BOOK of ABSTRACTS Young Researcher Symposium, Facing Environmental Challenges, South Africa, August 2014 page 3 Obed Phahlane and Hannes Rautenbach - Communicating climate change with the agricultural sector 4 Lindeque du Toit and Willem Van Niekerk - Potential of mitigating methane and nitrous oxide emissions from Southern African ruminant production systems 5 Riana Reinecke and Norman Casey - Green house gas emissions on South African dairy production systems 6 Bronwyn Reijnders and Norman Casey - The Bromide challenge in groundwater 7 Connie Frank Matthiesen and Anne-Helene Tauson - Dietary supplements to a low protein diet may affect the incidence of hepatic lipidosis in mink 8 Christine Brøkner, Julia L. Milling, Hanne H. Hansen - In vitro fermentation of various feed incubated with Warmblood or Icelandic horse faecal inoculum 9 Ewa Sawosz, Slawomir Jaworski, Marta Kutwin, Anna Hotowy, Mateusz Wierzbicki, Marta Grodzik, Natalia Kurantowicz, Barbara Strojny, André Chwalibog - Graphene toxicity in experiments in a chicken embryo model 10 Slawomir Jaworski, Ewa Sawosz, Marta Kutwin, Mateusz Wierzbicki, Marta Grodzik, Mateusz Hinzmann André Chwalibog - In vitro and in vivo evaluation of the effects of graphene nanoplatelets on glioblastoma multiforme 11 Barbara Strojny, Anna Grabowska, Marta Grodzik, Mateusz Hinzmann, Adam Lewera, Aleksandra Lisek, Barbara Mielcarek, Elżbieta Migurska, Sandra Sierankowska, Klaudia Strucińska, Anna Tyszko - The influence of newly produced platinum nanoparticles on model organism development 12 Marta Kutwin, Ewa Sawosz, Sławomir Jaworski, Mateusz Wierzbicki, Mateusz Hinzmann, André Chwalibog - Comparison of in vitro cytotoxicity of platinum nanoparticles and cisplatin against human glioma cells line 1 13 Natalia Kurantowicz, Sławomir Jaworski, Ewa Sawosz, Marta Kutwin, Barbara Strojny, Mateusz Wierzbicki, Jacek Szeliga, André Chwalibog - How graphene family materials affect Listeria monocytogenes and Salmonella Enteritidis strains 14 R. K. Tewo, J. P. Maree, S.J Ruto, B. S Xalakashe, A.V Kolesnikov, G.B Mohajane Validation of gypsum reduction process using the mintek pyrosim model 15 S. Nomfazi, J. Maree, L. Cele - Processing of calcium sulphide to calcium carbonate during batch studies 16 Maria A. Tejada, Karen Molbaek, Martias E. Martin , Dan A. Klaerke - Oocytes of the South-African clawed frog, Xenopus laevis, as a model for characterizing K+ channels 17 Rion Lerm, Gerard Malan, Ray Jansen - The conservation value of the avifauna of the Kathu Forest, South Africa 18 Carina Visser - Angora goats: genetics and product quality 19 Wanda Olech - Overcoming the genetic bottleneck – the European bison case 20 Eleen Strydom - Wakkerstroom Secetarybird Project: Diet and productivity 21 Ulrich Mwimbi, Francois Siebrits, Rebecca Mbatha - Evaluation of chimpanzee (pan troglodytes) diet nutrient composition based on the South African Medical Research Council Food Composition Tables 22 Rasmus Bovbjerg Jensen, Peter Junghans , Anne-Helene Tauson - Energy expenditure in horses measured with the 13C-bicarbonate tracer technique 23 Caroline Larsson, Øystein Ahlstrøm, Peter Junghans, Anne-Helene Tauson - The oral 13 C bicarbonate technique for measurement of energy expenditure in dogs under field conditions 24 Krzysztof Klimaszewski - Species management plan for the Great Crested Newt Triturus cristatus (Laurenti 1768) in Poland 2 Communicating climate change with the agricultural sector Obed Phahlane1 and Hannes Rautenbach2 1 Institute for Soil, Climate and Water, Agricultural Research Council, Private Bag X79, Pretoria, 0001, Email: mogohlwana@gmail.com 2 Department of Geography, Geoinformatics and Meteorology, University of Pretoria, Private Bag X20, Hatfield, 0028 Efficient communication of uncertainty of the seasonal forecast is important in improving the livelihood and climate vulnerability of poor rural communities in South Africa. It allows them to make better decisions that are attuned to the reliability of the forecast. Well-timed and correct dissemination method also helps to manage the expectations of users for accurate forecasts received. In an attempt to disseminate weather forecast information to communities in the Limpopo and North West Provinces, two radio stations were selected, namely Capricorn FM which broadcasts in English and Sepedi in Limpopo, and North West FM which broadcasts in English and Setswana in North West. A 30-minute talk show was broadcast during the second week of each month for a period of 6 months on each station. The efficiency of using radio as one of the methods to disseminate weather forecast information was evaluated. A web-based questionnaire was developed to evaluate the impact of the information discussed on the radio. The questionnaire was hosted on the ARC’s website and the seasonal weather forecast discussed was also uploaded after the show. A total of 96 correctly completed questionnaires were captured on the system. The number of people accessing the website and the questionnaire increased during the week of the talk show, because of the continuous of awareness on the radio. The percentage of questionnaire respondents endorsing television as a preferred method of weather forecast communication were 43%, secondly radio at 35% and thirdly cell phone at 22%. Radio as a method of weather forecast information dissemination indicted a high percentage of weather and agricultural content relevance as compared to television. The understanding of meteorological phenomena was clearly a limiting factor to many users, as a high number of respondents did not understand what “normal rainfall” is. In order to communicate weather forecasts effectively so that benefit is derived from such information, basic training in the form of radio programmes is recommended. 3 Potential of mitigating methane and nitrous oxide emissions from Southern African ruminant production systems Lindeque du Toit1,2 and Willem Van Niekerk2 1 Department of Animal Science, Tshwane University of Technology, Private Bag X680, Pretoria, 0001, South Africa, Email: dutoitcjl@tut.ac.za 2 Department of Animal and Wildlife Sciences, University of Pretoria, 0002, South Africa There are increasing concerns regarding the impact of livestock production on the global environment. The objective of this paper is to review possible mitigation potions or strategies, which could impact on the emissions reductions from ruminant production systems in Southern Africa. Livestock produce greenhouse gas (GHG) emissions in the form of methane (CH4) from enteric fermentation, and nitrous oxide (N2O) and methane from manure management and manure deposited on pastures and rangeland by grazing animals. Methane emissions from South African livestock are estimated at 1328 Giga gram (Gg) with extensive beef cattle (60.5%) the largest contributor followed by sheep (12.6%), privately owned game (9.9%), dairy cattle (9.8%) and the smaller livestock industries. Mitigation of greenhouse gas emissions can be targeted at three areas of livestock production, enteric