Genomic architecture of phenotypic extremes in a wild cervid.

Identifying the genes underlying fitness-related traits such as body size and male ornamentation can provide tools for conservation and management and are often subject to various selective pressures. Here we performed high-depth whole genome re-sequencing of pools of individuals representing the phenotypic extremes for antler and body size in white-tailed deer (Odocoileus virginianus). Samples were selected from a tissue repository containing phenotypic data for 4,466 male white-tailed deer from Anticosti Island, Quebec, with four pools representing the extreme phenotypes for antler and body size after controlling for age. Our results revealed a largely homogenous population but detected highly divergent windows between pools for both traits, with the mean allele frequency difference of 14% for and 13% for antler and body SNPs in outlier windows, respectively. Genes in outlier antler windows were enriched for pathways associated with cell death and protein metabolism and some of the most differentiated windows included genes associated with oncogenic pathways and reproduction, processes consistent with antler evolution and growth. Genes associated with body size were more nuanced, suggestive of a highly complex trait. Overall, this study revealed the complex genomic make-up of both antler morphology and body size in free-ranging white-tailed deer and identified target loci for additional analyses.

Demographic histories and genetic diversity across pinnipeds are shaped by human exploitation, ecology and life-history.

A central paradigm in conservation biology is that population bottlenecks reduce genetic diversity and population viability. In an era of biodiversity loss and climate change, understanding the determinants and consequences of bottlenecks is therefore an important challenge. However, as most studies focus on single species, the multitude of potential drivers and the consequences of bottlenecks remain elusive. Here, we combined genetic data from over 11,000 individuals of 30 pinniped species with demographic, ecological and life history data to evaluate the consequences of commercial exploitation by 18th and 19th century sealers. We show that around one third of these species exhibit strong signatures of recent population declines. Bottleneck strength is associated with breeding habitat and mating system variation, and together with global abundance explains much of the variation in genetic diversity across species. Overall, bottleneck intensity is unrelated to IUCN status, although the three most heavily bottlenecked species are endangered. Our study reveals an unforeseen interplay between human exploitation, animal biology, demographic declines and genetic diversity.

Measurement accuracy of heart rate and respiratory rate during graded exercise and sustained exercise in the heat using the Zephyr BioHarness.

The Zephyr BioHarness was tested to determine the accuracy of heart rate (HR) and respiratory rate (RR) measurements during 2 exercise protocols in conjunction with either a laboratory metabolic cart (Vmax) or a previously validated portable metabolic system (K4b2). In one protocol, HR and RR were measured using the BioHarness and Vmax during a graded exercise up to V˙O2max (n=12). In another protocol, HR and RR were measured using the BH and K4b2 during sustained exercise (30% and 50% V˙O2max for 20 min each) in a hot environment (30 °C, 50% relative humidity) (n=6). During the graded exercise, HR but not RR, obtained from the BioHarness was higher compared to the Vmax at baseline and 30% V˙O2max (p<0.05), but showed no significant difference at other stages with high correlation coefficients for both HR (r=0.87-0.96) and RR (r=0.90-0.99 above 30% V˙O2max). During the exercise in the heat, there were no significant differences between the BioHarness and K4b2 system. Correlation coefficients between the methods were low for HR but moderately to highly correlated (0.49-0.99) for RR. In conclusion, the BioHarness is comparable to Vmax and K4b2 over a wide range of V˙O2 during graded exercise and sustained exercise in the heat.

Genome-wide cross-amplification of domestic sheep microsatellites in bighorn sheep and mountain goats.

We tested for cross-species amplification of microsatellite loci located throughout the domestic sheep (Ovis aries) genome in two north American mountain ungulates (bighorn sheep, Ovis canadensis, and mountain goats, Oreamnos americanus). We identified 247 new polymorphic markers in bighorn sheep (≥ 3 alleles in one of two study populations) and 149 in mountain goats (≥ 2 alleles in a single study population) using 648 and 576 primer pairs, respectively. Our efforts increased the number of available polymorphic microsatellite markers to 327 for bighorn sheep and 180 for mountain goats. The average distance between successive polymorphic bighorn sheep and mountain goat markers inferred from the Australian domestic sheep genome linkage map (mean ± 1 SD) was 11.9 ± 9.2 and 15.8 ± 13.8 centimorgans, respectively. The development of genomic resources in these wildlife species enables future studies of the genetic architecture of trait variation.

