Cell wall enzymatic lysis of the yeast form of Pullularia pullulans and wall regeneration by protoplasts

During the process of degradation of the cell wall of the yeast form of Pullularia pullulans by the lytic system of Micromonospora chalcea samples were withdrawn at different times and observed under phase contrast and electron microscope. The progressive lysis of the walls reveals a fibrillar component inside the apparently amorphous wall. Freeze etched preparations of cells during the formation and regeneration of protoplasts show that the cellular membrane is split and this method allows the smooth external face of the membrane and other internal face covered by particles to be seen. The fact that the smooth face of the membrane is only visible during the preparation or the regeneration of protoplasts and very rarely when intact cells are fractured, suggests a strong adherence between cell wall and this external layer of the membrane. During the regeneration which takes place as in most of the yeasts and moulds, a special study of the extension of the cell wall is made and a possible mechanism for this extension of the regenerated cell wall is proposed.     

High survival rate of a critically endangered species, the Azores Bullfinch Pyrrhula murina, as a contribution to population recovery

This paper reports analyses of a capture–mark–recapture (CMR) dataset of 149 Azores Bullfinches ringed on S?o Miguel island (Azores) between 2005 and 2007, and recaptured–resighted on a monthly basis over a 4-year period (2005–2008) throughout their breeding range. We examined the effect of time, age (adults vs. juveniles), gender (adult males and females), and environmental covariates (temperature, rainfall, NAO index) on survival probabilities. The modelling found a high and constant monthly survival probability (mean ± SE) estimated at 0.96 ± 0.01, similar between both adults and juveniles and independent of environmental conditions and gender. These findings agree with expectations from island-based life-history theory where relatively mild conditions and lack of predators should favour high survival rates to compensate for the low reproductive output. The annual survival rate was estimated at 0.62, which was also consistent with this pattern when compared with survival estimates of mainland bullfinch and passerine species on other subtropical islands obtained in similar CMR studies. Based on a canonical estimator, the size of the studied population (mean ± SE) was estimated at 1608 ± 326 individuals. Given that the population size was only around 120–400 individuals in the early 1990s, we suggest that the high survival probabilities currently applying to this critically endangered species may have substantially contributed to the recent recovery of this population. Future research studies on the species’ demography should continue to monitor survival in order to measure the effect of management interventions currently taking place within the range of the Azores Bullfinch, including the restoration of the biodiversity rich laurel forest, but also focusing on nest success, which is important for understanding population dynamics.     

The Crown Pearl: a draft genome assembly of the European freshwater pearl mussel Margaritifera margaritifera (Linnaeus, 1758)

Since historical times, the inherent human fascination with pearls turned the freshwater pearl mussel Margaritifera margaritifera (Linnaeus, 1758) into a highly valuable cultural and economic resource. Although pearl harvesting in M. margaritifera is nowadays residual, other human threats have aggravated the species conservation status, especially in Europe. This mussel presents a myriad of rare biological features, e.g. high longevity coupled with low senescence and Doubly Uniparental Inheritance of mitochondrial DNA, for which the underlying molecular mechanisms are poorly known. Here, the first draft genome assembly of M. margaritifera was produced using a combination of Illumina Paired-end and Mate-pair approaches. The genome assembly was 2.4 Gb long, possessing 105,185 scaffolds and a scaffold N50 length of 288,726 bp. The ab initio gene prediction allowed the identification of 35,119 protein-coding genes. This genome represents an essential resource for studying this species’ unique biological and evolutionary features and ultimately will help to develop new tools to promote its conservation.     

