Acta Pædiatrica ISSN 0803-5253
MEDICAL ESSAY
The Swedish polio vaccine in memoriam Erik Lycke ([email protected]) €teborg, Sweden Sahlgren0 s Academy, University of Gothenburg, Go
Keywords Vaccine, Vaccination programmes, Vaccination Correspondence Erik Lycke MD, PhD, Aschebergsgatan 11A, 41127 €teborg, Sweden. Go Tel.: +46 31-121634 | Email: [email protected] Received 28 January 2014; accepted 29 January 2014. DOI:10.1111/apa.12584
INTRODUCTION This essay refers to personal memories of the 1950s and 1960s, when the Swedish polio vaccine was produced and we eradicated wild endemic polio virus. It is also a tribute to Sven Gard (1905–1998) (Figure 1), professor of virology, my mentor and scientific advisor, whose scientific contributions were crucial for the success of the vaccine and the vaccination campaigns.
THE EPIDEMICS It is difficult for the younger generations of today to image the fear in the past of contracting polio. The fear for polio had its roots in religious and superstitious believes, like the horror for plague in medieval times. Only a very select group of medical professionals knew anything about the nature of viruses and the epidemiology and the pathogenesis of poliomyelitis. One physician who was a patient during the 1953 epidemic said: The fear of polio was a reality, the treatment was a bed, nothing more, there were too many paralytic cases to take care of (1). The most severe epidemics, which resulted in thousands of cases of paralysis
Figure 1 Sven Gard (1905–1998), professor of virology at Karolinska Institutet in Stockholm.
and deaths hit Sweden in 1912, 1937, 1944, 1949 and 1953. It was not until 1963 that we had a polio free year. THE POLIO VACCINES For production of the inactivated polio vaccine (IPV) the virus suspensions are treated with formaldehyde, a process usually referred to as the inactivation of the virus, since after this treatment the vaccine shall no longer contain infectious virus. On the other hand, the viral proteins shall remain unaffected as much as possible during the inactivation process. The subtle balance between safety and immunogenicity is essential for the quality of the IPV, and consequently, the inactivation process must be elaborated with great care. The orally administered polio vaccine (OPV) contains attenuated, low-virulent but infectious and replicating virus. These fundamental differences between IPV and OPV are of importance when and how IPV or OPV are to be ideally used. I shall later in this essay discuss these matters in some detail. The first batches of the Swedish polio vaccine were produced already in 1954 but had to be discarded when safety tests revealed presence of infectious virus after the formaldehyde treatment. However, in 1957 and 1958, more than a million children were vaccinated with batches which had passed the many compulsory safety tests. And, in order to avoid further delays in the vaccination programme, vaccine imported from USA was used for the youngest age groups. From 1959 onwards, only Swedish vaccine was administered and in 1965 more than 4.5 million children and young adults had been vaccinated. The effect of the vaccinations was soon apparent. The epidemics disappeared and no more outbreaks were registered after 100 cases in Gothenburg in 1961. In the late 1960s, polio virus was no longer found in stool specimens or sewage samples, indicating that endemic wild polio virus was no longer circulating.
ª2014 Foundation Acta Pædiatrica. Published by John Wiley & Sons Ltd 2014 103, pp. 483–487
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SVEN GARD The Jubilee Fund, honoring King Gustav V’s 80th birthday, was established in 1938. And it was his wish that it should be used to finance the research and treatment of crippling diseases, in particular poliomyelitis and rheumatism. The Svedberg, head of the Department of Physical Chemistry at Uppsala University and a Nobel Prize laureate (1926) and another Nobel Prize recipient of the department, Arne Tiselius (1948) were asked to suggest the most urgent research project to overcome the poliomyelitis epidemics. They suggested studies on the chemical and physical structure of polio virus and approached Sven Gard, physician and scientist working with Svedberg0 s group in Uppsala. In addition, Gard received a grant to work at the Rockefeller Institute in New York. Staying there for almost 2 years Gard tried to purify mouse polio virus. And Albert Sabin – who later developed the OPV was to follow Gard to Uppsala for further studies. ‘We were supposed to solve all the riddles of polio’, as Gard later said. However, in 1939 the war made Sabin’s visit to Sweden impossible. Gard graduated in Uppsala but failed to purify the mouse polio virus. He later teamed up with H akan Leyon to present relevant data on the structure of polio virus, including electron microscopy images. But by then the findings of Schwerdt and Shaffer (2) were already published and confirmed. In 1948, Gard was appointed professor of virology at Karolinska Institutet (KI) in Stockholm, the first chair of virology in Sweden and probably also internationally. At Gard0 s own request the Department for Virus Research of KI was located at the National Bacteriological Laboratory (SBL). Gard had, for some years, collaborated with Carl Kling from SBL, and Kling had launched the hypothesis that polio was spread by fecal excretions. Kling and Gard gathered much of the information that would ultimately lead to important insights into the polio epidemiology. Both KI and SBL benefitted from the collaboration and in the years that followed Gard became an appreciated advisor to many of the physicians and chemists at SBL. The probably most impressive of Sven Gard0 s many talents as scientist was his analytical ability. He worked systematically according to a fixed protocol, was an advanced statistician and, fostered by the physical chemistry regime of the Uppsala department, he was not content until he could describe his findings with a valid formula. When I first met him he was already a living legend, dedicated to science, loved and feared. He was considered to be demanding while in reality he was generous and helpful. Just outside the door to his office there were three chairs. Once a week these chairs were occupied by doctors in spe, who were waiting with their dissertation manuscripts in trembling hands. Sven Gard was unpretentious, and always wore a grey suit and a black trench coat regardless of whether it was summer or winter. He walked from his home to the laboratory. His personal office in the department was small, nine square meters, with a desk, two chairs and bookshelves.
