Improving in sacco incubation technique to evaluate fresh forage for selecting fast‐degrading perennial ryegrass (Lolium perenne L.)

Grasses with fast fibre degradation are required by intensive pasture‐based animal production systems to maximize intakes and productivity. To select fast‐degrading elite cultivars, a repeatable, rapid and inexpensive screening method should be developed, so large numbers of samples can be evaluated. This study refined the experimental procedure for the in sacco incubation technique using fresh (not dried) perennial ryegrass (Lolium perenne L.). Pre‐ruminal incubation treatment and ratio of forage weight to the surface area of the in sacco bag have been tested to evaluate their effects on ryegrass degradation kinetic parameters in the bovine rumen. The timing of sampling and the number of sampling time points were also examined. Results indicated that warming the bags in water prior to incubation led to a faster dry‐matter (DM) degradation in the first 12 h. If ratio of forage to bag surface area was between 26 and 45 mg DM cm^?2, degradation parameters were not affected by bag fill. Sampling between 9 and 12 h was critical for determining degradation rate. From these results, an improved in sacco incubation procedure is recommended for screening of ryegrasses used for cultivar selection. The principles demonstrated here for ryegrass may be applicable to other forages, although the critical sampling times for measuring degradation rate are likely to differ.     

Effect of Zn-tolerant bacterial strains on growth and Zn accumulation in Orychophragmus violaceus

Several Zn-tolerant bacterial strains were isolated from heavy-metal contaminated sludge, and their effects on root elongation, mobility, and accumulation of Zn in Orychophragmus violaceus were studied. The isolated strains included Bacillus subtilis, B. cereus, Flavobacterium sp. and Pseudomonas aeruginosa which were capable of stimulating root elongation in O. violaceus seedlings either in the presence or absence of Zn. The four bacterial strains significantly increased the concentration of water-extractable Zn compared with axenic soil. In addition, the four Zn-tolerant bacteria significantly increased the shoot biomass and Zn accumulation in O. violaceus compared to non-inoculated plants. The bacterial strains displayed different capacities to enhance plant Zn accumulation. Flavobacterium sp. was identified as the best candidate for enhancing Zn accumulation in plants, increasing Zn accumulation up to 1.21- and 1.19-fold in shoots and roots, respectively, compared to non-inoculated plants. It was indicated that Zn-tolerant bacteria played an important role in influencing the availability of water-soluble Zn in soil and Zn accumulation by plants. This study provides insight into the development of plant–microbe systems for phytoremediation.     

Effect of Microbial Activity and Nitrogen Mineralization on Free-living Nitrogen Fixation in Permanent Grassland Soils

Free‐living nitrogen (N) fixation can be important for sustainable soil fertility, particularly in extensively managed soils with low abundance of leguminous plant species. However, the factors affecting free N₂‐fixation in situ are still poorly documented. We investigated the role of microbial active biomass activity, particularly substrate‐induced respiration (SIR) and net N mineralization, on the free‐living N₂ fixation in soils under a semi‐natural grassland ecosystem in France. Analysis of replicated bulk soil and rhizospheric soil samples obtained from sites experiencing contrasting grazing regimes revealed highly significant negative relationships (P < 0.01) between free‐living N₂‐fixation and SIR or N‐mineralization with a significant rhizosphere effect. The study has demonstrated that the activity of free‐living N₂‐fixers is more important in soils having low active microbial biomass and low N‐mineralization rates in these permanent grasslands.     

Hepatitis E virus: Animal reservoirs and zoonotic risk

Hepatitis E virus (HEV) is a small, non-enveloped, single-strand, positive-sense RNA virus of approximately 7.2 kb in size. HEV is classified in the family Hepeviridae consisting of four recognized major genotypes that infect humans and other animals. Genotypes 1 and 2 HEV are restricted to humans and often associated with large outbreaks and epidemics in developing countries with poor sanitation conditions, whereas genotypes 3 and 4 HEV infect humans, pigs and other animal species and are responsible for sporadic cases of hepatitis E in both developing and industrialized countries. The avian HEV associated with Hepatitis-Splenomegaly syndrome in chickens is genetically and antigenically related to mammalian HEV, and likely represents a new genus in the family. There exist three open reading frames in HEV genome: ORF1 encodes non-structural proteins, ORF2 encodes the capsid protein, and the ORF3 encodes a small phosphoprotein. ORF2 and ORF3 are translated from a single bicistronic mRNA, and overlap each other but neither overlaps ORF1. Due to the lack of an efficient cell culture system and a practical animal model for HEV, the mechanisms of HEV replication and pathogenesis are poorly understood. The recent identification and characterization of animal strains of HEV from pigs and chickens and the demonstrated ability of cross-species infection by these animal strains raise potential public health concerns for zoonotic HEV transmission. It has been shown that the genotypes 3 and 4 HEV strains from pigs can infect humans, and vice versa. Accumulating evidence indicated that hepatitis E is a zoonotic disease, and swine and perhaps other animal species are reservoirs for HEV. A vaccine against HEV is not yet available.     

