scholarly journals STUDIES ON HOST-VIRUS INTERACTIONS IN THE CHICK EMBRYO-INFLUENZA VIRUS SYSTEM

1955 ◽  
Vol 101 (5) ◽  
pp. 493-506 ◽  
Author(s):  
Kurt Paucker ◽  
Werner Henle
Keyword(s):  
Influenza Virus ◽  
Infectious Virus ◽  
Allantoic Fluid ◽  
Progressive Reduction ◽  
Virus Particles ◽  
Host Virus Interactions ◽  
Virus Interactions ◽  
Available Information ◽  
Virus System

An experimental analysis is here presented of the conditions that lead to the appearance of non-infectious hemagglutinins (NIHA) in the allantoic fluid of chick embryos injected with standard influenza virus (PR8 strain) which had been exposed to 37°C. in vitro for various periods of time. On progressive reduction of the infectivity of the undiluted inocula from about 109 to 103 ID50 (103.2 HA units) the yields of infectious virus in 24 hours decreased in straight correspondence 1 millionfold, but those of hemagglutinins only by a factor of 10. Thus the proportions of NIHA in the yields increased sharply but the total quantity obtained decreased gradually. The quantities of infectious virus produced per ID50 injected were the same throughout this range; i.e., between 50 and 100 ID50, regardless of increasing proportions of heat-inactivated virus in the seeds. This value agrees with previous estimates of yields under other conditions. Thus, initiation and completion of first cycles by the infectious virus remaining in the inocula were not, or at most, slightly inhibited. The inactivated virus, therefore, failed to establish immediate interference. It was capable, however, of holding the infectious process to one cycle. Upon 10-fold dilution of the seeds essentially similar results were obtained except that a slight loss in interfering activity could now be detected with an increase in exposure to 37°C. With further dilutions little or no interference was noted. The capacity to yield NIHA decreased slowly during exposure of the seeds to 37°C. over a period of 5 days, thereafter more rapidly. It could not be restored by addition of infectious virus. Furthermore, since NIHA was obtained when the seeds contained as little as 102 or 103 ID50, it is unlikely that it was derived from those cells which had adsorbed both infectious and inactivated seed virus. It is suggestive that multiple adsorption of inactivated virus particles per se will yield NIHA. The available information, as discussed, favors the view that the NIHA does not represent seed virus in some form but is newly produced.

scholarly journals STUDIES ON HOST-VIRUS INTERACTIONS IN THE CHICK EMBRYO-INFLUENZA VIRUS SYSTEM

1951 ◽  
Vol 94 (4) ◽  
pp. 269-289 ◽  
Author(s):  
Oscar C. Liu ◽  
Werner Henle
Keyword(s):  
Influenza Virus ◽  
Growth Curve ◽  
High Speed ◽  
Allantoic Fluid ◽  
In Vitro Tests ◽  
Host Virus Interactions ◽  
Virus Interactions ◽  
Virus System