Two similar enhanced root-colonizing Pseudomonas strains differ largely in their colonization strategies of avocado roots and Rosellinia necatrix hyphae

Pseudomonas alcaligenes AVO73 and Pseudomonas pseudoalcaligenes AVO110 were selected previously as efficient avocado root tip colonizers, displaying in vitro antagonism towards Rosellinia necatrix, causal agent of avocado white root rot. Despite the higher number of antagonistic properties shown in vitro by AVO73, only AVO110 demonstrated significant protection against avocado white root rot. As both strains are enhanced root colonizers, and as colonization is crucial for the most likely biocontrol mechanisms used by these strains, namely production of non-antibiotic antifungal compounds and competition for nutrients and niches, we decided to compare the interactions of the bacterial strains with avocado roots as well as with R. necatrix hyphae. The results indicate that strain AVO110 is superior in biocontrol trait swimming motility and establishes on the root tip of avocado plants faster than AVO73. Visualization studies, using Gfp-labelled derivatives of these strains, showed that AVO110, in contrast to AVO73, colonizes intercellular crevices between neighbouring plant root epidermal cells, a microhabitat of enhanced exudation. Moreover, AVO110, but not AVO73, also colonizes root wounds, described to be preferential penetration sites for R. necatrix infection. This result strongly suggests that AVO110 meets, and can attack, the pathogen on the root. Finally, when co-inoculated with the pathogen, AVO110 utilizes hyphal exudates more efficiently for proliferation than AVO73 does, and colonizes the hyphae more abundantly than AVO73. We conclude that the differences between the strains in colonization levels and strategies are likely to contribute to, and even can explain, the difference in disease-controlling abilities between the strains. This is the first report that shows that two similar bacterial strains, selected by their ability to colonize avocado root, use strongly different root colonization strategies and suggests that in addition to the total bacterial root colonization level, the sites occupied on the root are important for biocontrol.     

Mapping contacts between regulatory domains of skeletal muscle TnC and TnI by analyses of single-chain chimeras

The troponin (Tn) complex is formed by TnC, TnI and TnT and is responsible for the calcium-dependent inhibition of muscle contraction. TnC and TnI interact in an antiparallel fashion in which the N domain of TnC binds in a calcium-dependent manner to the C domain of TnI, releasing the inhibitory effect of the latter on the actomyosin interaction. While the crystal structure of the core cardiac muscle troponin complex has been determined, very little high resolution information is available regarding the skeletal muscle TnI-TnC complex. With the aim of obtaining structural information regarding specific contacts between skeletal muscle TnC and TnI regulatory domains, we have constructed two recombinant chimeric proteins composed of the residues 1-91 of TnC linked to residues 98-182 or 98-147 of TnI. The polypeptides were capable of binding to the thin filament in a calcium-dependent manner and to regulate the ATPase reaction of actomyosin. Small angle X-ray scattering results showed that these chimeras fold into compact structures in which the inhibitory plus the C domain of TnI, with the exception of residues 148-182, were in close contact with the N-terminal domain of TnC. CD and fluorescence analysis were consistent with the view that the last residues of TnI (148-182) are not well folded in the complex. MS analysis of fragments produced by limited trypsinolysis showed that the whole TnC N domain was resistant to proteolysis, both in the presence and in the absence of calcium. On the other hand the TnI inhibitory and C-terminal domains were completely digested by trypsin in the absence of calcium while the addition of calcium results in the protection of only residues 114-137.     

Methods for Screening Cloud Point Temperatures

A novel and simple method for the measurement of cloud point temperatures of solutions is presented. Cloud point determination, which is currently used to establish the phase diagrams of protein solutions, is indicative of proteins interactions and constitutes a useful tool for food products engineering. We describe a novel experimental setup that allows screening of a large number of physical-chemical conditions in one measurement and the determination of cloud point temperatures both above and below ambient temperature. We use a simple method to avoid solvent evaporation and condensation, so that the set-up can be used for solutions prepared with a volatile solvent. We present the operating parameter range and the precision of the measurement. The optical properties of the system are calibrated with solutions of known transmittance, and the determination of cloud point temperatures is validated on a standard non-ionic surfactant solution. Finally, we demonstrate the efficiency of the method by determining the phase diagram of a wheat protein extract, soluble in a water/ethanol mixture. Complemented with differential scanning calorimetry measurements, the liquid-liquid phase transition can be determined up to a protein concentration of 250 g/L, a range inaccessible with conventional methods for this protein extract.     

