Differential regulation of Sciaenops ocellatus viperin expression by intracellular and extracellular bacterial pathogens

Viperin is an antiviral protein that has been found to exist in diverse vertebrate organisms and is involved in innate immunity against the infection of a wide range of viruses. However, it is largely unclear as to the potential role played by viperin in bacterial infection. In this study, we identified the red drum Sciaenops ocellatus viperin gene (SoVip) and analyzed its expression in relation to bacterial challenge. The complete gene of SoVip is 2570 bp in length and contains six exons and five introns. The open reading frame of SoVip is 1065 bp, which is flanked by a 5'-untranslated region (UTR) of 34 bp and a 3'-UTR of 350 bp. The deduced amino acid sequence of SoVip shares extensive identities with the viperins of several fish species and possesses the conserved domain of the radical S-adenosylmethionine superfamily proteins. Expressional analysis showed that constitutive expression of SoVip was relatively high in blood, muscle, brain, spleen, and liver, and low in kidney, gill, and heart. Experimental challenges with poly(I:C) and bacterial pathogens indicated that SoVip expression in liver was significantly upregulated by poly(I:C) and the fish pathogen Edwardsiella tarda but down-regulated by the fish pathogens Listonella anguillarum and Streptococcus iniae. Similar differential induction patterns were also observed at cellular level with primary hepatocytes challenged with E. tarda, L anguillarum, and S. iniae. Infection study showed that all three bacterial pathogens could attach to cultured primary hepatocytes but only E. tarda was able to invade into and survive in hepatocytes. Together these results indicate that SoVip is involved in host immune response during bacterial infection and is differentially regulated at transcription level by different bacterial pathogens. (C) 2010 Elsevier Ltd. All rights reserved.

Dissection and localization of the immunostimulating domain of Edwardsiella tarda FliC

Bacterial flagellin is known to induce potent immune response in vertebrate systems via the toll-like receptor (TLR) 5. As a result, flagellin has been studied extensively as a vaccine adjuvant. In a previous study, we examined the vaccine and adjuvant potentials of the flagellin (FliC) of the fish pathogen Edwardsiella tarda. We found that E. tarda FliC induced low protective immunity by itself but could function as a molecular adjuvant and potentiate the specific immune response induced by the E. tarda antigen Eta6. Since FliC is a large protein and organized into distinct structural domains, we wondered whether the immunostimulating effect observed with the full-length protein could be localized to a certain region. To investigate this question, we in the present study dissected the FliC protein into several segments according to its structural features: (i) N163, which consists of the conserved N-terminal 163 residues of FliC; (ii) M160, which consists of the variable middle 160 residues; (iii) C94, which consists of the conserved C-terminal 94 residues; (iv) NC257, which is an artificial fusion of N163 and C94. To examine the adjuvanticity of the FliC fragments, DNA vaccine plasmids expressing FliC fragments in fusion with Eta6 were constructed and used to immunize Japanese flounder. The results showed that N163 produced the best adjuvant effect, which, in respect to improvement in the relative percent survival of the vaccinated fish, was comparable to that of the full-length FliC. None of the other FliC fragments exhibited apparent immunopotentiating effect. Further analysis showed that N163 enhanced the production of serum specific antibodies and, like full-length FliC, significantly upregulated the expression of the genes that are possibly involved in innate and adaptive immunity. These results indicate that N163 is the immunodominant region of FliC and suggest that E. tarda FliC may induce immune responses in Japanese flounder via mechanisms alternative to that involving TLR5. (C) 2010 Elsevier Ltd. All rights reserved.

