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R. Bešta-Gajević, E. Karalija, A. Jerković-Mujkić, Dženana Karadža, Lejla Smajlovic-Skenderagic, Sabina Dahija

Antifungal, antimicrobial, insecticidal and antioxidant activities of Origanum vulgare L. provide the basis for suggesting that oregano plant extracts may be useful for prevention and treatment of many infection. The main goal of this study was to determine antimicrobial and antioxidant activities of methanolic and aqueous extracts from the leaves and flowers of Origanum vulgare. Antimicrobial testing of plant extracts was done using well diffusion method. Activity of extracts were tested against Gram positive bacteria: Staphylococcus aureus ATCC 25923, methicillin-resistant Staphylococcus aureus (MRSA) ATCC 33591, Bacillus subtilis ATCC 6633, Enterococcus faecalis ATCC 29212 and five Gram-negative bacteria: Salmonella abony ATCC 6017, Salmonella enterica serovar Enteritidis ATCC 31194, Pseudomonas aeruginosa ATCC 9027, Escherichia coli ATCC 25922, extended-spectrum β-lactamase (ESBL) producing Escherichia coli ATCC 35218 and fungi Candida albicans ATCC 1023. Antibiotics ampicillin, streptomycin and antimycotic nystatin were used as positive controle. The antioxidant activity was determined by using the DPPH (1,1-diphenyl-2-picrylhydrazyl) method. The highest values for inhibition zone for methanolic and aqueous extracts were recorded for Gram positive Bacillus subtilis, Staphylococcus aureus and MRSA. Methanolic extracts exhibited antibacterial activity against tested Gram negative bacteria in variable degree while the growth of these bacteria was not inhibited by aqueous extracts. Tested fungi Candida albicans was not susceptible to investigated oregano extracts. All the extracts showed moderate to potent antioxidant activity, among which the methanolic flower extract demonstrated the strongest antioxidant activity with the IC50 value of 0.205 mg/mL. Therefore it can be concluded that flower and leaf oregano extracts have great antibacterial and antioxidant potential.

R. Bešta-Gajević, E. Karalija, A. Jerković-Mujkić, Dženana Karadža, Lejla, Smajlović-Skenderagić, Sabina Dahija

Antifungal, antimicrobial, insecticidal and antioxidant activities of Origanum vulgare L. provide the basis for suggesting that oregano plant extracts may be useful for prevention and treatment of infections. The main goal of this study was to determine antimicrobial and antioxidant activities of methanolic and aqueous extracts of the leaves and flowers of O. vulgare. Antimicrobial testing of plant extracts was done using well diffusion method. Activity of extracts was tested against Gram positive bacteria: Staphylococcus aureus ATCC 25923, methicillinresistant Staphylococcus aureus (MRSA) ATCC 33591, Bacillus subtilis ATCC 6633, Enterococcus faecalis ATCC 29212 and five Gram-negative bacteria: Salmonella abony ATCC 6017, Salmonella enterica serovar Enteritidis ATCC 31194, Pseudomonas aeruginosa ATCC 9027, Escherichia coli ATCC 25922, extended-spectrum β-lactamase (ESBL) producing Escherichia coli ATCC 35218 and fungus Candida albicans ATCC 1023. Antibiotic ampicillin and antimycotic nystatin were used as positive control. The antioxidant activity was determined using the DPPH (1,1-diphenyl-2-picrylhydrazyl) method. The highest values for inhibition zone for methanolic and aqueous extracts were recorded for Gram positive B. subtilis, S. aureus and MRSA. Methanolic extracts exhibited antibacterial activity against tested Gram negative bacteria in variable degree while the growth of these bacteria was not inhibited by aqueous extracts. Tested fungus C. albicans was not sensitive to investigated oregano extracts. All the extracts showed moderate to potent antioxidant activity with methanolic flower extract demonstrating the strongest antioxidative activity with the IC50 value of 0.205 mg/mL. Therefore it can be concluded that flower and leaf oregano extracts have significant antibacterial and antioxidant potential. *Correspondence E-mail: renatabestagajevic@gmail. com

