Assessment of s-Triazine Catabolic Potential in Soil Bacterial Isolates Applying atz Genes as Functional Biomarkers

Fluorescence in situ hybridization (FISH) technique and qPCR analyses, targeting atz genes, were applied to detect the presence of simazine-degrading bacteria in an agricultural soil with a history of herbicide application. atzB-targeted bacteria detected by FISH represented 5% of total soil bacteria with potential capability to metabolize the herbicide. The soil natural attenuation capacity was confirmed in soil microcosms by measuring simazine degradation. Moreover, four bacterial strains were isolated from the soil and identified as Acinetobacter lwoffii, Pseudomonas putida, Rhizobium sp. and Pseudomonas sp. The isolates were able to grow using different s-triazine compounds and related metabolites as the sole carbon source. Growth parameters in presence of simazine were calculated using the Gompertz model. Rhizobium sp. showed the highest simazine degradation (71.2%) and mineralization (38.7%) rates, whereas the lowest values were found to A. lwoffii??50.4% of degradation and 22.4% of mineralization. Results from qPCR analyses of atzA, atzB and atzC genes revealed their presence in Rhizobium sp. and A. lwoffii, being atzB and atzC the most abundant functional genes. Rhizobium sp. showed a higher amount of the three biomarkers compared to A. lwoffii: the atzA, atzB and atzC gene copy number per microlitre were, respectively, 10¹, 10² and 10³-fold higher in the former. Therefore the proposed molecular approaches based on the use of atz genes as biomarkers can be considered as useful tools to evaluate the presence and potential capability of degrading-s-triazines soil microorganisms.     

Effect of paratuberculosis on the diagnosis of bovine tuberculosis in a cattle herd with a mixed infection using interferon-gamma detection assay

Interferon-gamma (IFN-gamma) detection assay is being applied as an ancillary test to tuberculin tests in the diagnosis of bovine tuberculosis to detect the maximum number of infected animals. Among possible factors influencing the performance of tuberculosis-diagnostic tests, paratuberculosis, a widespread disease in Spain and other European countries, has been pointed out as a cause of false positive reactions. Still, its effect on the sensitivity of these tests in cattle has yet to be fully characterized. The impact of paratuberculosis in the apparent sensitivity of IFN-gamma assay was studied in a bullfighting cattle herd with a mixed tuberculosis-paratuberculosis infection, using culture of Mycobacterium bovis and Mycobacterium avium paratuberculosis as the gold standard to determine the infection status of every animal. A total of 218 animals were slaughtered and sampled for bacteriology after blood sampling. IFN-gamma assay showed a lower apparent sensitivity in animals with a mixed infection (50%) compared to all animals suffering tuberculosis (78.3%). This finding indicates that the presence of paratuberculosis in tuberculosis-infected herds could imply a serious impairment in the sensitivity of IFN-gamma detection test.     

Defining output-based standards to achieve and maintain tuberculosis freedom in farmed deer, with reference to member states of the European Union

Within the European Union (EU), detailed legislation has been developed for cattle, but not deer, to minimise disease risks associated with trade in animals and animal products. This legislation is expressed as input-based standards, providing a detailed outline of the activity required (for example, testing of animals and application of defined control measures), on the expectation that an adequate output (for example, confidence in freedom) will be achieved. Input-based standards are at odds with the increasing shift towards output-based standards, particularly in OIE rules governing international trade. In this paper, we define output-based standards to achieve and maintain freedom from tuberculosis (TB) in farmed deer, with reference to EU member states. After considering the probability of freedom achieved for cattle under existing EU legislation, we defined a ‘free farmed deer holding’ as one with a probability of freedom from infection of at least 99%. We then developed an epidemiological model of TB surveillance systems for deer holdings, incorporating different surveillance strategies, including combinations of diagnostic tests, and a variety of different scenarios relating to the potential for introduction of infection. A range of surveillance strategies were identified to achieve and maintain a free farmed deer holding, and worked examples are presented. The surveillance system sensitivity for varying combinations of screening and confirmatory tests in live animals, animals at slaughter and on-farm deaths is also presented. Using a single test at a single point in time, none of the TB tests routinely used in farmed deer is able to achieve an acceptable probability of TB freedom. If repeat testing were undertaken, an acceptable probability of TB freedom could be achieved, with differing combinations of the surveillance system sensitivity, frequency of testing and risk of introduction. The probability of introduction of infection through the importation of infected deer was influenced by the use of a pre-movement test (assumed 90% test sensitivity and negative test results), the TB prevalence in the source herd and the number of animals imported. A surveillance system sensitivity of at least 81% was achieved with different combinations of annual live animal surveillance and surveillance of animals at slaughter or on-farm deaths. This methodology has broad applicability and could also be extended to other diseases in both deer and other species with relevance to trade in animals and animal products.     

Characterization of exopolysaccharides produced by Bifidobacterium longum NB667 and its cholate-resistant derivative strain IPLA B667dCo

Bifidobacteria are natural members of the human intestinal microbiota and some strains are being used as probiotics. Adaptation to bile can allow them to increase survival in gastrointestinal conditions, thus improving their viability. Bifidobacterium longum NB667 and the cholate-resistant strain B. longum IPLA B667dCo produced exopolysaccharides (EPS) that were partially characterized. Analysis by size exclusion chromatography-multiangle laser light scattering indicated that the EPS crude fractions of both strains contained two polymer peaks of different molar mass. On the basis of chromatographic techniques both peaks appeared to be heteropolysaccharides. The smaller peak was mainly composed of glucose, galactose and rhamnose whose molar ratios and linkage types showed slight variations between the EPS fractions of both strains. The bigger peak consisted of glucose and galactose; the monosaccharide composition was identical in the EPS fractions of the two microorganisms, but their infrared spectra presented some differences regarding compounds other than carbohydrates that seem to be associated to the polymer. Differences in the composition of EPS fractions did not affect the capability of crude EPS from B. longum to be fermented by the human intestinal microbiota in fecal batch cultures.     

