Propionibacterium acnes meningitis in a previously normal adult.

A 25-year-old man was previously healthy until he contracted acute Propionibacterium acnes meningitis. Comparison with previous reports of de novo diphtheroid meningitis suggests that this entity can appear with features that are not characteristic of acute bacterial meningitis, including (1) stroke-like syndromes, (2) an afebrile course, and (3) a cerebrospinal fluid with a mononuclear pleocytosis and normal glucose level. The appropriate choice and dosage of antimicrobial agent must be guided by more than in vitro sensitivity data to prevent relapse and possible chronic meningitis. Although diphtheroids are as a rule exquisitely sensitive to penicillin, predictably high tissue levels of drug in diphtheroid meningitis are best achieved with chloramphenicol treatment. In the appropriate settling, the isolation of diphtheroids from cerebrospinal fluid should not be discounted as a "contaminant."

Myoglobinuria associated with herpes-group viral infections.

Two cases of myoglobinuria were associated with herpes-group viral infection (herpes simplex and Epstein-Barr). Muscle atrophy and acute renal failure were important complications. No evidence of direct invasion of muscle fibers by virus was found, nor was the severity of myoglobinuria correlated with histologic appearance of muscle.

Anaesthesia, surgery, obstetrics, and emergency care in Guyana.

The surgical and anaesthesia needs of low-income countries are mostly unknown due to the lack of data on surgical infrastructure and human resources. The goal of this study is to assess the surgical and anaesthesia capacity in Guyana. A survey tool adapted from the WHO Tool for Situational Analysis to Assess Emergency and Essential Surgical Care was used to survey nine regional and district hospitals within the Ministry of Health system in Guyana. In nine hospitals across Guyana, there were an average of 0.7 obstetricians/gynaecologists, 3.5 non-OB surgeons, and 1 anaesthesiologist per hospital. District and regional hospitals performed an annual total of 1520 and 10,340 surgical cases, respectively. All but 2 district hospitals reported the ability to perform surgery. An average hospital has two operating rooms; 6 out of 9 hospitals reported routine medication shortages, and 4 out of 9 hospitals reported routine water or electricity shortages. Amongst the three regional hospitals, 16.1% of pregnancies resulted in Caesarean section. Surgical capacity varies by hospital type, with district hospitals having the least surgical capacity and surgical volume. District level hospitals routinely do not perform surgery due to lack of basic infrastructure and human resources.

Brugia-like filarial infection acquired in the United States.

A mature male filarial worm was found in sections of an enlarged, painful retroauricular lymph node removed from an 18-year-old resident of New Jersey. On the basis of its morphology and location in an obstructed lymph vessel, the worm was identified as probably a species of Brugia, possibly Brugia beaveri, a parasite of the raccoom. The infection in this case resembled one reported earlier from the same general area of the country, New York City. In the absence of demonstrable microfilaremia in this and other cases of zoonotic filariasis acquired in the United States, specific drug treatment after surgical removal of the worm is usually unnecessary.

Yellow fever monoclonal antibodies: type-specific and cross-reactive determinants identified by immunofluorescence.

Monoclonal antibodies directed against the envelope glycoprotein and the NV3 non-structural viral protein of yellow fever (YF) were tested by the indirect fluorescent antibody technique against a variety of YF virus strains and heterologous flaviviruses. Monoclonal antibodies directed against the envelope glycoprotein exhibited YF strain-specificity, YF type-specificity, broad group cross-reactivity, or limited subgroup reactivity (YF + Banzi or YF + Koutango + Zika + Usutu + Uganda S). Monoclonal antibodies directed against NV3 reacted either with YF + Koutango or with YF + Banzi. These findings generally correlated with the results of biological tests reported previously. Monoclonal antibodies that were type-specific to YF will be useful for the rapid specific identification of YF virus isolates and are available from the Centers for Disease Control on request.

Homogeneity among Senegalese strains of yellow fever virus.

