Screening for Epstein-Barr virus (EBV) infection status in university freshmen: acceptability of a gingival swab method.

Prophylactic vaccines against Epstein-Barr virus (EBV) are under development. EBV-naïve college freshmen are ideal candidates for an efficacy trial, because their incidence of infectious mononucleosis (mono) during freshman year is as high as 20%. To assess perceptions about mono and a mono vaccine, and to learn if EBV immune status could be determined using a gingival swab rather than phlebotomy, we performed a cross-sectional study of 235 healthy students at the beginning of their freshman year. Subjects completed questionnaires and donated oral washes, gingival swabs and venous blood. Overall, 90% of students found the swab easy to use and 80% preferred the swab over venepuncture. Of the 193 students with sufficient samples, 108 (56%) had EBV antibodies in blood vs. 87 (45.1%) in the gingival swab. The sensitivity and specificity of the swab compared with blood for detecting EBV antibodies was 75.9% and 94.1%, respectively, with an accuracy of 89.3%. EBV DNA was detected in the oral wash and swab of 39.2% and 30.4% of blood-antibody-positive individuals, respectively. In conclusion, 44% of our freshmen were EBV-naïve and thus vaccine candidates, the gingival swab was an acceptable alternative to phlebotomy for detecting EBV antibody but needs improved sensitivity, and the perceived value of EBV vaccine was high (72% believed they would benefit).

The impact of recipient BKV shedding before transplant on BKV viruria, DNAemia, and nephropathy post-transplant: A prospective study.

We previously demonstrated that detectable BKV replication in donor urine pretransplant was significantly associated with post-transplant recipient BKV viremia. In this 4-year prospective study, we assessed whether recipient BKV replication pretransplant was associated with post-transplant viremia/BKV nephropathy. We studied 220 primary adult and pediatric organ transplant recipients for 490 person-years and 2100 clinical visits. BKV viruria was detectable in 28 (16%), 26 adults and two children; and viremia in none pretransplant. Post-transplant viruria occurred in all recipients with pretransplant BKV viruria, significantly more than in recipients without pretransplant viruria on univariate (P<.005) and multivariate analysis including type of organ transplanted and immunosuppression type (P .008). Time to post-transplant viruria was significantly shorter in recipients with pretransplant viruria (P .01). By univariate and multivariate analysis, BKV viruria in recipients pretransplant did not impact post-transplant BKV viremia (P=.97 and .97, respectively) even when stratified by type of organ transplant (kidney P=.6; liver P=.5). The peak serum and urine BKV PCR post-transplant were not significantly different in patients with pretransplant BKV viruria and no one developed BK nephropathy. In conclusion, recipient BKV viruria prior to transplant predicts post-transplant viruria but not viremia or BKV nephropathy.

Quantitative image analysis of HIV-1 infection in lymphoid tissue.

Tracking human immunodeficiency virus-type 1 (HIV-1) infection at the cellular level in tissue reservoirs provides opportunities to better understand the pathogenesis of infection and to rationally design and monitor therapy. A quantitative technique was developed to determine viral burden in two important cellular compartments in lymphoid tissues. Image analysis and in situ hybridization were combined to show that in the presymptomatic stages of infection there is a large, relatively stable pool of virions on the surfaces of follicular dendritic cells and a smaller pool of productively infected cells. Despite evidence of constraints on HIV-1 replication in the infected cell population in lymphoid tissues, estimates of the numbers of these cells and the virus they could produce are consistent with the quantities of virus that have been detected in the bloodstream. The cellular sources of virus production and storage in lymphoid tissues can now be studied with this approach over the course of infection and treatment.

Multicenter comparison of different real-time PCR assays for quantitative detection of Epstein-Barr virus.