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He was admired by many but also criticised. His exceptional personal integrity made him sensitive about everything that he felt harmed scientific honesty. If he saw that someone was not presenting scientific data honestly and correctly he could become an unforgiving and dangerous enemy. In his view, scientific observations should not be patented, they belonged to all mankind. And a true scientist should be personally responsible for the proper use of his discoveries. Sven Gard was well aware of the considerable challenge of producing a polio vaccine. During the first decades of the 20th century some attempts to produce a polio vaccine had been made in USA but all had failed and some had even been detrimental. However, the January 1949 issue of Science published results from the Children0 s Hospital in Boston. John Enders, Thomas Weller and Frederick Robbins reported that they had observed multiplication of polio virus in human fetal tissue fragments suspended in a nutrient medium (3). For years it had been considered as an axiom that polio virus only replicated in neural tissues. Now Enders and coworkers reported, most sensationally, that polio virus seemed to infect and replicate also in nonneuronal cells. Suddenly a vaccine against polio was no longer just wishful thinking, and a production of a vaccine seemed a real possibility. In 1952 Gard visited Enders in Boston, saw polio virus infected cultures, and was convinced. He nominated Enders for a Nobel Prize in medicine and physiology. However, in his evaluations in the Nobel committee he later suggested that the prize should be given jointly to Enders, Weller and Robbins since ‘no one of the group seemed to have contributed more than any of the others’. In 1954 they received the prestigious prize.* In 1951, I attended a seminar by Sven Gard who was an excellent lecturer, and became irresistibly interested in virology. For 4 months I practiced bacteriological techniques at a large diagnostic laboratory in Stockholm and with this ‘meritorious’ background I approached Gard, was interviewed and offered a post as his assistant. His working hours were from 10 am to 10 pm and somehow I managed to adapt myself to his routines. Generously, Sven allowed me to set my own schedules and was free to include the times when I had to be absent due to my clinical courses. I was still a student but also the assistant of the Department for Virus Research. In 1953 I published three papers, had learned to cultivate cells and tissue fragments and was fostered by Sven as if I was a son of the house.
THE CONTROVERSY WITH SALK Gunnar Olin, the director of SBL was also the administrative head for the polio vaccine project and Sven Gard was its scientific advisor. Tore Wessl en, a histologist with substantial knowledge in cell and tissue culturing, was recruited, and he set up a new vaccine production laboratory called Polio One. At Gard0 s request I moved to Polio One and we prepared with assistance of four technicians suspended tissue fragment cultures of human fetal tissues for production of virus suspensions. A mixture of formalin
ª2014 Foundation Acta Pædiatrica. Published by John Wiley & Sons Ltd 2014 103, pp. 483–487
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and glycine was added to the virus suspensions giving a final concentration of 0.006 M of formaldehyde and 0.02 M of glycine. The glycine-formaldehyde mixture was thought to function as a formaldehyde ‘buffer’ and probably this assumption was correct. The temperature was kept at 25°C to minimise the loss of immunogenicity of the viral proteins and samples were collected daily to assay remaining virus infectivity. Tore Wesslen, who had introduced the formaldehydeglycine mixture was keen to see this technical detail patented, Gard refused, and in 1955 Wesslen left for Uppsala. Jonas Salk (4) had repeatedly stated that the inactivation process of polio virus in presence of formaldehyde followed the kinetics of a first order chemical reaction, i.e. that for each day of treatment with formaldehyde the same amount of virus in log units lost its infectivity, or in other words that the log of remaining virus activity was a linear function of the time of treatment. Thus he claimed that it was possible to find the time of treatment when the vaccine was safe to use by simply extrapolate along a linear regression line. Our observations demonstrated, on the other hand, that Salk0 s statement must be wrong. In fact, the inactivation reaction graph deviated from a linear course, Figure 2 (5). At the Third International Poliomyelitis Conference in Rome in 1954, Gard pointed out that Salk0 s recommendations could mislead the US vaccine producers, but Salk was unwilling to discuss the matter. He simply meant that our observations were due to differences between American and Swedish inactivation techniques. Gard now asked me to test all possibly influential parameters such as temperature during the inactivation process, pH, formaldehyde concentration etc. In addition, I tested filtration, mechanical treatment and finally even purified virus, all to exclude that virus hidden in cellular debris or clusters possibly could escape exposure to the formaldehyde. However, in none of all our experiments the results supported the view that the deviation from a strict first order chemical reaction was a technical problem. In April of 1955, newspapers reported the so-called Cutter incident, 204 children in USA were infected by a vaccine produced by the Cutter factory following Salk0 s recommendations, and 11 children died. Gard0 s suspicions had been sadly verified.
