Assessment of surface water quality based on microbiological parameters and ecogenotoxicological and histopathological analysis of common bream Abramis brama (L., 1758), white bream Blicca bjoerkna (L., 1758) and white-eye bream Ballerus sapa (P., 1814) tissues

Biomonitoring predstavlja neophodnu komponentu tradicionalnih tehnika monitoringa, sa ciljem da se uspostavi veza izmeŤu spoljašnjih koncentracija zagaŤivaţa, koncentracija zagaŤivaţa u tkivima bioindikatora i ranih štetnih efekata po ispitivane organizme. Ribe su korisni bioindikatori i ţesto se primenjuju u ekogenotoksikološkim ispitivanjima. U ovoj studiji, procena stanja kvaliteta površinske vode vršena je na reci Savi na lokalitetu Duboko i na reci Dunav na lokalitetu Višnjica tokom 2014. godine, kako bi se procenio uticaj razliţitih sezona na variranje parametara kvaliteta i odgovor biomarkera deverike, krupatice i crnooke deverike. Procena kvaliteta vode vršena je merenjem fiziţko-hemijskih i mikrobioloških parametara fekalnog i organskog zagaŤenja. Procena genotoksiţnog potencijala vršena je primenom alkalnog komet testa za kvantifikaciju DNK oštešenja u šelijama krvi, jetre i škrga riba, kao biomarkera izlaganja. Histopatološke promene u jetri i škrgama prašene su kao biomarker efekta. Paralelno, prašene su koncentracije metala i metaloida u jetri, škrgama, gonadama i mišišu, primenom metode ICP-OES. Na lokalitetu Višnjica zabeleţen je viši nivo fekalnog i organskog zagaŤenja. Na lokalitetu Duboko zabeleţene su više koncentracije ispitivanih elemenata u sva ţetiri tkiva, u poreŤenju sa lokalitetom Višnjica. Na oba lokaliteta škrge su akumulirale najviše koncentracije metala i metaloida, a mišiš najniţe. Na oba lokaliteta, krv je bila tkivo sa najvišim nivoom DNK oštešenja, dok je jetra tokom vešine meseci imala najniţe DNK oštešenje. Na lokalitetu Višnjica zabeleţene su više vrednosti DNK oštešenja krvi, na lokalitetu Duboko više vrednosti DNK oštešenja škrga, dok je nivo DNK oštešenja u šelijama jetre na oba lokaliteta bio pribliţno jednak. Na lokalitetu Duboko pri najvešim vrednostima histopatološkog indeksa škrga zabeleţeno je blago do umereno oštešenje, a na lokalitetu Višnjica umereno do teško oštešenje škrga. Na oba lokaliteta, jetra je bila organ sa vešim obimom histopatoloških promena u odnosu na škrge. Sveukupno, škrge kao prvi organ u direktnom kontaktu sa zagaŤivaţima iz vode pokazale su vešu akumulaciju ispitivanih elemenata i viši nivo DNK oštešenja, dok je jetra kao glavni organ za procesuiranje zagaŤivaţa iz vode i hrane pokazala viši nivo histopatoloških promena. Odabrane vrste pokazale su se kao pogodni bioindikatori za in situ ispitivanja efekata zagaŤenja.Biomonitoring represents an essential part of traditional monitoring techniques with the aim to establish relationship between external concentrations of pollutants, concentrations of pollutants in the tissues of bioindicators and early adverse effects in examined organisms. Fish are useful bioindicators often used in ecogenotoxicological studies. In this study, assessment of the surface water quality was performed on the Sava River locality Duboko and on the Danube River locality Višnjica during 2014 in order to examine the impact of different seasons on the variation of the quality parameters and biomarker response in common bream, white bream and white-eye bream. Assessment of the water quality was performed based on the physico-chemical parameters and microbiological parameters of fecal and organic pollution. Genotoxic potential was assessed by using the alkaline comet assay in order to quantify DNA damage level, in blood, liver and gill cells, as a biomarker of exposure. Histopathological analyses of liver and gill tissue were monitored as a biomarker of effect. Additionally, concentrations of metals and metalloids in liver, gills, gonads and muscle were performed by the ICP-OES method. The higher level of fecal and organic pollution was present on the locality Višnjica. The higher concentrations of examined elements in all four tissues were present in fish from the locality Duboko, in comparison to the locality Višnjica. At both sites gills accumulated the highest concentrations of metals and metalloids, while muscle accumulated the lowest concentrations. Blood was the tissue with the highest level of DNA damage, while liver had the lowest level of DNA damage during most of the months at both sites. Higher DNA damage level in blood cells was observed at the locality Višnjica, in gill cells at the locality Duboko, while the level of DNA damage in liver was approximately the same at both localities. Based on the highest level of histopathological index of gills at the locality Duboko gills were slightly to moderately damaged, while at the locality Višnjica gills were moderately to heavily damaged. At both localities liver had higher level of histopathological alterations in comparison to gills. Overall, gills as the first organ in direct contact with contaminants form water showed higher level of examined elements and DNA damage, while the liver as the main organ for processing contaminants from water and food showed higher level of histopathological alterations. Examined fish species proved to be useful bioindicators for in situ assessment of the pollution effects