Expression and localization of a novel phosducin-like protein from amphioxus Branchiostoma belcheri

A full length amphioxus cDNA, encoding a novel phosducin-like protein (Amphi-PhLP), was identified for the first time from the gut cDNA library of Branchiostoma belcheri. It is comprised of 1 550 bp and an open reading frame (ORF) of 241 amino acids, with a predicted molecular mass of approximately 28 kDa. In situ hybridization histochemistry revealed a tissue-specific expression pattern of Amphi-PhLP with the high levels in the ovary, and at a lower level in the hind gut and testis, hepatic caecum, gill, endostyle, and epipharyngeal groove, while it was absent in the muscle, neural tube and notochord. In the Chinese Hamster Ovary (CHO) cells transfected with the expression plasmid pEGFP-N1/Amphi-PhLP, the fusion protein was targeted in the cytoplasm of CHO cells, suggesting that Amphi-PhLP is a cytosolic protein. This work may provide a framework for further understanding of the physiological function of Amphi-PhLP in B. belcheri.

Identification, expression, function and localization of a DUF985 domain-containing hypothetical gene from amphioxus Branchiostoma belcheri

The progress in genome sequencing has led to an increasing submission of uncharacterized hypothetical genes with the domain of unknown function, DUF985, in GenBank, and none of these genes is related to a known protein. We therefore underwent an experimental study to identify the function of a DUF985 domain-containing hypothetical gene BbDUF985 (GenBank Accession No. AY273818) isolated from amphioxus Branchiostoma belcheri (B. belcheri). BbDUF985 was successfully expressed in both prokaryotic and eukaryotic systems, and its recombinant proteins expressed in both systems definitely exhibited an activity of phosphoglucose isomerase (PGI). Both tissue-section in situ hybridization and immunohistochemistry demonstrated that BbDUF985 was expressed in a tissue-specific manner, with most abundant levels in the hepatic caecum and ovary. In CHO cells transfected with the expression plasmid pEGFP-N1/BbDUF985, the fusion protein was targeted in the cytoplasm of CHO cells, suggesting that BbDUF985 is a cytosolic protein. In contrast, Western blotting indicated that BbDUF985 was also present in amphioxus humoral fluids, suggesting that it exists as a secreted protein as well. Our study provided a framework for further understanding the biochemical properties and physiological function of DUF985-containing hypothetical proteins in other species. (c) 2008 Elsevier Inc. All rights reserved.

Localization of the electric-field distribution in graded core-shell metamaterials

The local electric-field distribution has been investigated in a core-shell cylindrical metamaterial structure under the illumination of a uniform incident optical field. The structure consists of a homogeneous dielectric core, a shell of graded metal-dielectric metamaterial, embedded in a uniform matrix. In the quasistatic limit, the permittivity of the metamaterial is given by the graded Drude model. The local electric potentials and hence the electric fields have been derived exactly and analytically in terms of hypergeometric functions. Our results showed that the peak of the electric field inside the cylindrical shell can be confined in a desired position by varying the frequency of the optical field and the parameters of the graded profiles. Thus, by fabricating graded metamaterials, it is possible to control electric-field distribution spatially. We offer an intuitive explanation for the gradation-controlled electric-field distribution.

Isolation and characterization of novel marine Roseobacter clade members producing unique intracellular chromium-rich aggregates

The marine Roseobacter clade comprises one of the largest fractions of heterotrophic marine bacteria and accounts for about 16% of 16S rRNA gene clones retrieved from marine bacterioplankton. Their global distribution seems to be related to oceanic water masses and their environmental and biogeochemical properties. In this study, we report isolation and characterization of novel Roseobacter clade members from the Yellow Sea, China. Phylogenetic analysis of 16S rRNA gene sequences reveals that the new isolates (YSCB1, YSCB2, YSCB3 and YSCB4) are closely related to uncultured Arctic seawater bacterium R7967 (99.57-100% sequence identity) and to the cultured Roseobacter sp. DSS-1 (99.27-99.76% sequence identity) isolated from the southeastern coastal water of the USA. Interestingly, YSCB strains possess unique intracellular chromium-containing aggregates. Therefore, these novel Roseobacter clade members exhibit a peculiar property in mineral biogeneration. (c) 2006 Elsevier SAS. All rights reserved.

