Vitellogenin transcytosis in follicular cells of the honeybee <Emphasis Type="Italic">Apis mellifera</Emphasis> and the wasp <Emphasis Type="Italic">Polistes simillimus</Emphasis>

Vitellogenin receptor (VgR) is a low-density lipoprotein receptor responsible for the mediated endocytosis of vitellogenin (Vg) during egg formation in insects. The maturing oocyte is enveloped by a follicular epithelium, which has large intercellular spaces during Vg accumulation (patency). However, Vg has been reported in the cytoplasm of follicular cells, indicating that there may be a transcellular route for its transport. This study verified the presence of VgR in the follicular cells of the ovaries of the honeybee Apis mellifera and the wasp Polistes simillimus in order to evaluate if Vg is transported via transcytosis in these insects. Antibodies specific for vitellogenin receptor (anti-VgR), vitellogenin (anti-Vg), and clathrin (anti-Clt) were used for immunolocalization. The results showed the presence of VgR on the apical and basal plasma membranes of follicular cells of the vitellogenic follicles in both species, indicating that VgR may have been transported from the basal to the apical cell domain, followed by its release into the perivitelline space, evidenced by the presence of apical plasma membrane projections containing VgR. Co-localization proved that Vg bind to VgR and that the transport of this protein is mediated by clathrin. These data suggest that, in these social insects, Vg is transported via clathrin-mediated VgR transcytosis in follicular cells.     

Selection and validation of reference genes for quantitative gene expression analyses in various tissues and seeds at different developmental stages in <Emphasis Type="Italic">Bixa orellana</Emphasis> L.

Bixa orellana L., popularly known as annatto, produces several secondary metabolites of pharmaceutical and industrial interest, including bixin, whose molecular basis of biosynthesis remain to be determined. Gene expression analysis by quantitative real-time PCR (qPCR) is an important tool to advance such knowledge. However, correct interpretation of qPCR data requires the use of suitable reference genes in order to reduce experimental variations. In the present study, we have selected four different candidates for reference genes in B. orellana, coding for 40S ribosomal protein S9 (RPS9), histone H4 (H4), 60S ribosomal protein L38 (RPL38) and 18S ribosomal RNA (18SrRNA). Their expression stabilities in different tissues (e.g. flower buds, flowers, leaves and seeds at different developmental stages) were analyzed using five statistical tools (NormFinder, geNorm, BestKeeper, ΔCt method and RefFinder). The results indicated that RPL38 is the most stable gene in different tissues and stages of seed development and 18SrRNA is the most unstable among the analyzed genes. In order to validate the candidate reference genes, we have analyzed the relative expression of a target gene coding for carotenoid cleavage dioxygenase 1 (CCD1) using the stable RPL38 and the least stable gene, 18SrRNA, for normalization of the qPCR data. The results demonstrated significant differences in the interpretation of the CCD1 gene expression data, depending on the reference gene used, reinforcing the importance of the correct selection of reference genes for normalization.     

New potent P-glycoprotein modulators with the cucurbitane scaffold and their synergistic interaction with doxorubicin on resistant cancer cells

The novel cucurbitacins, balsaminagenin A and B (1–2) and balsaminoside A (3) and the know cucurbitacin karavelagenin C (4), together with five new mono or diacylated derivatives (5–9) of karavelagenin C were evaluated for multidrug resistance reversing activity on human MDR1 gene transfected mouse lymphoma cells. Compounds 2–6 exhibited a strong activity compared with that of the positive control, verapamil. Structure–activity relationships are discussed. Moreover, in the checkerboard model of combination chemotherapy, the interaction between doxorubicin and compounds 2–5 synergistically enhanced the effect of the anticancer drug. Compounds 1–4 were isolated from the aerial parts of Momordica balsamina L. The structures of the compounds were established on the basis of spectroscopic methods including 2D NMR experiments (COSY, HMQC, HMBC and NOESY).     

