Decomposition rates and nutrient dynamics of Picea abies needles,twigs and fine roots after stem-only harvesting in eastern and western Norway

Objectives

Afforestation changes soil chemical properties over several decades. In contrast, microbial community structure can be shifted within the first decade and so, the direct effects of tree species can be revealed. The aim of this study was to determine the alteration of soil microbial community composition 10 years after afforestation by trees with contrasting functional traits.

Methods

The study was conducted at the BangorDIVERSE temperate forest experiment. Soil samples were collected under single, two and three species mixtures of alder and birch, beech and oak - early and secondary successional species, respectively, and contiguous agricultural field. Soil was analysed for total carbon (C) and nitrogen (N) contents, and microbial community structure (phospholipid fatty acids (PLFAs) analysis).

Results and conclusions

The total PLFAs content (370–640 nmol g^?1 soil) in forest plots increased for 30 to 110 % compared to the agricultural soil (290 nmol g^?1 soil). In contrast, soil C, N and C/N ratios were altered over 10 years much less - increased only up to 20 % or even decreased (for beech forest).

Afforestation increased bacterial PLFAs by 20–120 %, whereas it had stronger impact on the development of fungal communities (increased by 50–200 %). These effects were proved for all forests, but were more pronounced under the monocultures compared to mixtures. This indicates that species identity has a stronger effect than species diversity. Principal component analysis of PLFAs revealed that under mono and three species mixtures similar microbial communities were formed. In contrast, gram-positive PLFAs and actinomycete PLFAs contributed mainly to differentiation of two species mixtures from other forests. Thus, at the early afforestation stage: i) soil biological properties are altered more than chemical, and ii) tree species identity affects more than species amount on both processes.

     

Seasonal variation in the sex and age composition of the woodcock bag in Denmark

The Eurasian woodcock is a highly valued game bird in Western Europe from which c. 2.7 million individuals are harvested annually from an estimated population of 20–26 million birds. The population size and status remains uncertain due to the cryptic behaviour and widespread and solitary occurrence of woodcock, on breeding and wintering areas, making reliable population surveys difficult. Hunting bag records provide age ratios amongst bagged birds, but sex ratios remain poorly known because of the sexually monomorphic nature of this species. We used DNA analysis to determine sex ratios amongst 327 shot woodcocks from two hunting seasons in Denmark (1 October–31 January, 2012/13 and 2013/14). Based on bag totals, age ratios and sex ratios, juvenile females constituted 37%, juvenile males 27%, adult females 16% and adult males 20% of the annual woodcock bag. The female bias was related to a significant deviation from parity in the sex ratio amongst juvenile birds in October, although no such deviation was found at other times or amongst adults. Compared to limited data from other European countries, our data suggest that autumn migration of woodcock involves an initial wave of juvenile females followed by juvenile males and adults, and perhaps that males stay further north in Europe than females during autumn and winter. This migratory pattern would suggest that postponing the opening of the hunting season could reduce the hunting bag on reproductively valuable females in this polygamous species.     

Morphology of the archaellar motor and associated cytoplasmic cone in Thermococcus kodakaraensis

Archaeal swimming motility is driven by archaella: rotary motors attached to long extracellular filaments. The structure of these motors, and particularly how they are anchored in the absence of a peptidoglycan cell wall, is unknown. Here, we use electron cryotomography to visualize the archaellar basal body in vivo in Thermococcus kodakaraensis KOD1. Compared to the homologous bacterial type IV pilus (T4P), we observe structural similarities as well as several unique features. While the position of the cytoplasmic ATPase appears conserved, it is not braced by linkages that extend upward through the cell envelope as in the T4P, but rather by cytoplasmic components that attach it to a large conical frustum up to 500 nm in diameter at its base. In addition to anchoring the lophotrichous bundle of archaella, the conical frustum associates with chemosensory arrays and ribosome‐excluding material and may function as a polar organizing center for the coccoid cells.     

