. Experimental values are expressed as IC50 values (mg/ml), and show that the tested extracts have high antioxidant activities, which range in scope from 341.08 mg/ml to 29.46 mg/ml. In the tested extracts of plant species Teucrium chamaedys L. quantitative composition of total phenols and flavonoids were determined by spectrophotometric method. Total phenols determined by Folin-Ciocalteu reagent and their amounts in the range of 30.39 mg/g to 169.50 mg/g (expressed as gallic acid equivalent, mg GA/g of extract). The amounts of flavonoids in plant extracts of Teucrium chamaedys L. are in the range of 16.67 mg/g to 87.17 mg/g (expressed as rutin equivalent, mg RU/g of extract).

The influence of Cd2+ on the antioxidant system of Shizosaccharomyces pombe DSMZ 70576 was studied during batch cultivation on YPD medium containing 1.0 mM Cd(NO3)2 versus a control variant without Cd(NO3)2. The Cd2+ was added to the medium on the 12th h of the process. Samples for determination of the cells' growth, carbon source consumption, ethanol production and antioxidant system's status, represented by the enzymes superoxide dismutase (SOD) and catalase (CAT), and the intracellular principal antioxidant glutathione, were taken along the 48 h cultivation. The presence of 1.0 mM Cd2+ under the test conditions repressed the cell growth and resulted in less production of ethanol. The SOD and CAT specific enzyme activities and glutathione concentration were estimated as 11.8 U mg−1, 1.7 U mg−1 and 0.57 mM mg−1 respectively. The SOD and glutathione values were 1,6 and 1.7 fold higher than the corresponding ones measured during growth without Cd2+, while the CAT activity decreased by 40%. These results indicate that glutathione obviously had scavenged efficiently the H2O2 produced, thus compensated the diminished CAT activity and probably maintained the H2O2 homeostasis of the culture under the stress conditions. Electrophoretical analysis of SOD and CAT in cell free extracts obtained from the strain cultivated in the presence of Cd2+ and in norma indicated the absence of Mn SOD enzyme under the stress conditions. Apparently, the presence of 1.0 mM Cd(NO3)2 leaded to inhibition of mitochondrial functions and Mn SOD operation in the cells of Shizosaccharomyces pombe.

The aphid transmissibility of seven Plum pox virus (PPV) isolates and the amino acid sequences of their coat proteins were analysed. Two aphid transmissible isolates PPV-A and PPV-P contained the DAG amino triplet, while DAL or NAG replaced this triplet in the coat proteins of non-aphid transmissible isolates. In addition, a sequence of 15 or 17 amino acids was deleted in the coat proteins of PPV-P48, PPV-P45 and PPV-II/7. Helper components with Mr. of 52 kDa were successfully purified from N. benthamiana plants inoculated with aphid transmissible and non-aphid transmissible isolates. The helper components of two coat-proteins-defective isolates mediated aphid transmission of non-coat-protein defective isolates, indicating that the helper components were biologically active.

The impact of programmed cell death (PCD) inhibitors on lesion formation and biochemical events in transgenic (ttr line) and non-transgenic (Nevrokop 1164) tobacco infected with Pseudomonas syringae pv. tabaci was tested. Programmed cell death in tomato cell culture was induced by Fumonisin B1 (FUM) and effectively abolished by the administration of protease inhibitors and lanthanum. Caspase inhibitors Ac-YVAD-CMK and Z-asp-CH2-DCB, serine protease inhibitor TLCK and LaCl3 were inoculated together with P. syringae pv tabaci in detached tobacco leaves or applied simultaneously with fungal toxin FUM in the tomato cell suspension. The results illustrate that cell death in normal Nevrokop 1164 and ttr transgenic tobacco at sites infected with P. syringae pv tabaci is apoptotic-like and caspase-like proteases and serine proteases are involved in lesion formation. The cell death in the lesions was accompanied with enhanced H2O2 accumulation and MDA production in, the wild type and with reduced levels of both metabolites in the transgenic line. Lanthanum, caspase inhibitors and TLCK reduced the amount of H2O2 and MDA in the affected lesions. FUM-induced cell death in tomato suspension cells was greatly suppressed by the application of protease inhibitors and lanthanum. The results indicated that apoptotic cell death occurs at bacteria inoculation of tobacco leaves, and in fumonisin treated tomato cells

