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Seprehvir (HSV1716) is an oncolytic herpes simplex virus-1 (HSV-1) previously demonstrated to be well tolerated in pediatric patients when administered intratumorally. To determine the safety of administering Seprehvir systemically, we conducted the first-in-human phase I trial of intravenous injection in young patients with relapsed or refractory extra-cranial solid cancers. We delivered a single dose of 5 × 10 4 infectious units (iu)/kg (maximum dose of 2 × 10 6 ) or 2.5 × 10 5 iu/kg (maximum dose of 1 × 10 7 iu) of Seprehvir via the peripheral vein, monitored adverse events, and measured tumor responses by imaging. We monitored HSV-1 serology as well as viremia and shedding by PCR and culture. We administered a single dose of Seprehvir to seven patients and multiple doses to two patients. We did not observe any dose-limiting toxicities. All five HSV-1 seronegative patients seroconverted by day 28. Four of nine patients had detectable HSV-1 genomes in peripheral blood appearing on day +4 consistent with de novo virus replication. Two patients had stable disease in response to Seprehvir. Intravenous Seprehvir is well tolerated without viral shedding in children and young adults with late-stage cancer. Viremia consistent with virus replication holds promise for future Seprehvir studies at higher doses and/or in combination with other anti-neoplastic therapies., (Copyright © 2019 The Authors. Published by Elsevier Inc. All rights reserved.)

Purpose: HSV1716 is an oncolytic herpes simplex virus-1 (HSV-1) studied in adults via injection into the brain and superficial tumors. To determine the safety of administering HSV1716 to pediatric patients with cancer, we conducted a phase I trial of image-guided injection in young patients with relapsed or refractory extracranial cancers. Experimental Design: We delivered a single dose of 10 5 to 10 7 infectious units of HSV1716 via computed tomography-guided intratumoral injection and measured tumor responses by imaging. Patients were eligible for up to three more doses if they achieved stable disease. We monitored HSV-1 serum titers and shedding by PCR and culture. Results: We administered a single dose of HSV1716 to eight patients and two doses to one patient. We did not observe any dose-limiting toxicities. Adverse events attributed to virus included low-grade fever, chills, and mild cytopenias. Six of eight HSV-1 seronegative patients at baseline showed seroconversion on day 28. Six of nine patients had detectable HSV-1 genomes by PCR in peripheral blood appearing on day +4 consistent with de novo virus replication. Two patients had transient focal increases in metabolic activity on 18 fluorine-deoxyglucose PET, consistent with inflammatory reactions. In one case, the same geographic region that flared later appeared necrotic on imaging. No patient had an objective response to HSV1716. Conclusions: Intratumoral HSV1716 is safe and well-tolerated without shedding in children and young adults with late-stage, aggressive cancer. Viremia consistent with virus replication and transient inflammatory reactions hold promise for future HSV1716 studies. Clin Cancer Res; 23(14); 3566-74. ©2017 AACR ., (©2017 American Association for Cancer Research.)

Recovery of large carnivores remains a challenge because complex spatial dynamics that facilitate population persistence are poorly understood. In particular, recovery of the critically endangered red wolf (Canis rufus) has been challenging because of its vulnerability to extinction via human-caused mortality and hybridization with coyotes (Canis latrans). Therefore, understanding red wolf space use and habitat selection is important to assist recovery because key aspects of wolf ecology such as interspecific competition, foraging, and habitat selection are well-known to influence population dynamics and persistence. During 2009-2011, we used global positioning system (GPS) radio-telemetry to quantify space use and 3rd-order habitat selection for resident and transient red wolves on the Albemarle Peninsula of eastern North Carolina. The Albemarle Peninsula was a predominantly agricultural landscape in which red wolves maintained spatially stable home ranges that varied between 25 km2 and 190 km2. Conversely, transient red wolves did not maintain home ranges and traversed areas between 122 km2 and 681 km2. Space use by transient red wolves was not spatially stable and exhibited shifting patterns until residency was achieved by individual wolves. Habitat selection was similar between resident and transient red wolves in which agricultural habitats were selected over forested habitats. However, transients showed stronger selection for edges and roads than resident red wolves. Behaviors of transient wolves are rarely reported in studies of space use and habitat selection because of technological limitations to observed extensive space use and because they do not contribute reproductively to populations. Transients in our study comprised displaced red wolves and younger dispersers that competed for limited space and mating opportunities. Therefore, our results suggest that transiency is likely an important life-history strategy for red wolves that facilitates metapopulation dynamics through short- and long-distance movements and eventual replacement of breeding residents lost to mortality., Competing Interests: The authors have declared that no competing interests exist.