Quantification of Epstein-Barr virus (EBV) in peripheral blood is important for the diagnosis and management of serious EBV diseases, including posttransplant lymphoproliferative disorder. A variety of PCR-based methods are currently in use; however, there is little information on their comparability. This study assessed the relative performance of different quantitative assays. A multicenter comparative study was performed at eight sites using three panels consisting of serial dilutions of quantified EBV DNA and extracts from a total of 19 whole-blood specimens. Samples were distributed and tested blindly. Instrumentation, probe chemistries, amplification targets, and other test-related aspects varied considerably between laboratories. Each laboratory's calibration curve indicated strong evidence of a consistent log-linear relationship between viral load and cycle threshold, suggesting that intralaboratory tracking of a given patient would yield similar relative quantitative trends among the participating test sites. There was strong concordance among laboratories with respect to qualitative test results; however, marked quantitative discordance was seen. For most samples, the across-laboratory interquartile range of the reported viral load (in copies/microl) was roughly 0.6 log-units, and for one sample the overall range was approximately 4.2 log-units. While intralaboratory tracking of patients may yield similar results, these data indicate a need for caution when attempting to compare clinical results obtained at different institutions and suggest the potential value to be gained by more standardized testing methodology.

Human cytomegalovirus-stimulated peripheral blood mononuclear cells induce HIV-1 replication via a tumor necrosis factor-alpha-mediated mechanism.

Human cytomegalovirus (HCMV) is a potential cofactor in HIV-1 infection. To investigate the mechanism whereby HCMV promotes HIV-1 replication, a PBMC coculture assay which measures HIV-1 p24 antigen release was used as an index of viral replication. HCMV-stimulated PBMC were capable of inducing HIV-1 replication in cocultures with acutely infected PBMC; however, this occurred only when the PBMC were from HCMV-seropositive donors (598 +/- 207 versus 27 +/- 10 pg/ml p24 antigen with PBMC from HCMV-seronegative donors on day 6 of coculture). Upon stimulation with HCMV, PBMC obtained exclusively from HCMV-seropositive donors released tumor necrosis factor (TNF)-alpha (270 +/- 79 pg/ml at 18 h of culture). Monoclonal antibodies to TNF-alpha blocked the activity of HCMV-stimulated PBMC in cocultures both with acutely HIV-1-infected PBMC and with the chronically infected promonocytic line U1. Also, treatment of HCMV-stimulated PBMC with pentoxifylline, an inhibitor of TNF-alpha mRNA, markedly reduced HIV-1 replication in cocultures both with acutely and chronically infected cells. These results indicate that TNF-alpha is a key mediator of HIV-1 replication induced by HCMV-stimulated PBMC and support the concept that this cytokine plays an important role in the pathogenesis of HIV-1 infection.

Cytomegalovirus vaccine. Specific humoral and cellular immune responses in human volunteers.

Four seronegative adult male volunteers were immunized with Towne strain cytomegalovirus (CMV) vaccine. The only reaction was transient pain and swelling at the inoculation site. Viral cultures were performed during the first 12 weeks after immunization, and CMV was not recovered from throat, urine, or peripheral blood mononuclear cells. Both CMV-specific humoral and cellular immunity developed within three weeks of vaccination. Whereas humoral antibody titers declined steadily with time, the cellular immune responses seemed biphasic. An early peak in lymphocyte proliferation to CMV antigen occurred three to six weeks after immunization. Responses then diminished but increased again six to ten months after immunization. This study in a small group of normal male volunteers indicated that CMV vaccine was safe and immunogenic. That CMV vaccine elicited CMV-specific humoral and cell-mediated immunity is important, because there is evidence that both are necessary for protection from CMV infections.

A trial of vidarabine for cytomegalovirus infection in renal transplant patients.

Vidarabine was evaluated in renal transplant patients as a potential therapeutic agent in cytomegalovirus (CMV) infection. Four patients received vidarabine on an open protocol, then ten additional patients were enrolled in a double-blind protocol. Among the nine patients who received vidarabine, no notable clinical improvement occurred in either the vidarabine- or placebo-treated groups. Thus, vidarabine showed no therapeutic effect in the treatment of CMV infections at the dosages used. Four patients showed dramatic CNS deterioration within several days of the onset of vidarabine therapy. Tremors and myoclonus were common, and one patient had unusual brain pathologic changes with widespread neuronal chromatolysis. The pathologic findings in the brain in the other three patients were complex and included intracerebral hemorrhage, Fabry's disease, coccidioidomycosis meningitis, and cerebral vascular occlusion. Thus, there was no conclusive proof that vidarabine contributed to the sudden neurologic deterioration of these patients.

Morphine promotes the growth of HIV-1 in human peripheral blood mononuclear cell cocultures.

