Construction and characterization of a BAC library for sunflower (Helianthus annuus L.)

A bacterial artificial chromosome (BAC) library was constructed using the sunflower (Helianthus annuus L.) restorer line RHA325, which carries the restorer gene Rf1 and the Pl2-gene conferring resistance to downy mildew. High molecular weight DNA was prepared from nuclei using leaf material from two-week old seedlings. The library was constructed using the HindIII site of pBeloBAC11. The current BAC library comprises 104,736 clones. The insert size of the clones varied between 20 and 270 kb, with an average insert size of 60 kb. The whole 1.9× sunflower BAC library was spotted in duplicate on four high-density filters, each carrying 55,296 clones. The content of organellar DNA, which was estimated by colony hybridisation against the mitochondrial probe coxI and the chloroplast probe rbcL, proved to be less than 0.03 and 0.1%, respectively. BAC pools, allowing PCR-based screening, were made and used to identify positive BAC clones for the markers OP-K13_454, closely linked to the restorer gene Rf1. The PCR-based screening was verified by the results obtained for this marker by colony hybridisation.     

Antioxidative properties of the essential oil from Pinus mugo

The essential oil from Pinus mugo (PMEO) was tested on its antioxidative capacity. For this purpose, several biochemical test systems were chosen (e.g., the Fenton System, the xanthine oxidase assay, or the copper-induced oxidation of low-density lipoprotein (LDL)). The results show that there is moderate or weak antioxidative activity when tested in aqueous environments, like in the Fenton system, xanthine oxidase induced superoxide radical formation, or in the HOCl driven fragmentation of 1-aminocyclopropane-1-carboxylic acid (ACC). In contrast, when tested in more lipophilic environments (e.g., the ACC-cleavage by activated neutrophils in whole blood) the PMEO exhibits good antioxidative activity. PMEO does also show good antioxidative capacity in another lipophilic test system (i.e., the copper induced oxidation of LDL). Some components of PMEO (i.e., Delta(3)-carene, camphene, alpha-pinene, (+)-limonene and terpinolene) were also tested. As the PMEO, they showed weak or no antioxidant activity in aqueous environments, but some of them were effective antioxidants regarding ACC-cleavage by activated neutrophils in whole blood or copper-induced LDL-oxidation. Terpinolene, a minor component of PMEO, exhibited remarkable protection against LDL-oxidation.     

Bioaccumulation of melamine in catfish muscle following continuous, low-dose, oral administration

In this study, catfish muscle was analyzed for melamine (MEL) and cyanuric acid (CYA) residues following experimental feeding with low doses of MEL and MEL and CYA (MEL+CYA) and with the insoluble melamine-cyanurate complex (MEL=CYA). Catfish were daily fed 0.1 mg/kg BW of MEL for 15, 28, or 42 days, 0.1 mg/kg BW of MEL+CYA for 28 days, 2.5 mg/kg BW of MEL+CYA for 14 days, or 400 mg/kg BW of MEL=CYA for 3 days. Residues in the tissue were determined by LC-MS/MS. MEL was extracted with acidic acetonitrile, followed by defatting with dichloromethane, and isolated with cation exchange solid phase extraction (SPE). For CYA analysis, fish were extracted with dilute acetic acid, defatted with hexane, and cleaned up with a graphitic carbon SPE. Catfish fed 0.1 mg/kg BW of MEL reached a maximum muscle residue concentration of 0.33 ± 0.04 mg/kg (ppm) after 28 days of continuous feeding. The same concentration was found for MEL+CYA feeding at the 0.1 mg/kg BW level for 28 days. Feeding at 2.5 mg/kg BW of MEL+CYA yielded muscle concentrations above the 2.5 mg/kg level of concern for most of the study fish. Finally, catfish fed high levels of the MEL=CYA complex (400 mg/kg BW) accumulated relatively little MEL in the muscle (0.14 ± 0.07 mg/kg) and, unlike treatment with MEL+CYA, did not form renal melamine-cyanurate crystals. Appreciable concentrations of CYA were not detected in any of the muscles tested. These studies provide data to model the bioaccumulation of triazine residues into edible fish tissue as a result of the continuous consumption of adulterated feed.     

