Conspecific "gaze following" in bottlenose dolphins.

"Gaze following"-when one individual witnesses another shift its orientation, and then re-orients in the same direction-has been observed in a wide range of species. Related work with dolphins has to date focused on human-dolphin interactions. In this conspecific study, we examined a group of dolphins orienting, in passing, to gateways between their pools, as opportunities for witnesses to demonstrate "gaze following". Seven bottlenose dolphins were synchronously videotaped on six underwater cameras, for 21 h over three days, and the recordings analyzed by trained observers. The identities of all animals present, their partner state, and whether and to what degree they had altered their access to the gate (e.g., from Monocular to Binocular, or Binocular to Visio-Echoic) was recorded. Compared to animals that did not witness such a change, witnesses of an increase in access by another dolphin were significantly more likely to also act to increase their own access. We observed 460 such cases of "gaze following" in these animals. Dolphins who were partnered (showed sustained swimming within 1 body length) were significantly more likely, than non-partnered animals, to "gaze follow". Dolphins also showed a significant tendency toward matching the kind of access they observed. No significant difference was found in the presence of animals in the back pools, during changes in orientation that were followed, versus in those that were not. These findings support adding bottlenose dolphins to the growing list of species that display conspecific "gaze following".

Possible impacts of molten salt reactors on the International Monitoring System.

Molten salt reactors (MSRs) are gaining support as many countries look for ways to increase power generation and replace aging nuclear energy production facilities. MSRs have inherently safe designs, are scalable in size, can burn transuranic wastes from traditional solid fuel nuclear reactors, can store excess heat in thermal reservoirs for water desalination, and can be used to produce medical isotopes as part of the real-time liquid-fuel recycling process. The ability to remove 135Xe in real time from the fuel improves the power production in an MSR because 135Xe is the most significant neutron-absorbing isotope generated by nuclear fission. Xenon-135, and other radioactive gases, are removed by sparging the fuel with an inert gas while the liquid fuel is recirculated from the reactor inner core through the heat exchangers. Without effective abatement technologies, large amounts of radioactive gas could be released during the sparging process. This work examines the potential impact of radioxenon releases on samplers used by the International Monitoring System (IMS) to detect nuclear explosions. Atmospheric transport simulations from seven hypothetical MSRs on different continents were used to evaluate the holdup time needed before release of radioxenon so IMS samplers would register few detections. Abatement technologies that retain radioxenon isotopes for at least 120 d before their release will be needed to mitigate the impacts from a molten salt breeder reactor used to replace a nuclear power plant. A holdup time of about 150 d is needed to reduce emissions to the average level of current nuclear power plants.

The Evolving Landscape of Institutional Biosafety Committees and Biosafety Programs: Results from a National Survey on Organizational Structure, Resources, and Practices.

Introduction: There are vast differences in the size, scope, and needs of institutions that conduct research involving biohazardous materials, thus resulting in vast differences among Institutional Biosafety Committees (IBCs) and biosafety programs. Methods: A benchmarking survey of IBC and biosafety programs was conducted in an effort to identify common practices in the field and compare this information with that of the other institutional bioethics committees, namely, Institutional Animal Care and Use Committees (IACUCs) and Institutional Review Boards (IRBs). Objectives: The primary objectives of the survey were to assess the organizational structure of IBC and biosafety programs, determine the scope of IBC review, and compare the size of IBC and biosafety programs with that of IACUCs and IRBs. Results: The survey results showed that IBCs most commonly reside under the same administrative unit as the IACUC and IRB, while the majority of institutions' biosafety officers report to a different unit. The majority of respondents indicated their IBC reviews research utilizing biological hazards beyond what is required by the National Institutes of Health Guidelines. The survey data suggest that IBCs have fewer support staff than the other bioethics committees; 57% of institutions report one or more full-time employee (FTE) dedicated to support the IBC, compared to 86%, 85%, and 83% of institutions that reported one or more FTE to support the IACUC, the IRB, and the biosafety program, respectively. Conclusion: Data from the survey identified common practices among IBCs and provides institutions a tool to compare their program with others.

Prosocial Predictions by Bottlenose Dolphins ( Tursiops spp.) Based on Motion Patterns in Visual Stimuli.

In this study, paradigms that test whether human infants make social attributions to simple moving shapes were adapted for use with bottlenose dolphins. The dolphins observed animated displays in which a target oval would falter while moving upward, and then either a "prosocial" oval would enter and help or caress it or an "antisocial" oval would enter and hinder or hit it. In subsequent displays involving all three shapes, when the pro- and antisocial ovals moved offscreen in opposite directions, the dolphins reliably predicted-based on anticipatory head turns when the target briefly moved behind an occluder-that the target oval would follow the prosocial one. When the roles of the pro- and antisocial ovals were reversed toward a new target, the animals' continued success suggests that such attributions may be dyad specific. Some of the dolphins also directed high arousal behaviors toward these displays, further supporting that they were socially interpreted.

