Detecting immigration by using multilocus genotypes

Immigration is an important force shaping the social structure, evolution, and genetics of populations. A statistical method is presented that uses multilocus genotypes to identify individuals who are immigrants, or have recent immigrant ancestry. The method is appropriate for use with allozymes, microsatellites, or restriction fragment length polymorphisms (RFLPs) and assumes linkage equilibrium among loci. Potential applications include studies of dispersal among natural populations of animals and plants, human evolutionary studies, and typing zoo animals of unknown origin (for use in captive breeding programs). The method is illustrated by analyzing RFLP genotypes in samples of humans from Australian, Japanese, New Guinean, and Senegalese populations. The test has power to detect immigrant ancestors, for these data, up to two generations in the past even though the overall differentiation of allele frequencies among populations is low.     

Functional measurements of [Ca2+] in the endoplasmic reticulum using a herpes virus to deliver targeted aequorin

Nitric oxide (NO) is a free radical gas that is synthesized from L-arginine by NO synthase (NOS). Activation of NMDA, non-NMDA or metabotropic glutamate receptors causes NO formation through NOS activation. From data obtained in experiments performed by microdialysis together with nitrate assay, we have proposed that NO production in the cerebellum following non-NMDA and metabotropic glutamate receptor activation may be independent of NOS activity, while NMDA receptor-mediated NO production depends on its activity. Glial cells appear to play a role in modulating NO production by regulating L-arginine availability. Activation of NMDA receptors and the increase in intracellular calcium concentration is a trigger for the long-term potentiation (LTP). NO acts as a retrograde messenger in the hippocampal LTP to enhance glutamate release from presynaptic nerve terminal, in which cyclic GMP may be involved. Behavioral studies demonstrate that NO is involved in some forms of learning and memory. Our studies suggest that NMDA/NO/cyclic GMP signaling plays a role in spatial working memory. Further, it is suggested that NO production in the brain is altered by aging. These results support the hypothesis that NO plays a role in mechanism of synaptic plasticity.     

A bioluminescence assay using Nitrosomonas europaea for rapid and sensitive detection of nitrification inhibitors

An expression vector for the luxAB genes, derived from Vibrio harveyi, was introduced into Nitrosomonas europaea. Although the recombinant strain produced bioluminescence due to the expression of the luxAB genes under normal growing conditions, the intensity of the light emission decreased immediately, in a time-and dose-dependent manner, with the addition of ammonia monooxygenase inhibitors, such as allylthiourea, phenol, and nitrapyrin. When whole cells were challenged with several nitrification inhibitors and toxic compounds, a close relationship was found between the change in the intensity of the light emission and the level of ammonia-oxidizing activity. The response of bioluminescence to the addition of allylthiourea was considerably faster than the change in the ammonia-oxidizing rate, measured as both the O2 uptake and NO2- production rates. The bioluminescence of cells inactivated by ammonia monooxygenase inhibitor was recovered rapidly by the addition of certain substrates for hydroxylamine oxidoreductase. These results suggested that the inhibition of bioluminescence was caused by the immediate decrease of reducing power in the cell due to the inactivation of ammonia monooxygenase, as well as by the destruction of other cellular metabolic pathways. We conclude that the assay system using luminous Nitrosomonas can be applied as a rapid and sensitive detection test for nitrification inhibitors, and it will be used to monitor the nitrification process in wastewater treatment plants.     

Determination of fast proton exchange rates of biomolecules by NMR using water selective diffusion experiments

We present a new water selective pulse sequence allowing rapid determination of exchange rates of labile protons on the millisecond time scale. Using diffusion measurements, exchange rates of resolved protons can be determined without prior knowledge of relaxation parameters in a short overnight experiment. The use of a sensitive, highly selective and easy to implement water excitation scheme allows for its straightforward application to a wide range of biomolecules. The results obtained for the imino proton exchange rates of a 16 bp DNA are in strong agreement with values obtained by the classical approach of two-dimensional exchange spectroscopy.     