Hibernation is an adaptation to conserve energy in the face of extreme environmental conditions and low food availability that has risen in several animal phyla. This phenomenon is characterized by reduced metabolic rate (∼25% of the active basal metabolic rate in hibernating bears) and energy demand, while other physiological adjustments are far from clear. The profiling of the serum proteome of the American black bear (Ursus americanus) may reveal specific proteins that are differentially modulated by hibernation, and provide insight into the remarkable physiological adaptations that characterize ursid hibernation. In this study, we used differential gel electrophoresis (DIGE) analysis, liquid chromatography coupled to tandem mass spectrometry, and subsequent MASCOT analysis of the mass spectra to identify candidate proteins that are differentially expressed during hibernation in captive black bears. Seventy serum proteins were identified as changing by ±1.5 fold or more, out of which 34 proteins increased expression during hibernation. The majority of identified proteins are involved in immune system processes. These included α2-macroglobulin, complement components C1s and C4, immunoglobulin μ and J chains, clusterin, haptoglobin, C4b binding protein, kininogen 1, α2-HS-glycoprotein, and apoplipoproteins A-I and A-IV. Differential expression of a subset of these proteins identified by proteomic analysis was also confirmed by immunodetection. We propose that the observed serum protein changes contribute to the maintenance of the hibernation phenotype and health, including increased capacities for bone maintenance and wound healing during hibernation in bears.

Background/aims: A decrease in glomerular podocyte number in membranous nephropathy and focal segmental glomerulosclerosis (FSGS) ultimately underlines glomerulosclerosis and the decrease in kidney function. Recent studies have shown that in these diseases, glomerular parietal epithelial cells begin to express proteins considered unique to podocytes, and that these glomerular epithelial transition cells might serve as podocyte progenitors. Because retinoids improve many forms of experimental glomerular disease characterized by podocyte injury and loss, we asked if all-trans retinoic acid (ATRA) induces parietal epithelial cells to express podocyte proteins., Methods: ATRA or vehicle was administered to rats with experimental membranous nephropathy (passive Heymann nephritis model) and mice with experimental FSGS (anti-glomerular antibody model) following the onset of proteinuria. Immunohistochemistry staining of PAX2 (parietal epithelial cell marker), WT-1 (podocyte cell marker), and Ki-67 (proliferation marker) were performed on kidney tissues., Results: Compared to diseased animals receiving vehicle, ATRA statistically significantly increased the number of glomerular transition cells, defined as cells double-staining for PAX2 and WT-1, in membranous nephropathy at weeks 2, 5 and 16, and in FSGS at weeks 1 and 2. This was accompanied by an increase in the number of podocytes compared to diseased controls receiving vehicle., Conclusion: ATRA increases the number of glomerular epithelial transition cells in experimental proteinuric glomerular diseases. Thus, ATRA may provide a useful pharmacologic approach to decipher the mechanisms underlying the possible progenitor role of parietal epithelial cells., (Copyright © 2012 S. Karger AG, Basel.)

Studies have shown that certain cells of the glomerular tuft begin to express proteins considered unique to other cell types upon injury. Little is known about the response of parietal epithelial cells (PEC) to injury. To determine whether PECs change their phenotype upon injury to also express proteins traditionally considered podocyte specific, the following four models of glomerular disease were studied: the transforming growth factor (TGF)-beta1 transgenic mouse model of global glomerulosclerosis, the adriamycin model of focal segmental glomerulosclerosis (FSGS), the anti-glomerular basement membrane (GBM) model of crescentic glomerulonephritis, and the passive Heymann nephritis model of membranous nephropathy. Double immunostaining was performed with antibodies to podocyte-specific proteins (synaptopodin and Wilms' tumor 1) and antibodies to PEC specific proteins (paired box gene 8 and claudin-1). No double staining was detected in normal mice. In contrast, the results showed a statistical increase in the number of cells attached to Bowman basement membrane that were double-positive for both podocyte/PEC proteins in TGF-beta1 transgenic, anti-GBM, and membranous animals. Double-positive cells for both podocyte and PEC proteins were also statistically increased in the glomerular tuft in TGF-beta1 transgenic, anti-GBM, and FSGS mice. These results are consistent with glomerular cells coexpressing podocyte and PEC proteins in experimental glomerular disease, but not under normal circumstances.

