Cytomegalovirus Antigenemia Assay Research Articles

Comparison of cytomegalovirus antigenemia and culture assays in patients on and off antiviral therapy

We examined 1,869 consecutive blood specimens from 529 patients (>80% organ transplant recipients) for detection of CMV by antigenemia and culture assays, and compared results between patients on and off antiviral therapy. All 1,869 specimens were tested by the shell vial assay and antigenemia, and 503 were also tested by standard tube culture. The overall positivity rate for each test was 17.0% for antigenemia, 1.8% for shell vial culture assay, and 0.7% by tube culture. No specimens were positive by either shell vial or tube culture, while negative by antigenemia. These findings were consistent across all organ transplant and other patient types. Shell vial positivity was associated with higher antigenemia levels in patients either on or off anti-CMV drug therapy. Among the shell vial positive specimens, the antigenemia counts were higher in patients on antiviral drug therapy as compared to those not on therapy. We conclude that the pp65 antigenemia assay is superior to culture methods for detection of CMV in blood, particularly for patients on anti-CMV drug treatment. Additionally, its quantitative nature renders the antigenemia assay an excellent tracking tool for both resolution of asymptomatic, low level CMV reactivations and response of CMV infection to antiviral treatment. J. Med. Virol. 59:91–97, 1999. © 1999 Wiley-Liss, Inc.

Evaluation of the PrimeCapture CMV DNA detection plate system for detection of cytomegalovirus in clinical specimens.

With the availability of anticytomegalovirus (CMV) therapeutic agents, rapid detection of CMV is important in the care and management of the immunosuppressed patient. The PrimeCapture CMV DNA Detection Plate System (PC-PCR) was evaluated for the detection of CMV in blood and cerebrospinal fluid (CSF). The resolution of discordant results was performed by consensus testing utilizing a combination of conventional cell culture (TC-CPE), the CMV-antigenemia (CMV-Ag) assay, one or more in-house CMV nested PCR assays, and/or patient evaluation and follow-up. Of 51 blood specimens from 34 patients, 23 (45%) were identified as true positives. PC-PCR was significantly more sensitive than the CMV-Ag assay, TC-CPE, or a combination of both tests. The sensitivities, specificities, positive predictive values (PPV), and negative predictive values (NPV) for PC-PCR, the CMV-Ag assay, TC-CPE, and a combination of CMV-Ag and TC-CPE were 78, 75, 72, 81%; 46, 100, 100, 70%; 39, 100, 100, 67%; and 58, 100, 100, 73%, respectively. CMV was not detected or isolated in CSF, resulting in a combined PC-PCR sensitivity, specificity, PPV, and NPV of 77, 90, 68, and 93%, respectively. Among those laboratorians considering the incorporation of molecular CMV diagnostics into their clinical microbiology or virology laboratories, the CMV PC-PCR offers a relatively simple-to-perform and sensitive assay system.

Usefulness of the cytomegalovirus (CMV) antigenemia assay for predicting the occurrence of CMV disease and death in patients with AIDS.

A cohort study of 214 human immunodeficiency virus (HIV)-infected patients was performed to assess the usefulness of the cytomegalovirus (CMV) antigenemia assay for predicting the occurrence of CMV disease and death. Multivariate analysis revealed that only positive baseline CMV antigenemia assays (relative risk [RR], 7.2; 95% confidence interval [CI], 3.7-14.2; P = .0001) and CD4 cell counts (RR, 0.98; 95% CI, 0.97-0.99; P = .009) were associated with CMV disease. A positive baseline CMV antigenemia assay was also associated with death by multivariate analysis (RR, 2.2; 95% CI, 1.5-3.4; P = .0003). Increasing levels of CMV antigenemia during follow-up were associated with increased risks of CMV disease and death. A positive CMV antigenemia assay that showed > 10 cells per 2 x 10(5) polymorphonuclear leukocytes during follow-up was 91% sensitive and 84% specific for predicting a diagnosis of CMV disease; the negative predictive value for this positive test was high (97%). Therefore, the CMV antigenemia assay appears to be a simple, rapid, and inexpensive test for predicting the occurrence of CMV disease and death in patients with advanced HIV infection.

