Rubella reimmunization: comparative analysis of the immunoglobulin G response to rubella virus vaccine in previously seronegative and seropositive individuals.

Rubella virus (RV)-specific immunoglobulin G (IgG) antibodies were studied in military recruits undergoing unselected immunization with live attenuated measles, mumps, and rubella virus (MMR) vaccine. Three different whole-RV enzyme immunoassays (EIAs) and an epitope-specific EIA with a synthetic peptide (BCH-178c) representing a heutralization domain on the RV E1 envelope protein were used. Before vaccination, 84.2, 87.7, and 84.5% of the subjects tested (n = 399) were found to be seropositive (> 10 IU/ml or assay equivalent) by the three whole-RV EIAs, respectively, while only 82.5% were seropositive by the BCH-178c EIA. Although prevaccination seropositivity rates were similar for the whole-RV EIAs (sensitivity, 94 to 100%), many sera considered seropositive by the whole-RV EIAs had E1 peptide EIA antibody levels of < 10 IU/ml (sensitivity, 77.4 to 80.7%). One month after vaccination, 97.8, 97.2, and 93.5% of the subjects who were followed (n = 356) were seropositive by the three whole-RV EIAs, respectively, while 89% had BCH-178c peptide-specific IgG titers of > 10 IU/ml. After vaccination, depending on the assay used, up to 20.6% of initially seropositive individuals exhibited a greater than fourfold increase in RV-specific IgG, while up to 47.3% showed a greater than twofold increase. Increased antibody titers after vaccination (seroboosting) were most frequently associated with low levels of BCH-178c peptide-specific IgG before vaccination. RV protein-specific IgG was also studied by immunoblot assays in a subset (n = 56) of individuals receiving the MMR vaccine. Of these, 89.4 and 91.1% exhibited RV protein (E1, E2, and C protein)-specific IgG before and after vaccination, respectively. Seroboosting (two- to fourfold increase in EIA titers of individuals seropositive by the whole-RV EIA before vaccination) was usually accompanied by a shift in the IgG immunoblot pattern from a single (E1) to multiple (E1-E1, E1-C, or E1-E2-C) specificities, suggesting exposure of new epitopes as a result of viral replication.

Replication of measles virus in human monocytes and T cells.

Replication of measles virus (MV) in populations of peripheral blood mononuclear cells enriched for T cells and monocytes was studied using a temperature-sensitive mutant, MV ts38, and the parent counterpart, MV Lec. Stimulation of the cells was required for a full cycle of virus replication in both cell types. More infectious virus was released after stimulation from MV-infected populations enriched for T cells, T cell-enriched than from monocyte-enriched populations. However, similar amounts of viral mRNA, genomic RNA, and viral proteins of the expected size were found in both cell populations. The results indicate that MV-specific macromolecular synthesis is similar in both T cells and monocytes, but the assembly and (or) release of infectious virus is greatly reduced in monocytes as compared with T cells.

Seroprevalence of measles- and rubella-specific antibodies among military recruits, Canada, 1991.

A study of the seroprevalence of measles- and rubella-specific antibodies among military recruits in Canada in 1991 was undertaken to: 1) determine the proportion of military recruits who are measles and/or rubella seropositive when they enter the military; 2) detect general problems in the immune coverage in the young adult population; and 3) determine the proportion of measles seronegativity attributable to non-response, waning immunity or lack of exposure to either the disease or the vaccine. One initial blood sample was collected from all 399 recruits enrolled in basic training during the month of January 1991, prior to immunization with measles-mumps-rubella vaccine (MMR). Another sample was obtained from 354 of these recruits 3 to 5 weeks following this immunization. Only 18 (4.5%) recruits had negative measles-specific neutralization on the first sample. Only 12 (3.0%) recruits had negative measles-specific EIA on the first sample. All recruits had neutralization titres 40 or higher on the second sample. A total of 43 (10.8%) individuals had negative results for rubella EIA before immunization, 35 of which (81.4%) tested positive on the second sample.

Measles vaccine immunogenicity in 6- versus 15-month-old infants born to mothers in the measles vaccine era.