Although the normal glomerulus comprises four resident cell types, least is known about the parietal epithelial cells (PECs). This comprehensive review addresses the cellular origin of PECs, discusses the normal structure and protein makeup of PECs, describes PEC function, and defines the responses to injury in disease and how these events lead to clinical events. The data show that PECs have unique properties and that new functions are being recognized such as their role in differentiating into podocytes during disease.

Parietal epithelial cells (PEC) are major constituents of crescents in crescentic glomerulonephritis. The purpose of these studies was to establish an immortalized PEC cell line with similar characteristics to PEC in vivo for use in future mechanistic studies. Glomeruli were isolated from H-2Kb tsA58 transgenic mice (ImmortoMouse) by standard differential sieving, and several candidate PEC cell lines were obtained by subcloning outgrowths of cells from capsulated glomeruli. One clone, designated mouse PEC (mPEC), was extensively characterized. mPEC exhibited a compact cell body with typical epithelial morphology when grown in permissive conditions, but the cell shape changed to polygonal after 14 d in growth-restrictive conditions. mPEC but not podocytes used as a negative control expressed claudin-1, claudin-2, and protein gene product 9.5, which are proteins specific to PEC in vivo, and did not express the podocyte-specific proteins synaptopodin and nephrin. The junctional proteins zonula occludens-1 and beta-catenin stained positively in both mPEC and podocytes, but the staining pattern at cell-cell contacts was intermittent in mPEC and linear in podocytes. Finally, mPEC had thin bundled cortical F-actin filaments and no F-actin projections compared with podocytes, which exhibited thick bundled cortical F-actin filaments and interdigitating F-actin projections at cell-cell contacts. We conclude that immortalized mPEC in culture exhibit specific features of PEC in vivo and that these cells are distinct from podocytes, despite having the same mesenchymal origin. This mPEC line will assist in future mechanistic studies of PEC and enhance our understanding of glomerular injury.

The glomerular capillary tuft is a highly intricate and specialized microvascular bed that filters plasma water and solute to form urine. The mature glomerulus contains four cell types: Parietal epithelial cells that form Bowman's capsule, podocytes that cover the outermost layer of the glomerular filtration barrier, glycocalyx-coated fenestrated endothelial cells that are in direct contact with blood, and mesangial cells that sit between the capillary loops. Filtration begins only after the influx and organization of endothelial and mesangial cells in the developing glomerulus. Tightly coordinated movement and cross-talk between these cell types is required for the formation of a functional glomerular filtration barrier, and disruption of these processes has devastating consequences for early life. Current concepts of the role of mesangial and endothelial cells in formation of the capillary tuft are reviewed here.

