Immunization Against Swine Influenza in the Yale University Community

Nineteen percent of the approximately 30,000 members of the Yale community aged 18 through 59 received swine influenza monovalent vaccine (A/New Jersey/1976) during the three days of a mass immunization program in Nov. 1976. Based on 1508 card questionnaires received, 71.2 percent of the vaccine recipients experienced a sore arm, 23.4 percent headache, 13.4 percent chilliness, and 9.7 percent feverishness or fever. The sore arm was judged as severe in 5.9 percent as was the headache in 4.2 percent. Other reactions were regarded as severe in less than 2 percent. All reactions were reported more commonly by women than mean and all decreased with age.Serologic tests carried out at the start of the immunization period revealed that influenza A/New Jersey/1976 antibody was absent from 78.6 percent of the recipients; almost all persons under 25 lacked this antibody. A significant antibody rise occurred in 78.3 percent of those receiving a single dose of monovalent vaccine. Somewhat better antibody responses occurred in 36-59 year olds than in those age 17-25 (84.9 vs 75.5 percent); the geometric mean antibody titer was also much higher (1:136.8 vs 1:31.2). However, the presence of pre-existing homologous antibody did not significantly improve the antibody response to the vaccine. Cross-reacting antibody rises to A/Victoria/1975 were found in 16.2 percent of the recipients of monovalent vaccine.     

Cellular immunity in infectious mononucleosis. II. Specific reactivity to Epstein-Barr Virus antigens and correlation with clinical and hematologic parameters.

Infectious mononucleosis (IM) patients, Epstein-Barr virus (EBV)-seropositive and seronegative healthy donors, and patients with other viral infections were tested for lymphocyte blastogenesis (LB) with phytohemagglutinin and six EBV (virus concentrate, culture supernatant, and soluble [S] antigen) or control antigens. Fluorescent antibodies to EBV viral capsid antigen of IgG, IgM, IgA specificities, to nuclear antigen (EBNA), and heterophile antibodies were also assayed. These were correlated with clinical parameters (fever, pharyngitis, adenopathy, hepatitis, splenomegaly, atypical lymphocytes, and total mononuclear cell counts). EBV viral and S antigen-induced LB was significantly greater in seropositive donors. IM patients had antigenspecific LB below that of seropositive donors initially and low responses for the acute phase of illness when clinical symptoms were present and antibody titers were maximal. Specific LB rose to a peak at 3.5 to 9 weeks when the patients had recovered, most laboratory findings had returned to normal, and antibodies had declined. At peak, specific LB in IM patients exceeded that of seropositive donors, but later declined. These results demonstrate specific cell-mediated immunity (CMI) to EBV, and indicate that this develops slowly in IM and contrasts with the evolution of the clinical events and humoral immunity. This correlation supports the hypothesis that CMI is the mechanism of terminating lymphoproliferation in IM.     

Immunoblotting reactivity of human sera from various sources against purified epstein-barr virus

The immunoblotting technique was used to analyse polypeptides of purified Epstein-Barr virus reacting with antibodies present in sera from clinically healthy individuals, from patients with infectious mononucleosis (IM) or AIDS, and from renal transplant recipients with molecular sizes in the range of 40–290 kDa were detected.The 47- and 160-kDa nucleocapsid polypeptides, as well as the 72-, 74-, 140-, 220- and 290-kDa membrane polypeptides were the major viral proteins detected in the sera. Sera from clinically healthy individuals contained antibodies directed against all EBV membrane and nucleocapsid antigens. Sera from renal transplant recipients, from patients with IM and from patients with AIDS failed to react with certain nucleocapsid and membrane antigens; in particular, sera from AIDS patients and renal transplant recipients did not react with the 220-kDa polypeptide, one of the major membrane antigens, while sera from subjects with IM and from healthy individuals did.A high proportion of sera from patients with IM (38% vs 5% of clinically healthy individuals and 0–5% of the AIDS patients and renal transplant recipients) reacted with a 42-kDa polypeptide, suggesting its possible role in acute EBV infection.     