CH4 mitigation practices, CH4 and N2O mitigation practices from manure management, and animal management-based mitigation practices. Improving forage quality and the efficiency of nutrient use is an effective strategy of decreasing GHG emissions per unit of animal product. Dietary supplements such as lipids, concentrates, nitrates, ionophores, and plant derived secondary compounds have shown potential to reduce enteric CH4 emissions from ruminants. The long-term effects of many of these supplements have not been established and may not be adaptable or economically viable in developing countries. There are several management practices that have a significant potential for reducing emissions from manure although these are more focused on intensive management systems such as dairy production systems and feedlots. Increasing animal productivity is a very effective strategy for reducing GHG emissions per unit of livestock product. This can be achieved through improving genetic potential of animals, strategic supplementation, improved reproductive efficiency as well as improved animal health. Interactions among components in livestock production systems are complex and must be considered when recommending mitigation strategies. One practice may successfully mitigate enteric emissions, but increase emissions from manure management systems through increased fermentable substrate content of manure. Emission mitigation strategies must be practical, economically feasible, and take into account the natural resources and the socio-economic livelihoods of farmers in order to be adopted into agricultural best practice of a region. 4 Green house gas emissions on South African dairy production systems Riana Reinecke and Norman Casey Department of Animal and Wildlife Sciences, Faculty of Natural and Agricultural Sciences, University of Pretoria, Pretoria 0028, South Africa, Email: rianareinecke@telkomsa.net The study estimated individual and total GHG emissions in South African dairy production systems by applying different accounting methodologies. The data used to quantify GHG emissions originated from five different accounting methodologies (M1 to M5) that were applied to six different farm management systems (FMS). Results were largely dictated by data availability from FMS to quantify individual GHG sources for methane (CH4), nitrous oxide (N2O), carbon dioxide (CO2) and net GHG emissions. Accounting methodologies depending on larger data availability and more detailed input parameters (M 1 and M2) were preferred when mitigation options were considered. M3 predicted enteric CH4 emissions from metabolizable energy intake (MEI) and results showed this method might over-estimate emissions, while the M5 demonstrated under-prediction. Similar results between M1, M2 and M4 were observed for enteric CH4 production. Differences in CH4 and N2O from manure management were essentially affected by the chosen prediction methodology to calculate enteric CH4, while the methodology to calculate N2O emissions from soil management and fertiliser application used default emission factors as guided by the Intergovernmental Panel on Climate Change (IPCC). Distinguishing between different functional units was critical, since different functional units ranked dairy systems differently. The results suggested that dairy systems that emitted the lowest GHG per animal or unit area did not necessarily emit lowest GHG emissions for a given level of production. Emissions expressed per kg fat and protein corrected milk (FPCM) across all farms ranged from 1.12 – 1.56 kg CO2-eq per kg FPCM, which was lower compared to the FAO estimate of 2.4 kg CO2-eq per kg FPCM. The results suggested further that the most important parameters to consider in estimating GHG emissions would be dry matter intake (DMI), feed quality and emission factors for CH4 from enteric fermentation and soil management, amount of N applied as fertiliser and emissions from purchased feed. A benchmarking model may be devised for South African dairy production systems. 5 The Bromide challenge in groundwater Bronwyn Reijnders and Norman Casey Department of Animal and Wildlife Sciences, University of Pretoria, South Africa E-mail: bronwynreijnders88@gmail.com Bromide (Br-) in ground water has emerged as a water quality constituent (WQC) of concern (COC) over the past decade. Water quality standards published in the late 20th century did not list Br as a water quality constituent let alone a COC. Research into groundwater chemistry since 1990 has shown the wide distribution and the varying levels of Br in groundwater. Since southern Africa is largely a surface water scarce region, many livestock farmers and game ranchers are dependent on groundwater for their animals and domestic supplies. This paper presents the collated results of wide-ranging monitoring of Br in groundwater over a fifteen year period and the results of controlled research into the potential endocrine disruptive effect of the anion. Recent research has demonstrated the potential of Br to disrupt the thyroid endocrine system resulting in adverse livestock production and subclinical pathology. The effects however, would be determined by the rates of intake and clearance, which is the focus of current research. 6 Dietary supplements to a low protein diet may affect the incidence of hepatic lipidosis in mink Connie Frank Matthiesen and Anne-Helene Tauson Department of Veterinary Clinical and Animal Sciences, Faculty of Health and Medical Sciences, University of Copenhagen, Grønnegårdsvej 3, DK-1870 Frederiksberg C, Denmark, Email: cmt@sund.ku.dk Hepatic lipidosis, i.e. hepatic lipid infiltration, is multifactorial and it may be caused by a number of factors such as low protein provision, feed deprivation, rapid accretion or mobilisation of body fat, all resulting in metabolic and nutritional imbalances. Our objectives were to investigate if supplementation of a low protein diet with nutrients acting as methyl donors, antioxidants or having insulinogenic properties could lower the incidence of hepatic lipidosis in growing mink from August to November when mortality, caused by hepatic lipid infiltration, often is high. Seventy-two young mink, 36 females and 36 males, were used in the study. The animals were allocated to six treatment groups each consisting of 6 males and 6 females. The controls were fed a conventional feed (29.6:54.2:16.2% of metabolizable energy (ME) from protein: fat: carbohydrate) whereas the 5 experimental groups were fed a low protein diet (20% of ME from protein, prepared by adding 5.5% corn starch, 3.5 corn oil, 6% potato mash powder, 1% sugar beet pulp and 8% water to the control diet) and supplemented with the crystalline amino acids alanine (0.8% - negative control - group 1), taurine (0.5% - group 2), arginine (0.5% group 3), and methionine (0.5% - group 4) or dextrose (2.5% of the corn starch was replaced by dextrose - group 5). Feed intake was recorded as an average per group per day. Animals were weighed and blood sampled once