Measurement of platelet aggregation in ovine blood using a new impedance aggregometer

Background: Whole blood platelet aggregometry (impedance) is an important method to investigate platelet function disorders. Examination of hemostatic function in sheep is important with respect to their role as an animal model of human disease. Objective: The aim of this study was to evaluate and optimize selected methodological aspects (anticoagulant, agonist concentration) of impedance aggregometry in ovine blood using the new Multiplate 5.0 analyzer. Methods: Blood samples were collected in hirudin anticoagulant from 40 clinically healthy sheep. Samples from selected sheep were collected in citrate, with or without the addition of calcium chloride. The agonists adenosine diphosphate (ADP), collagen, ristocetin, arachidonic acid, and thrombin receptor‐activating peptide (TRAP) were added in several concentrations to induce aggregation. Results: Based on maximum aggregation values and internal precision, no significant difference was found between ADP concentrations of 3–10 μmol/L and collagen concentrations of 3–5 μg/mL (P>.05). The lowest interindividual variation of approximately 3–4‐fold was seen with 4 and 5 μmol/L ADP and 4 and 5 μg/mL collagen. Ristocetin, arachidonic acid, and TRAP did not induce significant aggregation at any concentration. Aggregation results were significantly lower when measured in citrate‐ vs hirudin‐anticoagulated blood, regardless of the presence of calcium chloride. Conclusions: Our results indicate that the multiplate impedance aggregometer is suitable for the measurement of platelet aggregation in sheep using optimal agonist concentrations of 4–5 μmol/L ADP and 4–5 μg/mL collagen. Hirudin‐anticoagulated blood is the preferred sample material.     

The effect of macro-invertebrates and plant litter of different quality on the release of N from litter to plant on alpine pastureland

The effect of ¹⁵N-labelled litter of different quality (Luzula sylvatica, a grass species, Vaccinium gaultheroides, a deciduous dwarf shrub, and Calluna vulgaris, a hardy dwarf shrub) and the presence of macro-decomposers (Lumbricus rubellus, Lumbricidae, and Enantiulus nanus, Diplopoda) on the growth of Dactylis glomerata (Poaceae), a grass species abundant on alpine pastureland, was investigated. After 4 months, the presence of soil animals significantly increased litter mass loss of L. sylvatica, V. gaultheroides and C. vulgaris by 27%, 11% and 40%, respectively. Soil animals generally reduced microbial biomass but significantly increased it in treatments where either L. sylvatica or C. vulgaris was present. The presence of soil animals significantly increased shoot and root biomass of D. glomerata by 48% and 64%, respectively. L. rubellus increased the transfer of ¹⁵N from the litter into plants. We conclude that macro-decomposers increased nutrient mobilization and plant uptake of nutrients mineralized from recalcitrant litter materials. Litter of L. sylvatica contributed most to the ¹⁵N uptake by D. glomerata, suggesting that litter quality is crucial for the cycling of nutrients on abandoned alpine pastureland.     

Small nuclear ribonucleoprotein remodeling during catalytic activation of the spliceosome

Major structural changes occur in the spliceosome during its activation just before catalyzing the splicing of pre-messenger RNAs (pre-mRNAs). Whereas changes in small nuclear RNA (snRNA) conformation are well documented, little is known about remodeling of small nuclear ribonucleoprotein (snRNP) structures during spliceosome activation. Here, human 45S activated spliceosomes and a previously unknown 35S U5 snRNP were isolated by immunoaffinity selection and were characterized by mass spectrometry. Comparison of their protein components with those of other snRNP and spliceosomal complexes revealed a major change in protein composition during spliceosome activation. Our data also suggest that the U5 snRNP is dramatically remodeled at this stage, with the Prp19 complex and other factors tightly associating, possibly in exchange for other U5 proteins, and suggest that after catalysis the remodeled U5 is eventually released from the postsplicing complex as a 35S snRNP particle.     

Microbial activity,organic C accumulation and <Superscript>13</Superscript>C abundance in soils under alley cropping systems after 9 years of recultivation of quaternary deposits

The impact of alley cropping on post-lignite mine soils developing from quaternary deposits after 9 years of recultivation was evaluated on the basis of microbial indicators, organic C and total N contents, and the isotope characteristics of soil C. Soils were sampled at the 0 to 3, 3 to 10, and 10 to 30 cm depths under black locust (Robinia pseudoacacia L.), poplar (Populus spp.), the transition zone and in the middle of alley under rye (Secale cereale). There was no significant effect of vegetation on microbial properties presumably, due to the high variability, whereas organic C and total N contents at the 0- to 3-cm layer were significantly higher under black locust and poplar than in the transition zone and rye field. Organic C total N contents, and basal respiration, microbial biomass, and microbial quotient decreased with soil depth. Soil organic C and total N contents were more than doubled after 9 years of recultivation, with annual C and N accretion rate of 162 g C _org m⁻² year⁻¹ and 6 g N _t m⁻² year⁻¹. Microbial properties indicated that the soils are in early stages of development; the C isotope characteristics confirmed that the sequestered C was predominantly from C3 plants of the alley cropping.     

