PR8 Strain Research Articles

Influence of Non-Specific Inhibitor on the Diagnostic Hemagglutination-Inhibition Test for Influenza

Summary Human sera contained heat stable (56 C) nonspecific inhibitor for influenza A and B hemagglutinin but not for influenza C. This inhibitor appeared to be of the alpha type and was destroyed by treating with filtrate from cultures of Vibrio cholerae which were rich in Burnet's receptor destroying enzyme. This procedure did not have a demonstrable effect on antibody. The PR8, FM1 and Lee strains of virus used in the Standard Hemagglutination-Inhibition Reference Test in Influenza Studies were highly sensitive to the alpha inhibitor. The inhibitor content for these and other strains often exceeded the amount of antibody. The reliability and sensitivity of the hemagglutination-inhibition technique for demonstrating antibody increase in paired serum specimens from cases of influenza and from vaccinated persons using the PR8, FM1, Lee and IB1 viruses was markedly improved by treating to remove inhibitor. This resulted primarily from the marked reduction in titer of the first serum specimen. The sensitivity of the test also depended on test virus; thus, the type A prime strains FM1 (1947) and FW-1-50 (1950) were superior to the 1934 PR8 agent for detecting infection with the A prime viruses of contemporary prevalence. The FW-1-50 virus was especially useful for diagnosis since it was essentially unaffected by the alpha inhibitor in human serum and the need for treatment with cholera filtrate was obviated. The importance of removing non-specific inhibitor from sera tested to determine the basic antibody levels in the human population for the various influenza strains was discussed.

Production of fever by influenzal viruses. IV. Tolerance to mumps and influenza pyrogens.

SummaryIntravenous injection of mumps virus causes a febrile response in rabbits similar to that produced by influenza or Newcastle disease viruses. Animals are partially or completely resistant to a second injection of mumps virus 24 hours later. Pretreatment with the PR8 strain of influenza A virus also inhibits the febrile response to mumps virus. Mumps virus itself is equally effective in producing tolerance to the PR8 strain.

Suspended cell tissue for study of virus growth kinetics.

The embryonated hen's egg, the deembry-onated egg, and, more recently, tissue cultures using the chick embryo chorioallantoic membrane have been widely used in the study of the growth of influenza virus (1-13). The present investigation was undertaken to appraise the suspended cell tissue culture for the quantitative determination of the multiplication of this agent. Attention has been focused on early virus production as measured by titration of infectivity in tissue culture flasks.Materials and methods. Virus. The PR8 strain of influenza A virus propagated in chick chorioallantoic membrane tissue culture has been used in all experiments. This agent was received from Dr. Frank Horsfall 15 years ago and has been maintained since then by allantoic passage in the chick embryo. The stock virus consisted of the fluid removed from cultures inoculated 36 to 48 hours previously with 0.1 ml of infected allantoic fluid diluted 10-3. It was stored at −70°C in glass-sealed ampules.“Titration” flasks. For titration o...

Reliability of 50% egg infectivity titrations with suspensions of influenza viruses prepared from mouse lungs.

Summary1. The use of 2 different methods for analyzing the reliability of 50% end point determination has been illustrated. The data were available because duplicate titrations of a number of virus suspensions were required for the purposes of other experiments. Data of comparable significance could have been obtained by a similar number of replicate titrations of the same virus suspension. The latter type of procedure is tedious, expensive, and is seldom carried out. It would seem likely that the records of other investigators would contain data suitable for analysis by the methods used in this report. 2. In the present study the reliability of 50% egg infectivity titrations was determined with crude suspensions of lungs infected with recently isolated strains of Type A-prime and Type B influenza viruses. The reliability of these determinations was similar to that previously described for titrations of infectivity for mice or for eggs with purified suspensions of the PR8 strain of influenza virus.

Effect of sodium monofluoroacetate on the multiplication of influenza viruses, mumps virus and pneumonia virus of mice (PVM).