Systemic and mucosal immune responses to sublingual or intramuscular human papilloma virus antigens in healthy female volunteers

The sublingual route has been proposed as a needle-free option to induce systemic and mucosal immune protection against viral infections. In a translational study of systemic and mucosal humoral immune responses to sublingual or systemically administered viral antigens, eighteen healthy female volunteers aged 19-31 years received three immunizations with a quadravalent Human Papilloma Virus vaccine at 0, 4 and 16 weeks as sublingual drops (SL, n = 12) or intramuscular injection (IM, n = 6). IM antigen delivery induced or boosted HPV-specific serum IgG and pseudovirus-neutralizing antibodies, HPV-specific cervical and vaginal IgG, and elicited circulating IgG and IgA antibody secreting cells. SL antigens induced ~38-fold lower serum and ~2-fold lower cervical/vaginal IgG than IM delivery, and induced or boosted serum virus neutralizing antibody in only 3/12 subjects. Neither route reproducibly induced HPV-specific mucosal IgA. Alternative delivery systems and adjuvants will be required to enhance and evaluate immune responses following sublingual immunization in humans. TRIAL REGISTRATION: ClinicalTrials.govNCT00949572.     

Characterization of IM virus, which is frequently isolated from cerebrospinal fluid of patients with multiple sclerosis and other chronic diseases of the central nervous system.

A transmissible agent, the IM virus, antigenically related to the Japanese subacute myelo-optico-neuropathy virus, has been isolated from several human cerebrospinal fluids obtained from American patients with multiple sclerosis and other chronic diseases of the central nervous system. The isolates were propagated in human diploid fibroblast (MRC5) cells, and virus was released into the culture medium in the absence of overt cytolysis. Infection of MRC5 cells resulted in a subtle alteration in the normal growth pattern of the cells. In unstained cultures, the cell changes were so mild that it was necessary to carry out all virus assays under code to eliminate bias. Cells in late passages were more susceptible than vigorously growing cells in early passages. Analysis of the kinetics of replication revealed that newly synthesized progeny virus was first detected about 12 h postinfection, that maximal virus release occurred by 48 h postinfection, and that virus production was persistent throughout an 8-day period. Several inhibitors of DNA synthesis were effective in blocking viral replication, including cytosine arabinoside, iododeoxyuridine, and phosphonoacetic acid. A substantial decrease in infectivity was observed upon treatment of IM virus with ether, suggesting that a lipid-containing structure is essential for infectivity. Ultrafiltration studies approximated the size (diameter) of IM virus to be between 100 and 200 nm.     

Immune response in subacute sclerosing panencephalitis: reduced antibody response to the matrix protein of measles virus.

Immune precipitation was used to study the humoral immune response of patients with subacute sclerosing panencephalitis (SSPE). Patients with SSPE have a progressive infection of the CNS by measles or a measles variant despite high serum antibody levels to measles virus as measured by standard serologic techniques. However, when the antibody response to individual measles virus proteins was measured, we found a striking reduction in the ability of sera from patients with SSPE to precipitate the matrix (M) protein as compared to the precipitation of the M protein by sera from normal adults who had natural measles infection in childhood, or by convalescent sera obtained 3 to 5 weeks after a naturally occurring measles infection. The decreased antibody response to the M protein in sera from patients with SSPE occurred despite a vigorous antibody response to the other viral proteins, suggesting a selective defect in the production of antibody to a single viral protein. The reduced anti-M antibody in sera from patients with SSPE was demonstrated whether immune precipitation was performed with wild-type measles virus or SSPE virus proteins. These results suggest that in SSPE only small amounts of the M protein are produced. This result may help explain how measles virus persists in the central nervous system of patients with SSPE.     

