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Immunization with inactivated poliovirus vaccine and attenuated type 3 poliovirus. II. Parallel vaccinations and cross-wise revaccination in school children with the Sabin and WM3 attenuated strains
August, 1969 T h e Journal o[ P E D I A T R I C S
273
Immunization ~vith inactivated poliovirus vaccine and attenuated type 3 poliovirus. II. Parallel vaccinations and cross-Mse reraccination in school children Mth the Sabin and WM3 attenuated strains Parallel vaccinations with the WM3 and Sabin type 3 poliovirus vaccine in a boarding school did not disclose any differences between the strains as regards their infectivity and antibody stimulating capacity. Cross-wise re[ceding, however, suggested that some antigenic variations might be present. The advantage o[ vaccination with several antigenieaUy varying type 3 attenuated strains appears to be worth [urther investigation.
Margareta BSttiger, M.D. S T O C K H O L M , SWEDEN
I N T H ~. first paper of this series of studies on vaccination with live type 3 vaccines, 1 the effect of vaccination and revaccination with the WM3 strain was investigated. In the present study comparative vaccinations with WM3 and Sabin type 3 were performed, followed by revaccination with the heterologous type 3 strain. The purpose of the study was : (1) to obtain preliminary information on the cross-wlse protection conferred by these two strains; (2) to study the antibody response after immunization with inactivated followed by live type 3 From the Department of Virology, The Karolinska Institute, and the National Bacteriological Laboratory.
poliovirus vaccines, and to obtain information on the duration of immunity; (3) to investigate the tendency of the vaccine virus to spread from person to person; and (4) to furnish material for marker studies on virus isolates. MATERIALS AND M E T H O D S Poliovlrus vaccines. vaccine. Immunization in 1957 were performed with vaccine manufactured by Eli Lilly Pharmaceuticals, U.S.A. For all subsequent immunizations, vacci!ae produced by the National BacteriologiCal Laboratory, Stockholm, was employed. 2 Live vaccine. The WM3 vaccine was Inactivated
Vol. 75, No. 2, pp. 273-28.0
274
Bi~tt{ger
prepared by the National Laboratory, Stockholm, from seed lot No. 17, supplied by Dr. Koprowski? The Sabin type 3 vaccine was purchased in 1966 from Pfizer Lab., England. The vaccine was given in a half glass of lemonade; each dose contained 4.5 to 5.0 log10 TCID~0. Tests for neutralizing antibodies and isolations of virus. These tests were performed as described in earlier publications?, 4 Plan of study. The vaccination trials with the attenuated type 3 poliovirus strains were performed in a boarding school for blind children. 5 The school had 135 children, of whom 120 were boarders; the rest were day pupils. Their ages ranged from 7 to 17 years. Previous vaccinations. Almost all children had already received 3 or 4 injections of 1 ml. of inactivated vaccine during the period between 1957 and the beginning of the current studies. For 48 of the children who had taken part in the earlier trial with type 1 vaccine, data on type 3 poliovirus antibody levels in January, 1960, were available. During the period from the spring of 1960 to the spring of 1965, the majority of the children who had received 3 injections in 195758 received a fourth injection--a few in 1960, the others in 1964. All children who had been given 2 injections in 1959 received a third in 1960 and a fourth in 1964. The rest, who were unvaccinated when they entered the trial in January, 1960, received 2 injections in February, 1960, and 1 or 2 doses in the spring of 1964. A number of new children with varying vaccination patterns entered the school during the period 1960 to 1965. The present study. The present trial started with blood sampling in February, 1965. Eight children who lacked demonstrable antibodies to type 3 poliovirus were given an injection of inactivated vaccine, after which they were retested and found positive. In the beginning of May, 1965, 30 children were given 5 log10 TCID~o of the WM3 vaccine; another 30 children were given the same dose of the Sabin type 3 vaccine. The vaccinees were evenly distributed among the different classes and dormitories.
The Journal of Pediatrics August 1969
Virus excretion was followed during the first month after feeding by investigation of fecal samples collected weekly from all inmates. Blood samples were taken again in September, 1965. In November, 1965, live virus vaccine was given again. A number of those fed with WM3 strain earlier were given the Sabin type 3 strain, and vice versa. The children who had not received live vaccine during the first phase of the study were now also included. Half of these children were fed the Sabin strain and the other half the WM3 strain. This time virus excretion was followed only in 2 fecal samples collected 1 and 2 weeks after the feeding. Blood samples were collected in February and in November, 1966. RESULTS Antibody status after vaccination with inactivated vaccines. The sera of 124 children were investigated for type 3 poliovirus antibodies in January, 1960. Their antibody levels in 1960 are correlated with their vaccination history in Table I. The levels of the younger age group at that time (born between 1948 and 1952) are represented in group 1, and the levels of the older children are shown in group 2. Among the unvaccinated, the proportion of seronegafives in 1960 was almost the same in the younger and older age groups; approximately two thirds lacked demonstrable antibodies at a serum dilution of 1 to 10. The children given 3 injections in 1957 and 1958 of a vaccine with comparatively poor immunogenicity (as estimated in a previous study 2) had generally lower titer levels than those who had received 2 injections of vaccine of better quality in the spring of 1959. At the beginning of the study of live type 3 vaccination in 1965, there were 3 unvaccinated seronegative children, 5 children who had received 2 injections, and 137 children who had received 3 or more injections of inactivated vaccine. Six of the 137 vaccinees were found to lack type 3 antibodies, even though 5 h a d been given 4 injections of inactivated vaccine. These 6 children all
Volume 75 Number 2
Poliovirus. I I
2 75
Table I. Distribution of antibody titers to type 3 poliovirus in January, 1960. Titers given as the reciprocal of highest neutralizing serum dilution. All vaccinations were performed during the spring months
