Ascaris suum: suppression of reaginic and hemagglutinating antibody responses in the mouse by crude extract and maintenance fluid.

Intraperitoneal administration in mice of crude extract (CE) or maintenance fluid (MF) of Ascaris suum in Freund's incomplete adjuvant (FICA) in doses of 200 and 2 (CE) and 4 μg (MF) on Days ?4, 0, and +4 relative to the day of the immunization with 10 μg of hen egg white lysozyme (HL) resulted in the suppression of anti-HL reaginic antibody responses at varying degrees depending on the dose and their time of administration. Hemagglutinating antibody responses were also affected but in a different manner. Treatment with CE on Day ?4 resulted in complete suppression of reaginic antibody responses and some degree of suppression of hemagglutinating antibody responses depending on the size of the CE dose. In mice pretreated with MF, transient suppression was found only for reaginic antibody responses. In mice receiving the treatment of CE on Day 0, 200 μg of CE caused complete suppression of reaginic antibody responses, while 2 μg was less effective. Hemagglutinating antibody responses were also suppressed in proportion to the dose. Simultaneous treatment with MF did not cause any suppression of either reaginic or hemagglutinating antibody responses. In mice treated with CE on Day +4, reaginic antibody responses were not markedly suppressed and hemagglutinating antibody responses were also not altered. In contrast, treatment with MF on Day +4 resulted in suppression of reaginic antibody responses during the whole course of the primary response, but had no effect on hemagglutinating antibody responses. When MF was administered 7 days after the priming, no suppressive effect on the antibody responses was demonstrated. On the other hand, if a lower dose (1 μg of HL) was used for the priming, the effect of MF treatment with Day +4 was more pronounced in the primary reaginic antibody response and the secondary response was also affected. A comparable suppression of hemagglutinating antibody responses also was observed.     

Effects of feeding transgenic corn with mCry1Ac or maroACC gene to laying hens for 12 weeks on growth,egg quality and organ health

The objective of the present study was to investigate the effect of feeding two transgenic corn lines containing the mCry1Ac gene from Bacillus thuringiensis strain (BT-799) and the maroACC gene from Agrobacterium tumefaciens strain (CC-2), respectively, on growth, egg quality and organ health indicators. Expression of the mCry1Ac gene confers resistance to Pyrausta nubilalis and the maroACC gene confers tolerance to herbicides. Healthy hens (n=96 placed in cages; 3 hens/cage) were randomly assigned to one of four corn–soybean meal dietary treatments (8 cages/treatment) formulated with the following corn: non-transgenic near-isoline control corn (control), BT-799 corn, CC-2 corn and commercially available non-transgenic reference corn (reference). The experiment was divided into three 4-week phases (week 1 to 4, week 5 to 8 and week 9 to 12), during which hens were fed mash diets. Performance (BW, feed intake and egg production) and egg quality were determined. Following slaughter at the end of 12 weeks of feeding (n=8/treatment), carcass yield and organ weights (heart, liver, spleen, lung, kidneys, stomach and ovary) were recorded; organs and intestines were sampled for histological analysis. Analysis of serum biochemistry parameters to assess the liver and kidney function were performed. No differences in BW, egg production and production efficiency were observed between hens consuming the control diet and hens consuming the BT-799 or CC-2 diet. Haugh unit measures and egg component weights were similar between the control and test groups. Carcass yield was not affected by the diet treatment. Similar organosomatic indices and serum parameters did not indicate the characteristics of organ dysfunction. All observed values of the BT-799 and CC-2 groups were within the calculated tolerance intervals. This research indicates that the performance, egg quality, organ health and carcass yield of laying hens fed diets containing the BT-799 or CC-2 corn line were similar to that of laying hens fed diets formulated with the non-transgenic near-isoline corn with comparable genetic backgrounds.     

Outdoor stocking density in free-range laying hens: radio-frequency identification of impacts on range use

