Maurice R. Hilleman

Live attenuated varicella virus vaccine. Efficacy trial in healthy children.

We conducted a double-blind, placebo-controlled efficacy trial of the live attenuated Oka/Merck varicella vaccine among 956 children between the ages of 1 and 14 years, with a negative clinical history of varicella. Of the 914 children who were serologically confirmed to be susceptible to varicella, 468 received vaccine and 446 received placebo. The vaccine produced few clinical reactions and was well tolerated. There was no clinical evidence of viral spread from vaccinated children to sibling controls. Approximately eight weeks after vaccination, 94 per cent of the initially seronegative children who received vaccine had detectable antibody to varicella. During the nine-month surveillance period, 39 clinically diagnosed cases of varicella, 38 of which were confirmed by laboratory tests, occurred among study participants. All 39 cases occurred in placebo recipients; no child who received vaccine contracted varicella. The vaccine was 100 per cent efficacious in preventing varicella in this population of healthy children (P less than 10(-9).

The Vacuolating Virus, S.V.40.

Summary The newly discovered vacuolating virus, S.V.40, appears to be a common and essentially ubiquitous contaminant of rhesus monkey kidney cell cultures and a likely common contaminant of cynomolgus kidney cultures in which it develops to high titer without evident cytopathic change. The virus grows readily in grivet, vervet and patas kidney and in rhesus testicle cultures causing a unique cytopathology characterized by ballooning and intense vacuolation. Some biological and biophysical properties of the virus are described. Principal interest in and importance of the vacuolating virus arise from its demonstrated presence in all 3 types of Sabins live poliovirus vaccines and in numerous respiratory virus seed lines employed in human volunteer inoculation experiments. Demonstration of the virus has raised the question of safety following administration to human subjects, especially infants, and of possible lack of validity of conclusions reached from experiments to measure neurovirulence or “genetic” markers of attenuated poliovirus vaccine strains and to determine the pathogenicity of respiratory viruses for man. The vacuolating virus is effectively dealt with in killed virus vaccines by virtue of its susceptibility to formalin. The optimal solution to the live virus vaccine problem appears to lie in elimination of the virus from such preparations.

Propagation of Human Hepatitis A Virus in Cell Culture in Vitro

Summary Human hepatitis A virus was reliably and repeatedly propagated in primary explant cell cultures of marmoset livers and in the normal fetal rhesus kidney cell line (FRhK6). Identity of virus was established in immunofluorescence, immunofluorescence blockade, serum neutralization, immune adherence, radioimmunoassay, immune electron microscopy, and marmoset inoculation tests. The virus propagated to greatest extent in FRhK6 cells. No cytopathology was observed. These studies represent the first reliable propagation of human hepatitis A virus in vitro and point the way to the eventual means for detection and quantification of live virus in vitro and of production in cell culture of virus for diagnostic antigen and vaccine preparation.

Realities and enigmas of human viral influenza: pathogenesis, epidemiology and control

Influenza A is a viral disease of global dimension, presenting with high morbidity and mortality in annual epidemics, and in pandemics which are of infrequent occurrence but which have very high attack rates. Influenza probes reveal a continuing battle for survival between host and parasite in which the host population updates the specificity of its pool of humoral immunity by contact with and response to infection with the most recent viruses which possess altered antigenic specificity in their hemagglutinin (HA) ligand. HA ligand binds the virus to the cell to bring about infection. Viral survival relies on escape from host immunity through antigenic alterations in nature which arise through genetic drift by point mutation principally of the HA gene, or through genetic shift by reassortment exchange of the HA ligand with that of viruses retained in avian species. Partial control of influenza is by use of killed whole, subunit, or possible live virus vaccines, all of which rely on worldwide surveillance to provide early detection of the altered immunologic specificity of the next virus to come. Future global surveillance may be aided by studies of sampled viral isolates in laboratories having capabilities for accelerated genetic sequencing and for automated rapid throughput analyses as well. Influenza vaccines of the future must be directed toward use of conserved group-specific viral antigens, such as are present in transitional proteins which are exposed during the fusion of virus to the host cell. Chemotherapy, though still primordial, must eventually provide the ultimate solution to vaccine failures. Probing the enigma of the severe influenza pandemic of 1918-1919 is an exciting contemporary venture in which genetic reconstruction of the viral genome from surviving archival RNA is being conducted with great success. Present evidence reveals successive recycling in pandemics, of only 3 of the 15 possible avian viral HAs. Pandemics are believed, conventionally, to be derived solely by rare events in which wild viruses of man acquire a new HA ligand of avian origin. There might be an alternative possibility involving a periodicity in selective control by the host population itself, in its receptivity or rejection at a particular time of particular reassortant viruses which might be created more frequently in nature than we are presently aware. This hypothesis, though remote, provides a different way to view and to probe the enigma of pandemic influenza.

Vaccination and Revaccination with Polyvalent Pneumococcal Polysaccharide Vaccines in Adults and Infants

Summary Adult persons developed substantial antibody increases against essentially all pneumococcal capsular types following injection of polyvalent pneumococcal vaccine containing 50 μg of each capsular polysaccharide per dose. Revaccination 13 months after the previous immunization did not evoke important further increases in antibodies and there was substantially greater local reaction at the injection site than when the previous dose was given. This finding appeared due to local reaction of antigens with circulating antibodies in the area of injection, since there was a correlation between the measured amount of circulating pneumococcal antibodies and the degree of reaction. Infants less than 2 years of age who were given a half-dose of vaccine generally responded poorly when compared with adults. In studies of 3- to 5-month-old infants, there was some increase _in antibodies when a booster dose of vaccine was given 6 months after the first. Very high level antibody responses against all capsular types were obtained when revaccination was delayed until 2 years of age.