Assessment of surface water quality based on microbiological parameters and ecogenotoxicological and histopathological analysis of common bream Abramis brama (L., 1758), white bream Blicca bjoerkna (L., 1758) and white-eye bream Ballerus sapa (P., 1814) tissues

Biomonitoring predstavlja neophodnu komponentu tradicionalnih tehnika monitoringa, sa ciljem da se uspostavi veza izmeŤu spoljašnjih koncentracija zagaŤivaţa, koncentracija zagaŤivaţa u tkivima bioindikatora i ranih štetnih efekata po ispitivane organizme. Ribe su korisni bioindikatori i ţesto se primenjuju u ekogenotoksikološkim ispitivanjima. U ovoj studiji, procena stanja kvaliteta površinske vode vršena je na reci Savi na lokalitetu Duboko i na reci Dunav na lokalitetu Višnjica tokom 2014. godine, kako bi se procenio uticaj razliţitih sezona na variranje parametara kvaliteta i odgovor biomarkera deverike, krupatice i crnooke deverike. Procena kvaliteta vode vršena je merenjem fiziţko-hemijskih i mikrobioloških parametara fekalnog i organskog zagaŤenja. Procena genotoksiţnog potencijala vršena je primenom alkalnog komet testa za kvantifikaciju DNK oštešenja u šelijama krvi, jetre i škrga riba, kao biomarkera izlaganja. Histopatološke promene u jetri i škrgama prašene su kao biomarker efekta. Paralelno, prašene su koncentracije metala i metaloida u jetri, škrgama, gonadama i mišišu, primenom metode ICP-OES. Na lokalitetu Višnjica zabeleţen je viši nivo fekalnog i organskog zagaŤenja. Na lokalitetu Duboko zabeleţene su više koncentracije ispitivanih elemenata u sva ţetiri tkiva, u poreŤenju sa lokalitetom Višnjica. Na oba lokaliteta škrge su akumulirale najviše koncentracije metala i metaloida, a mišiš najniţe. Na oba lokaliteta, krv je bila tkivo sa najvišim nivoom DNK oštešenja, dok je jetra tokom vešine meseci imala najniţe DNK oštešenje. Na lokalitetu Višnjica zabeleţene su više vrednosti DNK oštešenja krvi, na lokalitetu Duboko više vrednosti DNK oštešenja škrga, dok je nivo DNK oštešenja u šelijama jetre na oba lokaliteta bio pribliţno jednak. Na lokalitetu Duboko pri najvešim vrednostima histopatološkog indeksa škrga zabeleţeno je blago do umereno oštešenje, a na lokalitetu Višnjica umereno do teško oštešenje škrga. Na oba lokaliteta, jetra je bila organ sa vešim obimom histopatoloških promena u odnosu na škrge. Sveukupno, škrge kao prvi organ u direktnom kontaktu sa zagaŤivaţima iz vode pokazale su vešu akumulaciju ispitivanih elemenata i viši nivo DNK oštešenja, dok je jetra kao glavni organ za procesuiranje zagaŤivaţa iz vode i hrane pokazala viši nivo histopatoloških promena. Odabrane vrste pokazale su se kao pogodni bioindikatori za in situ ispitivanja efekata zagaŤenja.Biomonitoring represents an essential part of traditional monitoring techniques with the aim to establish relationship between external concentrations of pollutants, concentrations of pollutants in the tissues of bioindicators and early adverse effects in examined organisms. Fish are useful bioindicators often used in ecogenotoxicological studies. In this study, assessment of the surface water quality was performed on the Sava River locality Duboko and on the Danube River locality Višnjica during 2014 in order to examine the impact of different seasons on the variation of the quality parameters and biomarker response in common bream, white bream and white-eye bream. Assessment of the water quality was performed based on the physico-chemical parameters and microbiological parameters of fecal and organic pollution. Genotoxic potential was assessed by using the alkaline comet assay in order to quantify DNA damage level, in blood, liver and gill cells, as a biomarker of exposure. Histopathological analyses of liver and gill tissue were monitored as a biomarker of effect. Additionally, concentrations of metals and metalloids in liver, gills, gonads and muscle were performed by the ICP-OES method. The higher level of fecal and organic pollution was present on the locality Višnjica. The higher concentrations of examined elements in all four tissues were present in fish from the locality Duboko, in comparison to the locality Višnjica. At both sites gills accumulated the highest concentrations of metals and metalloids, while muscle accumulated the lowest concentrations. Blood was the tissue with the highest level of DNA damage, while liver had the lowest level of DNA damage during most of the months at both sites. Higher DNA damage level in blood cells was observed at the locality Višnjica, in gill cells at the locality Duboko, while the level of DNA damage in liver was approximately the same at both localities. Based on the highest level of histopathological index of gills at the locality Duboko gills were slightly to moderately damaged, while at the locality Višnjica gills were moderately to heavily damaged. At both localities liver had higher level of histopathological alterations in comparison to gills. Overall, gills as the first organ in direct contact with contaminants form water showed higher level of examined elements and DNA damage, while the liver as the main organ for processing contaminants from water and food showed higher level of histopathological alterations. Examined fish species proved to be useful bioindicators for in situ assessment of the pollution effects

