Mammalian Adaptation of an Avian Influenza A Virus Involves Stepwise Changes in NS1.

Influenza A viruses (IAVs) are common pathogens of birds that occasionally establish endemic infections in mammals. The processes and mechanisms that result in IAV mammalian adaptation are poorly understood. The viral nonstructural 1 (NS1) protein counteracts the interferon (IFN) response, a central component of the host species barrier. We characterized the NS1 proteins of equine influenza virus (EIV), a mammalian IAV lineage of avian origin. We showed that evolutionarily distinct NS1 proteins counteract the IFN response using different and mutually exclusive mechanisms: while the NS1 proteins of early EIVs block general gene expression by binding to cellular polyadenylation-specific factor 30 (CPSF30), NS1 proteins from more evolved EIVs specifically block the induction of IFN-stimulated genes by interfering with the JAK/STAT pathway. These contrasting anti-IFN strategies are associated with two mutations that appeared sequentially and were rapidly selected for during EIV evolution, highlighting the importance of evolutionary processes in immune evasion mechanisms during IAV adaptation.IMPORTANCE Influenza A viruses (IAVs) infect certain avian reservoir species and occasionally transfer to and cause epidemics of infections in some mammalian hosts. However, the processes by which IAVs gain the ability to efficiently infect and transmit in mammals remain unclear. H3N8 equine influenza virus (EIV) is an avian-origin virus that successfully established a new lineage in horses in the early 1960s and is currently circulating worldwide in the equine population. Here, we analyzed the molecular evolution of the virulence factor nonstructural protein 1 (NS1) and show that NS1 proteins from different time periods after EIV emergence counteract the host innate immune response using contrasting strategies, which are associated with two mutations that appeared sequentially during EIV evolution. The results shown here indicate that the interplay between virus evolution and immune evasion plays a key role in IAV mammalian adaptation.

Polarization signaling in swordtails alters female mate preference.

Polarization of light, and visual sensitivity to it, is pervasive across aquatic and terrestrial environments. Documentation of invertebrate use of polarized light is widespread from navigation and foraging to species recognition. However, studies demonstrating that polarization body patterning serves as a communication signal (e.g., with evidence of changes in receiver behavior) are rare among invertebrate taxa and conspicuously absent among vertebrates. Here, we investigate polarization-mediated communication by northern swordtails, Xiphophorus nigrensis, using a custom-built videopolarimeter to measure polarization signals and an experimental paradigm that manipulates polarization signals without modifying their brightness or color. We conducted mate choice trials in an experimental tank that illuminates a pair of males with light passed through a polarization filter and a diffusion filter. By alternating the order of these filters between males, we presented females with live males that differed in polarization reflectance by >200% but with intensity and color differences below detection thresholds (∼5%). Combining videopolarimetry and polarization-manipulated mate choice trials, we found sexually dimorphic polarized reflectance and polarization-dependent female mate choice behavior with no polarization-dependent courtship behavior by males. Male swordtails exhibit greater within-body and body-to-background polarization contrast than females, and females preferentially associate with high-polarization-reflecting males. We also found limited support that males increase polarization contrast in social conditions over asocial conditions. Polarization cues in mate choice contexts may provide aquatic vertebrates with enhanced detection of specific display features (e.g., movements, angular information), as well as a signaling mechanism that may enhance detection by intended viewers while minimizing detection by others.

Polarimetric imaging and retrieval of target polarization characteristics in underwater environment.

Polarized light fields contain more information than simple irradiance and such capabilities provide an advanced tool for underwater imaging. The concept of the beam spread function (BSF) for analysis of scalar underwater imaging was extended to a polarized BSF which considers polarization. The following studies of the polarized BSF in an underwater environment through Monte Carlo simulations and experiments led to a simplified underwater polarimetric imaging model. With the knowledge acquired in the analysis of the polarimetric imaging formation process of a manmade underwater target with known polarization properties, a method to extract the inherent optical properties of the water and to retrieve polarization characteristics of the target was explored. The proposed method for retrieval of underwater target polarization characteristics should contribute to future efforts to reveal the underlying mechanism of polarization camouflage possessed by marine animals and finally to generalize guidelines for creating engineered surfaces capable of similar polarization camouflage abilities in an underwater environment.