Number o/children with antibody titer Vaccination < 1 0 [ 10 50 250 I 1,250 I 6,250 history Group 1, Unvaccinated 13 1 1 4 2 1 6 3 3 3 1948- 2 injections in 1959 3 1 3 6 5 I I952 3 injections in 1957-58 8 Year Of birth
Group 2, Unvaccinated 1942- 2 injections in 1959 1947 3 injections in 1957-58
10 1 9
Number of children with antibody titer < 10 • 10 13 8 3 16 8 16
1
3
1
10
5
6
6
1
3
2
1
1
7
3
6
0
9
18 17
Total number 21 19 24
15 19 26
Table II. Mean titers to type 3 poliovirus prior to and after feeding with the WM3 vaccine and the Sabin vaccine. Titers are given in logarithms and expressed as the reciprocal of the highest neutralizing serum dilution
Feeding date
May, 1965
WM3 vaccines Number Titer prior Titer alter Year o[ Feeding o[ children to [ceding feeding birth 5 months alter 18 2.6 2.9 1948-53First 12 2.6 2.9 1954-57 feeding
Nov., 1965
1948-53First 1954-57 feeding
17 15
2.4 2.5
2 months after 3.2 3.1
Nov., 1965
1948-57Refeeding
22
3.1
2.8
received their primary vaccination in 195758 when the vaccine of inferior antigenic quality was used. All children vaccinated since 1959 were found to be seropositive to type 3, whether they had received 3 or 4 injections. Antibody response to oral vaccination. The antibody titers of the individual children prior to and after feeding with the WM3 and Sabin type 3 strains have been tabulated on checkerboard figures (Figs. 1 and 2). Mean titers are recorded in Table II. Quadrangles 1 and 2 of Fig. 1 give the data from the children vaccinated for the first time in the spring of 1965 with the W M 3 strain and the Sabin type 3 strain, respectively. The antibody titers of these
Sabin vaecinees Number o[ Titer prior Titer alter children ~o feeding [eeding 5 months alter 17 2.7 3.1 13 2.8 3.1
17 12
2.6 3.0
2 months alter 3.1 3.3
22
2.9
2.8
children were measured 2 ~ months before and 5 months after the feeding. Quadrangles 3 and 4 of Fig. 1 give the titers 2 months before and 2 months after oral vaccination of the children who received live type 3 vaccine for the first time in November of 1965. The 4 groups were fairly equal in size, ranging from 27 to 34 children. The data in Fig. 1 and Table I I indicate that 2 months after feeding the mean antibody level was 3 times as high (0.5 logz0) as the prefeeding level. In the group tested 5 months after the feeding, the postfeeding level was 0.3 log10, or twice as high. C~Idren with low prefeeding titers responded with comparatively higher rises of tlter than those with higher prefeeding titers. No sta-
276
The Journal o[ Pediatrics August 1969
BSttiger
WM 3. May 1965
U
/
1
3]250
E .9 E 6250
Sabin 3. May 1965-
/ 9gO0 /
ooO009~/~ 9
9
1250
9 9149O ~ O0 0oo
.~. //.o .
250
.7=
5O
o ,
10
/
/
/ WM 3, Nov. 1965
C ,9
3
31250
o
iooele
/,
99
25o
~
4
oo|
9
/
o9 /
""'"~7"" 9 oo/Io / /
=o/ooo
/
~00_o /~
~,0/
10
Sabin, Nov. 1965
o9 /
~~176 -
02/
9
/
50
250 1250 6250 31250 10 50 prior to oral vaccination
250 1250 6250 31250
Fig. 1. Correlation between pre- and postfeeding antibody titers to type 3 poliovirus in children fed the WM3 vaccine or Sabin type 3 vaccine for the first time, i.e., in May or in November 1965. Filled circles represent virus excretors, open circles nonexcretors, and circles with a vertical line vaccinees whose feces were not tested.
3]250
5
/
/
6
oo;/
6250
~
~ 1250 ~
25O
~~ E
.
100 0
9
o
9
/..oOO~oo
V
50
10
50
/ 250
1250 6250 31250
10
/~ 50
250
1250 6250
,1250
prior to feeding Fig. 2. Correlation between pre- and postfeeding antibody titers to type 3 poliovirus after refeeding with live vaccine. T h e quadrangIe 5 illustrates findings in W M 3 vaccines refed with Sabin type 3 virus; quadrangle 6 illustrates the reverse sequence. Symbols as in Fig. 1.
Volume 75 Number 2
Poliovirus. H
tistical differences between the antibody responses after feeding with either the WM3 strain or the Sabin type 3 strain were found. The Sabin vaccinees by chance happened to have slightly higher prefeeding titers than the WM3 vaccinees. There was no marked difference between the response of the younger and the older children. The antibody response to a second feeding with live virus 6 months after the first, but with the other type 3 strain, is shown in Fig. 2. Titers were measured 2 months prior to and 2 months after the feeding. The second feeding appeared to have no antibody stimulating effect, even in those with the lower prefeeding antibody titers. Virus excretion. The results of the virus isolation tests on stool samples collected weekly after feeding are given in Table I I I . Forty-five of 62 WM3 vaccinees (73 per cent) and 44 of 57 Sabin type 3 vaccinees (77 per cent) were found to excrete virus. With both vaccines, the excretion rate was higher in the younger age groups. Forty-five of 50 children (90 per cent) born between 1954 and 1958 were found to be virus excretors; among those born between 1948 and 1953, 44 of 67 (66 per cent) excreted
277
virus. Since circulating wild type 3 poliovirus has been found only rarely since 1953 in Sweden, few of the children born after 1954 had presumably been exposed to natural type 3 infection. Sixteen children who were naturally nonimmune in 1960, i.e., seronegative prior to parenteral vaccination, were available. Fourteen of these children were found to excrete virus. In another group of 8 children presumably naturally immune in 1960, 2 excreted virus after the first feeding. The children fed live vaccine a second time had an excretion rate of 21 per cent (bottom of Table I I I ) . Seven of 16 vaccinees in the younger, and 2 of 24 vaccinees in the older age group were found to excrete virus. Furthermore, 7 of 22 subjects vaccinated earlier with the WM3 strain and who were refed the Sabin type 3 strain became excretors, while 2 of 20 belonging to the group given Sabin strain first and the WM3 strain later were found to excrete virus. Eight of the 9 vaccinees excreting virus after the second challenge also had virus-positive stools after the first feeding. Of the other 33 found negative following the second feeding, 24 had been virus positive and 9 virus
Table I l L The proportion of vaccinees excreting virus in the feces
Age group 1954-58 1954-58 1948-53 1948-53
Date of [eedlng Number o[ Numbertested o[ samples [ (1965) children tested from each May 12 4 Nov. 15 2 May 18 4 Nov. 17 2
Vaccine WM3 WM3 WM3 WM3 Total excretion rate
1954-58 1954-58 1948-53 1948-53
May Nov. May Nov.