Clonal growth and fragment regeneration of Rumex obtusifolius L.

Clonal growth and fragment regeneration of Rumex obtusifolius L. were analysed in two dif ferent studies. Clonal growth system was des cribed by a morphological study of underground structure of different-aged individuals, using maximal branching order in the stem system as an age estimator. Glasshouse experiments were conducted, testing the regenerative capacity of different below-ground parts and the estab lishment of above root-collar fragments planted at different depths under contrasting water regimes. Results showed the presence of a ‘phalanx’ type clonal growth system in grassland populations of Rumex. The main structure in volved in clonal growth proved to be the stem system; the region above the root collar was also the only part able to regrow after damage. Stem fragment regeneration occurred to depths of 15 cm, but was prevented in soils maintained at waterlogging and field capacity. The significance of these results in relation to nonchemical con trol of Rumex populations in grasslands is discussed.     

Expression of recombinant Toxoplasma gondii P24

The gene encoding Toxoplasma gondii P24 has been reported previously. To determine the function of P24 against immune systems in the near future, we prepared recombinant P24 antigens using Escherichia coli, insect cells infected with recombinant baculovirus and mammalian cells infected with recombinant vaccinia virus. The P24 antigens derived from E. coli, insect cells and mammalian cells were detected with mouse immune sera against P24 or T. gondii homogenates by Western blot analysis; these corresponded to the authentic P24 and secreted into the supernatants of the insect and mammalian cell cultures. These proteins were not effected by tunicamycin treatment in cultured cells, indicating that recombinant P24 did not contain N-linked sugars. Recombinant P24 was separated by two-dimensional electrophoresis and analyzed by Western blotting. From these results, P24 was acidic protein and had identical isoelectric point with the authentic P24.     

Use of a von Frey device for evaluation of pharmacokinetics and pharmacodynamics of morphine after intravenous administration as an infusion or multiple doses in dogs

OBJECTIVE: To evaluate the pharmacokinetics and pharmacodynamics of morphine after IV administration as an infusion or multiple doses in dogs by use of a von Frey (vF) device. ANIMALS: 6 dogs. PROCEDURE: In the first 2 crossover experiments of a 3-way crossover study, morphine or saline (0.9%) solution was administered via IV infusion. Loading doses and infusion rates were administered to attain targeted plasma concentrations of 10, 20, 30, and 40 ng/mL. In the third experiment, morphine (0.5 mg/kg) was administered IV every 2 hours for 3 doses. The vF thresholds were measured hourly for 8 hours. Plasma concentrations of morphine were measured by high-pressure liquid chromatography. RESULTS: No significant changes in vF thresholds were observed during infusion of saline solution. The vF thresholds were significantly increased from 5 to 8 hours during the infusion phase, corresponding to targeted morphine plasma concentrations > 30 ng/mL and infusion rates > or = 0.15 +/- 0.02 mg/kg/h.The maximal effect (EMAX) was 78 +/- 11% (percentage change from baseline), and the effective concentration to attain a 50% maximal response (EC50) was 29.5 +/- 5.4 ng/mL. The vF thresholds were significantly increased from 1 to 7 hours during the multiple-dose phase; the EC50 and EMAX were 23.9 +/- 4.7 ng/mL and 173 +/- 58%, respectively. No significant differences in half-life, volume of distribution, or clearance between the first and last dose of morphine were detected. CONCLUSIONS AND CLINICAL RELEVANCE: Morphine administered via IV infusion (0.15 +/- 0.02 mg/kg/h) and multiple doses (0.5 mg/kg, IV, every 2 hours for 3 doses) maintained significant antinociception in dogs.