TNFalpha, interferon, and stress response induction as a function of age-related susceptibility to fatal Sindbis virus infection of mice.
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The age-related acquisition of resistance to fatal Sindbis virus infection was examined using a molecularly cloned laboratory strain of the AR339 isolate designated TRSB. TRSB caused 100% mortality in mice up to 5 days of age. Resistance to fatal infection developed abruptly between 5 and 9 days of age. Lethal Sindbis virus infection of mice inoculated at 4 days of age was characterized by high levels of virus replication, induction of high levels of interferon-alpha/beta and TNF-alpha and severe thymic involution indicative of a systemic stress response. These changes correlated with predominantly noninflammatory lesions. In contrast, TRSB infection of older mice was characterized by survival, more limited virus replication, reduced cytokine induction, and the development of inflammatory responses leading to encephalitis, myositis, and myocarditis. Previous studies utilized infections of neonatal mice with TRSB and an attenuated mutant of TRSB to compare fatal and nonfatal Sindbis infection (Trgovcich et al., 1996. Virology 224, 73-83). The experiments reported here utilize mouse age at the time of infection to create conditions for examination of fatal and nonfatal TRSB infections. Both experiments suggest that fatal infection is associated with a shock-like syndrome and little or no inflammatory pathology, while survival is correlated with greatly reduced cytokine levels and inflammatory lesions. (+info)
Infection of neonatal mice with sindbis virus results in a systemic inflammatory response syndrome.
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Laboratory strains of viruses may contain cell culture-adaptive mutations which result in significant quantitative and qualitative alterations in pathogenesis compared to natural virus isolates. This report suggests that this is the case with Sindbis virus strain AR339. A cDNA clone comprising a consensus sequence of Sindbis virus strain AR339 has been constructed (W. B. Klimstra, K. D. Ryman, and R. E. Johnston, J. Virol. 72:7357-7366, 1998). This clone (pTR339) regenerates a sequence predicted to be very close to that of the original AR339 isolate by eliminating several cell culture-adaptive mutations present in individual laboratory strains of the virus (K. L. McKnight et al., J. Virol. 70:1981-1989, 1996). It thus provides a unique reagent for study of the pathogenesis of Sindbis virus strain AR339 in mice. Neonatal mouse pathogenesis of virus (TR339) generated from the pTR339 clone was compared with that of virus from a cDNA clone of the cell culture-passaged laboratory AR339 strain, TRSB, and virus from a clone of a more highly cell culture-adapted strain, HR(sp) (Toto 50). The sequence of TRSB differs from the consensus at three coding positions, while Toto 50 differs at eight codons and one nucleotide in the 5' nontranslated region. Both cell culture-adapted strains contain mutations associated with heparan sulfate (HS)-dependent attachment to cells (W.B. Klimstra, K. D. Ryman, and R. E. Johnston, J. Virol. 72:7357-7366, 1998). TR339 caused 100% mortality with an average survival time (AST) of 1.7 +/- 0.25 days. While TRSB also caused 100% mortality, the AST was extended to 2.9 +/- 0.52 days. The more extensively cell culture-adapted virus Toto 50 caused only 30% mortality with an AST extended to 11.0 +/- 4.8 days. TRSB and TR339 induced high serum levels of alpha/beta interferon, gamma interferon, tumor necrosis factor alpha, interleukin-6, and corticosterone and induced pathology reminiscent of lipopolysaccharide-induced endotoxic shock, a type of systemic inflammatory response syndrome. However, the reduced intensity of this response in TRSB-infected mice correlated with the increased AST. Toto 50 failed to induce the shock-like cytokine cascade. In situ hybridization studies indicated that TR339 and TRSB replicated in identical tissues, but the TRSB signal was less widespread at early times postinfection. While Toto 50 also replicated in similar tissues, the extent of replication was severely restricted and mice developed lesions characteristic of encephalitis. A single mutation in TRSB at E2 position 1 (Arg) conferred HS-dependent attachment to cells and was associated with reduced cytokine induction and extended AST in vivo. (+info)
Macrophage-induced muscle pathology results in morbidity and mortality for Ross River virus-infected mice.