GARD0 S FORMULA We wanted a better understanding of the kinetics of the inactivation process and Gard went to Besken, the first Swedish-made computer at KTH, the technical university of Stockholm. Besken was a huge monster, occupying alone a whole room, and Gard spent a week feeding the computer all the data we possessed about the inactivation process. However, Besken was reluctant to reveal the mystery of the kinetics. Nevertheless, Gard returned with a formula of his own which described the course of the inactivation remarkably well (6). Gard’s formula read:
Figure 2 Kinetics of the inactivation of polio virus infectivity by treatment with 0.006 M formaldehyde at 25°C. Shaded area: mean values of 15 assays and statistical confidence intervals. Heavy line: best fitting linear regression line. Ordinate: log virus activity. Abscissa: time of treatment in days.
log y0 log y ¼ a log (1+bt) log y refers to the remaining infectivity at time t according to the assays of viral activity and deducted from the initial virus activity, log y0, the amount of inactivated virus is obtained. Of the two parameters, a and b, a determines the shape of the curvature of the regression line, i.e. the deviation from linearity, and b its position in relation to origin. The two parameters can be statistically estimated from a series of inactivation data. As shown by Figure 3 there was an extraordinary good fit when German and Swedish results were compared giving us an indication that Gard0 s formula was a reliable instrument. Gard0 s formula provided us with the necessary information to produce a safe vaccine but one more quality guarantee was essential. Earlier in this essay I have touched upon the crucial balance between inactivation of viral infectivity and maintenance of the immunising capacity of the vaccine. A method was required to ascertain this balance so that the relative immunogenicity of the different
ª2014 Foundation Acta Pædiatrica. Published by John Wiley & Sons Ltd 2014 103, pp. 483–487
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at the dinner table he suddenly said: ‘do you hate me as much as Sven Gard does?’ We all laughed and I did not have to answer.
Figure 3 Fitness of observations to Gard’s formula. Comparison of German (filled) circles and Swedish (open circles). German data: means of 60 batches. Swedish data: means of 21 batches. Parameters: a = 12.5, b = 0.779 and 0.339. Ordinate: log y0 – log y. Abscissa: log (1 + bt).
batches could be estimated. Using the so-called guinea pig test (7) we were able to estimate the antigenic extinction limit for each batch of vaccine. Together with our in-house references the guinea pig test soon provided us with a reliable control of the immunisation capacity of individual vaccine batches, and the test became an appreciated quality marker (8). The final protocol for the vaccine production was gradually developed. Because monkey kidney cell cultures produced ten times more virus than human fetal cultures a regular import of monkeys was organised and a second vaccine producing unit was established, Polio Two. More people was recruited, and the evaluation of vaccination results and surveillance of immunity became important issues. I graduated at KI in 1958 and in my thesis I discussed the results of my systematic studies on the inactivation of polio virus by formaldehyde. I received a Rockefeller scholarship in 1959 and spent a year at the University of Michigan. Before returning home I visited, among others, Jonas Salk and Julius Youngner, in Pittsburgh. Youngner, who was Salk0 s closest collaborator was interested in discussing various aspects of the inactivation process. While talking to him I became fully aware of that also he had noted the deviation from linearity of the inactivation kinetics and that it had worried him. Salk had invited me and my wife to his home and when we sat down
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NO NOBEL PRIZE FOR THE POLIO VACCINE The polio vaccines and the masterful accomplishment of the mass vaccination campaigns represent important achievements in the history of medicine. In fact they have been described as ‘milestones of progress’. Yet none of the scientists who were prominently involved in the development of the vaccines has been honored with a Nobel Prize. Salk was nominated for the first time in 1955, but in the preliminary evaluation Gard delivered to the Nobel committee he said that he was unable to reach a conclusion and wanted an ‘exhaustive analysis.’ (9). The following year when Salk was nominated for a second time Gard0 s thorough analysis for the Nobel committee commented on Salk’s misinterpretation of the inactivation kinetics. Gard argued that Salk0 s incorrect recommendations to the manufacturers were the cause to the Cutter incident. He ended his evaluation as follows: ‘According to mine opinion Salk has not demonstrated the cautiousness that one would expect to be applied in this context. It is my view, based on these conclusions, that Salk0 s publications on the poliomyelitis vaccine cannot be considered as prize worthy.’ Salk was nominated in 10 proposals in 1958 and seven times in 1959 but he was never considered worthy of a Nobel Prize, probably due to his misinterpretation of the inactivation kinetics and the Cutter incident. Koprowski and Sabin were nominated in 1958 for their attenuation of the polio strains used in OPV. Again Gard felt it premature to take a firm stand. In the late 1960s, I was approached by Rune Grubb, professor of bacteriology at Lund University. He wanted to nominate Salk and Gard for their achievements with the IPV and Koprowski and Sabin for the OPV. The draft nomination proposal was signed by many Swedish virologists, bacteriologists and immunologists and by their colleagues in the other Scandinavian countries. But the nomination never reached the Nobel Committee. When Gard heard about the proposal he vetoed it and said that he would not accept the nomination. His explanation was that the primary scientific achievement, the demonstration of the multiplication of polio virus in non-neuronal cells, already had been acknowledged, when Enders, Weller and Robbins received the prize. According to the Nobel statutes, secondary applications of already awarded accomplishments are not to be awarded. Sven Gard was a man of exceptional integrity.
