Does monensin in chicken manure from poultry farms pose a threat to soil invertebrates?

Monensin is a carboxylic polyether ionophore used in the poultry industry as a coccidiostat. It enters the environment via manure from broiler farms. In spite of its potential presence in the environment, information concerning monensin residues in manure and soil and its toxicity to soil organisms are insufficient. In the present study, two beneficial soil invertebrate species, earthworms (Eisenia andrei) and woodlice (Porcellio scaber), were used to assess the toxicity of monensin. Animals were exposed to a range of monensin concentrations via soil or food. Earthworm reproduction was found to be the most susceptible endpoint (NOEC=3.5 mg kg(-1) dry soil; EC(50)=12.7 mg kg(-1) dry soil), while no adverse effects were recorded in isopods (NOEC?849mgkg(-1) dry soil, NOEC?357mgkg(-1) dry food). The obtained toxicity data were compared with potential concentrations of monensin in soil. In view of this, manure from broiler chickens treated with monensin at a poultry farm was sampled. According to monensin and nitrogen concentrations in the chicken manure and the degradation time of monensin, the predicted environmental concentration (PEC) was calculated. PEC of monensin is around 0.013 mg kg(-1) soil if manure is used after 3 months of composting and 0.05 mg kg(-1) soil if used without storage. Data for earthworm reproduction was used to estimate the predicted no-effect concentration (PNEC). If fresh chicken manure is applied to terrestrial ecosystems, the risk quotient (PEC/PNEC ratio) is above 1, which indicates that monensin might pose an environmental risk under certain conditions. To prevent this, it is strongly recommended to compost chicken manure for several months before using it as fertiliser.     

The Properties of Poly(<Emphasis Type="SmallCaps">l</Emphasis>-Lactide) Prepared by Different Synthesis Procedure

Different synthesis methods were applied to determine optimal conditions for polymerization of (3S)-cis-3,6-dimethyl-1,4-dioxane-2,5-dione (l-lactide), in order to obtain poly(l-lactide) (PLLA). Bulk polymerizations (in vacuum sealed vessel, high pressure reactor and in microwave field) were performed with tin(II) 2-ethylhexanoate as the initiator. Synthesis in the vacuum sealed vessel was carried out at the temperature of 150 °C. To reduce the reaction time second polymerization process was carried out in the high pressure reactor at 100 °C and at the pressure of 138 kPa. The third type of rapid synthesis was done in the microwave reactor at 100 °C, using frequency of 2.45 GHz and power of 150 W at the temperature of 100 °C. The temperature in this method was controlled via infrared system for in-bulk measuring. The solution polymerization (with trifluoromethanesulfonic acid as initiator) was possible even at the temperature of 40 °C, yielding PLLA with narrow molecular weight distribution in a very short period of time (less than 6 h). The obtained polymers had the number-average molecular weights ranging from 43,000 to 178,000 g mol⁻¹ (polydispersity index ranging from 1 to 3) according to the gel permeation chromatography measurements. The polymer structure was characterized by Fourier transform infrared and NMR spectroscopy. Thermal properties of the obtained polymers were investigated using thermogravimetry and differential scanning calorimetry.     

Determination of microbiological contamination,antibacterial and antioxidant activities of natural plant hazelnut (Corylus avellana L.) pollen