The non-phagocytic route of collagen uptake: a distinct degradation pathway

The degradation of collagens, the most abundant proteins of the extracellular matrix, is involved in numerous physiological and pathological conditions including cancer invasion. An important turnover pathway involves cellular internalization and degradation of large, soluble collagen fragments, generated by initial cleavage of the insoluble collagen fibers. We have previously observed that in primary mouse fibroblasts, this endocytosis of collagen fragments is dependent on the receptor urokinase plasminogen activator receptor-associated protein (uPARAP)/Endo180. Others have identified additional mechanisms of collagen uptake, with different associated receptors, in other cell types. These receptors include β1-integrins, being responsible for collagen phagocytosis, and the mannose receptor. We have now utilized a newly developed monoclonal antibody against uPARAP/Endo180, which down-regulates the receptor protein level on treated cells, to examine the role of uPARAP/Endo180 as a mediator of collagen internalization by a wide range of cultured cell types. With the exception of macrophages, all cells that proved capable of efficient collagen internalization were of mesenchymal origin and all of these utilized uPARAP/Endo180 for their collagen uptake process. Macrophages internalized collagen in a process mediated by the mannose receptor, a protein belonging to the same protein family as uPARAP/Endo180. β1-Integrins were found not to be involved in the endocytosis of soluble collagen, irrespectively of whether this was mediated by uPARAP/Endo180 or the mannose receptor. This further distinguishes these pathways from the phagocytic uptake of particulate collagen.     

A Versatile System for USER Cloning-Based Assembly of Expression Vectors for Mammalian Cell Engineering

A new versatile mammalian vector system for protein production, cell biology analyses, and cell factory engineering was developed. The vector system applies the ligation-free uracil-excision based technique – USER cloning – to rapidly construct mammalian expression vectors of multiple DNA fragments and with maximum flexibility, both for choice of vector backbone and cargo. The vector system includes a set of basic vectors and a toolbox containing a multitude of DNA building blocks including promoters, terminators, selectable marker- and reporter genes, and sequences encoding an internal ribosome entry site, cellular localization signals and epitope- and purification tags. Building blocks in the toolbox can be easily combined as they contain defined and tested Flexible Assembly Sequence Tags, FASTs. USER cloning with FASTs allows rapid swaps of gene, promoter or selection marker in existing plasmids and simple construction of vectors encoding proteins, which are fused to fluorescence-, purification-, localization-, or epitope tags. The mammalian expression vector assembly platform currently allows for the assembly of up to seven fragments in a single cloning step with correct directionality and with a cloning efficiency above 90%. The functionality of basic vectors for FAST assembly was tested and validated by transient expression of fluorescent model proteins in CHO, U-2-OS and HEK293 cell lines. In this test, we included many of the most common vector elements for heterologous gene expression in mammalian cells, in addition the system is fully extendable by other users. The vector system is designed to facilitate high-throughput genome-scale studies of mammalian cells, such as the newly sequenced CHO cell lines, through the ability to rapidly generate high-fidelity assembly of customizable gene expression vectors.     

Extending Serum Half-life of Albumin by Engineering Neonatal Fc Receptor (FcRn) Binding

A major challenge for the therapeutic use of many peptides and proteins is their short circulatory half-life. Albumin has an extended serum half-life of 3 weeks because of its size and FcRn-mediated recycling that prevents intracellular degradation, properties shared with IgG antibodies. Engineering the strictly pH-dependent IgG-FcRn interaction is known to extend IgG half-life. However, this principle has not been extensively explored for albumin. We have engineered human albumin by introducing single point mutations in the C-terminal end that generated a panel of variants with greatly improved affinities for FcRn. One variant (K573P) with 12-fold improved affinity showed extended serum half-life in normal mice, mice transgenic for human FcRn, and cynomolgus monkeys. Importantly, favorable binding to FcRn was maintained when a single-chain fragment variable antibody was genetically fused to either the N- or the C-terminal end. The engineered albumin variants may be attractive for improving the serum half-life of biopharmaceuticals.     

Detection of arbuscular mycorrhizal fungi (Glomales) in roots by nested PCR and SSCP (Single Stranded Conformation Polymorphism)

PCR amplification of a region of the large subunit ribosomal DNA sequence with Glomus specific primers was used to detect arbuscular mycorrhizal fungi in root tissue of four plant species. The primers were specific to Glomus mosseae, Glomus caledonium, Glomus geosporum, Glomus coronatum, Glomus fragilistratum and Glomus constrictum, and did not recognise sequences from Glomus claroideum. Sequence differences between isolates were detected by Single Stranded Conformation Polymorphisms (SSCPs) in polyacrylamide gels under non-denaturing conditions. Isolates of G. mosseae, G. caledonium and G. coronatum could be separated by their SSCP patterns, while three isolates of G. geosporumshowed no variation. Specific SSCP patterns from isolates of G. mosseae and G. caledonium allowed detection of both fungi in the same root segment. Sequence differences leading to variations in SSCP patterns were confirmed by direct sequencing. This revised version was published online in June 2006 with corrections to the Cover Date.