every third week. The liver was removed and liver and body weights were recorded of all animals that died during the experiment. All remaining animals were euthanized at the end of the experiment when liver, kidney and body weights were recorded. All livers were frozen for later analysis. Liver weight was significantly affected (P<0.001) by dietary treatment, status (dead or alive) (P<0.001) and interactions between dietary treatment and status (p<0.001). This resulted in significantly heavier livers among animals diagnosed with hepatic lipidosis compared to livers from euthanized animals without hepatic lipidosis. The survival rate was numerically higher for control (100%) and group 4 (methionine –100%) animals compared to group 1 (alanine –males 84%, females 100%), group 2 (taurine – males 84%, females 100%), group 3 (arginine – males 84%, females 84%) and group 5 (dextrose – males 67%, females 84%). It can be concluded that our preliminary results indicate that the methionine level in a low protein diet plays an important role for the incidence of hepatic lipidosis. 7 In vitro fermentation of various feed incubated with Warmblood or Icelandic horse faecal inoculum Christine Brøkner1, Julia L. Milling2, Hanne H. Hansen2 1 Department of Veterinary Clinical and Animal Sciences, Faculty of Health and Medical Sciences, University of Copenhagen, Grønnegårdsvej 3,1870 Frederiksberg C, Denmark. 2 Department of Large Animal Sciences, Faculty of Health and Medical Sciences, University of Copenhagen, Grønnegårdsvej 2, DK-1870 Frederiksberg C, Denmark. Email: christine.broekner@sund.ku.dk Alternative energy sources for sport horses are needed (Brøkner et al., 2012). High intake of sugar and starch affects blood plasma metabolites and regulatory hormones, which consequently results in health problems (Kronfeld et al., 2005). The aim of this study was to determine the in vitro digestibility and digestion kinetics of feeds of various fibre fractions in Warmblood and Icelandic horses. The following feeds were analysed: Micronized maize, oats, sugar beet pulp, potato pulp, wheat bran, meadow and standard hay. The feeds were milled in Cyclotec 1093 sample mill (FossTM TecatorTM CyclotecTM Sample Mill, Hillerød, Denmark) to 1 mm and 0.5 g sample was placed in each incubation jar. Three replicates were made of all 7 feeds along with three blanks. Fresh faeces from 5 horses of each breed from the same farm and fed the same diet mainly consisting of roughages were used. Faeces were transported directly to the lab in airtight and preheated (38°C) thermos. Faeces were diluted in preheated (38°C) deionised water and filtered through double layered cheesecloth. The incubation media consisted of liquid faecal solution mixed with buffer solutions. Feed samples were incubated in 90 ml media in a 38°C shaking heat chamber at 0.75 psi. Cumulative gas production i.e. primarily CO2 and CH4 were recorded continuously at every 5 min. for 48 h using the ANKOMRF system as described by Cattani et al. (2014). After end of incubation, the fermentation was stopped by placing the jars on ice. PH was measured immediately after and directly in the jar. All samples were filtrated to a fibre bag with water suction and 10 ml faecal medium were pipette into prepared SCFA vials and frozen at -20°C until analysis. All fibre bags were put in the heat oven at 60°C for 48 h. Feeds were analysed for DM, Ash, CP, CF, NDF, ADF and ADL, fibre bags for DM, NDF and ADF and faecal media for SCFA. The results are currently being processed and await further analysis. The final results and conclusion will be presented at the conference. 8 Graphene toxicity in experiments in a chicken embryo model Ewa Sawosz1, Slawomir Jaworski1, Marta Kutwin1, Anna Hotowy1, Mateusz Wierzbicki1, Marta Grodzik1, Natalia Kurantowicz1, Barbara Strojny1, André Chwalibog2 1 Division of Nanobiotechnology, Faculty of Animal Science, Warsaw University of Life Sciences, Ciszewskiego 8, 02-786 Warsaw, Poland, Email: ewa_sawosz@sggw.pl 2 Department of Veterinary Clinical and Animal Sciences, Faculty of Health and Medical Sciences, University of Copenhagen, Grønnegårdsvej 3, 1870 Frederiksberg, Denmark Graphene is a single atomic layer of a sp2-bonded carbon nanostructure. It is so far the thinnest two-dimensional allotrope of carbon that can freely exist. Graphene exhibits unusual mechanical, electronic and thermal properties that influence its biocompatibility. Moreover, the physically active surface of graphene may bind and activate cell receptors, especially located in cell membranes. Evaluation of the potential cytotoxicity of graphene is a key factor for medical applications, where flakes or a surface of graphene may be used as bioactive molecules, drug carriers, or biosensors. In the present work, effects of pristine graphene (pG) on the development of a living organism, with an emphasis on morphological and molecular states of the brain, were investigated using a chicken embryo model. Fertilized chicken eggs were divided into the control group and groups administered with pG suspended in milli-Q water at the concentrations of 50, 100, 500, 1000, 5000 and 10000 μg/L (n=30 per group). The experimental solutions were injected in ovo into the albumen, and then the eggs were incubated. After 19 days of incubation, the survival, weight of the body and organs, and blood serum biochemical indices were measured. The brain samples were collected for microscopic examination of brain ultrastructure and measurements of gene and protein expression. Survival of embryos was significantly decreased after treatment with pG, but the body and organ weights as well as biochemical indices were not affected. In all treatment groups, some atypical ultrastructures of the brain were observed, but they were not enhanced by the increasing concentrations of pG. Expression of proliferating cell nuclear antigen (PCNA) at the mRNA level was down-regulated, and the number of PCNA positive nuclei was significantly reduced in the 500 – 10000 μg/L groups comparing with the control group, indicating a decreased rate of DNA synthesis in the brain. The present results demonstrate some harmful effects of the applied pG flakes on the developing organism, including brain tissue, which ought to be considered prior to any medical applications. 