Sensorimotor Decoupling Contributes to Triadic Attention: A Longitudinal Investigation of Mother-Infant-Object Interactions.

Previous developmental accounts of joint object activity identify a qualitative "shift" around 9-12 months. In a longitudinal study of 26 dyads, videos of joint object interactions at 4, 6, 9, and 12 months were coded for all targets of gaze and manual activity (at 10 Hz). At 12 months, infants distribute their sensorimotor modalities between objects handled by the parent and others controlled by the infant. Analyses reveal novel trajectories in distributed joint object activity across the 1st year. At 4 months, infants predominantly look at and manipulate a single object, typically held by their mothers. Between 6 and 9 months, infants increasingly decouple their visual and haptic modalities and distribute their attention between objects held by their mothers and by themselves. These previously unreported developments in the distribution of multimodal object activity might "bridge the gap" to coordinated joint activity between 6 and 12 months.

Visible and invisible displacement with dynamic visual occlusion in bottlenose dolphins (Tursiops spp).

Anticipating the location of a temporarily obscured target-what Piaget (the construction of reality in the child. Basic Books, New York, 1954) called "object permanence"-is a critical skill, especially in hunters of mobile prey. Previous research with bottlenose dolphins found they could predict the location of a target that had been visibly displaced into an opaque container, but not one that was first placed in an opaque container and then invisibly displaced to another container. We tested whether, by altering the task to involve occlusion rather than containment, these animals could show more advanced object permanence skills. We projected dynamic visual displays at an underwater-viewing window and videotaped the animals' head moves while observing these displays. In Experiment 1, the animals observed a small black disk moving behind occluders that shifted in size, ultimately forming one large occluder. Nine out of ten subjects "tracked" the presumed movement of the disk behind this occluder on their first trial-and in a statistically significant number of subsequent trials-confirming their visible displacement abilities. In Experiment 2, we tested their invisible displacement abilities. The disk first disappeared behind a pair of moving occluders, which then moved behind a stationary occluder. The moving occluders then reappeared and separated, revealing that the disk was no longer behind them. The subjects subsequently looked to the correct stationary occluder on eight of their ten first trials, and in a statistically significant number of subsequent trials. Thus, by altering the stimuli to be more ecologically valid, we were able to show that the dolphins could indeed succeed at an invisible displacement task.

IVMBIX-01294, an inhibitor of the histone methyltransferase EHMT2, disrupts histone H3 lysine 9 (H3K9) dimethylation in the cleavage-stage porcine embryo.

Global patterns of histone methylation are remodelled during cleavage development. Of the five histone methyltransferases known to mediate methylation of the lysine 9 residue of histone H3 (H3K9), euchromatic histone-lysine N-methyltransferase 2 (EHMT2; also known as G9a) has been shown to be a primary mediator of H3K9 dimethylation; BIX-01294 has been shown to be a specific inhibitor of EHMT2. The objective of the present study was to determine the effect of BIX-01294 treatment on global H3K9 dimethylation in porcine embryos. We hypothesised that inhibition of EHMT2 by BIX-01294 would result in reduced levels of H3K9 dimethylation and compromised embryo development. Our results showed that incubation in 5µM BIX-01294 markedly reduced global levels of H3K9 dimethylation at the pronuclear, 2-cell and 4-cell stages of development and resulted in developmental arrest before blastocyst formation. Although transient exposure of embryos to BIX-01294 did not alter in vitro development, embryos transiently exposed to BIX-01294 did not establish pregnancy. These data demonstrate that BIX-01294 is a potent inhibitor of H3K9 dimethylation and that transient alterations in global histone modifications can have profound effects on embryo developmental potential.

Differential developmental requirements for individual histone H3K9 methyltransferases in cleavage-stage porcine embryos.

