Does a pleiotropic gene explain deafness and blue irises in white cats?

The prevalence of deafness is high in cat populations in which the dominant white gene is segregating. The objective of this study was to investigate whether there is a gene that is responsible for deafness as well as for blue eyes and to establish a plausible mode of inheritance. For this purpose, data from an experimental colony with deaf cats were analyzed. The hearing status was determined by acoustically evoked brain stem responses (BAER). Complex segregation analyses were conducted to find out the most probable mode of inheritance using maximum likelihood procedures. The prevalence of deafness and partial hearing in the experimental colony was 67% and 29%, respectively. The results of the bivariate segregation analysis support the hypothesis of a pleiotropic major gene segregating for deafness and blue iris colour. The high heritability coefficients for both traits, 0.55 and 0.75 respectively, indicate that beside the major gene there is an important influence of polygenic effects.     

Phosphodiesterase isoenzymes in equine platelets and their influence on platelet adhesion

OBJECTIVE: To determine the phosphodiesterase (PDE) isoenzymes in equine platelets and evaluate their influence on platelet adhesion. SAMPLE POPULATION: Platelets obtained from healthy New Forest Pony geldings that ranged from 12 to 20 years of age (mean +/- SEM, 17.3 +/- 1.1 years). PROCEDURES: PDE isoenzyme activity in equine platelets was determined by use of a 2-step radioactive assay. Functional importance of PDE isoenzymes was established by use of selective inhibitors in a colorimetric adhesion assay. RESULTS: PDE1, PDE2, PDE3, and PDE5 and small amounts of PDE4 were found in equine platelets. Inhibition of PDE3 abolished platelet adhesion almost completely, whereas inhibition of PDE4 and PDE5 had little effect. CONCLUSIONS AND CLINICAL RELEVANCE: Function of equine platelets can be influenced by inhibition of PDE3. Selective PDE3 inhibitors may be clinically useful to regulate platelet function. They offer the advantage of increased potency with fewer adverse effects, compared with those for nonselective PDE inhibitors.     

Molecular subtype-specific clinical diagnosis of prion diseases

Sporadic Creutzfeldt-Jakob disease (sCJD) is a rare transmissible disease caused by accumulation of pathological prion protein (PrP(sc)) in the CNS. According to the codon 129 polymorphism (methionine or valine) and the prion protein type 1 or 2, a classification into distinct subtypes was established. Further analysis of these subtypes detected atypical clinical forms with longer disease duration or younger age at onset. The CJD subtype influences sensitivity of the technical investigations such as 14-3-3 in CSF, periodic sharp wave complexes in the EEG or hyperintense basal ganglia in MRI. A further characterization of these subtypes is important for reliable diagnosis and identification of rare disease variants. The aim is to establish specific patterns of test results and clinical findings. These improvements in diagnostics may be the reason for the apparent increase in sCJD incidence in Germany from 0.9 in 1994 to 1.6 in a million in 2005. Despite careful surveillance, no patient with variant CJD has been detected to date in Germany. Here we present the data of the CJD surveillance of the last 13 years. Additionally, the improvements in diagnostics and differential diagnosis are discussed.     

Reduced incidence of insect-bite hypersensitivity in Icelandic horses is associated with a down-regulation of interleukin-4 by interleukin-10 and transforming growth factor-beta1