Zastupljenost razvojnih anomalija zuba kod ortodontskih pacijenata u Srbiji

Introduction/Objective the aim of this study was to evaluate the prevalence of developmental dental anomalies (DDA) in Serbian orthodontic patients. Methods the sample was composed of 1,001 panoramic radiographs of orthodontic patients, older than seven years, taken as a part of the initial diagnostic procedure at the Clinic of Orthodontics, School of Dental medicine in Belgrade. The DDA that could be diagnosed accurately on panoramic X-rays were documented. Descriptive analysis was used to determine prevalence and sex distribution of DDA. The Pearson ch2 test and Fisher's exact test were used to compare number of affected teeth in males and females (level of significance was 95%). Results the prevalence of DDA in Serbian orthodontic patients was 34.8% (15.5% males and 19.3% females). Impactions were present in 16.5%, hypodontia in 12.9%, hyperdontia in 4.4%, microdontia in 2.9%, macrodontia in 1.8% and transposition in 0.8% of patients. Maxillary canines were the most frequently impacted teeth. Maxillary second molars were more prone to impaction in females (p lt 0.05). Impacted incisors were more prevalent in maxilla, premolars, and second molars in mandible. The most commonly missing teeth were upper left second premolars. Mesiodens was the most frequently found supernumerary tooth. Conclusion We reported a high a rate of DDA in Serbian orthodontic patients, more in females than males. The most frequently observed DDA were impaction, tooth agenesis, hyperdontia, microdontia, macrodontia, and transposition. All investigated DDA were more frequently present in females, except hyperdontia. Current findings could offer a foundation for epidemiological studies on DDA prevalence.Uvod/Cilj Cilj ovog istraživanja je bio da se ispita zastupljenost razvojnih anomalija zuba kod ortodontskih pacijenata u Srbiji. Metode Uzorak je činio 1001 ortopantomografski snimak ortodontskih pacijenata starijih od sedam godina sa Klinike za ortopediju vilica Stomatološkog fakulteta u Beogradu. Beleženo je prisustvo razvojnih anomalija za čiju dijagnostiku je potreban samo ortopantomografski snimak. Za ispitivanje zastupljenosti razvojnih anomalija zuba korišćena je deskriptivna statistička analiza. ch2 test je korišćen radi poređenja broja zuba sa anomalijom između polova (stepen značajnosti 95%). Rezultati Razvojne anomalije zuba su bile zastupljene kod 34,8% ortodontskih pacijenata (15,5% muškaraca i 19,3% žena). Impakcije zuba su bile prisutne kod 16,5% pacijenata, hipodoncija kod 12,9%, prekobrojni zubi kod 4,4%, mikrodoncija kod 2,9%, makrodoncija kod 1,8% i transpozicija kod 0,8% pacijenata. Očnjaci u gornjoj vilici su bili najčešće impaktirani zubi. Gornji drugi kutnjaci su bili skloniji impakciji kod žena (p lt 0,05). Dokumentovano je više impaktiranih sekutića u gornjoj vilici, a pretkutnjaka i drugih kutnjaka u donjoj vilici. Najčešće su nedostajali gornji levi pretkutnjaci. Od svih prekobrojnih zuba najčešće je bio uočavan meziodens. Zaključak Prikazali smo postojanje visoke učestalosti razvojnih anomalija zuba kod ortodontskih pacijenata u Srbiji sa većom izraženošću kod osoba ženskog pola. Najčešće anomalije bile su impakcija, hipodoncija, hiperdoncija, mikrodoncija, makrodoncija i transpozicija. Sve anomalije su bile učestalije kod žena, osim u slučaju prekobrojnih zuba. Rezultati sadašnje studije mogu biti polazna tačka za epidemiološke studije o učestalosti razvojnih anomalija zuba