HYPOTHESIS: The low titer of measles antibody in infants of mothers with vaccine-induced immunity may allow immunization against measles before 15 months of age. METHODS: Six- and 15-month-old infants born to mothers < or = 30 years of age with no history of measles were recruited. Infants enrolled at 6 months of age were immunized with monovalent measles vaccine (Attenuvax), and maternal serum and infant pre- and postvaccination sera were obtained. Those enrolled for primary vaccination at 15 months of age received either Attenuvax (N = 12) or M-M-RII (N = 3). Six-month-old infants were revaccinated with M-M-RII at 15 months of age; pre- and postrevaccination sera were again obtained. Three antibody assays were used: a measles neutralizing assay (NT) and two enzyme immunoassays (EIA) for measles IgG and measles IgM. RESULTS: Among primary vaccinees, 14 of 19 infants aged 6 months (74%) developed NT antibody, as did 15 of 15 infants aged 15 months (100%). The reciprocal geometric mean titer of 6-month-old seroresponders was 23.3, significantly lower than that of the 15-month-old primary vaccinees (87.7, P < .001). Primary seroconversion rates by EIA were 53% for 6-month-old infants and 100% for those aged 15 months. Revaccination of infants who had received Attenuvax at 6 months of age resulted in 100% NT positivity; the geometric mean titer rose to equal that of the group given primary immunization at 15 months of age. Measles IgM antibody was detected in 10 of 14 infants tested 1 month after primary vaccination at 15 months, but was not detected in any of the revaccinated infants after the second dose at 15 months of age (P < .001). CONCLUSIONS: 1) Immunization with measles vaccine in infants born to vaccine-immune mothers at 6 months of age induced NT antibody in 74% of infants. 2) Revaccination of prior 6-month-old vaccinees at 15 months resulted in antibody titers equivalent to 15-month-old vaccinees. 3) Lack of an IgM response following revaccination suggests that even seronegative infants may be primed to respond on re-exposure to measles.

Major outbreak of pertussis in northern Alberta, Canada: analysis of discrepant direct fluorescent-antibody and culture results by using polymerase chain reaction methodology.

A major outbreak of 5,683 cases of pertussis occurred in northern Alberta, Canada, from December 1989 to January 1991. The outbreak highlighted a number of problems with current methods of pertussis diagnosis. In particular, an exceptionally high proportion of direct fluorescent-antibody (DFA)-positive, culture-negative specimens (88.4%) was identified. We took this opportunity to use polymerase chain reaction (PCR) methodology to examine whether the low culture rates were due to specimens containing dead organisms or whether the DFA results represented high numbers of false-positive results. A set of primer sequences within a Bordetella pertussis-specific repetitive element was used to amplify proteinase K extracts of B. pertussis DNA recovered from 279 submitted slides inoculated at the point of collection with nasopharyngeal material obtained from pernasal swabs. The PCR data corroborated the culture results: 84.6% of DFA-positive, culture-negative specimens were similarly PCR negative. At least three different bacterial species that were significantly cross-reactive with the commercial DFA reagent were identified in clinical specimens and in pure culture, providing one possible explanation for the false-positive DFA results. These results and other limitations of current diagnostic techniques underline the urgent need for a new DFA reagent with improved specificity and a standardized means of measuring the patient antibody response for the diagnosis of pertussis.

Reduced measles immunity in infants in a well-vaccinated population.

The recommended age for measles vaccination is based in part on information gathered when most mothers had natural measles. Nowadays many mothers have received measles vaccine. To assess this change measles antibody neutralization titers (NT) were determined for 278 mother-infant pairs. One hundred sixty-four mothers, born before 1958, likely had had natural measles (Group 1). Sixty mothers received one to three killed plus one attenuated measles vaccination (Group 2) and 54 received 1 attenuated measles vaccination only (Group 3). NT were determined for the mother and for the infant at birth and in the infant during the fourth and sixth months. Group 1 mothers and infants at every age had higher geometric mean NT than those in Groups 2 or 3 (P less than 0.05). By 7 months 65% of Group 1 infants and greater than 90% of Group 2 and 3 infants had an NT less than 1:10. The rate of antibody decay was significantly faster for Group 1 infants (P less than 0.05). Earlier vaccination in the infant should be considered.

