Quantifying species composition in root mixtures using two methods: near-infrared reflectance spectroscopy and plant wax markers

Understanding of plant interactions is greatly limited by our ability to identify and quantify roots belonging to different species. We proposed and compared two methods for estimating the root biomass proportion of each species in artificial mixtures: near-infrared reflectance spectroscopy (NIRS) and plant wax markers. Two sets of artificial root mixtures composed of two or three herbaceous species were prepared. The proportion of root material of each species in mixtures was estimated from NIRS spectral data (i) and the concentration patterns of n-alkanes (ii), n-alcohols (iii), and n-alkanes +n-alcohols combined (iv). For each data set, calibration equations were developed using multivariate statistical models. The botanical composition of root mixtures was predicted well for all the species considered. The accuracy varied slightly among methods: alkanes < alcohols = alkanes + alcohols < NIRS. Correlation coefficients between predicted and actual root proportions ranged from 0.89 to 0.99 for alkanes + alcohols predictions and from 0.97 to 0.99 for NIRS predictions. These two methods provide promising potential for understanding allocation patterns and competitive interactions.     

Fate of carbon in upland grassland subjected to liming using in situ 13CO2 pulse-labelling

Knowledge of the fate of plant assimilate is fundamental to our understanding of the terrestrial carbon cycle, particularly if we are to predict the effects of changes in climate and land management practices on agroecosystems. Pulse-labelling experiments have revealed that some of the carbon fixed by plants is rapidly allocated below-ground and released back into the atmosphere in respiration. However, little is known about the fate of plant assimilate, not accounted for in soil respiration, in the longer term and how current management practices such as liming may affect this. In southern Scotland, UK, limed and unlimed acid grassland plots were pulse-labelled with ¹³CO₂ and the turnover of ¹³C was studied one and two years after labelling. In this study the amount of labelled carbon remaining in shoot, root, and bulk soil pools, and how this differed between limed and unlimed plots was investigated. The results indicated that plant-root turnover was faster, and plants invested less nitrogen in the roots in the limed plots than in the unlimed plots. More ¹³C remained in the soil in the unlimed treatment compared to the limed treatment, but the main difference was found in the particulate organic matter, which turned over relatively quickly. The label was still above natural abundance one and two years after labelling in many cases. In addition, the results demonstrate that a ¹³CO₂ pulse-label administered for only a few hours can be a useful approach for investigating turnover of carbon several years later.     

Folding kinetics and thermodynamics of Pseudomonas syringae effector protein AvrPto provide insight into translocation via the type III secretion system

In order to infect their hosts, many Gram-negative bacteria translocate agents of infection, called effector proteins, through the type III secretion system (TTSS) into the host cytoplasm. This process is thought to require at least partial unfolding of these agents, raising the question of how an effector protein might unfold to enable its translocation and then refold once it reaches the host cytoplasm. AvrPto is a well-studied effector protein of Pseudomonas syringae pv tomato. The presence of a readily observed unfolded population of AvrPto in aqueous solution and the lack of a known secretion chaperone make it ideal for studying the kinetic and thermodynamic characteristics that facilitate translocation. Application of Nzz exchange spectroscopy revealed a global, two-state folding equilibrium with 16% unfolded population, a folding rate of 1.8 s(-1), and an unfolding rate of 0.33 s(-1) at pH 6.1. TrAvrPto stability increases with increasing pH, with only 2% unfolded population observed at pH 7.0. The R(1) relaxation of TrAvrPto, which is sensitive to both the global anisotropy of folded TrAvrPto and slow exchange between folded and unfolded conformations, provided independent verification of the global kinetic rate constants. Given the acidic apoplast in which the pathogen resides and the more basic host cytoplasm, these results offer an intriguing mechanism by which the pH dependence of stability and slow folding kinetics of AvrPto would allow efficient translocation of the unfolded form through the TTSS and refolding into its functional folded form once inside the host.     

A model for the solution structure of the rod arrestin tetramer

Visual rod arrestin has the ability to self-associate at physiological concentrations. We previously demonstrated that only monomeric arrestin can bind the receptor and that the arrestin tetramer in solution differs from that in the crystal. We employed the Rosetta docking software to generate molecular models of the physiologically relevant solution tetramer based on the monomeric arrestin crystal structure. The resulting models were filtered using the Rosetta energy function, experimental intersubunit distances measured with DEER spectroscopy, and intersubunit contact sites identified by mutagenesis and site-directed spin labeling. This resulted in a unique model for subsequent evaluation. The validity of the model is strongly supported by model-directed crosslinking and targeted mutagenesis that yields arrestin variants deficient in self-association. The structure of the solution tetramer explains its inability to bind rhodopsin and paves the way for experimental studies of the physiological role of rod arrestin self-association.     

