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Between January and March 1971, an unusual outbreak of hemorrhagic fever occurred in Cochabamba, Bolivia. The index case was apparently infected elsewhere, but became ill in Cochabamba, where hemorrhagic fever had never been reported. Four secondary cases occurred among hospital contacts of the index case, and a pathologist was infected after cutting himself during an autopsy of one of the secondary cases. Clinical manifestations were typical of Bolivian hemorrhagic fever, except that jaundice occurred in three cases. An arenavirus of the Tacaribe complex was isolated from three cases. One isolate was studied in detail and shown to be closely related antigenically to the prototype strains of Machupo virus.

The dihydrochloride salt of ( S,S )-1,2-bis(5-methoxy-2-benzimidazolyl)-1,2-ethandiol (A37536) inhibits the synthesis of lymphocytic choriomeningitis (LCM), Parana, and Pichinde viruses in L-929 cells. The compound has no direct inactivating effect on LCM virus nor does it affect the adsorption of LCM virus to L cells. The drug-cell interaction is slow. Maximal activity is observed only by exposing cells to the drug at least 8 h prior to LCM virus infection, or by concomitant drug treatment and infection at a low multiplicity. Addition of serum-free media to L cells after LCM virus infection diminishes the activity of A37536. Whereas A37536 exhibits its antiviral activity at concentrations that have little or no effect on L cell division rate, a marked change can be noted in the cell's sensitivity to lysis by standard trypsin dispersal procedures. A37536 has no specific antiviral activity in LCM virus-infected BHK, HeLa, or Vero cells. All of the four tested derivatives of A37536 showed antiviral activity against LCM virus but only at concentrations that reduced the growth rate or were toxic to L cells.

SummaryPichinde virus, a virus capable of producing a chronic infection, was found to replicate in Vero cells and to form plaques in these cells. The virus had a latent period of approximately 8 hr and maximum titers were reached by 24 to 36 hr after infection. Adsorption of the virus was enhanced in the presence of 10 to 100 μg/ml of DEAE-dextran. Pichinde virus was found to be relatively thermostable, stable between pH 5.5 to 9.0 and sensitive to ethyl ether. Virus replication was not effected by the DNA inhibitors, IudR and ara-C; however, virus replication was inhibited by actinomycin D at concentrations of 1 and 4 μg/ml. The virus was stable in sucrose and cesium chloride and densities of 1.18 and 1.19 to 1.20 g/cm3, respectively, were obtained in gradients made of these two substances.

Summary The existence of defective interfering (DI) particles has been well documented for a number of virus systems (Huang & Baltimore, 1970). Such particles are currently of great interest since their ability to cause homologous interference has been implicated as a possible regulatory factor in the establishment of chronic or persistent virus disease (Huang & Baltimore, 1970). We have recently reported that about 50 cell generations after infection of BHK 21/13S cells with either lymphocytic choriomeningitis virus (LCMV) or Parana virus, infective virus could no longer be detected in either the cells or the culture medium (Staneck et al. 1972). However, LCMV-infected cultures continued to produce particles with an interference activity which closely resembled DI virus (Welsh, O'Connell & Pfau, 1972). The data presented here indicate that Parana, another arenavirus (Rowe et al. 1970), can initiate the nearly exclusive synthesis of DI virus under the same conditions.

Use of recently developed plaque assays for LCM virus-infected cells and free virus has led to a better understanding of the in vitro host-virus interactions. Many LCM virus stocks, even at high input multiplicities of infection, have very low efficiencies of infecting cells. Under these conditions where multiple cycles of infection are necessary to achieve total infectivity of a BHK21/13 S monolayer there appears to be a preferential cell to cell transfer of the viral genome. The low efficiency of virus stocks in forming infective centers is due to an interference phenomenon that can be readily demonstrated using an L cell monolayer plaque assay. The properties of the interfering agent in recently infected cultures as well as in long-term infections are those of defective interfering virus. Further studies to elucidate the host-virus relationship in the regulation of virus production have shown that conditions favoring cell replication enhance virus replication or maturation.