Intracellular copper/zinc superoxide dismutase from bay scallop Argopecten irradians: Its gene structure, mRNA expression and recombinant protein

Superoxide dismutases are an ubiquitous family of enzymes that function to efficiently catalyze the dismutation of superoxide anions. Two unique and highly compartmentalized bay scallop Argopecten irradians superoxide dismutases: MnSOD and ecCuZnSOD, have been molecularly characterized in our previous study. To complete characterize the SOD family in A. irradians, a novel intracellular copper/zinc SOD from the A. irradians (Ai-icCuZnSOD) was obtained and characterized. The full-length cDNA of Ai-icCuZnSOD was 1047 bp with a 459 bp open reading frame encoding 152 amino acids. The genomic length of the Ai-icCuZnSOD gene was about 4279 bp containing 4 exons and 3 introns. The promoter region containing many putative transcription factor binding sites were analyzed. Furthermore, quantitative reverse transcriptase real-time PCR (qRT-PCR) analysis indicated that the highest expression of the Ai-icCuZnSOD was detected in gill and the expression profiles in hemocytes of bay scallops challenged with bacteria Vibrio anguillarum and lipopolysaccharide (LPS) were different. The result presented an increased expression after injection with LPS whereas no significant changes were observed after V. anguillarum injection. A fusion protein containing Ai-icCuZnSOD was produced in vitro. The rAi-icCuZnSOD is a stable enzyme, retaining more than 80% of its activity between 10 and 60 degrees C and keeping above 88% of its activity at pH values between 5.8 and 9. Ai-icCuZnSOD is more stable under alkaline than acidic conditions. Crown Copyright (C) 2009 Published by Elsevier Ltd. All rights reserved.

Feature Matching for Object Localization in the Presence of Uncertainty

We consider the problem of matching model and sensory data features in the presence of geometric uncertainty, for the purpose of object localization and identification. The problem is to construct sets of model feature and sensory data feature pairs that are geometrically consistent given that there is uncertainty in the geometry of the sensory data features. If there is no geometric uncertainty, polynomial-time algorithms are possible for feature matching, yet these approaches can fail when there is uncertainty in the geometry of data features. Existing matching and recognition techniques which account for the geometric uncertainty in features either cannot guarantee finding a correct solution, or can construct geometrically consistent sets of feature pairs yet have worst case exponential complexity in terms of the number of features. The major new contribution of this work is to demonstrate a polynomial-time algorithm for constructing sets of geometrically consistent feature pairs given uncertainty in the geometry of the data features. We show that under a certain model of geometric uncertainty the feature matching problem in the presence of uncertainty is of polynomial complexity. This has important theoretical implications by demonstrating an upper bound on the complexity of the matching problem, an by offering insight into the nature of the matching problem itself. These insights prove useful in the solution to the matching problem in higher dimensional cases as well, such as matching three-dimensional models to either two or three-dimensional sensory data. The approach is based on an analysis of the space of feasible transformation parameters. This paper outlines the mathematical basis for the method, and describes the implementation of an algorithm for the procedure. Experiments demonstrating the method are reported.

Localization and Positioning Using Combinations of Model Views

A method for localization and positioning in an indoor environment is presented. The method is based on representing the scene as a set of 2D views and predicting the appearances of novel views by linear combinations of the model views. The method is accurate under weak perspective projection. Analysis of this projection as well as experimental results demonstrate that in many cases it is sufficient to accurately describe the scene. When weak perspective approximation is invalid, an iterative solution to account for the perspective distortions can be employed. A simple algorithm for repositioning, the task of returning to a previously visited position defined by a single view, is derived from this method.

Dependency-Directed Localization of Software Bugs

Software bugs are violated specifications. Debugging is the process that culminates in repairing a program so that it satisfies its specification. An important part of debugging is localization, whereby the smallest region of the program that manifests the bug is found. The Debugging Assistant (DEBUSSI) localizes bugs by reasoning about logical dependencies. DEBUSSI manipulates the assumptions that underlie a bug manifestation, eventually localizing the bug to one particular assumption. At the same time, DEBUSSI acquires specification information, thereby extending its understanding of the buggy program. The techniques used for debugging fully implemented code are also appropriate for validating partial designs.