Polaro-cryptic mirror of the lookdown as a biological model for open ocean camouflage.

With no object to hide behind in 3D space, the open ocean represents a challenging environment for camouflage. Conventional strategies for reflective crypsis (e.g., standard mirror) are effective against axially symmetric radiance fields associated with high solar altitudes, yet ineffective against asymmetric polarized radiance fields associated with low solar inclinations. Here we identify a biological model for polaro-crypsis. We measured the surface-reflectance Mueller matrix of live open ocean fish (lookdown, Selene vomer) and seagrass-dwelling fish (pinfish, Lagodon rhomboides) using polarization-imaging and modeling polarization camouflage for the open ocean. Lookdowns occupy the minimization basin of our polarization-contrast space, while pinfish and standard mirror measurements exhibit higher contrast values than optimal. The lookdown reflective strategy achieves significant gains in polaro-crypsis (up to 80%) in comparison with nonpolarization sensitive strategies, such as a vertical mirror. Lookdowns achieve polaro-crypsis across solar altitudes by varying reflective properties (described by 16 Mueller matrix elements m(ij)) with incident illumination. Lookdowns preserve reflected polarization aligned with principle axes (dorsal-ventral and anterior-posterior, m(22) = 0.64), while randomizing incident polarization 45° from principle axes (m(33) = -0.05). These reflectance properties allow lookdowns to reflect the uniform degree and angle of polarization associated with high-noon conditions due to alignment of the principle axes and the sun, and reflect a more complex polarization pattern at asymmetrical light fields associated with lower solar elevations. Our results suggest that polaro-cryptic strategies vary by habitat, and require context-specific depolarization and angle alteration for effective concealment in the complex open ocean environment.

Three-dimensional measurements of animal paths using handheld unconstrained GoPro cameras and VSLAM software.

I present the system PATMOS (paths and tessellated meshes from ORB_SLAM2) for measuring three-dimensional paths of animalsin situusing two handheld GoPro cameras and a small spatial reference object. Animal paths were triangulated from mobile camera positions obtained from a modified version of ORB_SLAM2, an open-source visual simultaneous localization and mapping software package. In addition to path calculation, this process provided a virtual three-dimensional surface approximation to the environment from which path to environment distances can be quantified. PATMOS can also fit a tranquil water's surface to an analytic plane if there are a sufficient number of visible objects intersecting the water's surface and can track objects over the water's surfaces with a single camera by measuring the object with its reflection. This technology was highly portable, could follow moving animals, and gave comparable spatial and temporal resolutions to fixed camera systems that use commercial cameras. An investigation of falling objects yielded a gravitational constant measurement of 978 ± 40 cm s-2. I demonstrated PATMOS's utility in terrestrial and aquatic environments by quantifying dragonfly flight characteristics and the inter-spatial distances between substrate and damselfish.

Determination of Total Lipid and Lipid Classes in Marine Samples.

Lipids are largely composed of carbon and hydrogen and, therefore, provide a greater specific energy than other organic macromolecules in the sea. Being carbon- and hydrogen-rich they are also hydrophobic and can act as a solvent and absorption carrier for organic contaminants and thus can be drivers of pollutant bioaccumulation in marine ecosystems. Their hydrophobic nature facilitates their isolation from seawater or biological specimens: marine lipid analysis begins with sampling and then extraction in non-polar organic solvents, providing a convenient method for their separation from other substances in an aquatic matrix. If seawater has been sampled, the first step usually involves separation into operationally defined 'dissolved' and 'particulate' factions by filtration. Samples are collected and lipids isolated from the sample matrix typically with chloroform for truly dissolved matter and colloids, and with mixtures of chloroform and methanol for solids and biological specimens. Such extracts may contain several classes from biogenic and anthropogenic sources. At this time, total lipids and lipid classes may be determined. Total lipid can be measured by summing individually determined lipid classes which customarily have been chromatographically separated. Thin-layer chromatography (TLC) with flame ionization detection (FID) is regularly used for the quantitative analysis of lipids from marine samples. TLC-FID furnishes synoptic lipid class information and, by summing classes, a total lipid measurement. Lipid class information is especially useful when combined with measurements of individual components e.g., fatty acids and/or sterols, after their release from lipid extracts. The wide variety of lipid structures and functions means they are used broadly in ecological and biogeochemical research assessing ecosystem health and the degree of influence by anthropogenic impacts. They have been employed to measure substances of dietary value to marine fauna (e.g., aquafeeds and/or prey), and as an indicator of water quality (e.g., hydrocarbons).