Mechanical unloading of bone causes an imbalance in bone formation and resorption leading to bone loss and increased fracture risk. Black bears (Ursus americanus) are inactive for up to six months during hibernation, yet bone mineral content and strength do not decrease with disuse or aging. To test whether hibernating bears have biological mechanisms to prevent disuse osteoporosis, we measured the serum concentrations of hormones and growth factors involved in bone metabolism and correlated them with the serum concentration of a bone formation marker (osteocalcin). Serum was obtained from black bears over a 7-month duration that included periods of activity and inactivity. Both resorption and formation markers increased during hibernation, suggesting high bone turnover occurred during inactivity. However, bone formation appeared to be balanced with bone resorption. The serum concentration of parathyroid hormone (PTH) was higher in the hibernation (P=0.35) and post-hibernation (P=0.006) seasons relative to pre-hibernation levels. Serum leptin was lower (P<0.004) post-hibernation relative to pre-hibernation and hibernation periods. Insulin-like growth factor I (IGF-I) decreased (P<0.0001) during hibernation relative to pre-hibernation and reached its highest value during remobilization. There was no difference (P=0.64) in 25-OH vitamin D between the three seasons. Serum osteocalcin (bone formation marker) was significantly correlated with PTH, but not with leptin, IGF-I or 25-OH vitamin D. Osteocalcin and PTH were positively correlated when samples from all seasons were pooled and when only hibernation samples were considered, raising the possibility that the anabolic actions of PTH help maintain bone formation to prevent disuse osteoporosis. Prostaglandin E(2) (PGE(2)) release from MC3T3 osteoblastic cells was significantly affected by treatment with bear serum from different seasons (i.e. hibernation versus active periods). The seasonal changes in PGE(2) release showed trends similar to the seasonal changes in serum IGF-I. Since both PGE(2) and IGF-I are associated with collagenous bone formation, it is possible that seasonal changes in a circulating factor influence IGF-I levels in vivo in bears and PGE(2) release in osteoblastic cells in vitro. The significant decrease in serum leptin following arousal from hibernation may promote bone formation during remobilization, assuming there is a similar decrease in intracerebroventricular leptin. These findings support the idea that seasonal changes in the concentration of circulating molecules help regulate bone formation activity and may be important for preventing disuse osteoporosis in bears.

The hibernating bear is an excellent model for disuse osteoporosis in humans because it is a naturally occurring large animal model. Furthermore, bears and humans have similar lower limb skeletal morphology, and bears walk plantigrade like humans. Black bears (Ursus americanus) may not develop disuse osteoporosis during long periods of disuse (i.e. hibernation) because they maintain osteoblastic bone formation during hibernation. As a consequence, bone volume, mineral content, porosity, and strength are not adversely affected by annual periods of disuse. In fact, cortical bone bending strength has been shown to increase with age in hibernating black bears without a significant change in porosity. Other animals require remobilization periods 2-3 times longer than the immobilization period to recover the bone lost during disuse. Our findings support the hypothesis that black bears, which hibernate for as long as 5-7 months annually, have evolved biological mechanisms to mitigate the adverse effects of disuse on bone porosity and strength.

Background: Podocytes are terminally differentiated and highly specialized epithelial cells. The factors governing podocyte differentiation are poorly understood. We tested the hypothesis that all-trans retinoic acid (ATRA), a vitamin A derivative, induces podocyte differentiation in vitro and in vivo., Methods: We tested the effects of ATRA on podocytes. Primary rat, primary mouse, and immortalized mouse podocytes were exposed to ATRA (1, 5, 10, 20, 40, 50, 80, 160, and 200 micromol/L) or control (ethanol) for 72 hours. Cell morphology was examined by electron microscopy, the expression of podocyte specific proteins was measured by immunoflourescence and Western blot analysis, cell number and apoptosis were measured by 3-[4,5] dimethylthiazol-2,5-diphenyltetrazolium bromide (MTT) assay and Hoechst staining, respectively. To determine if ATRA alters podocyte differentiation in vivo, experimental injury was induced in C57BL6 mice using the antiglomerular antibody. Animals were given either daily intraperitoneal ATRA (16 mg/kg) or vehicle (corn oil). For end points, we measured proteinuria, podocyte-specific protein immunostaining, and proliferation [proliferating cell nuclear antigen (PCNA)] at days 5 and 14 (N= 5/group/time point)., Results: ATRA induced podocyte process formation in vitro, and significantly increased the expression of nephrin and podocin. This coincided with a reduction in proliferation. ATRA also significantly prevented the decrease in staining for synaptopodin, nephrin, and podocin in experimental animals (P < 0.05 vs. control). This was accompanied by reduced proteinuria and decreased podocyte proliferation (P < 0.05 vs. control)., Conclusion: ATRA induces podocyte differentiation in vitro and in vivo and alters the expression of certain podocyte-specific proteins. Further studies are ongoing to delineate the mechanism of this effect.