The effects of deafferentation and exogenous NGF on neurotrophins and neurotrophin receptor mRNA expression in the adult superior cervical ganglion.

Levels of nerve growth factor (NGF) and neurotrophin-3 (NT-3) protein and neurotrophin receptor mRNA in adult sympathetic neurons were investigated following surgical removal of preganglionic input and/or in vivo administration of NGF. Expression of trkC and p75, but not trkA, was significantly decreased following a 3-week deafferentation of the superior cervical ganglion (SCG). Protein levels of NGF and NT-3 in the SCG were unchanged by deafferentation. A 2-week intracerebroventricular infusion of NGF without deafferentation resulted in enhanced mRNA levels of trkA, trkC, and p75 as well as significantly increased NGF and NT-3 protein in the SCG. When NGF infusion followed deafferentation, both trkA and p75 showed significant increases while trkC levels were similar to control values. NGF protein was not increased in the SCG when deafferentation preceded exogenous NGF, yet NT-3 was elevated and levels were similar to cases receiving NGF infusion only. These results support a role for preganglionic input in trkC and p75 expression in adult sympathetic neurons. The increased levels of NT-3 protein and trkC gene expression observed following NGF infusion suggest that NGF influences NT-3 regulation in adult sympathetic neurons. In addition, the present findings provide evidence that, when preganglionic input is removed prior to the NGF infusion, NT-3 effectively competes with NGF for trkA binding. Taken together, we propose that NT-3 may play a role in the robust sprouting of sympathetic cerebrovascular axons previously observed following NGF administration, particularly when deafferentation precedes the NGF infusion period.

Researching medicine in context: the Arts and Humanities Medical Scholars Program.

In 2000, the Arts and Humanities Medical Scholars Program at Stanford University School of Medicine issued its first grants to medical students interested in researching an area of the medical arts or humanities in depth. To date, 34 projects have been funded, including renewals. The projects encompass a range of genres and topics, from a website on Asian American health and culture to an ethnographic study of women physicians in training in Spain. Two projects are highlighted here: an online history of medicine course and a poetry project. Students are mentored by faculty from a wide array of university departments and centres and submit completion documents to the committee overseeing the programme. Students are encouraged to present their work at conferences, such as the programme's annual symposium, as well as in publication or other appropriate formats. Future directions include integration with the scholarly concentrations initiative at the medical school.

Patient threats present an ethical dilemma for the anesthesiologist.

IMPLICATIONS: Patients who receive sedation occasionally divulge thoughts that they would not usually express. This report describes a sedated patient who threatened to murder two family members. Immediate consultation with an attorney and psychiatrist is recommended when the anesthesiologist may be required to breach patient confidentiality to warn potential victims.

Remodeling of adult sensory axons in the superior cervical ganglion in response to exogenous nerve growth factor.

In previous studies, we found that a 2-week in vivo intracerebroventricular infusion of nerve growth factor (NGF) elicited a sprouting response by sympathetic perivascular axons associated with the intradural segment of the internal carotid artery. We hypothesized that NGF infused into the ventricular system would be internalized by responsive sympathetic cerebrovascular axons, retrogradely transported to parent cell bodies in the superior cervical ganglion (SCG), and subsequently released into the local ganglionic environment. Because fibers exhibiting immunoreactivity for calcitonin gene related peptide (CGRP) have been localized in the SCG, we used immunohistochemical methods to investigate whether a response by CGRP-immunoreactive axons in the SCG occurred following the proposed transport to and release of exogenous NGF in the ganglion. In consecutive tissue sections of the SCG stained for either CGRP or NGF, we found CGRP pericellular 'baskets' surrounding identified NGF-immunoreactive cell bodies. Nerve growth factor infusion resulted in a significant increase both in the number of CGRP pericellular baskets and in NGF-immunoreactive cell bodies. A significant positive correlation (r=0.95, P<0.05) between the pericellular baskets and NGF-immunoreactive cell bodies was observed, suggesting that intracranial projection neurons in the SCG released infused NGF (or possibly a converted signal) into the local ganglionic environment to elicit remodeling of CGRP fibers to form pericellular baskets. These findings were confirmed in sections double labeled for NGF and CGRP immunoreactivity. This remodeling suggests that exogenous NGF may mediate retrograde transneuronal plasticity, allowing for future in vivo examinations of the mechanisms involved in neurotrophin transport and release.