9 In vitro and in vivo evaluation of the effects of graphene nanoplatelets on glioblastoma multiforme Slawomir Jaworski1, Ewa Sawosz 1, Marta Kutwin1, Mateusz Wierzbicki1, Marta Grodzik1, Mateusz Hinzmann1 André Chwalibog2 1 Division of Nanobiotechnology, Faculty of Animal Science, Warsaw University of Life Sciences, Ciszewskiego 8, 02-786 Warsaw, Poland, Email: jaworski.slawek@gmail.com 2 Department of Veterinary Clinical and Animal Sciences, Faculty of Health and Medical Sciences, University of Copenhagen, Grønnegårdsvej 3, 1870 Frederiksberg, Denmark Glioblastoma multiforme (GBM) is the most common and the most aggressive primary malignant brain tumour. It comes from astrogial cells and is characterized by infiltrative growth, intensive migration and fast spread of cancer among brain tissue. Glioblastoma models existing in vivo are based on the inoculation of glioma cells into the rodent brain or the use of transgenic mice causing spontaneous tumours. However, these models are weak because are characterized by variable growth rate and poor penetration, and difficulty of obtaining morphological data. The chicken embryo chorioallantoic membrane (CAM) model is a well-known method for studying cancer’s biology. The CAM model is well-established method to keep explanted material alive and supply it with oxygen and nourishment. Graphene is a single atom-thick material with exciting potential. Graphene can be used in many industries, from electronics to biomedical areas. However, we still know little about its toxicity and biocompatibility. Herein, we performed a study on the toxicity of graphene nanoparticles (GN) by examining the influence of GN on the morphology, mortality, viability, membrane integrity and type of cell death of U87 and U118 glioma cells We also checked impact of GN on glioma tumours (U87 and U118) cultured on the chorioallantoic membrane (CAM) in the area of formed blood vessels. The present results demonstrate that graphene is toxic to glioma cells, but activated apoptosis only in the U118 cell line without inducing necrosis, indicating the potential applicability of GN in anticancer therapy. 10 The influence of newly produced platinum nanoparticles on model organism development Barbara Strojny1, Anna Grabowska1, Marta Grodzik1, Mateusz Hinzmann1, Adam Lewera2, Aleksandra Lisek1, Barbara Mielcarek2, Elżbieta Migurska 1, Sandra Sierankowska1, Klaudia Strucińska1, Anna Tyszko1 1 Division of Nanobiotechnology, Faculty of Animal Sciences, Warsaw University of Life Sciences, Ciszewskiego 8, 02-786 Warsaw, Poland, Email: barbara_strojny@sggw.pl 2 Laboratory of Electroanalytical Chemistry, Faculty of Chemistry, University of Warsaw, Pasteura 1, Warsaw, Poland Nanoparticles, due to their small size and enormous surface, have unique physicochemical properties. Platinum has been widely used in chemotherapy as a salt-based agent known as cisplatin. However, its main disadvantage is a high toxicity to an organism. Platinum nanoparticles have different features than Pt ions in salts and may, therefore, prove to be useful in medical application, though the toxicity of Pt nanoparticles on living organism has to be investigated. Recently produced platinum nanoparticles are characterized by very clear surface and narrow range of size, because of the patented synthesis method, based on mechanical separation of platinum particles. There are no additional surfactants in the reaction mixture, only the substrate (platinum chloride II) and the reducing agent (polyethylene glycol). The objective of this study was to evaluate the influence and potential toxicity of bare platinum nanoparticles on model organism development - chicken (Gallus gallus) embryo. Fertilized eggs (n=80) were divided into 4 equal groups: Control (not treated) and treated with the increasing concentration of platinum nanoparticles, Pt 10 (10 μg/ml), Pt 20 (20 μg/ml) and Pt 40 (40 μg/ml). The suspension of nanoparticles was given by injection into the albumen at the first day of incubation. The injection consisted of 0.3 ml nanoparticles hydrocolloid. On the 19th day of incubation, the embryos were sacrificed and weighed. Their development was compared to Hamilton and Hamburger stages. Blood samples were taken to prepare blood smears and organs (brain, heart, liver and spleen) were weighed. No significant differences in weight of embryos and their organs between the groups were measured. All embryos were developed normally, however, in Pt-treated groups the mortality in the beginning of incubation period was higher than in the control group. Analysis of blood showed increased level of leukocytes (granulocytes and lymphocytes) and thrombocytes in the Pt 40 group, as well as slight haemolysis of erythrocytes. The results suggest that investigated nanoparticles of platinum may have toxic effects on living organisms. Pure nanoparticles may affect red blood cells, causing haemolysis, and may have impact on immunological system. Therefore, further investigations concerning platinum nanoparticles should be performed. 11 Comparison of in vitro cytotoxicity of platinum nanoparticles and cisplatin against human glioma cells line Marta Kutwin1*, Ewa Sawosz1, Sławomir Jaworski1, Mateusz Wierzbicki1, Mateusz Hinzmann1, André Chwalibog2 1 Division of Nanobiotechnology, Faculty of Animal Science, Warsaw University of Life Sciences, Ciszewskiego 8, 02-786 Warsaw, Poland, *Email: marta_prasek@sggw.pl 2 Department of Veterinary Clinical and Animal Sciences, Faculty of Health and Medical Sciences, University of Copenhagen, Grønnegårdsvej 3, 1870 Frederiksberg, Denmark Gliomas are the most aggressive and common primary tumours of central nervous system (CNS). Platinum based drugs, like cisplatin, are widely used in medicine as chemotherapeutic agents for the treatment of tumours. However, the poor penetration of CNS is characteristic for drugs containing platinum and the chemotherapy based on platinum salt can be ineffective against brain tumours. Recently the new biologically active platinum nanoparticles (NP-Pt) can be useful therapeutics in gliomas cancer therapy. The objective of the study was to investigate and compare the toxicity of NP-Pt and cisplatin and their proapoptotic or necrotic properties in examination with glioma cells line (U87). NP-Pt were characterized by electron transmission microscopy. The NP-PT and cisplatin were incubated with glioma cells for 24h. The biological properties of nano-Pt and cisplatin were evaluated through morphology, metabolic activity, viability, mortality and the type of cell death of U87 glioma cell line. NP-Pt and cisplatin are toxic to glioma cells and activated apoptosis in the U87 cell line. NP-Pt treatment activated programmed cell death without activating necrosis. The effectiveness of cisplatin and NP-Pt treatment against glioma cells were mostly at the same level. The results suggest not only genotoxic and cytotoxic effect caused by NP-Pt, but also the cell cycle arrest and apoptosis in NP-Pt treated cells. The comparison between effectiveness of cancer cell treatment by NP-Pt and cisplatin showed promising results for future studies. The results indicate that properties of NP-Pt might be utilized for the brain cancer therapy. 