Routine use of positive controls in the daily performance of the human cytomegalovirus antigenemia assay

Few reports exist on the influence of humoral immune responses, against microorganisms involved in infections preceding Guillain–Barré syndrome (GBS) and GM1, on clinical outcome. Nor is there any data on the relation between anti-Helicobacter pylori antibodies and prognosis in patients with GBS. To address these questions, we assayed and correlated serum anti-GM1 IgG and IgM and anti-H. pylori, anti-Campylobacter jejuni and anti-cytomegalovirus (CMV) IgG with duration of hospitalization of GBS patients and prognosis at discharge. Patients with anti-GM1 alone or associated with anti-H. pylori antibodies had significant longer hospitalization to reach a low clinical score at discharge than those without (P=0.004). A significant difference was also found for the association of anti-GM1 with anti-CMV antibodies (P=0.019). A weak but significant association of anti-GM1 and anti-C. jejuni antibodies with long hospitalization and worse prognosis at discharge was also found (P=0.02). The statistical significance increased when patients with anti-GM1 and anti-microorganism antibodies were compared with those displaying anti-H. pylori or anti-CMV only. These findings provide further evidence that the level of circulating anti-GM1 IgG plays a role in determining recovery from disability in GBS patients irrespective of other IgG against microorganisms causing infections preceding GBS.

EFFECTS OF PREEMPTIVE THERAPY IN HEART TRANSPLANT PATIENTS WITH CYTOMEGALOVIRUS - ANTIGENEMIA

392 Preemptive antiviral therapy in patients with cytomegalovirus (CMV) -antigenemia offers a new alternative in prevention of CMV disease, one of the major complications after heart transplant. We reviewed a study group of 22 heart transplant patients who received preemptive therapy for positive CMV antigenemia in comparison to a historical control group of 20 heart transplant patients followed prior to the availability of CMV antigenemia assay. Study group patients received ganciclovir, 5 mg/kg/day bid IV for 7 days or Cytogam 200 mg/kg as a single dose, upon detection of one or more antigen-positive cells per 5 × 104 peripheral blood leukocytes examined. Patients were monitored weekly after heart transplant. Among control group patients, only CMV seronegative recipient of heart from CMV seropositive donor (R−D+) received prophylactic therapy. All patients received ganciclovir for CMV disease diagnosed by biopsy or clinical presentation. The groups were comparable with respect to donor CMV serostatus, age, sex, induction therapy, immunosuppression regimen and incidence of rejection. There were seven R−D+ patients in the study group vs. two R−D+ patients in control group. Only one patient of the seven R−D+ in study group developed CMV disease. CMV antigenemia was detected in 21 of 22 study group patients. Two study group patients were hospitalized for CMV disease (total 30 days), compared to seven control group patients (total 97 days), p=0.04. Patient charges (antigen test + therapy + room charge) in study group were 43% less than in control group. This preliminary data suggests that the use of preemptive antiviral therapy in heart transplant patients may decrease the incidence of CMV disease and result in reduced costs. A further analysis of a larger patient population is warranted.

Diagnosis of Cytomegalovirus Infection by the Detection of Early Antigen (pp65) in Leukocytes of Kidney Transplant Patients

Objective: Cytomegalovirus (CMV) infection is a frequent complication of kidney transplantation contributing substantially to morbidity and mortality. Early diagnosis of the infection is essential to implement an effective antiviral therapy. This study is aimed at establishing the CMV antigenemia assay for the diagnosis of CMV infection, and for monitoring the effectiveness of ganciclovir therapy in kidney recipients in Kuwait. Methods: Leukocyte-enriched fractions of 215 blood specimens from 92 kidney recipients were processed for the detection of CMV-specific protein (pp65) by using a commercial kit (Incstar Corp., USA). Plasma fractions were also tested for the presence of CMV IgM antibodies, and selected samples (n = 15) were processed for virus isolation in MRC-5 diploid cell culture. Results: Regarding symptomatic (n = 35) and asymptomatic patients (n = 57) the assay showed an 86% positive and a 79% negative predictive value, compared to the 55% and 70% values, respectively, for CMV IgM antibodies. The number of positive cells was parallel with the severity of symptoms. Using a cutoff level of >5 antigen-positive leukocytes/50,000 cells, the assay reached 93% positive and 96% negative predictive values. There was a very good agreement between the assay and virus isolation. Ganciclovir therapy could be monitored effectively with the assay, and in 11 symptomatic patients treated with ganciclovir, a prompt decrease in the number of antigen-positive cells was noted. Conclusion: The CMV antigenemia assay is a valuable tool for the early diagnosis of symptomatic CMV infection and monitoring antiviral therapy in kidney transplant patients.