Insect bite hypersensitivity (IBH) is an allergic dermatitis of horses caused by IgE-mediated reactions to bites of insects of the genus Culicoides. IBH does not occur in Iceland due to the absence of Culicoides. However, Icelandic horses exported to mainland Europe as adults (1st generation) have a > or =50% incidence of developing IBH. In contrast, their progeny (2nd generation) has a <10% incidence of IBH. Here we show that peripheral blood mononuclear cells (PBMC) from Icelandic horses born in mainland Europe and belonging either to the IBH or healthy subgroup produce less interleukin (IL)-4 after polyclonal or allergen-specific stimulation when compared with counterparts from horses born in Iceland. We examined a role of IL-10 and transforming growth factor (TGF)-beta1 in down-regulation of IL-4 in healthy 2nd generation Icelandic horses. Supernatants of PBMC from 2nd generation healthy horses down-regulated the proportion of IL-4-producing cells and IL-4 production in stimulated cultures of PBMC from 1st generation IBH. This inhibition was mimicked by a combination of IL-10 and TGF-beta1 but not by the single cytokines. Cultures of stimulated PBMC of healthy 2nd generation horses produced a low level of IL-4, but IL-4 production was increased by anti-equine IL-10 and anti-human TGF-beta1. This shows for the first time that in horses, IL-10 and TGF-beta1 combined regulate IL-4 production in vitro. It is suggested that in this naturally occurring IgE-mediated allergy, IL-10 and TGF-beta1 have a role in the down-regulation of IL-4-induced allergen-specific Th2 cells, thereby reducing the incidence of IBH.     

A histamine release assay to identify sensitization to Culicoides allergens in horses with skin hypersensitivity

Skin hypersensitivity is an allergic disease induced in horses by allergens of Culicoides midges. The condition is typically diagnosed by clinical signs and in some horses in combination with allergy testing such as intradermal skin testing or serological allergen-specific IgE determination. Here, we describe an alternative method for allergy testing: a histamine release assay (HRA) that combines the functional aspects of skin testing with the convenience of submitting a blood sample. The assay is based on the principle that crosslinking of allergen-specific IgE bound via high-affinity IgE receptors to the surfaces of mast cells and basophils induces the release of inflammatory mediators. One of these mediators is histamine. The histamine was then detected by a colorimetric enzyme-linked immunosorbent assay. The histamine assay was used to test 33 horses with skin hypersensitivity and 20 clinically healthy control animals for histamine release from their peripheral blood basophils after stimulation with Culicoides allergen extract or monoclonal anti-IgE antibody. An increased histamine release was observed in the horses with skin hypersensitivity compared to the control group after allergen-specific stimulation with Culicoides extract (p=0.023). In contrast, stimulation with anti-IgE induced similar amounts of released histamine in both groups (p=0.46). For further evaluation of the HRA, we prepared a receiver operating-characteristic (ROC) curve and performed a likelihood-ratio analysis for assay interpretation. Our results suggested that the assay is a valuable diagnostic tool to identify sensitization to Culicoides allergens in horses. Because some of the clinically healthy horses also showed sensitization to Culicoides extract, the assay cannot be used to distinguish allergic from non-allergic animals. The observation that sensitization is sometimes detectable in non-affected animals suggested that clinically healthy horses use immune mechanisms to control the reaction to Culicoides allergens that are different or absent in allergic horses.     

Characterization of Streptococcus equi subsp. ruminatorum isolated from spotted hyenas (Crocuta crocuta) and plains zebras (Equus burchelli), and identification of a M-like protein (SrM) encoding gene

Thirteen strains of Streptococcus equi subsp. ruminatorum from free-ranging spotted hyenas (Crocuta crocuta) and plains zebras (Equus burchelli) in Tanzania were characterized by biochemical and molecular-biological methods. Although the colony appearance of the S.e. ruminatorum wildlife strains differed from that of the S.e. ruminatorum type strain CECT 5772(T), all biochemical reactions of the wildlife strains were similar to those of the type strain. In addition, all wildlife strains produced hyaluronidase and were capable of hydrolysing arginine, three strains (23%) synthesized acetoin, but only eight strains (62%) produced acid from ribose. rep-PCR indicated that different clones of S.e. ruminatorum were distributed among the hyena and zebra populations in the study area. Identical rep-PCR patterns in hyena and zebra strains suggest that a direct transmission of S.e. ruminatorum between these species may occur. The presence of a M-like protein (SrM) gene was demonstrated in all S.e. ruminatorum strains including the type strain. Sequencing of the M-like protein gene revealed a hypervariable region within the deduced amino acid sequence. Most of the strains clustered with previously described strains based on the hypervariable region of the S.e. zooepidemicus SzP protein. Sequencing also demonstrated that identical SrM protein sequences were shared among S.e. ruminatorum strains from different host species.     