12 How graphene family materials affect Listeria monocytogenes and Salmonella Enteritidis strains Natalia Kurantowicz1, Sławomir Jaworski1, Ewa Sawosz1, Marta Kutwin1, Barbara Strojny1, Mateusz Wierzbicki1, Jacek Szeliga1, André Chwalibog2 1 Division of Nanobiotechnology, Faculty of Animal Science, Warsaw University of Life Sciences, Ciszewskiego 8, 02-786 Warsaw, Poland, Email: natalia_kurantowicz@sggw.pl 2 Department of Veterinary Clinical and Animal Sciences, Faculty of Health and Medical Sciences, University of Copenhagen, Grønnegårdsvej 3, 1870 Frederiksberg, Denmark This work examined the effect of three different graphene nanostructures (GFMs) on chosen food-borne bacteria strains: Gram-positive (G+) – Listeria monocytogenes ATCC 1911, and Gram-negative (G-) –Salmonella enterica subspecies enterica serovar Enteritidis ATCC 13076. Both were obtained from LGC Standards (Lomianki, Poland). Chosen bacteria are pathogenic and morphologically different. The nature of the cell wall is the key difference between G+ and G− bacteria. In general, G+ bacteria have a thick peptidoglycan layer outside cells, while G− strains have a much thinner peptidoglycan layer between the inner and outer membrane. High thermal stability, high mechanical strength and relatively good biocompatibility for humans suggest that graphene structures can be effective antibacterial agents. The study compared the toxicity of different forms of graphene; pristine graphene (pG), graphene oxide (GO) and reduced graphene oxide (rGO) towards bacteria strains with special emphasis on visualization of their interaction. All forms of graphene decreased number of bacteria colonies in probes with graphene concentration of 250 mg/L. The high concentration of graphene, reduced growth of both bacteria strains 100 %. Moreover, as low concentration of GO as 25 mg/L caused a drop in the number of bacteria colonies, as well and reduced bacteria growth by 99.9% and 100% for Listeria monocytogenes and Salmonella E. respectively. To sum up, graphene oxide gave the best result. The shape and size of graphene particles and their interactions with bacteria strains were inspected using a JEM-1220 transmission electron microscope at 80 KeV. All forms of graphene immobilized bacteria. Bacteria were aggregated, and a strong affinity occurred between bacteria and edges of pG and rGO, even small pieces of nanoparticle edges bound bacteria. Bacteria strains attached only to GO surfaces. There were no bacteria cells outside the GO layers and there were no graphene particles inside bacteria cells so it is extremely probably that graphene has bacteriostatic properties. Bacteria-graphene suspensions have higher zeta potential than bacteria strains. The raised zeta potential could be useful to measure antibacterial activity of graphene towards different bacterial strains. A strong correlation exists between reduced bacteria growth and the zeta potential. Probably, the ability of bacteria and graphene to form colloidal bonds is related to exposure time. The higher the absolute value of the zeta potential, the more stable the colloidal bond, so the exposure time is longer and the antibacterial activity of the agent is more potent. 13 Validation of gypsum reduction process using the mintek pyrosim model R. K. Tewo1, J. P. Maree1, S.J Ruto2, B. S Xalakashe3, A.V Kolesnikov2, G.B Mohajane1 1 Department of Environmental, Water and Earth Sciences, Tshwane University of Technology, Private Bag X680, Pretoria, 0001, South Africa, Email: kimuaitewo@gmail.com 2 Department of Chemical and Metallurgical Engineering, Tshwane University of Technology, Private Bag X680, Pretoria, 0001, South Africa 3 Pyrometallurgy Division, Mintek, Private Bag X3015, Randburg, 2125, South Africa The validation of the reduction process of waste gypsum using Pyrosim Mintek simulink gives better understanding of its thermodynamic parameters. The process of validation is one of the steps closer to realizing downstream gypsum waste beneficiation for the recovery of valuable materials. The comparison of the simulated and the experimental data forms the basis of the study. In this study, Mintek Pyrosim model was used to validate results from the simulated and the experimental data. The effects of temperature, coal to gypsum ratio in the reduction of waste to calcium sulphide, energy requirements and different coals were investigated. Thermodynamic parameters such as (ΔGo), (ΔHo) and (ΔSo) have also been simulated and has confirmed the reaction is an endothermic process. The process conditions found to be suitable for a good conversion were: a temperature range from 1100-12000C, gypsum to coal ratio 4:1 and for 100 kg/hr gypsum and 25 kg/hr coal the energy required was 79.9 kWh. 14 Processing of calcium sulphide to calcium carbonate during batch studies S. Nomfazi1*, J. Maree1, L. Cele2 1 Department of Chemistry; 2Department of Environmental, Water and Earth Sciences * Faculty of Science, Tshwane University of Technology, Private bag X680, Pretoria, 0001, South Africa, *Email: princessmyoli@gmail.com. This paper reports a process for converting calcium sulphide into calcium carbonate in a pilot plant. Calcium sulphide was suspended in water to form slurry. The reaction of gaseous carbon dioxide with the calcium sulphide slurry led to the formation of calcium carbonate and hydrogen sulphide gas. XRD analysis of calcium carbonate produced during batch studies showed > 98% purity. The hydrogen sulphide gas generated was absorbed in a ferric iron solution to precipitate elemental sulphur. The effects of various parameters such as stirring rate, temperature and calcium sulphide concentration on reaction time were investigated. The quality of the final calcium carbonate produced was strongly dependent on the purity of calcium sulphide utilized and was ascribed to the filter step in the configuration. The predominant crystal structure was found to be calcite, which has wide applications in industry. The calcium sulphide was derived from the calcining of waste gypsum in the presence of coal. If the process is viable on large scale, it would mean that the problem of stockpiled gypsum could see a partial solution. The recovery of useful by products calcium carbonate and sulphur from waste gypsum could be very beneficial to South African industry. 15 Oocytes of the South-African clawed frog, Xenopus laevis, as a model for characterizing K+ channels Maria A. Tejada, Karen Molbaek, Martias E. Martin and Dan A. Klaerke Department of Veterinary Clinical and Animal Sciences. Faculty of Health and Medical Sciences, University of Copenhagen. Dyrlaegevej 100, 1870 Frederiksberg, Denmark. E-mail: tejada@sund.ku.dk The wild South-African clawed frog Xenopus laevis has been used as a laboratory animal for the study of reproductive physiology, pregnancy tests, as well as studies on muscle and kidney physiology, since the beginning of the twentieth century. It was only in the 70’s Xenopus oocytes were introduced as an expression system for proteins. It was demonstrated that microinjection of mRNA into the cytoplasm of oocytes, resulted in the synthesis of human haemoglobin. Ever since, Xenopus oocytes have been used worldwide, given their efficiency in translating foreign mRNA into proteins. Advantages of this expression system include: easy handling of the relatively large oocytes (~1 mm in diameter); frogs are easily kept and bred in captivity; oocytes are resistant and fully equipped with an efficient translating machinery ready to synthesize proteins; oocytes have a low expression of endogenous membrane proteins, which