A series of 16 yellow fever (YF) viruses isolated from mosquitoes, monkeys and humans in different epidemiological contexts in Senegal and The Gambia between 1976 and 1983, was analyzed by T1 RNase oligonucleotide fingerprints of the genomic 32P-labeled RNA, by SDS-polyacrylamide gel electrophoresis of the intracellular virus-specified polypeptides, by peptide mapping of the envelope E glycoprotein and by immunological reactivities with monoclonal antibody fluids (MAF's) against the E glycoprotein. These strains had not been passed in suckling mice and were isolated in Aedes pseudoscutellaris Mos 61 cultured cells. These strains showed no virulence in three-week-old Swiss mice when injected intraperitoneally. Direct comparison of the large T1 RNase-resistant oligonucleotide maps indicated a relative genetic stability (92%-100%). A greater change was observed when these strains were compared with an epidemic YF strain isolated in 1965 with an oligonucleotide fingerprint map sharing 82%-88% similarity. The YF-specified proteins were identical in their molecular weight, and the fragments obtained after limited proteolysis of the envelope protein using protease V8 or alphachymotrypsine indicated that the strains were chemically similar. Only a few differences were observed when the strains were seroneutralized with MAF's, but no relation could be made with genetic or biological data. This suggested that the YF virus strains isolated from the same geographic area and during a short period of time had evolved slowly. Moreover, all the viruses were closely related and no correlation could be established with the apparent variations in virulence in nature.

Immunization of monkeys with baculovirus-dengue type-4 recombinants containing envelope and nonstructural proteins: evidence of priming and partial protection.

Groups of rhesus monkeys were immunized with baculovirus-dengue type-4 (DEN-4) recombinant-infected cell extracts. One recombinant contained all of the DEN-4 structural proteins and two nonstructural (NS) proteins (C-M-E-NS1-NS2a), while the other was a fusion protein containing a portion of the respiratory syncytial virus G glycoprotein and DEN-4 envelope glycoprotein (RSVG-E). Both preparations were immunogenic; all monkeys receiving either immunogen responded with the production of antivirion antibodies in enzyme immunoassays. All except one monkey receiving the recombinant b(C-M-E-NS1-NS2a) made antibodies to NS1. One monkey that received b(RSVG-E) showed the production of low levels of neutralizing antibodies. Following challenge with unmodified DEN-4 virus, seven of nine monkeys in the immunized group became infected and were viremic for a mean of 4.1 days. The control, sham-inoculated monkeys were also viremic; the mean number of days of viremia in this group was 4.7 days. The remaining monkeys in the immunized group (n = 7), although not protected, had evidence of priming. Hemagglutination inhibition antibody responses following challenge indicated an anamnestic response in this group of animals. Based on these results, it was concluded that future immunization schedules should be altered to optimize immune responses and that immunization with more potent and purified immunogens would probably result in higher seroconversion rates and antibody levels in monkeys.

Difference between the abilities of human Fcgamma receptor-expressing CV-1 cells to neutralize American and Asian genotypes of dengue virus 2.

Sera from patients involved in a Peruvian outbreak of dengue virus serotype 1 infection cross-neutralized the American genotype of dengue virus serotype 2 up to 100-fold more efficiently than they did the virulent Asian genotype of dengue virus serotype 2, as determined by a plaque reduction neutralization test (PRNT) with CV-1 fibroblasts modified to express human Fcgamma receptor CD32. The concordant preferential immune enhancement of the Asian genotype of dengue virus serotype 2 in human monocytes suggests that such a modification might strengthen the correlation between the PRNT titer and protection.

17D yellow fever virus infection of P388D1 cells mediated by monoclonal antibodies: properties of the macrophage Fc receptor.

Thirteen IgG monoclonal antibodies to the envelope protein of 17D yellow fever virus (17D YF) were produced. All of the antibodies, whether type-specific to 17D YF or flavivirus cross-reactive, mediated antibody-dependent enhancement (ADE) of virus growth in P388D1 cells. There was no consistent relationship between ADE titres and the degree or pattern of neutralizing and/or haemagglutination inhibition activity. Monoclonal antibodies of different isotypes were used to investigate further the properties of P388D1 Fc receptors. The effects of trypsin treatment of P388D1 on ADE were similar to those previously described in experiments measuring direct binding of IgG proteins or rosetting of sheep red blood cells (SRBC) by macrophages, demonstrating sensitivity to digestion by trypsin of the Fc receptor for monomeric IgG2a but not for IgG2b. Aggregated myeloma proteins of IgG2a and IgG2b isotypes competed equally well with either IgG2a or IgG2b monoclonal antibodies to 17D YF in inhibition of ADE. However, selective inhibition by the homologous isotype was observed when rosetting by P388D1 of SRBC coated with IgG2a or IgG2b monoclonal antibodies was examined. These results may help to explain apparent discrepancies previously reported between experiments utilizing direct binding of IgG proteins and those using rosetting of antibody-coated SRBC to examine Fc receptor properties and indicate that immune complexes of virus and antibody resemble aggregated immunoglobulins with respect to macrophage Fc receptor function and differ from antibody-coated SRBCs.