Comet assay – a sensitive method for detection DNA damage and primary monitoring of ecosystem pollution pressure

Untreated municipal wastewaters are one of the major negative contributors on freshwater quality, and consequently on ecosystem balance. Some compounds such as pharmaceuticals, cosmetic products, etc. could be genotoxic and lead to somatic and/or germinative mutations. This mutation should be reflected on organisms’ health and reproductive potential. Comet assay is a widely used test in ecogenotoxicology for detection of primary DNA damage. In that way, this sensitive test could provide information about early warning signs of potential stressors effects before those have an impact on the population or ecosystem level. The scope of our study was to assess the level of DNA damage of Alburnus alburnus specimens’ gill cells and erythrocytes. Three sites on the Sava river were chosen: reference site Zabran suited upstream the municipal discharging; second at the confluence of the Kolubara river and the Sava river; and the third at the confluence of the Barička river and the Sava river. The Kolubara river and the Barička river are chosen as important recipients of untreated wastewaters. From each site, 5 bleak specimens were collected and blood and gills were taken. Tail Intensity (TI%) was chosen as a parameter for evaluating the level of DNA damage. Besides that, cell viability and extremely damaged cells - hedgehogs (HH) were determined. The significant difference in TI% values was recorded in erythrocytes and gills cells in comparison between the Zabran (TI%=14.01±0.61 in erythrocytes, TI%=15.25±1.03 in gills cells) and the second site (TI%=18.53±0.65 in erythrocytes, TI%=22.78±1.89 in gills cells). Correlation between cell viability and HH frequency was not observed. According to the results, we could conclude that the second site is the most affected by pollutants. Also, usage of comet assay on freshwater organisms could be appropriate for preliminary screening of ecosystem state

Simultaneous determination of mesotrione and nicosulfuron in OD formulations

In this study an isocratic high-performance liquid chromatographic method with diode array detection was developed for simultaneous determination of mesotrione and nicosulfuron active ingredients content in pesticide products formulated as oil dispersion (OD). For the analysis, LC system an Agilent Technologies 1100 was used. Good separation was achieved on a Zorbax SB-C18 column using a mobile phase consisting of 0.1% CH3COOH/acetonitrile (75:25), at a flow rate of 0.9 ml/minute and UV detection at 245 nm. Column temperature was 40 °C, injected volume was 1 µl. Retention times for nicosulfuron and mesotrione were 3.009 min and 4.363 min, respectively. Validation of the method was performed according to IAEA guidelines. The obtained results showed that the linear coefficients were 0.9999 for both analyzed herbicides in the mixture. The repeatability of the method expressed as relative standard deviations (%RSDr) was 0.33% for mesotrione and 0.18% for nicosulfuron. The accuracy of the proposed method was determined from recovery experiments. The obtained results for mesotrione and nicosulfuron were 100.5-102.9% and 99.3-103.9%, respectively, proved to be acceptable. Precision of the method expressed as %RSD was 0.38% for mesotrione and 0.99% for nicosulfuron and are lower than the values calculated by the Horwitz equation