Scaling of species distribution models across spatial resolutions and extents along a biogeographic gradient. The case of the Iberian mole Talpa occidentalis

Scaling is a key process in modelling approaches since it allows for translating information from one scale to another. However, the success of this procedure may depend on ‘source’ and ‘target’ scales, but also on the biogeographic/ecological context of the study area. We aimed to quantify the performance and success of scaling species distribution model (SDM) predictions across spatial resolution and extent along a biogeographic gradient using the Iberian mole as study case. We ran separate MaxEnt models at two extents (national and regional) using independent datasets (species locations and environmental predictors) collected at 10 km and 50 m resolutions respectively. Model performance and success of scaling SDMs were quantified on the basis of accuracy measures and spatial predictions. Complementarily, we calculated marginality and tolerance as indicators of habitat availability and niche truncation along the biogeographic gradient. Model performance increased with resolution and extent, as well as from north to south (mainly for high resolution models). When regional models were validated at different scales, their performance reduced severely, particularly in the case of coarse resolution models (some of them performed worse than random). However, when the 10 km‐national model was downscaled within regions, it performed better (AUC_test: 0.82, 0.85 and 0.55 respectively for Galicia, Madrid and Granada) than models specifically calibrated within each region at 10 km (0.47, 0.65, 0.44). Indeed, it also had a better accuracy when projected at 50 m (0.77, 0.91, 0.79) than models fitted at that resolution (0.62, 0.83, 0.96) in two of the three cases. The success of scaling model predictions decreased along the biogeographic gradient, being these differences associated to niche truncation. Models representing non‐truncated niches were more successfully scaled across resolutions and extents (particularly in areas not offering all possible habitats for species), which has important implications for SDM applications.     

Life and death in aluminium-exposed cultures of rat lactotrophs studied by flow cytometry

Prolonged exposure to aluminium may impact health. Aluminium’s deleterious effects are mostly attributed to its selective accumulation in particular organs and cell types. Occupational exposure to aluminium is allied with a reduced level of serum prolactin, a stress peptide hormone mainly synthesised and secreted by the anterior pituitary lactotrophs. Our aim was to study the effect of aluminium on the viability of rat lactotrophs in primary suspension cultures where multicellular aggregates tend to form, comprising approximately two thirds of the total cell population as confirmed by confocal microscopy. Flow cytometric light scattering of calcein acetoxymethyl ester and ethidium homodimer-1 labelled cells was used to define subpopulations of live and dead cells in heterogeneous suspensions comprised of single cells and multicellular aggregates of distinct size. Concentration-dependent effects of AlCl₃ were observed on aggregate size and cell survival. After 24-h exposure to 3 mM AlCl₃, viability of single cells declined from 5% to 3%, while in multicellular aggregates, viability declined from 23% to 20%. The proportion of single cells increased from 30% to 42% within the same concentration range, while in large aggregates, the proportion remained approximately constant representing 35% of the cell suspension. In large aggregates, cell viability (75%) remained unaltered after exposure to AlCl₃ concentrations up to 300 μM, while in single cells, viability was halved at 30 μM. In conclusion, our finding indicates that prolonged exposure to aluminium may lead to significant loss of pituitary cells.     

Infections Caused by <Emphasis Type="Italic">Fusarium</Emphasis> Species in Pediatric Cancer Patients and Review of Published Literature

Fusarium species have emerged as responsible for a broad spectrum of infections, including superficial, locally invasive and disseminated ones, especially in the hospital environment. Since there are few reports of invasive and disseminated fusariosis in children, the aim of this study was to report four cases of nosocomial infection caused by this microorganism in children with cancer hospitalized in a public children’s hospital located in Brazil. Two of these patients were female and two were male. All patients presented febrile neutropenia, while three patients had acute lymphocytic leukemia and one patient had Wilms’ tumor as underlying disease. In two cases, fungi were isolated from blood and identified as Fusarium oxysporum species complex after phenotypic and genotypic studies, while in two other cases fungi were isolated from skin biopsies and identified as Fusarium solani species complex. One patient died 12 days after the onset of cutaneous lesions. All isolates, after susceptibility testing, presented high levels of minimum inhibitory concentration for itraconazole, voriconazole and amphotericin B. Considering the emergence of filamentous fungi as etiologic agents of nosocomial infections, health professionals should be aware of the problems these infections, especially fungal ones, may cause to debilitated patients.     

Life cycle assessment of a steam thermolysis process to recover carbon fibers from carbon fiber-reinforced polymer waste

Purpose

Carbon fibers have been widely used in composite materials, such as carbon fiber-reinforced polymer (CFRP). Therefore, a considerable amount of CFRP waste has been generated. Different recycling technologies have been proposed to treat the CFRP waste and recover carbon fibers for reuse in other applications. This study aims to perform a life cycle assessment (LCA) to evaluate the environmental impacts of recycling carbon fibers from CFRP waste by steam thermolysis, which is a recycling process developed in France.