Comparison of the polarization contrast of gelatinous zooplankton and a transparent single-use plastic bag-Implications for marine animals.

Plastic pollution in the ocean is an increasingly detrimental issue for marine organisms. As a form of polarized light pollution, transparent plastic debris may be more visible and pose additional threats to organisms that can detect and interpret polarized light. Plastic can mimic the visual features of common marine prey items, such as transparent gelatinous zooplankton, which may lead to more significant plastic ingestion. We measured, in situ, the polarization and radiance contrast between a transparent plastic bag and gelatinous zooplankton with an underwater video polarimeter. The plastic bag had significantly higher polarization contrast than the gelatinous zooplankton, yet both shared similar radiance contrasts. This higher polarization contrast may contribute to the observed high ingestion rates of transparent plastic by marine organisms. Further study into the connection between polarization-sensitive organisms and plastic ingestion is recommended.

Natural variation of canine parvovirus.

Canine parvovirus was first recognized during 1978. Analysis of isolates collected since its emergence revealed that viruses circulating after 1980 were antigenically different from earlier isolates. Monoclonal antibodies clearly distinguished the two strains, some being specific for either the old or the new viruses. Restriction enzyme analysis of viral DNA's showed that the post-1980 viruses were similar to earlier isolates, but some restriction site differences were present in the new strain. These results suggest that the canine parvoviruses infecting dogs in the seven areas of the United States that were sampled derive from a variant virus that replaced the original strain during 1980.

Genetic Analysis of Feline Panleukopenia Virus Full-length VP2 Gene in Domestic Cats Between 2006-2008 and 2012-2014, Portugal.

Feline panleukopenia virus (FPV) and canine parvovirus (CPV) are two closely related viruses, which cause acute gastroenteritis in carnivores, and cats may be infected by strains of both viruses. The viruses are found worldwide and may have changing host ranges and genetic variation that can be found around the world in some cases. Here, we screened a Portuguese population of cats by a conventional PCR assay for the presence of FPV/CPV viruses in faecal samples and tissues between 2006-2008 and 2012-2014. The sequence analysis of the complete VP2 gene showed that 18 of 31 animals tested were positive for FPV DNA, and no case of CPV infection was detected. The analysis of specific DNA detected three new non-synonymous substitutions in the VP2 gene that were found in single groups and were related to viruses reported elsewhere by phylogenetic analysis - some were related to Italian isolates, one was closely related to isolates from Vietnam and China, and two were related with older strains from the USA. The results of our study show that FPV circulated in the Portuguese cat population and as expected the found strains are slightly divergent from those reported previously.

Treatment of gastroparesis: a multidisciplinary clinical review.

This clinical review on the treatment of patients with gastroparesis is a consensus document developed by the American Motility Society Task Force on Gastroparesis. It is a multidisciplinary effort with input from gastroenterologists and other specialists who are involved in the care of patients with gastroparesis. To provide practical guidelines for treatment, this document covers results of published research studies in the literature and areas developed by consensus agreement where clinical research trials remain lacking in the field of gastroparesis.

The structure of a neutralized virus: canine parvovirus complexed with neutralizing antibody fragment.