Cryopreservation of fish blood – useful tool for assessing genotoxic potential of aquatic ecosystems

One of the major limitations in performing ecogenotoxicological studies is the distance between research field and the laboratory. As some of the methods used in ecogenotoxicology require fresh biological material with intact cell viability, transfer of samples to the laboratory within a few hours after sampling is usually required. To overcome this issue, we have introduced cryopreservation in our research as a possible solution. Cryopreservation is a method which includes preservation of intact, living cells at low temperature for a long time. In natural conditions freezing, forming of ice crystals and dehydration could destroy cell structures. To avoid this consequence, specific compounds were introduced, cryoprotective agents, in the method of cryopreservation. The main characteristic of these compounds is their ability to reduce ice crystal formation in cells at any temperature. We have applied cryopreservation in the evaluation of genotoxic potential along different river streams (the Adige River, the Sava River and the Velika Morava River basin). For this purpose, we focused on the level of DNA damage of cryopreserved fish blood cells (Salmo cenerinus, Salmo marmoratus, Alburnus alburnus) by using the comet assay. To test whether cryopreservation has the impact on cell viability, or that it induces additional DNA damage, we employed preliminary experiments in 4 Abramis brama and 8 A. alburnus specimens. Namely, from every specimen two blood samples were taken: one for analyzing cells viability and the level of DNA damage of fresh blood, and another for observing cell viability and DNA damage of cryopreserved samples. The viability of cell blood was determined by using acridine orange/ethidium bromide differential staining. For analyzing the level of DNA damage alkaline comet assay was used. Obtained results indicated that cryopreserved blood cells had approximately the same viability and the level of DNA damage as nonpreserved blood samples. According to our results, cryopreservation is a very useful method in genotoxicology and could have many benefits: blood samples should not be analyzed immediately after sampling; samples could be transported in liquid nitrogen without concern about additional DNA damage

Particle size effects on the structure and emission of Eu3+:LaPO4 and EuPO4 phosphors

The authors acknowledge the financial support of the Ministry of Education, Science and Technological Development of the Republic of Serbia (Projects nos. 45020 and 172056). T.G acknowledges to the ERDF PostDoc project No. 1.1.1.2/VIAA/1/16/215 (1.1.1.2/16/I/001).This paper provides the detailed study of (nano)particle's size effect on structural and luminescent properties of LaPO4:Eu3+ synthesized by four different methods: high temperature solid-state, co-precipitation, reverse micelle and colloidal. These methods delivered monoclinic monazite-phase submicron particles (> 100 nm), 4 × 20 nm nanorods and 5 nm spheres (depending on the annealing temperature), 2 × 15 nm nanorods, and ultra-small spheres (2 nm), respectively. The analysis of emission intensity dependence on Eu3+ concentration showed that quenching concentration increases with a decrease of the particle size. The critical distance for energy transfer between Eu3+ ions is found to be 18.2 Å, and the dipole-dipole interaction is the dominant mechanism responsible for the concentration quenching of emission. With the increase in Eu3+ concentration, the unit-cell parameter slightly increases to accommodate larger Eu3+ ions at sites of smaller La3+ ions. Photoluminescent emission spectra presented four characteristic bands in the red spectral region: at 592 nm (5D0→7F1), at 612 nm (5D0→7F2), at 652 nm (5D0→7F3) and at 684 nm (5D0→7F4), while in small colloidal nanoparticles additional emission bands from host defects appear at shorter wavelengths. Intensities of f-f electronic transitions change with particles size due to small changes in symmetry around europium sites, while emission bandwidths increase with the reduction of particle size due to increased structural disorder. Judd-Ofelt analysis showed that internal quantum yield of Eu3+ emission is strongly influenced by particle's morphology.Ministry of Education, Science and Technological Development of the Republic of Serbia (Projects nos. 45020 and 172056); ERDF PostDoc project No. 1.1.1.2/VIAA/1/16/215 (1.1.1.2/16/I/001); Institute of Solid State Physics, University of Latvia as the Center of Excellence has received funding from the European Union’s Horizon 2020 Framework Programme H2020-WIDESPREAD-01-2016-2017-TeamingPhase2 under grant agreement No. 739508, project CAMART

