Cell wall composition and virulence in Escherichia coli.

Uridine diphosphate galactose 4-epimerase and phosphomannose isomerase-deficient mutants of Escherichia coli O111:B4 were studied to test the hypothesis that in E. coli a specific relationship exists between O antigenicity, virulence, and capacity to resist phagocytosis. The first mutant, designated J-5, produces a cell wall lipopolysaccharide, the side chains of which do not contain galactose, glucose, N-acetylglucosamine, or colitose. The second mutant produces a cell wall lipopolysaccharide which lacks only colitose. The capacity of these various organisms to kill mice was strikingly different. E. coli O111 was 1000 times as virulent as J-5, and 100 times as virulent as L-2. The capacity of the organisms to kill mice was correlated with their ability to resist phagocytosis and to persist in the peritoneal cavity. The parent strain of O111 resisted phagocytosis by macrophages in vivo and polymorphonuclear leukocytes in vitro. The mutants did not, and the organism most deficient in the saccharide component of its LPS was most susceptible to phagocytosis and least virulent. These results were corroborated by growing the mutants in appropriately supplemented media which permitted the synthesis of complete LPS, reversed the susceptibility to phagocytosis, and restored virulence. Finally, serological reactivity was consistent with previous observations which had demonstrated that the O antigenicity of E. coli is determined by the saccharide composition of its cell wall lipopolysaccharide. Despite the difference in the capacity of the various log-phase organisms to kill mice when injected intraperitoneally, purified lipopolysaccharides extracted from them did not differ significantly in their capacity to kill or produce fever. Thus virulence was shown to be independent of endotoxin activity which in turn seemed to be unrelated to the saccharide composition of the cell wall LPS. Collectively, these data provide at least a partial molecular definition of virulence in E. coli by demonstrating that the presence or absence of specific sugars in its cell wall lipopolysaccharide is a determinant of its antiphagocytic capacity and its virulence.

Primary cytomegalovirus infection following cardiac transplantation in a murine model.

A murine cytomegalovirus (CMV) model was utilized to determine the source of primary CMV infection in cardiac transplantation. Hearts were taken from actively or latently infected BALB/C mice and then transplanted as primary, heterotopic isografts into CMV-negative BALB/C recipients. The transplantation of hearts from acutely infected donors into nonimmunosuppressed recipients resulted in asymptomatic primary infection as manifested by detectable virus in both donor and recipient hearts, liver, spleen, and salivary glands and by the development of anti-CMV antibody. When hearts from latently infected animals were transplanted into nonimmunosuppressed recipients, a transient primary infection occurred that was manifested by detectable virus in the spleen and salivary glands and the appearance of anti-CMV antibody. When recipient animals were immunosuppressed with cortisone acetate (125 mg/kg/day i.p.) and rabbit antimouse thymocyte globulin (0.2 ml i.p. twice weekly), after transplantation of hearts from acutely and latently infected mice, lethal primary CMV infection developed. High titers of virus were recovered in all organs tested in these animals, including both the donor and recipient hearts. We conclude that the heart is infected during the course of primary murine CMV infection, and that hearts from latently infected animals are a source of serious primary infection in immunosuppressed recipients. This experimental system should be a useful model relevant to human cardiac transplantation.

Ethical dilemmas in child and adolescent consultation psychiatry.

Ethical issues in child and adolescent psychiatry consultation arise frequently but seldom are discussed in a public setting. This case of an adolescent victim of a surgical accident illustrates many aspects of consultation psychiatry. The consult question itself, of behavior management, is not unusual, although in this case the question is complicated by the sequelae of trauma, psychosocial chaos, and the staff's angry feelings toward the patient. In addition, potential surgical wrongdoing at the referring hospital brings up the more difficult ethical questions of the consultant's responsibilities, which must be to the patient and his family, as well as to the attending and referring physicians.

Cell-mediated immunity in experimental cytomegalovirus infections: a perspective.