The number and size of free-range laying hen (Gallus gallus domesticus) production systems are increasing within Australia in response to consumer demand for perceived improvement in hen welfare. However, variation in outdoor stocking density has generated consumer dissatisfaction leading to the development of a national information standard on free-range egg labelling by the Australian Consumer Affairs Ministers. The current Australian Model Code of Practice for Domestic Poultry states a guideline of 1500 hens/ha, but no maximum density is set. Radio-frequency identification (RFID) tracking technology was used to measure daily range usage by individual ISA Brown hens housed in six small flocks (150 hens/flock – 50% of hens tagged), each with access to one of three outdoor stocking density treatments (two replicates per treatment: 2000, 10 000, 20 000 hens/ha), from 22 to 26, 27 to 31 and 32 to 36 weeks of age. There was some variation in range usage across the sampling periods and by weeks 32 to 36 individual hens from the lowest stocking density on average used the range for longer each day (P<0.001), with fewer visits and longer maximum durations per visit (P<0.001). Individual hens within all stocking densities varied in the percentage of days they accessed the range with 2% of tagged hens in each treatment never venturing outdoors and a large proportion that accessed the range daily (2000 hens/ha: 80.5%; 10 000 hens/ha: 66.5%; 20 000 hens/ha: 71.4%). On average, 38% to 48% of hens were seen on the range simultaneously and used all available areas of all ranges. These results of experimental-sized flocks have implications for determining optimal outdoor stocking densities for commercial free-range laying hens but further research would be needed to determine the effects of increased range usage on hen welfare.     

Computational Design of a Protein-Based Enzyme Inhibitor

While there has been considerable progress in designing protein–protein interactions, the design of proteins that bind polar surfaces is an unmet challenge. We describe the computational design of a protein that binds the acidic active site of hen egg lysozyme and inhibits the enzyme. The design process starts with two polar amino acids that fit deep into the enzyme active site, identifies a protein scaffold that supports these residues and is complementary in shape to the lysozyme active-site region, and finally optimizes the surrounding contact surface for high-affinity binding. Following affinity maturation, a protein designed using this method bound lysozyme with low nanomolar affinity, and a combination of NMR studies, crystallography, and knockout mutagenesis confirmed the designed binding surface and orientation. Saturation mutagenesis with selection and deep sequencing demonstrated that specific designed interactions extending well beyond the centrally grafted polar residues are critical for high-affinity binding.     

Multiple Forms of Rice Seeds β-Amylases on Zymogram

Hen lysozyme modified with histamine (HML) and Japanese quail lysozyme (JQL) were treated with immobilized metal ion affinity chromatography to analyze the states of their imidazole groups. When Ni(II) was used as the metal ion immobilized, JQL was strongly retained in a Ni(II)-chelating Sepharose column, while hen lysozyme and HML were hardly retained in the same column. All of these lysozymes have a histidine imidazole group at the 15th position, while JQL has an additional histidine imidazole group at the 103rd position and HML has an additional imidazole group covalently attached to Asp101. Thus, I concluded that the imidazole group at the 103rd position of JQL is exposed to the solvent and recognized by the metal ion, but that the imidazole group attached to Asp101 in HML is localized to a hydrophobic region and not recognized by the metal ion.     

Untersuchungen über die Wirkung von Vitamin B12 und behandeltem Sojaschrot bei der Mast von Küken

To examine the effects of dietary conjugated linoleic acids (CLA) on lipid metabolism and antioxidant capacity in laying hens, Hy-Line Brown layers (n = 384, 52 weeks old) were randomly allocated to one of four dietary treatments. Each treatment had six replicates of 16 hens each. All birds were assigned to acorn-soybean meal-based diet containing a mixture of CLA at 0%, 1%, 2% or 4% for six weeks. With increasing dietary CLA, egg weight and feed intake decreased, and yolk colour was darkened. Feed efficiency was improved at 1% and 2% dietary CLA. Serum triglyceride concentration was significantly reduced by CLA in a dose dependent manner. A linear decrease in total cholesterol and low-density lipoprotein cholesterol, and an increase in high-density lipoprotein cholesterol levels were observed after CLA supplementation. With increasing dietary CLA, the deposition of two major isomers of CLA (c9, t11; t10, c12) in yolk lipids increased linearly, the proportion of saturated fatty acids increased and monounsaturated fatty acids decreased significantly. The proportion of polyunsaturated fatty acids was highest at 1% CLA. Compared to the control, CLA supplementation significantly increased the activities of superoxide dismutase and glutathione peroxidase, inhibited hydroxyl radicals and superoxide anion production, and decreased the malonaldehyde concentrations in both serum and liver. The results demonstrated that dietary CLA meliorated serum lipid profiles and enhanced the antioxidant capacity of laying hens.     