Dimethylated H3K9 is a heritable epigenetic mark that is closely linked with transcriptional silencing and known to undergo global remodelling during cleavage development. Five mammalian histone methyltransferases (HMTases), namely Suv39H1, Suv39H2, SetDB1, EHMT1 and EHMT2, have been shown to mediate the methylation of H3K9. The aim of the present study was to determine the developmental requirements of these HMTases during cleavage development in porcine embryos. We hypothesised that knockdown of the abovementioned HMTases would differentially affect porcine cleavage development. To test this hypothesis, IVM and IVF porcine oocytes were divided into one of three treatment groups, including non-injected controls, oocytes injected with a double-stranded interfering RNA molecule specific for one of the HMTases and oocytes injected with a corresponding mutated (control) double-stranded RNA molecule. Nuclei were counted in all embryos 6 days after fertilisation. Although no significant difference in total cell number was detected in embryos injected with EHMT1 and EHMT2 interfering RNAs (compared with their respective control groups), embryos injected with interfering RNAs that targeted Suv39H1, Suv39H2 and SetDB1had significantly lower cell numbers than their respective control groups (P<0.05). This suggests that individual HMTases differentially affect in vitro developmental potential.

Global H3K9 dimethylation status is not affected by transcription, translation, or DNA replication in porcine zygotes.

Methylation of the lysine 9 residue of histone H3 (H3K9) is linked to transcriptional repression. The observed structure of chromatin in porcine and murine embryos is different with regard to H3K9 dimethylation status, leading to our hypothesis that the intracellular mechanisms responsible for H3K9 methylation would also differ between these two species. The objectives of this study were: (1) to determine the extent that DNA, mRNA, and protein synthesis serve in maintaining the asymmetrical distribution of dimethylated H3K9 in porcine zygotes, (2) determine the extent to which the intracellular localization of individual pronuclei correlated with H3K9 dimethylation status, and (3) to determine the abundance of transcripts encoding the histone methyltransferases, with H3K9 methylation activity, in porcine oocytes and embryos. Our findings are that (1) H3K9 dimethylation status is not affected by DNA replication, transcription, or protein synthesis, (2) the location of a pronucleus does not significantly affect the H3K9 dimethylation status of the chromatin within that pronucleus, and (3) the histone methyltransferases with activity for H3K9 differ in transcript abundance in porcine oocytes and cleavage stage embyros. These results support our hypothesis that there is a difference in intracellular mechanisms affecting dimethylation status of H3K9 between porcine and murine embryos.

Cholesterol control beyond the clinic: New York City's trans fat restriction.

Decades after key modifiable risk factors were identified, cardiovascular disease remains the leading cause of preventable death, and only one quarter of persons with high cholesterol levels have attained recommended levels of control. Cholesterol control efforts have focused on consumer education and medical treatment. A powerful, complementary approach is to change the makeup of food, a route the New York City Department of Health and Mental Hygiene took when it restricted artificial trans fat--a contributor to coronary heart disease--in restaurants. The Department first undertook a voluntary campaign, but this effort did not decrease the proportion of restaurants that used artificial trans fat. In December 2006, the Board of Health required that artificial trans fat be phased out of restaurant food. To support implementation, the Department provided technical assistance to restaurants. By November 2008, the restriction was in full effect in all New York City restaurants and estimated restaurant use of artificial trans fat for frying, baking, or cooking or in spreads had decreased from 50% to less than 2%. Preliminary analyses suggest that replacement of artificial trans fat has resulted in products with more healthful fatty acid profiles. For example, in major restaurant chains, total saturated fat plus trans fat in French fries decreased by more than 50%. At 2 years, dozens of national chains had removed artificial trans fat, and 13 jurisdictions, including California, had adopted similar laws. Public health efforts that change food content to make default choices healthier enable consumers to more successfully meet dietary recommendations and reduce their cardiovascular risk.

Modeling the emergence of language as an embodied collective cognitive activity.

Two decades of attempts to model the emergence of language as a collective cognitive activity have demonstrated a number of principles that might have been part of the historical process that led to language. Several models have demonstrated the emergence of structure in a symbolic medium, but none has demonstrated the emergence of the capacity for symbolic representation. The current shift in cognitive science toward theoretical frameworks based on embodiment is already furnishing computational models with additional mechanisms relevant to the emergence of symbolic language. An analysis of embodied interaction among captive, but not human-enculturated, bonobo chimpanzees reveals a number of additional features of embodiment that are relevant to the emergence of symbolic language, but that have not yet been explored in computational simulation models; for example, complementarity of action in addition to imitation, iconic in addition to indexical gesture, coordination among multiple sensory and perceptual modalities, and the orchestration of intra- and inter-individual motor coordination. The bonobos provide an evolutionarily plausible intermediate stage in the development of symbolic expression that can inform efforts to model the emergence of symbolic language.

Expression of eukaryotic elongation initiation factor 1A differentially marks zygotic genome activation in biparental and parthenogenetic porcine embryos and correlates with in vitro developmental potential.