The role of inhibitors of hemagglutination in the evaluation of host-virus interactions in the chick embryo-influenza virus system has been analyzed. Comparisons were made between materials (allantoic fluids and membrane suspensions) derived from in vivo (growth curve) experiments at hourly intervals after inoculation, and from in vitro tests in which normal allantoic fluids and membrane suspensions were incubated with virus at 37°C. for various periods of time. In both instances large amounts of virus were added to the systems, resulting in comparable concentrations of the agent. The seeds employed were either fully active or irradiated by ultraviolet light to the extent that the virus lost its capacity to increase but kept its interfering and hemagglutinating properties. The various materials were assayed for (a) the hemagglutinating titers of the virus present in the systems before and after heating to 56°C.; (b) the concentration of inhibitor in the materials at various stages of incubation after heating to 70°C. for 30 minutes as measured by the hemagglutination-inhibition reaction with native or heated test virus (30 minutes 56°C.); and (c) the degree of adsorption of the hemagglutinins present in the materials onto chicken red cells at 0°C. and their subsequent elution at 37°C. The effects of receptor-destroying enzyme (RDE), treatment with sodium periodate, or high speed centrifugation on the inhibitory activities were studied in some of the tests. The essential results which indicate certain sources of error in the evaluation of host-virus interactions as well as means for studying virus activity at the early stages of the infectious process, were as follows: 1. Though some inhibitory effects on hemagglutination were noticeable in the allantoic fluid during the 1st hour after inoculation they were, as a rule, no longer apparent after this interval, and treatment with RDE did not increase the hemagglutinin titers. Thus, the interpretation of growth curve data concerning allantoic fluids hardly seems to be affected by inhibitor. On the other hand, striking effects were noted with the membrane suspensions of growth curve experiments in that RDE shortened the latent period to 2 hours and the titers in the first few positive samples (4 to 5 hours) increased) whereas in later harvests no such effect was noted. Under these conditions complement-fixation antigens and hemagglutinins made their appearance in the tissues simultaneously and not as previously reported the former prior to the latter. However, the infectivity showed increments only several hours after these two activities had become measurable. Thus the hypothesis of the stagewise development of influenza virus is still supported by these data. 2. Using the inhibition of hemagglutination technic it was found that the inhibitor in allantoic fluid rapidly decreased as a result of the action of active and irradiated virus, but destruction was never complete. In the membranes of the in vivo series only active seed led to loss of inhibitor, again without complete destruction, beginning at the time complement-fixing antigen and hemagglutinins became measurable. Irradiated seed was without effect in vivo whereas, in the in vitro tests it equalled the activity of the active virus. The implications of this difference in the effectiveness of active and irradiated seed in vivo with regard to the understanding of the mode of viral multiplication are discussed. 3. Although many factors may influence the shape of adsorption-elution curves it is felt that at 0°C. the extent of adsorption is directly related to the amount of inhibitor present in the systems. In the early hours after inoculation the degree of adsorption was relatively small but it increased gradually with the time of incubation. The inhibitor of adsorption was destroyed by RDE and NaIO4 and was only partially sedimentable by high speed centrifugation. In every respect studied its properties corresponded with the findings obtained with inhibitors in the hemagglutination-inhibition technic. Although the difference in the rapidity of inhibitor destruction as measured by the various technics might suggest a multiplicity of inhibitors it is felt that it rather denotes a greater sensitivity of the adsorption technic as compared to the others.

scholarly journals STUDIES ON HOST-VIRUS INTERACTIONS IN THE CHICK EMBRYO-INFLUENZA VIRUS SYSTEM

1955 ◽  
Vol 101 (5) ◽  
pp. 461-478 ◽  
Author(s):  
Norman B. Finter ◽  
Oscar C. Liu ◽  
Werner Henle
Keyword(s):  
Influenza Virus ◽  
Chick Embryo ◽  
Host Cell ◽  
Infectious Virus ◽  
Chick Embryos ◽  
In Ovo ◽  
Virus Particles ◽  
Host Virus Interactions ◽  
Virus Interactions ◽  
Virus System

An analysis has been made of factors contributing to the von Magnus phenomenon; i.e., the emergence of increasing quantities of non-infectious hemagglutinins (NIHA) in successive passages in the allantois of chick embryos of undiluted allantoic fluids infected with influenza virus. Using the PR8 strain, the von Magnus phenomenon was pronounced when the serial seeds were obtained under conditions which permitted extensive inactivation of infectious virus during individual passages. Correspondingly, it was reduced but not abolished when precautions were taken to avoid accumulation of inactivated virus in the inocula. Thus, inactivated virus may be taken as a contributing factor. Preparations of infectious virus obtained under conditions largely excluding the presence of inactivated virus were capable of yielding some NIHA on passage as long as sufficient amounts were injected to permit each host cell to adsorb several infectious virus particles. However, the fact remains that more NIHA was found in the harvests when the inocula contained a large proportion of non-infectious virus material. Following injection of various types of seeds NIHA appeared in the allantoic fluids as soon as liberation of virus became detectable. This time relationship and the rates of release of non-infectious virus components seemed to exclude that the NIHA obtained consisted entirely of infectious virus which had been inactivated during incubation in ovo. It was apparent rather that NIHA other than that due to heat-inactivated virus was released. Correlations between the infectivities and hemagglutinating capacities of over 50 standard and undiluted passage seeds and the compositions of the harvests derived therefrom on passage without dilution indicated that the corresponding activities in the yields did not depend entirely upon the relative concentrations of infectious virus and non-infectious hemagglutinins in the inocula but that apparently different forms of NIHA were obtained in successive undiluted passages.