Because morphine has been shown to alter the function of human T lymphocytes and monocytes, we postulated that morphine would promote the growth of HIV-1 in these cells. To test this hypothesis, a coculture assay was used consisting of phytohemagglutinin (PHA)-activated peripheral blood mononuclear cells (PBMC) from normal donors and PBMC which had been infected with a viral isolate from an asymptomatic patient, HIV-1AT. The growth of HIV-1AT, as reflected by the concentration of p24 antigen in coculture supernatants, was markedly increased in cocultures that contained morphine. A bell-shaped dose-response curve was observed with three- to fourfold increased growth at a morphine concentration of 10(-12) M. Augmentation of HIV-1AT growth by morphine required an interaction with the PHA-activated donor PBMC. Furthermore, potentiation of HIV-1AT growth by morphine was stereospecific and was antagonized by naloxone and beta-funaltrexamine indicating involvement of an opiate receptor mechanism. These findings provide an additional explanation of how opiates could act as a cofactor in the pathogenesis of HIV-1 in intravenous drug users.

Pharmacokinetics and safety of high-dose oral acyclovir for suppression of cytomegalovirus disease after renal transplantation.

The pharmacokinetics and safety of high-dose oral acyclovir for suppression of cytomegalovirus disease were evaluated in 12 patients undergoing renal transplantation. A 12-week course beginning 24 hours before transplantation was administered in doses of 800 to 3200 mg/day based on renal function. Acyclovir plasma concentrations were measured by RIA on posttransplant days 1 or 2 and 5, 6, or 7. Mean peak and trough concentrations on days 5, 6 or 7 were 25 and 18 mumol/L, respectively. The pharmacokinetic model predicted acyclovir concentrations with a precision of 4.1 mumol/L and bias of -1.19 mumol/L. Estimates of individual pharmacokinetic parameters were consistent with literature and a priori values. Two of six adverse events were attributable to acyclovir; both resolved with dose modification. The dosage adjustment scheme and pharmacokinetic model performed well, allowing us to safely administer high-dose oral acyclovir immediately after renal transplantation. We are proceeding with a placebo-controlled study to assess efficacy for suppression of posttransplant cytomegalovirus disease.

Human pharmacokinetics of the antiviral drug DHPG.

The pharmacokinetics of the antiviral drug 9-[2-hydroxy-1-(hydroxymethyl) ethoxymethyl]guanine (DHPG) were examined in six patients receiving 2.5 or 5.0 mg/kg every 8 or 12 hours for human cytomegalovirus (HCMV) pneumonitis or retinitis. Biexponential decay with a mean distribution t1/2 of 0.23 hours and terminal t1/2 of 2.53 hours was observed. Total clearance correlated well with and exceeded creatinine clearance by a factor of 2.4. Mean volume of the central compartment was 15.26 L/1.73 m2 and the volume of distribution at steady state was 32.8 L/1.73 m2. Peak (model predicted) and trough plasma concentrations were 4.75 to 6.20 micrograms/ml and less than 0.25 to 0.63 microgram/ml, respectively, in patients receiving 2.5 mg/kg. Peak concentrations are well above those needed to inhibit HCMV at the 50% level (ID50) and troughs are near this ID50. Cerebrospinal fluid concentrations of DHPG indicate a penetration of 24% to 67%. No accumulation of DHPG was apparent in these patients. However, dosage reduction is necessary in renal insufficiency. Neutropenia occurred in one patient. The plasma concentration profile of DHPG suggests potential beneficial activity against HCMV.

Concentration-controlled zidovudine therapy.

