Host-Adaptation of Francisella tularensis Alters the Bacterium's Surface-Carbohydrates to Hinder Effectors of Innate and Adaptive Immunity

The gram-negative bacterium Francisella tularensis survives in arthropods, fresh water amoeba, and mammals with both intracellular and extracellular phases and could reasonably be expected to express distinct phenotypes in these environments. The presence of a capsule on this bacterium has been controversial with some groups finding such a structure while other groups report that no capsule could be identified. Previously we reported in vitro culture conditions for this bacterium which, in contrast to typical methods, yielded a bacterial phenotype that mimics that of the bacterium's mammalian, extracellular phase.SDS-PAGE and carbohydrate analysis of differentially-cultivated F. tularensis LVS revealed that bacteria displaying the host-adapted phenotype produce both longer polymers of LPS O-antigen (OAg) and additional HMW carbohydrates/glycoproteins that are reduced/absent in non-host-adapted bacteria. Analysis of wildtype and OAg-mutant bacteria indicated that the induced changes in surface carbohydrates involved both OAg and non-OAg species. To assess the impact of these HMW carbohydrates on the access of outer membrane constituents to antibody we used differentially-cultivated bacteria in vitro to immunoprecipitate antibodies directed against outer membrane moieties. We observed that the surface-carbohydrates induced during host-adaptation shield many outer membrane antigens from binding by antibody. Similar assays with normal mouse serum indicate that the induced HMW carbohydrates also impede complement deposition. Using an in vitro macrophage infection assay, we find that the bacterial HMW carbohydrate impedes TLR2-dependent, pro-inflammatory cytokine production by macrophages. Lastly we show that upon host-adaptation, the human-virulent strain, F. tularensis SchuS4 also induces capsule production with the effect of reducing macrophage-activation and accelerating tularemia pathogenesis in mice.F. tularensis undergoes host-adaptation which includes production of multiple capsular materials. These capsules impede recognition of bacterial outer membrane constituents by antibody, complement, and Toll-Like Receptor 2. These changes in the host-pathogen interface have profound implications for pathogenesis and vaccine development

CASE Tool Integration and Standardization

CASE tool users are faced with the task of coordinating tools and data from a variety of sources spanning the entire software development life cycle. Despite much discussion and increased standardization activity, complete, transparent CASE tool integration is still a long way from realization. There are a number of factors which have complicated the tool integration scenario and a number of actions being taken in an attempt to resolve the problems. The implications of these concerns can be examined from the perspectives of single-vendor, multiple-vendor, operating environment, development process, and end-user integration. In addition to specific technical and methodological solutions, standards efforts are viewed as a possible path to tool integration. To date, formal efforts have done little to resolve the integration problems, but de facto standards may well become the cornerstone of future CASE tool evolution

Replacing the Message Service Component in an Integration Framework

In an on-going set of commercial off-the-shelf (COTS) tool integration experiments being conducted by the CASE Environments Project, we have integrated a set of CASE tools using a combination of data integration mechanisms (PCTE Object Management System (QMS) and UNIX file system) and control integration mechanisms (Broadcast Message Server (BMS) of HP SoftBench). One of the key issues addressed in our work is the extent to which the integration of CASE tools can be independent of particular integration framework technology products. This report describes a task to examine interoperability aspects of the control integration component of the integration framework. The major conclusion from our work is that it is possible to integrate CASE tools using a message-passing approach that is independent of the integration framework product used. This report describes the activities an organization must undertake to integrate CASE tools in order to ensure this interoperation of message-passing integration products. The report also includes a set of lessons learned concerning the experiments we carried out

FOR THE COMMANDER

The ideas and findings in this report should not be construed as an official DoD position. It is published in the interest of scientific and technical information exchange

Issues in Tool Acquisition

This technical report identifies issues involved in the acquisition of Computer Aided Software Engineering (CASE) tools. Among the issues identified and discussed are cost, performance, process support, maintenance, data management, tool integration, and standardization. The report concludes with recommendations intended for individuals or groups responsible for acquiring CASE tools

Experiences with a federated environment testbed

In order to address the question, “What tool integrations are possible for third party tool users given the current state of Commercial Off The Shelf (COTS) tools and integration technology?”, a set of tool integration experiments were undertaken. These experiments used existing COTS tool and integration technology to create a loosely integrated toolset supporting a simple process scenario. The experiments demonstrated that tool integration is possible by third party tool users with moderate effort. A set of lessons learned from the experiments is provided, along with potential future directions for this work.</p