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Antigenicity of Beta-propiolactone Inactivated Newcastle Disease Virus Vaccines Prepared from Various Strains*
Antigenicity of Beta-propiolactone Inactivated Newcastle Disease Virus Vaccines Prepared from Various Strains* JOHN GRUN Department of Animal Sciences, Rutgers, The State University, New Brunswick, New Jersey (Received for publication October 6, 1965)
B
* Paper of the Journal Series, New Jersey Agricultural Experiment Station, Rutgers, The State University of New Jersey, New Brunswick, New Jersey.
minimum concentration of the inactivating agent acting on a virus unencumbered by excessive tissue for the shortest period at the lowest effective temperature constitutes ideal environmental conditions for the preparation of an inactivated Newcastle disease vaccine." The effects of BPL inactivated whole embryo ingredients and AA fluid NDV vaccines are described by Grun (1966). When Grun and Hudson (1966) compared a BPL inactivated whole embryo NDV vaccine containing one strain of virus (GB) with a BPL inactivated AA fluid NDV vaccine containing four strains of virus, the AA fluid preparation appeared to be the superior vaccine. This report presents the results of experiments with several BPL inactivated AA fluid NDV vaccines containing from one to four strains of virus. The NDV vaccines were all prepared in the same manner and their immunogenicity was determined by challenging groups of birds at 4, 8, 12, and 16 weeks after vaccination. Also included in this report were the effects on mortality of the BPL inactivated ingredients of these vaccines (no virus). This corroborates data reported earlier (Grun, 1966). EXPERIMENTAL PROCEDURE
Unless otherwise indicated, the following procedures were used throughout our experiments. S.C. White Leghorn chicks were brooded in isolation until 4 weeks of age. One day before vaccination a number of birds (10) were randomly selected and bled and hemagglutination-inhibition (HI) tests were carried out on the individual serum samples to test for parental im-
492
Downloaded from http://ps.oxfordjournals.org/ at Florida Atlantic University on November 7, 2014
ASED on the work of Kelly (1052), Kelly and Hartman (1952), and Lo Grippo and Hartman (1954, 1955), Mack and Chotisen (1955) showed that beta-propiolactone (BPL) inactivated Newcastle disease virus (NDV) vaccines are feasible. Their study lacked, however, experiments to determine which strains would produce the strongest immunity over the longest period of time. This stimulated further work in that direction by Sullivan et al. (1958). Chicks treated with a single inoculation of BPL inactivated NDV (George Boney) vaccine at 2 weeks of age were quite resistant to challenge up to 12 weeks of age. When birds were treated at 2 weeks and again at 6 weeks of age, no differences were found between the following NDV strains: George Boney (GB), Roakin-N.J., and Manhattan-Kansas. Hofstad et al. (1963) evaluated the immunogenicity of NDV preparations inactivated with gamma radiation, formalin, and BPL at 6 weeks post treatment. This 1963 work corroborated in part that of Winmill and Weddell (1961), where local NDV strains were used and the challenge virus was a homologous strain. It was also suggested that infected embryonic fluid was superior to whole egg contents for the preparation of a BPL-NDV vaccine. Although Appleton et al. (1963) thought that a critical balance may exist between the concentration of BPL and amnio-allantoic (AA) fluid, this point was not adequately explored. They also stated that "a
493
NEWCASTLE DISEASE VIRUS loon
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Foxnlol Immwnily Unlraotcd Conlfdit AA Fluid • 4 NOV AA Fluid - 1 ml AA Fluid - OS ml toS- 1 ml G l - OS ml
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A GB, LaS and Ho - OS ml A Ho - 0 . 5 ml 9 GB and la S - OS ml
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INOCULATION
FIG. 1. Response of chickens to intranasal challenge with virulent NDV (GB) at various time intervals after treatment with the following BPL inactivated amnio-allantoic fluid vaccines: 4 NDV (GB, I Pa, LaS, Ho (0.5 ml.)); AA fluid (0.5, 1 ml.); LaS (1 ml.); GB (0.5 ml.); 3 NDV (GB, LaS, Ho (0.5 ml.)); Ho (0.5 ml.); and 2 NDV (GB, LaS (0.5 ml.)). RESULTS