Синтеза и биолошка активност алкилтио и арилтио деривата терц-бутилхинона

Biological activity of 2-tert-butyl-1,4-benzoquinone (TBQ) and its derivatives, 2-tert-butyl-5-(2-propylthio)-1,4-benzoquinone, 2-tert-butyl-5- -(propylthio)-1,4-benzoquinone, 2-tert-butyl-5,6-(ethylenedithio)-1,4-benzoquinone, 2-tert-butyl-5-(phenylthio)-1,4-benzoquinone and 2-tert-butyl-6-(phenylthio)- 1,4-benzoquinone, were tested for their antioxidant, antibacterial, toxic, cytotoxic and genotoxic potential. Using the DPPH test, all derivatives showed good antioxidant activity, better than ascorbic acid, and the 2-tert- -butyl-5-(propylthio)-1,4-benzoquinone derivative showed the strongest effect. Better antibacterial potential was observed against Gram-positive bacteria in the broth microdilution method in which the 2-tert-butyl-5-(phenylthio)-1,4- -benzoquinone derivative showed the strongest activity (MIC = 15.6 μM). The results of toxicity tests, using the Brine shrimp test, indicated that the derivatives lose their toxic potential compared to TBQ, except for 2-tert-butyl-6- -(phenylthio)-1,4-benzoquinone, which showed a 3 times stronger effect. Cytotoxicity was assessed by the MTT assay in 24 and 72 h treatments in MRC-5, HS 294T and A549 cell lines in threefold decreasing gradient (11, 33 and 100 μM). Modifications potentiate the cytotoxic effect, and the strongest effect was observed with the 2-tert-butyl-5,6-(ethylendithio)-1,4-benzoquinone derivative. In addition, the genotoxic potential was examined in the MRC-5 cell line using the comet assay. All tested derivatives of TBQ showed a genotoxic effect at all applied subtoxic concentrations. In general, the chemical modifications of TBQ enhanced its biological activity.Испитана је биолошка активност 2-терц-бутил-1,4-бензохинона (TBQ) и његових деривата: 2-терц-бутил-5-(изопропилтио)-1,4-бензохинона, 2-терц-бутил-5-(пропилтио)-1,4-бензохинона, 2-терц-бутил-5,6-(етиленедитио)-1,4-бензохинона, 2-терц-бутил- -5-(фенилтио)-1,4-бензохинона и 2-терц-бутил-6-(фенилтио)-1,4-бензохинона укључујући њихов антиоксидативни, антибактеријски, токсични, цитотоксични и генотоксични потенцијал. Применом DPPH теста, сви деривати су показали добру антиоксидативну активност, бољу од аскорбинске киселине, а најјаче дејство показао је дериват 2-терц-бутил-5-(пропилтио)-1,4-бензохинон. Бољи антимикробни потенцијал је примећен против Грам-позитивних бактерија методом микродилуције у бујону, где је дериват 2-терц-бутил-5-(фенилтио)-1,4-бензохинон показао најјачу активност (MIC = 15,6 μМ). Резултати испитивања токсичности, применом теста на Artemia salina, показују да деривати губе токсични потенцијал у односу на TBQ, осим 2-терц-бутил-6-(фенилтио)- -1,4-бензохинона, који је показао 3 пута јачи ефекат. Цитотоксичност је испитана МТТ тестом у третманима од 24 и 72 h на ћелијским линијама MRC-5, HS 294T и A549 у троструко опадајућем градијенту (11, 33 и 100 μМ). Модификације појачавају цитотоксични ефекат, а најјачи ефекат је примећен код деривата 2-терц-бутил-5,6-(етилендитио)-1,4-бензохинона. Поред тога, генотоксични потенцијал је испитан на ћелијској линији MRC-5 комет тестом. Сви испитивани деривати су показали генотоксични ефекат при свим примењеним субтоксичним концентрацијама. Генерално, хемијске модификације побољшавају биолошку активност 2-терц-бутил-1,4-бензохинона