Antibody-mediated infection of P388D1 cells with 17D yellow fever virus: effects of chloroquine and cytochalasin B.

Antibody-mediated 17D yellow fever virus infection of a murine macrophage-like cell line, P388D1, was not inhibited by concentrations of cytochalasin B which did inhibit phagocytosis of antibody-coated sheep red blood cells. Chloroquine did not affect antibody-mediated enhancement of viral growth but inhibited viral replication equally whether infection was carried out in the presence or absence of antibody. The results suggest that the route of internalization of virus and subsequent intracellular pathways of replication are similar for both conditions of infection.

Evidence for transmission of lymphocyte responses to tuberculin by breast-feeding.

The possibility that cell-mediated immunity could be acquired by breast-feeding was evaluated in a prospective study of 26 tuberculin positive and 9 negative puerperal mothers and their infants. 13 infants of the positive and all the infants of negative mothers were breast-fed. Tuberculin-reactive T cells were found in colostrum and early milk of most positive but in none of the negative nursing mothers. A significant number (8/13) of infants born to positive mothers had tuberculin-reactive peripheral blood T cells after 4 weeks of breast-feeding compared with bottle-fed infants (1/13) of positive mothers or breast-fed infants (0/9) of negative mothers. Examination of cord blood for tuberculin-reactive T cells provided no significant evidence of transplacental transmission of responsiveness to tuberculin. The results suggest that breast-fed infants may passively acquire T cell responsiveness to a specific antigen by ingestion of breast milk.

Neutralizing F(ab')2 fragments of protective monoclonal antibodies to yellow fever virus (YF) envelope protein fail to protect mice against lethal YF encephalitis.

Monoclonal antibodies (MAbs) prepared against the yellow fever virus (YF) vaccine strain 17D (17D YF) envelope E protein were used to investigate Fc piece involvement in antibody-mediated protection against YF encephalitis in mice 17D YF passaged either in Vero cells or in mouse brain (P-YF) to increase neurovirulence was used. To avoid uncertainty concerning antibody clearance and blood-brain barrier penetration, and to directly compare protective activity with neutralization in vitro, pre-formed antibody-virus complexes were injected intracerebrally or assayed for plaque formation in parallel. F(ab')2 fragments of an IgG2a MAb that strongly neutralized both YF strains retained molar equivalent neutralizing activity in vitro, but did not protect. However, further incubation of such F(ab')2-virus antibody complexes with rabbit IgG, but not F(ab')2 anti-mouse IgG resulted in protection. To unambiguously test for Fc piece involvement in this model we derived an IgG2a isotype switch variant from a protective IgG1 MAb-secreting hybridoma and prepared F(ab')2 fragments of the derivative. Intact and fragmented antibodies exhibited weak neutralizing activity. The variant antibody failed to protect against P-YF, but against considerably less neurovirulent 17D YF its protective capacity was 10-fold higher than that of its IgG1 parent. F(ab')2 fragments of the variant did not protect. Together, these results provide strong evidence of an in vivo protective function for the anti-virion antibody Fc piece and indicate that in vitro neutralizing activity as a predictor of antibody protective capacity is dependent on Fc piece integrity and isotype.

Protection of mice against yellow fever virus encephalitis by immunization with a vaccinia virus recombinant encoding the yellow fever virus non-structural proteins, NS1, NS2a and NS2b.

Recent evidence of a protective immune response to the flavivirus non-structural protein, NS1, has suggested its incorporation into possible recombinant vaccines. The region of the 17D yellow fever virus (YFV) genome encoding the C terminus of envelope glycoprotein and extending to the N terminus of non-structural protein NS3 (NS1-NS2a-NS2b; nucleotides 2030 to 4940) was expressed in vaccinia virus and physical and immunogenic properties of the NS1 moiety were studied. Recombinant NS1 protein, and native YFV NS1, was detected at the surface of infected cells by immunofluorescence and by immune cytolysis after treatment with anti-NS1 antibody and complement. NS1 was also detected in the extracellular medium as a secreted form. Recombinant NS1 was immunoprecipitated as a single protein of approximately the same size as native 17D YFV NS1. Unboiled, both recombinant and native NS1 formed polymers of high Mr. Immunization of mice with this recombinant vaccinia virus stimulated production of non-neutralizing, complement-fixing cytolytic antibody and conferred partial protection against lethal intracerebral inoculation of mice with live 17D YFV.

Antibody-mediated infection of macrophages and macrophage-like cell lines with 17D-yellow fever virus.

