T Cell Responses Are Required for Protection from Clinical Disease and for Virus Clearance in Severe Acute Respiratory Syndrome Coronavirus-Infected Mice

A dysregulated innate immune response and exuberant cytokine/chemokine expression are believed to be critical factors in the pathogenesis of severe acute respiratory syndrome (SARS), caused by a coronavirus (SARS-CoV). However, we recently showed that inefficient immune activation and a poor virus-specific T cell response underlie severe disease in SARS-CoV-infected mice. Here, we extend these results to show that virus-specific T cells, in the absence of activation of the innate immune response, were sufficient to significantly enhance survival and diminish clinical disease. We demonstrated that T cells are responsible for virus clearance, as intravenous adoptive transfer of SARS-CoV-immune splenocytes or in vitro-generated T cells to SCID or BALB/c mice enhanced survival and reduced virus titers in the lung. Enhancement of the number of virus-specific CD8 T cells by immunization with SARS-CoV peptide-pulsed dendritic cells also resulted in a robust T cell response, earlier virus clearance, and increased survival. These studies are the first to show that T cells play a crucial role in SARS-CoV clearance and that a suboptimal T cell response contributes to the pathological changes observed in SARS. They also provide a new approach to SARS vaccine design.Severe acute respiratory syndrome (SARS), caused by a novel coronavirus (SARS-CoV), resulted in over 8,000 cases of respiratory disease, with 10% mortality, in 2002 and 2003 (18). Patients with severe disease developed acute lung injury (ALI), concomitant with neutrophilia, lymphopenia, and prolonged expression of several proinflammatory cytokines (4, 17, 18, 32). Virus was cleared slowly from these patients and could be detected in respiratory secretions for as long as 21 days postinfection (p.i.) (14, 17).SARS has not recurred to a significant extent since 2003, so most studies have used animal infections to investigate the mechanism of severe disease. These studies have been facilitated by the isolation and characterization of SARS-CoV variants that cause severe respiratory disease in mice or rats after adaptation to these hosts by serial passage (15, 16, 22). One strain, isolated after 15 passages through the lungs of BALB/c mice, caused respiratory disease in young BALB/c mice (MA15 virus) (22). As in humans with severe disease, cytokine expression is elevated and prolonged in animals with severe disease, whether caused by the human Urbani or rodent-adapted strains of SARS-CoV, and is accompanied by delayed kinetics of virus clearance (1, 5, 15, 23, 26, 35). This elevated expression of proinflammatory mediators is believed to be a major contributory factor in the development of pneumonia and, subsequently, ALI.These studies did not address the role of the antivirus T cell response in disease. T cell responses are critical for virus clearance and protection from clinical disease in mice infected with other coronaviruses, such as mouse hepatitis virus (MHV) (31), or other pulmonary pathogens, such as influenza A virus or Sendai virus (7, 29). Although some studies using the Urbani strain showed that virus clearance was T cell independent (5, 34), mice infected with this strain develop only mild pneumonitis, so that the requirement for a virus-specific T cell response for protection in SARS-CoV-infected animals remains unclear.Young BALB/c mice infected with MA15 virus develop severe respiratory disease, with substantial mortality, dependent upon virus dosage (22). We recently showed that the pulmonary immune response is inefficiently activated in these mice at early times after infection, resulting in a barely detectable antivirus T cell response in mice with a lethal infection (35). Survival correlated with the development of a SARS-CoV-specific CD4 and CD8 T cell response. Poor activation of the immune response could be reversed by depletion of inhibitory alveolar macrophages with clodronate, a drug that depletes phagocytic cells (28, 30), or by treatment with poly(I-C) or CpG, both of which activate macrophages and dendritic cells (DCs). Disease could also be ameliorated by adoptive transfer of activated DCs, which are able to traffic to the lung draining lymph nodes (DLN) and prime the antivirus T cell response. These interventions effect two changes in the host immune response: they result in activation of the innate immune system and facilitate the development of a robust virus-specific T cell response. From these results, we could not determine the relative importance of these two limbs of the immune response in protection. Here, we examined whether a robust antivirus T cell response was sufficient to protect mice in the absence of interventions that activated the innate immune response. Our results indicate that virus-specific T cells are necessary and sufficient for virus clearance and for protection from clinical disease in MA15 virus-infected mice.     

Pervasive associations between Cybaeus spiders and the bacterial symbiont Cardinium

Cardinium is a recently discovered maternally transmitted bacterial endosymbiont in the Bacteroidetes that has thus far been documented in five arthropod orders. While its effects on his hosts are largely unknown, a few strains have been shown to manipulate host reproduction in parasitic wasps and in mites, either by transforming males into females, or by causing mating incompatibilities between infected males and uninfected males. Cardinium has recently been reported to be widespread in spiders, and in this study, we document pervasive infections in Cybaeus spiders, which are some of the most abundant yet understudied spiders in the understory of moist Western North American forests. 12/20 species, as well as 96% of individuals in a local population of Cybaeus signifer were infected. Phylogenetic analysis revealed three closely related symbiont haplotypes within Cybaeus. Haplotypes clustered within geographically close species, suggesting that horizontal transmission might be quite high in this symbiont lineage.     

