Development of a panel of genome-wide ancestry informative markers to study admixture throughout the Americas

Most individuals throughout the Americas are admixed descendants of Native American, European, and African ancestors. Complex historical factors have resulted in varying proportions of ancestral contributions between individuals within and among ethnic groups. We developed a panel of 446 ancestry informative markers (AIMs) optimized to estimate ancestral proportions in individuals and populations throughout Latin America. We used genome-wide data from 953 individuals from diverse African, European, and Native American populations to select AIMs optimized for each of the three main continental populations that form the basis of modern Latin American populations. We selected markers on the basis of locus-specific branch length to be informative, well distributed throughout the genome, capable of being genotyped on widely available commercial platforms, and applicable throughout the Americas by minimizing within-continent heterogeneity. We then validated the panel in samples from four admixed populations by comparing ancestry estimates based on the AIMs panel to estimates based on genome-wide association study (GWAS) data. The panel provided balanced discriminatory power among the three ancestral populations and accurate estimates of individual ancestry proportions (R2 > 0.9 for ancestral components with significant between-subject variance). Finally, we genotyped samples from 18 populations from Latin America using the AIMs panel and estimated variability in ancestry within and between these populations. This panel and its reference genotype information will be useful resources to explore population history of admixture in Latin America and to correct for the potential effects of population stratification in admixed samples in the region.     

Membrane Potential and Catecholamine Secretion by Bovine Adrenal Chromaffin Cells: Use of Tetraphenylphosphonium Distribution and Carbocyanine Dye Fluorescence

Changes in plasma membrane potential of isolated bovine adrenal chromaffin cells were measured independently by two chemical probe methods and related to corresponding effects on catecholamine secretion. The lipophilic cation tetraphenylphosphonium (TPP+) and the carbocyanine dye 3,3'-dipropylthiadicarbocyanine [DiS-C3-(5)] were used. The necessity of evaluating the subcellular distribution of TPP+ among cytoplasmic, mitochondrial, secretory granule, and bound compartments was demonstrated and the resting plasma membrane potential determined to be -55 mV. The relationship between membrane potential and catecholamine secretion was determined in response to variations in extracellular K+ and to the presence of several secretagogues including cholinergic receptor ligands, veratridine, and ionophores for Na+ and K+. The dependence of potential on K+ concentration fit the Goldman constant field equation with a Na/K permeability ratio of 0.1. The dependence of both K+- and veratridine-evoked catecholamine secretion on membrane potential exhibited a potential threshold of about -40 mV before a significant rise in secretion occurred. This is likely related to the threshold for opening of voltage-sensitive Ca2+ channels. Acetylcholine and nicotine evoked a large secretory response without a sufficiently sustained depolarization to be detectable by the relatively slow potential sensitive chemical probes. Decamethonium induced a detectable depolarization of the chromaffin cells. Veratridine and gramicidin evoked both membrane depolarization and catecholamine release. By contrast the K ionophore valinomycin evoked significant levels of secretion without any depolarization. This is consistent with its utilization of an intracellular source of Ca2+ and the independence of its measured secretory response on extracellular Ca2+.     

Reduced expression of selected FASCICLIN‐LIKE ARABINOGALACTAN PROTEIN genes associates with the abortion of kernels in field crops of Zea mays (maize) and of Arabidopsis seeds

Abortion of fertilized ovaries at the tip of the ear can generate significant yield losses in maize crops. To investigate the mechanisms involved in this process, 2 maize hybrids were grown in field crops at 2 sowing densities and under 3 irrigation regimes (well‐watered control, drought before pollination, and drought during pollination), in all possible combinations. Samples of ear tips were taken 2–6 days after synchronous hand pollination and used for the analysis of gene expression and sugars. Glucose and fructose levels increased in kernels with high abortion risk. Several FASCICLIN‐LIKE ARABINOGALACTAN PROTEIN (FLA) genes showed negative correlation with abortion. The expression of ZmFLA7 responded to drought only at the tip of the ear. The abundance of arabinogalactan protein (AGP) glycan epitopes decreased with drought and pharmacological treatments that reduce AGP activity enhanced the abortion of fertilized ovaries. Drought also reduced the expression of AthFLA9 in the siliques of Arabidopsis thaliana. Gain‐ and loss‐of‐function mutants of Arabidopsis showed a negative correlation between AthFLA9 and seed abortion. On the basis of gene expression patterns, pharmacological, and genetic evidence, we propose that stress‐induced reductions in the expression of selected FLA genes enhance abortion of fertilized ovaries in maize and Arabidopsis.     