scholarly journals STUDIES ON HOST-VIRUS INTERACTIONS IN THE CHICK EMBRYO-INFLUENZA VIRUS SYSTEM

1955 ◽  
Vol 101 (5) ◽  
pp. 479-492 ◽  
Author(s):  
Kurt Paucker ◽  
Werner Henle
Keyword(s):  
Influenza Virus ◽  
Infectious Virus ◽  
Growth Curves ◽  
Chick Embryos ◽  
Differential Growth ◽  
Host Virus Interactions ◽  
Virus Interactions ◽  
Allantoic Cavity

The role of inactivated influenza virus in the von Magnus phenomenon has been studied by exposing standard virus preparations in vitro to 37°C. for periods up to 6 or more days. The rate of inactivation of the infectious property of the line of PR8 virus employed was found to be approximately 1.1 log10 unit per day, denoting a half-life of 6½ hours. The rate of inactivation was similar in the allantoic cavity of chick embryos. On allantoic passage of such heated seeds without dilution it was seen that with a decrease in the infectivity of the inocula proportionately less infectious virus was found in the harvests. The yields of hemagglutinins were much less affected, and thus ID50/HA ratios of as low as 101.0 were observed in the 24 hour harvests. The ratios of the yields always equalled or were higher than those of the inocula. On 100- or 1000-fold dilution of the heated seeds standard virus was obtained. Growth curves in intact chick embryos or in deembryonated eggs (differential yields) revealed that non-infectious hemagglutinins appeared in the tissues or were liberated therefrom as soon as any virus activity became detectable. Furthermore, once maximal liberation had been established, infectious virus and non-infectious hemagglutinins were released for extended periods of time at nearly constant rates and in unchanging proportions, the latter depending upon the seed employed. Heated standard virus and undiluted passage seeds (von Magnus), selected on the basis of similar ID50 and HA concentrations, failed to yield similar results in differential growth curves in deembryonated eggs. Although the hemagglutinin titers in the 2-hourly harvests were nearly identical, the undiluted passage seeds produced as little as 1 per cent of the infectious virus which was derived from the heated inocula. Thus considerable differences exist between the 2 types of seeds.

scholarly journals STUDIES ON HOST-VIRUS INTERACTIONS IN THE CHICK EMBRYO-INFLUENZA VIRUS SYSTEM

1956 ◽  
Vol 103 (6) ◽  
pp. 799-822 ◽  
Author(s):  
Werner Henle ◽  
Oscar C. Liu ◽  
Kurt Paucker ◽  
Florence S. Lief
Keyword(s):  
Chick Embryo ◽  
Infectious Virus ◽  
Potassium Cyanide ◽  
Cold Room ◽  
The Status ◽  
The Media ◽  
Host Virus Interactions ◽  
Virus Interactions ◽  
Virus System ◽  
Made In