13 10 17 17
4 2 4 2
Sabln 3 Sabln 3 Sabin 3 Sabin 3 Total excretion rate
1954-57 1948-53 1954-57 1948-53
Nov.
8 12 8 14
2 2 2 2
WM3 WM3 Sabin 3 Sabin 3 Total excretion rate
Nov.
Nov. Nov.
Number of children tested with Positive Negative isolation . isolation 11 I 13 2 13 5 8 9
45/62 12 8 9 15
1 2 8 2
44/57 2 5 2 9/42
6 12 3 12
2'7 8
BSttiger
The Journal of Pediatrics August 1969
Table IV. Excretion pattern of children fed live vaccine in May, 1965, and followed by weekly investigations of fecal specimens for 4 weeks
Vaccine WM3 Sabin type 3
Non- . exeretors 4 9
Number o[ children excreting virus for given period 1 week [ 2 weeks I 3 weeks I ~ 4 weeks 5 4 3 11 6 4 2 7
negative after the first feeding. All vaccinees found resistant (nonexcretors) to both feedings belonged to the older age group born between 1948 and 1953. T h e proportion of these nonexcretors (27 per cent) was almost the same as the proportion of naturally immune children in this age group (33 per cent) as observed in the earlier investigation in 1960. 5, 6 After the May vaccination, the excretion of virus was followed for 4 weeks (Table I V ) . No major differences between the two vaccine strains were observed. Spread of virus. After the first oral vaccination in May, 1965, when approximately half of the children in the study were fed live vaccine, 4 first grade children belonging to the 61 unvaccinated controls were found to excrete virus. Forty-four of these unvaccinated controls could be considered susceptible to the virus infection as judged by the results of the challenge with live virus performed later. The virus isolated from the 4 contactinfected children were tested against immune sera prepared against the WM3 or Sabin type 3 strain. All 4 isolates resembled the WM3 strain in this test. Three of them also formed plaques resembling those formed by the WM3 strain (fairly small with uneven outlines) while one of them formed larger and more distinct plaques.
DISCUSSION Since the introduction of poliovirus vaccination in Sweden in 1957, the main interest has been study of the immunity to poliovirus type 1 as this type was the dominant cause of disease. Moreover, in the early inactivated vaccines the type 1 component also was the least immunogenic.
Total number 27 28
During recent years more attention has been directed toward type 3, especially with regard to the live attenuated vaccine? The type 3 antigen of the inactivated vaccine produced in Sweden has since 1959 induced antibodies in all vaccinees tested, with mean titers of about 1 to 6,250. 2 However, several follow-up studies 2, 7. s indicate that the type 3 antibody level might be less persistent than that of the other types.
Immunity after parenteral vaccination. The present study stresses the importance of an initially strong response to immunization with inactivated vaccine. Thus, those chiL dren who took part in the vaccination carried out in 1957 and 1958 with a less potent vaccine 2 than used later seemed to retain lower antibody levels in spite of the fact that they later received a third and even a fourth injection with a more potent vaccine. I n contrast, all children who were given their primary vaccination in 1959 or later had demonstrable antibody titers whether they had received 3 or 4 doses.
Antibody response to oral vaccination. As was experienced with type 1 oral vaccination, vaccinees with low prefeeding titers reacted with relatively higher rises of titers to the live antigenic stimulus than did those with higher prefeeding titers. The general mean level reached (3.1 log~o) agrees well with the titers observed in school-age children in the first study 1 in this series. The second feeding a half year later with the heterologous type 3 strain had apparently little or no antibody stimulating effect. It must be noted, however, that the mean antibody level prior to refeeding was relatively high, approximately 3 log~0. According to the earlier vaccination trials, this level corresponds to the maximum attainable by oral
Volume 75 Number 2
vaccination in this age group, and thus no further rises might be expected. Vonka and co-workers9 obtained considerable titer rises after refeeding with the U S O L - D bac strain in children vaccinated previously with the Sabin strain. The two studies cannot be directly compared, however, as the prefeeding titers of the children studied by Vonka and co-workers were approximately between 1 and 2 log~0 lower than those of the children in this study. However, after the refeeding with the U S O L - D bac strain, mean titer values roughly estimated to be 2.8 loga0 were obtained; these agree fairly well with the levels obtained in this study after the combined vaccinations with inactivated and live vaccines. Virus exeertlon. Ninety per cent of the younger and 66 per cent of the older children were found to excrete virus after the first feeding in the present study. The corresponding figures with the type 1 C H A T strain were 89 and 61 per cent, respectively. When comparing the data from all those fed oral type 3 vaccine for the first time, there was no difference in excretion ratios between the two type 3 strains. It appears likely that part of the resistance to virus infection was due to previously induced immunity by natural infection. However, such immunity did not induce a 100 per cent resistance to infection with attenuated poliovirus. The group of 22 WM3 vaccinees refed with the Sabin type 3 strain had 7 excretors, while the group vaccinated in reverse sequence had 2 excretors among 20 revaceinated. Prefeeding antibody titers differed very little in the two groups. I n the earlier study in which W M 3 vaecinees were refed after 3 months with the same vaccine, 3 of 21 children were found to be excretors. No significant differences between the strains were thus disclosed in these 3 trials with refeedings. T h e observation might be made, however, that prevaccination with the Sabin type 3 strain seemed to prevent virus excretion after challenge with WM3 vaccine as effectively as prevaccination with the homologous vac-