A method for detection of hydroxyl radicals in the vicinity of biomolecules using radiation-induced fluorescence of coumarin

A novel method is described to quantitate radiation-induced hydroxyl radicals in the vicinity of biomolecules in aqueous solutions. Coumarin-3-carboxylic acid (CCA) is a non-fluorescent molecule that, upon interaction with radiation in aqueous solution, produces fluorescent products. CCA was derivatized to its succinimidyl ester (SECCA) and coupled to free primary amines of albumin, avidin, histone-H1, polylysine, and an oligonucleotide. When SECCA-biomolecule conjugates were irradiated, the relationship between induced fluorescence and dose was linear in the dose range examined (0.01-10 Gy). The fluorescence excitation spectrum of irradiated SECCA-biomolecule conjugates was very similar to that of 7-hydroxy-SECCA-biomolecule conjugates, indicating the conversion of SECCA to 7-hydroxy-SECCA following irradiation. Control studies in environments that excluded certain radiation-induced water radicals for both the conjugated and unconjugated forms of irradiated SECCA demonstrated that: (1) the induction of fluorescence is mediated by the hydroxyl radical; (2) the presence of oxygen enhances induced fluorescence by a factor of about 1.4, and (3) other primary water radicals and secondary radicals caused by interaction of primary water radicals with biomolecules do not significantly influence the induced fluorescence. The data indicate that the induction of fluorescence on SECCA-biomolecule conjugates records specifically the presence of the hydroxyl radical in the immediate vicinity of the irradiated biomolecule. The method is rapid and sensitive, uses standard instrumentation, and the sample remains available for further studies.     

Polarization-enhanced NMR spectroscopy of biomolecules in frozen solution

Large dynamic nuclear polarization signal enhancements (up to a factor of 100) were obtained in the solid-state magic-angle spinning nuclear magnetic resonance (NMR) spectra of arginine and the protein T4 lysozyme in frozen glycerol-water solutions with the use of dynamic nuclear polarization. Polarization was transferred from the unpaired electrons of nitroxide free radicals to nuclear spins through microwave irradiation near the electron paramagnetic resonance frequency. This approach may be a generally applicable signal enhancement scheme for the high-resolution solid-state NMR spectroscopy of biomolecules.     

Remarkable preservation of biomolecules in ancient radish seeds

Desiccated seeds from a 6th century AD storage vessel recovered from Qasr Ibr?m, Egypt, were examined for the presence of lipids and nucleic acids. A remarkable degree of lipid preservation was discovered, the fatty acid and sterol profiles being very similar to those of modern radish seeds. The only significant differences were hydrolysis of triacylglycerols and depletion of the polyunsaturated fatty acids (C18:2 and C18:3). The delta 13 C values of the principal fatty acids were in the range -25.4 to -29.2/1000, which is congruent with modern radish (C3 seeds) taking account of isotopic shifts caused by recent changes in atmospheric CO2. Deoxyribonucleosides and nucleic acid bases were detected by direct chemical analysis, and polymerase chain reactions gave products with sequences comparable to those from modern radish. The degree of lipid preservation, which was much greater than that reported for other archaeological remains, suggests that the microenvironment within desiccated seeds retards biomolecular decay. The results illustrate the utility of combined lipid-nucleic acid analysis in chemotaxonomic and genotypic studies of archaeobotanical remains.     

Lumazine protein and the excitation mechanism in bacterial bioluminescence

The spectral properties of lumazine protein and mixtures with the intermediates of the bacterial luciferase reaction, are reviewed. Measurements of fluorescence dynamics in particular have been employed with the aim of elucidating the mechanism by which lumazine protein functions in the bioluminescence of the bacteria of the type Photobacterium. The reaction of bacterial luciferase with its substrates produces bioluminescence emission with a spectral maximum at 496 nm. This spectrum is the same as the fluorescence of a luciferase flavin intermediate in the reaction, called the Fluorescent Transient. When lumazine protein is also present in the reaction; however, the bioluminescence emission now corresponds to the fluorescence of lumazine protein, which has a maximum at 475 nm. From measurements of the decay of fluorescence anisotropy of lumazine protein alone and in mixtures with the luciferase fluorescent transient, it is shown that a protein-protein complex is formed and that there is rapid energy transfer between the flavin on the luciferase and the lumazine derivative bound to its protein. An approximate calculation estimates the rate of this energy transfer to be faster than 10(9) s-1, and this would account for the efficient transfer of excitation from the flavin on the associated luciferase in the mixed protein bioluminescence reaction.     