Multi‐spectroscopic and molecular dynamics simulations investigation of the binding mechanism of polybrominated diphenyl ethers to hen egg white lysozyme

Three PBDEs (BDE25, BDE47, and BDE154) were selected to investigate the interactions between PBDEs and hen egg white lysozyme (HEWL) by molecular modeling, fluorescence spectroscopy, and FT‐IR spectra. The docking results showed that hydrogen bonds were formed between BDE25 and residue TRP63 and between BDE47 and TRP63 with bond lengths of 2.178 Å and 2.146 Å, respectively. The molecular dynamics simulations indicated that van der Waals forces played a predominant role in the binding of three PBDEs to HEWL. The observed fluorescence quenching can be attributed to the formation of complexes between HEWL and PBDEs, and the quenching mechanism is a static quenching. According to Förster's non‐radiative energy transfer theory, the binding distances r were < 7 nm, indicating a high probability of energy transfer from HEWL to the three PBDEs. The synchronous fluorescence showed that the emission maximum wavelength of tryptophan (TRP) residues emerged a red‐shift. FT‐IR spectra indicated that BDE25, BDE47 and BDE154 induced the α‐helix percentage of HEWL decreased from 32.70% ± 1.64% to 28.27% ± 1.41%, 27.50% ± 1.38% and 29.78% ± 1.49%, respectively, whereas the percentage of random coil increased from 26.67% ± 1.33% to 27.60% ± 1.38%, 29.18% ± 1.46% and 30.59% ± 1.53%, respectively.     

Validation of the GROMOS force-field parameter set 45A3 against nuclear magnetic resonance data of hen egg lysozyme

The quality of molecular dynamics (MD) simulations of proteins depends critically on the biomolecular force field that is used. Such force fields are defined by force-field parameter sets, which are generally determined and improved through calibration of properties of small molecules against experimental or theoretical data. By application to large molecules such as proteins, a new force-field parameter set can be validated. We report two 3.5 ns molecular dynamics simulations of hen egg white lysozyme in water applying the widely used GROMOS force-field parameter set 43A1 and a new set 45A3. The two MD ensembles are evaluated against NMR spectroscopic data NOE atom–atom distance bounds, ³J_NH and ³J coupling constants, and ¹5N relaxation data. It is shown that the two sets reproduce structural properties about equally well. The 45A3 ensemble fulfills the atom–atom distance bounds derived from NMR spectroscopy slightly less well than the 43A1 ensemble, with most of the NOE distance violations in both ensembles involving residues located in loops or flexible regions of the protein. Convergence patterns are very similar in both simulations atom-positional root-mean-square differences (RMSD) with respect to the X-ray and NMR model structures and NOE inter-proton distances converge within 1.0–1.5 ns while backbone ³J_HN-coupling constants and ¹H– ¹5N order parameters take slightly longer, 1.0–2.0 ns. As expected, side-chain ³J-coupling constants and ¹H– ¹5N order parameters do not reach full convergence for all residues in the time period simulated. This is particularly noticeable for side chains which display rare structural transitions. When comparing each simulation trajectory with an older and a newer set of experimental NOE data on lysozyme, it is found that the newer, larger, set of experimental data agrees as well with each of the simulations. In other words, the experimental data converged towards the theoretical result.     

Solid-Phase Artificial Chaperone-Assisted Refolding using Insoluble β-Cyclodextrin–Acrylamide Copolymer Beads

Solid-phase refolding methods are advantageous since they facilitate both separation of solid additives from the refolded protein and recycling of the additives. -Cyclodextrin–acrylamide copolymer hydrogel beads were used as a matrix for detergents in solid-phase artificial chaperone-assisted refolding and improved the yield of lysozyme (up to 65%) and carbonic anhydrase B (up to 80%), compared with conventional solid host matrices.Revisions received 29 September 2004     

A QTL for osteoporosis detected in an F2 population derived from White Leghorn chicken lines divergently selected for bone index

Osteoporosis, resulting from progressive loss of structural bone during the period of egg-laying in hens, is associated with an increased susceptibility to bone breakage. To study the genetic basis of bone strength, an F(2) cross was produced from lines of hens that had been divergently selected for bone index from a commercial pedigreed White Leghorn population. Quantitative trait loci (QTL) affecting the bone index and component traits of the index (tibiotarsal and humeral strength and keel radiographic density) were mapped using phenotypic data from 372 F(2) individuals in 32 F(1) families. Genotypes for 136 microsatellite markers in 27 linkage groups covering approximately 80% of the genome were analysed for association with phenotypes using within-family regression analyses. There was one significant QTL on chromosome 1 for bone index and the component traits of tibiotarsal and humeral breaking strength. Additive effects for tibiotarsal breaking strength represented 34% of the trait standard deviation and 7.6% of the phenotypic variance of the trait. These QTL for bone quality in poultry are directly relevant to commercial populations.