Bovine Respiratory Syncytial Virus

A large epizootic of an acute respiratory disease of cattle occurred in Japan during the months from October 1968 to May 1969. A virus was recovered in primary cultures of calf kidney and testicle cells from nasal swabs of affected cattle. Neutralization tests revealed the virus to be closely related to the Long strain of human respiratory syncytial virus. The virus induced cytopathic changes including the formation of syncytia and acidophilic-cytoplasmic inclusions in calf kidney and testicle cell cultures. A calf inoculated with the virus by the respiratory route developed an illness resembling the natural disease. Most cattle clinically diagnosed as having the disease showed significant rises of neutralizing antibody titer for the isolated virus, whereas none or only small fractions of those animals showed serological evidence for recent infection with bovine ephemeral fever virus, infectious bovine rhinotracheitis virus, Ibaraki virus, bovine diarrhea virus, bovine adenovirus Type 7 and parainfluenza virus Type 3. Neutralization tests on paired sera revealed a wide dissemination of the isolated virus among cattle in many areas of the country during the epizootic. All these findings leave no doubt that the epizootic was caused by bovine respiratory syncytial virus. This is the first study that ever shows the presence of infection of cattle with this virus in Japan.     

Quasi-immune response of Penaeus japonicus to penaeid rod-shaped DNA virus (PRDV)

A quasi-immune response was demonstrated in kuruma prawn Penaeus japonicus infected naturally or experimentally with PRDV (penaeid rod-shaped DNA virus, also called white spot syndrome virus or WSSV), the causative agent of PAV (penaeid acute viremia). In the first step of this study, natural survivors 4 mo after a PAV outbreak demonstrated 94 % relative percent survival (RPS) upon experimental PRDV challenge. Mortalities after challenge were confirmed by PRDV detection to be due to PAV using a PCR method. In the second step, experimental PAV survivors were produced by intramuscular (IM) injection of PRDV into naive shrimp subsequently reared collectively in a tank (A group) or individually in chamber units (B group). Survival was 41 and 90% in the A and B groups, respectively. A subsequent IM re-challenge of these PRDV survivor groups with PRDV made 32 d after the first challenge revealed a protective response with high RPS of 77 and 64%, respectively. These high survival rates suggested that PAV survivors (natural or experimental) were able to resist PRDV infection and that the resistance was not due to selection of naturally resistant shrimp during a PAV outbreak, but due to enhancement of an immune-like system (quasi-immune response) after exposure to PRDV. No PRDV neutralizing activity was revealed in the serum of the 4 mo natural survivors of the PRDV outbreak. However, it was found in their serum 17 d after they had been experimentally challenged with PRDV.     

Microprecipitation test for the detection of adenovirus antibody

A microprecipitation test (MPT) for the detection of adenovirus antibody has been developed. The new procedure combines precipitation of virus particles with specific antibody, separation of unreacted components from the resulting electroneutral virus-antibody complexes by electrophoresis, and detection of these complexes with a protein stain. Type-specific antibody was detected in rabbit antisera but, under similar conditions, antibody in convalescent human sera reacted with adnovirus antigens of types 4 and 7. Paired sera from 57 patients with suspected adenovirus infection were examined for significant rises in antibody activity by the microprecipitation test and by complement fixation.     

Immune response to a hepatitis B DNA vaccine in Aotus monkeys: a comparison of vaccine formulation, route, and method of administration.