Treatment of cytomegalovirus (CMV) infections has assumed increasing importance in the care of immunocompromised patients, particularly allograft recipients and patients with symptomatic infection with the human immunodeficiency virus. The crucial role of T cells in the host defense against CMV may be inferred by the expression of life-threatening CMV disease in these restricted patient subpopulations. However, detailed understanding of the pathogenesis of CMV disease in the compromised host is lacking, and the mechanisms by which T cells confer protection are unknown. Insight into the pathogenesis of CMV in these patients is crucial for the development of rational therapeutic strategies. A considerable body of data has been generated to explore the role of T cell-mediated immunity in a number of model viral infections as well as in experimental CMV infections in both normal and immunocompromised mice. This experience will be critically reviewed to clarify the often conflicting observations regarding a protective or, conversely, a pathogenic role for T cells in producing CMV disease.

The protective effects of hyperimmune anti-murine cytomegalovirus antiserum against lethal viral challenge: the case for passive-active immunization.

The administration of 0.2 ml of hyperimmune anti-mouse cytomegalovirus (CMV) antiserum intraperitoneally (ip) or intravenously provided complete protection against lethal challenge (10(5.8) PFU ip) with murine CMV. Antiserum protection was complete when the antiserum was administered as long as 24 hr after viral challenge. The administration of antiserum had little effect on the titers of virus in the organs of these animals. Ammonium sulfate-treated antiserum provided similar complete protection. Animals rechallenged with 10(6)-10(6.5) PFU of murine CMV 1 month after initial challenge, at a time when the administrated antiserum was no longer detectable, all survived. We conclude that hyperimmune antiserum can provide significant protection against otherwise lethal murine CMV infection, that the protecting material lies within the immunoglobulin fraction, and that long-term immunity results from the combined exposure to virus and antiserum. Such passive-active protection could be useful in protecting against human CMV infection.

9-(1-3-Dihydroxy-2-propoxymethyl)guanine prevents death but not immunity in murine cytomegalovirus-infected normal and immunosuppressed BALB/c mice.

Immunosuppressed (from treatment with cortisone acetate and anti-thymocyte globulin) and control adult female BALB/c mice, latently infected with murine cytomegalovirus (MCMV) or lethally challenged (10(6) PFU) with MCMV intraperitoneally, were treated with 9-(1-3-dihydroxy-2-propoxymethyl)guanine (DHPG) intraperitoneally. A dose of 3 mg/kg reduced mortality by 50% in lethally challenged normal mice; 10 mg/kg was required in immunosuppressed mice. When 15 mg/kg was given, the onset of treatment could be delayed for 64 h after lethal challenge. DHPG did not prevent the establishment of latent MCMV infection or immunosuppression-induced reactivation. The antibody titer to MCMV in DHPG-treated mice which survived lethal challenge was 41 (reciprocal of geometric mean) 4 to 5 weeks after inoculation; such mice survived a second lethal challenge. When antiserum treatment was begun 64 h and DHPG was begun 72 h after a lethal challenge, most mice survived; most did not when either treatment alone was begun at those times. In summary, DHPG effectively treated lethal MCMV infection even in immunosuppressed mice and even when treatment onset was delayed for 64 h. Treatment did not alter the establishment or reactivation of latent infections or the induction of effective immunity. The administration of DHPG coupled with antiserum treatment may be even more effective than the administration of either alone.

Combined antibody and ganciclovir treatment of murine cytomegalovirus-infected normal and immunosuppressed BALB/c mice.

