Repetitive Ag encounter, coupled with dynamic changes in Ag density and inflammation, imparts phenotypic and functional heterogeneity to memory virus-specific CD8 T cells in persistently infected hosts. For herpesvirus infections, which cycle between latency and reactivation, recent studies demonstrate that virus-specific T cell memory is predominantly derived from naive precursors recruited during acute infection. Whether functional memory T cells to viruses that persist in a nonlatent, low-level infectious state (smoldering infection) originate from acute infection-recruited naive T cells is not known. Using mouse polyomavirus (MPyV) infection, we previously showed that virus-specific CD8 T cells in persistently infected mice are stably maintained and functionally competent; however, a sizeable fraction of these memory T cells are short-lived. Further, we found that naive anti-MPyV CD8 T cells are primed de novo during persistent infection and contribute to maintenance of the virus-specific CD8 T cell population and its phenotypic heterogeneity. Using a new MPyV-specific TCR-transgenic system, we now demonstrate that virus-specific CD8 T cells recruited during persistent infection possess multicytokine effector function, have strong replication potential, express a phenotype profile indicative of authentic memory capability, and are stably maintained. In contrast, CD8 T cells recruited early in MPyV infection express phenotypic and functional attributes of clonal exhaustion, including attrition from the memory pool. These findings indicate that naive virus-specific CD8 T cells recruited during persistent infection contribute to preservation of functional memory against a smoldering viral infection.

Nephropathy associated with BK polyomavirus causes kidney allograft dysfunction and failure. Understanding the pathogenesis of polyomavirus‐associated allograft nephropathy (PVAN) is hampered by the species specificity of Polyomaviridae family members. Using a mouse polyomavirus (MPyV) kidney transplant model, we investigated clinically relevant variables that may contribute to PVAN. We found that the timing and source (i.e. donor vs. recipient) of MPyV infection and the titer of the viral inoculum have significant effects on the extent of allograft injury, with acute infection of the recipient by high‐titer MPyV inoculums producing the most profound PVAN. In contrast, altering the degree of MHC matching or increasing ischemia/reperfusion injury by prolonging the cold ischemic time of the allograft did not affect the severity of PVAN. Survival correlated positively with serum creatinine levels, but not with viral loads in the kidney allograft. Using splenectomized alymphoplasia mice, which are unable to mount primary adaptive immune responses, we further demonstrate that persistent high viral loads in the kidney are not sufficient to cause advanced PVAN. These findings suggest that the mechanism of PVAN in mice is not a direct consequence of viral cytopathology, but rather involves interplay between viral infection and the recipient antidonor immune response.

We describe the use of flow cytofluorometry to type lymphocytes for HLA-B27 antigens. We modified the cytotoxicity test of Terasaki and McClelland [Nature (London) 1964;204:998] for use with a flow cytofluorometer. In a second assay we used a commercially available monoclonal anti-HLA-B27 antibody and indirect immunofluorescence staining of cell surfaces. Although this antibody cross reacts with B7 and Bw22 antigens, the results for HLA-B27 and the cross-reacting antigens can be separated. Comparison of our results with those by a HLA-typing laboratory for 100 patients showed total agreement of the assigned typings.

Flow cytometry and monoclonal antibodies are promising tools for HLA-antigen detection. Previous approaches have been hampered by the lack of a carefully standardized system for calibration and sample analysis. A new system for HLA-B27 screening was developed using a FACScan flow cytometer, software for automated calibration and analysis, calibration beads, and the anti-HLA-B27-FITC/anti-Leu4-PE (CD3) monoclonal antibodies. The median fluorescence channel result for the HLA-B27-FITC signal of CD3+ T lymphocytes is compared to a decision marker. Values lower than this threshold are read as HLA-B27 negative and those above are recommended for retesting with the classic microcytotoxicity assay on the presumption of HLA-B27 positivity. The anti-HLA-B27 antibody reacts with all six HLA-B27 subtypes and shows a weaker binding to HLA-B7. The screening test results were compared with those from the microcytotoxicity assay for HLA-typing in studies involving several European centers. The observed sensitivity was 100% (95% CI: 98.6-100) and the specificity was 97.4% (95% CI: 96.4-98.3). Other performance studies verified the reproducibility and reliability of results obtained with the screening system. o 1994 Wiley-Liss, inc.

BackgroundThe interplay between viral infection and alloimmunity is known to influence the fate of transplanted organs. Clarifying how local virus-associated inflammation/injury and antiviral immunity can alter host alloimmune responses in transplantation remains a critical question.MethodsWe used a mouse model of polyomavirus (PyV) infection and kidney transplantation to investigate the roles of direct viral pathology, the antiviral immune response, and alloimmunity in the pathogenesis of PyV-associated allograft injury. We have previously shown that an effective primary T cell response is required in PyV-associated graft injury.ResultsHere we show that the transfer of primed antidonor, but not antiviral, T cells results in PyV-associated allograft injury. In further studies, we use a surrogate minor antigen model (ovalbumin) and show that only antidonor specific T cells and not antiviral specific T cells are sufficient to mediate injury. Lastly, we demonstrate that local but not systemic virus-mediated inflammation and injury within the graft itself are required.ConclusionsThese data suggest that in this mouse model, the predominant mechanism of allograft injury in PyV-associated injury is due to an augmented alloimmune T cell response driven by virus-induced inflammation/injury within the graft. These studies highlight the important interplay between viral infection and alloimmunity in a model system.