BACKGROUND: Heterogeneity in the response to antiretroviral therapy has been attributed to pharmacologic, immunologic, and virologic differences between patients. Currently available antiretroviral agents used for the treatment of human immunodeficiency virus (HIV) infection in adults are administered in standard fixed doses. The active moiety of nucleoside anti-HIV drugs is the intracellular anabolite. Therefore the heterogeneity in response to nucleoside agents may arise as a result of pharmacologic variability at both the systemic and cellular level. OBJECTIVES: To determine whether a novel concentration-controlled zidovudine regimen could improve anti-HIV response compared with the standard fixed-dose approach. DESIGN: At the Outpatient Clinic of the General Clinical Research Center at the University of Minnesota, 20 persons with HIV infection received an oral regimen of zidovudine designed to achieve a target concentration in plasma of 0.7 mumol/L and the 500 mg/day standard dose in a randomized, crossover 24-week study. RESULTS: The concentration-controlled regimen achieved overall higher systemic concentrations with reduced interpatient variability: steady-state average zidovudine plasma concentrations were 0.76 mumol/L (coefficient of variation, 12%) versus 0.62 mumol/L (coefficient of variation, 32%) for the standard regimen. There was no difference in safety and tolerance between regimens. Intracellular zidovudine triphosphate concentrations averaged 160 fmol/10(6) peripheral blood mononuclear cells (PBMCs) with concentration-controlled versus 92 fmol/10(6) PBMCs for standard therapy. The percentage change from baseline in CD4 cells was a 22% increase for the concentration-controlled regimen versus a 7% decrease with standard therapy. CONCLUSIONS: These data indicate that pharmacologic variability affects antiretroviral response. Furthermore, these findings provide a framework to characterize the pharmacologic determinants of effect and quantitate their contribution to the heterogeneity in clinical response to optimize therapeutic benefit.

Clinical and laboratory evaluation of cytomegalovirus-induced mononucleosis in previously healthy individuals. Report of 82 cases.

The present report describes the clinical and laboratory profile of 82 previously healthy individuals who developed cytomegalovirus (CMV)-induced mononucleosis. Many of these patients posed initial diagnostic problems and were hospitalized with diagnoses such as fever of undetermined origin, active viral hepatitis, acute leukemia, probable systemic lupus erythematosus, autoimmune hemolytic anemia, and severe pancytopenia. These patients underwent a variety of diagnostic biopsies, including liver biopsies (6) and bone marrow aspirations (9). Four patients had exploratory laparotomies, 1 for a ruptured spleen, and another had a splenectomy following an erroneous initial diagnosis of agnogenic myeloid metaplasia. There was no apparent clinical response to a short course of steroid therapy in 3 of 5 cases and acyclovir in another. The vast majority of these patients demonstrated infectious mononucleosis-type reactive blood smears, negative heterophil antibody studies, mildly or moderately elevated aspartate aminotransferase activity, and evidence for subclinical hemolysis on serial specimens. The peak serum bilirubin levels were above 2.0 mg/dl in only 2 of 71 cases tested, both of the latter patients having significant hemolysis (hemoglobin values 8.6-9.3 g/dl). The CMV-IgM test had a high sensitivity for detection of CMV macroglobulins (positive in 81 of 82 cases). In contrast, complement-fixing antibodies to CMV showed diagnostic four-fold titer changes in only 39/82 cases (47.6%). Despite its great sensitivity, the CMV-IgM test is limited by a one-way crossreaction of acute Epstein-Barr virus (EBV)-IM sera and spurious positive reactions in some sera due to the presence of rheumatoid factors. Based on EBV-specific serologic studies, the 82 patients with CMV-IM could be divided into 4 groups: 3 patients without antibodies to EBV; 2) 69 patients with uncomplicated serologic data indicative of long-past EBV infections; (3) 6 patients with unusual antibody profiles, e.g., anti-D responses; and (4) 5 patients, including 1 originally susceptible to EBV, with apparent dual CMV/EBV infections. At the conclusion of our study, final diagnoses and initial hematologic data were correlated in 750 cases in which CMV macroglobulins were searched for. The vast majority of patients with active CMV infections initially demonstrated either markedly or moderately reactive peripheral blood smears. These data support our impression that diagnostic tests for CMV, as well as for EBV, are seldom indicated in symptomatic previously healthy patients whose blood smears during the acute phase (first several weeks) of their illnesses are either nonreactive or minimally reactive.

Cytomegalovirus disease in renal allograft recipients: a prospective study of the clinical features, risk factors and impact on renal transplantation.

