George F. Grady

Use of Cytomegalovirus Immune Globulin to Prevent Cytomegalovirus Disease in Renal-Transplant Recipients

We undertook a prospective randomized trial to examine whether an intravenous cytomegalovirus (CMV) immune globulin would prevent primary CMV disease in renal-transplant recipients. Fifty-nine CMV-seronegative patients who received kidneys from donors who had antibodies against CMV were assigned to receive either intravenous CMV immune globulin or no treatment. The immune globulin was administered in multiple doses over the first four months after transplantation. The incidence of virologically confirmed CMV-associated syndromes was reduced from 60 percent in controls to 21 percent in recipients of CMV immune globulin (P less than 0.01). Fungal or parasitic superinfections were not seen in globulin recipients but occurred in 20 percent of controls (P = 0.05). Only 4 percent of globulin recipients had marked leukopenia (reflecting serious CMV disease), as compared with 37 percent of the controls (P less than 0.01). There was a concomitant but not statistically significant reduction in the incidence of CMV pneumonia (17 percent of controls as compared with 4 percent of globulin recipients). A significant reduction in serious CMV-associated disease was observed even when patients were stratified according to therapy for transplant rejection (P = 0.04). We observed no effect of immune globulin on rates of viral isolation or seroconversion, suggesting that treated patients often harbored the virus but that clinically evident disease was much less likely to develop in them. We conclude that CMV immune globulin provides effective prophylaxis in renal-transplant recipients at risk for primary CMV disease.

The pathogenesis of Shigella Diarrhea: I. Enterotoxin production by Shigella dysenteriae 1

A strain of Shigella dysenteriae 1, freshly isolated from a patient with dysentery in Guatemala in August 1969, was found to elaborate an enterotoxin into the liquid of broth cultures. Partial purification of the enterotoxin by ultrafiltration on graded polymeric membranes and Sephadex gel filtration (Pharmacia Fine Chemicals, Inc., Piscataway, N. J.) suggested an approximate molecular weight of 55,000-60,000. The partially purified toxin was heat-labile, pronase sensitive, and activated by alkaline pH, and it elicited fluid production in ligated rabbit ileal segments; it failed, however, to cause increased vascular permeability in skin. When the activities of equal weights of identically prepared Vibrio cholerae and S. dysenteriae enterotoxins were compared in the rabbit ileum the latter caused a significantly smaller volume response with increased concentrations of potassium, chloride, and protein. The previously described neurotoxic (mouse lethal) factor was also present and eluted from Sephadex G-150 with the enterotoxin. If these biological activities prove to be possessed by a single molecular species, it is suggested that it be renamed Shigella enterotoxin in recognition of the physiologically more relevant biological action.

Accidental Hepatitis-B-Surface-Antigen-Positive Inoculations: Use of e Antigen to Estimate Infectivity

We assessed the ability of radioimmunoassay for hepatitis B e antigen (HBeAg) to predict infectivity in exposed medical personnel by analyzing 390 samples of sera positive for hepatitis B surface antigen (HBsAg) that were implicated in accidental inoculations of known outcome. The radioimmunoassay detected HBeAg or its antibody (anti-HBe) in 91% of the donor sera. The incidence of hepatitis B was 19% (44 of 234) in recipients of HBeAg-positive sera but was only 2.5% (three of 121) in recipients of sera positive for anti-HBe, and nil (none of 35) in recipients of sera negative for HBeAg and anti-HBe. The known relation of HBeAg and infectivity was quantified by radioimmunoassay as a risk ratio of 10:1 (HBeAg-positive to HBeAg-negative) for this type of exposure. The sensitivity of the radioimmunoassay also showed that a large proportion (55%) of donor sera not producing hepatitis were positive for HBeAg; therefore, even the most flagrant needlestick exposures to HBsAg-positive sera often must involve subthreshold amounts of infective material.

Seroprevalence of human immunodeficiency virus among childbearing women: estimation by testing samples of blood from newborns

Attempts to predict the course of the epidemic of acquired immunodeficiency syndrome (AIDS) have been hampered by the lack of an objective, practical way to estimate the prevalence of infection with the human immunodeficiency virus (HIV) in the general population. Testing for the prevalence of HIV infection in women should be a sensitive means to track the epidemic and to study the potential for perinatal transmission. Antibodies in maternal blood are contained in neonatal blood specimens routinely collected on absorbent paper for other purposes, such as screening for phenylketonuria; we therefore tested for HIV antibody in these specimens. Analysis of batches of individually blinded specimens from selected hospitals protected the anonymity of the mothers and babies and was cost efficient. Using the newborns blood as an indicator of the mothers serologic status, we concluded that 1 of every 476 women (2.1 per 1000) giving birth in Massachusetts was positive for HIV antibody by immunofluorescence assay or enzyme-linked immunosorbent assay, both confirmed by immunoblot (Western blot) testing. The prevalence of HIV infection varied according to the type and location of the maternity hospitals; rates of seropositivity were highest in inner-city hospitals (8.0 per 1000), lower in mixed urban and suburban hospitals (2.5 per 1000), and lowest in suburban and rural hospitals (0.9 per 1000). This method is useful for collecting data needed to plan and evaluate prevention strategies and to predict the health care resources that will be needed to care for women and children who contract AIDS. Because other states have newborn screening programs similar to the Massachusetts program, this approach can be used for national surveillance of AIDS in women.