Studies have been reported concerning the relationships between virus materials found in the allantoic membranes and media of eggs deembryonated after injection of Standard (ST), heat-inactivated (37°C.) standard (ΔST), and undiluted passage (UP) seeds. It was found that the membranes always contained relatively more non-infectious hemagglutinins (NIHA) than the media and, correspondingly, the ratios between infectious virus and hemagglutinin units (ID50/HA) in the tissues were up to 1.5 log10 units lower than in the liberated progeny. These differences were seen not only following inoculation of undiluted ST, ΔST, and UP seeds, the progenies of which always contain considerable proportions of NIHA, but also when dilute ST inocula were employed which lead to the liberation of only infectious virus. Essentially similar differences in the ID50/HA ratios were observed also in the allantoic membranes and fluids obtained from growth curve experiments in the intact chick embryo employing the various types of seeds. In correlating the liberated virus materials in the media of deembryonated eggs to those in the membranes it was noted that in any given 2 hour interval during the phase of nearly constant production and release up to 10 times the quantity of infectious virus was shed as was present in the tissues at the onset of that period. In contrast, only about ¼ of the hemagglutinins were released during the same time. The viral (V) and soluble (S) complement-fixing antigens were found in the tissues but no detectable quantities were released during any 2 hour interval. The NIHA in the membranes apparently is located within the cells since it could not be released by the action of RDE. Intracellular inhibitors of hemagglutination were readily inactivated following inoculation of undiluted ST, ΔST, or UP seeds but not when ultraviolet-inactivated virus was used. The inhibitor activity decreased in proportion to the hemagglutinins produced. Transfer of infected deembryonated eggs to the cold room after production and liberation of progeny were well under way immediately halted further release but in the tissues the status quo was maintained and release was resumed on return to the 37°C. incubator. The addition of potassium cyanide to the medium of deembryonated eggs at 37°C. during the period of nearly constant production and release of virus material reduced immediately and to comparable extents the ID50 and HA titers in the tissues and liberation decreased in proportion. On removal of the cyanide 2 hours later, both titers in the tissues gradually returned to those of the untreated control eggs with a corresponding increase in liberation. The ID50/HA ratios were not affected by these manipulations. It is concluded that the NIHA in the membranes forms part of a dynamic process. An attempt has been made in the discussion to integrate the present results with previous observations concerning the formation of incomplete forms of virus and their nature and role in the infectious process.

scholarly journals STUDIES ON HOST-VIRUS INTERACTIONS IN THE CHICK EMBRYO-INFLUENZA VIRUS SYSTEM

1949 ◽  
Vol 90 (1) ◽  
pp. 23-37 ◽  
Author(s):  
Werner Henle ◽  
Gertrude Henle
Keyword(s):  
Influenza A ◽  
Allantoic Fluid ◽  
Experimental Period ◽  
Virus Antigen ◽  
Immune Serum ◽  
Host Virus Interactions ◽  
Virus Interactions ◽  
Infectivity Titer ◽  
Infectious Cycle ◽  
Virus System

In agreement with earlier observations the infectivity titer in the allantoic fluids of chick embryos injected with influenza A virus remains constant for 5 to 6 hours before an increase in this activity can be noted. In contrast, the titers of hemagglutinin and complement-fixing antigen (virus antigen) have already begun to rise after 3 hours. The origin of the hemagglutinating and complement-fixing but non-infectious material is still obscure. In the allantoic membrane development of both the soluble and virus antigens can be demonstrated after the 2nd hour of incubation and 1 hour prior to an increase in hemagglutinins and 2 hours prior to a rise in infectivity. Thus there remain the first 2 hours during which no record of virus activity in the tissues can be obtained. Similar relationships are noted on dilution of the seed both in the allantoic fluids and membranes as long as the various properties reach measurable levels during the experimental period of one infectious cycle. Injection of high titered immune serum following infection with about 109 ID50 reduces the amount of demonstrable seed virus in the allantoic fluid and membrane without significantly affecting propagation of the agent in the tissues as measured by infectivity titrations. The production of hemagglutinins appears markedly reduced under these conditions whereas formation of complement-fixing antigen is only slightly delayed and decreased. No definite explanations for the various discrepancies between the infectivity and hemagglutination can be given at present.

scholarly journals Role of viral infectivity in the induction of influenza virus-specific cytotoxic T cells.