Ultrasound-Assisted Water Extraction of Mastocarpus stellatus Carrageenan with Adequate Mechanical and Antiproliferative Properties

Ultrasound-assisted water extraction was optimized to recover gelling biopolymers and antioxidant compounds from Mastocarpus stellatus. A set of experiments following a Box–Behnken design was proposed to study the influence of extraction time, solid liquid ratio, and ultrasound amplitude on the yield, sulfate content, and thermo-rheological properties (viscoelasticity and gelling temperature) of the carrageenan fraction, as well as the composition (protein and phenolic content) and antiradical capacity of the soluble extracts. Operating at 80 °C and 80 kHz, the models predicted a compromise optimum extraction conditions at ~35 min, solid liquid ratio of ~2 g/100 g, and ultrasound amplitude of ~79%. Under these conditions, 40.3% carrageenan yield was attained and this product presented 46% sulfate and good mechanical properties, a viscoelastic modulus of 741.4 Pa, with the lowest gelling temperatures of 39.4 °C. The carrageenans also exhibited promising antiproliferative properties on selected human cancer cellular lines, A-549, A-2780, HeLa 229, and HT-29 with EC₅₀ under 51.9 μg/mL. The dried soluble extract contained 20.4 mg protein/g, 11.3 mg gallic acid eq/g, and the antiradical potency was equivalent to 59 mg Trolox/g.     

Collection and morphological characterization of Panicum coloratum L. in Argentina

Panicum coloratum (Poaceae) is a perennial grass native to South Africa, distributed all over the world, but poorly adopted by ranchers in Argentina. Adapted to a wide range of soil and climatic conditions, it constitutes a good forage resource for zones with edaphic and climatic restrictions, where cattle rising activities are here displaced. The high polymorphism in the species has made it difficult to delineate sub specific taxonomic categories. It has been classified in botanical varieties, types and/or cultivars. In order to document the morphological variability in P. coloratum and to investigate the differences between var. makarikariense and var. coloratum, a collection was gathered in INTA Rafaela composed by fourteen accessions from different provinces of Argentina: six accessions of var. makarikariense and eight from var. coloratum. The evaluation of 152 individuals gives evidence of a wide morphological variation in the collection in traits related to forage and caryopses production; seven out of nine traits showed significant differences between var. makarikariense and var. coloratum. Based on the study of local type specimens, a morphological key to distinguish between P. coloratum varieties is provided. These results report new information about the diversity in P. coloratum in Argentina and provide valuable insights for future use of the collection as the base material of a breeding program.     

Rheological characterization of refrigerated and frozen non-fermented gluten-free dough: Effect of hydrocolloids and lipid phase

This work intended to study the rheological and textural behavior of gluten-free dough for “empanadas” and pie-crusts production. Traditionally, these products are made with non-fermented wheat-based dough. They are highly consumed in Latin America, but unsuitable for celiac people. Gluten matrix is a major determinant of the properties of dough, thus it must be replaced with other network forming components, such as hydrocolloids. Different hydocolloids were tested: hydroxypropylmethylcellulose (HPMC) and mixtures of xanthan/guar, and xathan/HPMC gums. Three different kinds of lipid phase were also studied; i.e. sunflower oil and low and high solid content margarine at two different levels (20–30%). Changes in rheological and textural properties during refrigerated storage were evaluated by dynamic oscillatory measurements and puncture and elongation tests on the unbaked dough. Best results were obtained using either of the hydrocolloid mixtures. Besides, the textural characteristics of cooked empanadas were also studied. Freezing before baking did not alter the quality of the crust in the products. ESEM micrographs revealed a continuous matrix formed by hydrocolloid entanglements. Starch granules were homogenously distributed in the dough and acted as inactive fillers. An untrained panel accepted the xanthan/HPMC dough with a 74/90 score and it was significantly preferred over a commercial gluten-free dough.     

Selection for ovulation rate in rabbits

An experiment of selection for ovulation rate was carried out. Animals were derived from a synthetic line first selected 12 generations for litter size, then 10 generations for uterine capacity. Selection was relaxed for 6 generations. Selection was based on the phenotypic value of ovulation rate with a selection pressure on does of 30%. Males were selected from litters of does with the highest ovulation rate. Males were selected within sire families in order to reduce inbreeding. Ovulation rate was measured in the second gestation by a laparoscopy, 12 days after mating. Each generation had about 80 females and 20 males. Results of three generations of selection were analyzed using Bayesian methods. Marginal posterior distributions of all unknowns were estimated by Gibbs sampling. Heritabilities of ovulation rate (OR), number of implanted embryos (IE), litter size (LS), embryo survival (ES), fetal survival (FS), and prenatal survival (PS) were 0.44, 0.32, 0.11, 0.26, 0.35, and 0.14, respectively. Genetic correlation between OR and LS was 0.56, indicating that selection for ovulation rate can augment litter size. Response to selection for OR was 1.80 ova. Correlated responses in IE and LS were 1.44 and 0.49, respectively. Selection for ovulation rate may be an alternative to improve litter size.