Detecting mild brain impairment using the Luria-Nebraska Neuropsychological Battery.

Demonstrated the validity of the standardized Luria-Nebraska Neuropsychological Battery with 36 pseudoneurologic, borderline, or brain-damaged Ss (average ages 41, 36, and 35 yrs). A correct overall classification rate of 80% was obtained, with relatively low false positive and false negative rates. (4 ref) (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Fluorescence resonance energy transfer in studies of inter-chromophoric distances in biomolecules

Fluorescence resonance energy transfer (FRET) is a technique widely used in studies of interchromophoric distances in biomolecules such as peptides, proteins and nucleic acids. FRET is especially useful in determination of conformational changes caused by a solvent, presence of denaturing agents, diffusion and other external factors. Precision of interchromophoric distances obtained using the FRET technique is comparable with that of low-resolution X-ray diffraction and NMR data. Comparison of FRET results with the crystal structure for several proteins is reviewed. Moreover, the effect of the orientation factor kappa2 value on FRET results and determinants of kappa2 are discussed.     

Utilization of glass HPLC autosampler vials as cuvettes in an iodometric assay of hydroperoxides

Bromide treatment was successful in controlling seizures in an 11-year-old Dachshund with epilepsy and presumptive phenobarbital-associated hepatopathy. Because bromide does not induce liver enzyme activity and does not seem to be hepatotoxic, it can be used to control seizures in dogs with concurrent epilepsy and hepatic disease. In this dog, institution of a special calculolytic diet with high chloride content was associated with a decrease in serum bromide concentrations and the recurrence of seizures. High chloride intake increases the elimination of bromide in dogs, leading to higher dosage requirements for bromide in dogs fed high-chloride diets.     

Regeneration and luminescence of aequorin in Chinese hamster ovary cells transformed with cDNA for apoaequorin

Aequorin, a photoprotein which is regenerated from apoaequorin by incubation with coelenterazine, emits light when it binds Ca2+. The aim of this study was to determine if apoaequorin could be used in adherent mammalian cells for measuring cytosolic Ca2+, and imaging Ca2+, at the single cell level. Chinese hamster ovary (CHO-K1) cells were stably transformed with apoaequorin cDNA and expressed apoaequorin while attached to the culture dishes. Maximal luminescence intensity was obtained when 0.5 x 10(6) cells/ml were grown and incubated with 2.5 microM coelenterazine for 4 hr at 20 degrees C. Ca2+ mobilizing agents (ionomycin and maitotoxin) induced luminescence in CHO-K1 transformed cells. However, imaging of light emission from single cells proved to be unsuccessful. Ca2+ could be readily measured in the adherent CHO-K1 cells, but imaging was not possible at the single cell level.     

Detecting undetected HIV-1 variants in African children using degenerate polymerase chain reaction and sequence analyses

CONTEXT: Scientific journals issue press releases to disseminate scientific news about articles they publish. OBJECTIVE: To assess whether press releases about journal articles were associated with publication of subsequent newspaper stories. DESIGN: Retrospective content analysis of newspaper stories, journal press releases, and journal tables of contents. From December 1, 1996, to February 28, 1997, press releases and tables of contents were collected from BMJ, Nature, Science, and The Lancet, along with newspaper stories on scientific research published in The New York Times (United States), Le Figaro and Le Monde (France), El País and La Vanguardia (Spain), La Repubblica (Italy), and the International Herald Tribune. MAIN OUTCOME MEASUREMENTS: Number of newspaper stories that contained reference to articles appearing in the 4 scientific journals, number of newspaper stories that referred to journal articles described in press releases, and the order in which journal articles were mentioned in press releases. RESULTS: Of the 1060 newspaper stories analyzed, 142 referred to journal articles; of these, 119 (84%) referred to articles mentioned in press releases and 23 (16%) referred to journal articles not mentioned in press releases (comparison of proportions, P=.03). Articles described first or second were referenced in more newspapers than articles described later in the press release (P=.01 by chi2 analysis). CONCLUSIONS: Journal articles described in press releases, in particular those described first or second in the press release, are associated with the subsequent publication of newspaper stories on the same topic.     