The 17D vaccine strain of yellow fever virus (17D-YF) produces a safe human arboviral infection that can provide antisera of well-defined specificity under chronologically defined conditions. We studied 17D-YF growth in human peripheral blood macrophages and in two continuous Fc receptor-bearing, macrophage-like cell lines, P388D1 of mouse origin and U937 of human origin. Cells were infected with virus in the presence or absence of antibody to 17D-YF and to two related flaviviruses, St. Louis encephalitis (SLE) and dengue 2 (D2V). The virus 17D-YF grew in the three cell types when infection was established without antibody; viral yields were increased by addition of antibody to 17D-YF, SLE, and D2V. Increased titers of virus were accompanied by an increased number of infected cells by immunofluorescent assay. Enhancing activity was present in the IgG but not the IgM fractions of immune sera. Infection without cytopathic effect was observed in U937.

Growth of 17D yellow fever virus in a macrophage-like cell line, U937: role of Fc and viral receptors in antibody-mediated infection.

Growth characteristics of 17D yellow fever virus (17D-YF) and conditions for infection were studied in U937, a macrophage-like, Fc receptor-bearing continuous human cell line. Antibody to 17D-YF was obtained by immunization of normal subjects with 17D-YF vaccine. Cells were infected in the presence or absence of immune whole sera or immunoglobulin fractions. Infection of U937 was temperature dependent; the yield of virus was variable but at low temperature viral titers were consistently higher when infection was established in the presence of antibody. Results of infectious center assays indicated that the increased yield of virus was largely or entirely due to an increase in the number of cells producing virus early in the course of infection. Enhancement of viral growth was mediated by IgG but not IgM fractions of immune sera. Trypsinization of U937 resulted in a 90 to 95% reduction of infection in the absence of antibody but in the presence of antibody viral titers were higher in trypsinized than in nontrypsinized cells. Antibody to 17D-YF, contained in the whole IgG fraction of sera, bound to U937 to mediate infection without first being complexed to virus. Preincubation of U937 with IgG1 but not IgG2 myeloma proteins abrogated antibody-mediated infection. This result is compatible with the known affinities of U937 Fc receptors for specific subclasses of IgG and provides evidence for the role of the Fc receptor in antibody-mediated enhancement of viral growth. Persistent infection characterized by a lack of detectable cytopathogenic effect was established in long-term cultures of U937. This pattern of infection might be related to the unique effectiveness of the 17D-YF vaccine.

Influence of the human high-affinity IgG receptor FcgammaRI (CD64) on residual infectivity of neutralized dengue virus.

We examined dengue virus immune complex-phagocyte interaction with respect to a single Fc receptor class using a transient expression system involving the high-affinity human macrophage receptor, FcgammaRI. We found that New Guinea C strain dengue 2 virus formed well-defined plaques in normal and transfected COS cells and we analyzed the structural determinants of FcgammaRI-mediated binding and internalization of dengue 2 virus immune complexes by expressing native or truncated forms of the receptor in COS cells, alone or with its accessory gamma chain signaling unit, which bears an immunoreceptor tyrosine-based activation motif (ITAM). The residual infectivity of dengue 2 virus treated with neutralizing human antiserum was strikingly higher in FcgammaRI-bearing COS cells than in controls. Compatible with the IgG subclass specificity of FcgammaRI, this difference was abrogated quantitatively by treatment of FcgammaRI-transfected cells with human IgG1 but not IgG2 myeloma protein. The magnitude of receptor-mediated plaque formation after cotransfection with gamma chain was also significantly higher than in controls but was less than that observed with FcgammaRI transfection only, a difference probably explained by reduced levels of FcgammaRI expression in gamma chain cotransfectants. Deletion of the FcgammaRI cytoplasmic domain had no effect on receptor-mediated immune complex infectivity. We conclude that the FcgammaRI extracellular domain is sufficient for internalization of infectious dengue virus immune complexes through a mechanism that does not involve classical ITAM-dependent signaling.

Immunogenicity of a purified fragment of 17D yellow fever envelope protein.

Information on the immunogenic properties of purified flavivirus proteins may be useful in the development of recombinant or synthetic peptide vaccines. Using a monoclonal antibody, an attempt was made to purify the envelope (E) protein of 17D yellow fever virus (17D YF) by affinity chromatography. The purified material could not be identified as intact E protein but it did bear antigenic determinants of E as determined by selective reactivity with anti-E monoclonal antibodies. Rabbits immunized with this material produced antibodies that neutralized 17D YF and dengue-2 viruses in comparable titers, indicating that cross-reactive antigenic determinants were preserved. Immunization of mice resulted in protection against intracerebral challenge with 17D YF.