Development of Tumors in Hamsters Inoculated in the Neonatal Period with Vacuolating Virus, SV40

Summary Intracerebral and subcutaneous injection into newborn hamsters of vacuolating virus, SV40, grown in renal cell cultures of grivet monkey resulted in single or multiple fibrosarcomas at site of injection which were histologically of varying degree of malignancy. These occurred 3 1/2 to 8 months post-inoculation and in both sexes. Animals injected with appropriate control materials or held uninoculated failed to develop tumors. Tests to exclude mouse polyoma virus as a factor were clearly negative. Evidence for the role of SV40 virus as a primary oncogenic agent was provided by recovery of the virus from the tumor and by demonstration of SV40 antigen in tumors by fluorescent antibody staining. The agent appeared to be localized in the tumors since the virus was not detected in blood or excretions, since antibody response was minimal or lacking, and since gross metastases were lacking. Transplantation and serial passage of SV40-induced tumors were accomplished with ease. The data represent the first definitive evidence implicating a virus of primate origin as a malignant oncogenic agent in experimental animals. The findings relate to observations of oncogenesis in hamsters and do not warrant extrapolation to oncogenesis in other mammalian species.

Vaccines in historic evolution and perspective: a narrative of vaccine discoveries

The sciences of vaccinology and of immunology were created just two centuries ago by Jenners scientific studies of prevention of smallpox through inoculation with cowpox virus. This rudimentary beginning was expanded greatly by the giants of late 19th and early twentieth centuries biomedical sciences. The period from 1930 to 1950 was a transitional era with the introduction of chick embryos and minced tissues for propagating viruses and Rickettsiae in vitro for vaccines. Modern era vaccinology began about 1950 as a continuum following notable advances made during the 1940s and World War II. Its pursuit has been based largely on breakthroughs in cell culture, bacterial polysaccharide chemistry, molecular biology and immunology, which have yielded many live and killed viral and bacterial vaccines plus the recombinant-expressed hepatitis B vaccine. The present paper was presented as a lecture given(1) on August 30, 1999 and recounts, by invitation, more than five-and-half decades of vaccine research from the venue of personal experience and attainment by the author. The paper is intentionally brief and truncated with focus only on highlights and limited referencing. Detailed recounting and referencing are given elsewhere in text references [Hilleman MR. Six decades of vaccine development - a personal history. Nat. Med. 1998;4 (Vaccine Suppl.): 507-14] and [Hilleman MR. Personal historical chronicle of six decades of basic and applied research in virology, immunology and vaccinology. Immunol. Rev. (in press)]. This narration will have achieved its purpose if it provides a background of understanding and guidelines that will assist others who seek to engage in creation of new vaccines.

Purification and Characterization of Chick Embryo Interferon

Summary Chick embryo interferon was purified from allantoic fluid of embryonated eggs infected with influenza A virus. Purification consisted essentially of acid precipitation to remove virus and extraneous protein, concentration and purification by precipitation with Zn++, column chromatography on CM-cellulose, and zone ionophoresis on pevikon. The interferon was purified 4500 times with respect to initial protein and one unit of interferon activity was 0.0042 μg protein in the chick embryo cell-EEE virus assay system used. Interferon was found to be a slightly basic protein of low molecular weight and free of nucleic acid as well as constituents giving absorption in the visible spectrum. A trace amount of carbohydrate was found. Detailed findings in the analyses are presented. The striking difference in properties of interferon from those described heretofore by other workers appears to lie in previous failure of purification. Both crude and purified chick interferon were active in suppressing Rous sarcoma development and NDV virus infection in susceptible chicks but were without therapeutic effect.

Specific Immune Adherence Assay for Human Hepatitis A Antibody. Application to Diagnostic and Epidemiologic Investigations

Summary A specific immune adherence (IA) test for hepatitis A antibody in human serum was described employing liver extract of marmosets infected with CR326 strain human hepatitis A virus. Persons with hepatitis A, but not hepatitis B, developed hepatitis A IA antibody soon after onset of the acute illness and this persisted thereafter. There was very close agreement in the tests for human hepatitis A immune adherence, complement fixing (CF) and neutralizing antibodies. IA antibodies appeared to develop somewhat later than CF or neutralizing antibody. A limited epidemiologic study of a family outbreak of hepatitis A and B in Costa Rica showed simultaneous occurrence of the two diseases and was supportive of the concept that susceptible persons in a country with high hepatitis A prevalence generally acquire their infections at an early age and are immune thereafter. Most persons of high socioeconomic level in an area of low hepatitis A incidence may proceed to adulthood without experience with hepatitis A. Persons of low socioeconomic level, however, such as commercial blood bank donors and prisoners, show high incidence of hepatitis A antibody. Hepatitis IA and CF antibodies persisted in human subjects for at least 7 hr after hepatitis A virus infection. Captive chimpanzees and grivet and rhesus monkeys, not given hepatitis A virus, showed evidence of previous experience with human hepatitis A or an antigenically related virus based on tests for hepatitis A antibody. Other subhuman primates, rodents, and swine, not given hepatitis A virus, were without hepatitis A antibody. The IA test provides an excellent tool for diagnostic and epidemiologic investigations of hepatitis A and should be of considerable value to detect hepatitis A virus in attempts to propagate the virus in cell culture. There was considerable difference in hepatitis A IA antibody content of different lots of commercial human immune globulin, though the majority titered 1:4000 or 1:8000.

Live, Attenuated Mumps-Virus Vaccine

MUMPS is a common childhood disease that may be severely and even permanently crippling when it involves the brain, testis, ovary, auditory nerves or pancreas. Adult males may be permanently steril...