Kinetics of jump landing in agility dogs

A recent survey reported an increased risk of injury in dogs participating in agility, a competitive canine sport involving different jumping activities. The aim of this study was to quantify the kinetic parameters during jump landing for commonly used obstacle types. It was hypothesised that with increasing obstacle height, the vertical force and vertical and accelerative horizontal impulse will increase as a result of a lengthened aerial phase, a more acute landing angle and the need to convert potential into forwards kinetic energy. Simultaneous kinetic and kinematic data were recorded from 11 competition agility dogs jumping over obstacle combinations of different height and inter-obstacle distance. Speed and landing angle of the second of the two consecutive jumps were successfully controlled by obstacle height and distance between obstacles. Statistical analysis showed differences between obstacles for peak vertical force, vertical impulse and accelerative horizontal impulse (increasing values with more acute landing angles). Extremely high peak vertical force was observed in the forelimbs (4.5 times bodyweight) when landing from a hurdle jump at high speed. Further detailed studies into the consequences for internal limb structures are warranted in order to clarify how this might be related to injury.     

The environmental safety of eprinomectin to earthworms

A study was conducted to assess the environmental safety of the endectocide eprinomectin to the earthworm Lumbricus terrestris under conditions mimicking typical product use on pasture. The LC50 value of eprinomectin in artificial soil after 28 days of exposure is higher than the levels expected in feces from dosed cattle or in soil fertilized with manure from dosed cattle, which indicates a wide margin of safety for this compound to earthworms. However, the no-observed-effect concentration has not been established. Therefore, the current study was conducted to determine whether there would be any effects on earthworms from feces from cattle treated with the commercial formulation of eprinomectin. Feces were collected rectally from grazing cattle on Day 0 before treatment and on Days 2, 4, 7 and 14 after treatment with EPRINEX (eprinomectin) Pour-On for Beef and Dairy Cattle (Merial Limited) at 0.5 mg eprinomectin per kg bodyweight. Assays of eprinomectin B1a (the major component of eprinomectin) were 0, 0.427, 0.152, 0.0512 and 0.00185 mg kg-1 wet weight of feces (equivalent to 0, 3.34, 1.19, 0.40 and 0.010 mg kg-1 on a dry weight basis, respectively). No significant differences (p>0.05) were observed at any day post-treatment in the survival or behavioral effects of any worms fed post-dose feces relative to the worms fed control feces. All post-dose comparisons of weight changes of living earthworms to the control group were not significantly different (p>0.05), indicating that treatment of cattle with EPRINEX (eprinomectin) Pour-On for Beef and Dairy Cattle did not affect feeding or weight gain of the earthworms. The LC50 value and the results of this study establish the wide margin of safety afforded to earthworms by eprinomectin under typical usage conditions.     

Routes of transmission and consequences of small ruminant lentiviruses (SRLVs) infection and eradication schemes

Small ruminant lentiviruses (SRLV = maedi-visna in sheep and caprine arthritis encephalitis in goats) are distributed throughout most countries of the world, particularly Europe. Laboratories from 16 European countries established collaborations within the framework of a COST (CO-operation in the field of Scientific and Technical Research) action sponsored by the European Union in order to (i) better organize their research programmes on SRLVs and (ii) to coordinate efforts to combat these two diseases. After five years, a consensus conference--the first one in the veterinary medicine field--concluded the work of this network of laboratories by reviewing the present position and discussing three important questions in the field of SRLVs: routes of transmission, consequences of infection and potential role of eradication programmes at either a European or local level, according to the situation in each country or region. This paper brings together existing information regarding these questions and identifies areas for future research.     

Inheritance of rol-genes from Agrobacterium rhizogenes through two generations in Kalancho?

Transformation of Kalancho? blossfeldiana ‘Molly’ using the naturally occurring bacterium Agrobacterium rhizogenes is a non-GMO strategy to breed compact plants. In the present study, crosses resembling a commercial breeding strategy were made to determine if the improved ornamental quality observed in the T1 generation was inherited to the offspring. F1 lines were produced by crossing the commercial Kalancho? cultivar ‘Sarah’ with a selected T1 line. Subsequently, F2 populations were produced by self-pollination of individual selected lines. The rol-genes were inherited to the progeny and the presence of rol-genes was confirmed in all F1 and many F2 plant lines exhibiting dwarfism. Screening of F1 and F2 plants showed that the rol-genes were inherited together. Besides decreased plant height, several F1 and F2 lines containing rol-genes exhibited changes in plant diameter, number of branches, flower diameter and time to first open flower and duration of flowering compared to the WT ‘Molly’. Furthermore, increased ethylene tolerance was observed in several lines containing rol-genes compared to the WT ‘Sarah’. Screening of three selected F2 populations derived from self-pollination of F1 lines containing rol-genes indicated a shift in distribution towards a lower mean plant height within the entire plant population compared to a control population of plants without rol-genes. Moreover, one of these F2 populations also exhibited earlier flowering compared to the control population. Compact potted plants and lines without delayed flowering and with improved ethylene tolerance were obtained and are valuable in commercial breeding programmes without using recombinant DNA technology.