The first animal with black traits to be studied was the Black-boned chicken. In China, we identified a flock of sheep with black traits. In the present study, the complete coding sequences of three genes, PSPH, SNRPA1 and TPM1, of Black-boned sheep (Ovis aries) were amplified using the reverse transcription polymerase chain reaction, according to the conserved sequence information for cattle and other mammals and known highly homologous sheep ESTs. The results showed that the Black-boned sheep PSPH gene encodes a protein of 225 amino acids, which has high homology with the PSPH proteins of seven species: cattle (99%), humans (94%), rat (93%), mouse (93%), African clawed frog (77%), Atlantic salmon (70%) and zebrafish (68%). The Black-boned sheep SNRPA1 gene encodes a protein of 255 amino acids, which has high identity with the SNRPA1 proteins of four species: cattle (99%), humans (99%), mouse (97%) and chicken (91%). The Black-boned sheep TPM1 gene encodes a protein of 284 amino acids, which has high homology with the TPM1 proteins of four species: cattle (82%), rabbit (82%), mouse (82%) and humans (81%). Using phylogenetic analysis, it was shown that the Black-boned sheep PSPH, SNRPA1 and TPM1 proteins are closely related to their cattle counterparts. The tissue expression analyses revealed that the Black-boned sheep PSPH, SNRPA1 and TPM1 genes were expressed in a range of tissues, including leg muscles, the kidneys, the skin, the longissimus dorsi muscle, the spleen, the heart and the liver. These data serve as a foundation for further insight into these three genes in this rare sheep breed.

More than 30 lactic acid bacteria strains were isolated from different sources (matsuns, cheeses) collected in various regions of Armenia. Four of them inhibit the growth of different test-organisms. Treatment of antibacterial agent with various enzymes established their nature of inhibition by protein bacteriocin or by hydrogen peroxide production. The isolated strains were evaluated according to their growth, acidity and antibacterial activities as well as according to their survival under simulated gastrointestinal conditions. All tested strains during both “stomach “phase and in gastro-intestinal tract phase kept large number of viable cells.

The production of glycosyltransferases (GTFs) from constitutive mutant strain Leuconostoc mesenteroides M2860 was studied. When grown in glucose medium in the absence of sucrose Leuc. mesenteroides M2860 produced low, but detectable GTF activity. Much of the GTF activity (81%) in sucrose grown cultures was located in the cell pellets. Extracellular and cell associated activities were determined when run on SDS-PAGE for in situ activity detection by periodic acid-Schiff s staining. In situ analysis showed single band corresponding to 180 kDa molecular size in supernatant and enzyme concentrate, received by cultivation on glucose media. The enzyme concentrate and supernatant fraction obtained by fermentation on sucrose media showed three bands corresponding to 180 kDa, 120 kDa and 86 kDa.

Research has been carried out of the main characteristics of phytoplankton communities in order to assess the ecological status and ecological potential of the types of lakes in Bulgaria, according to the requirements of WFD 2000/60/EC. Eighty lakes/reservoirs have been researched on the territory of the Republic of Bulgaria. The assessment was made on the basis of four main metrics (phytoplankton biovolume; Algae Groups Index; transparency, chlorophyll a) and three additional metrics (% Cyanobacteria; intensity of algal “bloom” and presence of toxic species). More than half of the researched lakes in Bulgaria are in compliance with the WFD requirements for good ecological state (high and good ecological status, maximum and good potential). A classification system for assessment of ecological status or potential has been developed using above-mentioned phytoplankton metrics. The existing 17 types of lake in Bulgaria are classified in two main groups: oligotrophic lake types and mesotrophic lake types.