Sodium fluoroacetate, given after virus inoculation in doses of 3 to 4 mg. per kg. in mice or 2 mg. in chick embryos, caused only a slight delay in the multiplication of the PR8 strain of influenza A virus in the mouse lung and of PR8 or the Lee strain of influenza B virus in the allantoic sac. The quantities of the compound used were sufficient to cause approximately 10 to 20 per cent mortality in mice and 100 per cent in chick embryos. The use of small virus inocula did not markedly increase the effect of sodium fluoroacetate on the multiplication of PR8 or Lee in the chick embryo and maximal titers were obtained in all cases. In contrast to the findings in the chick embryo, sodium fluoroacetate caused a definite delay in the multiplication of Lee virus in the mouse lung but did not affect the final virus titer. Sodium fluoroacetate in like amounts caused only a minimal delay in the multiplication of pneumonia virus of mice (PVM) in the mouse lung or of mumps virus in the chick embryo. With both PVM and mumps virus, maximal titers were obtained almost simultaneously in fluoroacetate and control animals. When three daily injections of the compound were given to mice infected previously with PVM, a definite diminution in the virus titer was demonstrable. However, pretreatment with three daily injections of the compound caused no alteration in the capacity of mice to support the multiplication of PVM. From the results of these experiments, it appears that the cellular metabolic processes blocked by sodium fluoroacetate are not essential for the multiplication of influenza viruses, mumps virus, or pneumonia virus of mice (PVM).

Counts of virus particles by sedimentation on agar and electron micrography.

SummaryA method has been devised for counting virus particles in electron micrographs of the virus sedimented on an agar surface and removed quantitatively in a collodion pseudoreplica. With influenza virus A (PR8 strain) as test virus, the accuracy of the counts, under conditions practical for routine studies, was the same as that observed with the spray technic or by sedimentation of the agent on collodion. In common with the latter method, sedimentation on agar is applicable to materials of relatively low virus content and, since salt diffuses into the agar, to viruses harmed either by drying in the presence of salt or by salt-free water. The usefulness of this procedure is limited to the study of virus particles which can be recognized either because of characteristic morphology or of other properties that can be related quantitatively to virus behavior.

Concentration of influenza virus (PR8 strain) by a cation exchange resin.

SummaryA simple method has been described for the rapid concentration and partial purification of the PR8 strain of influenza virus by means of an ion exchange resin.

Quantitative distribution of phosphorus in chorioallantoic membrane as affected by infection with influenza virus.

SummaryIt has been shown that infection of the chorioallantoic membrane by the PR-8 strain of influenza virus results in: 1. The decrease in the phospholipid phosphorus concentration of the membrane; 2. The presence of large amounts of lipid-bound phosphorus in the fluid medium; and 3. A more rapid uptake of P32 into the membrane.

Quantitative aspects of the multiplication of influenza A virus in the mouse lung; relation between the degree of viral multiplication and the extent of pneumonia.

Influenza A virus, PR8 strain, increases in amount in the infected mouse lung at a relatively constant rate. When more than 25 M.S.50 doses of virus is inoculated, the rate of multiplication appears to be independent of the amount of virus introduced; has a value of 1,100-fold increase per day. The rate of increase in the pulmonary lesions induced by infection of the mouse lung with PR8 also appears to be relatively constant and independent of the amount of virus inoculated; has a value of 8.5-fold increase per day. The essential variables in the PR8-mouse lung system appear to be equated satisfactorily by functions which were derived previously (4) during a similar quantitative investigation on pneumonia virus of mice (PVM). Evidence in support of the hypothesis that the processes of multiplication of PR8 and PVM are different in the mouse lung is presented.

