Isolation and characterization of microsatellites in Pacific white shrimp Penaeus (Litopenaeus) vannamei

Five polymorphic microsatellite loci were characterized for Penaeus (Litopenaeus) vannamei. Loci were isolated using a partial Sau3A1 genomic library by the sequencing of randomly selected clones and by a biotinylated (CT)₁₀ and (GT)₁₀ probes screening procedure. The last strategy resulted in the most useful data. About 40% of the clones showed a previously reported satellite/microsatellite (PVS1), reducing the chance of finding new microsatellite regions. Whereas two of the microsatellite loci with more than 10 alleles will be useful for mating analysis in a breeding program, the others might prove useful for population genetic studies.     

Intracellular Ca2+ concentration and the antidiuretic hormone-induced increase in water permeability: effects of ionophore A23187 and quinidine

The hydroosmotic responses induced by oxytocin and 8-bromo-cyclic AMP, in frog and toad urinary bladders, were recorded minute by minute. 3HHO and 45Ca unidirectional fluxes as well as prostaglandin B2 liberation were also measured. It was observed that: (1) Addition of the calcium ionophore A23187 or quinidine to the serosal bath inhibited the response to oxytocin, but not to 8-bromo-cyclic AMP, while increasing prostaglandin E1 liberation into the serosal but not into the mucosal bath. (2) Addition of A23187 to the mucosal bath induced a transient and temperature-dependent inhibition of the response elicited by 8-bromo-cyclic AMP. The time-course of this reduction in water permeability and its sensitivity to medium temperature were similar to those observed after the withdrawal of agonist, but clearly different of those observed after intracellular acidification. (3) The hydroosmotic response was also transitorily inhibited when the Ca2+ concentration was step-changed in the mucosal bath. (4) When added to the mucosal or to the serosal baths, the ionophore increased either the apical or the laterobasal Ca2+ permeabilities. It is concluded that manipulation of intracellular Ca2+ interferes with the hydroosmotic response at two different levels. (1) A first target point located 'pre-cyclic-AMP production'. This effect would be mediated by prostaglandin liberation. (2) A second target point located after cyclic AMP production and before the 'temperature-dependent rate-limiting step'. This effect is probably related to the mechanism controlling the insertion and removal of water channels.     

Testosterone alters duodenal calcium transport and longitudinal bone growth rate in parallel in the male rat.

Duodenal active calcium transport and longitudinal bone growth rate have been shown previously to be regulated in parallel by alteration of gonadal hormone status in sexually maturing female rats. The present study was designed to extend these observations to the sexually maturing male rat. Male rats were orchidectomized (ORX) and given Silastic implants containing either testosterone or estradiol at 6 weeks of age. At 9 weeks of age, duodenal active calcium transport was measured by the everted gut sac method and longitudinal bone growth rate was determined by tetracycline labeling. Decreases in body weight, longitudinal bone growth rate, duodenal calcium transport, and serum Ca and P were exhibited by ORX animals as compared with age-matched control animals. Testosterone administration to ORX animals resulted in an increase in body weight, longitudinal bone growth rate, duodenal calcium transport, and serum Ca and P as compared with ORX animals to a level not significantly different from that of age-matched control animals. Estradiol administration to ORX animals resulted in an additional decrease in body weight, although no significant effect on duodenal calcium transport, serum Ca, or P was noted as compared with ORX animals. There were no statistically significant alterations in the circulating levels of 1,25-dihydroxyvitamin D, parathyroid hormone, or osteocalcin in response to any of the experimental manipulations of gonadal status. These results indicate that, as in the female, gonadal hormone status affects intestinal calcium transport in sexually maturing male rats in parallel with changes in bone growth rate by mechanisms that are independent of circulating levels of 1,25-dihydroxyvitamin D.     

Enumeration of potentially pathogenic bacteria from sewage sludges.

To ascertain the health risks that may be posed by the land application of sewage sludges, a scheme was devised to determine the types and numbers of pathogenic and potentially pathogenic bacteria present in sludges. A processing treatment was adapted to sludge to give a homogenate which yielded the greatest numbers of viable bacteria. Conventional methods were successful in enumerating Klebsiella, Staphylococcus, gram-negative enteric bacteria, and commonly used indicator organisms. Modifications of conventional methods improved the enumeration of Salmonella, Mycobacterium sp., fluorescent Pseudomonas sp., and Clostridium perfringens. However, Shigella methodology yielded only one isolate. Utilizing the proposed scheme, the population densities of these organisms were estimated in three domestic wastewater sludges. In light of these results, the potential impact of land application of sewage sludges is discussed.     

