Validation of a second generation competitive enzyme immunoassay (CELISA) for the diagnosis of brucellosis in various species of domestic animals

A second generation competitive enzyme immunoassay (CELISA) for detection of bovine antibody to Brucella abortus was developed to eliminate reagent variables in the assay. This assay was different from earlier CELISA formats in that it used recombinant protein A and protein G immunoglobulin receptors (PAG), labelled with horseradish peroxidase, thus eliminating the requirement for polyclonal anti-mouse-enzyme conjugate for detection. This allowed standardization of the assay. The CELISA uses a monoclonal antibody specific for a common epitope of the O-polysaccharide (OPS) of smooth lipopolysaccharide (SLPS) derived from B. abortus S1119.3. This antibody did not react with PAG. This monoclonal antibody was used to compete with antibody in the bovine test serum to the smooth lipopolysaccharide (SLPS) antigen. Reaction of bovine antibody was then measured directly with the PAG enzyme conjugate. In this case, development of colour in the reaction indicated a positive reaction. The performance characteristics of the new CELISA, sensitivity, specificity and exclusion of antibody of B. abortus S19 vaccinated animals, were very similar to those of the classical CELISA and to the indirect enzyme immunoassay (IELISA) when using sera deemed positive by isolation of the bacterium, either from individual animals or from some animals on the premises. All sera were tested by the buffered antigen plate agglutination test (BPAT) and the complement fixation test (CFT). Only samples positive on both BPAT and CFT were considered as positive and only samples negative on both tests were used considered negative. Sufficient samples from cattle, swine, sheep and goats to validate the test were included based on OIE guidelines suggesting inclusion of a minimum of 300 positive and 1000 negative samples.     

In vivo and in vitro treatments against Sparicotyle chrysophrii (Monogenea: Microcotylidae) parasitizing the gills of gilthead sea bream (Sparus aurata L.)

The effect of in vitro and in vivo treatments against Sparicotyle chrysophrii, a microcotylid parasite of gilthead sea bream (Sparus aurata L.), was studied. In vitro chemical treatments were targeted to eggs, oncomiracidia and adults, and were tested both as disinfectants and therapeutics for infected animals. The compounds were: distilled water, formalin, limoseptic ®, hydrogen peroxide, chlorine, and praziquantel (PZQ). Larvae were sensitive to all the treatments, but adults were more resistant, as chlorine (60 ppm - 1 h), hydrogen peroxide (100 ppm - 30 min) and PZQ (50 ppm - 30 min) produced only 10% mortality. All adults were killed only with distilled water, limoseptic (0.1% - 5 min), formalin (300 ppm - 30 min), or hydrogen peroxide (200 ppm - 30 min). Eggs were the most resistant stage, as only 30 min in limoseptic (0.1% in distilled water) or in formalin (300 ppm) prevented hatching. PZQ was used in vivo either as a curative or preventive treatment. The highest dose tested (400 mg kg^− 1 BW; effective dose 116.3 mg kg^− 1 BW due to palatability problems leading to 45% reduction in host food intake) did not significantly decrease prevalence of infection when given for 6 consecutive days. A lower dose (200 mg kg^− 1 BW) (effective dose 158.1 mg kg^− 1 BW) was rejected to a lower degree and decreased the prevalence of infection from 90% to 40%. When a lower dose (40 mg kg^− 1 BW) was administered for longer periods (20 days), food intake was reduced slightly, but the infection did not decrease significantly. The oral intubation with PZQ (200 mg kg^− 1 BW) once a week for 4 weeks did not prevent the infection of fish by cohabitation. However, a significant reduction in the abundance of the parasite was registered. In view of the results, recommendations for fish treatment and disinfection of aquaculture facilities are discussed.     

The impacts of inorganic nitrogen application on mineralization of C-labelled maize and glucose, and on priming effect in saline alkaline soil