The data in Figure 1 are expressed as the medians of mortality percentages resulting from the IN GB-NDV challenge at 4, 8, 12, and 16 weeks after treatment. Each point represents a composite of mortality percentages of 10 to 70 birds derived from one to six experiments; each group, therefore, has been duplicated at least once, except preparations No. 6 and No. 7, which contained only the Ho strain and the GB and LaS strains, respectively. The two groups that had no mortalities throughout the challenge period of 16 weeks were those treated with the vaccines containing all four strains (No. 1, given in 0.5 and 1 ml. amounts) and three strains (No. 5, given
Downloaded from http://ps.oxfordjournals.org/ at Florida Atlantic University on November 7, 2014
munity. These birds were then challenged intranasally with 0.1 ml. of George Boney (GB) NDV. This dosage of the GB strain (titer 10s ELD50/O.I ml.) was the standard challenge used in every experiment. After vaccination, all birds were maintained in isolation quarters until just prior to their exposure to the GB challenge. A number of birds from each group were selected at random to be bled for individual HI testing and then challenged at specific time intervals. The experiments discussed in this publication were run concurrently and intercurrently, depending on the amount of space available. The protocols were as similar as possible in order that valid conclusions could be derived from the comparison of one experiment to another. This laboratory's initial 0.2% BPL inactivated preparation was an equal mixture of 4 NDV strains (Preparation No. 1) in AA fluid and, after testing for live virus by chicken embryo inoculation, 50% by volume of amphojel was added. The four virus strains were: GB, La Sota (LaS), Indian Partridge (I Pa), and the Horowitz (Ho) isolate of NDV. All strains except GBNDV were isolated in this laboratory and maintained here for some period of time. Our no-virus preparation (No. 2) was normal AA fluid treated with 0.2% BPL and given at two dosage levels, 0.5 ml. and 1 ml. All the inoculations were given in either 0.5 ml. or 1 ml. amounts subcutaneously in the neck region. Preparation No. 3 contained only the LaS strain; No. 4 the GB strain; No. 5 the GB, LaS and Ho; No. 6 the Ho; and No. 7 the GB and LaS. A number of randomly selected birds from each of the experimental and control groups were challenged IN with 0.1 ml. live GB virus at 4, 8, 12, and 16 week intervals after treatment. The birds were observed for 2 weeks after exposure to the challenge virus; those exhibiting CNS involvement were also counted as mortalities.
494
J. GRUN
DISCUSSION
The data suggest that under the conditions outlined for these experiments, previously established mortality patterns can be reproduced. Essentially, the experiments were concerned with the response of groups of chickens to GB challenge after having undergone one of the following treatment regimens: no treatment; BPL-inactivated AA fluid in 0.5 ml. and 1 ml. doses; BPLinactivated vaccines composed of from 1 to 4 strains of NDV (GB, LaS, I Pa, Ho) in 0.5 ml. doses, with the exception of LaS alone (1 ml.). Although the first two NDV strains are familiar to most investigators, the latter two strains (I Pa and Ho) may
need some description. The I Pa strain given IN is similar in neurotropic and other properties to the GB strain, with the exception that the onset of symptoms is delayed for approximately 24 hours in birds less than 12 weeks of age with no parental immunity. Although the Ho isolate is similar to the I Pa strain, only 30 to 50% of the birds will show CNS involvement. CNS involvement is evident in 95 to 100% of the birds infected with the GB and I Pa strains. The LaSota strain very rarely, if ever, produces CNS involvement when given intranasally. The GB challenge was at times homologous, heterologous, or both, depending on the composition of the NDV vaccines. From our data, therefore, we can conclude that it is the neurotropic virus particles present in the vaccines which determine the antigenicity and thus the amount