Measles virus specific antibody in infants in a highly vaccinated society.

Measles virus specific antibody levels were measured in infants from 2 to 12 months of age. The sera were tested by hemagglutination-inhibition (HI), neutralization (NT), and enzyme immunoassay (EIA) methods. The results of this study indicate that in the population examined, infants at an early age have very low or no immunity of maternal origin to measles virus-93% of the infants were without detectable neutralizing antibody (NT titer less than or equal to 10) at 6 months of age, and by the end of the first year of life 100% had no neutralizing antibody.

Antibodies to measles virus surface polypeptides in an immunized student population before and after booster vaccination.

Serum specimens collected from 82 students before and after booster immunization with live measles virus (MV) vaccine were tested for MV surface protein-specific antibodies using a previously developed competitive enzyme immunoassay (EIA). The specificity of the assay in measuring antibodies against three sites on the haemagglutinin (H) and two on the fusion (F) protein was demonstrated. All subjects in this study were vaccinated, as one to two year-olds, three times with inactivated killed vaccine and later once or more with live vaccine. Fifty-three students were administered only live measles virus vaccine (M), whereas 29 were vaccinated with the MV combined with mumps and rubella (MMR). There was a clear tendency to a lower increase in antibody titres when MMR vaccine was used. This difference between the groups was most pronounced in antibodies against the site defined by a monoclonal antibody (mAb) I29. Antibodies to the site defined by the mAb 16AG5 on the F protein had no correlation with any of the other tests which suggests that subjects originally vaccinated with killed vaccine may have developed a distinct response to this site.

Isolation and partial characterization of a human adenovirus type 4 temperature-sensitive mutant (Mastadenovirus h 4 tsl).

A random selection procedure was used to isolate a temperature-sensitive (ts) mutant of human adenovirus type 4 (Mastadenovirus h 4 tsl) from nitrous acid mutagenized virus stock. The mutant displayed restricted growth at the nonpermissive temperature of 39 degrees C. Analysis of the mutant grown at 39 degrees C, by two-dimensional immunoelectrophoretic analysis, showed the mutant to be defective in the expression of the penton base and fibre structural components. The mutant was, however, capable of synthesizing immunologically reactive hexon components. Temperature-shift experiments revealed detectable fibre and penton to be present following shift-down from 39 to 32 degrees C. Time-sequence analysis of shift-down experiments suggested a possible defect in processing of the components, as indicated by an increase of immunologically detectable penton base. The ability of the mutant to assemble viruslike particles at 39 degrees C was confirmed by electron microscopy. Though the particles assembled appeared as mature virions, crystalline arrays of packed particles were less in number and somewhat smaller in size than those observed at 32 degrees C.

Studies on the multiplication of a porcine parvovirus.

A porcine parvovirus has been characterized with regard to its replication in foetal porcine kidney cells and certain biophysical properties. Electron microscopy of infected cells at selected times postinfection revealed that porcine parvovirus replication took place within or near a series of granular intranuclear inclusions which may be contiguous with cellular heterochromatin. Developing virions were observed to aggregate into a nucleolar-like amorphous mass which gradually disrupted as cellular integrity was lost. Purified virions were found to have a buoyant density in CsCl of 1.38 g/ml, while 'empty' particles had a buoyant density of 1.29 g/ml. The particle diameter was calculated to be approximately 22 nm.

An interference phenomenon associated with a measles virus SSPE isolate (Halle).

A measles virus (Halle SSPE isolate) induced ring plaque phenomenon (concentric rings of living and dead cells) has been shown to be associated with the temperature-dependent production of interfering particles. The interfering particles have been purified by potassium tartrate linear gradient centrifugation and have buoyant densities of 1.15 g/ml (M particle) and 1.06 g/ml (L particle) respectively. Both interfering particle population have been shown to decrease the yield of wild-type measles virus from infected cells by 50% when co-infection experiments were performed. Neither the M or L particle population interfered with the growth of VSV in the same host-cell system.