多重聚合酶链反应检测环境中产气荚膜杆菌

产气荚膜杆菌根据产生4种主要毒素（α－β、ε、τ－毒素）可分为5种类型A－E型。在该研究中依据产气荚膜杆菌的5种毒素基因的序列，设计了5对PCR引物，建立了多重PCR方法，该方法可以快速区分5种类型的产气荚膜杆菌。并对68份环境样品（生活污水、生物肥料）进行检测和基因分型，共检出55株产气荚膜杆菌，其中A型产气荚膜杆菌50株，占总数的90％以上，B型、C型、D型只占5％，未检出E型产气荚膜杆菌，所有的分离株没有发现带有肠毒素。结果表明，目前环境中主要存在的菌型为A型菌，其他菌型的产气荚膜杆菌很少，而多数是非致病性产气荚膜杆菌。     

PCR-based monitoring of recent isolates of tobacco blue mold from Europe reveals the presence of two genetically distinct phenotypes differing in fungicide sensitivity

Bioassays testing the fungicide sensitivity against metalaxyl of Peronospora tabacina isolates collected in German tobacco fields in 2005 revealed the presence of two phenotypes, resistant and sensitive. DNA fingerprints using SSR and minisatellite primers allowed separation of the samples into two groups. The differences in amplification patterns coincided with the sensitive and resistant reaction of the isolates in metalaxyl bioassays. New primers were developed which allowed PCR-based detection of P. tabacina and differentiation of the metalaxyl-sensitive and the metalaxyl-resistant phenotype, respectively. Screening of recent blue mold isolates from Germany and other European countries for metalaxyl sensitivity with leaf disk bioassays coincided completely with the PCR-based identification of the two phenotypes. In Germany, exclusively resistant isolates were found between 2002 and 2004. These still dominate. Since 2005 the co-occurrence of sensitive isolates has been shown. No similar monitoring has been done in any other European country. However, we found the resistant phenotype reaction in a French isolate of 2004 and in two out of three Italian isolates in 2007. Two isolates from Poland and Bulgaria in 2007 were sensitive to metalaxyl. For all 58 isolates tested since 2002 the metalaxyl bioassay-based resistance type and the PCR-based tests for sensitive and resistant genotype coincided. Using genotype-specific primers for future population studies may help to trace the sources of the pathogen from which yearly propagation starts.     

The prevalence of antimicrobial susceptibility of veterinary pathogens isolated from cattle and pigs: national antibiotic resistance monitoring 2002/2003 of the BVL

The national antimicrobial resistance monitoring determines the current quantitative resistance level of life-stock pathogens, in order to permit the evaluation and surveillance of the distribution of resistances on a valid basis. During the examination period from June 2002 to July 2003, a total of 1849 pathogens was collected, following a representative German-wide pattern in collaboration with 29 laboratories. The selection of examined bacterial strains included different specimen causing respiratory diseases in fattening pigs (Pasteurella multocida, Bordetella bronchiseptica) and cattle (Pasteurella multocida, Mannheimia haemolytica), respectively, as well as strains causing mastitis in dairy cows (Staphylococcus spp., Streptococcus spp., E. coli). Determination of the in-vitro susceptibility (minimal inhibitory concentration) to at least 17 antimicrobial agents was performed centrally by the BVL using the microdilution broth method. The findings approximately match results of an analogous study conducted in 2001, and correspondingly revealed significantly lower resistance-values in comparison to data published for Germany so far. No correlation could be established between the incidence of resistance and differing stock densities. By means of the resistance monitoring data gathered by the BVL, the risk potential of antimicrobials applied in Germany can be reliably defined. This valuable information about the epidemiological situation of resistance in Germany can be helpful to veterinarians as a decision guidance when choosing appropriate means of therapy. Accordingly, the results stated by the BVL are apt to provide an important contribution to improving the safety of food-animal products. The experiences obtained from the monitoring also show that valid data about the antimicrobial susceptibility can only be raised on an interdisciplinary approach (federal agencies, county diagnostic laboratories, universities, industry). Currently, the BVL put into practice a study with an extended spectrum of bacterial species and indications.     

Eine objektschonende Methode zur Herstellung mikroskopischer Dauerquetschpräparate

Zusammenfassung Es wird eine Methode beschrieben, die es erlaubt, Chromosomen-Quetschpräparate ohne Ablösen des Deckglases haltbar zu machen. Diese Methode arbeitet materialschonend und ist deshalb für besonders empfindliche Objekte geeignet.

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Summary It is described a new method for making smear slides of chromosomes permanent without removing the cover slip from the slide. This method is working without any damage or loss of material and therefore suitable for sensible objects.

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