Standardization of the human cytomegalovirus antigenemia assay by means of in vitro-generated pp65-positive peripheral blood polymorphonuclear leukocytes.

We generated in vitro human cytomegalovirus (HCMV) pp65-positive polymorphonuclear leukocytes (PMN) resembling those detected in vivo, following cocultivation of PMN from healthy donors and wild-type HCMV-infected endothelial cells or fibroblasts. After purification, PMN are suitable for preparation of cytospots which can be used for the antigenemia assay. Cytospin preparations containing a predetermined number of in vitro-generated pp65-positive PMN were used to test some of the major parameters involved in performing the antigenemia assay. The results showed or confirmed that (i) formalin fixation followed by permeabilization is the best fixation procedure developed to date, (ii) the test performance levels provided by different pools of pp65-specific monoclonal antibodies may be significantly different, and (iii) long-term storage (for an unlimited time) is best achieved by keeping fixed slides at -80 degreesC, whereas short-term storage (for up to 1 month) is best achieved by keeping unfixed slides at room temperature. This finding signifies that slides can be shipped all over the world at room temperature. In conclusion, the newly developed procedure for in vitro generation of pp65-positive PMN will provide the basis for standardization of the HCMV antigenemia assay and development of quality control programs.

Application of a standardized cytomegalovirus antigenemia assay in the management of patients with AIDS

One-hundred twenty AIDS patients, tested by an optimized cytomegalovirus antigenemia (CMV-Ag) assay, were followed over a 6-month period in an attempt to correlate viral (CMV) load with severe CMV organ-specific disease. Among patients available for follow-up, CMV organ-specific disease was present in seven of eight (88%) with pp65 antigen levels ≥50 cells per 4 × 10 5 polymorphonuclear leukocytes. One-hundred six patients of 107 patients with levels <50 pp65 reactive cells did not develop CMV organ-specific disease within our 6-month follow-up period. The CMV-Ag assay, as standardized in our clinical setting, served as a marker and predictor of CMV organ-specific disease in our AIDS patient population.

Improved outcome of cytomegalovirus retinitis in AIDS patients after introduction of protease inhibitors.

The objective of this study was to evaluate the influence of protease inhibitor therapy on the rate of progression and survival of 17 AIDS patients with stable Cytomegalovirus retinitis, who were receiving anti-CMV therapy. CD4+ count, HIV load, and CMV antigenemia assay were determined at baseline, at the first month, and every 3 months thereafter. Median CD4+ count increased from 11 to 87 cells/mm3, and median HIV RNA level decreased from 4.96 to 3.28 log10 copies/ml, after 6 months on therapy. Although 9 patients (53%) relapsed in a median time of 97 days (range, 15-152 days), no further episodes were observed during a median follow-up of 17 months (range, 5-18 months). Thus, the probability of remaining free of relapse was twofold higher than that observed in matched patients who did not receive protease inhibitors. Median CD4+ count at the 3rd month was higher in those individuals who went on to progress (p = .03), and a positive result to a CMV antigenemia test was associated with progression of retinitis (relative hazard, 4.45; p = .04). Survival rate was 94% at 17 months (89% increase). Therefore, protease inhibitor therapy reduces retinitis progression and improves survival. However, the immunologic response may not provide initial sufficient protection to avoid, or even may play a role on, early CMV progression.