High-resolution crystal structure and in vivo function of a kinesin-2 homologue in Giardia intestinalis

A critical component of flagellar assembly, the kinesin-2 heterotrimeric complex powers the anterograde movement of proteinaceous rafts along the outer doublet of axonemes in intraflagellar transport (IFT). We present the first high-resolution structures of a kinesin-2 motor domain and an ATP hydrolysis-deficient motor domain mutant from the parasitic protist Giardia intestinalis. The high-resolution crystal structures of G. intestinalis wild-type kinesin-2 (GiKIN2a) motor domain, with its docked neck linker and the hydrolysis-deficient mutant GiKIN2aT104N were solved in a complex with ADP and Mg(2+) at 1.6 and 1.8 A resolutions, respectively. These high-resolution structures provide unique insight into the nucleotide coordination within the active site. G. intestinalis has eight flagella, and we demonstrate that both kinesin-2 homologues and IFT proteins localize to both cytoplasmic and membrane-bound regions of axonemes, with foci at cell body exit points and the distal flagellar tips. We demonstrate that the T104N mutation causes GiKIN2a to act as a rigor mutant in vitro. Overexpression of GiKIN2aT104N results in significant inhibition of flagellar assembly in the caudal, ventral, and posterolateral flagellar pairs. Thus we confirm the conserved evolutionary structure and functional role of kinesin-2 as the anterograde IFT motor in G. intestinalis.     

AERIAL SURVEYS FOR COASTAL DOLPHINS: ABUNDANCE OF HECTOR'S DOLPHINS OFF THE SOUTH ISLAND WEST COAST, NEW ZEALAND

Line-transect surveys of abundance depend critically on the estimation of detection probability, which includes corrections for availability and visibility. In this aerial line-transect survey for Hector's dolphin ( Cephalorhynchus hectori ) we recorded dive times from a helicopter to estimate the proportion of time that dolphins could be seen at or near the water surface and were therefore "available" to be counted. The proportion of available sightings that were counted on transect lines was estimated by comparing sightings by two independent teams of two observers in the fixed-wing plane. The survey covered the area between Farewell Spit and Milford Sound, off the west coast of the South Island of New Zealand. Survey effort was stratified according to existing distribution data. A total of 142 separate sightings was made on 1,355 km of trackline. Average availability for fifty Hector's dolphin groups observed from the helicopter (161 dive/surface cycles) was 46.3% (CV = 4.2%). Data from the two independent observer teams indicated that 96.2% (CV = 2.3%) of the dolphin groups that were at the surface and on the trackline were seen. The abundance estimate for the South Island west coast, corrected for visibility and availability, was 5,388 Hector's dolphins (CV = 20.6%). The total population estimate for South Island Hector's dolphins is 7,270 (CV = 16.2%).     

Modelling species distributions in Britain: a hierarchical integration of climate and land-cover data

A modelling framework for studying the combined effects of climate and land-cover changes on the distribution of species is presented. The model integrates land-cover data into a correlative bioclimatic model in a scale-dependent hierarchical manner, whereby Artificial Neural Networks are used to characterise species' climatic requirements at the European scale and land-cover requirements at the British scale. The model has been tested against an alternative non-hierarchical approach and has been applied to four plant species in Britain: Rhynchospora alba , Erica tetralix , Salix herbacea and Geranium sylvaticum . Predictive performance has been evaluated using Cohen's Kappa statistic and the area under the Receiver Operating Characteristic curve, and a novel approach to identifying thresholds of occurrence which utilises three levels of confidence has been applied. Results demonstrate reasonable to good predictive performance for each species, with the main patterns of distribution simulated at both 10 km and 1 km resolutions. The incorporation of land-cover data was found to significantly improve purely climate-driven predictions for R. alba and E. tetralix , enabling regions with suitable climate but unsuitable land-cover to be identified. The study thus provides an insight into the roles of climate and land-cover as determinants of species' distributions and it is demonstrated that the modelling approach presented can provide a useful framework for making predictions of distributions under scenarios of changing climate and land-cover type. The paper confirms the potential utility of multi-scale approaches for understanding environmental limitations to species' distributions, and demonstrates that the search for environmental correlates with species' distributions must be addressed at an appropriate spatial scale. Our study contributes to the mounting evidence that hierarchical schemes are characteristic of ecological systems.     

Phenotypic and molecular characterisation of <Emphasis Type="Italic">Brucella</Emphasis> isolates from marine mammals

Background  

Bacteria of the genus Brucella are the causative organisms of brucellosis in animals and man. Previous characterisation of Brucella strains originating from marine mammals showed them to be distinct from the terrestrial species and likely to comprise one or more new taxa. Recently two new species comprising Brucella isolates from marine mammals, B. pinnipedialis and B. ceti, were validly published. Here we report on an extensive study of the molecular and phenotypic characteristics of marine mammal Brucella isolates and on how these characteristics relate to the newly described species.