Three Schiff bases were synthesized by reaction of different benzaldehydes with amino acids. The characterization of these compounds was performed using IR spectroscopy, molecular calculations, thin-layer chromatography, determining the melting point and other physical characteristics. IR spectra for imino groups (C=N), which are characteristic of Schiff bases, show stretching frequency from 1629 to 1654 cm-1. The obtained spectral results were confirmed by molecular calculations using the density functional theory (DFT) and were performed before experimental work. The DFT global chemical reactivity descriptors were calculated and used to predict their relative stability and reactivity of synthesized compounds. The antimicrobial assay of all compounds were screened for Grampositive bacteria species: Staphylococcus aureus ATCC 25923; Methicillin-resistant Staphylococcus aureus: MRSA ATCC 33591; Bacillus subtilis ATCC 6633; and Enterococcus faecalis ATCC 29212, Gram-negative: Salmonella enterica ATCC 31194; Pseudomonas aeruginosa ATCC 9027; Escherichia coli ATCC 25922; Extended Spectrum Beta-Lactamase producing E. coli: ESBL E. coli ATCC 35218, and one yeast Candida albicans ATCC 1023.The highest values of inhibition zones were recorded for compound1, followed by the compound 3, while compound 2 performed inhibitory effect just in case of MRSA. DFT calculations show that antimicrobial activity has a good correlation with chemical reactivity descriptors obtained Schiff bases.

A. Dedić, H. Džudžević-Čančar, T. Stanojković, M. Roje, Damjanović, A. Alispahić, A. Jerković-Mujkić

Sloe ( Prunus spinosa L.) extracts are a good source of natural bioactive compounds, including phytosterols. Phytosterols are known to be applied in the treatment of various prostate diseases. The in vitro antiproliferative activity of sloe ethanolic extracts (flower, leaf, and fruit), collected from three areas in Bosnia and Herzegovina, were investigated against human prostate cancer cell lines PC-3 and DU145 using MTT assay. β-sitosterol, campesterol and stigmasterol were quantified by HPLC-PDA analysis using Symmetry C18 chromatographic column. The results of the analysis proved the presence of phytosterols, mostly β-sitos - terol in all extracts. All extracts possess antiproliferative activity. The highest activity against PC-3 and DU145 was gathered from leaf extracts obtained by different extraction methods (microwave-assisted extraction and ultrasound-assisted extraction). To the best of our knowledge, no other studies have presented results on antiproliferative activity of ethanol sloe extracts. Based on these results, further investigation should be recommended on other cancer cell lines as well.

A. Jerković-Mujkić, I. Mahmutović, R. Bešta-Gajević

UDK 665.52:635.7]:632.38           632.38:633.71 Numerous studies have found that essential oils have significant inhibitory activity against human viruses, but there is scarce information on the effects of essential oils on plant viruses. In this study, effects of cajuput (Melaleuca leucadendron (L.) L.), common myrtus (Myrtus communis L.) and winter savory (Satureja montana L.) essential oils were assayed against Tobacco mosaic virus (TMV) by inoculation in different plant host systems. Antiphytoviral activity of the essential oils was determined by the half-leaf method using test plants: Chenopodium quinoa Willd., Cucumis sativus L. 'Cornishon' and Phaseolus vulgaris L. 'Top Crop'. Results showed that investigated oils posess strong antiviral activities. Among tested oils, the highest and broadest acitivity was shown by winter savory essential oil. Evidence suggests that investigated essential oils are possible antiphytoviral agents.