Molecular Modeling of the N-terminal Regions of High Molecular Weight Glutenin Subunits 7 and 5 in Relation to Intramolecular Disulfide Bond Formation

Analyses of cystine peptides derived from the high molecular weight glutenin subunits (HMW-GS) 5 and 7 indicate that, in spite of a distinct sequence homology between the two subunits in the N-terminal region, different disulfide linkages of cysteine residues are present in these regions. To investigate the structural basis for these experimental results, the conformational structures of the polypeptide chains corresponding to the N-terminal regions (first 50 amino acids) of the wheat HMW-GS 5 and 7 were modeled by computer methods. Secondary structures were predicted by the method of Rost and Sander (1993) and, to the extent appropriate, applied to the constructed polypeptide chains. The resulting structures were energy-minimized and subjected to simulated heating and dynamic equilibration. In the final structure of subunit 5, the first two cysteines were located in a region of continuous α-helix. If folding to the helical form occurs rapidly during biosynthesis as expected, the distance between the sulfhydryl groups of these two cysteines would be great enough (≈2.2 nm) to make intramolecular disulfide bond formation unlikely. Although a somewhat similar region of α-helix was predicted for the subunit 7, in some predictions the helix was interrupted between the first two cysteines, and this break was assigned either extended structure or arbitrarily modeled as an inverse γ-turn. In the final structure of subunit 7 with the assigned inverseγ-turn, after energy minimization, heating, and dynamics, the two cysteines approached one another closely (≈0.4 nm). Formation of an intramolecular disulfide bond appeared a likely possibility. This model is in accord with experimental evidence for this latter intramolecular bond (Köhler et al 1993). In agreement with the modeling, an equivalent intramolecular disulfide bond of subunit 5 has not been found and experimental evidence for a different arrangement is presented.     

Rice roots and CH4 oxidation: the activity of bacteria, their distribution and the microenvironment

Irrigated rice fields account for 10–30% of global methane emissions. Rice plants ventilate the soil and enlarge the oxic–anoxic interface by their root system, thus supplying the necessary O₂ to aerobic CH₄ oxidizing bacteria (MOB). Rice plants (Oryza sativa type japonica var. Roma) were grown in microcosms in a greenhouse. The roots were sandwiched between two blocks of flooded rice field soil separated by a nylon gauze bag. A root mat developed which mimicked the dense root texture in the upper layer of a natural rice field. Flux measurements under oxic and anoxic conditions showed that CH₄ was oxidized with a constant rate of 19% of the anoxically emitted CH₄, suggesting that CH₄ oxidation in the rhizosphere was at least sometimes limited by CH₄ availability. Washed rice roots could both produce and oxidize CH₄, depending upon incubation conditions. CH₄ production by washed rice roots accounted for at most 10% of the CH₄ emitted under anoxic conditions. Initial CH₄ oxidation rates of washed roots equaled oxidation rates calculated from the difference between oxic and anoxic fluxes in situ. Oxidation rates became twice as high after an induction period of 20 h, indicating a limitation by O₂ or CH₄ in situ. The micro-environmental conditions near to the root mat were measured using microelectrodes for O₂, redox potential and NH₄⁺ and diffusion probes for CH₄. Up to 42 μM O₂ was detected in the root mat and concentrations were >2.5 μM in 45% of all measurements. In the bulk soil, no O₂ was detected below 2 mm depth, but the root mat significantly increased the redox potential. Plant roots and associated bacteria decreased porewater CH₄ and NH₄⁺ concentrations. In the root mat, concentrations of dissolved CH₄ were below the detection limit of our probes (<5 μM). Cell numbers of MOB increased with time in the rhizosphere and in the rhizoplane. MOB and aerobic heterotrophic bacteria (AHB) each numbered from 10⁶ to 10⁸ cells g⁻¹ dry weight of soil or root biomass). Active MOB occurred near to a root mat similar to the dense root texture in the upper layer of rice fields. We speculate about O₂ or CH₄ limitation of MOB.