gives a low background to the heterologously expressed proteins. Given these characteristics, Xenopus oocytes have been widely used for the functional expression of membrane proteins such as potassium channels. Xenopus oocytes are virtually impermeable to water, as they do not express endogenous aquaporins (water channels). Therefore we have used the oocytes to develop an assay for identification of K+ channels, which are regulated by cell volume changes. If aquaporins are co-expressed with K+ channels, the oocytes promptly change their volume, when the extracellular osmolality is changed, and our setup allows simultaneous and precise measurements of the oocyte volume changes and the activity of the expressed K+ channels. In absence of co-expressed aquaporins possible effects of changes in osmolality without changes in cell volume may be observed. A number of K+ channels examined in this way, have proven to be either strictly regulated by small changes in cell volume, or totally insensitive. In this way we have been able to identify at number of K+ channels, which act as cell volume sensors through a “functional interaction” with aquaporins. Another unique feature of the Xenopus oocytes is that vesicles that are injected into the oocytes cytoplasm will fuse with the plasma membrane. That means that purified protein after reconstitution into phospholipid vesicles may be “transplanted” to Xenopus oocytes for electrophysiological characterization. At present, we are developing such procedures for Plasmodium falciparum K+ channels, which can only be expressed in yeast cells; a cell type, which is in itself quite unsuitable for electrophysiological approaches and thus for characterization of ion channels. 16 The conservation value of the avifauna of the Kathu Forest, South Africa Rion Lerm*, Gerard Malan, Ray Jansen Faculty of Science, Tshwane University of Technology, Private Bag X680, Pretoria 0001, South Africa, *Email: austringerever@yahoo.co.uk Prioritising areas for conservation requires the need for species diversity information. The protected Kathu Forest in the semi-arid Northern Cape Province of South Africa is characterised by unusually large Camel Thorn (Vachellia erioloba) trees. Local mining activities, urbanisation and timber harvest threaten this unique forest and its protected keystone structures, the Camel thorns. Since little is known about the avifauna occupying the forest, this study investigated the avian diversity and conservation value in various study sites of the Kathu Forest. Avian surveys were carried out during the breeding and non-breeding seasons and point counts were employed to record avian species, abundances, behaviours and microsite uses. Species were subsequently divided into foraging strategies and nesting categories. Counts were carried out at four study sites: urban, golf course, game farm and a control site (located outside the forest). For both seasons, strongly significant differences were observed between the avian abundances and –alpha diversities of the study sites with values descending from the urban site to the control. Resting behaviour showed the strongest significant difference between study sites for the breeding season as well as crown- and canopy use. Avian foraging strategies and –nesting categories were significantly associated to the study sites for the breeding season only. Only the breeding season delivered indicator species, the Orange River White-eye (Zosterops pallidus) being the strongest indicator overall, but specifically to the urban study site. The results suggest that the man-made study sites (urban and golf course) in the forest are hosts to a larger and more diverse avian community than the natural sites (game farm and control). Apart from extensive disturbance from humans and pollution and contrary to expectation, avifauna survive in the urban and golf course sites and seem to perform far better than those communities occupying more natural habitats such as the game farm and control site. Furthermore, areas outside the forest do contain unique avian communities, but as with the forest, none of the species are of conservation concern. Nonetheless, it is necessary for conservation authorities to maintain the vegetation structure of the Kathu Forest, but also realize the importance of avian communities outside the forest. 17 Angora goats: genetics and product quality Carina Visser Department of Animal and Wildlife Sciences, University of Pretoria, Private Bag X20, Hatfield, 0028, South Africa, Email: carina.visser@up.ac.za South Africa is a major role player in the global mohair industry, producing almost 50% of the world product. Apart from being the largest producer, South Africa is also widely considered as the most reliable producer of a good quality clip of mohair worldwide, with a consistently fine clip showing little variation. This was achieved by intense selection for increased mohair production from 1970 to 1990, with no selection directed towards weaning weight. Body weight was compromised leading to smaller, unthrifty animals with an inability to survive sub-optimum conditions. It has been suggested that there is a genetic basis for the negative association observed between selection for fine hair production and fitness traits, but this hypothesis has not been proven yet. Molecular genetics has provided an opportunity to investigate the genetic diversity of superior and inferior producing Angora goats on a DNA level. Whole blood samples of 48 unrelated Angora goats were collected from the DNA biobank for small stock (GADI, NDAFF). The animals were selected based on two important phenotypic traits, namely fibre diameter (FD) and fleece weight (FW). Animals with both inferior and superior performance were chosen in order to ensure a diverse group of animals. The animals were investigated using two approaches. Firstly, they were genotyped with the Illumina goat SNP50 beadchip that features 53 347 SNP probes distributed across the whole goat genome. Sample-based quality control was performed, and no individuals were removed for having a call rate below 98%. A total of 9 588 SNP were removed after marker-based quality control filtering, including: 2 279 SNP with low call rate (<98%), 6364 SNP for having a MAF below 5% and 1681 SNP for violating HWE (p<0.001). The remaining 46 983 (88.1%) SNP were polymorphic for the sampled individuals. Secondly, it was postulated that variations in genes associated with keratin-associated proteins (KAP) and intermediate filament proteins (KIF) should play a functional role in the expression of fibre quality. Identification of favourable alleles affecting mohair production will assist in accelerating genetic progress. Characterization of KAP 1.1, 8.1 and 13.3 have shown promising results with regards to their effects on fibre formation, and quality traits of cashmere. Variation in KAP1.1, KAP8.1, KAP13.3 were investigated in South African Angora goats, to identify possible association with quality traits in mohair. This should lead to the identification of candidate genes that affect economically important traits. 