Fate of prions in soil: Degradation of recombinant prion in aqueous extracts from soil and casts of two earthworm species

Prions represent the active agent in transmissible spongiform encephalopathy (TSE) diseases and can remain infective to mammals even after prolonged periods in soil. The influence of mesofauna on prion dispersal and degradation in soil, however, remains unknown. In this study the effect of earthworms on the retention/dissemination of TSEs in soil was evaluated using a model recombinant prion protein (recPrP) and aqueous extracts from soil and fresh casts of two earthworm species, Lumbricus terrestris and Aporrectodea caliginosa. Our results showed that earthworm gut-derived enzymes did not enhance the degradation of recPrP in comparison to soil, even though non-prion related proteolytic activity was higher in fresh worm excrements than in soil samples. Complete degradation of recPrP occurred in the aqueous extracts from all samples within up to 6 days at +15 °C. The proteolytic enzymes responsible for degrading recPrP were inhibited by aprotinin and leupeptin and studies in pure cultures suggested these were most probably of soil microbial origin.     

Producing progeny from endangered birds of prey: treatment of urine-contaminated semen and a novel intramagnal insemination approach.

Wild raptors brought into an ex situ environment often have poor semen quality that is further compromised by urine contamination. Generally, it is believed that in birds, artificial insemination into the cloaca or caudal vagina of females requires large doses of high-quality spermatozoa to maximize fertility. In an effort to define and overcome some of the challenges associated with reproduction in wild raptors, the objectives of this study were to 1) evaluate the frequency, impact, and remediation of urine contamination in fresh ejaculates for the purpose of maintaining sperm motility and viability in vitro, and 2) develop a deep insemination method that allows low numbers of washed sperm to be placed directly into the magnum to increase the probability of producing fertilized eggs. The species evaluated include golden eagle (Aquila chrysoetos), imperial eagle (A. adalberti), Bonelli's eagle (Hiernaetus fasciatus), and peregrine falcon (Falco peregrinus). Semen samples were collected and pooled by species, and a minimum of 25 pooled ejaculates per species were evaluated for urine contamination, pH, sperm viability, and sperm motility; the samples were either unwashed or washed in neutral (pH 7.0) or alkaline (pH 8.0) modified Lake's diluent. Female golden eagles and peregrine falcons were inseminated via transjunctional, intramagnal insemination with washed spermatozoa from urine-contaminated samples. Urine contamination occurred in 36.8 +/- 12.8% (mean +/- SEM) golden eagle, 43.1 +/- 9.1% imperial eagle. 28.7 +/- 16.1% Bonelli's eagle, and 48.2 +/- 17.3% peregrine falcon ejaculates. The pH in urine-contaminated semen samples ranged from 6.48 +/- 0.3 to 6.86 +/- 0.2, and in noncontaminated samples it ranged from from 7.17 +/- 0.1 to 7.56 +/- 0.1. Sperm viability and motility were reduced (P < 0.05) in all species for unwashed vs. washed sperm after 30 min incubation at room temperature. Two peregrine falcon chicks and one golden eagle chick hatched after intramagnal insemination. This study demonstrates that urine contamination, a common and lethal acidifier in manually collected raptor ejaculates, can be circumvented by immediate, gentle seminal washing. Furthermore, these processed sperm, when deposited by transjunctional intramagnal insemination, can produce live young.     

Changes in Cd,Cu, Ni,Pb and Zn Fractionation and Liberation Due to Mussel Shell Amendment on a Mine Soil

Mining activities are related to relevant environmental pollution issues that should be controlled. We used sequential extractions to fractionate Cd, Cu, Ni, Pb and Zn retained on unamended or mussel shell‐amended mine soil samples, all of them treated with a mixture of the five heavy metals (total metal concentration of 1·57 mmol L⁻¹), after 1, 7 and 30 days of incubation. In addition, we used the stirred flow chamber technique to study the release of each of the five heavy metals from these different unamended and shell‐amended soil samples. The results indicate that the shell amendment caused a decrease in the most soluble fraction, while increasing the most recalcitrant (least mobile) fraction. With equivalent implications, the stirred flow chamber experiments showed that mussel shell amendment was associated to a decrease in heavy metal release and increased retention. The highest mussel shell dose and incubation time caused the most relevant changes in pH values and thus in metal retention, also indicating the importance of pH modifications in the mechanism of retention acting in the amended samples. In view of these results, the use of mussel shell amendment can be encouraged to increase heavy metal retention in acid mine soils, in order to minimise risks of environmental pollution. Copyright © 2016 John Wiley & Sons, Ltd.     