Poliovirus. I I
2 79
cine strain. If these two groups refed with the WM3 strain (i.e., the group prevaccinated with Sabin type 3 and presented in this paper and the W M 3 vaccinees from the earlier study) are analyzed together, there is a suggestively lower excretion rate after the second feeding than in the group prevaccinated with WM3 and refed with Sabin type 3 (significance level approximately 95 per cent). Spread of virus. Although a massive infection with virus was initiated, the spread of virus observed was limited, especially considering the blind children's handicap that favors virus transmission through contact. The observation that the virus is spread mainly by the youngest children is in agreement with earlier experiences? ~ General comment. The present study on 135 children did not reveal any major differences between the two type 3 strains. Both appeared acceptable as immunizing agents. Although the WM3 strain is a "cold mutant," i.e., it had been adapted to growth at low temperature, its infectivity did not appear to be inferior to that of the Sabin type 3 strain. Neither was it inferior from an antibody-stimulating point of view. Both strains seemed to possess a fairly limited tendency to spread to contacts of school age. Parallel testings for neurovirulence of the two type 3 vaccine strains have been carried out by a number of laboratories; the results summarized by Pagano and Hoskins al did not reveal any major differences between the two strains. Whether the WM3 strain might be more suitable than the Sabin type 3 strain from a safety point of view can, however, only be determined after mass vaccination. The WM3 strain has been used for more than 1.2 million people, mainly children in Switzerland and Yugoslaviaa2,~3; no vaccine-associated paralytic cases have been reported. SUMMARY
Parallel vaccinations with the live WM3 and the Sabin type 3 poliovirus vaccines were performed in a boarding school for
2 80
B6ttiger
blind children. All 135 children p a r t i c i p a t i n g were p r e i m m u n i z e d parenterally. Doses of the two live vaccines containing a p p r o x imately the same a m o u n t of virus (5 log10 T C I D s 0 ) p r o d u c e d the same p r o p o r t i o n of excretors a n d h a d a similar a n t i b o d y stimulating effect. Excretion rates were higher a m o n g the younger (90 p e r cent) t h a n a m o n g the older (66 p e r cent) children. W h e n 60 of the 135 children were given live virus, 4 u n v a c c i n a t e d inmates in close contact were f o u n d to excrete virus; transmission of virus was observed in the first g r a d e only. Children given W M 3 vaccine earlier a n d refed with the Sabin type 3 strain m a y have a higher rate of excretion after the second vaccination t h a n either Sabin type 3 or W M 3 vaccinees refed with the W M 3 strain. E x t e n d e d trials will have to be p e r f o r m e d to evaluate these observations. T h e a d v a n tage of i m m u n i z a t i o n with several antigenically different type 3 a t t e n u a t e d strains appears to be w o r t h f u r t h e r investigation. REFERENCES
1. BSttlger, M., and Lagercrantz, R.: Immunization with inactivated poliovirus vaccine and attenuated type 3 pollovirus. I. Vaccination with the WM3 strain in 20 families, J. PEmAT. 75: 30, 1969. 2. Btittiger, M., Arro, L., Lundbeck, H., and Salenstedt, C. -R.: The immune response to vaccination with inactivated poliovlrus vaccine in Sweden, Acta path. et microbiol. scandinav. 66: 239, 1966. 3. Plotkin, S. A., Norton, T. W., Cohen, B. J., and Koprowski, H.: A type 3 attenuated poliovirus genetically stable after human intestinal passage, Proe. Soe. Exper. Biol. & Med. 107: 829, 1961. 4. B6ttlger, M., Gard, S., and Lagercrantz, R.: Vaccination with attenuated type 1 poliovirus, the CHAT strain. I. A study in 20 families, Acta paedlat, scandinav. 55: 405, 1966.
The Journal o/ Pediatrics August 1969
5. B6ttiger, M., B6ttiger, 2., and Zetterberg, B.: Vaccination with attenuated type 1 poliovirus, the CHAT strain. III. Antibody response and spread of virus in school children, Acta paediat, scandinav. 55: 422, 1966. 6. Olin, G., and Wesslen, T.: Seroimmune patterns for poliomyelitis in Sweden, Arch. ges. Virusforsch. 7: 191, 1957. 7. Zacek, K., Adam, E., Adamova, V., Burian, V., Rozacova, P., Skridlowska, E., Vanerkova, N., Vonka, V., and Zackova, Z.: Mass oral (Sabin) poliomyelitis vaccination. Virological and serological surveillance in Czechoslovakia 1958-59 and 1960-61, VIIIth Syrup. Europ. Ass. poliomyelitis and allied diseases, Prague, 1962, p. 219. 8. Buser, F., Fleury, C., and Rohner, F.: Status of immunity 5 years after the oral vaccination against poliomyelitis, IXth Syrup. Europ. Ass. poliomyelitis and allied diseases, Stockholm, 1963, p. 129. 9. Vonka, V., Janda, Z., Simon, J., Adam, E., Zavadova, H., and Starek, M.: Experiences with the new type 3 attenuated poliovirus USOL-D bac developed in Czechoslovakia, Xth Symp. Europ. Ass. poliomyelitis and allied diseases, Warsaw, 1964, p. 413. 10. BSttiger, M., Gard, S., and Zetterberg, B.: Vaccination with attenuated type 1 poliovirus, the CHAT strain. II. Transmission of virus in relation to age, Acta paediat, seandinav. 55: 416, 1966. 11. Pagano, J. S., and Hoskins, J. iV[.: Variations in the assessment of the comparative neurovirulence of type 3 attenuated poliovirus vaccines, Conference on testing for neurovirulence of virus vaccines, Permanent section of microbiological standardization, Munchen, 1965, p. 221. 12. Ikie, D., and Janikic, B.: Epldemiologlcal data on efficacy of live oral pollovaccine Koprowsky, Proceedings of symposium on the characterization and uses of human diploid cell strains, Opatija, 1963, p. 541. 13. Buser, F., Fleury, C., Martin du Pan, R., and Rohner, E.: Immunization with live attenuated poliovirus prepared in human diploid cell strains, with special reference to the WM3 strain. Proceedings of symposium on characterization and uses of human diploid cell strains, Opatija, 1963, p. 381.