Tetracycline-regulatable expression vectors tightly regulate in vitro gene expression of secreted proteins.

The regulation of gene expression by the tetracycline system has attracted a high level of interest in the recent past. However, expression of secreted proteins has not been evaluated precisely. In this study, we constructed two versions of a one-plasmid system containing the elements necessary for the regulation of gene expression. The regulatable elements and the selectable marker (Neor) were set up in two different configurations, pTRIN31 and pTRIN76. With these two regulatable versions, the levels of protein expression after transfection into the NIH/3T3 cell line were measured by insertion of three different genes encoding the secreted proteins (hGH, ApoE3, hGM-CSF). The maximum levels of gene expression obtained with the pTRIN76-derived plasmids were 100ng/24h/106 cells for hGH, 427ng/24h/106 cells for ApoE3 and 108ng/24h/106 cells for hGM-CSF. For the pTRIN31-derived plasmids the maximum levels were 2.7ng/24h/106 cells for hGH and 47ng/24h/106 for ApoE3. Both plasmids give rise to an expression of the transfected gene that can be tightly regulated by three different molecules: tetracycline, minocycline and doxycycline. The levels of the secreted proteins are below the detectable level when the reporter genes are repressed. This repression is reversible within 48h after the regulator has been removed from the medium.

Complications of sedation with midazolam in the intensive care unit and a comparison with other sedative regimens.

OBJECTIVE: To describe the various complications that have been reported with use of midazolam for sedation in the intensive care unit (ICU). DATA SOURCES: Publications in scientific literature. DATA EXTRACTION: Computer search of the literature. SYNTHESIS: Sedation is required in the ICU in order for patients to tolerate noxious stimuli, particularly mechanical ventilation. Under- and oversedation can lead to complications. To sedate patients in the ICU, midazolam is commonly administered via titrated, continuous infusions. Cardiorespiratory effects tend to be minimal; however, hypotension can occur in hypovolemic patients. Prolonged sedation after cessation of the midazolam infusion may be caused by altered kinetics of the drug in critically ill patients or by accumulation of active metabolites. In addition, paradoxical and psychotic reactions have been rarely reported. Tolerance and tachyphylaxis may occur, particularly with longer-term infusions (> or = 3 days). Benzodiazepine withdrawal syndrome has also been associated with high dose/long-term midazolam infusions. Compared with propofol infusions, midazolam infusions have been associated with a decreased occurrence of hypotension but a more variable time course for recovery of function after the cessation of the infusion. Lorazepam is a more cost-effective choice for long-term (> 24 hrs) sedation. CONCLUSION: Continuous infusion midazolam provides effective sedation in the ICU with few complications overall, especially when the dose is titrated.

Vancomycin control measures at a tertiary-care hospital: impact of interventions on volume and patterns of use.

OBJECTIVE: Evaluate vancomycin prescribing patterns in a tertiary-care hospital before and after interventions to decrease vancomycin utilization. DESIGN: Before/after analysis of interventions to limit vancomycin use. SETTING: 420-bed academic tertiary-care center. INTERVENTIONS: Educational efforts began August 10, 1994, and involved lectures to medical house staff followed by mailings to all physicians and posting of guidelines for vancomycin use on hospital information systems. Active interventions began November 15, 1994, and included automatic stop orders for vancomycin at 72 hours, alerts attached to the medical record, and, for 2 weeks only, computer alerts to physicians following each vancomycin order. Parenteral vancomycin use was estimated from the hospital pharmacy database of all medication orders. Records of a random sample of 344 patients receiving vancomycin between May 1, 1994, and April 30, 1995, were reviewed for an indication meeting published guidelines. RESULTS: Vancomycin prescribing decreased by 22% following interventions, from 8.5 to 6.8 courses per 100 discharges (P<.05). The estimated proportion of vancomycin ordered for an indication meeting published guidelines was 36.6% overall, with no significant change following interventions. However, during the 2 weeks that computer alerts were in place, 60% of vancomycin use was for an approved indication. CONCLUSIONS: Parenteral vancomycin prescribing decreased significantly following interventions, but the majority of orders still were not for an indication meeting published guidelines. Further improvement in the appropriateness of vancomycin prescribing potentially could be accomplished by more aggressive interventions, such as computer alerts, or by targeting specific aspects of prescribing patterns.