The aim of our study is to determine microbial contamination, antibacterial and antioxidant activities of 14 pollen samples of Corylus avellana collected from different locations in Slovakia. Microbiological analysis was carried out in two steps: microbiological assays and studies of antibacterial activity of pollen extracts. The antimicrobial properties of pollen extracts were carried out with the disc-diffusion method. Methanol (70%), ethanol (70%) and distilled water were used for pollen extracts. Five strains of bacteria such as gram-negative (Salmonella enterica subsp. enterica CCM 3807, Escherichia coli CCM 2024, and Yersinia enterocolitica CCM 5671) and gram-positive (Staphylococcus aureus CCM 2461 and Bacillus thuringiensis CCM 19T) were tested. Antioxidant activity of pollen extracts was determined by the DPPH method. Bacterial analysis includes the determination of the total bacterial count ranged from 4.08 to 4.61 log CFU g^?1, mesophilic aerobic bacteria ranged from 3.40 to 4.89 log CFU g^?1, mesophilic anaerobic bacteria ranged from 3.20 to 4.52 log CFU g^?1, coliform bacteria ranged from 3.30 to 4.55 log CFU g^?1, yeasts and filamentous fungi ranged from 3.00 to 3.56 log CFU g^?1. Microscopic filamentous fungi Aspergillus spp., Alternaria spp., Penicillium spp., Cladosporium spp., Rhizopus spp., and Paecylomyces spp. were isolated from hazelnut pollen. Yersinia enterocolitica was the most sensitive strain among ethanolic and methanolic pollen hazelnut extracts. Staphylococcus aureus was the most sensitive strain against aqueous hazelnut pollen extracts. We determined the following sensitivity against ethanol pollen extracts respectively: Yersinia enterocolitica?>?Salmonella enterica?>?Staphylococcus aureus?>?Bacillus thuringiensis?>?Escherichia coli. Methanol pollen extracts had shown following sensitivity: Yersinia enterocolitica?>?Salmonella enterica?>?Escherichia coli?>?Staphylococcus aureus?>?Bacillus thuringiensis. Aqueous extracts had shown the following sensitivity: Staphylococcus aureus?>?Salmonella enterica?>?Escherichia coli?>?Bacillus thuringiensis?>?Yersinia enterocolitica. Hazelnut pollen extracts have over 82% antioxidant capacity in samples from non-urban zones. An elevated level of antioxidant potential in the pollen is determined by its biological properties conditioned by biologically active substances. DPPH method allowed characterizing pollen as a source of antioxidants.     

Stakeholders' expectations from the agri-environmental programme in Slovenia and Croatia

The agri-environmental programme (AEP) is the European Union policy instrument used for the delivery of environmental services expected by the society, but societal expectations for these environmental services are insufficiently assessed. In order to realistically meet the expectations of AEP, this research utilises analytical hierarchy process based web survey to assess the importance that various societal stakeholders in the European countries of Slovenia and Croatia give to specific environmental services and to also identify the agricultural practices that have the greatest potential to realise the expectations that society values the most. All stakeholders (overall group) from Slovenia and Croatia ranked water quality and availability as the first most important environmental service, and reduction of pesticides as the most important agricultural practice to deliver societal expectations. The results indicate that there is similar demand for environmental services in these two countries that differ in their agricultural settings.     

Evaluation and validation of two different chromatographic methods (HPLC and LC-MS/MS) for the determination and confirmation of ochratoxin A in pig tissues

This study was undertaken to compare two different analytical methods for the determination and confirmation of ochratoxin A (OTA) in blood serum, kidney and liver of pigs. Sample clean-up was based on liquid-liquid phase extraction. The detection of OTA was accomplished with high-performance liquid chromatography (HPLC) combined either with fluorescence detection (FL) or electro spray ionization (ESI+) tandem mass spectrometry (MS–MS). Comparative method evaluation was based on the investigation of 90 samples of blood serum, kidney and liver per animal originating from different regions of Serbia. The analytical results are discussed in view of the respective method validation data and the corresponding experimental protocols. In general, analytical data obtained with (LC–MS–MS) liquid chromatography electro spray tandem mass spectro metry detection offered comparable good results at the sub-ppb concentration level. The results indicate that the liquid chromatography electro spray tandem mass spectrometric (LC-MS/MS) method was more specific and sensitive for the analysis and confirmation of ochratoxin A in pig tissues then high pressure liquid chromatography (HPLC) method after methylation of OTA.     

