A novel approach to an old problem: tracking dispersed seeds

Animals are the principal vectors of dispersal for a large number of plant species. Unfortunately it is not easy to discern their movement patterns or the fate of their dispersed seeds. Many animals transport seeds by consuming them and then, some time later, defecating them. Others gather seeds and then store them for later consumption. Both circumstances lead to a set of seeds that have been dispersed in a clumped pattern, which offers a unique opportunity to assess seed movements. We introduce a novel approach that uses maternally inherited seed tissue to quantify the genetic structure of dispersed seed pools. This direct approach measures the genetic variability within and among seed pools, and estimates the scale of seed movement, without requiring a highly polymorphic battery of markers or the location and genotypes of all possible seed parents. We demonstrate this approach with the specific case of seed transport of valley oak (Quercus lobata) acorns by acorn woodpeckers (Melanerpes formicivorus). These territorial birds store acorns in drilled holes in the bark of trees, called granaries. We sampled stored acorns from different granaries, extracted DNA from the maternally inherited pericarp, and then assessed individuals for three microsatellite markers. We found extremely high genetic structure among granaries, a low number of effective seed donors per granary, and restricted seed movement. A maternity analysis performed on the same sample with seven microsatellites confirms acorn transport is limited to approximately 100-m radius. Our findings provide insight into the foraging and seed-dispersal behaviour of acorn woodpeckers, with an approach that can be widely extended to other systems.     

A Bacillus thuringiensis delta-endotoxin induces programmed cell death in mosquito larvae

We present evidence that a delta-endotoxin isolated from Bacillus thuringiensis subsp.israelensis induces programmed cell death in polytene midgut cells of Culex pipiens larvae. After exposure to toxin, polytene nuclei in the anterior region of the larval midgut undergo many of the morphological and physiological changes which are characteristic of apoptosis, including the ability to stain with the vital dye, acridine orange, and fragmentation of nuclear DNA as demonstrated by agarose gel electrophoresis and in situ TUNEL labeling. The temporal sequence of toxin ingestion, acridine orange staining and larval death suggests a cause and effect relationship between programmed cell death and larval death. Amino sugars that interfere with toxicity also interfere with the time course of acridine orange staining of larval polytene nuclei. The toxin first causes programmed cell death of anterior midgut and gastric caeca cells and, subsequently, posterior midgut cells. This pattern is similar to the temporal sequence of larval polytene cell death that occurs during metamorphosis. From the size and distribution of the nuclei that are stained with acridine orange, it appears that only polytene midgut cells are affected by toxin and that the diploid regenerative cell are not affected.     

N2 Fixation (C2H2-Reducing Activity) and Leghaemoglobin Content during Nitrate- and Water-Stress-Induced Senescence of Medicago sativa Root Nodules

Nitrate and water stress were used to induce senescence in rootnodules of alfalfa (Medicago sativa L. cv. Aragon). Nodule senescencewas assessed by determinations of the nitrogenase (C₂H₂-reducing)activity, and the leghaemoglobin (LHb) and total soluble proteincontents of the nodules. Nodules responded similarly to and water stress in many respects, but there was a significant difference.All parameters of nodule activity, expressed on the basis ofnodule dry weight (DW), consistently decreased following treatmentwith or during drought; there was a significant interaction (synergism) between the inhibitory effects of and water stress on nitrogenase activity, but sucheffects were merely additive in the case of LHb content or LHb/solubleprotein ratio. However, caused the selective decay of LHb with respect to other nodular soluble proteins,whereas the decrease of LHb during water stress was due to ageneral inhibition of protein synthesis and to an increasedproteolytic activity in the nodule cytosol rather than to aspecific proteolysis of LHb. Key words: Leghaemoglobin, Medicago saliva, nitrogen fixation, root nodule senescence, water stress     

Morphology agrees with molecular data: phylogenetic affinities of Euptychiina butterflies (Nymphalidae: Satyrinae)

Euptychiina is the most species‐rich subtribe of Neotropical Satyrinae, with over 450 known species in 47 genera (14 monotypic). Here, we use morphological characters to examine the phylogenetic relationships within Euptychiina. Taxonomic sampling included 105 species representing the majority of the genera, as well as five outgroups. A total of 103 characters were obtained: 45 from wing pattern, 48 from genitalia and 10 from wing venation. The data matrix was analysed using maximum parsimony under both equal and extended implied weights. Euptychiina was recovered as monophyletic with ten monophyletic genera, contrasting previous DNA sequence‐based phylogenies that did not recover the monophyly of the group. In agreement with sequence‐based hypotheses, however, three main clades were recognized: the ‘Megisto clade’ with six monophyletic and three polyphyletic genera, the ‘Taygetis clade’ with nine genera of which three were monophyletic, and the ‘Pareuptyhia clade’ with four monophyletic and two polyphyletic genera. This is the first morphology‐based phylogenetic hypothesis for Euptychiina and the results will be used to complement molecular data in a combined analysis and to provide critical synapomorphies for clades and genera in this taxonomically confused group.     