Three dimensional typological studies using scanning electron microscopy for characterization of Termitomyces pellets obtained from submerged growth conditions

There are gaps in existing understanding of fungal pellet growth dynamics. We used scanning electron microscopy (SEM) for morphological characterization of the biomass organization of Termitomyces pellets for seven species: T. microcarpus (TMI1), T. albuminosus (TAL1, TAL2), T. striatus (TSTR), T. aurantiacus (TAUR), T. heimii (THE1, THE2), T. globulus (TGLO) and T. clypeatus (TCL1, TCL2, TCL3, TCL4, TCL5). We assessed the utility of SEM for morphological and structural characterization of Termitomyces spp. in three dimensional (3D) pellet form to identify ideal pellet morphology for industrial use. Typological classification of Termitomyces species was based on furrows, isotropy, total motifs and fractal dimensions. The pellets formed were entangled and exhibited highly compacted mycelial mass with microheterogeneity and microporosity. The mean density of furrows of Termitomyces species was between 10,000 and 11,300 cm/cm², percentage isotropy was 30?80 and total motifs varied from 300 to 2500. TGLO exhibited the highest furrow mean density, 11243 cm/cm², which indicated a compact, cerebroid structure with complex ridges and furrows, whereas TAL2 exhibited the lowest furrow density. TMI1a exhibited a high percentage isotropic value, 74.6, TSTR exhibited the lowest, 30.9. Total motif number also was used as a typological classification parameter. Fractal values were 2.64?2.78 for various submerged conditions of Termitomyces species. TAL1 exhibited the highest fractal dimension and TAL2 the lowest, which indicates the complexity of branching patterns. Three-dimensional SEM image analysis can provide insight into pellet micromorphology and is a powerful tool for exploring topographical details of pellets.     

Sea urchin Hox genes: insights into the ancestral Hox cluster

We describe the Hox cluster in the radially symmetric sea urchin and compare our findings to what is known from clusters in bilaterally symmetric animals. Several Hox genes from the direct-developing sea urchin Heliocidaris erythrogramma are described. CHEF gel analysis shows that the Hox genes are clustered on a < or = 300 kilobase (kb) fragment of DNA, and only a single cluster is present, as in lower chordates and other nonvertebrate metazoans. Phylogenetic analyses of sea urchin, amphioxus, Drosophila, and selected vertebrate Hox genes confirm that the H. erythrogramma genes, and others previously cloned from other sea urchins, belong to anterior, central, and posterior groups. Despite their radial body plan and lack of cephalization, echinoderms retain at least one of the anterior group Hox genes, an orthologue of Hox3. The structure of the echinoderm Hox cluster suggests that the ancestral deuterostome had a Hox cluster more similar to the current chordate cluster than was expected Sea urchins have at least three Abd-B type genes, suggesting that Abd-B expansion began before the radiation of deuterostomes.      

Cytolytic activity and immunological similarity of the Bacillus thuringiensis subsp. israelensis and Bacillus thuringiensis subsp. morrisoni isolate PG-14 toxins