Purification and partial characterization of a 33 000 molecular weight endonuclease associated with human adenovirus type 5.

An endonuclease activity has been purified from human adenovirus type 5 (HAd5) virions and HAd5-infected cell extracts. The endonuclease activity is associated with a monomeric protein of molecular weight approximately 33 000. The endonuclease activity is more active at pH 4.5 than pH 7.2. Incubation of the enzyme at room temperature for periods of longer than 96 h results in a substantial increase in activity. The endonuclease activity is sensitive to EDTA at concentrations 10mM or greater but is insensitive to 500 mM NaCl. Immunological analysis with endonuclease-specific antiserum indicates that the endonuclease may be a host cell derived, viral modified, and incorporated protein.

Comparative polypeptide patterns of representative human and nonhuman adenoviruses.

SDS-polyacrylamide slab gel analysis of selected primate and nonprimate adenoviruses has shown these viruses to differ significantly at the polypeptide molecular level. It follows that data derived from study of a single adenovirus serotype are unique to that serotype and are not representative of the adenovirus group as a whole.

Characterization of the protein and nucleic acid of Aleutian disease virus.

Aleutian disease virus (ADV) was extracted and purified from infected mink. Nucleic acid extracted from the virus was examined in an electron microscope. Three different sizes of molecule, with approximate lengths of 1.2, 0.55, and 0.25 micron, were observed. The ratios of the large molecules to the small molecules were similar in all the particles prepared under different conditions. Equilibrium CsCl density gradient centrifugation showed that ADV nucleic acid had a buoyant density of 1.733 g/cm3. In Cs2SO4, ADV had a lower buoyant density than that of double-stranded RNA. These properties and its sensitivity to DNase suggested that ADV contains DNA. Thermal denaturation curves revealed that the DNA of ADV had a single-stranded configuration. Polypeptide analysis of ADV by polyacrylamide gel electrophoresis revealed the presence of four polypepties, with molecular weights of 30,000, 27,000, 20,500, and 14,000. These polypeptides were present in a ratio of 10:3:10:1, respectively. The data suggested that ADV is closely related to the members of the parvovirus groups.

Electron-microscopic study of the development of an equine adenovirus in cultured fetal equine kidney cells.

Sequential changes induced by an equine adenovirus in cultured fetal equine kidney cells were studied by electron microscopy. The first morphological change was the appearance of type I inclusions. These inclusions developed to type II inclusions which appeared as ring forms. Type III inclusions were formed within the central part of type II inclusions and finally filled up most of the nuclear space. As the infection proceeded, type IV inclusions which appeared as dense dark-staining spheres were formed at the center of the type III inclusions and also inside the cytoplasm. These dark-staining spheres developed and their center was filed with light-staining material and virus particles which eventually resulted in the formation of type V inclusions. Autoradiography study showed that types I, II, and III were composed of nucleoprotein and type IV was composed of protein.

Crystallization and reaggregation of adenovirus type 5 structural components from infected cell extracts.

Treatment of adenovirus type 5-infected cells at late times after infection (48 to 72 h p.i.) by hypotonic buffer and subsequent manipulation of the crude extract has resulted in the crystallization and reaggregation of the adenovirus type 5 fibre and hexon components.

An image analysis study of adenovirus type 5-induced crystalline inclusions.

A combined electron microscopic and optical diffractometric study of Ad5-induced non-virion protein crystals found in the nuclei of infected KB cells at late times (48 to 72 h p.i.) after infection has been carried out. Data obtained have indicated that the unit cell of the crystal is rectangular and not hexagonal as previously presumed.

Association of endonuclease activity with serotypes belonging to the three subgroups of human adenoviruses.

Endonuclease activity has been detected in association with highly purified virions, pentons, and/or dodecons of adenovirus types 2, 3, 5, 9, 12, 15, and 16. Only single-strand scissions in substrate DNA were detected. The nuclease activity was detected by a highly sensitive ethidium bromide fluorimetric assay procedure.