The meningococcal vaccine candidate GNA1870 binds the complement regulatory protein factor H and enhances serum resistance

Neisseria meningitidis binds factor H (fH), a key regulator of the alternative complement pathway. A approximately 29 kD fH-binding protein expressed in the meningococcal outer membrane was identified by mass spectrometry as GNA1870, a lipoprotein currently under evaluation as a broad-spectrum meningococcal vaccine candidate. GNA1870 was confirmed as the fH ligand on intact bacteria by 1) abrogation of fH binding upon deleting GNA1870, and 2) blocking fH binding by anti-GNA1870 mAbs. fH bound to whole bacteria and purified rGNA1870 representing each of the three variant GNA1870 families. We showed that the amount of fH binding correlated with the level of bacterial GNA1870 expression. High levels of variant 1 GNA1870 expression (either by allelic replacement of gna1870 or by plasmid-driven high-level expression) in strains that otherwise were low-level GNA1870 expressers (and bound low amounts of fH by flow cytometry) restored high levels of fH binding. Diminished fH binding to the GNA1870 deletion mutants was accompanied by enhanced C3 binding and increased killing of the mutants. Conversely, high levels of GNA1870 expression and fH binding enhanced serum resistance. Our findings support the hypothesis that inhibiting the binding of a complement down-regulator protein to the neisserial surface by specific Ab may enhance intrinsic bactericidal activity of the Ab, resulting in two distinct mechanisms of Ab-mediated vaccine efficacy. These data provide further support for inclusion of this molecule in a meningococcal vaccine. To reflect the critical function of this molecule, we suggest calling it fH-binding protein.     

Positive reinforcement training affects hematologic and serum chemistry values in captive chimpanzees (Pan troglodytes)

Positive reinforcement training (PRT) techniques have received considerable attention for their stress reduction potential in the behavioral management of captive nonhuman primates. However, few published empirical studies have provided physiological data to support this position. To address this issue, PRT techniques were used to train chimpanzees (Pan troglodytes) to voluntarily present a leg for an intramuscular (IM) injection of anesthetic. Hematology and serum chemistry profiles were collected from healthy chimpanzees (n=128) of both sexes and various ages during their routine annual physical examinations over a 7-year period. Specific variables potentially indicative of acute stress (i.e., total white blood cell (WBC) counts, absolute segmented neutrophils (SEG), glucose (GLU) levels, and hematocrit (HCT) levels) were analyzed to determine whether the method used to administer the anesthetic (voluntary present for injection vs. involuntary injection) affected the physiological parameters. Subjects that voluntarily presented for an anesthetic injection had significantly lower mean total WBC counts, SEG, and GLU levels than subjects that were involuntarily anesthetized by more traditional means. Within-subjects analyses revealed the same pattern of results. This is one of the first data sets to objectively demonstrate that PRT for voluntary presentation of IM injections of anesthetic can significantly affect some of the physiological measures correlated with stress responses to chemical restraint in captive chimpanzees.     

A novel Ly6C/Ly6G-based strategy to analyze the mouse splenic myeloid compartment

Currently, there is no standardized panel for immunophenotyping myeloid cells in mouse spleen using flow cytometry. Markers such as CD11b, CD11c, F4/80, Gr-1, Ly6C, and Ly6G have long been used to identify various splenic cell myeloid populations. Flow cytometry and fluorescence-activated cell sorting (FACS) analysis demonstrated that Ly6G/Ly6C markers are superior to Gr-1 for identifying splenic neutrophils, eosinophils, and subsets of monocytes/macrophages. Moreover, these experiments showed that F4/80 is not required for identifying these myeloid subsets and that many of the commercially available preparations of anti-F4/80 antibodies stain poorly for this antigen in spleen. Taken together, we have now developed an informative flow cytometry panel that can be combined with other cell markers to further delineate subpopulations of mouse splenic myeloid cells. This panel will be highly useful to investigators in the flow cytometry field, as there is a critical need to standardize the analysis of myeloid cell subsets.     

Global and Local Concerns: What Attitudes and Beliefs Motivate Farmers to Mitigate and Adapt to Climate Change?

In response to agriculture''s vulnerability and contribution to climate change, many governments are developing initiatives that promote the adoption of mitigation and adaptation practices among farmers. Since most climate policies affecting agriculture rely on voluntary efforts by individual farmers, success requires a sound understanding of the factors that motivate farmers to change practices. Recent evidence suggests that past experience with the effects of climate change and the psychological distance associated with people''s concern for global and local impacts can influence environmental behavior. Here we surveyed farmers in a representative rural county in California''s Central Valley to examine how their intention to adopt mitigation and adaptation practices is influenced by previous climate experiences and their global and local concerns about climate change. Perceived changes in water availability had significant effects on farmers'' intention to adopt mitigation and adaptation strategies, which were mediated through global and local concerns respectively. This suggests that mitigation is largely motivated by psychologically distant concerns and beliefs about climate change, while adaptation is driven by psychologically proximate concerns for local impacts. This match between attitudes and behaviors according to the psychological distance at which they are cognitively construed indicates that policy and outreach initiatives may benefit by framing climate impacts and behavioral goals concordantly; either in a global context for mitigation or a local context for adaptation.     

Localization of the gene encoding insulin-degrading enzyme to human chromosome 10, bands q23----q25.

Insulin-degrading enzyme (IDE) is a cytosolic proteinase involved in the cellular processing of insulin. Using somatic cell hybrid analysis and in situ chromosomal hybridization, we have localized the gene encoding IDE to human chromosome 10, bands q23----q25. The murine Ide gene was previously mapped to Chromosome 19; together, these results suggest that the IDE gene is a member of a conserved syntenic group on human chromosome 10, bands q23----q25 and mouse Chromosome 19.