The efficacy of treatment with ganciclovir (DHPG) and antibody activity-containing ascitic fluid (AF) separately and in combination was studied in normal and immunosuppressed BALB/c mice challenged intraperitoneally with a lethal dose (10(6) PFU) of murine cytomegalovirus (CMV). With combination therapy, lower doses of both DHPG and AF were often as effective as a higher dose of either agent given singly. For instance, the survival rate of murine CMV-challenged immunosuppressed mice was doubled when 4 mg of DHPG per kg and a 1:16 dilution of AF were both administered in contrast to when each was used alone. In both groups of animals, combination therapy was shown to be more effective than either therapy individually, even when initiation of therapy was delayed as long as 48 h. Such an approach holds promise for decreasing the expense associated with antibody use and the dose-related toxicity associated with DHPG use while maintaining or possibly increasing the efficacy of prophylaxis and therapy of serious CMV disease in humans.

Changes in the phenotype of T-cell subset determinants following murine cytomegalovirus infection.

The murine model provides a particularly apt experimental system in which to evaluate the effects of cytomegalovirus (CMV) infection. CMV exerts a profound "suppression" of the immune response in the mouse and in humans; the infected animal is no longer able to mediate an appropriate response to mitogens or alloantigens. Using fluorocytometry and fluoresceinated monoclonal antibodies directed against the Thy-1.2, Lyt-1, and Lyt-2 cell membrane determinants following a nonlethal intraperitoneal inoculation of weanling BALB/c mice with Smith strain murine cytomegalovirus, significant changes in T-cell subsets were found that are consistent with findings described in the clinical situation. These ratio changes are temporally consistent with the cytotoxic T-lymphocyte population described by others. Finally, novel changes in the antigenic determinant distribution is found which may reflect the appearance of an antigen-committed cytotoxic T-lymphocyte population. This population which peaks at the ninth postinfection day may consist of 20-47% of the T-lymphocyte population and may offer an explanation for the cellular hyporesponsiveness seen following CMV infection.

Myopericarditis and enhanced dystrophic cardiac calcification in murine cytomegalovirus infection.

In mice inoculated with murine cytomegalovirus (MCMV) an acute myopericarditis developed which varied from a focal lymphohistiocytic inflammation to intense inflammation with necrosis and cytomegalic inclusion-bearing cells. Sublethal doses caused focal transient nonspecific chronic inflammation, followed months later by an increased frequency and extent of dystrophic cardiac calcification. When such latently infected hearts were heterotopically transplanted into uninfected animals which were then immunosuppressed (IS), a fatal generalized CMV infection followed. Cytomegalic inclusion-bearing endothelial, fibroblastic, and myocardial cells were seen in the intense inflammation found in hearts taken from mice 4 days after lethal inoculation and transplanted into uninfected mice, which were then IS. These findings may be relevant to human cardiac transplantation because they show that MCMV regularly causes cardiac infection with both acute and chronic consequences; chronic injury may follow a morphologically nonspecific myopericarditis which might not be attributed to CMV infection.

The in vivo and in vitro effects of interleukin-1 and tumor necrosis factor on murine cytomegalovirus infection.

The effects of tumor necrosis factor (TNF) and interleukin-1 (IL-1) on infection with murine cytomegalovirus (MCMV) were investigated in vitro and in vivo. The addition of each of these cytokines (at 1 ng/ml) to tissue culture monolayers 24 hr prior to MCMV challenge produced a reproducible decrease in vital titer (from 1 x 10(8) pfu to approximately 4 x 10(6) pfu for both cytokines). There was no further increase in this effect when a 10 or 100 ng/ml of each of these cytokines was employed. Despite these in vitro effects, the pretreatment of suckling, weanling, or adult mice with 80 or 400 ng of TNF or IL-1 alone, or 80 ng of each cytokine together, had no effect on the survival of mice following MCMV. Similarly, neither of these cytokines adversely influenced the protective effects of hyperimmune anti-MCMV antiserum; that is, they did not attenuate the protection conferred by the antiserum nor affect the protective effects of subtherapeutic doses of the antiserum. We conclude that despite promising antiviral effects against MCMV in vitro, these agents do not result in a useful therapeutic effect in vivo. Moreover, despite the ability of IL-1 to induce ACTH and corticosterone in mice, IL-1 treatment did not increase the mortality to CMV.