Zygotic genome activation (ZGA) is a major event during cleavage development. In vitro manipulation of mammalian embryos (including embryo culture) can result in developmental arrest around the time of ZGA. Eukaryotic elongation initiation factor 1A (eIF1A) has been used as a marker for ZGA in some mammalian species. We hypothesised expression of eIF1A can be used to assess ZGA in the pig; we also hypothesised that the expression profile of eIF1A can be used to assess developmental potential in vitro. The aims of the present study were to determine the expression pattern of eIF1A during porcine cleavage development and to assess its expression levels in embryos of different quality. We used a real-time reverse transcription-polymerase chain reaction assay to quantify eIF1A transcripts at different time points during cleavage development in porcine embryos produced by parthenogenetic activation (PA) and in vitro fertilisation (IVF). We found that eIF1A is activated at the two-cell stage in IVF embryos and at the four-cell stage in PA embryos. We showed that the increase in transcript levels observed in parthenogenetic embryos is dependent on de novo transcription. We found altered levels of eIF1A transcripts in parthenogenetic embryos that presented as either two- or eight-cell embryos 48 h after activation compared with four-cell embryos at the same time point. Our work supports the hypothesis that eIF1A is a marker of porcine ZGA and its expression profile can be used to assess embryo quality.

Evaluation of a 2-pyridone, KRQ-10018, against Mycobacterium tuberculosis in vitro and in vivo.

A novel subclass of quinolones, 2-pyridones, showed potent activity against Mycobacterium tuberculosis, with KRQ-10018 being an early lead. KRQ-10018 showed better activity in vitro against M. tuberculosis versus moxifloxacin. In vivo efficacy of KRQ-10018 at 300 mg/kg of body weight was similar to that of isoniazid at 25 mg/kg, but showed less activity than moxifloxacin at 300 mg/kg.

Rapid microbiologic and pharmacologic evaluation of experimental compounds against Mycobacterium tuberculosis.

The assessment of physiochemical and pharmacological properties at early stages of drug discovery can accelerate the conversion of hits and leads into candidates for further development. A strategy for streamlined evaluation of compounds against Mycobacterium tuberculosis in the early preclinical stage is presented in this report. As a primary assay to rapidly select experimental compounds with sufficient in vitro activity, the growth inhibition microtiter plate assay was devised as an alternative to current methods. This microdilution plate assay is a liquid culture method based on spectrophotometric readings of the bacillary growth. The performance of this method was compared to the performance of two established susceptibility methods using clinical available tuberculosis (TB) drugs. Data generated from all three assays were similar for all of the tested compounds. A second simple bioassay was devised to assess the oral bioavailability of compounds prior to extensive in vivo efficacy testing. The bioassay estimates drug concentrations in collected serum samples by a microdilution MIC plate method using M. tuberculosis. In the same assay, the MIC of the compound is also determined in the presence of 10% mouse serum as an indication of protein binding. The method was validated using different clinically available TB drugs, and results are discussed in this report. With these methodological advances, screening of compounds against tuberculosis in the preclinical phase will be rapid, can be adapted to semi-high-throughput screening, and will add relevant physicochemical and basic pharmacological criteria to the decision process of drug discovery.

Oral therapy using nanoparticle-encapsulated antituberculosis drugs in guinea pigs infected with Mycobacterium tuberculosis.

We evaluated the efficacy of nanoparticle-encapsulated antituberculosis drugs administered every 10 days versus that of daily nonencapsulated drugs against Mycobacterium tuberculosis aerosol infection in guinea pigs. Both treatments significantly reduced the bacterial count and lung histopathology, suggesting that the nanoparticle drug delivery system has potential in intermitted treatment of tuberculosis.

Significant increases in the levels of liver enzymes in mice treated with anti-tuberculosis drugs.

Besides the long-term effectiveness of a given compound, safety is a very important feature to consider when developing new compounds for chemotherapy against tuberculosis. Reports of fatal and severe liver injury associated with rifampicin-pyrazinamide (RIF-PZA) treatment regimens for latent tuberculosis infections prompted this study to evaluate whether a mouse model has any potential as a tool to assess liver injury following extensive exposure to tuberculosis drugs. Mice were administered high doses of existing drug regimens for latent tuberculosis over a relatively short time period. Alanine aminotransferase (ALT), aspartate aminotransferase and bilirubin levels were determined after 2 weeks and 4 weeks of treatment in serum samples collected from uninfected mice as well as mice infected with Mycobacterium tuberculosis. ALT levels increased significantly after a RIF-PZA treatment regimen for 4 weeks in uninfected mice and after 2 weeks in infected mice. Bilirubin serum levels were also significantly elevated in the M. tuberculosis-infected mice after 4 weeks of RIF-PZA treatment. The data obtained indicate that changes in serum enzyme levels in mice after extensive exposure to tuberculosis drugs could be useful as an initial indicator of drug-related hepatotoxicity.

