Antiviral roles of natural killer (NK) cell subsets were examined in C57BL/6 mice infected with murine cytomegalovirus (MCMV) and other viruses, including lymphocytic choriomeningitis virus (LCMV), vaccinia virus (VV), and mouse hepatitis virus (MHV). Each virus vigorously induced an NK cell infiltrate into the peritoneal cavity and liver, causing some redistributions of NK cell subsets defined by monoclonal antibody (mAb) directed against Ly49A, C/I, D, and G2. Striking results were seen with a mAb (1F8) reactive with the positively signaling molecule Ly49H, present in MCMV-resistant C57BL/6 mice. mAb 1F8 also stains Ly49 C and I, but exclusion of those reactivities with mAb 5E6, which recognizes Ly49 C and I, indicated that Ly49H+ cells infiltrated the peritoneal cavity and liver and were particularly effective at synthesizing interferon γ. Depletion of 1F8+ but not 5E6+ cells in vivo by mAb injections enhanced MCMV titers by 20-1,000-fold in the spleen and approximately fivefold in the liver. Titers of LCMV or VV were not enhanced. These anti-MCMV effects were attributed to prototypical NK1.1+CD3− NK cells and not to NK1.1+CD3+ “NK/T” cells. This is the first evidence that control of a virus infection in vivo is mediated by a distinct NK cell subset.

As is evident from the human immunodeficiency virus epidemic, there is no systematic method for producing a vaccine. Genetic immunization is a new approach to vaccine production that has many of the advantages of live/attenuated pathogens but no risk of infection. It involves introducing DNA encoding a pathogen protein into host cells and has shown promise in several disease models. Here we describe a new method for vaccine development, expression-library immunization, which makes use of the technique of genetic immunization and the fact that all the antigens of a pathogen are encoded in its DNA. An expression library of pathogen DNA is used to immunize a host thereby producing the effects of antigen presentation of a live vaccine without the risk. We show that even partial expression libraries made from the DNA of Mycoplasma pulmonis, a natural pathogen in rodents, provide protection against challenge from the pathogen. Expression library immunization may prove to be a general method for vaccination against any pathogen.

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We evaluated a molecular subtyping system for Treponema pallidum for its ability to differentiate between strains obtained from male patients with primary syphilis in South Africa. Of 201 T. pallidum-positive specimens, 161 were typeable, revealing 35 subtypes. The unique subtypes identified in Durban, Cape Town, and Carletonville and the total number of subtypes suggested that the strain population was very diverse and varied geographically.

The Ly49 family of genes encode NK cell receptors that bind class I MHC Ags and transmit negative signals if the cytoplasmic domains have immunoregulatory tyrosine-based inhibitory motifs (ITIMs). 5E6 mAbs recognize Ly49C and Ly49I receptors and depletion of 5E6+ NK cells prevents rejection of allogeneic or parental-strain H2d bone marrow cell (BMC) grafts. To determine the function of the Ly49I gene in the rejection of BMC grafts, we transfected fertilized eggs of FVB mice with a vector containing DNA for B6 strain Ly49I (Ly49IB6). Ly49IB6 is ITIM+ and is recognized by 5E6 as well as Ly49I-specific 8H7 mAbs. Normal FVB H2q mice reject H2b but not H2d BMC allografts, and the rejection of H2b BMC was inhibited partially by anti-NK1.1 and completely by anti-asialo GM1, but not by anti-CD8, Abs. In FVB mice, NK1.1 is expressed on only 60% NK cells. FVB.Ly49IB6 hosts failed to reject H2d or H2b BMC, but did reject class I-deficient TAP-1−/− BMC, indicating that NK cells were functional. Nondepleting doses of anti-Ly49I Abs reversed the acceptance of H2b BMC by FVB.Ly49IB6 mice. FVB.Ly49IB6+/− mice were crossed and back-crossed with 129 mice—H2b, 5E6−, poor responders to H2d BMC grafts. While transgene-negative H2b/q F1 or first-generation back-crossed mice rejected H2b marrow grafts (hybrid resistance), transgene-positive mice did not. Thus B6 strain Ly49I receptors transmit inhibitory signals from H2b MHC class I molecules. Moreover, Ly49IB6 has no positive influence on the rejection of H2d allografts.

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Infection of C57BL/6J mice with Mycoplasma pulmonis (MP) enhanced NK cell function 3-7 days later, as detected by in vitro and in vivo assays. Moreover, spleen and lung cells of acutely infected C57BL/6J mice inhibited MP growth in vitro. The effectors were eliminated by treatment with anti-NK antibody in vivo and anti-asialo GM1 serum or anti-3A4 antibody plus complement in vitro. Clearance of viable and radiolabeled MP from the lungs was also enhanced in acutely infected mice. Acutely infected mice with severe combined immunodeficiency (SCID) eliminated viable MP faster than did uninfected mice. Antibodies to interferon-gamma (IFN-gamma) impaired clearance of MP from the lungs of SCID mice and decreased their survival times. Activated NK cells can function in resistance to early stages of infection with MP. NK cells directly inhibit MP with secrete IFN-gamma, which may activate macrophages or inhibit the growth of MP or both.