18 Overcoming the genetic bottleneck – the European bison case Wanda Olech Faculty of Animal Science, Warsaw University of Life Sciences, Ciszewskiego 8, 02-786 Warsaw, Poland, Email: wanda_olech@sggw.pl Species Bison bonasus that ninety years ago was on the verge of extinction, has been saved first through captive breeding, and since 1952 by gradual releasing the animals to the wild. All those efforts have brought present world population of the wisent to over 5000 individuals, out of which over 1600 remains in more than 200 herds in captivity and other enjoys freedom in 39 free ranging or semi-free herds. For a long time, the only countries with free ranging European bison were: Poland (having the largest total population of 1300 individuals), Lithuania, Byelorussia, Russia and Ukraine, but since 2004 and 2012 respectively, wild herds have been established also in Slovakia and Romania and Germany. Reintroduced wisents enriched the local fauna and through their foraging patterns contributed to an increase of biodiversity within their home ranges. The presence of wisents is also a major tourist attraction that helps to create jobs, and provides opportunities for a local business. There are, however, also certain problems that are connected with restitution of this species and one of most important is proper management of its genetic pool. The gene pool of the European bison is very small because it consists only of 12 sets of genotypes (founders). Additionally the species is divided into two genetic lines - Lowland line (LB) was derived from seven founders and Lowland Caucasian line (LC) from all 12 ancestors. A large difference is observed between genetic structures of two lines. Significantly different are values of inbreeding and founder’s contribution as well as the distribution of paternal and maternal lines. Within LB line all males carry Y chromosome of one founder Plebejer but within LC line still small percentage of males are carriers of other Y-chromosomes. The maternal influence in LB line belongs to the founder 42 Planta but in LC line to 89 Bilma. The future breeding of the species should be oriented towards the separation of both lines, and higher amount of exchanges between the captive and free ranging part within every genetic line. Important for the future is an increased use for reproduction animals that are carriers of rare paternal or maternal lines. Nevertheless, the loss of the wisent would be an irreversible impoverishment of European fauna, and serious setback for nature conservation. Its salvage against all odds can become a symbol of a success in preventing further decline of our natural resources. 19 Wakkerstroom Secetarybird Project: Diet and productivity Eleen Strydom Department Nature Conservation, Faculty of Science, Tshwane University of Technology, 129 de Bruyn Street, Weavindpark, Pretoria, 0184, South Africa, Email: strydommme@tut.ac.za The Secretarybird (Sagittarius serpentarius) is uncommon to locally fairly common in southern Africa. A decrease in reporting rates between SABAP1 (1985-1993) and SABAP2 (2007 - present day) shows a drastic, but unconfirmed reduction in Secretarybird distribution. Not surprisingly, the IUCN recently upgraded the Secretarybird's conservation status to Vulnerable. Reasons for the above decline may include: (1) habitat degradation due to overgrazing and bush encroachment, (2) disturbance and loss of habitat due to afforestation and crop cultivation, (3) human disturbance, (4) poisoning, (5) road mortalities, and (6) extreme and regular veld fires. To study some of the possible reasons for the decline in the bird’s distribution, the Department of Nature Conservation and BirdLife South Africa came together to start the Secretarybird Project that will be conducted in the Greater Wakkerstroom area over a 17month period. The project has two legs namely diet and productivity. The Secretarybird’s diet consists of a wide variety of prey taxa and will be studied employing photographs of regurgitation remains and the examination of pellets. Although the Secretarybird is locally common in southern Africa, productivity is largely unknown due to unpredictable breeding patterns and difficulty in quantifying non-breeding years. Productivity, as the number of young (brood size) that fledge per nest per year, will firstly be calculated. Secondly, regular visits to nests will help to determine what number of pairs succeeds in each stage of the nesting period, i.e. laying, hatching and fledging. Nesting success and productivity has been investigated for half of the breeding season in the Greater Wakkerstroom area. 20 Evaluation of chimpanzee (pan troglodytes) diet nutrient composition based on the South African Medical Research Council Food Composition Tables Ulrich Mwimbi1, Francois Siebrits1 & Rebecca Mbatha2 1 Faculty of Science, Tshwane University of Technology, South Africa, Email: mwimbiku@tut.ac.za,2University of South Africa Studies of captive animals have the advantage of being in a controlled environment. However, they may not adequately represent the physiology of free-ranging animals. The feeding strategies, gut morphology, and nutrient requirements of primates are challenges faced in zoological institutions. In free ranging, the resultant multidirectional of primate's eating behaviour and nutritional requirements, the plants in its diet (with their very specific carbohydrates), and the animal's gut function and symbionts are essential to the animal's health. The specific nutritional requirements for chimpanzees are still not completely understood. Most diets analyses are based on human recommended daily allowances (RDA). However the questions, which are the objectives, are to know (1) if the chimpanzees are as humans and could be sustained by the same diet as human, (2) the result of nutrition imprinting on the conservation of the captive diets to diets in natural environment. We used the Medical Research Council Food Nutritional Tables for South Africa to determine their nutritional composition; International Network of Feed Information Centres (INFIC) nomenclature system to classify the feeding items and Zootrition™ program to compare the chimpanzee’s National Zoological Gardens of South Africa diet sheet to natural feeding stuffs of chimpanzee and to the Primates requirements (Nonhuman and human, from National Research Council). The natural feed of chimpanzees falls in class 4, 5, 6 and 7 of the INFIC nomenclature while that of the National Zoological Gardens diet falls in class 2, 4, 5, 6, and 7.The nutritional composition of the diet showed unbalanced nutrients in terms of protein and vitamins and minerals. The results on CP content of the chimpanzee’s diet showed that protein level was high, but less than the critical limit of 20%. The Ca, Mg and Fe concentrations were high but Zn was deficient in the NZG diet. In the case of vitamins, vitamin A was in high contents and vitamins E and D deficient. 