of protection to a virulent neurotropic NDV challenge for a period of at least 16 weeks. The mortality in the Ho-treated group at 4 weeks may have been the result of any number of circumstances, but the results with this vaccine indicate it has some merit for at least 16 weeks after vaccination. The La Sota strain evidently is not a good BPL-inactivated vaccine strain, alone or in conjunction with GB (Preparation No. 7). Whether this is due to its reducing the concentration of neurotropic particles or to a form of inhibition or interference cannot be determined by these data. The group treated with the GB strain alone also had some mortalities at 16 weeks after treatment; this may have been the consequence of an interaction between the ingredients and the homologous challenge virus. Those vaccines containing the greatest concentration of neurotropic particles tend to make the best BPL inactivated NDV vaccines. These vaccines were given in a single dose of 0.5 ml. or 1 ml. and were challenged with a large concentration of virulent GB
Downloaded from http://ps.oxfordjournals.org/ at Florida Atlantic University on November 7, 2014
in 0.5 ml. amounts and in which the I Pa strain was omitted). The group treated with preparation No. 6 had some mortalities on challenge at 4 weeks after treatment but none for the remainder of the 16 week period. Groups receiving No. 4 (GB) and No. 7 (GB and LaS) were protected for 12 weeks; at 16 weeks, however, they had mortalities of 7% and 25%, respectively. These latter preparations (No. 6, No. 4, and No. 7) were given at the 0.5 ml. dosage level. The magnitude of the mortalities, beginning at 4 weeks and continuing for 8, 12, and 16 weeks, would preclude preparation No 3 (LaS) from consideration as the only viral component of a BPL inactivated NDV vaccine. Further, the La Sota preparation was given in 1 ml. amounts and contained more virus protein (as shown by Kjeldahl analysis) than the 4-strain vaccine. The results with BPL inactivated AA fluid (No. 2) given in 0.5 ml. and 1 ml. inoculations were similar to the results described previously (Grun, 1966), except for a decrease in mortality in the 0.5 ml. AA fluid group at the 16 week challenge. It should again be noted that the ingredients of these vaccines can influence the course of the challenge infection.
NEWCASTLE DISEASE VIRUS
SUMMARY
In experiments involving six BPL inactivated NDV vaccines inoculated subcutaneously in the neck region and containing one or more strains, with characteristics ranging from non-neurotropic to almost completely so, it was found that those containing two or more strains with neurotropic tendencies (0.5 ml. dose) were the most effective during the 16 week experimental period. The GB strain vaccine may be better than can be concluded from the data; this may have been due to the homologous challenge virus and the influence of the vaccine ingredients. The La Sota strain alone, given in 1 ml. amounts, a dosage in which the vaccine ingredient influence tends to favor the host, is not sufficiently antigenic to be considered as the only virus component of a BPL inactivated NDV vaccine. It is evident that more work is needed in regard to the influence of the ingredients of an AA fluid NDV vaccine, the combina-
tion of strains used, and the effect of homologous vs. heterologous challenges. ACKNOWLEDGMENTS
The author is grateful for the technical assistance and the graphics of Miss E. DeBolle and for assistance with the manuscript from Dr. E. V. Adams. REFERENCES Appleton, G. S., S. B. Hitchner and R. W. Winterfield, 1963. A comparison of the immune response of chickens vaccinated with formalinand beta-propiolactone-inactivated Newcastle disease vaccines. Am. J. Vet. Res. 24: 827-831. Grun, J., 1966. Synergistic and antagonistic reactions with beta-propiolactone inactivated disease virus vaccine ingredients. Poultry Sci. 45: 126-131. Grun, J., and C. B. Hudson, 1966. Comparison of a beta-propiolactone whole embryo Newcastle disease vaccine (one strain) with a beta-propiolactone amnio-allantoic Newcastle disease vaccine (four strain). Poultry Sci. 4 5 : 159-164. Hofstad, M. S., J. C. Picken, Jr., K. E. Collins and H. W. Yoder, Jr., 1963. Immunogenicity of inactivated Newcastle disease virus preparations. Avian Dis. 7 : : 435-445. Kelly, A. R., 1952. Effects of virucide beta-propiolactone and its degradation products on blood platelets. Fed. Proc. 11: 419. Kelly, A. R., and F. W. Hartman, 1952. Biological effects of beta-propiolactone. Fed. Proc. 11: 419. Lo Grippo, G. A., and F. W. Hartman, 1954. Antigenicity of beta-propiolactone inactivated virus. Fed. Proc. 13 : 503. Lo Grippo, G. A., and F. W. Hartman, 1955. Antigenicity of |3-propiolactone inactivated virus vaccines. J. Immunol. 