Fifty-nine renal transplant recipients with overt CMV disease were treated at the University of Minnesota Health Sciences Center between October 1, 1977 and November 15, 1978. In a group of 141 consecutive transplant patients, the incidence of overt CMV disease was 31%. Fifty-three patients (90%) developed clinical manifestations of CMV disease within 4 months of transplantation, and it was during this time period that overt CMV disease was associated with a significantly increased incidence of transplant nephrectomy and death. Fever was the most common presenting symptom (95% of patients), and overt CMV disease was found to be the single most common cause of fever in all hospitalized transplant recipients. Prolonged fever, diffuse pulmonary infiltrates, gastrointestinal bleeding, pancreatitis, transplant nephrectomy and development of other systemic infections were clinical features used to categorize patients according to disease severity. A number of these features were found to be significantly associated with the diagnosis of overt CMV disease. Twelve patients (20%) developed lethal CMV disease characterized by the presence of most of these features, 6 (10%) had severe disease, 9 (15%) had disease of moderate severity and 32 patients (54%) had mild CMV disease with fever being essentially their only clinical finding. Development of secondary systemic infection was most ominous, and occurred before death in 10 of the 12 patients with lethal CMV disease. The only patients to die with serious bacterial, fungal or protozoan infection during the period of this study had concomitant overt CMV disease. Abnormal liver function tests and leukopenia were common, and the degree of abnormality correlated with the severity of CMV disease. Of the multiple factors analyzed for their influence on the risk of developing overt CMV disease, several factors related to the kidney donor (the relationship of the donor to the recipient, HLA matching and CMV serology) appeared to be most important.

Comparison of two serum HIV antigen assays for selection of asymptomatic antigenemic individuals into clinical trials.

The detection and recruitment of HIV antigen-positive asymptomatic individuals for clinical trials is important. Two commercial enzyme-linked immunosorbent assays (ELISA) for the detection and quantitation of human immunodeficiency virus (HIV) antigens were evaluated for sensitivity by testing serum samples from 155 asymptomatic HIV Western blot positive individuals. The Abbott HIV antigen ELISA detected HIV antigen in the serum of 17 (11.0%) of 155 patients compared with 18 (11.6%) of 155 by the Coulter HIV antigen ELISA. In serial twofold dilution experiments, there was no significant difference in sensitivity between these two assays in the detection of HIV serum antigen. However, both assays are limited in their ability to detect HIV antigen in most asymptomatic HIV-infected patients. This low detection rate should be taken into account in the design of clinical trials involving asymptomatic infected patients.

Therapeutic algorithm for treatment of cytomegalovirus retinitis in persons with AIDS. A roundtable summary.

Foscarnet and ganciclovir appear to be of similar effectiveness in halting active infection when given as induction therapy and in forestalling progression of disease when given as maintenance therapy in persons with AIDS who have cytomegalovirus (CMV) retinitis. The primary dose-limiting toxicity of foscarnet is nephrotoxicity, whereas that of ganciclovir is neutropenia. The availability of two effective agents with different toxicities permits selection of initial treatment for CMV retinitis based on individual patient characteristics and provides an alternative for therapy if drug intolerance or viral resistance develops. An approach to treatment of first-episode and recurrent CMV retinitis based on patient and drug characteristics is presented. Case reports detailing the use of foscarnet and ganciclovir and problems encountered in patient management are discussed.

Management of acyclovir-resistant herpes simplex and varicella-zoster virus infections.

Persons with AIDS who have CD4+ counts < or = 100 and transplant patients, especially bone marrow allograft recipients, may experience clinically significant infections with acyclovir-resistant herpes simplex virus (HSV) or varicella-zoster virus (VZV). Patients who have received prior repeated acyclovir treatment appear to be at the highest risk of harboring acyclovir-resistant strains. Algorithms for the management of these infections were developed at a recent roundtable symposium. The consensus of the panelists was that treatment with foscarnet should be initiated within 7-10 days in patients suspected to have acyclovir-resistant HSV or VZV infections. Foscarnet therapy should be continued for at least 10 days or until lesions are completely healed.

Increasing CD8+ T lymphocytes predict subsequent development of intraoral lesions among individuals in the early stages of infection by the human immunodeficiency virus.