Establishment of polarized endocytosis in differentiable intestinal HT29-18 subclones

Subclones of the HT29-18 clone, derived from a human adenocarcinoma, are able to acquire an enterocyte-like phenotype depending on the culture conditions. To investigate fluid-phase and receptor-mediated endocytosis in the polarized subclone HT29-18-C1, we established culture conditions that allowed cell growth on permeable supports. HT29-18-C1 monolayers had an electrical resistance of 43 ohms.cm2 and developed a transepithelial potential of about 2 mV. Transferrin receptors were uniformly distributed on the entire cell surface of undifferentiated HT29-18 cells but were located on the basolateral membrane of differentiated cells. Transferrin had a high affinity (Kd = 2.5 x 10(-9) M) for its receptor independent of the state of differentiation. The number of transferrin receptors and the mRNA amounts encoding them were comparable in the undifferentiated and differentiated HT29-18 cells. Transferrin was quickly internalized and recycled back to the cell surface of undifferentiated HT29-18 cells. The same phenomenon also occurred in differentiated HT29-18 cells, but the receptors were limited to the basolateral membrane. In the presence of ammonium chloride, the process was slower but remained polarized. Fluid-phase uptake was also investigated with horseradish peroxidase (HRP) in differentiated HT29-18 C1 cells. HRP that was internalized in 1 hour from a given membrane domain preferentially recycled back to the same membrane domain. No significant accumulation of the enzyme in the late endosomes and lysosomes of the differentiated HT29-18-C1 cells was observed.     

The Sabinas syndrome

We have defined a new autosomal recessive disorder in patients stemming from a small community in northern Mexico. Diagnosable at birth, its major symptoms include brittle hair, mental retardation, and nail dysplasia. Structural hair abnormalities are seen by both light and electron microscopy. Hair cystine content is reduced while the copper/zinc ratio in hair is increased.     

Discrimination of coloured patterns by honeybees through chromatic and achromatic cues

We investigated pattern discrimination by worker honeybees, Apis mellifera, focusing on the roles of spectral cues and the angular size of patterns. Free-flying bees were trained to discriminate concentric patterns in a Y-maze. The rewarded pattern could be composed of either a cyan and a yellow colour, which presented both different chromatic and achromatic L-receptor contrast, or an orange and a blue colour, which presented different chromatic cues, but the same L-receptor contrast. The non-rewarded alternative was either a single-coloured disc with the colour of the central disc or the surrounding ring of the pattern, a checkerboard pattern with non-resolvable squares, the reversed pattern, or the elements of the training pattern (disc or ring alone). Bees resolved and learned both colour elements in the rewarded patterns and their spatial properties. When the patterns subtended large visual angles, this discrimination used chromatic cues only. Patterns with yellow or orange central discs were generalised toward the yellow and orange colours, respectively. When the patterns subtended a visual angle close to the detection limit and L-receptor contrast was mediating discrimination, pattern perception was reduced: bees perceived only the pattern element with higher contrast.     

Antagonism between Cry1Ac1 and Cyt1A1 Toxins of Bacillus thuringiensis

Most strains of the insecticidal bacterium Bacillus thuringiensis have a combination of different protoxins in their parasporal crystals. Some of the combinations clearly interact synergistically, like the toxins present in B. thuringiensis subsp. israelensis. In this paper we describe a novel joint activity of toxins from different strains of B. thuringiensis. In vitro bioassays in which we used pure, trypsin-activated Cry1Ac1 proteins from B. thuringiensis subsp. kurstaki, Cyt1A1 from B. thuringiensis subsp. israelensis, and Trichoplusia ni BTI-Tn5B1-4 cells revealed contrasting susceptibility characteristics. The 50% lethal concentrations (LC₅₀s) were estimated to be 4,967 of Cry1Ac1 per ml of medium and 11.69 ng of Cyt1A1 per ml of medium. When mixtures of these toxins in different proportions were assayed, eight different LC₅₀s were obtained. All of these LC₅₀s were significantly higher than the expected LC₅₀s of the mixtures. In addition, a series of bioassays were performed with late first-instar larvae of the cabbage looper and pure Cry1Ac1 and Cyt1A1 crystals, as well as two different combinations of the two toxins. The estimated mean LC₅₀ of Cry1Ac1 was 2.46 ng/cm² of diet, while Cyt1A1 crystals exhibited no toxicity, even at very high concentrations. The estimated mean LC₅₀s of Cry1Ac1 crystals were 15.69 and 19.05 ng per cm² of diet when these crystals were mixed with 100 and 1,000 ng of Cyt1A1 crystals per cm² of diet, respectively. These results indicate that there is clear antagonism between the two toxins both in vitro and in vivo. Other joint-action analyses corroborated these results. Although this is the second report of antagonism between B. thuringiensis toxins, our evidence is the first evidence of antagonism between toxins from different subspecies of B. thuringiensis (B. thuringiensis subsp. kurstaki and B. thuringiensis subsp. israelensis) detected both in vivo and in vitro. Some possible explanations for this relationship are discussed.     

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.