Recombinant Subgroup B Human Respiratory Syncytial Virus Expressing Enhanced Green Fluorescent Protein Efficiently Replicates in Primary Human Cells and Is Virulent in Cotton Rats

Human respiratory syncytial virus (HRSV) is the most important viral cause of severe respiratory tract disease in infants. Two subgroups (A and B) have been identified, which cocirculate during, or alternate between, yearly epidemics and cause indistinguishable disease. Existing in vitro and in vivo models of HRSV focus almost exclusively on subgroup A viruses. Here, a recombinant (r) subgroup B virus (rHRSV(B05)) was generated based on a consensus genome sequence obtained directly from an unpassaged clinical specimen from a hospitalized infant. An additional transcription unit containing the gene encoding enhanced green fluorescent protein (EGFP) was introduced between the phosphoprotein and matrix genes (position 5) of the genome to generate rHRSV(B05)EGFP(5). The recombinant viruses replicated efficiently in both HEp-2 cells and in well-differentiated normal human bronchial cells grown at air-liquid interface. Intranasal infection of cotton rats (Sigmodon hispidus) resulted in high numbers of EGFP(+) cells in epithelia of the nasal septum and conchae. When administered in a relatively large inoculum volume, the virus also replicated efficiently in bronchiolar epithelial cells and spread extensively in both the upper and lower respiratory tracts. Virus replication was not observed in ciliated epithelial cells of the trachea. This is the first virulent rHRSV strain with the genetic composition of a currently circulating wild-type virus. In vivo tracking of infected cells by means of EGFP fluorescence in the absence of cytopathic changes increases the sensitivity of virus detection in HRSV pathogenesis studies.

IMPORTANCE

Virology as a discipline has depended on monitoring cytopathic effects following virus culture in vitro. However, wild-type viruses isolated from patients often do not cause significant changes to infected cells, necessitating blind passage. This can lead to genetic and phenotypic changes and the generation of high-titer, laboratory-adapted viruses with diminished virulence in animal models of disease. To address this, we determined the genome sequence of an unpassaged human respiratory syncytial virus from a sample obtained directly from an infected infant, assembled a molecular clone, and recovered a wild-type recombinant virus. Addition of a gene encoding enhanced green fluorescent protein allowed this wild-type virus to be tracked in primary human cells and living animals in the absence of significant cytopathic effects. Imaging of fluorescent cells proved to be a highly valuable tool for monitoring the spread of virus and may help improve assays for evaluating novel intervention strategies.

Characterization and Modeling of the Indoor Radio Channel at 868 MHz for a Mobile Bodyworn Wireless Personal Area Network

This letter reports the statistical characterization and modeling of the indoor radio channel for a mobile wireless personal area network operating at 868 MHz. Line of sight (LOS) and non-LOS conditions were considered for three environments: anechoic chamber, open office area and hallway. Overall, the Nakagami-m cdf best described fading for bodyworn operation in 60% of all measured channels in anechoic chamber and open office area environments. The Nakagami distribution was also found to provide a good description of Rician distributed channels which predominated in the hallway. Multipath played an important role in channel statistics with the mean recorded m value being reduced from 7.8 in the anechoic chamber to 1.3 in both the open office area and hallway.

Higher order statistics for lognormal small-scale fading in mobile radio channels

This letter reports the statistical characterization and modeling of the indoor radio channel for a mobile wireless personal area network operating at 868 MHz. Line of sight (LOS) and non-LOS conditions were considered for three environments: anechoic chamber, open office area and hallway. Overall, the Nakagami-m cdf best described fading for bodyworn operation in 60% of all measured channels in anechoic chamber and open office area environments. The Nakagami distribution was also found to provide a good description of Rician distributed channels which predominated in the hallway. Multipath played an important role in channel statistics with the mean recorded m value being reduced from 7.8 in the anechoic chamber to 1.3 in both the open office area and hallway.

Higher order statistics for the κ–μ distribution

Closed-form expressions for the level crossing rate and average fade duration of a kappa–mu distributed fading signal envelope are presented. The proposed equations are validated by reduction to known Rice, Rayleigh and Nakagami-m special cases. They are also compared with measured data obtained from field trials analysing human body to body radio channels and shown to provide good agreement.

