g Boid inclusion body disease (BIBD) is a progressive, usually fatal disease of constrictor snakes, characterized by cytoplasmic inclusion bodies (IB) in a wide range of cell types. To identify the causative agent of the disease, we established cell cultures from BIBD-positive and -negative boa constrictors. The IB phenotype was maintained in cultured cells of affected animals, and supernatants from these cultures caused the phenotype in cultures originating from BIBD-negative snakes. Viruses were purified from the supernatants by ultracentrifugation and subsequently identified as arenaviruses. Purified virus also induced the IB phenotype in naive cells, which fulfilled Koch's postulates in vitro. One isolate, tentatively designated University of Helsinki virus (UHV), was studied in depth. Sequencing confirmed that UHV is a novel arenavirus species that is distinct from other known arenaviruses including those recently identified in snakes with BIBD. The morphology of UHV was established by cryoelectron tomography and subtomographic averaging, revealing the trimeric arenavirus spike structure at 3.2-nm resolution. Immunofluorescence, immunohistochemistry, and immunoblotting with a polyclonal rabbit antiserum against UHV and reverse transcription-PCR (RT-PCR) revealed the presence of genetically diverse arenaviruses in a large cohort of snakes with BIBD, confirming the causative role of arenaviruses. Some snakes were also found to carry arenavirus antibodies. Furthermore, mammalian cells (Vero E6) were productively infected with UHV, demonstrating the potential of arenaviruses to cross species barriers. In conclusion, we propose the newly identified lineage of arenaviruses associated with BIBD as a novel taxonomic entity, boid inclusion body disease-associated arenaviruses (BIBDAV), in the family Arenaviridae.

Human respiratory syncytial virus is a human pathogen that causes severe infection of the respiratory tract. Current information about the structure of the virus and its interaction with host cells is limited. We carried out an electron cryotomographic characterization of cell culture-grown human respiratory syncytial virus to determine the architecture of the virion. The particles ranged from 100 nm to 1,000 nm in diameter and were spherical, filamentous, or a combination of the two. The filamentous morphology correlated with the presence of a cylindrical matrix protein layer linked to the inner leaflet of the viral envelope and with local ordering of the glycoprotein spikes. Recombinant viruses with only the fusion protein in their envelope showed that these glycoproteins were predominantly in the postfusion conformation, but some were also in the prefusion form. The ribonucleocapsids were left-handed, randomly oriented, and curved inside the virions. In filamentous particles, they were often adjacent to an intermediate layer of protein assigned to M2-1 (an envelopeassociated protein known to mediate association of ribonucleocapsids with the matrix protein). Our results indicate important differences in structure between the Paramyxovirinae and Pneumovirinae subfamilies within the Paramyxoviridae, and provide fresh insights into host cell exit of a serious pathogen.cryo-ET | paramyxovirus | virus structure H uman respiratory syncytial virus (HRSV) causes severe disease, especially in children and the elderly (1, 2). In a study covering data from 2005, it was found that HRSV was responsible for 22% of acute lower respiratory infections worldwide and 66,000-199,000 deaths in children under the age of 5 y (2). Treatment is currently limited to purine analogs (Ribavirin) and passive immune-prophylaxis with humanized monoclonal antibodies (Palivizumab) (3).RSV belongs to the Paramyxoviridae, a large family of enveloped, negative-sense RNA viruses with many members from humans and animals. Together with human metapneumovirus, it constitutes the Pneumovirinae subfamily. Like other paramyxoviruses, it has a plasma-membrane-derived lipid envelope and a helical ribonucleocapsid (RNP) that contains a single viral RNA molecule associated with nucleoproteins (N), and an RNA-dependent RNA polymerase. The virion contains, in addition, a matrix protein (M) (4-7) and a transcription antiterminator, M2-1 (an envelopeassociated protein known to mediate association of RNPs with the M protein) (8-11). The lipid envelope contains two transmembrane glycoproteins: the fusion protein, F, and a glycoprotein, G, used for attachment to host cells. In addition, HRSV contains a small hydrophobic protein, SH, of unclear function and proteins of cellular origin, such as actin and chaperones (12). The RNP and F have been analyzed using 3D-EM and X-ray crystallography (13,14).HRSV particles are pleomorphic with both spherical and filamentous particles of different sizes (15). Early electron microscopy studies showed the presence of a helical p...

bOur understanding of the third domain of life, Archaea, has greatly increased since its establishment some 20 years ago. The increasing information on archaea has also brought their viruses into the limelight. Today, about 100 archaeal viruses are known, which is a low number compared to the numbers of characterized bacterial or eukaryotic viruses. Here, we have performed a comparative biological and structural study of seven pleomorphic viruses infecting extremely halophilic archaea. The pleomorphic nature of this novel virion type was established by sedimentation analysis and cryo-electron microscopy. These nonlytic viruses form virions characterized by a lipid vesicle enclosing the genome, without any nucleoproteins. The viral lipids are unselectively acquired from host cell membranes. The virions contain two to three major structural proteins, which either are embedded in the membrane or form spikes distributed randomly on the external membrane surface. Thus, the most important step during virion assembly is most likely the interaction of the membrane proteins with the genome. The interaction can be driven by single-stranded or double-stranded DNA, resulting in the virions having similar architectures but different genome types. Based on our comparative study, these viruses probably form a novel group, which we define as pleolipoviruses.

Measles virus is a highly infectious, enveloped, pleomorphic virus. We combined electron cryotomography with subvolume averaging and immunosorbent electron microscopy to characterize the 3D ultrastructure of the virion. We show that the matrix protein forms helices coating the helical ribonucleocapsid rather than coating the inner leaflet of the membrane, as previously thought. The ribonucleocapsid is folded into tight bundles through matrixmatrix interactions. The implications for virus assembly are that the matrix already tightly interacts with the ribonucleocapsid in the cytoplasm, providing a structural basis for the previously observed regulation of RNA transcription by the matrix protein. Next, the matrix-covered ribonucleocapsids are transported to the plasma membrane, where the matrix interacts with the envelope glycoproteins during budding. These results are relevant to the nucleocapsid organization and budding of other paramyxoviruses, where isolated matrix has been observed to form helices.image reconstruction | subtomogram averaging