A new method of estimating the pollen dispersal curve independently of effective density

We introduce a novel indirect method of estimating the pollen dispersal curve from mother-offspring genotypic data. Unlike an earlier indirect approach (TwoGener), this method is based on a normalized measure of correlated paternity between female pairs whose expectation does not explicitly depend on the unknown effective male population density (d(e)). We investigate the statistical properties of the new method, by comparison with those of TwoGener, considering the sensitivity to reductions of d(e), relative to census density, resulting from unequal male fecundity and asynchronous flowering. Our main results are: (i) it is possible to obtain reliable estimates of the average distance of pollen dispersal, delta, from indirect methods, even under nonuniform male fecundity and variable flowering phenology; (ii) the new method yields more accurate and more precise delta-estimates than TwoGener under a wide range of sampling and flowering scenarios; and (iii) TwoGener can be used to obtain approximate d(e) estimates, if needed for other purposes. Our results also show that accurately estimating the shape of the tail of the pollen dispersal function by means of indirect methods remains a very difficult challenge.     

Carbon and nitrogen content based estimation of the fat content of animal carcasses in various species

The objective of this study was to explore whether the C and N content can be used to estimate the fat content of animal carcasses. Considering the mean C and N contents of body fat and body protein, the fat content (EE) [%] can be predicted from C and N values [%] according to the generally valid equation EE=1.3038·C – 4.237·N. The application of this equation to estimate the total fat content of all animal carcasses results in significant differences in fat content between predicted and measured values. Therefore, we derived specific equations for rats, pigs, cattle, sheep, broilers and mice to predict the fat content by dual linear regression analysis (y=EE [% DM], x₁=C [% DM], x₂=N [% DM]) based on measured fat, C, and N contents of animal body samples. The specific equations for different animals showed residual standard deviations of 1.55, 1.63, 1.12, 1.35, 1.85 and 0.92% fat for rats, pigs, cattle, sheep, broilers and mice, respectively.     

Diversity in Mediterranean farm ponds: trade‐offs and synergies between irrigation modernisation and biodiversity conservation

1. Agricultural intensification has caused dramatic biodiversity loss in many agricultural landscapes over the last century. Here, we investigated whether new types of farm ponds (made of artificial substrata) in intensive systems and natural‐substratum ponds in traditional farming systems differ in their value for aquatic biodiversity conservation. 2. We analysed the main patterns of environmental variation, compared α‐, β‐ and γ‐diversity of macroinvertebrates between ponds types and evaluated the role of submerged aquatic vegetation (SAV). Generalised additive models (GAM) were used to analyse the relationships of α‐ and β‐diversity with environmental predictors, and variation partitioning to separate the effect of environmental and spatial characteristics on the variation in macroinvertebrate assemblages. Moran’s eigenvector maps (MEMs) were used to define spatial variables. 3. A principal coordinate analysis (PCoA) detected a primary environmental gradient that separated nutrient‐rich ponds from those dominated by SAV; a secondary morphometric gradient distinguished natural‐substratum ponds, with large surface area and structural complexity, from artificial‐substratum ponds with steeper slopes. Natural‐substratum ponds had almost twice the α‐ and γ‐diversity of artificial‐substratum ponds, and diversity significantly increased when SAV was present, particularly in artificial‐substratum ponds. Total phosphorus (TP) strongly contributed to explain the patterns in diversity, while SAV was a significant predictor of assemblage composition and diversity. GAMs revealed optima of both α‐diversity at intermediate SAV covers and β‐diversity at intermediate–high TP concentrations. 4. These findings have important implications for conservation planning. Adaptation of artificial‐substratum ponds by adding natural substratum and smoothing the gradient of pond margins would improve their conservation value. Development of SAV with occasional harvests and certain cautionary measures to control nutrient levels may also improve both the agronomical and environmental function of ponds.     