Rarity and decline in bumblebees - A test of causes and correlates in the Irish fauna

Bees are believed to be in decline across many of the world's ecosystems. Recent studies on British bumblebees proposed alternative theories to explain declines. One study suggested that greater dietary specialization among the rarer bumblebee species makes them more susceptible to decline. A second study disputed this theory and found that declines in British bumblebees were correlated with the size of species' European ranges, leading to the suggestion that climate and habitat specialization may be better indicators of the risk of decline. Here we use a new and independent dataset based on Irish bumblebees to test the generality of these theories. We found that most of the same bumblebee species are declining across the British Isles, but that, within Ireland, a simple food-plant specialization model is inadequate to explain these declines. Furthermore, we found no evidence of a relationship between declines in Irish bumblebees and the size of species' European ranges. However, we demonstrate that the late emerging species have declined in Ireland (and in Britain), and that these species show a statistically significant westward shift to the extremity of their range, probably as a result of changing land use. Irish data support the finding that rare and declining bumblebees are later nesting species, associated with open grassy habitats. We suggest that the widespread replacement of hay with silage in the agricultural landscape, which results in earlier and more frequent mowing and a reduction in late summer wildflowers, has played a major role in bumblebee declines. (C) 2006 Elsevier Ltd. All rights reserved.

Absolute configuration of conformationally flexible cis-dihydrodiol metabolites by the method of confrontation of experimental and calculated electronic CD spectra and optical rotations

We have determined the absolute configurations of conformationally flexible cis-dihydrodiol metabolites (cis-1,2-dihydroxy-3,5-cyclohexadienes), bearing different substituents (e.g., Br, F, CF3, CN, Me) in 3- and 5-positions, by the method of confrontation of experimental and calculated electronic CD spectra and optical rotations. Convergent results were obtained by both methods in eight out of ten cases. For the difficult cases, where either conformer population and/or chiroptical properties (calculated rotational strengths of the long-wavelength Cotton effect or optical rotations) of contributing conformers remain inconclusive, the absolute configuration could still be correctly assigned based on one of the biased properties (either ECD or optical rotation). This approach appears well-suited for a broad spectrum of conformationally flexible chiral molecules.

Genome sequence of the metazoan plant-parasitic nematode Meloidogyne incognita

Plant-parasitic nematodes are major agricultural pests worldwide and novel approaches to control them are sorely needed. We report the draft genome sequence of the root-knot nematode Meloidogyne incognita, a biotrophic parasite of many crops, including tomato, cotton and coffee. Most of the assembled sequence of this asexually reproducing nematode, totaling 86 Mb, exists in pairs of homologous but divergent segments. This suggests that ancient allelic regions in M. incognita are evolving toward effective haploidy, permitting new mechanisms of adaptation. The number and diversity of plant cell wall-degrading enzymes in M. incognita is unprecedented in any animal for which a genome sequence is available, and may derive from multiple horizontal gene transfers from bacterial sources. Our results provide insights into the adaptations required by metazoans to successfully parasitize immunocompetent plants, and open the way for discovering new antiparasitic strategies.

Integrated structure selection and parameter optimisation for eng-genes neural models

The eng-genes concept involves the use of fundamental known system functions as activation functions in a neural model to create a 'grey-box' neural network. One of the main issues in eng-genes modelling is to produce a parsimonious model given a model construction criterion. The challenges are that (1) the eng-genes model in most cases is a heterogenous network consisting of more than one type of nonlinear basis functions, and each basis function may have different set of parameters to be optimised; (2) the number of hidden nodes has to be chosen based on a model selection criterion. This is a mixed integer hard problem and this paper investigates the use of a forward selection algorithm to optimise both the network structure and the parameters of the system-derived activation functions. Results are included from case studies performed on a simulated continuously stirred tank reactor process, and using actual data from a pH neutralisation plant. The resulting eng-genes networks demonstrate superior simulation performance and transparency over a range of network sizes when compared to conventional neural models. (c) 2007 Elsevier B.V. All rights reserved.

Millimeter-Wave Soldier-to-Soldier Communications for Covert Battlefield Operations

Mobile ad hoc networking of dismounted combat personnel is expected to play an important role in the future of network-centric operations. High-speed, short-range, soldier-to-soldier wireless communications will be required to relay information on situational awareness, tactical instructions, and covert surveillance related data during special operations reconnaissance and other missions. This article presents some of the work commissioned by the U. K. Ministry of Defence to assess the feasibility of using 60 GHz millimeter-wave smart antenna technology to provide covert communications capable of meeting these stringent networking needs. Recent advances in RF front-end technology, alongside physical layer transmission schemes that could be employed in millimeter-wave soldier-mounted radio, are discussed. The introduction of covert communications between soldiers will require the development of a bespoke directive medium access layer. A number of adjustments to the IEEE 802.11 distribution coordination function that will enable directional communications are suggested. The successful implementation of future smart antenna technologies and direction of arrival-based protocols will be highly dependent on thorough knowledge of transmission channel characteristics prior to deployment. A novel approach to simulating dynamic soldier-to-soldier signal propagation using state-of-the-art animation-based technology developed for computer game design is described, and important channel metrics such as root mean square angle and delay spread for a team of four networked infantry soldiers over a range of indoor and outdoor environments is reported.