BACKGROUND: Members of the Parvovirus genus cause a variety of diseases in mammals, including humans. One of the major defences against viral infection is the presence of neutralizing antibodies that prevent virus particles from infecting target cells. The mechanism of neutralization is not well understood. We therefore studied the structure of canine parvovirus (CPV) complexed with the Fab fragment of a neutralizing antibody, A3B10, using image reconstruction of electron micrographs of vitrified samples, together with the already known structure of CPV from X-ray crystallographic data. RESULTS: The structure of the complex of CPV with Fab A3B10 has been determined to 23 A resolution. The known CPV atomic structure was subtracted from the electron density of the complex, and the difference map was used to fit the atomic coordinates of a known Fab fragment, HyHEL-5. The long axis of each Fab molecule is oriented in a near radial direction, inclined away from the two-fold axes. The viral epitope consists of 14 amino acid residues found in loops 1, 2 and 3 on the capsid surface, which include previously identified escape mutations. CONCLUSIONS: The mode of Fab binding suggests that the A3B10 neutralizing antibody cannot bind bivalently to the capsid across the two-fold axes, consistent with the observation that whole A3B10 antibody readily precipitates CPV. Since Fab A3B10 can also neutralize the virus, mechanisms of neutralization such as interference with cell attachment, cell entry, or uncoating, must be operative.

Lipid structure of rat adipocyte plasma membranes following dietary lard and fish oil.

We have determined the changes in the lipid structure of the adipocyte plasma membranes of rats receiving lard or fish oil in their diet. For this purpose, mature Wistar rats were fed 20% (w/w) lard or fish oil diets for 22 days, when the plasma membranes of the epididymal and perirenal adipocytes were prepared. Detailed analysis of the membrane lipids by chromatographic methods showed that dietary fat exerted a major effect on the lipid class and molecular species composition of the phospholipids. As a result of fish oil feeding, significant increases in the 20:5(n-3), 22:5(n-3) and 22:6(n-3) were detected in all glycerophospholipid classes, while the 18:1(n-9) and 18:2(n-6) and to a lesser extent 20:4(n-6) decreased. Incorporation of n-3 fatty acids increased the phosphatidylcholine/sphingomyelin ratio without changing the total phospholipid or free cholesterol content of the membrane. Fish oil feeding also caused a marked increase in the proportion of 24:1 in sphingomyelins, which occurred mainly at the expense of 18:0 and 24:0. New n-3 fatty acid-containing species appeared in the choline and ethanolamine glycerophospholipids, when compared to membrane lipids from lard-fed rats. Membranes from fish oil fed rats also had moderately higher levels of ether lipids. Few differences were seen between the membranes of the epididymal and perirenal adipocytes. It is concluded that dietary fish oils modify the lipid structure of rat adipocyte plasma membranes by increasing the ratio of phosphatidylcholine to sphingomyelin and by increasing the proportion of molecular species with polyunsaturated fatty acids, which would be anticipated to increase the fluidity of the lipid bilayer of adipocyte plasma membranes.

Host range and variability of calcium binding by surface loops in the capsids of canine and feline parvoviruses.

Canine parvovirus (CPV) emerged in 1978 as a host range variant of feline panleukopenia virus (FPV). This change of host was mediated by the mutation of five residues on the surface of the capsid. CPV and FPV enter cells by endocytosis and can be taken up by many non-permissive cell lines, showing that their host range and tissue specificity are largely determined by events occurring after cell entry. We have determined the structures of a variety of strains of CPV and FPV at various pH values and in the presence or absence of Ca(2+). The largest structural difference was found to occur in a flexible surface loop, consisting of residues 359 to 375 of the capsid protein. This loop binds a divalent calcium ion in FPV and is adjacent to a double Ca(2+)-binding site, both in CPV and FPV. Residues within the loop and those associated with the double Ca(2+)-binding site were found to be essential for virus infectivity. The residues involved in the double Ca(2+)-binding site are conserved only in FPV and CPV. Our results show that the loop conformation and the associated Ca(2+)-binding are influenced by the Ca(2+) concentration, as well as pH. These changes are correlated with the ability of the virus to hemagglutinate erythrocytes. The co-localization of hemagglutinating activity and host range determinants on the virus surface implies that these properties may be functionally linked. We speculate that the flexible loop and surrounding regions are involved in binding an as yet unidentified host molecule and that this interaction influences host range.