273
Immunization ~vith inactivated poliovirus vaccine and attenuated type 3 poliovirus. II. Parallel vaccinations and cross-Mse reraccination in school children Mth the Sabin and WM3 attenuated strains Parallel vaccinations with the WM3 and Sabin type 3 poliovirus vaccine in a boarding school did not disclose any differences between the strains as regards their infectivity and antibody stimulating capacity. Cross-wise re[ceding, however, suggested that some antigenic variations might be present. The advantage o[ vaccination with several antigenieaUy varying type 3 attenuated strains appears to be worth [urther investigation.
Margareta BSttiger, M.D. S T O C K H O L M , SWEDEN
I N T H ~. first paper of this series of studies on vaccination with live type 3 vaccines, 1 the effect of vaccination and revaccination with the WM3 strain was investigated. In the present study comparative vaccinations with WM3 and Sabin type 3 were performed, followed by revaccination with the heterologous type 3 strain. The purpose of the study was : (1) to obtain preliminary information on the cross-wlse protection conferred by these two strains; (2) to study the antibody response after immunization with inactivated followed by live type 3 From the Department of Virology, The Karolinska Institute, and the National Bacteriological Laboratory.
poliovirus vaccines, and to obtain information on the duration of immunity; (3) to investigate the tendency of the vaccine virus to spread from person to person; and (4) to furnish material for marker studies on virus isolates. MATERIALS AND M E T H O D S Poliovlrus vaccines. vaccine. Immunization in 1957 were performed with vaccine manufactured by Eli Lilly Pharmaceuticals, U.S.A. For all subsequent immunizations, vacci!ae produced by the National BacteriologiCal Laboratory, Stockholm, was employed. 2 Live vaccine. The WM3 vaccine was Inactivated
Vol. 75, No. 2, pp. 273-28.0
274
Bi~tt{ger
prepared by the National Laboratory, Stockholm, from seed lot No. 17, supplied by Dr. Koprowski? The Sabin type 3 vaccine was purchased in 1966 from Pfizer Lab., England. The vaccine was given in a half glass of lemonade; each dose contained 4.5 to 5.0 log10 TCID~0. Tests for neutralizing antibodies and isolations of virus. These tests were performed as described in earlier publications?, 4 Plan of study. The vaccination trials with the attenuated type 3 poliovirus strains were performed in a boarding school for blind children. 5 The school had 135 children, of whom 120 were boarders; the rest were day pupils. Their ages ranged from 7 to 17 years. Previous vaccinations. Almost all children had already received 3 or 4 injections of 1 ml. of inactivated vaccine during the period between 1957 and the beginning of the current studies. For 48 of the children who had taken part in the earlier trial with type 1 vaccine, data on type 3 poliovirus antibody levels in January, 1960, were available. During the period from the spring of 1960 to the spring of 1965, the majority of the children who had received 3 injections in 195758 received a fourth injection--a few in 1960, the others in 1964. All children who had been given 2 injections in 1959 received a third in 1960 and a fourth in 1964. The rest, who were unvaccinated when they entered the trial in January, 1960, received 2 injections in February, 1960, and 1 or 2 doses in the spring of 1964. A number of new children with varying vaccination patterns entered the school during the period 1960 to 1965. The present study. The present trial started with blood sampling in February, 1965. Eight children who lacked demonstrable antibodies to type 3 poliovirus were given an injection of inactivated vaccine, after which they were retested and found positive. In the beginning of May, 1965, 30 children were given 5 log10 TCID~o of the WM3 vaccine; another 30 children were given the same dose of the Sabin type 3 vaccine. The vaccinees were evenly distributed among the different classes and dormitories.
The Journal of Pediatrics August 1969
Virus excretion was followed during the first month after feeding by investigation of fecal samples collected weekly from all inmates. Blood samples were taken again in September, 1965. In November, 1965, live virus vaccine was given again. A number of those fed with WM3 strain earlier were given the Sabin type 3 strain, and vice versa. The children who had not received live vaccine during the first phase of the study were now also included. Half of these children were fed the Sabin strain and the other half the WM3 strain. This time virus excretion was followed only in 2 fecal samples collected 1 and 2 weeks after the feeding. Blood samples were collected in February and in November, 1966. RESULTS Antibody status after vaccination with inactivated vaccines. The sera of 124 children were investigated for type 3 poliovirus antibodies in January, 1960. Their antibody levels in 1960 are correlated with their vaccination history in Table I. The levels of the younger age group at that time (born between 1948 and 1952) are represented in group 1, and the levels of the older children are shown in group 2. Among the unvaccinated, the proportion of seronegafives in 1960 was almost the same in the younger and older age groups; approximately two thirds lacked demonstrable antibodies at a serum dilution of 1 to 10. The children given 3 injections in 1957 and 1958 of a vaccine with comparatively poor immunogenicity (as estimated in a previous study 2) had generally lower titer levels than those who had received 2 injections of vaccine of better quality in the spring of 1959. At the beginning of the study of live type 3 vaccination in 1965, there were 3 unvaccinated seronegative children, 5 children who had received 2 injections, and 137 children who had received 3 or more injections of inactivated vaccine. Six of the 137 vaccinees were found to lack type 3 antibodies, even though 5 h a d been given 4 injections of inactivated vaccine. These 6 children all
Volume 75 Number 2
Poliovirus. I I
2 75
Table I. Distribution of antibody titers to type 3 poliovirus in January, 1960. Titers given as the reciprocal of highest neutralizing serum dilution. All vaccinations were performed during the spring months
Number o/children with antibody titer Vaccination < 1 0 [ 10 50 250 I 1,250 I 6,250 history Group 1, Unvaccinated 13 1 1 4 2 1 6 3 3 3 1948- 2 injections in 1959 3 1 3 6 5 I I952 3 injections in 1957-58 8 Year Of birth
Group 2, Unvaccinated 1942- 2 injections in 1959 1947 3 injections in 1957-58
10 1 9