Development of left ventricular hypertrophy (LVH) is a compensatory response to hemodynamic overload and echocardiography is a noninvasive, sensitive method to study the process of myocardial remodeling and functional changes. In this study we assessed in vivo the time-course of the left ventricular structural and functional changes after aortic banding and validated them by necropsy. Wistar rats were subjected to abdominal aorta banding (AAB) or sham operation. A control group of age- and sex-matched, were maintained and sacrificed in parallel with operated animals. Echocardiographic assessment was performed before and at days 10, 15, 20, 25, 35 and 45 post-operation. Some rats were euthanized at each time point to obtain body and organ weights. Echo left ventricular wall thickness (LVWT) increased progressively on the 10th and the 15th day, then there was a reduction on the 20th day, followed by a second marked increase on the 25th day and after 35th day it presented a plateau. On the 45th day LVWT in AAB rats increased to 35% compared with SH. The percentage of left ventricular fractional shortening was similar in different groups at each time point. These parameters remained stable in control groups. Echocardiographic parameters and actual whole heart mass were well coincided. Echo LVWT correlated well with actual whole heart mass (r=0.724, P < 0.001). We developed an easier model of significant LVH in rats, not requiring thoracotomy. The progression of myocardial hypertrophy is a time-dependent, stage developing process. The model is suitable to study the mechanisms governing the development of left ventricular hypertrophy for therapeutic benefit.

Different strains of phosphate-solubilizing bacteria (PSB) were isolated from the rhizosphere of different plants of Lahore District, Pakistan. The objective of the study was to explore the capabilities of PSB and evaluate their efficiency to enhance growth of sugarcane plants under greenhouse condition. The purified isolates were identified as Proteous vulgaris, Klebsiella pneumoniae, Enterobacter aerogenes, Burkholederi acepacia, Citrobacter freundii, Acinetobacter lwoffii, and Pseudomonas fluorescens, and the identification was based on the characteristic morphological and biochemical behavior. The efficiency of different PSB isolates for phosphate solubilization was evaluated from the zones they formed on agar plates of Pickovaskaya growth medium (PVK) by solubilizing the tricalcium phosphate of the medium. The efficiency of purified PSB was evaluated from pot experiments of two different genetically modified sugarcane varieties (SCMV resistant CAMB-I and CAMB-II) under greenhouse condition for their positive role in plant growth promotion. All the six PSB enhanced the growth rate significantly over that in the non-inoculated control. A significant increase in plant height, number of leaves, root length and dry matter contents was recorded and E. aerogenes and C. freundii were found significantly superior over the rest of the isolates for all the tested parameters. The efficiency gradient of different isolates for CAMB-I and CAMB-II varieties was recorded as C. freundii> K. pneumoniae> E. aerogenes> B. capacia> A. lwoffi> P. vulgaris and E. aerogenes> C. freundii> A. lwoffi> B. capacia> P. vulgaris, respectively. The results of this greenhouse evaluation are encouraging and need to be confirmed under field condition in combination with organic and chemical fertilizers.

The sequences of different genetic determinants of bacteriocins belonging to the classes of lantibiotics and pediocin-like bacteriocins were compared, and in a few cases it was possible to obtain consensus sequences for genes with identical functions. On the basis of these consensus sequences, fifteen primer sets were designed, and a screening of four bacteriocin-producing Enterococcus strains was performed, as well as of a collection of other Enterococci which have not been screened before for antibacterial activity. In the case of the four bacteriocinogenic strains, several amplifications with the expected sizes were obtained with some of the primers. A particular case represented two strains, one of them being bacteriocin producer, and the second one resistant to the bacteriocin produced by the first one. In this case, in both strains amplifications were obtained with primers specific to one of the immunity protein genes, a fact probably suggesting horizontal gene transfer of the immunity determinants.