Disuse by bed rest, limb immobilization or space flight causes rapid bone loss by arresting bone formation and accelerating bone resorption. This net bone loss increases the risk of fracture upon remobilization. Bone loss also occurs in hibernating ground squirrels, golden hamsters, and little brown bats by arresting bone formation and accelerating bone resorption. There is some histological evidence to suggest that black bears Ursus americanus do not lose bone mass during hibernation (i.e. disuse). There is also evidence suggesting that muscle mass and strength are preserved in black bears during hibernation. The question of whether bears can prevent bone loss during hibernation has not been conclusively answered. The goal of the current study was to further assess bone metabolism in hibernating black bears. Using the same serum markers of bone remodeling used to evaluate human patients with osteoporosis, we assayed serum from five black bears, collected every 10 days over a 196-day period, for bone resorption and formation markers. Here we show that bone resorption remains elevated over the entire hibernation period compared to the pre-hibernation period, but osteoblastic bone formation is not impaired by hibernation and is rapidly accelerated during remobilization following hibernation.

Disuse osteopenia was studied in hibernating black bears (Ursus americanus) using serum markers of bone metabolism. Blood samples were collected from male and female, wild black bears during winter denning and active summer periods. Radioimmunoassays were done to determine serum concentrations of cortisol, the carboxy-terminal cross-linked telopeptide, and the carboxy-terminal propeptide of Type I procollagen, which are markers of bone resorption and formation, respectively. The bone resorption marker was significantly higher during winter hibernation than it was in the active summer months, but the bone formation marker was unchanged, suggesting an imbalance in bone remodeling and a net bone loss during disuse. Serum cortisol was significantly correlated with the bone resorption marker, but not with the bone formation marker. The bone formation marker was four- to fivefold higher in an adolescent and a 17-year-old bear early in the remobilization period compared with the later summer months. These findings raise the possibility that hibernating black bears may minimize bone loss during disuse by maintaining osteoblastic function and have a more efficient compensatory mechanism for recovering immobilization-induced bone loss than that of humans or other animals.

Congenital hyperplastic goiter and cretinism were documented in a 16 month-old male North American black bear (Ursus americanus). The cub was captured at approximately 8 months of age and maintained for an additional 8 months in captivity. Clinical signs included growth retardation, clumsiness, and facial dysmorphism. Hypothyroidism was documented by determining serum triiodothyronine (T3) and thyroxine (T4) levels. Lysosomal storage disease was ruled out by measuring various lysosomal enzyme activities. Serologic, radiographic, computed tomographic, necropsy, and histopathologic findings were consistent with congenital hypothyroidism and cretinism.

Background: Among an independent group of subjects selected for their history of violent, impulsive behaviors and nonviolent control subjects, we attempted to replicate the finding that plasma docosahexaenoic acid concentrations were negatively correlated with cerebrospinal fluid 5-hydroxyindoleacetic acid (CSF 5-HIAA) concentrations., Methods: CSF 5-HIAA and homovanillic acid (HVA), fasting total cholesterol, and plasma fatty acid concentrations were examined in violent and nonviolent subjects matched for their severity of alcohol dependence., Results: Violent subjects had significantly higher lifetime violence and hostility ratings and lower concentrations of CSF 5-HIAA than nonviolent subjects. Plasma docosahexaenoic acid was negatively correlated with CSF 5-HIAA only among violent subjects., Conclusions: This observational study suggests that dietary essential fatty acids may change neurotransmitter concentrations. Prospective dietary intervention trials will be required to determine if increasing dietary intake of docosahexaenoic acid will increase or decrease either CSF 5-HIAA concentrations or impulsive and violent behaviors.