Effect of centrifuging shell vials at 3,500 x g on detection of viruses in clinical specimens

An increase in shell vial centrifugation force to 3,500 x g and a concomitant reduction in spin time to 15 min did not decrease the sensitivity of detecting viruses in clinical specimens compared with the accepted practice of using 700 x g for 40 min. No damage to the cell monolayer (ML) at the higher g force was observed. Toxicity to the ML is decreased with the shorter spin, probably because of reduced time of contact between the specimen and the ML.     

Measurement of intracellular calcium in cell populations loaded with aequorin: neurokinin-1 responses in U373MG cells

Changes in intracellular calcium concentration are important in mediating a wide variety of physiological responses. Recently there has been renewed interest in the use of aequorin, a protein from jellyfish that emits light when calcium is bound, to measure calcium levels in cells. We have loaded populations of cells from the human glioma line, U373MG, with aequorin. Lysis of aequorin-loaded but not control cells with detergent resulted in a luminescence signal that was dependent on extracellular calcium. Aequorin-loaded cells responded to substance P, histamine, or the calcium ionophore, ionomycin, with an increase in luminescence. Signals in response to detergent, ionomycin, or substance P could be detected up to 48 h after cells were loaded with aequorin. Other neurokinin-1 agonists but not agonists at neurokinin-2 or neurokinin-3 receptors produced luminescence signals. Neurokinin-1 antagonists inhibited the substance P-induced signal. The aequorin-loading procedure worked well with U373MG cells but not with AR42J, CHO, IMR-90, or WI-38 cells.     

Detecting cyclicity in social interaction.

Reviews spectral and cross-spectral analytic methods for detecting cyclicity, cross-cyclicity, and lead–lag relationships in continuous data derived from the observation of dyadic interaction. It is found that lead–lag relationships can be assessed using the phase spectrum. Spectral analytic methods are then generalized to categorical observational data (taken from a study by the author and his associates; see record 1978-20895-001). It is shown that by these methods one can derive the classical information theory definition of social communication and its distribution statistics. (27 ref) (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Ca2+-induced Ca2+ release in chromaffin cells seen from inside the ER with targeted aequorin

The presence and physiological role of Ca2+-induced Ca2+ release (CICR) in nonmuscle excitable cells has been investigated only indirectly through measurements of cytosolic [Ca2+] ([Ca2+]c). Using targeted aequorin, we have directly monitored [Ca2+] changes inside the ER ([Ca2+]ER) in bovine adrenal chromaffin cells. Ca2+ entry induced by cell depolarization triggered a transient Ca2+ release from the ER that was highly dependent on [Ca2+]ER and sensitized by low concentrations of caffeine. Caffeine-induced Ca2+ release was quantal in nature due to modulation by [Ca2+]ER. Whereas caffeine released essentially all the Ca2+ from the ER, inositol 1,4, 5-trisphosphate (InsP3)- producing agonists released only 60-80%. Both InsP3 and caffeine emptied completely the ER in digitonin-permeabilized cells whereas cyclic ADP-ribose had no effect. Ryanodine induced permanent emptying of the Ca2+ stores in a use-dependent manner after activation by caffeine. Fast confocal [Ca2+]c measurements showed that the wave of [Ca2+]c induced by 100-ms depolarizing pulses in voltage-clamped cells was delayed and reduced in intensity in ryanodine-treated cells. Our results indicate that the ER of chromaffin cells behaves mostly as a single homogeneous thapsigargin-sensitive Ca2+ pool that can release Ca2+ both via InsP3 receptors or CICR.