Micronucleus frequency in sheep lymphocytes after in vitro exposure to fungicide tolylfluanid

The fungicide tolylfluanid (N - dichlorofluoromethylthio-N′, N - dimethyl - N - p - tolylsulfamide), was investigated by cytokinesis-block micronucleus assay. Tolylfluanid at the lowest concentration (1 × 10^{? 6}mol L^{? 1})did not influence significantly the frequency of micronuclei in sheep lymphocyte cultures in comparison with control (32.33 ± 3.51/1000 binucleated cells versus 30.33 ± 2.82/1000 binucleated cells in dimethylsulfoxide (DMSO) control, P = 0.44). Higher tolylfluanid concentrations (1 × 10^{? 4} and, 1 × 10^{? 5} mol L^{? 1}) resulted in a significant dose-dependent increase in the number of micronuclei in comparison with control (74.00 ± 13.00/1000 binucleated cells and 52.67 ± 10.12/1000 binucleated cells versus 30.33 ± 2. 82/1000 binucleated cells in DMSO control, P = 0.005 and 0.02, respectively, ANOVA followed by Tukey test P < 0.05). Many of the treated cells also possessed multiple micronuclei. Tolylfluanid did not affect the nuclear division index at all treatment concentrations. Our in vitro results thus demonstrate that tolylfluanid had a significant genotoxic effect at only the highest concentration tested.     

Toxicity of metalaxyl, azoxystrobin, dimethomorph, cymoxanil, zoxamide and mancozeb to Phytophthora infestans isolates from Serbia

A study of the in vitro sensitivity of 12 isolates of Phytophthora infestans to metalaxyl, azoxystrobin, dimethomorph, cymoxanil, zoxamide and mancozeb, was conducted. The isolates derived from infected potato leaves collected at eight different localities in Serbia during 2005-2007. The widest range of EC(50) values for mycelial growth of the isolates was recorded for metalaxyl. They varied from 0.3 to 3.9 μg mL(-1) and were higher than those expected in a susceptible population of P. infestans. The EC(50) values of the isolates were 0.16-0.30 μg mL(-1) for dimethomorph, 0.27-0.57 μg mL(-1) for cymoxanil, 0.0026-0.0049 μg mL(-1) for zoxamide and 2.9-5.0 μg mL(-1) for mancozeb. The results indicated that according to effective concentration (EC(50)) the 12 isolates of P. infestans were sensitive to azoxystrobin (0.019-0.074 μg mL(-1)), and intermediate resistant to metalaxyl, dimethomorph and cymoxanil. According to resistance factor, all P. infestans isolates were sensitive to dimethomorph, cymoxanil, mancozeb and zoxamide, 58.3% of isolates were sensitive to azoxystrobin and 50% to metalaxyl. Gout's scale indicated that 41.7% isolates were moderately sensitive to azoxystrobin and 50% to metalaxyl.     

Antibacterial activity against Clostridium genus and antiradical activity of the essential oils from different origin

In the present study, the antimicrobial and antiradical activities of 15 essential oils were investigated. The antimicrobial activities were determined by using agar disc diffusion and broth microdilution methods against Clostridium genus and antioxidant properties of essential oils by testing their scavenging effect on DPPH radicals activities. We determined the antibacterial activity of Clostridium butyricum, Clostridium hystoliticum, Clostridium intestinale, Clostridium perfringens and Clostridium ramosum. We obtained the original commercial essential oils samples of Lavandula angustifolia, Carum carvi, Pinus montana, Mentha piperita, Foeniculum vulgare Mill., Pinus sylvestris, Satureia montana, Origanum vulgare L. (2 samples), Pimpinella anisum, Rosmarinus officinalis L., Salvia officinalis L., Abies alba Mill., Chamomilla recutita L. Rausch and Thymus vulgaris L. produced in Slovakia (Calendula a.s., Nova Lubovna, Slovakia). The results of the disk diffusion method showed very high essential oils activity against all tested strains of microorganisms. The best antimicrobial activity against C. butyricum was found at Pimpinella anisum, against C. hystoliticum was found at Pinus sylvestris, against C. intestinale was found at Satureia hortensis L., against C. perfringens was found at Origanum vulgare L. and against C. ramosum was found at Pinus sylvestris. The results of broth microdilution assay showed that none of the essential oils was active against C. hystoliticum. The best antimicrobial activity against C. butyricum was found at Abies alba Mill., against C. intestinale was found at Abies alba Mill., against C. perfringens was found at Satureia montana and against C. ramosum was found at Abius alba and Carum carvi. Antioxidant DPPH radical scavenging activity was determined at several solutions of oil samples (50 μL.mL^?1–0.39 μL.mL^?1) and the best scavenging effect for the highest concentration (50 μL.mL^?1) was observed. The antioxidant properties were different in particular plant species. The highest% of inhibition after 30 min. of reaction was observed at Origanum vulgare (93%), Satureia montana (90.66%) and Lavandula augustifolia (90.22%).     