Organic matter dynamics and nutrient availability in saline alkaline soil of the former lake Texcoco will determine the success of a planned reforestation program. Uniformly labelled ¹⁴C-maize (MAI-treatment) and glucose (GLU-treatment) with or without 200 mg  kg⁻¹ soil (MAI-N treatment and GLU-N treatment, respectively) were added to soils with electrolytic conductivity (EC) 56 dS m⁻¹ (soil A) and 12 dS m⁻¹ (soil B) to investigate the importance of N availability on decomposition of organic material. Production of CO₂ and and inorganic N dynamics were monitored. The amount of ¹⁴C-glucose mineralized increased 1.8-times in the soil A, but had no effect in the soil B when 200 mg  kg⁻¹ soil was added, while the amount of ¹⁴C-maize mineralized increased 1.7 and 1.3-times when 200  kg⁻¹ soil was added in the soils A and B, respectively. Application of increased priming effect 3.7-times in the MAI-treatment of the soil A and 3.4-times in the GLU-treatment, while in the soil B the increase of priming effect was 4.1-times in the MAI-treatment and 3.7-times in the GLU-treatment. Of the 200 mg  kg⁻¹ added to both soils less than 10 mg NH₃-N kg⁻¹ was volatilized within one day, while 22 and 44 mg  kg⁻¹ soil was fixed on the soil matrix in the soil A and the soil B, respectively. Therefore more than 100 mg −N kg⁻¹ was immobilized into the microbial biomass within the first day. Concentration of nitrite increased sharply in all the treatments of soil A at the onset of the incubation followed by a decrease. A similar pattern was observed in the GLU-N and MAI-N treatments of the soil B, but not in the other treatments. A decrease in concentration of was observed in both soils followed by an increase in the MAI-N and GLU-N treatments of the soil B. It was found that application of had a stimulating effect on the decomposition of maize and glucose, and on the priming effect, while assimilatory reduction of resulted in an increase of in the soil A, and nitrification in the soil B.     

Application of multivariate analysis and artificial neural networks for the differentiation of red wines from the Canary Islands according to the island of origin

Eleven metals (K, Na, Ca, Mg, Fe, Cu, Zn, Mn, Sr, Li, and Rb) were determined in 83 red wines from the Canary Islands. The wines presented high concentrations of Na, and the concentrations of Cu and Zn were much lower than the maximum concentrations established by the International Office of Vine and Wine (OIV). Applying principal component analysis, the dimension space was reduced to five principal components that explain 76.4% of the total variance, and the wines tend to separate on the basis of the island of production. Linear discriminant analysis (LDA) allowed a reasonable classification of wines according to the island of production. When artificial neural networks (Kohonen self-organizing maps and back-propagation feed-forward as unsupervised and supervised techniques, respectively) were applied on the matrix of data constituted by the analyzed metals, the results improved in relation to those obtained by other multivariate methods observing a differentiation of wines according to island of production.     

Effect of growth conditions on wood density of Spanish Pinus nigra

The natural forests of Laricio Pine (Pinus nigra Subsp. salzmannii) cover in Spain an area of 353,000 ha, producing annually 310,000 m³ of roundwood. This species is considered as the fourth, most important Spanish conifer, being its timber frequently used in construction. The natural area of distribution of Laricio pine in Spain is officially divided into ten different regions of provenance, being only four of them considered as productive. The present study on the influence of growth conditions on extracted wood basic density (EBD) is carried out on 89 P. nigra trees, sampled from the two most productive Spanish regions of provenance (R.7 and R.8) in seven different sites. Cross-sectional disks were cut at 3 m (in 89 trees) and every 3 m height (in 18 trees) from the butt up to 15 m. For assessing the pattern of radial variation of the properties, 20×20×20 mm³ wood samples were taken radially from every cross-sectional disk. In every 20×20×20 mm³ wood sample, age and distance to the pith of the central annual ring was recorded, together with the average growth rate. Extractive content and EBD were measured in every sample. The results lead to the conclusion that there is not a significant relationship between EBD and provenance or growth rate. On the contrary, a significant relationship has been found between EBD and site quality or cambial age. The results also demonstrate that in a tree EBD tends to diminish from the base to the top and from the pith outwards. The main result of the present study is related with the observation that for the same cambial age, annual ring width has no significant influence on EBD, confirming that its utility as a predictor of the quality of timber is highly questionable, at least if the ring curvature is not considered.     

Termite resistance of beech wood treated with zinc oxide and zinc borate nanocompounds*

The field of wood protection is currently changing due to the restrictions imposed by the Biocidal Products Regulation. The need for development of new wood protection technologies is therefore growing. In this work, the resistance of European beech (Fagus sylvatica L.) wood impregnated with nano-dispersions of zinc oxide (nano-ZnO) and zinc borate (nano-ZnB) against the termite Reticulitermes grassei Clement was investigated. Three different concentrations (0.5%, 1%, 2%) of the nanocompounds were tested. The effects of water leaching were also investigated. A severe effect in terms of termite workers mortality was induced by both dispersions, even at the lowest concentration. In terms of termite feeding, nano-ZnO dispersions appeared to be more efficient than nano-ZnB as they induced significant improvement at concentrations as low as 0.5%. Nano-ZnB was applied at 1% or higher concentrations in order to impart similar changes. Nano-ZnB dispersions were efficient in terms of termite resistance for a concentration at least 1%. Further increasing concentration to 2% resulted in a respective increase of wood efficacy against termites. For both tested nanocompounds, water leaching did not result in any significant increase of termite feeding.     