75: 123-128. Mack, W. N., and A. Chotisen, 1955. Beta-propiolactone as a virus altering agent for Newcastle disease vaccine. Poultry Sci. 34: 10101013. Sullivan, J. F., E. Gill and A. M. Somer, 1958. Immune responses of chickens to beta-propiolactone-killed Newcastle disease vaccines. Am. J. Vet. Res. 19: 483-488. Winmill, A. J., and W. Weddell. 1961. A Newcastle disease vaccine inactivated by betapropiolactone. Res. Vet. Sci. 2: 381-386.
Downloaded from http://ps.oxfordjournals.org/ at Florida Atlantic University on November 7, 2014
particles. They induced the strongest immunity up to the end of our experimental time (16 weeks). The GB strain alone may be better than the data reflects, since the homologous challenge virus may find the host in a specific hypersensitive condition and prone to synergism. Birds treated with vaccines containing two or more strains with neurotropic properties tend to survive the challenge best. All of the vaccines may be better than indicated by these data, since they were also subject to the synergistic reaction brought about by the vaccine ingredients (at the 0.5 ml. dosage level). This, of course, requires more experimentation in regard to the ingredient factors involved, the influence of these factors, and the magnitude of that influence with or without the presence of NDV particles.
495
B
* Paper of the Journal Series, New Jersey Agricultural Experiment Station, Rutgers, The State University of New Jersey, New Brunswick, New Jersey.
minimum concentration of the inactivating agent acting on a virus unencumbered by excessive tissue for the shortest period at the lowest effective temperature constitutes ideal environmental conditions for the preparation of an inactivated Newcastle disease vaccine." The effects of BPL inactivated whole embryo ingredients and AA fluid NDV vaccines are described by Grun (1966). When Grun and Hudson (1966) compared a BPL inactivated whole embryo NDV vaccine containing one strain of virus (GB) with a BPL inactivated AA fluid NDV vaccine containing four strains of virus, the AA fluid preparation appeared to be the superior vaccine. This report presents the results of experiments with several BPL inactivated AA fluid NDV vaccines containing from one to four strains of virus. The NDV vaccines were all prepared in the same manner and their immunogenicity was determined by challenging groups of birds at 4, 8, 12, and 16 weeks after vaccination. Also included in this report were the effects on mortality of the BPL inactivated ingredients of these vaccines (no virus). This corroborates data reported earlier (Grun, 1966). EXPERIMENTAL PROCEDURE
Unless otherwise indicated, the following procedures were used throughout our experiments. S.C. White Leghorn chicks were brooded in isolation until 4 weeks of age. One day before vaccination a number of birds (10) were randomly selected and bled and hemagglutination-inhibition (HI) tests were carried out on the individual serum samples to test for parental im-
492
Downloaded from http://ps.oxfordjournals.org/ at Florida Atlantic University on November 7, 2014