Contamination of wild-grown edible mushrooms by heavy metals in a former mercury-mining area

The aim of this study was to evaluate the contamination of six edible wild species of mushrooms (Boletus pulverulentus, Cantharellus cibarius, Lactarius quietus, Macrolepiota procera, Russula xerampelina and Suillus grevillei) by heavy metals (Hg, Cd, Pb, Zn, Cu, Ni, Cr, Co, Mn and Fe). Mushroom samples were collected from sites contaminated by emissions from mining and processing of polymetallic ores in operation during the period 1969–1993 in Rudňany, southeast Slovakia. The four study sites spanned up to a 5-km distance from the emission source. The collected mushroom samples were analyzed using Flame Atomic Absorption Spectrophotometry and/or Flame Atomic Absorption Spectrophotometry with graphite furnace. Mercury, Cd and, in some samples, also Pb present the highest risks in terms of contamination of the food chain following subsequent consumption. The content of two metals in the dry matter (dm) of the mushrooms exceeded the limits set by the European Union (EU; Cd: 0.5 mg/kg dm, Pb: 1.0 mg/kg dm). The highest mean contents of the eight metals recorded for S. grevillei were 52.2, 2.15, 107, 104, 2.27, 2.49, 81.6 and 434 mg/kg dm for Hg, Pb, Zn, Cu, Ni, Cr, Mn and Fe, respectively. The highest content of Cd was recorded in M. procera (3.05 mg/kg dm) and that of Co in L. quietus (0.90 mg/kg dm). The calculated weekly intake for Hg, Pb and Cd shows that regular consumption of mushrooms from the studied area poses risks to human health.     

Toxicological characterization of the landfill leachate prior/after chemical and electrochemical treatment: A study on human and plant cells

In this research, toxicological safety of two newly developed methods for the treatment of landfill leachate from the Piškornica (Croatia) sanitary landfill was investigated. Chemical treatment procedure combined chemical precipitation with CaO followed by coagulation with ferric chloride and final adsorption by clinoptilolite. Electrochemical treatment approach included pretreatment with ozone followed by electrooxidation/electrocoagulation and final polishing by microwave irradiation. Cell viability of untreated/treated landfill leachate was examined using fluorescence microscopy. Cytotoxic effect of the original leachate was obtained for both exposure periods (4 and 24 h) while treated samples showed no cytotoxic effect even after prolonged exposure time. The potential DNA damage of the untreated/treated landfill leachate was evaluated by the comet assay and cytokinesis-block micronucleus (CBMN) assay using either human or plant cells. The original leachate exhibited significantly higher comet assay parameters compared to negative control after 24 h exposure. On the contrary, there was no significant difference between negative control and chemically/electrochemically treated leachate for any of the parameters tested. There was also no significant increase in either CBMN assay parameter compared to the negative control following the exposure of the lymphocytes to the chemically or electrochemically treated landfill leachate for both exposure periods while the original sample showed significantly higher number of micronuclei, nucleoplasmic bridges and nuclear buds for both exposure times. Results suggest that both methods are suitable for the treatment of such complex waste effluent due to high removal efficiency of all measured parameters and toxicological safety of the treated effluent.