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Ross River virus (RRV) is an Australian alphavirus that is often responsible for chronic epidemic polyarthritis and myalgia in humans. Past studies have shown severe disruption of striated muscle fibers to be prominent in RRV pathology in mice; in the present study, macrophages were directly implicated as the primary mediators of muscle damage. General immunosuppressive therapies had only minor effects on mortality and morbidity in RRV-infected mice, with no inhibition of muscle damage. Treatment of mice with macrophage-toxic agents (e.g., silica) prior to RRV infection completely abrogated disease symptoms without significantly affecting titers of virus in organs. Further studies found that clinical signs of infection and muscle damage correlated with a massive influx of macrophages into hind leg muscle, whereas no such infiltrate or damage was observed for silica-treated mice. These observations are significant for the human disease context, as monocytic cells have been detected in the synovial effusions of persons with epidemic polyarthritis. (+info)
Large-plaque mutants of Sindbis virus show reduced binding to heparan sulfate, heightened viremia, and slower clearance from the circulation.
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Laboratory strains of Sindbis virus must bind to the negatively charged glycosaminoglycan heparan sulfate in order to efficiently infect cultured cells. During infection of mice, however, we have frequently observed the development of large-plaque viral mutants with a reduced ability to bind to heparan sulfate. Sequencing of these mutants revealed changes of positively charged amino acids in putative heparin-binding domains of the E2 glycoprotein. Recombinant viruses were constructed with these changes as single amino acid substitutions in a strain Toto 1101 background. All exhibited decreased binding to heparan sulfate and had larger plaques than Toto 1101. When injected subcutaneously into neonatal mice, large-plaque viruses produced higher-titer viremia and often caused higher mortality. Because circulating heparin-binding proteins are known to be rapidly sequestered by tissue heparan sulfate, we measured the kinetics of viral clearance following intravenous injection. Much of the parental small-plaque Toto 1101 strain of Sindbis virus was cleared from the circulation by the liver within minutes, in contrast to recombinant large-plaque viruses, which had longer circulating half-lives. These findings indicate that a decreased ability to bind to heparan sulfate allows more efficient viral production in vivo, which may in turn lead to increased mortality. Because Sindbis virus is only one of a growing number of viruses from many families which have been shown to bind to heparan sulfate, these results may be generally applicable to the pathogenesis of such viruses. (+info)
Re-emergence of Chikungunya and O'nyong-nyong viruses: evidence for distinct geographical lineages and distant evolutionary relationships.
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Chikungunya (CHIK) virus is a member of the genus Alphavirus in the family TOGAVIRIDAE: Serologically, it is most closely related to o'nyong-nyong (ONN) virus and is a member of the Semliki Forest antigenic complex. CHIK virus is believed to be enzootic throughout much of Africa and historical evidence indicates that it spread to other parts of the world from this origin. Strains from Africa and Asia are reported to differ biologically, indicating that distinct lineages may exist. To examine the relatedness of CHIK and ONN viruses using genetic data, we conducted phylogenetic studies on isolates obtained throughout Africa and Southeast Asia. Analyses revealed that ONN virus is indeed distinct from CHIK viruses, and these viruses probably diverged thousands of years ago. Two distinct CHIK virus lineages were delineated, one containing all isolates from western Africa and the second comprising all southern and East African strains, as well as isolates from Asia. Phylogenetic trees corroborated historical evidence that CHIK virus originated in Africa and subsequently was introduced into Asia. Within the eastern Africa and southern Africa/Asia lineage, Asian strains grouped together in a genotype distinct from the African groups. These different geographical genotypes exhibit differences in their transmission cycles: in Asia, the virus appears to be maintained in an urban cycle with Aedes aegypti mosquito vectors, while CHIK virus transmission in Africa involves a sylvatic cycle, primarily with AE: furcifer and AE: africanus mosquitoes. (+info)
Langerhans cells migrate to local lymph nodes following cutaneous infection with an arbovirus.