Seaweed Blends as a Valuable Source of Polyunsaturated and Healthy Fats for Nutritional and Food Applications

Seaweeds are considered healthy and sustainable food. Although their consumption is modest in Western countries, the demand for seaweed in food markets is increasing in Europe. Each seaweed species has unique nutritional and functional features. The preparation of blends, obtained by mixing several seaweeds species, allows the obtaining of maximum benefits and ingredients with single characteristics. In this work, five seaweed blends, commercially available and produced under organic conditions in Europe, were characterized. The proximal composition included contents of ash (20.28–28.68% DW), proteins (17.79–26.61% DW), lipids (0.55–1.50% DW), and total carbohydrates (39.47–47.37% DW). Fatty acid profiles were determined by gas chromatography–mass spectrometry (GC–MS), allowing quantification of healthy fatty acids, namely n-3 and n-6 polyunsaturated fatty acids (PUFA), and calculation of lipid quality indices. Each blend showed a characteristic PUFA content in the lipid pool (35.77–49.43% of total fatty acids) and the content in essential and healthy n-3 PUFA is highlighted. The atherogenicity (0.54–0.72) and thrombogenicity (0.23–0.45) indices evidenced a good nutritional value of lipid fractions. As nutritional and environmentally attractive products, the consumption of the studied seaweed blends can contribute to a healthy lifestyle.     

Influence of High‐Temperature Drying on Structural and Textural Properties of Durum Wheat Pasta

Starch and protein are the main polymeric ingredients of pasta and they determine the structural and textural properties of cooked pasta. The present investigation sought better understanding of the impact of high‐temperature (HT) drying on the starch and the protein fraction, and their role in structure and texture of pasta. Durum wheat spaghetti was prepared in a pilot‐plant installation. The drying conditions were selected for the HT phase at 80 or 100°C applied at high, intermediate, or low product moisture content. Spaghetti dried at 55°C served as a reference sample. The color of dry pasta was measured and the changes in the starch and protein fractions were determined by protein solubility, light microscopy, confocal scanning laser microscopy (CSLM), cooking tests, and texture measurements. HT drying at 100°C and low product moisture promoted browning of pasta. At the molecular level, HT drying promoted protein denaturation. At the microscopic level, HT drying contributed to a better preservation of the protein network and reduced swelling of starch and disintegration of granules. At the macroscopic level, HT drying enhanced the firmness of cooked pasta and reduced surface stickiness. In general, the changes were more pronounced by increasing the drying temperature from 80 to 100°C and by shifting the HT phase from an early to a late stage of the drying process. The drying conditions are determinant for the phase morphology of protein and starch in cooked pasta which, in turn, govern the textural properties of pasta.     

Structural Properties of Starch in Bread and Bread Model Systems: Influence of an Antistaling α‐Amylase

The influence of an antistaling α‐amylase on bread crumb and on wheat starch gels was investigated taking into account different levels of structural hierarchy. Bread was prepared by a conventional baking procedure. Starch gels were produced by heating a concentrated starch dispersion in closed molds. Bread and starch gels were characterized by compression tests, light microscopy (LM), differential scanning calorimetry, and X‐ray measurements. The α‐amylase enhanced the initial firmness of starch gels and reduced the firming rate of bread and starch gels on aging. LM revealed that amylose and amylopectin phase‐separated within the starch granules and that freshly baked control bread and starch gels showed weak birefringence which became more intense during aging. Amylase‐containing bread and starch gels exhibited strong birefringence in the amylose rich region of the granules directly after baking which did not significantly increase during aging. The enzyme hindered the retrogradation of amylopectin as detected by differential scanning calorimetry, whereas X‐ray diffraction indicated that the enzyme induced low levels of starch crystallinity which did not change during aging. It is hypothesized that the antistaling effect of the amylase is based on the capacity to partially degrade amylopectin and, by this, to hinder its recrystallization. On the other hand, the enzyme slightly degrades amylose by an endo‐mechanism which, in turn, promotes the rapid formation of a partly crystalline amylose network in fresh bread and hinders amylose rearrangements during aging.