Trk2 transporter is a relevant player in K<Superscript>+</Superscript> supply and plasma-membrane potential control in <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis>

In Saccharomyces cerevisiae, TRK1 and TRK2 genes encode partially redundant K⁺ transporters. Direct involvement in K⁺ uptake has been shown for Trk1p since cells growing under limiting environmental K⁺ concentrations demand its presence. The biological role of Trk2p is less understood. In our experiments, TRK2 overexpression improved the ability of trk1 cells to grow in low K⁺ and led to a higher accumulation of K⁺. Using diS-C₃(3) as a potentiometric probe, we revealed a higher hyperpolarization of trk2 cells compared to the wild type. In addition, the deletion of TRK2 in the trk1 genetic background increased the cell sensitivity to hygromycin B, spermine, and TMA. Our studies reinforced the conclusion that Trk1p is the prominent K⁺ uptake transporter and for the first time revealed that though Trk2p is much less effective, its activity contributes significantly to K⁺ supply and the maintenance of plasma-membrane potential.     

Oak trees and soil interactions in Mediterranean forests: a positive feedback model

Questions: What is the spectrum of variability of chemical elements in a Mediterranean forest ecosystem across the different compartments? Do co‐existing tree species with different leaf chemical composition and nutrient cycling distinctly modify soil conditions? Could these species‐specific, tree‐generated soil changes create a potential positive feedback by affecting long‐term species distribution? Location: Mixed oak forests of southern Spain, Los Alcornocales Natural Park. Methods: We sampled and chemically analysed five different ecosystem components: leaves, leaf fall, litter and superficial (0–25 cm) and sub‐superficial (25–50 cm) soil beneath the canopies of evergreen Quercus suber and deciduous Q. canariensis trees. We used multiple co‐inertia analysis (MCoA) to conjointly analyse the patterns of variability and covariation of eight macro‐ and micronutrients determined in each of the sampled ecological materials. We implemented a path analysis to investigate alternative causal models of relationships among the chemical properties of the different ecosystem components. Results: Variability in the concentration of chemical elements was related to the nature of their biogeochemical cycles. However, the rank of element concentration was consistent across ecosystem components. Analysis of co‐inertia (MCoA) revealed that there was a common underlying multivariate pattern of nutrient enrichment in the ecosystem, which supported the hypothesis of a separation in biogeochemical niches between the two co‐existing oak species, with Q. canariensis having richer plant tissues and more fertile soil directly under each tree than Q. suber. The feasibility of a potential tree–soil positive feedback model was the only statistically validated among several alternative (non‐feedback) models tested. Conclusions: In the studied Mediterranean forests, oak species distinctly modify soil fertility conditions through different nutrient return pathways. Further investigation is needed to address whether these tree‐generated soil changes could affect seedling establishment and ultimately influence species distribution.     

Throwing the baby out with the bathwater: does laurel forest restoration remove a critical winter food supply for the critically endangered Azores bullfinch?

The invasive Clethra arborea has a dual-role in the diet of the Azores bullfinch, a critically endangered bird species endemic to the island of São Miguel (Azores, Portugal). This is a crucial winter food resource but it lowers the availability of native laurel forest species that compose most of the bird’s diet throughout the year. The removal of this and other invasive alien species is part of current laurel forest habitat restoration programmes, disregarding the impact on the Azores bullfinch population. In order to evaluate the first responses of the Azores bullfinch to habitat restoration, we studied bird diet, foraging behaviour, food availability and habitat occupancy in managed (without C. arborea) and control areas. Significant increases in the availability of native food resources in managed areas were noticeable in the diet, particularly the intake of Ilex perado ssp. azorica and Prunus lusitanica ssp. azorica flower buds. In most of the studied months birds heavily used and foraged in managed over control areas. The one exception was in December, when a resource-gap occurred in managed areas, which may be overcome in the short-term due to re-establishment of native plants following removal of invasive aliens.