Determining the progression of human immunodeficiency virus (HIV) type 1 infection based on cellular and clinical markers has become increasingly important. Although a number of studies have shown a relationship between the presence of certain oral lesions and progression to AIDS, few data exist regarding the association with T lymphocyte counts. In this study, the question of whether intraoral lesions preceded or were the consequences of changes in T lymphocyte counts was examined. A total of 116 HIV-infected patients participating in two randomized double-blind placebo-controlled trials of zidovudine at the University of Minnesota AIDS Clinical Trials Unit (ACTU) were enrolled in a prospective dental study. Patients were examined for the presence of hairy leukoplakia, candidiasis, herpes simplex, herpes zoster, aphthae, atypical gingivitis, HIV-associated periodontitis, and necrotizing ulcerative gingivitis, as well as other oral lesions, every 3 months for a maximum of four examinations over a 1-year period. T lymphocyte counts before and after each patient's oral examination were obtained. No significant differences were found at examination 1 for differences in gender, race, age, education, tobacco smoking status, ethanol consumption habits, duration in ACTU drug protocol, duration in dental study protocol, or mean T lymphocyte counts between individuals with or without oral lesions at any time in the dental study.(ABSTRACT TRUNCATED AT 250 WORDS)

Varicella zoster virus infections in immunocompromised hosts. A review of the natural history and management.

Varicella is relatively mild in otherwise normal children, in whom new lesions form for a mean of four days after onset and heal 50 percent of their lesions in eight days. New lesions form in most immunocompromised children for longer than five days and those not treated with antiviral drugs have a 28 percent incidence of pneumonitis and a 7 percent mortality rate. Untreated immunocompromised adults with herpes zoster shed virus for longer (7.0 days) than otherwise normal adults (5.3 days). Herpes zoster is much more likely to disseminate cutaneously in immunocompromised than in immunocompetent hosts. Visceral dissemination, which is a rare event in immunocompetent patients, occurred in 8 percent of prospectively followed untreated immunocompromised hosts with herpes zoster. Acyclovir has been found to be superior to vidarabine for treatment of both chickenpox and herpes zoster. Whether or not steroids should be used to treat herpes zoster remains controversial. Concerns about the use of intravenous acyclovir include the side effects of renal and central nervous system dysfunction and the possibility of emergence of resistant viral strains. None of these concerns has proved to be an impediment to successful treatment of immunocompromised patients. The major future challenge is to find an optimal way to treat varicella zoster virus infections with oral formulations of acyclovir or its congeners.

Fever in renal transplant recipients: causes, prognostic significance and changing patterns at the University of Minnesota Hospital.

During a three year period in which 433 renal transplants were performed, 194 episodes of fever were documented in allograft recipients hospitalized at the University of Minnesota. Viral infections were responsible for over half of the febrile episodes, and 98 (51 percent) of the fevers were associated with cytomegalovirus (CMV), either occurring alone or in conjunction with allograft rejection or another systemic infection. Bacterial infections, fungal infections and rejection were other important causes of fever, accounting for 14 percent, 5 percent and 13 percent of the febrile episodes, respectively. Most fevers occurred in the first four months after transplantation; although about two thirds of these fevers were due to CMV, only 17 percent of fevers that occurred more than one year after the renal transplant were due to CMV. Bacterial and fungal infections and malignancy were important causes of these fevers. Of the febrile illnesses associated with transplant nephrectomy or death, a majority occurred in patients with CMV disease. Secondary bacterial and/or fungal infections were observed in a large majority of patients with lethal CMV disease. During the third year of this study there was a significant decrease in the proportion of febrile episodes due to CMV.

Therapy of herpes zoster with oral acyclovir.

Oral acyclovir therapy for herpes zoster has been studied in double-blind, placebo-controlled trials of two dosages, 400 mg and 800 mg five times per day for 10 days. Compared with placebo recipients, recipients of the high-dosage acyclovir experienced a significantly shortened period of viral shedding, significantly accelerated time to 50 percent scabbing, significantly accelerated time to 50 percent healing, and after two days of therapy, significantly less frequent formation of new lesions. The duration and severity of acute pain were less in acyclovir recipients, with differences in pain severity achieving statistical significance (p = 0.03) between Days 3 and 10 and correlating with the treatment differences in new lesion formation. In studies of the 400 mg five times per day dose schedule, differences between acyclovir and placebo recipients were not significant. In a six-month follow-up of recipients in the higher dosage study, the acyclovir recipients experienced less post-zoster pain than placebo recipients; differences in the prevalence of pain were most significant for the presence of a persistent pain in the first three months of follow-up. Oral acyclovir at these dosages appears to be free of adverse reactions. In summary, oral acyclovir at a dosage of 800 mg five times per day for 10 days for treatment of acute herpes zoster is superior to 400 mg five times per day and favorably alters the course of the disease.