Preclinical testing of the nitroimidazopyran PA-824 for activity against Mycobacterium tuberculosis in a series of in vitro and in vivo models.

This study extends earlier reports regarding the in vitro and in vivo efficacies of the nitroimidazopyran PA-824 against Mycobacterium tuberculosis. PA-824 was tested in vitro against a broad panel of multidrug-resistant clinical isolates and was found to be highly active against all isolates (MIC<1 microg/ml). The activity of PA-824 against M. tuberculosis was also assessed grown under conditions of oxygen depletion. PA-824 showed significant activity at 2, 10, and 50 microg/ml, similar to that of metronidazole, in a dose-dependent manner. In a short-course mouse infection model, the efficacy of PA-824 at 50, 100, and 300 mg/kg of body weight formulated in methylcellulose or cyclodextrin/lecithin after nine oral treatments was compared with those of isoniazid, rifampin, and moxifloxacin. PA-824 at 100 mg/kg in cyclodextrin/lecithin was as active as moxifloxacin at 100 mg/kg and isoniazid at 25 mg/kg and was slightly more active than rifampin at 20 mg/kg. Long-term treatment with PA-824 at 100 mg/kg in cyclodextrin/lecithin reduced the bacterial load below 500 CFU in the lungs and spleen. No significant differences in activity between PA-824 and the other single drug treatments tested (isoniazid at 25 mg/kg, rifampin at 10 mg/kg, gatifloxacin at 100 mg/kg, and moxifloxacin at 100 mg/kg) could be observed. In summary, its good activity in in vivo models, as well as its activity against multidrug-resistant M. tuberculosis and against M. tuberculosis isolates in a potentially latent state, makes PA-824 an attractive drug candidate for the therapy of tuberculosis. These data indicate that there is significant potential for effective oral delivery of PA-824 for the treatment of tuberculosis.

Selection of HPRT primers as controls for determination of mRNA expression in dogs by RT-PCR.

Reliable housekeeping gene controls are critical for measuring and comparing gene expression at the transcription level by Northern blot and RT-PCR. In order to develop such controls for studying cytokine mRNA expression in dogs, DNA sequence encoding a full-length canine HPRT protein has been obtained. Numerous primer pairs derived from the canine HPRT sequence have been tested on canine genomic DNA as well as cDNA. The data from the present study suggest that there may be processed HPRT pseudogenes in dogs. Three pairs of canine HPRT primers designed and tested in the present study were able to differentiate between cDNA and genomic DNA under specific PCR conditions. These primers would be useful controls for measurement of mRNA expression by RT-PCR in the dog.

Intralesional lipid-complexed cytokine/superantigen immunogene therapy for spontaneous canine tumors.

These studies sought to determine the gene expression and short-term effects of intralesional lipid-complexed immunogene therapy with constructs encoding Staphylococcus aureus enterotoxin A and canine interleukin-2 (L-SEA/cIL-2) in dogs with tumors of various histotypes, and then to assess the safety and efficacy of repeated L-SEA/cIL-2 injections in dogs with spontaneous soft tissue sarcomas (STS). In the first study, pet dogs with a variety of tumors received a single intralesional injection of L-SEA/cIL-2, and surgical excision was performed 48 h later. In the second study, dogs with histologically confirmed STS were treated weekly for a maximum of 12 weeks with escalating doses of L-SEA/cIL-2. Tumors were then surgically excised and assessed histologically and immunohistochemically. Overall, treatments were well tolerated, with no dose-limiting toxicities encountered. At 48 h, in the single injection study, plasmid DNA was detected in 14 of 16 tumor samples, and plasmid-specific mRNA was detected in 3 of 14. In the multiple injection study, the overall response rate in dogs with STS was 25%, consisting of 3 complete responses (CR) and 1 partial response (PR). Diffuse lymphoplasmacytic inflammation was observed in all tumors from patients experiencing CR or PR, whereas these changes were not evident in tumors from nonresponders. The infiltrate was composed primarily of CD3(+) cells at 48 h from the single-injection study, and was composed of both CD3(+) and CD79a(+) cells at 12 weeks in responding dogs from the multiple-injection study. In conclusion, these studies suggests that intralesional L-SEA/cIL-2 immunotherapy is well tolerated, results in detectable transgene expression in canine tumors, and has antitumor activity in dogs with spontaneous STS.