Oncolytic reovirus enhances rituximab-mediated antibody-dependent cellular cytotoxicity against chronic lymphocytic leukaemia.

The naturally occurring oncolytic virus (OV), reovirus, replicates in cancer cells causing direct cytotoxicity, and can activate innate and adaptive immune responses to facilitate tumour clearance. Reovirus is safe, well tolerated and currently in clinical testing for the treatment of multiple myeloma, in combination with dexamethasone/carfilzomib. Activation of natural killer (NK) cells has been observed after systemic delivery of reovirus to cancer patients; however, the ability of OV to potentiate NK cell-mediated antibody-dependent cellular cytotoxicity (ADCC) is unexplored. This study elucidates the potential of oncolytic reovirus for the treatment of chronic lymphocytic leukaemia (CLL), both as a direct cytotoxic agent and as an immunomodulator. We demonstrate that reovirus: (i) is directly cytotoxic against CLL, which requires replication-competent virus; (ii) phenotypically and functionally activates patient NK cells via a monocyte-derived interferon-α (IFNα)-dependent mechanism; and (iii) enhances ADCC-mediated killing of CLL in combination with anti-CD20 antibodies. Our data provide strong preclinical evidence to support the use of reovirus in combination with anti-CD20 immunotherapy for the treatment of CLL.

Open-ocean fish reveal an omnidirectional solution to camouflage in polarized environments.

Despite appearing featureless to our eyes, the open ocean is a highly variable environment for polarization-sensitive viewers. Dynamic visual backgrounds coupled with predator encounters from all possible directions make this habitat one of the most challenging for camouflage. We tested open-ocean crypsis in nature by collecting more than 1500 videopolarimetry measurements from live fish from distinct habitats under a variety of viewing conditions. Open-ocean fish species exhibited camouflage that was superior to that of both nearshore fish and mirrorlike surfaces, with significantly higher crypsis at angles associated with predator detection and pursuit. Histological measurements revealed that specific arrangements of reflective guanine platelets in the fish's skin produce angle-dependent polarization modifications for polarocrypsis in the open ocean, suggesting a mechanism for natural selection to shape reflectance properties in this complex environment.

Changes in dietary lipid saturation modify fatty acid composition and high-density-lipoprotein binding of adipocyte plasma membrane.

Binding of rat high-density lipoprotein (HDL) to adipocyte plasma membranes appears to be mediated by specific protein-recognition sites, but membrane lipids may also affect these interactions. To study the effect of dietary fatty acid composition on adipocyte-membrane phospholipid composition and HDL binding, male Wistar rats (250 +/- 10 g) were fed diets high in polyunsaturated (20% wt:wt sunflowerseed oil) or saturated and monounsaturated fatty acids (20% lard, by weight). Sunflower-oil vs lard feeding significantly increased the polyunsaturated fatty acid content of membrane diacylglycerophospholipids and the (22:0 and 24:0) content of long-chain fatty acids of sphingomyelin (p less than 0.05) from epididymal and perirenal adipocyte plasma membranes. Concomitant with these changes in membrane phospholipid composition, sunflower-oil vs lard treatment significantly increased the maximum binding capacity (Bmax) of 125I-labeled HDL2 (p less than 0.05). These results show that feeding polyunsaturated fatty acids enhanced HDL2 Bmax of adipocyte plasma membranes, possibly as a consequence of altering the fatty acid composition of membrane phospholipids.

Response to a hospitalized case of measles at a medical school affiliated hospital.