Intraspecific olive diversity assessed with AFLP

Amplified fragment length polymorphism (AFLP) was used to study diversity within and among Spanish olive varieties. A high degree of diversity was found among the varieties present in each growing region. Olive oil production and quality relies on appropriate cultivar selection as well as good orchard management. Production based only on a few superior cultivars would result in improved yield, oil quality, and production management. Amplified fragment length polymorphism were evaluated as a tool to identify the intraspecific and intravarietal diversity of olive. Amplified fragment length polymorphism analysis of 38 accessions belonging to 10 cultivars using six primer combinations produced 106 polymorphic bands. Results were analyzed for similarity among accessions via unweighted pair‐group means cluster analysis, resulting in 10 clusters corresponding to named variety designations. Similarity among varieties ranged from 0.60 to 0.72. Diversity within varieties was identified. Similarity within named varieties (accessions with the same varietal name) ranged from 0.75 to 0.96. Differences in several markers were found among 34 accessions. Intravarietal diversity was shown to exist within the Spanish olive varieties grown in the region surrounding Valencia.     

'Francolí', a late flowering almond cultivar re-classified as self-compatible

To verify the compatibility behaviour of the almond cultivar ‘Francolí’ and to clarify its S genotype a combination of pollination tests, stylar ribonuclease and allele specific PCR analysis was used. ‘Francolí’ was released from IRTA's breeding programme in 1994, having been putatively raised from the cross ‘Cristomorto’ (S₁S₂) × ‘Gabaix’ (S₁₀S₂₅). This cultivar was also reported to be self‐incompatible but revealing only one S band in the zymograms after S‐RNases analysis. ‘Francolí’ sets nuts after test crossing with two S₁S₂₅ cultivars, having a different genotype from that earlier reported. ‘Francolí’ was also observed to be self‐compatible after selfing flowers in the field and in the laboratory. ‘Francolí’ was re‐assigned the S₁S_f genotype after test crossing, stylar ribonuclease and PCR data analysis. After microsatellite analysis, the self‐compatible ‘Tuono’ (S₁S_f) cultivar is suggested as the male parent of ‘Francolí’ instead of the earlier reported ‘Gabaix’.     

Sunflower mutant containing high levels of palmitic acid in high oleic background

A new sunflower mutant, CAS-12, was obtained, which has both high palmitic (≈30%) and high oleic acid contents, and also a substantial amount of palmitoleic acid (≈7%). The mutant was selected after X-ray irradiation of dry seeds of the inbred line BSD-2-423, which had normal palmitic (≈3%) and high oleic (≈88%) acid levels. The increase of palmitic and palmitoleic acids occurred at the expense of the oleic acid content, which decreased to around 55% in respect to the original line. Linoleic acid content is always under 5%. Palmitic and palmitoleic acid levels were similar to those of the high palmitic mutant CAS-5 obtained in a previous programme from a low oleic line isogenic to BSD-2-423 using a similar mutagenic treatment. In that previous programme we also selected three high stearic acid mutants using chemical mutagenic treatment on the same sunflower line (RDF-1-532). We attempted to obtain mutants in other lines but were unsuccessful. The isolation of similar mutants in isogenic parental lines illustrates the importance of the genetic background in the development of specific mutants with an altered seed oil fatty acid composition. The oil of this mutant will increase the range of potential uses of sunflower oil. This revised version was published online in July 2006 with corrections to the Cover Date.     

Selection for seed yield in Andean intra-gene pool and Andean × Middle American inter-gene pool populations of common bean