Number of children with antibody titer < 10 • 10 13 8 3 16 8 16
1
3
1
10
5
6
6
1
3
2
1
1
7
3
6
0
9
18 17
Total number 21 19 24
15 19 26
Table II. Mean titers to type 3 poliovirus prior to and after feeding with the WM3 vaccine and the Sabin vaccine. Titers are given in logarithms and expressed as the reciprocal of the highest neutralizing serum dilution
Feeding date
May, 1965
WM3 vaccines Number Titer prior Titer alter Year o[ Feeding o[ children to [ceding feeding birth 5 months alter 18 2.6 2.9 1948-53First 12 2.6 2.9 1954-57 feeding
Nov., 1965
1948-53First 1954-57 feeding
17 15
2.4 2.5
2 months after 3.2 3.1
Nov., 1965
1948-57Refeeding
22
3.1
2.8
received their primary vaccination in 195758 when the vaccine of inferior antigenic quality was used. All children vaccinated since 1959 were found to be seropositive to type 3, whether they had received 3 or 4 injections. Antibody response to oral vaccination. The antibody titers of the individual children prior to and after feeding with the WM3 and Sabin type 3 strains have been tabulated on checkerboard figures (Figs. 1 and 2). Mean titers are recorded in Table II. Quadrangles 1 and 2 of Fig. 1 give the data from the children vaccinated for the first time in the spring of 1965 with the W M 3 strain and the Sabin type 3 strain, respectively. The antibody titers of these
Sabin vaecinees Number o[ Titer prior Titer alter children ~o feeding [eeding 5 months alter 17 2.7 3.1 13 2.8 3.1
17 12
2.6 3.0
2 months alter 3.1 3.3
22
2.9
2.8
children were measured 2 ~ months before and 5 months after the feeding. Quadrangles 3 and 4 of Fig. 1 give the titers 2 months before and 2 months after oral vaccination of the children who received live type 3 vaccine for the first time in November of 1965. The 4 groups were fairly equal in size, ranging from 27 to 34 children. The data in Fig. 1 and Table I I indicate that 2 months after feeding the mean antibody level was 3 times as high (0.5 logz0) as the prefeeding level. In the group tested 5 months after the feeding, the postfeeding level was 0.3 log10, or twice as high. C~Idren with low prefeeding titers responded with comparatively higher rises of tlter than those with higher prefeeding titers. No sta-
276
The Journal o[ Pediatrics August 1969
BSttiger
WM 3. May 1965
U
/
1
3]250
E .9 E 6250
Sabin 3. May 1965-
/ 9gO0 /
ooO009~/~ 9
9
1250
9 9149O ~ O0 0oo
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250
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o ,
10
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/
/ WM 3, Nov. 1965
C ,9
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31250
o
iooele
/,
99
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9
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~,0/
10
Sabin, Nov. 1965
o9 /
~~176 -
02/
9
/
50
250 1250 6250 31250 10 50 prior to oral vaccination
250 1250 6250 31250
Fig. 1. Correlation between pre- and postfeeding antibody titers to type 3 poliovirus in children fed the WM3 vaccine or Sabin type 3 vaccine for the first time, i.e., in May or in November 1965. Filled circles represent virus excretors, open circles nonexcretors, and circles with a vertical line vaccinees whose feces were not tested.
3]250
5
/
/
6
oo;/
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~
~ 1250 ~
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.
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9
o
9
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50
10
50
/ 250
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10
/~ 50
250
1250 6250
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prior to feeding Fig. 2. Correlation between pre- and postfeeding antibody titers to type 3 poliovirus after refeeding with live vaccine. T h e quadrangIe 5 illustrates findings in W M 3 vaccines refed with Sabin type 3 virus; quadrangle 6 illustrates the reverse sequence. Symbols as in Fig. 1.
Volume 75 Number 2
Poliovirus. H
tistical differences between the antibody responses after feeding with either the WM3 strain or the Sabin type 3 strain were found. The Sabin vaccinees by chance happened to have slightly higher prefeeding titers than the WM3 vaccinees. There was no marked difference between the response of the younger and the older children. The antibody response to a second feeding with live virus 6 months after the first, but with the other type 3 strain, is shown in Fig. 2. Titers were measured 2 months prior to and 2 months after the feeding. The second feeding appeared to have no antibody stimulating effect, even in those with the lower prefeeding antibody titers. Virus excretion. The results of the virus isolation tests on stool samples collected weekly after feeding are given in Table I I I . Forty-five of 62 WM3 vaccinees (73 per cent) and 44 of 57 Sabin type 3 vaccinees (77 per cent) were found to excrete virus. With both vaccines, the excretion rate was higher in the younger age groups. Forty-five of 50 children (90 per cent) born between 1954 and 1958 were found to be virus excretors; among those born between 1948 and 1953, 44 of 67 (66 per cent) excreted
277
virus. Since circulating wild type 3 poliovirus has been found only rarely since 1953 in Sweden, few of the children born after 1954 had presumably been exposed to natural type 3 infection. Sixteen children who were naturally nonimmune in 1960, i.e., seronegative prior to parenteral vaccination, were available. Fourteen of these children were found to excrete virus. In another group of 8 children presumably naturally immune in 1960, 2 excreted virus after the first feeding. The children fed live vaccine a second time had an excretion rate of 21 per cent (bottom of Table I I I ) . Seven of 16 vaccinees in the younger, and 2 of 24 vaccinees in the older age group were found to excrete virus. Furthermore, 7 of 22 subjects vaccinated earlier with the WM3 strain and who were refed the Sabin type 3 strain became excretors, while 2 of 20 belonging to the group given Sabin strain first and the WM3 strain later were found to excrete virus. Eight of the 9 vaccinees excreting virus after the second challenge also had virus-positive stools after the first feeding. Of the other 33 found negative following the second feeding, 24 had been virus positive and 9 virus
Table I l L The proportion of vaccinees excreting virus in the feces
Age group 1954-58 1954-58 1948-53 1948-53
Date of [eedlng Number o[ Numbertested o[ samples [ (1965) children tested from each May 12 4 Nov. 15 2 May 18 4 Nov. 17 2
Vaccine WM3 WM3 WM3 WM3 Total excretion rate
1954-58 1954-58 1948-53 1948-53
May Nov. May Nov.