1978 ◽  
Vol 147 (4) ◽  
pp. 1236-1252 ◽  
Author(s):  
T J Braciale ◽  
K L Yap
Keyword(s):  
T Cells ◽  
Influenza Virus ◽  
T Cell ◽  
Cell Response ◽  
Infectious Virus ◽  
Cytotoxic T Cells ◽  
T Cell Response ◽  
Cytotoxic T Cell

This report examines the requirement for infectious virus in the induction of influenza virus-specific cytotoxic T cells. Infectious influenza virus was found to be highly efficient at generating both primary and secondary cytotoxic T-cell response in vivo. Inactivated influenza virus however, failed to stimulate a detectable cytotoxic T-cell response in vivo even at immunizing doses 10(5)-10(6)-fold higher than the minimum stimulatory dose of infectious virus. Likewise inactivated virus failed to sensitize target cells for T cell-mediated lysis in vitro but could stimulate a specific cytotoxic response from primed cells in vitro. Possible requirements for the induction of virus-specific cytotoxic T-cell responses are discussed in light of these observations and those of other investigators.

scholarly journals B23/nucleophosmin is involved in regulation of adenovirus chromatin structure at late infection stages, but not in virus replication and transcription

2012 ◽  
Vol 93 (6) ◽  
pp. 1328-1338 ◽  
Author(s):  
Mohammad Abdus Samad ◽  
Tetsuro Komatsu ◽  
Mitsuru Okuwaki ◽  
Kyosuke Nagata
Keyword(s):  
Chromatin Immunoprecipitation ◽  
Infectious Virus ◽  
Core Protein ◽  
Virus Genome ◽  
Virus Particles ◽  
Core Proteins ◽  
Stimulatory Factor ◽  
Interfering Rna

B23/nucleophosmin has been identified in vitro as a stimulatory factor for replication of adenovirus DNA complexed with viral basic core proteins. In the present study, the in vivo function of B23 in the adenovirus life cycle was studied. It was found that both the expression of a decoy mutant derived from adenovirus core protein V that tightly associates with B23 and small interfering RNA-mediated depletion of B23 impeded the production of progeny virions. However, B23 depletion did not significantly affect the replication and transcription of the virus genome. Chromatin immunoprecipitation analyses revealed that B23 depletion significantly increased the association of viral DNA with viral core proteins and cellular histones. These results suggest that B23 is involved in the regulation of association and/or dissociation of core proteins and cellular histones with the virus genome. In addition, these results suggest that proper viral chromatin assembly, regulated in part by B23, is crucial for the maturation of infectious virus particles.

scholarly journals CENTRIFUGATION AND ULTRAFILTRATION STUDIES ON ALLANTOIC FLUID PREPARATIONS OF INFLUENZA VIRUS

1944 ◽  
Vol 79 (3) ◽  
pp. 301-317 ◽  
Author(s):  
William F. Friedewald ◽  
Edward G. Pickels
Keyword(s):  
Influenza Virus ◽  
Red Blood Cells ◽  
Blood Cells ◽  
Allantoic Fluid ◽  
Spherical Shape ◽  
Soluble Antigen ◽  
Chick Embryos ◽  
Virus Particles ◽  
Sedimentation Constant ◽  
Primary Virus

A synthetic density gradient technique has been applied to the study of the PR8 and Lee strains of influenza virus in the angle centrifuge. The method counteracted convective disturbances and permitted about a fiftyfold improvement in clearing supernatant fluids of virus. Sedimenting boundaries of infective virus particles, hemagglutinin, and complement-fixing antigen were obtained in the angle centrifuge and correlated with boundaries observed optically in the ultracentrifuge. The sedimentation constant of infective Lee virus particles is approximately 800 Svedberg units, while that of PR8 virus is only about 700. On the assumption of spherical shape, these values correspond to approximate diameters of 85 and 80 mµ respectively. These values agree with those obtained by filtration with graded collodion membranes. The concentration of primary virus particles in untreated allantoic fluid preparations of PR8 or Lee virus is of the order of 0.01 per cent. The primary infective particles are identical with the hemagglutinin and the complement-fixing antigen to a large extent. However, allantoic fluid preparations of PR8 virus also show a slightly inhomogeneous group of particles with an average sedimentation constant of 460 S, which are adsorbed by and eluted from red blood cells yet appear to be non-infective. In addition the virus preparations contain a small amount of "soluble antigen" which sediments more slowly than the virus and is not adsorbed by red blood cells. This soluble antigen is probably associated with material which was observed optically in the ultracentrifuge to sediment at rates ranging from very low values up to that characteristic of the primary virus boundary. This distribution of rate makes it seem likely that the material represents disintegrated virus particles. Calculations based on the experimental results obtained indicate that of the order of 10 influenza virus particles are required to produce infection of chick embryos or mice with the PR8 virus. While a comparable number is required with Lee virus for infection of chick embryos, about 10,000 particles are necessary for infection of mice. The ratio of hemagglutinin to red blood cells required to produce 50 per cent agglutination with dilute virus suspensions in the standard test is roughly 1.