Nonresponsiveness to hepatitis B vaccine in health care workers. Results of revaccination and genetic typings.

Twenty-eight health care workers who had a poor antibody response when initially vaccinated with hepatitis B vaccine were revaccinated with three additional 20-microgram doses. Eight of the twenty nonresponders, who had levels of antibody to hepatitis B surface antigen (anti-HBs) of less than 8 estimated radioimmunoassay (RIA) units, and all 8 of the hyporesponders, who had anti-HBs levels of 8 or 16 RIA units, attained anti-HBs levels of 36 RIA units or more after revaccination. Tests for HLA-A, B, C, and DR; for complement proteins C2, C4A, C4B, and BF; and for the erythrocyte enzyme glyoxalase I were done in 17 nonresponders and 3 hyporesponders. Nine (45%) had HLA-DR7 and 8 (40%) had HLA-DR3, compared with an expected rate of 23% in the general population. At least one of two extended haplotypes (B44, DR7, FC31 or B8, DR3, SCO1) were detected in 6 of the 9 who did not respond to revaccination, compared with 2 of 11 who responded to a second course of vaccine. Poor responders to vaccine may benefit from revaccination, and genetic factors may modulate the immune response to vaccination.

Hepatitis B Vaccine in Health Care Personnel: Safety, lmmunogenicity, and Indicators of Efficacy

In a double-blind trial, we randomly assigned 1330 high-risk health care personnel to receive three 20-micrograms doses of hepatitis B vaccine or placebo. Among vaccine recipients 58% responded within 1 month and 97% within 9 months; there was no difference in immune response to the vaccine between men and women. Efficacy was evaluated after a mean follow-up of only 13.2 months, just before the vaccine was released commercially. Five hepatitis B infections were identified in placebo recipients and one in a vaccine recipient. Although the number of infections was too small to allow confident conclusions about protective efficacy of the vaccine, we saw a 67% reduction in the need for hepatitis B immune globulin after accidental hepatitis B inoculation in the vaccine group (relative risk, 5.08; 95% confidence intervals, 1.3 to 19.9). Minor side effects occurred with equal frequency after vaccine (28.7%) and placebo (27.2%) injections; no participant had a severe adverse reaction. Vaccination with the 20-micrograms hepatitis B vaccine was highly immunogenic and safe in health care workers.

Cytomegalovirus immune globulin prophylaxis in liver transplantation. A randomized, double-blind, placebo-controlled trial.