A 27-year-old woman was diagnosed as having measles 2 days after being hospitalized for an unrelated complaint. Hospital personnel, medical housestaff, and students were considered at risk for developing measles infection. Over a 7-day period, measles vaccine was administered to individuals born in or after 1957. No secondary measles cases occurred in the hospital. An immediate response to the possibility of nosocomial measles transmission can be successfully undertaken, but ongoing preemployment or preenrollment school programs are more practical and potentially can have a higher rate of compliance.

Imaging of adenoviral-directed herpes simplex virus type 1 thymidine kinase reporter gene expression in mice with radiolabeled ganciclovir.

UNLABELLED: We are developing procedures to repeatedly and noninvasively image the expression of transplanted reporter genes in living animals and in patients, using PET. We have investigated the use of the Herpes Simplex Virus type 1 thymidine kinase gene (HSV1-tk) as a reporter gene and [8-14C]-ganciclovir as a reporter probe. HSV1-tk, when expressed, leads to phosphorylation of [8-14C]-ganciclovir. As a result, specific accumulation of phosphorylated [8-14C]-ganciclovir should occur almost exclusively in tissues expressing the HSV1-tk gene. METHODS: An adenoviral vector was constructed carrying the HSV1-tk gene along with a control vector. C6 rat glioma cells were infected with either viral vector and uptake of [8-3H]-ganciclovir was determined. In addition, 12 mice were injected with varying levels of either viral vector. Adenovirus administration in mice leads primarily to liver infection. Forty-eight hours later the mice were injected with [8-14C]-ganciclovir, and 1 hr later the mice were sacrificed and biodistribution studies performed. Digital whole-body autoradiography also was performed on separate animals. HSV1-tk expression was assayed, using both normalized HSV1-tk mRNA levels and relative HSV1-TK enzyme levels, in both the cell culture and murine studies. RESULTS: Cell culture, murine tissue biodistribution and murine in vivo digital whole-body autoradiography all demonstrate the feasibility of HSV1-tk as a reporter gene and [8-14C]-ganciclovir as an imaging reporter probe. A good correlation (r2 = 0.86) between the [8-14C]-ganciclovir percent injected dose per gram tissue from HSV1-tk positive tissues and HSV1-TK enzyme levels in vivo was found. An initial study in mice with [8-18F]-fluoroganciclovir and microPET imaging supports further investigation of [8-18F]-fluoroganciclovir as a PET reporter probe for imaging HSV1-tk gene expression. CONCLUSION: These results demonstrate the feasibility of using [8-14C]-ganciclovir as a reporter probe for the HSV1-tk reporter gene, using an in vivo adenoviral mediated gene delivery system in a murine model. The results form the foundation for further investigation of [8-18F]-fluoroganciclovir for noninvasive and repeated imaging of gene expression with PET.

Brown tumor of the orbit. Case report and review of the literature.

Brown tumors, focal bony lesions of hyperparathyroidism, result from the direct effect of parathyroid hormone on bone. While such lesions are not uncommon in primary hyperparathyroidism, brown tumors have been associated less frequently with secondary hyperparathyroidism and have rarely been described as involving the orbital bones. We have found only four such cases previously reported in the ophthalmic literature. We report a case of orbital involvement by brown tumor in a child with chronic renal failure and secondary hyperparathyroidism. Use of long-term hemodialysis has increased the life span of individuals with chronic renal failure and produced an increased population of patients with secondary hyperparathyroidism and resultant bony changes. The ophthalmologist should consider brown tumor in the differential diagnosis of a patient with chronic renal failure and ocular symptomatology.

Dietary fish oils limit adipose tissue hypertrophy in rats.

Total body mass, fat pad mass, and fat cell size were examined after feeding rats diets containing 20% triglycerides from fish oil or lard. Although food consumption, weight gain, and fat balance on the two diets were similar, lard-fed rats had 77% more fat in perirenal fat pads and 51% more fat in epididymal fat pads compared with fish oil-fed rats. There was no difference between the two groups in fat cell number in each region; however, adipocytes were significantly smaller in fish oil-fed rats. Thus dietary fish oil appears to limit triglyceride accumulation in adipose tissue and thereby limit fat cell trophic growth.