13 10 17 17
4 2 4 2
Sabln 3 Sabln 3 Sabin 3 Sabin 3 Total excretion rate
1954-57 1948-53 1954-57 1948-53
Nov.
8 12 8 14
2 2 2 2
WM3 WM3 Sabin 3 Sabin 3 Total excretion rate
Nov.
Nov. Nov.
Number of children tested with Positive Negative isolation . isolation 11 I 13 2 13 5 8 9
45/62 12 8 9 15
1 2 8 2
44/57 2 5 2 9/42
6 12 3 12
2'7 8
BSttiger
The Journal of Pediatrics August 1969
Table IV. Excretion pattern of children fed live vaccine in May, 1965, and followed by weekly investigations of fecal specimens for 4 weeks
Vaccine WM3 Sabin type 3
Non- . exeretors 4 9
Number o[ children excreting virus for given period 1 week [ 2 weeks I 3 weeks I ~ 4 weeks 5 4 3 11 6 4 2 7
negative after the first feeding. All vaccinees found resistant (nonexcretors) to both feedings belonged to the older age group born between 1948 and 1953. T h e proportion of these nonexcretors (27 per cent) was almost the same as the proportion of naturally immune children in this age group (33 per cent) as observed in the earlier investigation in 1960. 5, 6 After the May vaccination, the excretion of virus was followed for 4 weeks (Table I V ) . No major differences between the two vaccine strains were observed. Spread of virus. After the first oral vaccination in May, 1965, when approximately half of the children in the study were fed live vaccine, 4 first grade children belonging to the 61 unvaccinated controls were found to excrete virus. Forty-four of these unvaccinated controls could be considered susceptible to the virus infection as judged by the results of the challenge with live virus performed later. The virus isolated from the 4 contactinfected children were tested against immune sera prepared against the WM3 or Sabin type 3 strain. All 4 isolates resembled the WM3 strain in this test. Three of them also formed plaques resembling those formed by the WM3 strain (fairly small with uneven outlines) while one of them formed larger and more distinct plaques.
DISCUSSION Since the introduction of poliovirus vaccination in Sweden in 1957, the main interest has been study of the immunity to poliovirus type 1 as this type was the dominant cause of disease. Moreover, in the early inactivated vaccines the type 1 component also was the least immunogenic.
Total number 27 28
During recent years more attention has been directed toward type 3, especially with regard to the live attenuated vaccine? The type 3 antigen of the inactivated vaccine produced in Sweden has since 1959 induced antibodies in all vaccinees tested, with mean titers of about 1 to 6,250. 2 However, several follow-up studies 2, 7. s indicate that the type 3 antibody level might be less persistent than that of the other types.
Immunity after parenteral vaccination. The present study stresses the importance of an initially strong response to immunization with inactivated vaccine. Thus, those chiL dren who took part in the vaccination carried out in 1957 and 1958 with a less potent vaccine 2 than used later seemed to retain lower antibody levels in spite of the fact that they later received a third and even a fourth injection with a more potent vaccine. I n contrast, all children who were given their primary vaccination in 1959 or later had demonstrable antibody titers whether they had received 3 or 4 doses.
Antibody response to oral vaccination. As was experienced with type 1 oral vaccination, vaccinees with low prefeeding titers reacted with relatively higher rises of titers to the live antigenic stimulus than did those with higher prefeeding titers. The general mean level reached (3.1 log~o) agrees well with the titers observed in school-age children in the first study 1 in this series. The second feeding a half year later with the heterologous type 3 strain had apparently little or no antibody stimulating effect. It must be noted, however, that the mean antibody level prior to refeeding was relatively high, approximately 3 log~0. According to the earlier vaccination trials, this level corresponds to the maximum attainable by oral
Volume 75 Number 2
vaccination in this age group, and thus no further rises might be expected. Vonka and co-workers9 obtained considerable titer rises after refeeding with the U S O L - D bac strain in children vaccinated previously with the Sabin strain. The two studies cannot be directly compared, however, as the prefeeding titers of the children studied by Vonka and co-workers were approximately between 1 and 2 log~0 lower than those of the children in this study. However, after the refeeding with the U S O L - D bac strain, mean titer values roughly estimated to be 2.8 loga0 were obtained; these agree fairly well with the levels obtained in this study after the combined vaccinations with inactivated and live vaccines. Virus exeertlon. Ninety per cent of the younger and 66 per cent of the older children were found to excrete virus after the first feeding in the present study. The corresponding figures with the type 1 C H A T strain were 89 and 61 per cent, respectively. When comparing the data from all those fed oral type 3 vaccine for the first time, there was no difference in excretion ratios between the two type 3 strains. It appears likely that part of the resistance to virus infection was due to previously induced immunity by natural infection. However, such immunity did not induce a 100 per cent resistance to infection with attenuated poliovirus. The group of 22 WM3 vaccinees refed with the Sabin type 3 strain had 7 excretors, while the group vaccinated in reverse sequence had 2 excretors among 20 revaceinated. Prefeeding antibody titers differed very little in the two groups. I n the earlier study in which W M 3 vaecinees were refed after 3 months with the same vaccine, 3 of 21 children were found to be excretors. No significant differences between the strains were thus disclosed in these 3 trials with refeedings. T h e observation might be made, however, that prevaccination with the Sabin type 3 strain seemed to prevent virus excretion after challenge with WM3 vaccine as effectively as prevaccination with the homologous vac-