Cytomegalovirus (CMV) is the most frequent viral pathogen complicating organ transplantation [1]. In renal transplantation, prophylactic cytomegalovirus immune globulin (CMVIG) can reduce the rate of CMV-associated disease by 50% or more in kidney recipients at risk for primary CMV disease (that is, donor, CMV-seropositive; recipient, CMV seronegative) [2, 3]. Further, CMVIG has also been shown to reduce the risk for serious CMV disease even in those patients given additional immunosuppressive treatment for rejection [2, 3]. The widespread use of liver transplantation has been accompanied by the recognition of CMV disease as a serious complication in about 30% of patients [4]. Although some researchers have suggested that cytomegalovirus immune globulin or unselected immune globulin plus acyclovir may help prevent CMV in liver transplantation [5, 6], no single strategy has emerged as the standard of care. Furthermore, studies of CMV prevention in liver transplantation have been limited by the small number of patients studied and the lack of any randomized clinical studies [5-7]. Therefore, we performed a randomized, multicenter, placebo-controlled, double-blind trial of CMV disease prevention in liver transplant recipients. Methods Study Sample and Protocol Eligible patients included children and adults who had liver transplants at the participating centers of the Boston Center for Liver Transplantation: Childrens Hospital, Massachusetts General Hospital, New England Deaconess, and New England Medical Center. The study protocol was approved by the human-investigation review committee of each institution. Liver-transplant recipients received either prophylactic intravenous CMV immune globulin (CMVIG) or placebo. All candidates for transplantation and all liver donors were screened for CMV antibody. The CMV antibody status of donors and recipients did not influence eligibility for the trial. Based on the estimate that CMV disease would develop in 50% to 60% of liver transplant recipients [8], we tested the hypothesis that the rate of CMV disease could be reduced by 50% in globulin recipients. We anticipated that we would need to study 65 patients in each group to have an 80% chance of identifying a significant effect (P = 0.05) of CMV immune globulin. Cytomegalovirus Immune Globulin The CMVIG was produced by the Massachusetts Public Health Biological Laboratories, used in previous clinical trials [2, 3] and licensed for use in the United States [9]. Details regarding CMV antibody titers and production have been previously described [2, 3, 10]. Patients who were randomly assigned to receive intravenous CMVIG were given 150 mg/kg body weight within 72 hours of transplantation, repeated doses at 2, 4, 6, and 8 weeks after transplantation, and 100 mg/kg 12 and 16 weeks after transplantation. This regimen was chosen after a pilot study showed that this dose would maintain antibody titers of 8 or higher as measured by indirect hemagglutination for up to 5 months after transplantation [11]. The placebo was 1% serum albumin, packaged in a manner identical to that of CMVIG. Prenumbered vials for each patients series of infusions were used. Randomization was performed in blocks of four, with each center having a separate randomization code and its own series of vials. Clinical Management and Immunosuppression After enrollment, patients were followed weekly for the first 8 weeks after transplantation, monthly for the next 6 months, and at 1 year after transplantation. Clinical care was individualized by the transplantation team at each institution. Baseline laboratory studies were done at enrollment, weekly for 2 months, when special problems arose, and at scheduled follow-up visits. Standard immunosuppressive drugs included prophylactic cyclosporine, azathioprine, and steroid, given intravenously soon after transplantation, with a transition to oral administration when gastrointestinal function allowed. Azathioprine was withheld during periods of leukopenia. During the study, 35% of patients received prophylactic murine monoclonal antibody to T3 antigen (OKT3, Orthoclone, Ortho Biotech; Raritan, New Jersey) as part of a concurrent randomized trial comparing prophylactic OKT3 with standard triple immunosuppression. These patients received lower doses of cyclosporine. Acute rejection was treated primarily with intravenous bolus infusion of methylprednisolone; refractory or recurrent rejection was treated with OKT3. Virologic Studies Urine and throat-wash specimens (or throat swabs in young children) for viral isolation and serum for analysis of CMV antibody were collected before the first infusion, weekly for 2 months, and then monthly for 6 months. Peripheral-blood leukocytes were obtained for viral isolation every other week for the first 2 months and then every month for 6 months as well as at the time of clinical illness compatible with CMV disease and at the end of 1 year. In addition, tissue specimens obtained by biopsy and at autopsy were cultured for virus [12]. Postinfusion serum samples were also obtained from all patients. For routine CMV cultures, human foreskin fibroblasts and MRC-5 cells were used at the different centers and examined regularly for viral cytopathic effects for up to 6 weeks [13]. Cytomegalovirus culture with early antigen detection using the shell-vial assay was used at several centers to rapidly diagnose CMV infection on various specimen types when clinically indicated [14]. At the time of transplantation, CMV antibody was measured in all patients and liver donors serum samples by complement fixation, latex agglutination, and enzyme-linked immunosorbent assay [13]. The enrollment and termination samples and selected serum samples collected during the time of illness were tested by complement fixation, latex agglutination, indirect hemagglutination, and enzyme-linked immunosorbent assay. To confirm seroconversion in the presence of exogenously administered globulin, we tested selected serum samples for CMV-specific immunoglobulin M antibody by enzyme immunoassay (M.A. Bioproducts; Walkersville, Maryland). The serologic criteria were based on complement fixation as the primary test method for our analysis. We defined seronegativity as a titer of less than 1:8. We defined seroconversion as an increase in complement fixation titer from less than 1:8 to 1:16 or greater. We relied on complement fixation for determinations of seroconversion to avoid the confounding effect of exogenously administered antibody observed in other assays, whether from multiple transfusions administered during or after surgery or globulin administration. We used enzyme-linked immunosorbent assay, indirect hemagglutination, and latex agglutination to supplement the complement fixation assay when needed to clarify a lack of agreement with a donor procurement result or when anticomplementary activity was present. All decisions on serologic classification were made blinded as to randomization and before breaking the code. Antiviral Use High-dose acyclovir (>2 g/d for 30 days or more) was used in a few patients after the report by Balfour and colleagues was published [15]. Until May 1989 ganciclovir use was restricted to established CMV disease on a compassionate protocol. Throughout the study, use was determined at each institution on the basis of clinical judgment and availability from the manufacturer. For patients in whom CMV pneumonia or clinically severe CMV disease developed, we allowed combination therapy with ganciclovir and CMVIG according to a modification of combination regimens for treating CMV pneumonia after bone marrow transplantation [16-18]. For these patients a standard dose of ganciclovir was given along with CMVIG in a dose of 100 mg/kg every other day for 14 days. Decisions to use combination therapy were left to the discretion of each investigator. Analytical Framework and Case Definitions Before beginning the study, we defined as primary end points the effect of CMVIG on rates of CMV disease, CMV pneumonia, severe CMV-associated disease (as defined below), viremia, mortality, and graft survival. All case and severity of illness classifications were made by an outside group of three consultants blinded to the randomization code. Final classification required unanimity of opinion. The analytic framework and all outcome measures were agreed on before breaking the code. We considered as dropouts all patients who died fewer than 15 days after transplantation and all patients who received fewer than two doses of CMVIG or placebo. We defined CMV disease as the presence of clinical evidence of organ dysfunction along with biopsy proof of CMV in the affected organ, as documented either by virus isolation or histologic evidence of CMV in that organ. We only considered CMV disease to be present if disease was virologically confirmed, that is, at the time of the disease, either CMV was isolated from a clinical specimen, antibody seroconversion occurred within 1 month of the event, or histopathologic evidence of CMV infection was found. In addition, we defined a clinical CMV syndrome as illness having two or more of the features known to be related to CMV (unexplained fever for at least 3 days in association with one of the following: pneumonitis without other cause, leukopenia [<4.0 109/L leukocytes/mm3] or thrombocytopenia (platelets <100 109/L) on 3 or more consecutive days after withdrawal of azathioprine or ganciclovir, or atypical lymphocytosis [>5% of peripheral leukocytes]). Because liver transplant recipients frequently have abnormal liver biochemistry test results, chemical evidence of hepatitis was not included in the syndrome definition. We classified CMV pneumonia as definite if there was histologic, biopsy, or autopsy evidence of typical intranuclear inclusions or if a bronchoalveolar lavage specimen had cytologic evidence of intranuclear inclusions in a patient with organ dysfunction (hypoxia, in