Knowledge of the relative stabilities of S-DNA.RNA hybrids of different sequences is important for choosing RNA targets for hybridization with antisense phosphorothioate oligodeoxyribonucleotides (S-DNAs). It is also important to know how hybrid secondary structure varies with sequence, since different structures could influence thermal stability and the activity of RNase H. Our approach has been to study relatively simple sequences consisting of repeating di-, tri-, and tetranucleotides, which allow the maximum resolution of nearest-neighbor effects. Circular dichroism (CD) spectra and melting temperatures were acquired for 16 hybrid sequences that could be formed by mixing S-DNA and RNA oligomers of 24 nucleotides in length. CD spectra of S-DNA.RNA hybrids were sequence-dependent and were similar to those of analogous unmodified hybrids. From singular value decomposition, the major CD spectral component was like that of the A-conformation. Three nearest-neighbor relationships among the hybrid CD spectra were in as good agreement as are such relationships among spectra of duplex RNAs. Tm values ranged from 44.1 degrees C for S-d(ACT)8. r(AGU)8 to 66.6 degrees C for S-d(CCT)8.r(AGG)8 (in 0.15 M K+, phosphate buffer, pH 7). The S-DNA.RNA hybrids had a sequence-dependence of melting temperatures that was approximately the same as that calculated using published data for normal DNA.RNA hybrids [Sugimoto, N., Nakano, S., Katoh, M., Matsumura, A., Nakamuta, H., Ohmichi, T.,Yoneyama, M., & Sasaki, M. (1995) Biochemistry 34, 11211-11216]. In general, sequence-dependent CD spectra and Tm values of S-DNA.RNA hybrids appear to reflect the unique nearest-neighbor interactions of adjacent base pairs, where the S-DNA and RNA strands are in different, but relatively uniform, conformations.

The National Institutes of Health (NIH) and the National Aeronautics and Space Administration (NASA) are seeking solutions to the human problem of osteopenia, or immobility-induced bone loss. Bears, during winter dormancy, appear uniquely exempted from the debilitating effects of immobility osteopenia. NIH and ESA, Inc. are creating a large database of metabolic information on human ambulatory and bedrest plasma samples for comparison with metabolic data obtained from bear plasma samples collected in different seasons. The database generated from NASA's HR113 human bedrest study showed a clear difference between plasma samples of ambulatory and immobile subjects through cluster analysis using compounds determined by high performance liquid chromatography with coulometric electrochemical array detection (HPLC-EC). We collected plasma samples from black bears (Ursus americanus) across 4 seasons and from 3 areas and subjected them to similar analysis, with particular attention to compounds that changed significantly in the NASA human study. We found seasonal differences in 28 known compounds and 33 unknown compounds. A final database contained 40 known and 120 unknown peaks that were reliably assayed in all bear and human samples; these were the primary data set for interspecies comparison. Six unidentified compounds changed significantly but differentially in wintering bears and immobile humans. The data are discussed in light of current theories regarding dormancy, starvation, and anabolic metabolism. Work is in progress by ESA Laboratories on a larger database to confirm these findings prior to a chemical isolation and identification effort. This research could lead to new pharmaceuticals or dietary interventions for the treatment of immobility osteopenia.

Circular dichroism spectroscopy was used to study mutants of phage fd gene 5 protein (Y26F, Y34F, and Y41F) in which three of the five tyrosines, Tyr-26, Tyr-34, and Tyr-41, were individually substituted with phenylalanine. The tyrosine 229 nm CD bands of the wild type, Y26F, and Y41F gene 5 proteins decreased in magnitude during complex formation with either fd ssDNA or poly[d(A)]. However, the Y34F gene 5 protein showed no decrease in the 229 nm band during titrations of these nucleic acids. This suggested that Tyr-34 of the wild type gene 5 protein dominated the 229 nm CD changes upon binding to single-stranded DNA. Titrations of poly[d(A)] or fd ssDNA with wild type, Y26F, Y34F, or Y41F gene 5 proteins resulted in perturbations of the nucleic acid near-UV CD bands, specific for the particular nucleic acid, but similar for all four proteins (in 2 mM sodium phosphate buffer, pH 7.0). For both nucleic acids, the addition of protein beyond a certain [protein monomer]/[nucleotide] ratio (0.25 for poly[d(A)] or 0.33 for fd ssDNA) resulted in a partial reversal of the CD change of the nucleic acid. These data are interpreted to mean that, in addition to the two well-known n = 4 and n = 3 stoichiometric modes of binding, there is a third mode of binding in which the nucleic acid is in limited contact with the protein. As shown by salt dissociation studies of complexes with poly[d(A)], the binding affinities, K omega, of the proteins were in the order: wild type > Y26F >> Y34F > or = Y41F (for the n = 4 binding mode in 0.1-0.2 M NaCl). Our data indicate that Tyr-34 plays a more important role in forming a complex with ssDNA than is apparent in current models of the g5p.ssDNA complex. We suggest that the hydroxyl moieties of Tyr-34 and Tyr-41 are both somehow involved in stabilizing the interface of bound protein dimers.