ASED on the work of Kelly (1052), Kelly and Hartman (1952), and Lo Grippo and Hartman (1954, 1955), Mack and Chotisen (1955) showed that beta-propiolactone (BPL) inactivated Newcastle disease virus (NDV) vaccines are feasible. Their study lacked, however, experiments to determine which strains would produce the strongest immunity over the longest period of time. This stimulated further work in that direction by Sullivan et al. (1958). Chicks treated with a single inoculation of BPL inactivated NDV (George Boney) vaccine at 2 weeks of age were quite resistant to challenge up to 12 weeks of age. When birds were treated at 2 weeks and again at 6 weeks of age, no differences were found between the following NDV strains: George Boney (GB), Roakin-N.J., and Manhattan-Kansas. Hofstad et al. (1963) evaluated the immunogenicity of NDV preparations inactivated with gamma radiation, formalin, and BPL at 6 weeks post treatment. This 1963 work corroborated in part that of Winmill and Weddell (1961), where local NDV strains were used and the challenge virus was a homologous strain. It was also suggested that infected embryonic fluid was superior to whole egg contents for the preparation of a BPL-NDV vaccine. Although Appleton et al. (1963) thought that a critical balance may exist between the concentration of BPL and amnio-allantoic (AA) fluid, this point was not adequately explored. They also stated that "a
493
NEWCASTLE DISEASE VIRUS loon
•
90
• 0 80-
•
* •
:: 70
cP >- 60-
0
•
ALI
l—
O
1—
<* 50-
•
O S -Jo °
M 40-
n 30-
0
Foxnlol Immwnily Unlraotcd Conlfdit AA Fluid • 4 NOV AA Fluid - 1 ml AA Fluid - OS ml toS- 1 ml G l - OS ml
O
A GB, LaS and Ho - OS ml A Ho - 0 . 5 ml 9 GB and la S - OS ml
20-
10-
M • O O * O •
A OBAa
• OkfA
GIAAO
A*
U*f TIME-WEEKS-AFTER
INOCULATION
FIG. 1. Response of chickens to intranasal challenge with virulent NDV (GB) at various time intervals after treatment with the following BPL inactivated amnio-allantoic fluid vaccines: 4 NDV (GB, I Pa, LaS, Ho (0.5 ml.)); AA fluid (0.5, 1 ml.); LaS (1 ml.); GB (0.5 ml.); 3 NDV (GB, LaS, Ho (0.5 ml.)); Ho (0.5 ml.); and 2 NDV (GB, LaS (0.5 ml.)). RESULTS
The data in Figure 1 are expressed as the medians of mortality percentages resulting from the IN GB-NDV challenge at 4, 8, 12, and 16 weeks after treatment. Each point represents a composite of mortality percentages of 10 to 70 birds derived from one to six experiments; each group, therefore, has been duplicated at least once, except preparations No. 6 and No. 7, which contained only the Ho strain and the GB and LaS strains, respectively. The two groups that had no mortalities throughout the challenge period of 16 weeks were those treated with the vaccines containing all four strains (No. 1, given in 0.5 and 1 ml. amounts) and three strains (No. 5, given
Downloaded from http://ps.oxfordjournals.org/ at Florida Atlantic University on November 7, 2014
munity. These birds were then challenged intranasally with 0.1 ml. of George Boney (GB) NDV. This dosage of the GB strain (titer 10s ELD50/O.I ml.) was the standard challenge used in every experiment. After vaccination, all birds were maintained in isolation quarters until just prior to their exposure to the GB challenge. A number of birds from each group were selected at random to be bled for individual HI testing and then challenged at specific time intervals. The experiments discussed in this publication were run concurrently and intercurrently, depending on the amount of space available. The protocols were as similar as possible in order that valid conclusions could be derived from the comparison of one experiment to another. This laboratory's initial 0.2% BPL inactivated preparation was an equal mixture of 4 NDV strains (Preparation No. 1) in AA fluid and, after testing for live virus by chicken embryo inoculation, 50% by volume of amphojel was added. The four virus strains were: GB, La Sota (LaS), Indian Partridge (I Pa), and the Horowitz (Ho) isolate of NDV. All strains except GBNDV were isolated in this laboratory and maintained here for some period of time. Our no-virus preparation (No. 2) was normal AA fluid treated with 0.2% BPL and given at two dosage levels, 0.5 ml. and 1 ml. All the inoculations were given in either 0.5 ml. or 1 ml. amounts subcutaneously in the neck region. Preparation No. 3 contained only the LaS strain; No. 4 the GB strain; No. 5 the GB, LaS and Ho; No. 6 the Ho; and No. 7 the GB and LaS. A number of randomly selected birds from each of the experimental and control groups were challenged IN with 0.1 ml. live GB virus at 4, 8, 12, and 16 week intervals after treatment. The birds were observed for 2 weeks after exposure to the challenge virus; those exhibiting CNS involvement were also counted as mortalities.