BACKGROUND: Attempts to optimize DNA vaccines in mice include using different routes of administration and different formulations. It may be more relevant to human use to carry such studies out in nonhuman primates. Here we compare different approaches to delivery of a DNA vaccine against the hepatitis B virus (HBV) in Aotus monkeys. MATERIALS AND METHODS: Thirty-two adult Aotus l. lemurinus monkeys divided into 8 groups of four were immunized with 400 microg of a DNA vaccine which encoded hepatitis B surface antigen (HBsAg). DNA in saline was administered by intradermal (ID) or intramuscular (IM) injection with needle and syringe, IM injection with the Biojector needleless injection system or combined ID (needle) and IM (Biojector). DNA formulated with cationic liposomes (CellFECTIN) was injected IM with needle or Biojector. DNA with added E. coli DNA (100 microg) was injected IM with the Biojector or ID. A ninth group of 4 monkeys was injected IM (needle) with Engerix-B, a commercial vaccine containing recombinant HBsAg (10 microg) adsorbed onto alum. Monkeys were boosted in an identical fashion to their prime at 8 weeks, but all received the protein vaccine (Engerix-B) at 16 weeks. Sera was assessed for antibodies against HBsAg (anti-HBs) by enzyme-linked imunosorbent assay (ELISA). RESULTS: The primary humoral response induced by IM delivery of the DNA vaccine was very poor. In most cases there was no detectable anti-HBs even after 2 DNA doses but the kinetics of the response to subsequent protein indicated that a memory B cell response had been induced. In contrast, following IM-administration of DNA using the Biojector, detectable anti-HBs were observed in 3 of 8 animals and evidence for immunological priming was apparent in an additional 4 of the 8 monkeys. ID injection of DNA vaccine in saline induced a potent antibody response which was augmented 6-fold by the addition of E. coli DNA. Combining ID and IM administration did not improve humoral immunity over ID injection alone. CONCLUSIONS: For immunization of primates with DNA vaccines, ID may be a preferable route to IM, although it is not clear whether the Aotus monkey is a relevant model for humans in this respect. Nevertheless, the use of the Biojector needleless injection system may improve responses with IM delivery of DNA vaccines. As well, the immunostimulatory action of E. coli DNA may be used to augment the humoral response induced by a DNA vaccine.     

Acquisition of serum antibodies to specific viral glycoproteins of parainfluenza virus 3 in children.

A radioimmunoprecipitation assay was used to study antibody responses to parainfluenza virus 3 glycoproteins in human sera. The method was not only more sensitive than the neutralization test for the detection of antibody but also provided semiquantitative assessments of the antibody response to both glycoproteins in a single assay system. Anti-hemagglutinin-neuraminidase titers were consistently higher than anti-fusion levels in the same serum specimen. Thirteen children were monitored serologically and virologically from birth until 12 months or more after their primary infection with parainfluenza virus 3. At 1 to 3 months after infection, a significant increase in the level of antibody to the hemagglutinin-neuraminidase protein developed in 12 children; of these, 9 showed rises in the level of fusion protein. In 11 of the children, antibody titers continued to rise and the geometric mean titers to the hemagglutinin-neuraminidase protein was highest in sera collected 8 to 10 months after primary infection. Reinfection as the reason for these progressive increases in antibody levels could only be confirmed for four of the children. Three other children had reinfections after the 10-month sera were obtained; in each instance the only antibody responses were to the fusion protein.     

Viral antibodies in infectious mononucleosis

Abstract Patients with Epstein-Barr virus (EBV) infectious mononucleosis (IM) usually develop heterophilic antibodies and some autoantibodies. Antibodies to rubella, measles, adeno-, entero-, herpes simplex, cytomegalo- and varicella-zoster viruses were titrated in sera from IM patients and matched healthy controls using the complement fixation test (CFT) and the haemagglutination inhibition test. Except for herpes simplex virus and cytomegalovirus, the IM sera had significantly higher arithmetical and geometrical mean antibody titres and showed in most cases higher antibody prevalences in the CFT. The titre rise was most pronounced for rubella and measles antibodies, between 2- and 3-fold. There were no cases of very high titres occasionally seen in IM. The IM sera had higher total IgG serum levels than the controls, 17.27 g/1 and 11.8 g/1, respectively ( P < 0.001). The present data show that in addition to previously reported high levels of some autoantibodies and of heterophilic antibodies, there is a more general increase in IgG antibodies to commonly occurring viruses. This increase is most likely due to the polyclonal activation of B-lymphocytes following the binding of EBV to the complement receptor CR2 (CD21). When due consideration is given to the possible occasional occurrence of a false positive rubella IgM test, the raised antibody-titres will most likely not interfere with routine diagnostics.     