Clinical Course of Cytomegalovirus (CMV) Viremia with and without Ganciclovir Treatment in CMV-Seropositive Kidney Transplant Recipients

This study was designed to evaluate the longitudinal history of cytomegalovirus (CMV) infection and to test the capacity of ganciclovir as effective therapy in CMV-seropositive renal transplant recipients. The CMV viremia was detected with CMV pp65 antigenemia assay in 153 renal transplants. The recipients were classified as having low-grade and high-grade CMV infections according to the severity of CMV infection. The recipients with low-grade CMV infections were observed without ganciclovir treatment, and the recipients with high-grade CMV infection were randomly assigned to ganciclovir-treated and untreated groups. The clinical course between low-grade and high-grade CMV infections was evaluated. All recipients with low-grade CMV infection (n = 62) showed spontaneous remission regardless of immunosuppresants. In high-grade CMV infection (n = 31), the ciclosporin A treated group (n = 11) showed no evidence of CMV disease, and the methylprednisolone-treated group (n = 8) showed CMV disease in 1 (25%) of 4 ganciclovir-untreated recipients. In the OKT3 group (n = 12), symptomatic CMV infection was observed in 6 (100%) ganciclovir-untreated recipients contrary to no CMV disease in the ganciclovir-treated group (p < 0.05). In conclusion, the CMV antigenemia assay is effective in monitoring CMV viremia, and ganciclovir treatment should be done during early CMV viremia in OKT3-treated recipients.

Use of the human cytomegalovirus (HCMV) antigenemia assay for diagnosis and monitoring of HCMV infections and detection of antiviral drug resistance in the immunocompromised.

Quantification of viral load in blood has proven to be helpful in the follow-up of disseminated HCMV infections in immunocompromised patients. (i) To describe the antigenemia assay and its optimization and (ii) to analyze the use of the antigenemia assay for the diagnosis and monitoring of HCMV infections and for the detection of treatment failures. This article is intended to give an overview of our experience in the use of the antigenemia assay. In solid organ transplant recipients and patients with AIDS, HCMV symptomatic infections mostly appear when antigenemia values are > 300 pp65-positive PBL/2 x 10(5) examined. To avoid the appearance of overt HCMV disease antiviral treatment could be administered when antigenemia levels are > 100 pp65-positive PBL/2 x 10(5) examined. Bone marrow transplant recipients show symptomatic HCMV infections when antigenemia values are > 100 pp65-positive PBL/2 x 10(5) examined. This group of patients should be treated when antigenemia levels are < 10 pp65-positive PBL/2 x 10(5) examined due to the higher mortality rate associated with HCMV complications. A decrease in antigenemia levels during therapy indicates a good response to antiviral drug, while stable or increasing values document a treatment failure and the emergence of drug-resistant HCMV strains.

A COMPARATIVE STUDY OF 500 MG BID AND 250 MG BID OF PROPHYLACTIC ORAL GANCILOVIR IN POST-KIDNEY TRANSPLANT "CMV AT RISK" RECIPIENTS

596 Recently, in a prospective, controlled study we reported that oral ganciclovir 750 mg bid for 3 months was an effective CMV prophylaxis after kidney transplantation (Clin Transplantation 1997:11:633-39). However, controversy exists about the appropriate dose and duration of this therapy. This is a prospective, comparative study. Forty adult consecutive recipients of kidney transplant 'at risk' for CMV (D+R-, D+R+, D-R+) were entered into the study to receive: prophylactic oral ganciclovir 500 mg bid[OGC 500] (n=20) or 250 mg bid [OGC 250] (n=20) for 3 months. Brief therapy with oral acylovir was allowed in case of herpetic infection. Both groups received micro-emulsion cyclosporine, mycophenolate mofetil and prednisone based immunosuppression. The diagnosis of CMV infection at presentation included serological determination of CMV specific IgM antibodies, CMV antigenemia assay, CMV immunofluorescence assay with standard culture tube and centrifugation enhanced (in-house grown monolayer) shell vial techniques, and virus isolation from urine and tissue. Primary efficacy end point was episode of CMV infection and/or disease during the first 6 months after transplant. There were no statistically significant differences between OGC 500 vs. OGC 250 groups with respect to patient's age (48.7±2.8 vs. 49.8±3.7yr), gender (M/F 12/8 vs. 13/7), race (Caucasian 15 vs. 16), causes of ESRD, type of transplant (cadaver/living 17/3 vs. 16/4), CMV D+/R- status (6 vs. 6), and mean serum creatinine (mg/dl) at discharge(2.0±0.4 vs. 1.8±0.2). At the end of the 6 months of study, no patient in OGC 500 (0%) and 1 patient in OGC 250 (5%) groups developed CMV infections (p=ns). The episodes of biopsy proven rejections were 0% (OGC 500) vs. 16% (3/20) OGC 250 (p=ns). Except for 1 patient (OGC 250), none developed CMV infection either before or after rejection. No patient in either group developed herpetic infection. Our study showed that oral ganciclovir 250 mg twice a day for 3 months successfully prevented episodes of CMV infection after kidney transplantation and was well tolerated. We recommend prophylactic use of oral ganciclovir to lower the morbidity associated with CMV infections in organ transplant recipients.