インフルエンザの自然感染免疫

In order to understand the nature of influenza epidemics, it is necessary to investigate the susceptibility to the infection on the part of individuals as well as the very nature of influenza viruses. The most important factor which determines one's susceptibility to influential affection is the neutralizing capacity of one's serum against influenza viruses. According to the literature available on this subject, it is probable that human adults always possess a certain level of neutralization antibodies against influenza viruses A and B, and that children, except for a year after birth when an adult's level of antibodies transferred from mother can be demonstrated in them, show the lowest level of the neutralizing capacity which, as a result of repeated influenzal affection, gradually increases with age and reaches the adult's level before 10 to 15 years of age. However, our existing knowledge is not yet sufficient to make this point clear, and it is desirable to prosecute the study on a large number of individuals in many different areas.So we investigated the distribution of neutralization antibodies against influenza viruses A and B in different age groups of population, particularly school-aged (6 to 15 years of age) children, in different areas of Western Japan. The areas where the investigation was made numbered nineteen, including cities, rural districts, remote mountain regions and solitary islands, and the number of people whose sera was examined totaled more than 1500. Neutralization antibodies of serum against influenza virusee A (PR8 strain) and B (Lee strain) were estimated by the hemagglutination inhibition test, and the results obtained were expressed in terms of Fukumi-Yamamoto's “hemagglutination inhibition index”.The results obtained were as follows:1. The hemagglutination inhibition indices of adults were between 3.0 to 4.0, and this range of neutralizing capacity is maintained throughout life.2. In all areas examined, school-aged children showed considerable or an adult's level of neutralization antibodies. The fact indicated that infection of influenza viruses A and B were very common in those areas.3. In children, the adult's level of the neutralizing capacity against A virus was reached before the ages of 6 to 13. But against influenza B virus, this level war not yet reached untill 14 or 15 in many areas.4. The mode of distribution of the neutralization antibodies in different age groups of children can be classified into two different types. In the first type, the antibodies are distributed in a narrow range at the adult's level, showing no differences in different age groups. In the second type, the antibodies as a whole are distributed in a wide range, the levels varying from a very low value to that of the adult; furthermore, they show definite age differences in their distribution.5. As the state of age distribution of neutralization antibodies against influenza viruses A and B in a certain population clearly reflects the state of its resistance to influenza epidemics, the knowledge of the changes in the age distribution soon after outbreak of the epidemic will enable us to diagnose whether the causative agent was virus A or B. Some illustrative epidemics were descibed.

Propagation of the PR8 strain of influenza A virus in chick embryos. II. The formation of incomplete virus following inoculation of large doses of seed virus.

Propagation of the PR8 strain of influenza A virus in chick embryos. II. The formation of incomplete virus following inoculation of large doses of seed virus.

Propagation of the PR8 strain on influenza A virus in chick embryos. I The influence of various experimental conditions on virus multiplication.

Propagation of the PR8 strain on influenza A virus in chick embryos. I The influence of various experimental conditions on virus multiplication.

A Method for Estimating the Capacity of Influenza Virus to Effect Antigenic Modification of Red Blood Cells

Summary Following treatment with a vibrio filtrate or the PR8 or Lee strains of influenza virus human Group O red cells develop closely similar if not identical antigenic changes. A method for estimating the cell modifying activity (CMA) of virus fluids is described and it is shown that the titers obtained by this method are considerably higher than hemagglutination titers as determined by the pattern test. The capacity of PR8 allantoic fluid to effect antigenic modification of human Group O red cell is less susceptible to the action of heat or ultra-violet irradiation than is infectivity in eggs.

Air sterilization in an infants' ward; effect of triethylene glycol vapor and dust-suppressive measures on respiratory cross infection rate.

The control of respiratory cross infections in infants' wards has always presented a serious problem.¹Since the introduction of gamma globulin, sulfonamides and antibiotics this problem has become less acute. Nevertheless, there are still far too many respiratory tract infections—bacterial, viral and of unknown cause— which are acquired after admission to a hospital ward. The relative importance of contact versus air-borne infection has not been established. However, in recent years the air-borne mode of transmission has received much emphasis.²The study to be described is concerned with two measures for the control of air-borne infection—use of triethylene glycol vapor and dust suppression by oiling. Triethylene glycol vapor possesses marked bactericidal and virucidal properties against a variety of airborne micro-organisms. Robertson and his associates³demonstrated that it was rapidly lethal for hemolytic streptococci, pneumococci, staphylococci, influenza bacilli and the PR8 strain of influenza virus. Rosebury and his associates