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Whereas there has been recent interest in interactions between dendritic cells and pathogenic viruses, the role of dendritic cells in the initiation of protective immunity to such organisms has not been elucidated. The aim of this study was to examine whether a resident dendritic cell population in the skin, Langerhans cells, respond to cutaneous viral infections which are effectively cleared by the immune system. We therefore characterized the ability of Langerhans cells to migrate to local draining lymph nodes following infection with the arthropod-borne viruses, West Nile virus or Semliki Forest virus. The data show that major histocompatibility complex class II+/NLDC145+/E-cadherin+ Langerhans cell numbers are increased in the draining lymph nodes of infected mice and this increase is accompanied by a concomitant decrease in the Langerhans cell density in the epidermis. Langerhans cell migration is associated with an accumulation of leukocytes in the lymph node, which is one of the earliest events in the initiation of an immune response. Both the migratory response and the draining lymph node leukocyte accumulation were abrogated if ultraviolet-inactivated instead of live viruses were used, suggesting the activation and subsequent migration of Langerhans cells requires a live, replicating antigen. Our findings are likely to have wider implications for the development of epidermally delivered vaccines and suggest that mobilization of dendritic cells may be involved in the development of immune responses to arthropod-borne viruses. (+info)
Detection of viral ribonucleic acid and histologic analysis of inflamed synovium in Ross River virus infection.
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OBJECTIVE: To document the histology of Ross River virus (RRV) arthritis and to examine inflamed synovium for viral RNA. METHODS: Biopsy tissue from the inflamed knees of 12 patients with RRV infection was studied using conventional and immunostaining techniques. Reverse transcriptase-polymerase chain reaction technology was used to probe for the presence of viral RNA in the synovial biopsy samples and in serum. RESULTS: Hyperplasia of the synovial lining layer, vascular proliferation, and mononuclear cell infiltration were the main histologic changes. RRV RNA was found in knee biopsy tissue that was obtained from 2 patients at 5 weeks after the onset of symptoms. CONCLUSION: RRV RNA was identified in inflamed synovium more than a month after symptoms began. Inflammation was apparent in the absence of detectable virus in the majority of patients. (+info)
Alpha/beta interferon protects adult mice from fatal Sindbis virus infection and is an important determinant of cell and tissue tropism.
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Infection of adult 129 Sv/Ev mice with consensus Sindbis virus strain TR339 is subclinical due to an inherent restriction in early virus replication and viremic dissemination. By comparing the pathogenesis of TR339 in 129 Sv/Ev mice and alpha/beta interferon receptor null (IFN-alpha/betaR(-/-)) mice, we have assessed the contribution of IFN-alpha/beta in restricting virus replication and spread and in determining cell and tissue tropism. In adult 129 Sv/Ev mice, subcutaneous inoculation with 100 PFU of TR339 led to extremely low-level virus replication and viremia, with clearance under way by 96 h postinoculation (p.i.). In striking contrast, adult IFN-alpha/betaR(-/-) mice inoculated subcutaneously with 100 PFU of TR339 succumbed to the infection within 84 h. By 24 h p.i. a high-titer serum viremia had seeded infectious virus systemically, coincident with the systemic induction of the proinflammatory cytokines interleukin-12 (IL-12) p40, IFN-gamma, tumor necrosis factor alpha, and IL-6. Replicating virus was located in macrophage-dendritic cell (DC)-like cells at 24 h p.i. in the draining lymph node and in the splenic marginal zone. By 72 h p.i. virus replication was widespread in macrophage-DC-like cells in the spleen, liver, lung, thymus, and kidney and in fibroblast-connective tissue and periosteum, with sporadic neuroinvasion. IFN-alpha/beta-mediated restriction of TR339 infection was mimicked in vitro in peritoneal exudate cells from 129 Sv/Ev versus IFN-alpha/betaR(-/-) mice. Thus, IFN-alpha/beta protects the normal adult host from viral infection by rapidly conferring an antiviral state on otherwise permissive cell types, both locally and systemically. Ablation of the IFN-alpha/beta system alters the apparent cell and tissue tropism of the virus and renders macrophage-DC-lineage cells permissive to infection. (+info)