Use of the cytomegalovirus (CMV) antigenemia assay for the rapid diagnosis of primary CMV infection in hospitalized adults.

We assessed the value of the cytomegalovirus (CMV) antigenemia assay for diagnosing primary CMV infection in adults. The CMV antigenemia assay was performed for 40 patients admitted to our unit over a 2-year period with unexplained fever and suspected primary CMV infection. Nine of the 10 patients with primary CMV infection had positive CMV antigenemia assays, and the results were available within 5 hours. All 10 patients had a mononucleosis-like syndrome. All but one of the 30 other patients had negative CMV antigenemia assays. A false-positive result was obtained for a patient with systemic lupus erythematosus. Overall, the CMV antigenemia assay was 90% sensitive and 96% specific for the diagnosis of primary CMV infection. Therefore, the CMV antigenemia assay appears to be a simple, rapid, inexpensive test for the diagnosis of primary CMV infection in hospitalized adults.

Prevention and treatment of cytomegalovirus infection and disease in heart transplant recipients.

Despite advances in prophylaxis and treatment, cytomegalovirus (CMV) infection remains a significant problem in the solid-organ transplant recipient. In addition to the clinical manifestations of CMV infection, there is also the immunosuppressive effect of CMV, which confers increased risk for fungal and other opportunistic infections. In reference to heart transplant recipients, the possible connection between CMV infection and rejection or CMV infection and allograft vasculopathy are areas of active research. Recent diagnostic advances, such as the CMV antigenemia assay and CMV-DNA detection by polymerase chain reaction or direct hybrid capture, have enabled early detection and monitoring of CMV infection and have raised the question of the implications of asymptomatic viremia. A wide variety of prophylactic strategies have been evaluated in heart and other solid-organ transplant recipients, including antiviral agents, globulin preparations, combinations of these therapies, and pre-emptive treatment strategies based on early detection or identification of a high-risk subset of patients. Many of these regimens have demonstrated efficacy in certain groups of patients, but a consensus has yet to emerge in terms of a single preferable strategy. Future advances on the horizon include the development of newer antiviral agents and a vaccine.

Cytomegalovirus (CMV) retinitis and CMV antigenemia as a clue to impaired adrenocortical function in patients with AIDS.

To elucidate the relationship between the activity of CMV disease and adrenocortical function in patients with AIDS. CMV retinitis and CMV antigenemia assay (CMV-Ag: numbers of polymorphonuclear leukocytes positive for CMV pp65 antigen per 1.5 x 10(5) cells) are the least invasive and easily accessible examinations to assess the CMV disease activity. All HIV-infected patients with CD4+ lymphocyte counts < 50 x 10(6)/l who were admitted to the Research Hospital of the Institute of the Medical Science (University of Tokyo) between May 1995 to April 1996 were included in this study. Fundoscopic examination on CMV retinitis and CMV-Ag were chosen as methods to assess CMV activity because of their simplicity. Adrenocortical function was evaluated by basal plasma adrenocorticotropin, plasma cortisol, plasma aldosterone, plasma renin activity, and responses of plasma cortisol and plasma aldosterone to 250 micrograms intravenous cosyntropin [rapid adrenocorticotropin test (RAT)]. Thirty patients were enrolled in this study with a maximum CD4+ lymphocyte count of 32 x 10(6)/l. Eleven out of 30 patients showed impaired RAT response (37%). Fourteen out of 30 patients had CMV retinitis. A significant correlation was found between the presence of CMV retinitis and subnormal cortisol response (P < 0.005). Sixteen out of the 30 patients were CMV-Ag-positive. A significant correlation was found between CMV-Ag positivity and subnormal cortisol response to RAT (P < 0.005). CMV-Ag levels in the patients with subnormal cortisol response to RAT were significantly higher than those with normal response (P < 0.001). Importantly, five patients with subnormal cortisol response but not overt adrenal insufficiency at the time of RAT developed overt disease shortly afterwards. Autopsy was performed in six patients with subnormal cortisol response and showed multiple inclusion bodies indicative of CMV adrenitis. The adrenal gland is most frequently affected by CMV in AIDS patients. Our result suggests that CMV retinitis or CMV-Ag positivity independently serve as an indication of possible adrenal dysfunction.