Low plant density enhances gene dispersal in the Amazonian understory herb Heliconia acuminata

In theory, conservation genetics predicts that forest fragmentation will reduce gene dispersal, but in practice, genetic and ecological processes are also dependent on other population characteristics. We used Bayesian genetic analyses to characterize parentage and propagule dispersal in Heliconia acuminata L. C. Richard (Heliconiaceae), a common Amazonian understory plant that is pollinated and dispersed by birds. We studied these processes in two continuous forest sites and three 1‐ha fragments in Brazil's Biological Dynamics of Forest Fragments Project. These sites showed variation in the density of H. acuminata. Ten microsatellite markers were used to genotype flowering adults and seedling recruits and to quantify realized pollen and seed dispersal distances, immigration of propagules from outside populations, and reproductive dominance among parents. We tested whether gene dispersal is more dependent on fragmentation or density of reproductive plants. Low plant densities were associated with elevated immigration rates and greater propagule dispersal distances. Reproductive dominance among inside‐plot parents was higher for low‐density than for high‐density populations. Elevated local flower and fruit availability is probably leading to spatially more proximal bird foraging and propagule dispersal in areas with high density of reproductive plants. Nevertheless, genetic diversity, inbreeding coefficients and fine‐scale spatial genetic structure were similar across populations, despite differences in gene dispersal. This result may indicate that the opposing processes of longer dispersal events in low‐density populations vs. higher diversity of contributing parents in high‐density populations balance the resulting genetic outcomes and prevent genetic erosion in small populations and fragments.     

Characterization of a G Protein -Coupled Guanylyl Cyclase -B Receptor from Bovine Tracheal Smooth Muscle

A G protein-coupled natriuretic peptide-guanylyl cyclase receptor-B (NPR-B) located in plasma membranes from bovine tracheal smooth muscle shows complex kinetics and regulation. NPR-B was activated by natriuretic peptides (CNP-53 > ANP-28) at the ligand extracellular domain, stimulated by Gq-protein activators, such as mastoparan, and inhibited by Gi-sensitive chloride, interacting at the juxtamembrane domain. The kinase homology domain was evaluated by the ATP inhibition of Mn²⁺-activated NPR-B, which was partially reversed by mastoparan. The catalytic domain was studied by kinetics of Mn²⁺/Mg²⁺ and GTP, and the catalytic effect with GTP analogues with modifications of the /γ phosphates and ribose moieties. Most NPR-B biochemical properties remained after detergent solubilization but the mastoparan activation and chloride inhibition of NPR-B disappeared. Our results indicate that NPR-B is a highly regulated nano-machinery with domains acting at cross-talk points with other signal transducing cascades initiated by G protein-coupled receptors and affected by intracellular ligands such as chloride, Mn²⁺, Mg²⁺, ATP, and GTP.     

Seed germination temperatures of eight Mexican Agave species with economic importance

The genetic diversity of Agave plants is threatened by clonal commercial reproduction and climatic change. Sexual reproduction is uncommon and research on seed germination is scarce. The present study evaluated the seed germination of Agave lechuguilla, Agave striata, Agave americana var. marginata, Agave asperrima, Agave cupreata, Agave duranguesis, Agave angustifolia ssp. tequilana and Agave salmiana at constant temperatures (10, 15, 20, 25, 30, 35 and 40°C). Initial imbibition (after the first 12 h) was significantly variable among species, positively correlated with seed weight (r = 0.6560, P < 0.001) and increased with temperature (from 35% at 10°C to 66% at 40°C). Temperature affected maximum imbibition (83–150%) for A. asperrima, A. lechuguilla, A. salmiana and A. striata; other species averaged 110%. Most germination kinetics best fitted a logistic model, whereas only a few treatments fit a Weibull model. The time to germination onset diminished (P < 0.05) from 125–173 h at 15°C to 68–84 h at 25°C, and then ascended to 84–196 h at 35°C. The mean germination rate and seed germination percentage after 312 h peaked at 25°C (0.50–0.95% seeds/h and 85–99%, respectively) and fell (P < 0.05) to near zero at 10 and 40°C. Temperatures of 10, 35 and 40°C were partially lethal to A. asperrima, A. duranguensis and A. salmiana seeds. The time to germination onset, seed germination percentage after 312 h and mean germination rate are best described by a Gaussian distribution, with its optimum at approximately 25°C. Thus, optimum temperatures are related to the ecological characteristics of each species area.