Impaired serum antibody response to inactivated influenza A and B vaccine in cancer patients.

The serum antibody response to vaccination with bivalent inactivated influenza vaccine containing A/Port Chalmers/1/73 (H3N2) and B/Hong Kong/5/72 antigens was assessed in 44 patients with cancer and in 27 healthy control subjects. A fourfold or greater increase in antibody titre after vaccination occurred in 16 of the 44 cancer patients and 25 of the 27 controls for the A antigen, and in 14 of the 44 cancer patients and 20 of the 27 controls for the B antigen. Patients with lymphoma, who tended to have hypogammaglobulinemia, responded less well than did patients with solid tumours. Among the latter the failure to show a fourfold or greater increase in antibody titre correlated with a poorer 18-month survival.     

Lyme disease in Minnesota: epidemiologic and serologic findings

During the four years, 1980 to 1983, 83 Minnesota residents have been diagnosed with Lyme disease. Sixty-five of the patients were male. The median age of patients was 39 years with a range from one to 77 years. Seventy-five (90 percent) had onset in 1982 and 1983. Of these latter cases, 56 (75 percent) recalled a tick bite three to 27 days prior to the development of erythema chronicum migrans. Patients experienced possible exposure to Ixodes dammini in at least 24 (28 percent) of the 87 Minnesota counties; however, over 50 percent had reported exposure in one of eight east-central counties near or immediately west of the Wisconsin border. Serologic studies for antibody against the Ixodes dammini spirochete were completed on 30 patients with onset in 1982 and 1983. Of 28 patients with paired acute and convalescent serum samples, only two (7 percent) had fourfold rises in antibody titers. Lyme disease is an emerging public health problem in Minnesota. Additional studies are needed to define the risk of disease by geographic area within the state. Physicians statewide should be alert to the possibility of Lyme disease among their patients, since only 39 percent of patients with onset in 1982 and 1983 were exposed in their county of residence.     

Anti-pre-S2 antibody response in subjects vaccinated against hepatitis B and in naturally immunized subjects

Serum samples from individuals immunized with a pepsinized or non-pepsinized vaccine and from patients who had recovered from acute hepatitis B or who developed a chronic form of the disease, were analysed for the presence of antibody against the pre-S2 epitope of the hepatitis B virus.Anti-pre-S2 antibody was absent in all but one individual immunized with the pepsinized vaccine. Thirty-eight percent of the subjects who responded by anti-HBs production to the non-pepsinized preparation showed anti-pre-S2 antibody one year after complete vaccination. Among subjects who did not produce anti-HBs after immunization with this vaccine, 1 single individual produced anti-pre-S2 antibody. Anti-preS2 antibody was detectable after one year in 38% of the patients who recovered from acute hepatitis B, but in none of those with chronic hepatitis B. The kinetics of anti-pre-S2 antibody response to a booster injection was also analysed 1 month and 1 year after the 3rd injection and 1 month after the 4th injection of the non-pepsinized vaccine.     

Demonstration of EBNA (Epstein-Barr virus nuclear antigen) antibodies of different immunoglobulin classes.

Anit-EBNA IgM, a previously unknown antibody, was detected by the antihuman globulin anticomplement immunofluorescence (ACIF) method in serum samples from acute infectious mononucleosis (IM) of Epstein-Barr virus (EBV) origin. The antibody disappears from the serum in some weeks during convalescence. It was absent in anti-EBV=positive sera of healthy donors and in serum samples taken from patients with IM caused by cytomegalo-virus. The antibody appears simultaneously with anti-EBV IgmM and, reaching a lower titre than the latter, its titre curve runs parallel with the anti-EBV IgM curve. Since in acute EBV infections, anti-EBNA IgM always appeared, its presence may serve as an additional evidence of the acuteness of EBV infection. In EBV-seropositive healthy subjects, the bulk of antibodies belongs to the IgG class, non-complement-fixing IgA antibodies occur only sporadically.     

