Symptomatic cytomegalovirus (CMV) infections identified by image cytometry and other parameters for CMV infection.

Thirty-eight renal transplant recipients were followed during the first 3 months after transplantation. Once weekly, cultures of urine and buffy coat for cytomegalovirus (CMV) were taken and an immunocytochemical assay for immediate early antigens of CMV (IEA assay) was performed. Thirty patients had evidence of a CMV infection and 11 had a symptomatic CMV infection. All symptomatic patients had one or more positive urine cultures or a positive IEA assay. However, 15 patients with positive urine cultures and 12 patients with a positive IEA assay lacked any signs of symptomatic CMV disease. Moreover, 6 out of 15 patients with positive buffy coat cultures for CMV did not have symptomatic CMV disease. Using a computerized system to quantify IEA-positive granulocytes, we show that the absolute number of positive cells per million correlates very well with the occurrence of symptomatic CMV disease.

Rapid detection of human cytomegalovirus DNA in peripheral blood leukocytes of viremic transplant recipients by the polymerase chain reaction.

Peripheral blood leukocyte samples (n = 458) of 24 bone marrow transplant and 52 kidney transplant patients were examined weekly for the presence of human cytomegalovirus (HCMV) using an improved culture technique (DEAFF; detection of early antigen fluorescent foci). In total 5 (21%) bone marrow transplant and 11 (21%) kidney transplant patients developed a viremia. Patients' samples were investigated for the presence of HCMV DNA using an in vitro DNA amplification technique, the polymerase chain reaction (PCR). From the statistically evaluable viremic patients (n = 13), 110 blood samples were analyzed. In 5 of these patients, the DEAFF and PCR led to identical results. In 8 patients however the PCR was more sensitive, i.e. HCMV DNA was detected for a longer period of time. Applying statistical analysis using the McNemar test, this result was significant (P less than 0.05). The PCR applied on leukocyte samples did not detect HCMV DNA in viruric patients without viremia. Moreover, the current PCR never led to positive results with peripheral blood leukocyte samples of healthy seropositive or seronegative controls. Since the PCR can be performed in 6 hr, this technique will contribute to rapid detection of HCMV DNA in peripheral blood leukocytes and therefore to optimal clinical management of HCMV-infected transplant recipients.

Infections in renal transplant patients treated with cyclosporine or azathioprine.

The incidence and severity of infectious complications were retrospectively investigated in 80 renal transplant patients who had been selected randomly to receive either cyclosporine (CsA) or azathioprine (Aza) in combination with low doses of corticosteroids. In the first 3 months, the incidence of infections was twice as high in the Aza-treated patients (p less than 0.05) which was due to an increase of predominantly minor infections. This increased incidence of infections was related to the increased number of anti-rejection treatments in the Aza-treated group. The types of infections were not different between the 2 treatment groups nor did the incidence of CMV infections differ. In the CsA-treated group, patients were randomly assigned at 3 months after transplantation to either continuation of CsA therapy or conversion to Aza. After conversion a small but not significant increase in predominantly minor infections was observed, which may be attributable to increased doses of corticosteroids given during the conversion. We conclude that in the first 3 months following transplantation, CsA therapy is associated with significantly less infections than Aza therapy; following conversion of CsA to Aza at 3 months only a small increase in the infection incidence is found.

Perivascular deposits of IgM in the skin of transplant recipients during active cytomegalovirus infections. Correlation with IgM rheumatoid factors and IgM immune complexes.

Skin biopsies from 30 renal transplant patients were investigated for cellular infiltrates and deposits of IgM, IgA, IgG, C3, and C5-9 neoantigen. Granular perivascular deposits of IgM were detected in biopsies of 8 of 14 patients during active cytomegalovirus (CMV) infections and in none of 16 controls. In 5 biopsies, the IgM deposits were accompanied by little or no IgG, IgA, or C, while in 3 biopsies definite C3 deposits were present. One of the biopsies with C3 deposits also had C5-9 deposits and another had C5-9 and IgA deposits. Three monoclonal antibodies failed to detect early or late nuclear antigens of CMV in the deposits. These deposits were not associated with clinically evident manifestations of vasculitis. A strong correlation was found between IgM deposits in the skin and IgM circulating immune complexes (CIC) and also IgM rheumatoid factor (RF). The deposition of IgM was not more frequent in primary than in secondary CMV infections, and it did not correlate with the production of IgM antibodies that were specific for CMV antigens.

Association of rheumatoid factors in renal transplant recipients with cytomegalovirus infection and not with rejection.