Development and validation of a competitive enzyme-linked immunosorbent assay for detection of type A influenza antibodies in avian sera.

Serologic screening of avian sera for group-specific antibodies to type A influenza is currently accomplished by using the avian influenza (AI) agar gel immunodiffusion (AGID) test. A competitive enzyme-linked immunosorbent assay (CELISA) was developed using a baculovirus vector, Autographa californica nuclear polyhedrosis virus, expressing the nucleoprotein (NP) gene of A/Ann Arbor/6/60 influenza virus. The recombinant NP was obtained by inoculation of Spodoptera frugiperda (Sf9) insect cells or Trichoplusia ni insect larvae with the recombinant baculovirus. A hybridoma cell line producing monoclonal antibody against influenza virus A nucleoprotein was used to generate mouse ascitic fluid for the CELISA. The nucleoprotein and the monoclonal antibody were used without further purification in a CELISA for detection of avian-origin serum antibodies to type A influenza. The AI AGID and CELISA tests were compared for sensitivity and specificity using 1651 experimental and reference antisera. Samples discrepant in AGID and CELISA test results were further evaluated by the AI indirect fluorescent antibody (IFA), hemagglutination-inhibition (HI), and neuraminidase-inhibition (NI) tests. The results demonstrated a high degree of correlation between the AGID and CELISA test results, with the IFA, HI, and NI tests offering additional support of CELISA test specificity. The CELISA is a rapid, economical, sensitive, and specific serodiagnostic method for screening large numbers of avian sera for antibodies to avian influenza virus.

Cue reactivity in cocaine-dependent subjects: effects of cue type and cue modality.

Thirty cocaine-dependent subjects were enrolled into a cue-laboratory study to determine the specificity and sensitivity of this paradigm as a craving measure. Subjects experienced three cue types (i.e., cocaine, arousing, and neutral stimuli) in three cue modalities (i.e., audio, visual, and manual). Cue types were administered in different experimental sessions with a period of 2 to 3 days between sessions. Our results showed that subjective and physiological craving for cocaine was relatively specific for the cocaine cue as compared with either the arousing or the neutral cue. The relative sensitivities of the cue modalities in decreasing order was manual, audio, and visual. We suggest that a modified conditioned-cue paradigm could be a useful tool in the repeated assessment of craving during a clinical study.

Donor attitudes about exporting and importing blood.

BACKGROUND: The movement of blood among different areas of the United States and the collection of more blood than is needed locally in some areas are increasing. Little is known of donors' attitudes about this blood resource sharing. STUDY DESIGN AND METHODS: One thousand donors from five regions of the American Red Cross Blood Services were surveyed by telephone. Demographic information about the donors and the regions was obtained, and the donors were asked to describe their attitudes about blood resource sharing as well as other blood donation-related issues. RESULTS: Donors are not very knowledgeable about whether their community is self-sufficient in its blood supply. In regions that import blood, 29 to 43 percent of donors believed that enough blood was collected to meet all local needs, and, in regions that export blood, only 22 to 24 percent of donors believed that more than enough blood was collected. About three-fourths of the donors believed it acceptable to send their blood to other communities if it is needed there. However, this attitude was based on the premise that all local needs would be met first. Only 4 percent of donors would be less willing to donate if their blood was being sent to another community. CONCLUSION: Donors are not very aware of blood resource sharing but are willing, under certain circumstances, to donate blood for use outside their local communities.

The expression of leptin and its receptors in pre-ovulatory human follicles.

The expression of leptin and its receptors was examined by reverse transcriptase-polymerase chain reaction and immunofluorescence in granulosa and cumulus cells of pre-ovulatory follicles and in meiotically mature oocytes obtained from women undergoing in-vitro fertilization. Leptin concentrations were measured in newly aspirated follicular fluids and in maternal serum before and after the administration of an ovulatory dose of human chorionic gonadotrophin. The findings demonstrate leptin expression at the mRNA and protein levels by granulosa and cumulus cells, and the presence of leptin in mature human oocytes. While an association between follicular leptin concentration and embryo development was not observed, a post-ovulatory increase in serum leptin concentration was associated with implantation potential. The results are discussed with respect to possible roles of leptin in early human development.