Small-angle scattering and electron microscopy studies of fd gene 5 protein (g5p) and reconstituted g5p.nucleic acid complexes have been used to test models for the complexes and evaluate their uniqueness. In addition, we have obtained new information on the dependence of nucleotide type and protein/nucleotide (P/N) ratio on the structure of the complexes. Reconstituted complexes were made with single-stranded fd viral DNA (fd ssDNA), poly[d(A)] and poly[r(A)]. All complexes form similar left-handed, flexible superhelices having approximately the same diameter, but the pitch differs among these complexes. The g5p protein is a dimer in solution and the dimers associate to form a superhelical framework to which the polynucleotide is attached. The combined X-ray and neutron scattering data confirm the nucleic acid is inside the protein superhelix. A Monte Carlo integration modeling procedure applied to the scattering data was used to systematically test large numbers of possible models for each complex, and previously proposed models based on parameters obtained from electron microscopy were found to be essentially correct and unique. The data on the complexes with different P/N ratios showed that mass per unit length values decreased while the rise per dimer and pitch of the superhelix increased for g5p.fd-ssDNA complexes with decreasing P/N ratios.

This study evaluated the physiological condition and environmental contaminant concentrations in muskrats inhabiting the contaminated lower region of the Elizabeth River, Virginia, (USA). Muskrats trapped in the lower region of the Elizabeth River weighed less, had lower mean fat indexes, lower relative spleen weights, greater relative adrenal weights, and an increased incidence of disease and parasitism compared to muskrats trapped in the less contaminated upper region of the Elizabeth River or Nansemond River. Concentrations of the metals aluminum, cadmium, copper, nickel, and zinc and the number of polyaromatic hydrocarbon compounds were greater in lower Elizabeth River muskrats compared to upper region Elizabeth River or Nansemond River muskrats. These physiological data indicated that muskrats in the lower region of the Elizabeth River were in poor health relative to muskrats from the other regions studied. However, no significant difference in fecundity or density was observed among regions studied, suggesting that the level of environmental contaminant exposure was not sufficient to elicit a population level response. Greater population declines following severe natural environmental stresses (large storms, extreme cold, etc.) would be anticipated in lower region Elizabeth River muskrats; however, the muskrat population in this region does not now appear to be at risk of extirpation.

CD spectra were obtained for eight synthetic double-stranded DNA polymers down to at least 175 nm in the vacuum uv. Three sets of sequence isomers were studied: (a) poly[d(A-C).d(G-T)] and poly[d(A-G).d(C-T)], (b) poly[d(A-C-C).d(G-G-T)] and poly[d(A-C-G).d(C-G-T)], and (c) poly[d(A).d(T)], poly[d(A-T).d(A-T)], poly[d(A-A-T).d(A-T-T)], and poly[d(A-A-T-T).d(A-A-T-T)]. There were significant differences in the CD spectra at short wavelengths among each set of sequence isomers. The (G.C)-containing sequences had the largest vacuum uv bands, which were positive and in the wavelength range of 180-191 nm. There were no large negative bands at longer wavelengths, consistent with the polymers all being in right-handed conformations. Among the set of sequences containing only A.T base pairs, poly[d(A).d(T)] had the largest vacuum uv CD band, which was at 190 nm. This CD band was not present in the spectra of the other (A.T)-rich polymers and was absent from two first-neighbor estimations of the poly[d(A).d(T)] spectrum obtained from the other three sequences. We concluded that the sequence dependence of the vacuum uv spectra of the (A.T)-rich polymers was due in part to the fact that poly[d(A).d(T)] exists in a noncanonical B conformation.

Rectal temperature was determined for 84 black bears (Ursus americanus) during 99 handlings in Great Dismal Swamp, Virginia and North Carolina (USA). All bears had been trapped with cable snares and immobilized with a 2:1 ketamine hydrochloridexylazine hydrochloride mixture. Temperatures were significantly greater in males and varied significantly by season. Immobilized bears began panting at rectal temperatures greater than 42.0 C. One death occurred at 43.0 C. We recommended cooling measures on black bears at rectal temperatures of greater than or equal to 40.0 C.