Leptospirosis is a zoonosis of worldwide distribution. Current methods for the direct detection of leptospires are either slow or of limited reliability so that serology is often the most appropriate diagnostic method. The microscopic agglutination test (MAT) using live bacteria is the reference serological test. For long-term preservation leptospiras are usually maintained by periodic subculture into fresh media because the conventional methods such as freeze-drying have reported inconsistent results for storing leptospiras. Although this technique is very simple the question is to which extent such procedure affects the protein structure of the outer membrane with its key antigens. The aim of the study was to identify potential protein variations of serovars Hardjo and Grippotyphosa after thirty consecutive subcultivations by Sodium Dodecyl Sulfate Polyacrylamide Gel Electrophoresis (SDS). The overall analysis of the leptospiral proteins by SDS-PAGE demonstrated minor differences in the intensities of some bands, while changes in protein expression were not detected. Key words: leptospira, subcultivation, SDS-PAGE, protein variations

R. Bešta-Gajević, A. Jerković-Mujkić, S. Pilić, I. Stanković, A. Vučurović, A. Bulajić, B. Krstic

Lamium maculatum L. (spotted dead-nettle) is a flowering perennial ornamental that is commonly grown as a landscape plant for an effective ground cover. In June 2010, severe mosaic accompanied by reddish brown necrosis and leaf deformation was noticed on 80% of L. maculatum growing in shade under trees and shrubs in Sarajevo (Bosnia and Herzegovina). Leaves from 10 symptomatic L. maculatum plants were sampled and analyzed by double-antibody sandwich (DAS)-ELISA using commercial diagnostic kits (Bioreba AG, Reinach, Switzerland) against Cucumber mosaic virus (CMV), Tomato spotted wilt virus (TSWV), and Impatiens necrotic spot virus (INSV), the most important viral pathogens of ornamental plants (1,2). Commercial positive and negative controls and extracts from healthy L. maculatum leaves were included in each assay. All samples tested negative for TSWV and INSV and positive for CMV. The virus was mechanically transmitted to test plants and young virus-free plants of L. maculatum using 0.01 M phosphate buffer (pH 7). The virus caused chlorotic local lesions on Chenopodium quinoa, while systemic mosaic was observed on Capsicum annuum 'Rotund,' Nicotiana rustica, N. glutinosa, N. tabacum 'White Burley,' and Phaseolus vulgaris 'Top Crop.' The virus was transmitted mechanically to L. maculatum and induced symptoms resembling those observed on the source plants. Inoculated plants were assayed by DAS-ELISA and all five inoculated plants of each species tested positive for CMV. The presence of CMV in L. maculatum as well as mechanically infected N. glutinosa plants was further confirmed by RT-PCR. Total RNA from symptomatic leaves was isolated using RNeasy Plant Mini Kit (Qiagen, Hilden, Germany) and RT-PCR was performed with the One-Step RT-PCR Kit (Qiagen) following the manufacturer's instructions. The primer pair, CMVAu1u/CMVAu2d, that amplifies the entire coat protein (CP) gene and part of 3'- and 5'-UTRs was used for both amplification and sequencing (4). Total RNA obtained from the Serbian CMV isolate from pumpkin (GenBank Accession No. HM065510) and a healthy L. maculatum plant were used as positive and negative controls, respectively. All naturally and mechanically infected plants as well as the positive control yielded an amplicon of the expected size (850 bp). No amplicon was observed in the healthy control. The amplified product derived from isolate 3-Lam was purified (QIAquick PCR Purification Kit, Qiagen), directly sequenced in both directions and deposited in GenBank (JX436358). Sequence analysis of the CP open reading frame (657 nt), conducted with MEGA5 software, revealed that the isolate 3-Lam showed the highest nucleotide identity of 99.4% (99.1% amino acid identity) with CMV isolates from Serbia, Australia, and the USA (GQ340670, U22821, and U20668, respectively). To our knowledge, this is the first report of the natural occurrence of CMV on L. maculatum worldwide and it adds a new host to over 1,241 species (101 plant families) infected by this virus (3). This is also an important discovery for the ornamental industry since L. maculatum is commonly grown together with other ornamental hosts of CMV in nurseries and the urban environment as well as in natural ecosystems. References: (1) Y. K. Chen et al. Arch. Virol. 146:1631, 2001. (2) M. L. Daughtrey et al. Plant Dis. 81:1220, 1997. (3) M. Jacquemond. Adv. Virus Res. 84:439, 2012. (4) I. Stankovic et al. Acta Virol. 55:337, 2011.

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