Dynamics of Mycobacterium tuberculosis Ag85B Revealed by a Sensitive Enzyme-Linked Immunosorbent Assay.

Secretion of specific proteins contributes to pathogenesis and immune responses in tuberculosis and other bacterial infections, yet the kinetics of protein secretion and fate of secreted proteins in vivo are poorly understood. We generated new monoclonal antibodies that recognize the Mycobacterium tuberculosis secreted protein Ag85B and used them to establish and characterize a sensitive enzyme-linked immunosorbent assay (ELISA) to quantitate Ag85B in samples generated in vitro and in vivo We found that nutritional or culture conditions had little impact on the secretion of Ag85B and that there is considerable variation in Ag85B secretion by distinct strains in the M. tuberculosis complex: compared with the commonly used H37Rv strain (lineage 4), Mycobacterium africanum (lineage 6) secretes less Ag85B, and two strains from lineage 2 secrete more Ag85B. We also used the ELISA to determine that the rate of secretion of Ag85B is 10- to 100-fold lower than that of proteins secreted by Gram-negative and Gram-positive bacteria, respectively. ELISA quantitation of Ag85B in lung homogenates of M. tuberculosis H37Rv-infected mice revealed that although Ag85B accumulates in the lungs as the bacterial population expands, the amount of Ag85B per bacterium decreases nearly 10,000-fold at later stages of infection, coincident with the development of T cell responses and arrest of bacterial population growth. These results indicate that bacterial protein secretion in vivo is dynamic and regulated, and quantitation of secreted bacterial proteins can contribute to the understanding of pathogenesis and immunity in tuberculosis and other infections.IMPORTANCE Bacterial protein secretion contributes to host-pathogen interactions, yet the process and consequences of bacterial protein secretion during infection are poorly understood. We developed a sensitive ELISA to quantitate a protein (termed Ag85B) secreted by M. tuberculosis and used it to find that Ag85B secretion occurs with slower kinetics than for proteins secreted by Gram-positive and Gram-negative bacteria and that accumulation of Ag85B in the lungs is markedly regulated as a function of the bacterial population density. Our results demonstrate that quantitation of bacterial proteins during infection can reveal novel insights into host-pathogen interactions

Cell-to-Cell Transfer of M. tuberculosis Antigens Optimizes CD4 T Cell Priming

SummaryDuring Mycobacterium tuberculosis and other respiratory infections, optimal T cell activation requires pathogen transport from the lung to a local draining lymph node (LN). However, the infected inflammatory monocyte-derived dendritic cells (DCs) that transport M. tuberculosis to the local lymph node are relatively inefficient at activating CD4 T cells, possibly due to bacterial inhibition of antigen presentation. We found that infected migratory DCs release M. tuberculosis antigens as soluble, unprocessed proteins for uptake and presentation by uninfected resident lymph node DCs. This transfer of bacterial proteins from migratory to local DCs results in optimal priming of antigen-specific CD4 T cells, which are essential in controlling tuberculosis. Additionally, this mechanism does not involve transfer of the whole bacterium and is distinct from apoptosis or exosome shedding. These findings reveal a mechanism that bypasses pathogen inhibition of antigen presentation by infected cells and generates CD4 T cell responses that control the infection

Variation of Mycobacterium tuberculosis antigen-specific IFN-γ and IL-17 responses in healthy tuberculin skin test (TST)-positive human subjects.

ObjectiveTo determine the variation of IFN-γ and IL-17 responses to M. tuberculosis antigens in healthy TST+ humans.MethodsWe isolated peripheral blood mononuclear cells from 21 TST+ healthy adults, stimulated them with phytohemagglutinin (PHA), PPD, Ag85B, ESAT-6, and live M. bovis BCG, and assayed IFN-γ and IL-17 secretion by ELISA in supernatants after 24 or 72 hours of incubation respectively.ResultsAs in other studies, we found a wide range of IFN-γ responses to M. tuberculosis antigens; the variation significantly exceeded that observed in the same donors to the polyclonal T cell stimulus, phytohemagglutinin (PHA). In addition, we assayed IL-17 secretion in response to the same stimuli, and found less subject-to-subject variation. Analysis of the ratio of IFN-γ to IL-17 secretion on a subject-to-subject basis also revealed a wide range, with the majority of results distributed in a narrow range, and a minority with extreme results all of which were greater than that in the majority of subjects. The data suggest that study of exceptional responses to M. tuberculosis antigens may reveal immunologic correlates with specific outcomes of M. tuberculosis infection.ConclusionVariation of IFNγ and IFN-γ/IL-17 responses to mycobacterial antigens exceeds that of responses to the polyclonal stimulus, PHA, in TST positive healthy humans. This indicates a quantitative spectrum of human immune responses to infection with M. tuberculosis. Since the outcome of human infection with M. tuberculosis varies greatly, systematic study of multiple immune responses to multiple antigens is likely to reveal correlations between selected immune responses and the outcomes of infection

Sistem Informasi Strategis Mendayagunakan Sistem Informasi untuk Mencapai Keunggulan Kompetitif

While most companies are content to develop and implement computer-based information systems to improve their operational and managerial effectiveness, a few also rely on innovative systems to give them an edge over the competition. An information system that is specifically designed with the aim to create a competitive advantage for its firm is called a strategic information system.To build a strategic information system, one should have a good understanding as to what factors contribute towards competitive advantage. It is here that the analyses by Michael E. Porter on this subject are particularly invaluable. Furthermore, one should also realize what the modern computer excels at fast processing time, fast data retrieval, fast communications, and reliability.There are also risks inherent to strategic information systems. These include the large capital outlay involved, as well as the reality that any competitive advantage thus achieved is not sustainable for long

Anti-ganglioside antibodies in patients with Zika virus infection-associated Guillain-Barré Syndrome in Brazil.