Significance of leukocyte concentration in the performance of the quantitative cytomegalovirus (CMV) antigenemia assay

Background: The quantitative cytomegalovirus-antigenemia (CMV-Ag) assay is an important technology in the regimen of tests utilized in the care and management of acquired immunodeficiency syndrome (AIDS) and other immunocompromised patient groups. Performance of this assay is contingent upon the appropriate processing of the polymorphonuclear leukocyte (PMNL) compartment of the peripheral blood. However, a cell input standard in the performance of the CMV-Ag assay, has not been established. Interpretive differences between laboratories utilizing the CMV-Ag assay may reflect this lack of test uniformity. Objectives: To determine the effect of different PMNL concentrations on the quantitation of CMV in peripheral blood. The leukocyte concentration resulting in optimal rates of viral detection, will be compared with the shell vial assay-indirect immunofluorescent assay (SVA-IFA), and conventional tube culture (TC-CPE). Study design: A total of 74 freshly collected blood specimens were tested by the CMV-Ag assay, using cytospin preparations consisting of 2×10 5, 4×10 5 and in preliminary experiments, 8×10 5 PMNLs/slide. Data obtained from these studies were compared to SVA-IFA and TC-CPE. Viral load was monitored among 11 symptomatic patients through sequential testing of these patients at the start of ganciclovir (GCV), foscarnet (PFA), or combination drug therapy. Results: Among 74 blood specimens tested by the CMV-Ag assay, cytospin preparations consisting of 4×10 5 compared with 2×10 5 PMNLs/slide, affected a mean positive cell increase of 215% ( P=0.03). PMNL slide preparations consisting 8×10 5 cells produced background levels which prevented accurate reading of slides. The CMV-Ag assay was more sensitive than the SVA-IFA, but equivalent to TC-CPE. Among 11 patients started on drug therapy, viral load was markedly reduced in 8 within 2–3 weeks; three patients (2 deceased within 3 weeks after receiving therapy), showed no decrease in viral load. One patient was identified as harboring a PFA resistant strain. Conclusions: A PMNL concentration of 4×10 5 cells facilitated the reading of CMV-Ag assay slide preparations. The modified CMV-Ag assay furthermore, is applicable in the monitoring of viral load for the tracking of susceptible or resistant CMV strains.

Increased incidence of cytomegalovirus (CMV) infection and CMV-associated disease after allogeneic bone marrow transplantation from unrelated donors. The Fukuoka Bone Marrow Transplantation Group.

Cytomegalovirus (CMV) infection and CMV-associated disease were monitored using the CMV antigenemia assay in 72 patients who received allogeneic bone marrow transplantation (BMT), and their incidences were compared between related and unrelated donor transplant patients. The incidence of CMV infection after BMT was significantly higher in patients who received transplants from HLA-matched unrelated donors than from HLA-matched sibling donors (87% vs 53%, P < 0.05). CMV-associated disease developed in 73% of unrelated and in 14% of sibling donor transplant patients (P < 0.01). The peak levels of CMV antigenemia were significantly higher in unrelated donors than in sibling donor transplant patients (16 vs 1 CMV antigen-positive cells per 50000 WBCs, P < 0.01). The median number of CMV antigen-positive cells on first detection was also significantly higher in unrelated donor transplant patients (15 vs 1, P < 0.01). The detection of CMV antigen-positive cells preceded the development of CMV-associated disease in 18% of unrelated donor transplant patients, suggesting a lower predictive value of CMV antigenemia for subsequent CMV-associated disease in unrelated donor BMT. Careful monitoring and further studies are needed for the early diagnosis and prevention of CMV-associated disease in unrelated donor BMT.