494
J. GRUN
DISCUSSION
The data suggest that under the conditions outlined for these experiments, previously established mortality patterns can be reproduced. Essentially, the experiments were concerned with the response of groups of chickens to GB challenge after having undergone one of the following treatment regimens: no treatment; BPL-inactivated AA fluid in 0.5 ml. and 1 ml. doses; BPLinactivated vaccines composed of from 1 to 4 strains of NDV (GB, LaS, I Pa, Ho) in 0.5 ml. doses, with the exception of LaS alone (1 ml.). Although the first two NDV strains are familiar to most investigators, the latter two strains (I Pa and Ho) may
need some description. The I Pa strain given IN is similar in neurotropic and other properties to the GB strain, with the exception that the onset of symptoms is delayed for approximately 24 hours in birds less than 12 weeks of age with no parental immunity. Although the Ho isolate is similar to the I Pa strain, only 30 to 50% of the birds will show CNS involvement. CNS involvement is evident in 95 to 100% of the birds infected with the GB and I Pa strains. The LaSota strain very rarely, if ever, produces CNS involvement when given intranasally. The GB challenge was at times homologous, heterologous, or both, depending on the composition of the NDV vaccines. From our data, therefore, we can conclude that it is the neurotropic virus particles present in the vaccines which determine the antigenicity and thus the amount of protection to a virulent neurotropic NDV challenge for a period of at least 16 weeks. The mortality in the Ho-treated group at 4 weeks may have been the result of any number of circumstances, but the results with this vaccine indicate it has some merit for at least 16 weeks after vaccination. The La Sota strain evidently is not a good BPL-inactivated vaccine strain, alone or in conjunction with GB (Preparation No. 7). Whether this is due to its reducing the concentration of neurotropic particles or to a form of inhibition or interference cannot be determined by these data. The group treated with the GB strain alone also had some mortalities at 16 weeks after treatment; this may have been the consequence of an interaction between the ingredients and the homologous challenge virus. Those vaccines containing the greatest concentration of neurotropic particles tend to make the best BPL inactivated NDV vaccines. These vaccines were given in a single dose of 0.5 ml. or 1 ml. and were challenged with a large concentration of virulent GB
Downloaded from http://ps.oxfordjournals.org/ at Florida Atlantic University on November 7, 2014
in 0.5 ml. amounts and in which the I Pa strain was omitted). The group treated with preparation No. 6 had some mortalities on challenge at 4 weeks after treatment but none for the remainder of the 16 week period. Groups receiving No. 4 (GB) and No. 7 (GB and LaS) were protected for 12 weeks; at 16 weeks, however, they had mortalities of 7% and 25%, respectively. These latter preparations (No. 6, No. 4, and No. 7) were given at the 0.5 ml. dosage level. The magnitude of the mortalities, beginning at 4 weeks and continuing for 8, 12, and 16 weeks, would preclude preparation No 3 (LaS) from consideration as the only viral component of a BPL inactivated NDV vaccine. Further, the La Sota preparation was given in 1 ml. amounts and contained more virus protein (as shown by Kjeldahl analysis) than the 4-strain vaccine. The results with BPL inactivated AA fluid (No. 2) given in 0.5 ml. and 1 ml. inoculations were similar to the results described previously (Grun, 1966), except for a decrease in mortality in the 0.5 ml. AA fluid group at the 16 week challenge. It should again be noted that the ingredients of these vaccines can influence the course of the challenge infection.