Automated Assembly Using Feature Localization

Automated assembly of mechanical devices is studies by researching methods of operating assembly equipment in a variable manner; that is, systems which may be configured to perform many different assembly operations are studied. The general parts assembly operation involves the removal of alignment errors within some tolerance and without damaging the parts. Two methods for eliminating alignment errors are discussed: a priori suppression and measurement and removal. Both methods are studied with the more novel measurement and removal technique being studied in greater detail. During the study of this technique, a fast and accurate six degree-of-freedom position sensor based on a light-stripe vision technique was developed. Specifications for the sensor were derived from an assembly-system error analysis. Studies on extracting accurate information from the sensor by optimally reducing redundant information, filtering quantization noise, and careful calibration procedures were performed. Prototype assembly systems for both error elimination techniques were implemented and used to assemble several products. The assembly system based on the a priori suppression technique uses a number of mechanical assembly tools and software systems which extend the capabilities of industrial robots. The need for the tools was determined through an assembly task analysis of several consumer and automotive products. The assembly system based on the measurement and removal technique used the six degree-of-freedom position sensor to measure part misalignments. Robot commands for aligning the parts were automatically calculated based on the sensor data and executed.

Qualitative Knowledge, Casual Reasoning and the Localization of Failures

This report investigates some techinques appropriate to representing the knowledge necessary for understanding a class of electronic machines -- radio receivers. A computational performance model - WATSON - is presented. WATSONs task is to isolate failures in radio receivers whose principles of operation have been appropriately described in his knowledge base. The thesis of the report is that hierarchically organized representational structures are essential to the understanding of complex mechanisms. Such structures lead not only to descriptions of machine operation at many levels of detail, but also offer a powerful means of organizing "specialist" knowledge for the repair of machines when they are broken.

Simultaneous Localization and Recognition of Dynamic Hand Gestures

A framework for the simultaneous localization and recognition of dynamic hand gestures is proposed. At the core of this framework is a dynamic space-time warping (DSTW) algorithm, that aligns a pair of query and model gestures in both space and time. For every frame of the query sequence, feature detectors generate multiple hand region candidates. Dynamic programming is then used to compute both a global matching cost, which is used to recognize the query gesture, and a warping path, which aligns the query and model sequences in time, and also finds the best hand candidate region in every query frame. The proposed framework includes translation invariant recognition of gestures, a desirable property for many HCI systems. The performance of the approach is evaluated on a dataset of hand signed digits gestured by people wearing short sleeve shirts, in front of a background containing other non-hand skin-colored objects. The algorithm simultaneously localizes the gesturing hand and recognizes the hand-signed digit. Although DSTW is illustrated in a gesture recognition setting, the proposed algorithm is a general method for matching time series, that allows for multiple candidate feature vectors to be extracted at each time step.

Experimental impulse radio IEEE 802.15.4a UWB based wireless sensor localization technology: Characterization, reliability and ranging

Ultra Wide Band (UWB) transmission has recently been the object of considerable attention in the field of next generation location aware wireless sensor networks. This is due to its fine time resolution, energy efficient and robustness to interference in harsh environments. This paper presents a thorough applied examination of prototype IEEE 802.15.4a impulse UWB transceiver technology to quantify the effect of line of sight (LOS) and non line of sight (NLOS) ranging in real indoor and outdoor environments. Results included draw on an extensive array of experiments that fully characterize the 802.15.4a UWB transceiver technology, its reliability and ranging capabilities for the first time. A new two way (TW) ranging protocol is proposed. The goal of this work is to validate the technology as a dependable wireless communications mechanism for the subset of sensor network localization applications where reliability and precision positions are key concerns.