In the present study, an indigenous biosurfactant-producing bacterium, Rhodococcus sp. BAP-1, with effective fluoranthene- degrading ability was isolated from crude-oil contaminated soil. The biodegradation data indicated that Rhodococcus sp. BAP-1 grew well in the presence of 20 mg-L−1 of fluoranthene, whereas in the presence of 150 mg-L−1 of fluoranthene, the biodegradation rate almost remained at low levels. During growth on mineral salts medium, the bacterium could produce biosurfactants. As a result of biosurfactant synthesis, the surface tension of the culture broth was reduced, thus facilitating the contact between the bacterium and hydrocarbon. According to the experimental results, emulsification was probably not the leading mechanism that occurred simultaneously during the biodegradation process. Pollutants might be growing as a confluent biofilm and the uptake system might have a high specific affinity; an interfacial uptake mechanism may play a predominant role in the biodegradation process. Fourier transform infrared spectroscopy showed that the biosurfactant contained aliphatic hydrocarbons. A disruption of the cell surface in certain zones was observed by scanning electron microscopy in some of the cells grown in the presence of 150 mg-L−1 of fluoranthene, which indicated that drastic changes of the cell surface morphology might occur under different growth conditions. The results of this study collectively suggest that Rhodococcus sp. BAP-1 may be useful in the bioremediation of environments contaminated with polycyclic aromatic hydrocarbons.

In this paper, a hybrid scheme using Bat Algorithm (BA) and Sequential Quadratic Programming (SQP) method is introduced. In the hybrid BA—SQP, the role of BA is to generate feasible solutions to a problem. The role of SQP is to exploit the information gathered by BA. This process obtains a solution which is at least as good as—but usually better than—the best solution devised by BA. To demonstrate the usefulness of the presented approach, the hybrid scheme was applied to parameter identification of an E. coli MC4110 fed-batch cultivation process model. A comparison with both the conventional BA and SQP method is presented. The results showed that the hybrid BA-SQP has the advantages of both BA's global search ability and SQP's local search ability, thus enhancing the overall search ability and computational efficiency. For comparison, the results obtained by applying Ant colony optimization algorithm in conditions similar to those of BA are further shown.

This study aimed to address the problems of small towns, e.g. low economy, undeveloped technology and management level, and fluctuant water quality and water quantity. To develop an efficient low-cost small-town sewage treatment technology, the integration of sequencing batch type bioreactor/ecosystem hybrid treatment process was chosen. In this experiment, the bioreactor unit of the process included a sequencing batch biofilm reactor (SBBR), while the ecosystem unit applied a sequencing constructed wetland (SCW) based on a polyurethane foam filler in a matrix. By changing the operation model, tests were carried out to find the key parameters of the optimal operation model for the sequencing batch type bioreactor/ecosystem hybrid treatment process in different seasons. The experiment was conducted throughout a year of operation after setup. The results showed that when the bio- and ecosystem reactors ran together in combination at a temperature of 15 °C ∼ 25 °C in spring and autumn, The final effluent chemical oxygen demand (COD), ammonium nitrogen (NH4+-N), and total nitrogen (TN) concentrations of the hybrid reactors were 48 mg/L, 7 mg/L, and 16 mg/L, respectively, with a corresponding total removal efficiencies of 86%, 89%, and 79%. When the ecosystem reactors ran independently in the temperature range of 25 °C ∼ 35 °C in the summer, the effluent COD, NH4+-N, and TN concentrations were 47 mg/L, 7.3 mg/L, and 17.3 mg/L respectively, with a corresponding total removal rate of 84%, 87%, and 74%. The bioreactors and ecosystem reactors ran together in combination at 5 °C ∼ 15 °C in the winter and the final COD, NH4+-N, and TN concentrations of the hybrid reactors effluent were, respectively, 54 mg/L, 11 mg/L, and 18 mg/L, with a corresponding total removal of 86%, 84%, and 76%. The research developed an integration of sequencing batch type bioreactor/ecosystem hybrid treatment process with important realistic significance and practical value. The designed operation models are able to be used in guiding practical engineering.

The East Palaearctic seed beetle Bruchidius terrenus (Coleoptera: Chrysomelidae: Bruchinae) is recorded for the first time to the Bulgarian fauna. Larvae of the bruchid infest mature seeds of introduced mimosa Albizia julibrissin (Fabacea), an ornamental tree in many countries. The level of damage on seeds caused by bruchid larvae was examined.