21 Energy expenditure in horses measured with the 13C-bicarbonate tracer technique Rasmus Bovbjerg Jensen1*, Peter Junghans2 and Anne-Helene Tauson1 1 Department of Veterinary Clinical and Animal Sciences, Faculty of Health and Medical Sciences, University of Copenhagen, Grønnegaardsvej 3, 1., 1870 Frederiksberg C, Denmark, *Email: ralle@sund.ku.dk 2 Leibniz Institute for Farm Animal Biology, Institute of Nutritional Physiology ‘Oskar Kellner’, Wilhelm-Stahl-Allee 2, 18196 Dummerstorf, Germany The energy required for maintenance varies and some horses gain weight easily, even under conditions where other horses will lose weight if not fed additional feed. These horses, especially some pony breeds and Icelandic horses are commonly called “easy keepers”. The 13 C-bicarbonate tracer technique can be used for indirect determination of the CO2 production and to estimate energy expenditure. The technique is based on 13C kinetics in expired air after administration of a dose of 13C labelled bicarbonate. The aim of the present experiment was to test the 13C-bicarbonate tracer technique under field conditions, and to estimate energy expenditure in three groups of horses with expected differences in energy expenditure. Four Icelandic horses (initial body weight (BW): 347±5 kg), four sports ponies (initial BW: 429±38 kg) and four warmblood horses (initial BW: 546±37 kg) were used in this experiment. All horses were measured at three occasions approximately 4 weeks apart. A single dose of 13C-bicarbonate (500 mg/horse) was administered intravenously. Exhaled breath was collected in breath bags with a volume of approximately 1 litre by using a mask with a two-way non-rebreathing valve system. The mask covered one of the nostrils and a breath sample was taken within 30 seconds. Two baseline samples were taken before and 12 samples after (5, 10, 20, 30, 40, 60, 90, 120, 150, 180, 240 and 360 minutes) administration of 13 C-bicarbonate. Area under the 13C-enerichement curve was used to calculate CO2 production, when a recovery factor of 13C of 0.72 was used. Energy expenditure was calculated from Brouwer’s equation, and a respiratory quotient of 0.82 was used to estimate O2 consumption. Energy expenditure differed between groups of horses, with Icelandic horses having significantly lower energy expenditure (519 kJ/BW0.75 per day) than sports ponies (748 kJ/BW0.75 per day) and warmblood horses (754 kJ/BW0.75 per day). It was possible to measure differences in energy expenditure with the 13C-bicarbonate technique under field conditions. The difference between the warmblood and Icelandic horses in the present experiment was expected. The sports ponies might not be considered as “easy keepers”, as their energy requirement was similar to the warmblood horses. 22 The oral 13C bicarbonate technique for measurement of energy expenditure in dogs under field conditions Caroline Larsson1*, Øystein Ahlstrøm2, Peter Junghans3 and Anne-Helene Tauson1 1 Department of Veterinary Clinical and Animal Sciences, Faculty of Health and Medical Sciences, University of Copenhagen, Grønnegårdsvej 3, 1870 Frederiksberg, Denmark, *Email: cala@sund.ku.dk 2 Department of Animal and Aquacultural Sciences, Faculty of Veterinary Medicine and Biosciences, Norwegian University of Life Sciences, P. O. Box 5003, N-1432 Ås, Norway. 3 Leibniz Institute for Farm Animal Biology, Research Unit Nutritional Physiology ̒ Oskar Kellner ̓, Dummerstorf, Germany Matching the energy demands of the dog might be challenging. Factors such as breed, physical activity and ambient temperature may all have a great influence on the daily energy requirements. The objective with the present study was to assess the non-invasive oral 13Cbicarbonate technique (o13CBT) for estimation of CO2 production (RCO2) and energy expenditure (EE) in sled dogs under controlled conditions in the field. Eight Alaskan huskies (body weights, BW: 19.5 ±1.6 kg) were included in the study. The dogs were exercised in sled run, pulling a four-wheeler with a speed of approximately 18 km/h. The total distance was 146 km, spread out over two periods of each three weeks. Data was collected in fasted state during rest and during exercise in the third week of each period. The resting conditions were considered to be active rest, as the dogs were kept outside, chained to their individual doghouse placed side by side. After a baseline sample of expired breath air was collected into a breath bag by using a mask with a two-way non-rebreathing valve system, an oral dose (115 mg) of NaH13CO3 incorporated in a small piece of liver paté was administrated. Thereafter, breath samples were collected at frequent time intervals for the first hour (5, 10, 15, 20, 30, 40 and 60 minutes after 13 C administration), and then with 30 min intervals until 4 hours. During days with exercise, the dogs started their exercise session (12 km) immediately after the 13C-dosis was administrated and were quickly stopped when time for breath sampling. The 13C/12C ratio in collected breath bags was analysed with an IRIS infrared analyser and from the 13C kinetics RCO2 and EE were estimated. Energy expenditure measured during resting conditions was higher (p = 0.01) in period 2 (1598 kJ/kg BW0.75/d with the 95 % confidence interval, CI: 1209 – 1986), compared to period 1 (1151 kJ/kg BW0.75/d with 95 % CI: 762 – 1539). The 95 % CI´s show the great variation between individual dogs. Changed weather conditions (i.e. decreased temperature and snow) from period one to period two may likely explain the effect of period. Data collected during exercise could not be used for further analyses, as the 13C/12C ratio could not be measured. In conclusion, the o13CBT can be used in the field for estimation of EE during resting conditions, but not during exercise whit the experimental setup used in this study. 23 Species management plan for the Great Crested Newt Triturus cristatus (Laurenti 1768) in Poland Krzysztof Klimaszewski Faculty of Animal Sciences, Warsaw University of Life Sciences, Nowousrynowska 166, 02786 Warsaw, Poland, Email: krzysztof_klimaszewski@sggw.pl The plan was prepared for Polish government under the project “Development of renaturalisation plans for natural and species habitats in Natura 2000 sites and management plans for selected species covered by the Birds and Habitats Directives”. Great Crested Newt is an amphibian species widely distributed across most of Europe, but has suffered severe declines in most of the 37 known range countries. The main goal of a plan was to define threats and find measures to ensure the maintenance or restoration where necessary, of viable crested populations across Poland. A series of general objectives are outlined and specific actions were proposed. Among others the lack of knowledge about detailed distribution is main problem in conservation measures planning. Therefore plan suggests distribution surveys, population and conservation status monitoring, scientific research, habitat management recommendations and public awareness programmes. 24