Procedures for optimal infection of BHK21 cells by LCM virus are described. LCM virus could be purified by ammonium sulfate precipitation of the virus-containing medium followed by centrifugation in sucrose gradients, whereby the buoyant density of the virus was estimated to be 1.18 g/ml. The nucleic acids of the LCM virus were studied by centrifugation in sucrose gradients and by acrylamide gel electrophoresis. These analyses have led to the detection of 7 different RNA molecules in LCM virus with sedimentation values of 31S, 28S, 23S, 18S, 5.5S, 5S and 4S. Only the 31S and the 23S RNA’s appear to be virus-specific; the 28S and the 18S RNA‘s originate from ribosomes located inside the virion and the small 5.5S, 5S and 4S RNA’s are probably host RNA’s associated with these ribosomes. Preliminary protein analysis revealed 1 major and 5 smaller components. Thus, both the nucleic acid and the protein composition of the LCM virus, which is the prototype of the arenavirus group, seem to be different from other enveloped RNA viruses.

Pichinde virus, a member of the arenavirus group, was found to contain single-stranded RNA. The RNA consisted of 5 discernible components corresponding to 31S, 28S, 22S, 18S and 4 to 6S. The 28S and 18S RNA’s appeared to be of host cell origin and the 4 to 6S RNA was probably from the same source. The 31S and 22S RNA’s appeared to be components which coded for viral products. The virus was found to contain 4 structural polypeptides. Two were glycopeptides and 2 were polypeptides. One polypeptide (VI) and 1 glycopeptide (VII) remained associated with the viral RNA after treating the virus with nonionic detergent, while the other glycopeptide (VIII) was removed with such treatment. VIV was a minor peptide and its fate following treatment of the virus with nonionic detergent could not be determined.

L cells, infected with strain WE LCM virus, were examined by electron microscopy. Whereas the ultrastructure of nuclei was essentially normal, both cytoplasm and plasma membrane were markedly altered, which was demonstrable as early as 16 h after infection. Changes consisted of increasing density of mitochondria, the formation of membranous structures appearing as multilayered concentrically arrayed lamellae or as irregular convolutions, and the appearance of granules either singly or accumulated in masses which were sometimes as large and delineated as cytoplasmic inclusions. These granules resembled closely those which characteristically are found in the interior of arenavirus particles. Higher magnification revealed that they were composed of granular and/or rod-like subunits.

Bolivian hemorrhagic fever (BHF) was first recognized in the Province of Beni, Bolivia, in 1959 (1), and cases have continued to occur since then. Recent outbreaks have confirmed the fact that clinical manifestations can be quite nonspecific. Laboratory diagnosis is handicapped by a scarcity of facilities in the endemic region, as well as by the hazard to nonimmune personnel of working with the causative agent, Machupo virus. These problems also apply in varying degree to two other diseases caused by members of the arenavirus group (2, 3): Argentine hemorrhagic fever (AHF) caused by Junin virus (4) and Lassa fever caused by Lassa virus (5). The early recognition of outbreaks of these diseases is important since rodent control, immune serum therapy, and isolation of patients are thought to be of some use in treatment and prevention.A plaque-neutralization test for detection of Machupo virus antibodies has been developed and proved to be specific and sensitive (6). However, these advantages are posited on t...

The morphology and morphogenesis of all arenaviruses are remarkably similar: 1) virus particles are round or pleomorphic and vary widely in size (60 to 350 nm diameter), 2) they have a membranous envelope and surface projections, and 3) they have characteristic ribosome-like granules in varying numbers within an u~structured interior. Viral morphogenesis occurs via budding upon plasma membranes. Despite the distinctiveness of these virion characteristics, ultrastructural study of in vivo arenavirus infections proved difficult. In salivary gland cells of Calomys callosus, Machupo virus clearly matured upon plasma membranes and yielded salivary secretions containing large numbers of particles. However, in other tissues, such as lymph nodes, thymus, spleen, and central nervous system, where widespread infection of particular host species was anticipated from immunofluorescence, only modest numbers of virus particles were found. Inclusion bodies, easily identified in infected cells in culture, were impossible to discern in vivo. Some cytopathology was spatially associated with virus particles in lymphoid tissues. Ultrastructural study of in vivo arenavirus infections has proven excessively difficult without an immuno-electron microscopic contribution.

Tamiami virus, a member of the arenavirus group, produces an acute central nervous system disorder in suckling mice characterized by cerebellar ataxia and less frequently by paralysis, convulsions, and death. Histopathologic changes in acutely ill mice include vasculitis, meningitis, and impaired migration of the external granule cell layer of the cerebellum and, in survivors, asymptomatic cerebellar heterotopia. All animals carry virus in their brains for at least 2 months following infection, as detected by immunofluorescence and virus titration.