Fine specificity of the in vitro antibody response to influenza virus by human blood lymphocytes

The fine specificity of anti-influenza antibody produced in vitro by human PBM stimulated with different strains of influenza virus was examined by competition binding in solid phase enzyme immunoassay. Most of the antibody produced in vitro is directed to strain-specific or cross-reactive determinants on the hemagglutinin molecule. The extent of cross-reactivity is dependent on the strain of virus used to stimulate PBM as well as the individual tested and presumably on his previous exposure to influenza viruses. PBM from some individuals produced antibody that bound to the stimulating strain of influenza virus but not to other strains of the same subtype. In other individuals, antibody was produced in vitro that cross-reacted with all viruses in the same subtype (e.g., H3N2; A/X31, A/X47, and A/Texas) but did not bind to other (H2N1 or H1N1) subtypes, and in a few individuals, extensive cross-reaction between subtypes was seen. The presence of antibody to hemagglutinin in these culture supernatants was confirmed by competition binding to highly purified hemagglutinin. This in vitro culture system allows the immunologic memory of individuals to a wide range of stimulating virus strains to be examined simultaneously in terms of specificity of the antibody response by human PBM to influenza virus after natural infection or immunization.     

Clioquinol targets zinc to lysosomes in human cancer cells

A panel of 52 murine monoclonal antibodies was found to recognize antigenic determinants that had been conserved among all major genetic subgroups of the H5N1 avian influenza virus prevalent since 1997. We screened a phage display library for peptides recognized by one such antibody (8H5). We analysed the specificity of 8H5 for reactive peptides presented as fusion proteins of HBc (hepatitis B core protein) and HEV (hepatitis E virus) structural protein, p239. This was then related to the specificity of the native HA (haemagglutinin) molecule by virtue of the capacity of fusion proteins to compete for 8H5 binding with different strains of H5N1 virus and the reactivity of antisera generated against fusion proteins to bind native HA molecules, and to inhibit haemagglutination and arrest infection by the virus. Nine reactive peptides of different amino acid sequences were identified, six of which were also reactive with the antibody in association with HBc and four were in association with p239. Binding occurred with the dimeric form of the four p239-fusion proteins and one of the HBc-fusion proteins, but not with the monomeric form. The HBc-fusion proteins blocked 8H5 binding with four strains of H5N1 influenza virus. Mouse antisera generated against fusion proteins bound to HA molecules, but did not inhibit haemagglutination or arrest H5N1 infection. Our findings indicate that 8H5 recognizes discontinuous sites presented by secondary and possibly higher structural orders of the peptides in spatially favourable positions for binding with the antibody, and that the peptides partially mimic the native 8H5 epitopes on the H5N1 virus.     

Viral Infections of Toronto Children During 1965: I. Enteroviral Disease

Enteroviruses were isolated from feces and/or cerebrospinal fluid of 29 of 43 Toronto children who contracted aseptic meningitis, pleurodynia, abdominal pain or febrile upsets between June and October, 1965. Coxsackie A9 virus was the dominant agent in aseptic meningitis and Coxsackie B1 virus in pleurodynia and other syndromes. Sero-logical evidence of recent Coxsackie B1 and Echo 6 infection was obtained in two additional patients with aseptic meningitis who did not yield virus, and elevated Coxsackie B1 antibody titres were found in one patient with pericarditis. A newborn infant died with myocarditis due to Coxsackie B1 virus following infection of the mother during the immediate antenatal period. Paired sera collected only two to four days apart from patients with enteroviral syndromes or mumps meningoencephalitis frequently showed four-fold or greater increases of antibody levels.