Comparison of cytomegalovirus antigenemia and shell vial culture in allogeneic marrow transplantation recipients receiving ganciclovir prophylaxis.

We prospectively monitored 61 allogeneic BMT patients for evidence of CMV infection and disease starting 7 days prior to transplant until day 110 after transplant. Patients receiving pre- and post-transplantation ganciclovir prophylaxis were followed for the incidence of infection by the CMV antigenemia assay and shell vial cultures. The median age of all patients was 32 years (range 5-54 years). Fourteen (25%) of 57 evaluable patients became CMV antigenemia or culture positive. The incidence of culture or antigenemia positivity in CMV seropositive or seronegative patients with a seropositive donor was 29% (14 of 49 patients). The antigenemia assay became positive a median of 29 days (range 12-89 days) after BMT as compared to 46 days (range 26-98 days) by shell vial assay (P < 0.001). There were no cases of CMV disease in the first 110 days after transplant. This study demonstrates that despite the use of prophylactic ganciclovir, BMT patients developed CMV infection but did not progress to disease in this study, the CMV antigenemia assay may be used to monitor for CMV infection during prophylaxis, and the current regimens for CMV prophylaxis with ganciclovir may require further evaluation to determine an optimal regimen to prevent CMV infection.

Polymerase Chain Reaction Assays for the Detection of Cytomegalovirus in Organ and Bone Marrow Transplant Recipients

Cytomegalovirus (CMV) infection is ubiquitous and results in a wide spectrum of clinical manifestations ranging from asymptomatic infection to severe life threatening disease. Infection in normal children and adults usually causes no symptoms but in the immuncompromised host, CMV may result in severe opportunistic infections with high morbidity and mortality. Historically, virus detection was dependent on culture of the virus or on a centrifugation culture system referred to as a shell vial assay. The shell vial assay frequently lacked sensitivity and was unable to detect infection in its early phase. Also, as with culture assays, the results were affected by antiviral therapy. The CMV antigenemia assay was developed to provide more rapid results and has gained wide usage. This assay is limited to detection of the virus in white blood cells and is more sensitive than culture or the shell vial assay.Application of the polymerase chain reaction (PCR) to these problems has resulted in the development of assays for CMV which are more sensitive than previously available methods. This method employs liquid hybridization with 32P labeled probes and gel retardation analysis for detection of amplified DNA specific for each virus.A comparison of the detection of CMV by an antigenemia assay or the PCR method in the leukocytes of renal transplant patients revealed that the PCR assay detects cytomegalovirus earlier and more consistently than the antigenemia assay.Finally, the application of a fluorescent dye detection system and image analysis of the acrylamide gel with a laser scanner provides additional sensitivity to the detection of cytomegalovirus, as well as avoiding the use of radioactivity, making the assay more adaptable to the clinical laboratory.

Cytomegalovirus Antigenemia in Acquired Immunodeficiency Syndrome Patients With Untreated Cytomegalovirus Retinitis

To determine the frequency of cytomegalovirus (CMV) viremia in patients with acquired immunodeficiency syndrome (AIDS) and untreated CMV retinitis using conventional cell culture isolation and the sensitive CMV antigenemia assay. We examined 24 AIDS patients with ophthalmologic diagnosis of untreated CMV retinitis and 24 AIDS patients without present or past retinitis (control patients) from three medical centers between September 1992 and March 1994. Cytomegalovirus antigenemia was detected by an indirect peroxidase staining in 300,000 cytocentrifuged neutrophils, using a mixture of murine monoclonal antibodies directed against the pp65 lower matrix protein of CMV. Positive antigenemia was demonstrated in eight (33.3%) of the 24 retinitis patients and in none of the 24 control patients (P < .001). Only two of the eight antigenemia-positive patients had a concurrent positive CMV isolation from blood leukocytes by conventional cell culture assay. These results emphasize the risk of extraocular disease in AIDS patients with CMV retinitis because the virus is often present in peripheral blood leukocytes. The CMV antigenemia assay may be a simple and rapid means of identifying those patients with unilateral retinitis at highest risk of developing CMV retinitis of the fellow eye or of visceral CMV disease if intravitreal injections or implants are used as sole treatment for CMV retinitis.