On the global limits of bioenergy and land use for climate change mitigation

Across energy, agricultural and forestry landscapes, the production of biomass for energy has emerged as a controversial driver of land‐use change. We present a novel, simple methodology, to probe the potential global sustainability limits of bioenergy over time for energy provision and climate change mitigation using a complex‐systems approach for assessing land‐use dynamics. Primary biomass that could provide between 70 EJ year^?1 and 360 EJ year^?1, globally, by 2050 was simulated in the context of different land‐use futures, food diet patterns and climate change mitigation efforts. Our simulations also show ranges of potential greenhouse gas emissions for agriculture, forestry and other land uses by 2050, including not only above‐ground biomass‐related emissions, but also from changes in soil carbon, from as high as 24 GtCO₂eq year^?1 to as low as minus 21 GtCO₂eq year^?1, which would represent a significant source of negative emissions. Based on the modelling simulations, the discussions offer novel insights about bioenergy as part of a broader integrated system. Whilst there are sustainability limits to the scale of bioenergy provision, they are dynamic over time, being responsive to land management options deployed worldwide.     

Facilitation by pioneer shrubs for the ecological restoration of riparian forests in the Central Andes of Colombia

Riparian vegetation has great functional importance in agroscapes because it establishes physical connections between natural ecosystems embedded in an agricultural matrix. Throughout the tropics, the historical demand for cropland and pastures in the lands adjacent to streams has led to the replacement of forests by exotic grasses. In order to find new methods for initiating the restoration of riparian forests, we evaluated the use of the pioneer shrubs Tithonia diversifolia and Piper auritum as nurse species for their effects on (1) competition with dominant grasses; (2) natural regeneration and the ecological facilitation of six native tree species of different successional stages; and (3) influence on microclimate. Over a period of 15 months, 4.4% coverage of P. auritum was insufficient to inhibit grasses. In contrast, 81% coverage of T. diversifolia limited the growth of dominant grasses such as Cenchrus purpureus, Paspalum paniculatum, and Cynodon plectostachyus likely by intercepting more than 90% of photosynthetically active radiation (PAR), even though other factors cannot be ruled out. T. diversifolia showed simultaneous effects of facilitation and competition by promoting higher survival of the planted native trees while slowing the growth of pioneer species and inhibiting the regeneration of native woody plants probably as a result of high light interception. This study suggests that planting T. diversifolia as a nurse shrub may facilitate early stages of restoration by inhibiting the growth of pastures, but requires pruning, both to increase light availability and promote the growth of planted trees and the recruitment of woody plants.     

<Emphasis Type="Italic">IL4</Emphasis> in the 5q31 context: association studies of type 1 diabetes and rheumatoid arthritis in the Spanish population

IL4, the gene coding the prototypic Th2 cytokine, has been frequently studied in the context of several inflammatory conditions, but conclusive results have not been obtained. This gene is located in the 5q31-33 complex genetic region, which shows some susceptibility factors to type 1 diabetes (T1D) and rheumatoid arthritis (RA) among other inflammatory conditions. Our aim was to assess the involvement on T1D and RA of IL4 polymorphisms considered individually and in combination with other polymorphisms in 5q31-33, specifically in the OCTN locus, where the L503F polymorphism has been associated with Crohn's disease and other Th1 diseases. We performed a case-control study including 316 T1D patients, 599 RA patients and 540 healthy controls, all of them corresponding to white Spanish individuals. The IL4 single-nucleotide polymorphisms (SNPs) -590C/T (rs2243250) and the OCTN1 exonic SNP L503F (rs1050152) were analysed in all samples. Frequency comparisons of -590C/T and stratified analysis including both cited SNPs were performed using chi-square tests. The -590C/T IL4 SNP was not found associated with T1D or RA when individual analyses were performed. However, a significant association with T1D emerged after stratification by L503F [p=0.02, odds ratio=1.95, 95% CI=1.07-3.55]. The location of the IL4 gene in the complex 5q31-33 genetic region, which contains many genes involved in immunological responses and presents linkage disequilibrium extended along many kilobases, makes necessary to interpret cautiously the previous IL4-association studies.     