Yields of large-seeded Andean (A) common bean (Phaseolus vulgaris L.) cultivars of Chile and Nueva Granada races are 40 to 60% lower compared to their Middle American (M) counterparts of small-seeded Mesoamerica and medium-seeded Durango races. Our objective was to use the concept of congruity backcrossing between Andean x Middle American inter-gene pool [AM 11833 = A 483 (A)///// A 686 (M) //// PVA 800A(A)/// ‘Carioca’ (M)// Carioca (M)/ G 19833 (A)] and between races within Andean gene pool [i.e., intra-gene pool Andean, AA 11834 = A483 (A)//// ‘Cardinal’(A) /// ‘Blanco Español’(A) // BlancoEspañol (A) / ‘Taylor’ (A)] to compare selection for seed yield improvement of large-seeded Andean beans. Seven hundred sixty seven F₂-derived F₃ (F_2:3) families were produced for each population. Visual appraisal for total plant performance, combined with seed yield from non-replicated plots was used for selection of 551 families in F_2:3, 182 families in F_2:4, and 91 families in F_2:5 in each population. Eight hundred twenty three F_5:6 lines were developed from the 91 F_2:5 families in each population. Visual selection, combined with seed yield in non-replicated plots was again used to select 294 lines in F_5:6 in each population. Similarly, 44 highest yielding F_5:7 lines were selected in AM 11833 and 39 F_5:7 lines in AA 11834. Thus, single plant selections were made in the F₂ and F₅, and plants within each plot were harvested in bulk in F₃, F₄, F₆, and F₇. Thirty nine F_5:8 lines from AA 11834 and 44 lines from AM 11833, parents, and checks were evaluated at Popayán and Quilichao, Colombia in 1998 and 1999. Selected lines in both populations, on average, out-yielded the mean of their large-seeded Andean parents. Mean yield of the lines selected from AM 11833 was 50% higher than AA 11834 lines. Twelve F_5:8 lines out-yielded the highest yielding Andean parents G 19833 and A 483 in AM 11833, whereas only one line yielded significantly higher (p < 0.05) than the highest yielding parent A 483 in AA 11834. However, none of selected lines out-yielded small-seeded Middle American parents used in AM 11833 (A 686 and ‘Carioca’). The mean 100 seed-weight of AA 11834 was 36 g compared to 28 G for AM 11833 F_5:8 lines. Selected lines had similar days to maturity as parents in AM 11833, and matured 3 d later in AA11834. Correlation coefficients between yield and 100 seed-weight were negative in both populations. Yield and days to maturity were positively correlated in AA 11834.     

A methodology for evaluating a location as a selection site for an international plant breeding program

Summary A method for comparing locations as selection sites based upon their abilities to predict yield and disease reaction over a target region is proposed. The probability of coincidence in selection for a site is defined as the probability for a line selected at the site to be selected at other sites within the region. The probability of divergence in rejection is defined as that associated with regional selection of a line given that is discarded by the site where selection is being conducted. The ideal selection site would maximize the probability of coincidence in selection and minimize the probability of divergence in rejection. The method is illustrated using a set of data from the rice yield nurseries of the International Rice Testing Program for Latin America planted under the rainfed conditions of Central América and México during the period 1978–1984. Five locations were compared for their predictive ability in selecting for the rainfed rice growing region, based on yield and disease reaction. Selection for yield was defined as performance superior to the best check in each location. Selection for disease reaction was based on an index derived from the Standard Evaluation System for Rice for diseases of regional importance. Locations varies 10–15 percent in their selection coincidence with the region for both yield and disease selection criteria applied independently.     

Suitability of Wastes from Olive-Oil Industry for Initial Reclamation of a Pb/Zn Mine Tailing

An incubation experiment was conducted to evaluate the ameliorating role of two organic amendments—olive-mill solid wastes and compost from olive-mill solid wastes- in the ecological reclamation of a lead/Zn-mine tailing collected in southern Spain. Four enzymatic activities (dehydrogenase, β-glucosidase, urease and phosphatase) and soluble and AB-DTPA extractable Pb and Zn and were periodically determined. High concentrations of Pb (5394 mg kg^?1) and Zn (9607 mg kg^?1), mainly in insoluble forms, were recorded in the lead/zinc-mine tailing, as well as very low biochemical activity. Application of the compost from olive-mill solid waste stimulated microbial activity and the biogeochemical cycles into the mine tailing because of the initially increased dehydrogenase, β-glucosidase and urease activities, which tended to decline or remained constant during the incubation period. By contrast, these enzyme activities were scarcely affected by the incorporation of the olive-mill solid wastes because this olive-organic amendment contains extractable polyphenols (36 g kg^?1), which inhibit these enzyme activities. Phosphatase activity was enhanced by the application of both olive-organic amendments, especially when the olive-mill solid waste was added to the mine tailing. Amounts of soluble and AB-DTPA-extractable Pb and Zn in the mine tailing were increased by the application of the olive-mill solid waste, and to a lesser degree, by the compost from this olive waste. This fact could restrict the use of these olive-organic amendements as useful materials in reclamation of lead/zinc-mine tailings. Nevertheless, the increases of available lead and zinc would represent an advantage where Pb/Zn-mine tailings are reclaimed by phytoextraction, effectively reducing the metal pollution in these mining wastes.