The parasporal bodies of the mosquitocidal isolates of Bacillus thuringiensis subsp. israelensis and B. thuringiensis subsp. morrisoni isolate PG-14 were compared with regard to their hemolytic and cytolytic activities and the immunological relatedness of the 28- and 65-kilodalton (kDa) proteins that occur in both subspecies. The alkali-solubilized parasporal bodies of B. thuringiensis subsp. israelensis caused 50% lysis of human erythrocytes at 1.14 micrograms/ml, whereas those of B. thuringiensis subsp. morrisoni caused similar lysis at 1.84 micrograms/ml. Preincubation of solubilized parasporal bodies with dioleolyl phosphatidylcholine significantly inhibited the hemolytic activity of both supspecies. In cytolytic assays against Aedes albopictus cells, the toxin concentrations causing 50% lysis for B. thuringiensis subsp. israelensis and B. thuringiensis subsp. morrisoni were 1.87 and 11.98 micrograms/ml, respectively. Polyclonal antibodies raised separately against the 25-kDa protein (a tryptic digest of the 28-kDa protein) or the 65-kDa protein of B. thuringiensis subsp. israelensis cross-reacted, respectively, with the 28- and the 65-kDa proteins of B. thuringiensis subsp. morrisoni. However, neither of these antibodies cross-reacted with the 135-kDa protein of either subspecies. These results indicate that the mosquitocidal and hemolytic properties of B. thuringiensis subsp. israelensis and B. thuringiensis subsp. morrisoni isolate PG-14 are probably due to the biologically related proteins that are present in the parasporal bodies of both subspecies. The lower hemolytic activity of the B. thuringiensis subsp. morrisoni may be due to the presence of lower levels of the 28-kDa protein in that subspecies.(ABSTRACT TRUNCATED AT 250 WORDS)     

Isolation of a relatively nontoxic 65-kilodalton protein inclusion from the parasporal body of Bacillus thuringiensis subsp. israelensis.

Ultrastructural studies of the mosquitocidal bacterium Bacillus thuringiensis subsp. israelensis revealed that the parasporal body contained three major inclusion types, designated types 1, 2, and 3, which could be differentiated on the basis of electron opacity and size and, to some extent, shape. The type-2 inclusion, which was of moderate electron density and often appeared as a bar-shaped polyhedral body, was isolated on NaBr gradients from purified parasporal bodies and characterized by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, transmission electron microscopy, and bioassays against neonate larvae of Aedes aegypti. Purified inclusions averaged 150 to 200 nm by 500 to 700 nm in transverse sections and consisted almost exclusively of a 65-kilodalton (kDa) protein contaminated with minor quantities of 38- and 28-kDa proteins. Lethal concentration values at the 50% level for preparations of the purified parasporal body and the type-2 inclusion were, respectively, 0.66 and 43 ng/ml, indicating that the 65-kDa protein is only slightly toxic to mosquitoes in comparison to the intact parasporal body. Analysis of the type-2 polyhedral inclusion by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and bioassays during different stages of purification demonstrated a positive correlation between the toxicity of the preparation and the degree of contamination with the 28-kDa protein. These results indicate that the 65-kDa protein is not the primary larvicidal toxin, although it may act in conjunction with other parasporal body proteins to produce the high mosquitocidal toxicity characteristic of this bacterium.     

The relevance of in vitro anthelmintic screening tests employing the free-living stages of trichostrongylid nematodes

The response of the free-living stages of Nippostrongylus brasiliensis, Nematospiroides dubius, Haemonchus contortus, Trichostrongylus colubriformis and Ostertagia ostertagi to a wide variety of antiparasitic agents in vitro was investigated. All the major broad spectrum veterinary anthelmintics showed good activity against each of these worms with EC50 values varying from about 0.0002 mg/l for certain benzimidazoles and ivermectin to about 6.5 mg/l for febantel. Of 22 known narrow spectrum anthelmintics useful only against H. contortus and/or helminths other than trichostrongyles, only 10% showed good activity at concentrations equal to or less than 10.0 mg/l. Further, only one of 15 antiprotozoal agents showed good activity in these tests at the 10.0 mg/l level. The screening test employing free-living Nippostrongylus brasiliensis was selected for an extended trial where the evaluation of 1400 miscellaneous organic chemicals was undertaken. Approximately 10% of these showed activity at concentrations equal to or less than 10.0 mg/l. It is concluded that in vitro screening tests employing the free-living stages of these five genera of nematodes afford simple yet effective means for selecting relevant compounds for further evaluation as possible leads to new broad spectrum anthelmintics for use in ruminants. However, tests using the free-living stages of these worms, including H. contortus, are unsuitable for detecting narrow spectrum 'specifics', e.g., for the treatment of haemonchiasis.