NEWCASTLE DISEASE VIRUS
SUMMARY
In experiments involving six BPL inactivated NDV vaccines inoculated subcutaneously in the neck region and containing one or more strains, with characteristics ranging from non-neurotropic to almost completely so, it was found that those containing two or more strains with neurotropic tendencies (0.5 ml. dose) were the most effective during the 16 week experimental period. The GB strain vaccine may be better than can be concluded from the data; this may have been due to the homologous challenge virus and the influence of the vaccine ingredients. The La Sota strain alone, given in 1 ml. amounts, a dosage in which the vaccine ingredient influence tends to favor the host, is not sufficiently antigenic to be considered as the only virus component of a BPL inactivated NDV vaccine. It is evident that more work is needed in regard to the influence of the ingredients of an AA fluid NDV vaccine, the combina-
tion of strains used, and the effect of homologous vs. heterologous challenges. ACKNOWLEDGMENTS
The author is grateful for the technical assistance and the graphics of Miss E. DeBolle and for assistance with the manuscript from Dr. E. V. Adams. REFERENCES Appleton, G. S., S. B. Hitchner and R. W. Winterfield, 1963. A comparison of the immune response of chickens vaccinated with formalinand beta-propiolactone-inactivated Newcastle disease vaccines. Am. J. Vet. Res. 24: 827-831. Grun, J., 1966. Synergistic and antagonistic reactions with beta-propiolactone inactivated disease virus vaccine ingredients. Poultry Sci. 45: 126-131. Grun, J., and C. B. Hudson, 1966. Comparison of a beta-propiolactone whole embryo Newcastle disease vaccine (one strain) with a beta-propiolactone amnio-allantoic Newcastle disease vaccine (four strain). Poultry Sci. 4 5 : 159-164. Hofstad, M. S., J. C. Picken, Jr., K. E. Collins and H. W. Yoder, Jr., 1963. Immunogenicity of inactivated Newcastle disease virus preparations. Avian Dis. 7 : : 435-445. Kelly, A. R., 1952. Effects of virucide beta-propiolactone and its degradation products on blood platelets. Fed. Proc. 11: 419. Kelly, A. R., and F. W. Hartman, 1952. Biological effects of beta-propiolactone. Fed. Proc. 11: 419. Lo Grippo, G. A., and F. W. Hartman, 1954. Antigenicity of beta-propiolactone inactivated virus. Fed. Proc. 13 : 503. Lo Grippo, G. A., and F. W. Hartman, 1955. Antigenicity of |3-propiolactone inactivated virus vaccines. J. Immunol. 75: 123-128. Mack, W. N., and A. Chotisen, 1955. Beta-propiolactone as a virus altering agent for Newcastle disease vaccine. Poultry Sci. 34: 10101013. Sullivan, J. F., E. Gill and A. M. Somer, 1958. Immune responses of chickens to beta-propiolactone-killed Newcastle disease vaccines. Am. J. Vet. Res. 19: 483-488. Winmill, A. J., and W. Weddell. 1961. A Newcastle disease vaccine inactivated by betapropiolactone. Res. Vet. Sci. 2: 381-386.
Downloaded from http://ps.oxfordjournals.org/ at Florida Atlantic University on November 7, 2014
particles. They induced the strongest immunity up to the end of our experimental time (16 weeks). The GB strain alone may be better than the data reflects, since the homologous challenge virus may find the host in a specific hypersensitive condition and prone to synergism. Birds treated with vaccines containing two or more strains with neurotropic properties tend to survive the challenge best. All of the vaccines may be better than indicated by these data, since they were also subject to the synergistic reaction brought about by the vaccine ingredients (at the 0.5 ml. dosage level). This, of course, requires more experimentation in regard to the ingredient factors involved, the influence of these factors, and the magnitude of that influence with or without the presence of NDV particles.
495