Methods

The LCA is performed by comparing a scenario where the CFRP waste is recycled by steam-thermolysis with other where the CFRP waste is directly disposed in landfill and incineration. The functional unit set for this study is 2 kg of composite. The inventory analysis is established for the different phases of the two scenarios considered in the study, such as the manufacturing phase, the recycling phase, and the end-of-life phase. The input and output flows associated with each elementary process are standardized to the functional unit. The life cycle impact assessment (LCIA) is performed using the SimaPro software and the Ecoinvent 3 database by the implementation of the CML-IA baseline LCIA method and the ILCD 2011 midpoint LCIA method.

Results and discussion

Despite that the addition of recycling phase produces non-negligible environmental impacts, the impact assessment shows that, overall, the scenario with recycling is less impactful on the environment than the scenario without recycling. The recycling of CFRP waste reduces between 25 and 30% of the impacts and requires about 25% less energy. The two LCIA methods used, CML-IA baseline and ILCD 2011 midpoint, lead to similar results, allowing the verification of the robustness and reliability of the LCIA results.

Conclusions

The recycling of composite materials with recovery of carbon fibers brings evident advantages from an environmental point of view. Although this study presents some limitations, the LCA conducted allows the evaluation of potential environmental impacts of steam thermolysis recycling process in comparison with a scenario where the composites are directly sent to final disposal. The proposed approach can be scaled up to be used in other life cycle assessments, such as in industrial scales, and furthermore to compare the steam thermolysis to other recycling processes.

     

Response of Benthic Foraminifera to Organic Matter Quantity and Quality and Bioavailable Concentrations of Metals in Aveiro Lagoon (Portugal)

This work analyses the distribution of living benthic foraminiferal assemblages of surface sediments in different intertidal areas of Ria de Aveiro (Portugal), a polihaline and anthropized coastal lagoon. The relationships among foraminiferal assemblages in association with environmental parameters (temperature, salinity, Eh and pH), grain size, the quantity and quality of organic matter (enrichment in carbohydrates, proteins and lipids), pollution caused by metals, and mineralogical data are studied in an attempt to identify indicators of adaptability to environmental stress. In particular, concentrations of selected metals in the surficial sediment are investigated to assess environmental pollution levels that are further synthetically parameterised by the Pollution Load Index (PLI). The PLI variations allowed the identification of five main polluted areas. Concentrations of metals were also analysed in three extracted phases to evaluate their possible mobility, bioavailability and toxicity in the surficial sediment. Polluted sediment in the form of both organic matter and metals can be found in the most confined zones. Whereas enrichment in organic matter and related biopolymers causes an increase in foraminifera density, pollution by metals leads to a decline in foraminiferal abundance and diversity in those zones. The first situation may be justified by the existence of opportunistic species (with high reproduction rate) that can live in low oxic conditions. The second is explained by the sensitivity of some species to pressure caused by metals. The quality of the organic matter found in these places and the option of a different food source should also explain the tolerance of several species to pollution caused by metals, despite their low reproductive rate in the most polluted areas. In this study, species that are sensitive and tolerant to organic matter and metal enrichment are identified, as is the differential sensitivity/tolerance of some species to metals enrichment.     

Predator–prey relationships in a Mediterranean vertebrate system: Bonelli’s eagles,rabbits and partridges

How predators impact on prey population dynamics is still an unsolved issue for most wild predator–prey communities. When considering vertebrates, important concerns constrain a comprehensive understanding of the functioning of predator–prey relationships worldwide; e.g. studies simultaneously quantifying ‘functional’ and ‘numerical responses’ (i.e., the ‘total response’) are rare. The functional, the numerical, and the resulting total response (i.e., how the predator per capita intake, the population of predators and the total of prey eaten by the total predators vary with prey densities) are fundamental as they reveal the predator’s ability to regulate prey population dynamics. Here, we used a multi-spatio-temporal scale approach to simultaneously explore the functional and numerical responses of a territorial predator (Bonelli’s eagle Hieraaetus fasciatus) to its two main prey species (the rabbit Oryctolagus cuniculus and the red-legged partridge Alectoris rufa) during the breeding period in a Mediterranean system of south Spain. Bonelli’s eagle responded functionally, but not numerically, to rabbit/partridge density changes. Type II, non-regulatory, functional responses (typical of specialist predators) offered the best fitting models for both prey. In the absence of a numerical response, Bonelli’s eagle role as a regulating factor of rabbit and partridge populations seems to be weak in our study area. Simple (prey density-dependent) functional response models may well describe the short-term variation in a territorial predator’s consumption rate in complex ecosystems.