The Egg-White Inhibitor of Influenza Virus Hemagglutination

Summary Semipurified egg-white inhibitor was treated for various periods with influenza virus A (PR8 strain), B (Lee strain), or the swine influenza virus and the effects were tested with heated derivatives of each of the 3 viruses. The existence of an inhibitor gradient, written PR8, Lee, Swine, was established, such that each virus was capable of destroying completely the inhibitor for itself as well as for the preceding virus or viruses, but only partially the inhibitor for the virus or viruses which follow it in the gradient. The same gradient appeared to describe the order of increasing inhibition titers of untreated inhibitor against the corresponding heated viruses, as well as the order of increasing apparent complexity of the changes induced by active virus in the character of the inhibition curve. The results have been interpreted in terms of the structure of the inhibitor and the mechanism of destruction of the inhibitor by virus.

Further studies on serial passage of a mixture of influenza viruses in embryonated eggs.

SummaryUndiluted allantoic fluids were used as inocula for serial passage of a mixture of the PR8 and Lee strains of influenza virus in embryonated eggs. Although wide differences were found in the relative concentrations of the two viruses in the various fluids, the mixture was successfully carried through 52 consecutive passages, and each agent was recovered in an apparently pure culture at the end of the experimental period. Since the two viruses have different growth rates and are capable of interfering with the multiplication of each other, the persistence of both agents during serial passage of the mixture suggests that multiplication of the PR8 and of the Lee viruses may not require the same host factors.

Antigenic pattern of strains of influenza A and B.

SummaryInfluenza A viruses, as revealed by agglutination-inhibition tests with rooster antisera, comprised a graded series of antigenic forms in which precise subgrouping was not possible by the present method. The B strains examined fell into 3 subgroups. The reappearance from time to time of strains closely related to an earlier prototype, necessitates the selection of a group of strains for a polyvalent vaccine which possess broad immunogenic activity. Certain possible deficiencies in the present formula vaccine were mentioned. The recent outbreak of influenza among Eskimos in remote Victoria Island, Canada, in which there was mortality and morbidity approaching the pandemic form of disease, was apparently caused by 2 different viruses which resembled the well-studied PR8 and FM1 strains of influenza A.

The Sedimentable Components of Influenza Virus Propagated in Deembryonated Eggs

Summary and Conclusions The PR8 strain of type A influenza virus diluted to 10−1 or 10−5 with Tyrode's solution was propagated in deembryonated eggs. The preparations obtained in this manner were concentrated by ultracentrifugation, examined in the analytical ultracentrifuge and tested for hemagglutinating and infectious capacities. It was found that preparations obtained when the inocula were diluted 10 times were of low infectivity, moderately high hemagglutinating capacity and contained one major component sedimenting at approximately 480 S. Preparations obtained when the inocula were diluted to 10−5 also contained only one major component, but this exhibited a sedimentation constant of approximately 630 S, a high infectivity, and the hemagglutinating capacity was comparable to that of the 480 S preparations. It has been concluded from these and from other data that the infectious principle of the PR8 strain of Type A influenza virus resides primarily in the heavy component, while the hemagglutinative capacity is distributed between heavy and light components.

THE CHEMICAL ANALYSIS OF PURIFIED INFLUENZA VIRUS A (PR8 STRAIN) CONTAINING RADIOACTIVE PHOSPHORUS

Purified influenza virus A (PR8 strain) was found to contain about 11% phospholipid and 5% nucleic acid in agreement with previously reported work. The method of Schmidt and Thannhauser, applied to the nucleic acid fraction of the virus, indicated the presence of 4.5% pentose nucleic acid and 0.3% desoxypentose nucleic acid. When influenza virus was grown in the allantoic membrane of the embryonated egg in the presence of inorganic radioactive phosphorus both phospholipid and nucleic acid components of the virus were found to contain the isotope. The specific radioactivity of the nucleic acid fraction was about four times that of the phospholipid.

Observations on the toxin of influenza virus.

SummaryThe mouse toxic factor.of influenza virus, PR8 strain, originally reported by the Henles was reinvestigated, and similarities and differences between their observations and ours are reported and discussed. The toxin also was shown to affect ferrets.