Poliovirus. I I
2 79
cine strain. If these two groups refed with the WM3 strain (i.e., the group prevaccinated with Sabin type 3 and presented in this paper and the W M 3 vaccinees from the earlier study) are analyzed together, there is a suggestively lower excretion rate after the second feeding than in the group prevaccinated with WM3 and refed with Sabin type 3 (significance level approximately 95 per cent). Spread of virus. Although a massive infection with virus was initiated, the spread of virus observed was limited, especially considering the blind children's handicap that favors virus transmission through contact. The observation that the virus is spread mainly by the youngest children is in agreement with earlier experiences? ~ General comment. The present study on 135 children did not reveal any major differences between the two type 3 strains. Both appeared acceptable as immunizing agents. Although the WM3 strain is a "cold mutant," i.e., it had been adapted to growth at low temperature, its infectivity did not appear to be inferior to that of the Sabin type 3 strain. Neither was it inferior from an antibody-stimulating point of view. Both strains seemed to possess a fairly limited tendency to spread to contacts of school age. Parallel testings for neurovirulence of the two type 3 vaccine strains have been carried out by a number of laboratories; the results summarized by Pagano and Hoskins al did not reveal any major differences between the two strains. Whether the WM3 strain might be more suitable than the Sabin type 3 strain from a safety point of view can, however, only be determined after mass vaccination. The WM3 strain has been used for more than 1.2 million people, mainly children in Switzerland and Yugoslaviaa2,~3; no vaccine-associated paralytic cases have been reported. SUMMARY
Parallel vaccinations with the live WM3 and the Sabin type 3 poliovirus vaccines were performed in a boarding school for
2 80
B6ttiger
blind children. All 135 children p a r t i c i p a t i n g were p r e i m m u n i z e d parenterally. Doses of the two live vaccines containing a p p r o x imately the same a m o u n t of virus (5 log10 T C I D s 0 ) p r o d u c e d the same p r o p o r t i o n of excretors a n d h a d a similar a n t i b o d y stimulating effect. Excretion rates were higher a m o n g the younger (90 p e r cent) t h a n a m o n g the older (66 p e r cent) children. W h e n 60 of the 135 children were given live virus, 4 u n v a c c i n a t e d inmates in close contact were f o u n d to excrete virus; transmission of virus was observed in the first g r a d e only. Children given W M 3 vaccine earlier a n d refed with the Sabin type 3 strain m a y have a higher rate of excretion after the second vaccination t h a n either Sabin type 3 or W M 3 vaccinees refed with the W M 3 strain. E x t e n d e d trials will have to be p e r f o r m e d to evaluate these observations. T h e a d v a n tage of i m m u n i z a t i o n with several antigenically different type 3 a t t e n u a t e d strains appears to be w o r t h f u r t h e r investigation. REFERENCES
1. BSttlger, M., and Lagercrantz, R.: Immunization with inactivated poliovirus vaccine and attenuated type 3 pollovirus. I. Vaccination with the WM3 strain in 20 families, J. PEmAT. 75: 30, 1969. 2. Btittiger, M., Arro, L., Lundbeck, H., and Salenstedt, C. -R.: The immune response to vaccination with inactivated poliovlrus vaccine in Sweden, Acta path. et microbiol. scandinav. 66: 239, 1966. 3. Plotkin, S. A., Norton, T. W., Cohen, B. J., and Koprowski, H.: A type 3 attenuated poliovirus genetically stable after human intestinal passage, Proe. Soe. Exper. Biol. & Med. 107: 829, 1961. 4. B6ttlger, M., Gard, S., and Lagercrantz, R.: Vaccination with attenuated type 1 poliovirus, the CHAT strain. I. A study in 20 families, Acta paedlat, scandinav. 55: 405, 1966.
The Journal o/ Pediatrics August 1969
5. B6ttiger, M., B6ttiger, 2., and Zetterberg, B.: Vaccination with attenuated type 1 poliovirus, the CHAT strain. III. Antibody response and spread of virus in school children, Acta paediat, scandinav. 55: 422, 1966. 6. Olin, G., and Wesslen, T.: Seroimmune patterns for poliomyelitis in Sweden, Arch. ges. Virusforsch. 7: 191, 1957. 7. Zacek, K., Adam, E., Adamova, V., Burian, V., Rozacova, P., Skridlowska, E., Vanerkova, N., Vonka, V., and Zackova, Z.: Mass oral (Sabin) poliomyelitis vaccination. Virological and serological surveillance in Czechoslovakia 1958-59 and 1960-61, VIIIth Syrup. Europ. Ass. poliomyelitis and allied diseases, Prague, 1962, p. 219. 8. Buser, F., Fleury, C., and Rohner, F.: Status of immunity 5 years after the oral vaccination against poliomyelitis, IXth Syrup. Europ. Ass. poliomyelitis and allied diseases, Stockholm, 1963, p. 129. 9. Vonka, V., Janda, Z., Simon, J., Adam, E., Zavadova, H., and Starek, M.: Experiences with the new type 3 attenuated poliovirus USOL-D bac developed in Czechoslovakia, Xth Symp. Europ. Ass. poliomyelitis and allied diseases, Warsaw, 1964, p. 413. 10. BSttiger, M., Gard, S., and Zetterberg, B.: Vaccination with attenuated type 1 poliovirus, the CHAT strain. II. Transmission of virus in relation to age, Acta paediat, seandinav. 55: 416, 1966. 11. Pagano, J. S., and Hoskins, J. iV[.: Variations in the assessment of the comparative neurovirulence of type 3 attenuated poliovirus vaccines, Conference on testing for neurovirulence of virus vaccines, Permanent section of microbiological standardization, Munchen, 1965, p. 221. 12. Ikie, D., and Janikic, B.: Epldemiologlcal data on efficacy of live oral pollovaccine Koprowsky, Proceedings of symposium on the characterization and uses of human diploid cell strains, Opatija, 1963, p. 541. 13. Buser, F., Fleury, C., Martin du Pan, R., and Rohner, E.: Immunization with live attenuated poliovirus prepared in human diploid cell strains, with special reference to the WM3 strain. Proceedings of symposium on characterization and uses of human diploid cell strains, Opatija, 1963, p. 381.