Whether behavior is present in the science for many years, it is still young and very attractive discipline that needs interdisciplinar approach for studying. Large scale of experimental models makes experimental and scientific work more exciting. At the Faculty of Biology, University of Belgrade (Serbia), we have behavioral courses at all three levels of studies (bachelor, master and PhD). At the basic level, we have Animal Behavior course, than, at the master level, we offer to our students two courses, Biology of Behavior and Introduction into Behavioral Genetics. Finally, for PhD students of Neuroscience we have two courses—Cellular Basis of Behavior and Neurobiology of Behavior, while PhD students of Genetics attend course Behavior Genetics. In this way, all aspects and levels of understanding behavioral mechanisms are covered.

Molecular karyotyping is an extremely suitable method for genetic diagnosis of patients with unclear dysmorphic syndromes and intellectual disability. In this study we present our results from microarray analysis of 52 patients with developmental delay and congenital malformations. Our data revealed definite etiology in 9 out of 52 patients tested. Fifteen pathological aberrations were found in them. All pathological findings were validated by fluorescent in situ hybridization (FISH) analysis. In addition, the majority of the patients tested (41 patients) showed normal variations in the number of copies and variations of unknown clinical significance (34 patients). Analyses of the type and distribution of the different variations was performed and the clinical significance of variants of unknown nature was discussed. Our results show the advantages of high resolution microarrays for clinical diagnosis of patients with intellectual disability and congenital malformations, and also highlight the need for extensive population studies revealing the molecular nature and clinical significance of different copy number variations and for creation of detailed maps of variations in the Bulgarian population.

The immobilized lipase-catalyzed transesterification of Jatropha curcas L. seed oil and methanol for biodiesel production in tert-butanol was investigated. The effects of different tert-butanol volume, methanol molar ratio, reaction temperature, reaction time and immobilized lipase amount on the total conversion were systematically analyzed by response surface methodology (RSM). RSM analysis showed good correspondence between experimental and predicted values. The optimal conditions for the transesterification were a reaction time of 17.355 h, a reaction temperature of34.868°C, an immobilized lipase amount of 12.435%, a methanol molar ratio of5.282:1, a tert-butanol volume ratio of 0.577:1. The optimal predicted yield of fatty acid methyl esters (FAME) was 88.5% and the actual value was 88.1%. The predicted yield of fatty acid esters and the real one was very close, indicating that the RSM based on central composite design (CCD) was adaptable for a FAME study for the present transesterification system. Moreover, the infrared spectum of biodiesel showed the characteric bands of C=O, O—C—O, C=C and—(CH2)n—. Furthermore, GC-linked mass spectrometry showed that biodiesel was mainly composed of the methyl esters of hexadecanoic, 9,12-octadecadienoic and 9-octadecadienoic acid.

The aim of the present study was to investigate biofilm-forming capabilities of clinically isolated strains Escherichia coli, associated with different urinary tract infections. Biofilm production was detected in 36% of the isolates from UTI. Additionally some of virulence factors are estimated to find correlation between antibiotic resistance, hemolysins, morfotypes and biofilm production. Our data indicate that no combination of VFs was highly associated with biofilm production.

The results of a comparative study of two biogas production bioprocess systems are presented. The systems submitted to comparison are based on the suspended cells cultures and the biofilm formed on solid inert support. A comprehensive research concept is formulated and discussed. It includes the main considerations regarding the choice of substrate, bioagent as mixed microbial society, type of bioreactors, regimes of functioning, analytical determinations and method of comparison. The main requirements for efficient experimental activity in comparative investigations are formulated. Their satisfaction can grant correctness of the experimental design and data acquisition. On this basis the key parameter of comparison of the two systems is defined as the specific productivity of the bioprocess systems. Under these conditions series of preliminary experiments are carried out for testing the readiness of experimental set ups for long time stable functioning and monitoring devices capabilities to maintain the bioprocess parameters at the determined intervals. These tests grant continuous incessant experimentation with the investigated bioprocess systems. The results obtained show that biofilm bioprocess systems possess up to two and half time higher specific productivity in comparison with the bioprocess systems with the suspended cells. Some visions about the future developments of comparative research on the influence of additional parameters like the mixer rotation steed, organic loads, and higher values of dilution rates are outlined.