Leptin stimulates fetal and adult erythroid and myeloid development.

The ob gene product, leptin, has been shown in several studies to be involved in weight control and recombinant leptin recently has entered clinical trials to treat obesity. The leptin receptor (OB-R/B219) is expressed in a variety of protein isoforms not only in the central nervous system, but also in reproductive, and hematopoietic tissues. We reported recently that the OB-R/B219 was associated with a variety of hematopoietic lineages as well as the small fraction of cells containing the long-term reconstituting hematopoietic stem cells. Herein we report that leptin significantly stimulates the proliferation and differentiation of yolk sac cells and fetal liver cells and stimulates directly hematopoietic precursors. Leptin alone can increase the number of macrophage and granulocyte colonies, and leptin plus erythropoietin act synergistically to increase erythroid development. These data show that leptin has a significant, direct effect on early hematopoietic development and can stimulate the differentiation of lineage-restricted precursors of the erythrocytic and myelopoietic lineages. These observations along with a recent report strongly support our previous hypothesis that leptin has an unanticipated important role in hematopoietic and immune system development.

Preoperative anxiety and fear: a comparison of assessments by patients and anesthesia and surgery residents.

We sought to compare self-assessment of preoperative anxiety levels and selection of worst fears by surgical patients with the assessments made by the anesthesia and surgery residents providing intraoperative care for those patients. One hundred inpatients at a Veterans Affairs hospital (Group 1) and 45 patients at a University hospital (Group 2) were asked to complete a brief questionnaire; the residents were asked to complete the same questionnaire. Group 1 results showed that median patient visual analog scale (VAS) scores were lower for anxiety about anesthesia compared to surgery (16 vs 22, P < or = 0.05). Anesthesia resident VAS scores were higher than patient or surgery resident scores. Neither type of resident was able to predict their individual patient's VAS score (Kendall's tau). The fear chosen with the greatest incidence by Group 1 patients and residents was "whether surgery would work". A significant number of residents (34%, anesthesia or surgery, P < or = 0.05) matched their patient's fear choice. Residents commonly chose fears related to their specialty (e.g., anesthesia residents chose anesthesia-related fears more often than surgery residents, 50% vs 28%, P < or = 0.001). In Group 2, residents demonstrated an improved ability to predict patient scores. For instance, both surgery and anesthesia residents were able to predict individual University patient VAS scores (P < or = 0.01). The fear chosen with the greatest frequency by Group 2 patients was "pain after the operation". Sixty percent of anesthesia residents matched their patients' fear choice (P < or = 0.001). This study indicates a variable ability of anesthesia and surgery residents to predict patient anxiety and fear which may be due, in part, to difficulty in understanding a Veterans Affairs hospital patient population.

Susceptibility of pigeons to avian influenza.

Susceptibility to infection with avian influenza virus (AIV) was studied in pigeons inoculated via oculonasal (Experiment 1) or intravenous (Experiment 2) route. Chickens were included as susceptible hosts in both experiments. Two subtypes each of the highly pathogenic AIV (HPAIV; HP CK/PA H5N2 and HP CK/Australia H7N7) and non-pathogenic AIV (NPAIV; NP CK/PA H5N2 and NP emu/TX H7N1) at a dose of 10(5) embryo infective dose per bird were used as inoculum. The pigeons inoculated with HP CK/PA H5N2 or HP CK/Australia H7N7 remained apparently healthy throughout the 21-day observation period, did not shed viruses on 3, 7, 14, and 21 days postinoculation (DPI), and had no demonstrable levels of antibodies on 21 DPI. On the other hand, 9 of 12 chickens inoculated with the HPAIV died of highly pathogenic avian influenza; the viruses were recovered from their respiratory and intestinal tissues, and the surviving chickens had antibodies to AIV. Regarding responses of pigeons to inoculation with NP CK/PA H5N2 or NP emu/TX H7N1, the pigeons remained clinically healthy throughout the 21-day observation period and did not have detectable levels of antibodies on 21 DPI; only one pigeon yielded the NP emu/TX H7N1 on 3 DPI. The virus was isolated from a tracheal swab and was believed to be the residual inoculum virus. Based on the responses of pigeons to NPAIV and HPAIV, it was concluded that the pigeons were resistant or minimally susceptible to infection with HPAIV or NPAIV.