Dopamine D4 receptors are heterogeneously distributed in the striosomes/matrix compartments of the striatum

Two important aspects of striatal function, exploratory behaviour and motor co-ordination, require the integrity of the dopamine D4 receptor subtype. These receptors are also implicated in the pathophysiology of certain neuropsychiatric disorders. However, the distribution of D4 receptors in the striatum has not yet been described and this situation impairs our understanding of the anatomical substrate in which D4 receptors function. We developed a D4 receptor-specific antibody that has permitted us to investigate the regional and cellular localization of the receptor in the neostriatum of the rat, mouse, cat and monkey. The subcellular distribution and the synaptic organization of this receptor were also determined in the rat striatum. We found moderate levels of D4 receptor expression in the caudoputamen and lower levels in the nucleus accumbens. These receptors were expressed in cell bodies and in the neuropil and were heterogeneously distributed among different striatal compartments, being more abundant in striosomes than in the matrix. At the subcellular level, the receptor immunoreactivity was mainly localized to dendritic shafts and spines. The prominent immunoreactivity observed in the striosomes indicates that integrative processes involved in D4-mediated limbic behaviours occurs through the striosomes rather than accumbens, whereas the motor behaviour is based in the striatal matrix.     

Quantification of Extracellular Matrix Proteins from a Rat Lung Scaffold to Provide a Molecular Readout for Tissue Engineering

The use of extracellular matrix (ECM)¹ scaffolds, derived from decellularized tissues for engineered organ generation, holds enormous potential in the field of regenerative medicine. To support organ engineering efforts, we developed a targeted proteomics method to extract and quantify extracellular matrix components from tissues. Our method provides more complete and accurate protein characterization than traditional approaches. This is accomplished through the analysis of both the chaotrope-soluble and -insoluble protein fractions and using recombinantly generated stable isotope labeled peptides for endogenous protein quantification. Using this approach, we have generated 74 peptides, representing 56 proteins to quantify protein in native (nondecellularized) and decellularized lung matrices. We have focused on proteins of the ECM and additional intracellular proteins that are challenging to remove during the decellularization procedure. Results indicate that the acellular lung scaffold is predominantly composed of structural collagens, with the majority of these proteins found in the insoluble ECM, a fraction that is often discarded using widely accepted proteomic methods. The decellularization procedure removes over 98% of intracellular proteins evaluated and retains, to varying degrees, proteoglycans and glycoproteins of the ECM. Accurate characterization of ECM proteins from tissue samples will help advance organ engineering efforts by generating a molecular readout that can be correlated with functional outcome to drive the next generation of engineered organs.Organ transplantation is an established, lifesaving therapy for patients with chronic end-stage diseases. However, transplantation as a therapeutic option is limited by availability of suitable donor organs (1). Although advancements in surgical techniques, such as successful implementation of bilateral lung transplants and improved immunosuppressant treatments, have led to more successful outcomes in recent years, the percentage of people that die while on the transplant wait list has increased (2, 3). One attractive approach to meet this demand is the in vitro generation of organs using decellularized tissues as scaffolds for recellularization. For complex organs such as the lung, these tissue scaffolds can be derived from a donor organ that would have otherwise been unfit for transplantation. This whole organ scaffold can be recellularized using a patient''s own primary or stem-derived cells, thus eliminating many issues related to graft/host incompatibility. This approach was recently used to generate lungs that, when implanted in rat recipients, allowed for gas exchange (4, 5). However, examination of the lung indicated leakage of erythrocytes into the alveolar space, indicating a compromised capillary-endothelial barrier. These exciting results highlighted the potential of the method for organ transplantation but also the need for improved molecular readouts to guide engineering efforts.Efficient reseeding of decellularized scaffolds has been shown to be dependent on retaining native ECM structural integrity and elasticity (6). Local variations in expression of abundant proteins in the ECM scaffolding (collagens, laminins, fibronectins) have been correlated to variance in cell repopulation and subsequent proliferation (7). It is thought that retaining specific ECM components and architectures may allow cells to be directed back to a tissue-specific niche during reseeding and that small changes in abundance of these molecular cues can drastically affect the recellularization process (8). Current methods used to characterize the protein composition of native and acellular tissues involve antibody- or dye-based staining, hydroxyproline assays assessing collagen content, or relative quantification of proteins by liquid chromatography tandem mass spectrometry (LC-MS/MS) (9, 10). All of these methods either fall short in specificity, accurate quantification, or both. A more complete and accurate method for protein characterization would provide a valuable tool for tissue engineering efforts, while shedding light on the possible molecular mechanisms resulting in cell seeding variability and alterations in mechanical properties of engineered lung tissues.Current relative quantification strategies (iTRAQ, Spectral Counting, dimethyl labeling, others) (11–15) perform well when the majority of protein in samples does not change, there are approximately equal increases and decreases in protein levels, or in cases where proteins that are known not to change in abundance can be used for normalization. However, normalization steps often employed have the potential to introduce experimental bias (16). The decellularization process differentially removes and enriches proteins in the ECM scaffolding, depleting some proteins with high efficiency while leaving others mostly intact. This makes relative comparisons between native and decellularized lung challenging. Although strategies can be employed in an attempt to normalize data (17), there is a distinct advantage to quantification methods using stable isotope labeled (SIL) peptides in this application. Here, we developed ECM targeted, isotopically labeled peptides using the QconCAT approach first described by Beynon et al. (18). SIL quantification allows for intra- and intersample comparison of heterogeneous tissues, such as native organs and decellularized scaffolds, with high accuracy and precision.The ECM is largely responsible for defining the biomechanical properties of organs. Maintaining structural rigidity and native microarchitecture through the decellularization process makes an acellular organ a good candidate to serve as a tissue scaffold (19, 20). These same characteristics are a central reason why the ECM is challenging to characterize using common bottom-up proteomics approaches (21). Currently accepted and widely used digestion methods require proteins to be solubilized for bottom-up proteomic analysis (22). Recent papers have reported characterization of the ECM fraction from tissues through the use of strong chaotropes (11, 21, 23–27) or cellular fractionation followed by strong detergent (10, 28, 29). However, in our experience, these protocols invariably yield various sizes of an insoluble protein-containing pellet when applied to a variety of tissue samples (heart, lung, and mammary gland). On one end of the spectrum, methods utilizing deglycosylation and enzymatic digestions for clarification of partial solubilized protein slurries yields good ECM coverage with a high number of spectral matches for collagen alpha-1(I), a highly abundant ECM protein in lung (28). On the other end of the spectrum, methods using only detergents or chaotropes for solubilization result in protein pellets that are generally removed prior to LC-MS/MS analysis. These pellets often contained a majority of fibrillar proteins, resulting in quantitative errors. Consistent with this finding, several of these studies characterizing tissue engineered lungs do not report the identification of collagen alpha-1(I) (8, 10, 30). We believe these observations result from a failure to solubilize and enzymatically digest insoluble ECM proteins. To this end, we explored the use of chemical digestion of the insoluble pellet to improve coverage of the ECM proteome from tissue. This method has been used to quantify protein levels from native and decellularized lungs to determine decellularization specificity and efficiency. The accurate characterization of ECM proteins from lung samples should advance tissue engineering efforts by yielding a readout that can be correlated with functional outcome to drive further development.     