Because rheumatoid factors (RF) were detected in the circulation of the majority of early renal transplant recipients and could be eluted from rejected transplants, RF were hypothesized to be related to antibody responses to the histoincompatible graft. The possibility that RF production might have been related to infection and not rejection has not been considered previously. Therefore, we investigated serial serum samples from 147 adult renal transplant recipients for RF with latex agglutination and radioimmune assays. RF were detected in the sera of 32 patients, 30 of whom had coincident active cytomegalovirus (CMV) infections. Another 45 patients with active CMV infections did not have detectable circulating RF. In contrast, of 74 patients who experienced a total of 103 treated reversible or irreversible rejection episodes in the absence of evidence of active CMV infections, only 2 patients produced RF during their rejection episodes. Nine of the patients who did not produce RF during a rejection episode subsequently produced RF during a later CMV infection. These data indicate that RF production in renal transplant recipients is associated with CMV infection and not rejection. Moreover, RF production was found to be more frequently associated with primary and severe CMV infections than with secondary or milder CMV infections. RF production was not more frequent in patients who were HLA-DR-4-positive., older, or female, characteristics that have been associated with RF production in other populations. All of the sera with detectable RF contained IgM antibodies that were directed to the Fc portion of human IgG, and about half contained additional IgM antibodies directed to Fab. Thus CMV infections may be the stimuli for the IgM anti-Fab antibodies that have been reported in pretransplant serum samples. Eleven patients produced IgG or IgA RF in addition to IgM RF during CMV infections.

IgM immune complexes, lymphocytotoxins, and rheumatoid factors in renal transplant recipients with CMV disease.

In studying 127 consecutive adult recipients of cadaver renal transplants, we found that the 23 patients who developed CMV disease produced IgM immune complexes as measured by a polyethyleneglycol precipitation (PEG) assay which coincided with symptoms of their illness. In addition to anti-CMV antibodies, PEG precipitated apparently non-specific antibodies such as lymphocytotoxins and rheumatoid factor (RF). The lymphocytotoxins were IgM antibodies that were not directed against HLA antigens and lysed granulocytes as well as lymphocytes but not platelets at 22 degrees C. Lymphocytotoxin production was correlated with HLA-DR 3 and 7 and with graft dysfunction during the CMV disease. The RF also were predominantly IgM antibodies that were detectable for only 3-8 weeks. The production of RF coincided with the initial rise in IgG anti-CMV antibody activity and some reacted with the Fab fragments of IgG raising the possibility that they could modulate the cellular or humoral immune response to CMV. Patients with RF tended to have severe CMV infections with pneumonia and graft dysfunction.

Increased IgM and IgM immune complex-like material in the circulation of renal transplant recipients with primary cytomegalovirus infections.

Twelve of 60 consecutive adult recipients of cadaver kidney transplants had increased polyethyleneglycol (PEG) precipitable IgM immune complex-like material in their circulation in the first 4 months after transplantation. All 10 recipients with primary CMV and two of four with secondary CMV infections had significant elevations in PEG precipitable IgM that coincided with rises in their CMV antibody titres. Ultracentrifuge analysis demonstrated two peaks of PEG precipitable material with sedimentation rates of about 20S and 40S. Total IgM immunoglobulin levels also were increased in transplant recipients with CMV infections, but this was less specific and occurred in patients without CMV infections. The Clq binding assay, which is more sensitive for IgG than IgM containing complexes, was positive in only three of 10 patients with primary CMV and none of four with secondary CMV. Granular deposits of IgM, but not IgG, were detected in the glomeruli of six of seven transplants biopsied during CMV infection. The PEG-IgM assay was not influenced by rejection or prednisone therapy. Thus, transplant patients, who develop primary CMV infections, produce elevated levels of circulating IgM and IgM immune complex-like material. These findings may help to differentiate CMV infection from transplant rejection as well as to increase our understanding of the special pathogenic properties of CMV in transplant recipients.

Anti-thymocyte globulin (ATG) can eliminate platelet refractoriness.

Of the 14 patients with aplastic anaemia treated in our hospital with anti-thymocyte globulin (ATG), four were refractory to random platelets before therapy due to the presence of leucocyte antibodies. In contrast to the ten nonrefractory patients in whom no success was obtained, three of the four refractory patients showed haematological improvement after ATG. Additionally, two patients could be substituted again with random platelets. The other two hardly needed platelet-transfusions after ATG, and they were given HLA-compatible platelets. To determine the degree of immunosuppression, these four patients were tested for the presence of antibodies against leucocytes and two endemic viruses, i.e., mumps and rubella virus. Before ATG, all sera reacted with almost the whole leucocyte testpanel. After treatment, the leucocyte antibodies disappeared completely in three patients. In one patient there was no dramatic change. In all patients, however, the antibody-titre against the mumps and rubella viruses remained constant and there was only a slight decrease in total IgG content in the three "suppressed" patients. We conclude that it might be worthwhile to study systematically the selective immuno-suppressive effect of ATG.