scholarly journals STUDIES ON HOST-VIRUS INTERACTIONS IN THE CHICK EMBRYO-INFLUENZA VIRUS SYSTEM

1949 ◽  
Vol 90 (1) ◽  
pp. 1-11 ◽  
Author(s):  
Werner Henle
Keyword(s):  
Chick Embryo ◽  
Influenza A ◽  
Allantoic Fluid ◽  
Active Virus ◽  
B Virus ◽  
The Difference ◽  
Host Virus Interactions ◽  
Virus Interactions ◽  
Allantoic Cavity ◽  
Partial Success

Upon injection of active influenza A or B virus into the allantoic cavity of the developing chick embryo, an average of only 70 per cent of the agent was adsorbed onto the tissue, as measured by the difference between the quantity of virus injected and that found free in the allantoic fluid of the injected eggs during the constant period. The degree of adsorption was similar, regardless of whether 109 or 102 ID50 of active virus was injected. Attempts to demonstrate the adsorbed virus in suspensions of the infected tissue met with partial success only in that not more than 1 to 5 per cent of the amount calculated to be adsorbed was actually found. All efforts to increase the yield of virus have failed. These results led to the suggestion that the seed virus, which participates in the propagation, becomes altered in such a way that it no longer may be demonstrated by infectivity titrations, whereas the active virus found represents superficially adsorbed virus, which does not multiply.

scholarly journals Heat-Aggregated Noninfectious Influenza Virus Induces a More Balanced CD8+-T-Lymphocyte Immunodominance Hierarchy Than Infectious Virus

2003 ◽  
Vol 77 (8) ◽  
pp. 4679-4684 ◽  
Author(s):  
Yunjung Cho ◽  
Sameh Basta ◽  
Weisan Chen ◽  
Jack R. Bennink ◽  
Jonathan W. Yewdell
Keyword(s):  
Influenza Virus ◽  
T Lymphocyte ◽  
Infectious Virus ◽  
Ex Vivo ◽  
Intracellular Cytokine Staining ◽  
Cd8 T Lymphocyte ◽  
Immunogenic Peptides ◽  
Exogenous Antigen ◽  
Antigen Presenting

ABSTRACT CD8+-T-cell (TCD8+) responses to infectious viruses are characterized by an immunodominance hierarchy in which the majority of TCD8+ respond to one or a few immunodominant determinants, with a minority of TCD8+ responding to a number of subdominant determinants. It is now well established that exogenous antigens are capable of inducing TCD8+ to such immunodominant determinants, but the diversity of the response and the nature of the immunodominance hierarchy have not been examined. We addressed this issue by characterizing TCD8+ responses to influenza virus preparations rendered inert by incubation for 10 min at 100°C, as first reported by Speidel et al. (Eur. J. Immunol. 27:2391-2399, 1997). Extending these findings, we show that the primary TCD8+ response to boiled virus can be sufficiently robust to be detected ex vivo by intracellular cytokine staining and that the response encompasses many of the peptides recognized by TCD8+ induced by infectious virus. Importantly, the immunodominance hierarchy elicited was leveled, and we were unable to detect TCD8+ that were specific for boiled virus. We used peritoneal exudate cells as antigen-presenting cells in vitro, and a number of observations indicated that boiled virus is processed via a phagocytic route that is likely to be endosomal in nature. These findings suggest that the repertoires of immunogenic peptides generated by endosomes and cytosolic processes overlap to a surprising degree. Furthermore, they demonstrate that the form of antigen administered can influence immunodominance hierarchies and that exogenous-antigen vaccines can induce broad and balanced TCD8+ responses.

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