Molecular differentiation of Anopheles (Nyssorhynchus) benarrochi and An. (N.) oswaldoi from southern Colombia

Anopheles (Nyssorhynchus) benarrochi, An. (N.) oswaldoi, and An. (N.) rangeli are the most common anthropophilic mosquitoes in the southern Colombian state of Putumayo. Adult females are most commonly collected in epidemiological studies, and this stage poses significant problems for correct identification, due to overlapping inter-specific morphological characters. Although An. rangeli is easy to identify, the morphological variant of An. benarrochi found in the region and An. oswaldoi are not always easy to separate. Herein we provide a rapid molecular method to distinguish these two species in Southern Colombia. Sequence data for the second internal transcribed spacer (ITS2) region of rDNA was generated for link-reared progeny of An. benarrochi and An. oswaldoi, that had been identified using all life stages. ITS2 sequences were 540 bp in length in An. benarrochi (n = 9) and 531 bp in An. oswaldoi (n = 7). Sequences showed no intra-specific variation and ungapped inter-specific sequence divergence was 6.4%. Species diagnostic banding patterns were recovered following digestion of the ITS2 amplicons with the enzyme Hae III as follows: An. benarrochi (365, 137, and 38 bp) and An. oswaldoi (493 and 38 bp). This polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) assay provides rapid, accurate, and inexpensive species diagnosis of adult females. This will benefit future epidemiological studies and, as PCR amplification can be achieved using a single mosquito leg, the remaining specimen can be either retained as a morphological voucher or further used in vector incrimination studies. That An. benarrochi comprises a complex of at least two species across Latin America is discussed.     

Epistasis in the expression of relevant traits in cassava (Manihot esculenta Crantz) for subhumid conditions

There is limited knowledge on the inheritance of agronomic traits in cassava and the importance of epistasis for most crops. A nine-parent diallel study was conducted in subhumid environments. Thirty clones were obtained from each F1 cross. Each clone was represented by six plants, which were distributed in three replications at two locations. Therefore the same 30 genotypes of each F1 cross were planted in the three replications at the two locations. Analysis of variance suggested significant genetic effects for all variables analyzed (reaction to thrips, fresh root and foliage yields, harvest index, dry matter content, and root dry matter yield). Significant epistatic effects were observed for all variables, except harvest index. Dominance variance was always significant, except for dry matter content and dry matter yield. Additive variance was significant only for reaction to thrips. Results suggested that dominance plays an important role in complex traits such as root yield. The significance of epistasis can help us understand the difficulties of quantitative genetics models and QTLs in satisfactorily explaining phenotypic variation in traits with complex inheritance. Significant epistasis would justify the production of inbred parental lines to fix favorable allele combinations in the production of hybrid cassava cultivars.