Outbreak of Severe Hepatitis Due to Delta and Hepatitis B Viruses in Parenteral Drug Abusers and Their Contacts

We investigated an unusually large and severe outbreak of hepatitis B, primarily involving parenteral drug abusers and their sexual contacts, in Worcester, Massachusetts, over a 21-month period from 1983 to 1985. Of 135 patients with drug-related acute hepatitis B, 81 percent were parenteral drug abusers and 19 percent had sexual contact with drug abusers; 13 fulminant cases resulted in 11 deaths. Among the patients with hepatitis B, evidence of delta virus infection was found in 54 percent of drug abusers, 33 percent of their sexual contacts, and 9 percent of other patients with acute hepatitis B (P less than 0.001). Most of the delta infections (86 percent) were coinfections with hepatitis B virus; the balance were superinfections. Delta infection was strongly associated with fulminant hepatitis: 91 percent of patients with a fulminant outcome had delta infection, as compared with 45 percent of less severely ill drug abusers and their contacts (P = 0.0037). Alcohol, other drugs, and other hepatitis viruses could not be implicated as hepatotoxic cofactors for fulminant disease. This outbreak appeared to result from the concurrent spread of hepatitis B and delta viruses among new drug users. Control measures included the distribution to physicians of guidelines on prophylaxis in contacts of patients with hepatitis B, health education for drug abusers, and a hepatitis B vaccination program. Despite these efforts, the outbreak continued unabated until the number of new cases began to decline slowly in late 1986.

Pathogenesis of Bacterial Diarrheas

NO less than a renaissance has emerged in studies of diarrhea, in spite of the unattractive attributes of feces, the difficulty in identifying pathogens among a sea of other bacteria, and the conse...

Hepatitis B immune globulin-prevention of hepatitis from accidental exposure among medical personnel.

The role of anti-HBs antibody in reducing the probability of hepatitis after accidental exposure to serum from patients with hepatitis or carriers of HBs Ag was studied prospectively among 712 medical workers. One fourth of the workers were anti-HBs positive and less than one per cent of them developed hepatitis, in contrast to 11 per cent among those who were anti-HBs negative. Three coded immune-serum globulin preparations of varying anti-HBs titer were randomly assigned. Among 251 persons passively immunized with globulin having a conventionally low anti-HBs titer, hepatitis developed in 17 (seven per cent) within six months. Comparative rates among those receiving intermediately high titer and high titer globulin, respectively, were five per cent (11 of 208) and two per cent (5 of 253). The significantly lower incidence among the latter (P less than 0.05) was offset by six additional cases, all in recipients of high titer globulin, detected when follow-up was extended to nine months.