Zika virus infection is associated with the development of Guillain-Barré syndrome (GBS), a neurological autoimmune disorder caused by immune recognition of gangliosides and other components at nerve membranes. Using a high-throughput ELISA, we have analyzed the anti-glycolipid antibody profile, including gangliosides, of plasma samples from patients with Zika infections associated or not with GBS in Salvador, Brazil. We have observed that Zika patients that develop GBS present higher levels of anti-ganglioside antibodies when compared to Zika patients without GBS. We also observed that a broad repertoire of gangliosides was targeted by both IgM and IgG anti-self antibodies in these patients. Since Zika virus infects neurons, which contain membrane gangliosides, antigen presentation of these infected cells may trigger the observed autoimmune anti-ganglioside antibodies suggesting direct infection-induced autoantibodies as a cause leading to GBS development. Collectively, our results establish a link between anti-ganglioside antibodies and Zika-associated GBS in patients

Initiation of the adaptive immune response to Mycobacterium tuberculosis depends on antigen production in the local lymph node, not the lungs

The onset of the adaptive immune response to Mycobacterium tuberculosis is delayed compared with that of other infections or immunization, and allows the bacterial population in the lungs to expand markedly during the preimmune phase of infection. We used adoptive transfer of M. tuberculosis Ag85B-specific CD4+ T cells to determine that the delayed adaptive response is caused by a delay in initial activation of CD4+ T cells, which occurs earliest in the local lung-draining mediastinal lymph node. We also found that initial activation of Ag85B-specific T cells depends on production of antigen by bacteria in the lymph node, despite the presence of 100-fold more bacteria in the lungs. Although dendritic cells have been found to transport M. tuberculosis from the lungs to the local lymph node, airway administration of LPS did not accelerate transport of bacteria to the lymph node and did not accelerate activation of Ag85B-specific T cells. These results indicate that delayed initial activation of CD4+ T cells in tuberculosis is caused by the presence of the bacteria in a compartment that cannot be mobilized from the lungs to the lymph node, where initial T cell activation occurs

Disinfection of football protective equipment using chlorine dioxide produced by the ICA TriNova system

<p>Abstract</p> <p>Backround</p> <p>Community-associated methicillin-resistant <it>Staphylococcus aureus </it>outbreaks have occurred in individuals engaged in athletic activities such as wrestling and football. Potential disease reduction interventions include the reduction or elimination of bacteria on common use items such as equipment. Chlorine dioxide has a long history of use as a disinfectant. The purpose of this investigation was to evaluate the ability of novel portable chlorine dioxide generation devices to eliminate bacteria contamination of helmets and pads used by individuals engaged in football.</p> <p>Methods</p> <p>In field studies, the number of bacteria associated with heavily used football helmets and shoulder pads was determined before and after overnight treatment with chlorine dioxide gas. Bacteria were recovered using cotton swabs and plated onto trypticase soy agar plates. In laboratory studies, <it>Staphylococcus aureus </it>was applied directly to pads. The penetration of bacteria into the pads was determined by inoculating agar plates with portions of the pads taken from the different layers of padding. The ability to eliminate bacteria on the pad surface and underlying foam layers after treatment with chlorine dioxide was also determined.</p> <p>Results</p> <p>Rates of recovery of bacteria after treatment clearly demonstrated that chlorine dioxide significantly (p < 0.001) reduce and eliminated bacteria found on the surface of pads. For example, the soft surface of shoulder pads from a university averaged 2.7 × 10³recoverable bacteria colonies before chlorine dioxide treatment and 1.3 × 10²recoverable colonies after treatment. In addition, the gas was capable of penetrating the mesh surface layer and killing bacteria in the underlying foam pad layers. Here, 7 × 10³to 4.5 × 10³laboratory applied <it>S. aureus </it>colonies were recovered from underlying layers before treatment and 0 colonies were present after treatment. Both naturally occurring bacteria and <it>S. aureus </it>were susceptible to the treatment process.</p> <p>Conclusion</p> <p>Results of this study have shown that chlorine dioxide can easily and safely be used to eliminate bacteria contamination of protective pads used by football players. This could serve to reduce exposure to potential pathogens such as the methicillin-resistant <it>Staphylococcus aureus </it>among this group of individuals.</p

Phase Separation of Crystal Surfaces: A Lattice Gas Approach

We consider both equilibrium and kinetic aspects of the phase separation (``thermal faceting") of thermodynamically unstable crystal surfaces into a hill--valley structure. The model we study is an Ising lattice gas for a simple cubic crystal with nearest--neighbor attractive interactions and weak next--nearest--neighbor repulsive interactions. It is likely applicable to alkali halides with the sodium chloride structure. Emphasis is placed on the fact that the equilibrium crystal shape can be interpreted as a phase diagram and that the details of its structure tell us into which surface orientations an unstable surface will decompose. We find that, depending on the temperature and growth conditions, a number of interesting behaviors are expected. For a crystal in equilibrium with its vapor, these include a low temperature regime with logarithmically--slow separation into three symmetrically--equivalent facets, and a higher temperature regime where separation proceeds as a power law in time into an entire one--parameter family of surface orientations. For a crystal slightly out of equilibrium with its vapor (slow crystal growth or etching), power--law growth should be the rule at late enough times. However, in the low temperature regime, the rate of separation rapidly decreases as the chemical potential difference between crystal and vapor phases goes to zero.Comment: 16 pages (RevTex 3.0); 12 postscript figures available on request ([email protected]). Submitted to Physical Review E. SFU-JDSDJB-94-0