IPV OR OPV The vaccination campaigns administrated by the World Health Organisation (WHO) have successfully annihilated epidemic polio globally. However, the eradication of wild polio virus which was the ultimate goal has not been achieved despite costly mass vaccinations since 1988. Polio
ª2014 Foundation Acta Pædiatrica. Published by John Wiley & Sons Ltd 2014 103, pp. 483–487
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is still endemic in Afghanistan, Nigeria and Pakistan and minor outbreaks have been reported also from Syria to Israel. In his article ‘Polio risk looms over Europe’ (10), senior reporter Declan Butler quotes the concerns of Marc Sprenger, director of the European Center for Disease Prevention and Control (ECDC) that surveillance systems and vaccination rates are suboptimal in many European countries. There is a risk, Sprenger says, that wild polio virus will be transmitted to Europe. Wild polio virus seems to circulate in both Syria and Israel. OPV vaccinations have already been administered in extensive campaigns further campaigns are also planned for the neighboring countries. It is likely that OPV vaccinations will be recommended also for Europe. If so, the recommendation might be a cause for concern. The OPV stimulates protective intestinal immune responses but does not eliminate fecal excretion and transmission. And circulation of the vaccine virus complicates surveillance systems. Moreover, there is always the risk that OPV virus reverts to virulence. In fact true polio eradication demands the elimination of both wild and vaccine- derived virus. The WHO OPV vaccinations have faced these and many other problems in attempts to eradicate polio by OPV vaccination. Most European countries have been primarily vaccinated with IPV, and Finland, Holland and Sweden have exclusively used IPV. Europe has been reportedly free of polio since 2002. The IPV vaccination provides good individual protection against polio and if the vaccination rate is sufficiently high the unvaccinated individuals of a population are also protected by herd immunity. But individuals protected against polio might if exposed to infection excrete wild endemic virus, as was recently observed in Israel (11). They obviously represent a risk for the spread of the infection. However, the duration of the virus excretion period depends on the level of the systemic immunity induced by the IPV, thus the higher the titers of virus neutralising serum antibodies, the shorter is the excretion period (12,13). The experiences from the 1950s and 1960s
should not be neglected. IPV vaccinations and surveillance of subsequent immunity seems the preferable choice. And as Sven Gard said: ‘the only good virus is a dead virus’.
References 1. Olsson TH. Polioepidemin 1953 – ett outpl anligt patientminne. €kartidningen 2006; 103: 30–1. La 2. Schwerdt CE, Schaffer FL. Some physical and chemical properties of purified poliomyelitis virus preparations. Ann N Y Acad Sci 1955; 61: 740–50. 3. Enders J, Weller TH, Robbins FC. Cultivation of the Lansing strain of poliomyelitis virus in cultures of various human embryonic tissues. Science 1949; 109: 85–7. 4. Salk J. Poliomyelitis vaccine in the fall of 1955. Amer Health 1955; 46: 1–14. 5. Wesslen T, Lycke E, Gard S, Olin G. Inactivation of poliomyelitis virus by formaldehyde. Arch ges Virusforsch 1957; VII: 125–35. 6. Gard S, Lycke E. Inactivation of poliomyelitis virus by formaldehyde. Analysis of inactivation curves. Arch ges Virusforsch 1957; VII: 471–82. 7. Gard S, Wesslen T, Fagraeus A, Svedmyr A, Olin G. Arch ges Virusforsch 1956; 6: 401. 8. Gard S, Johnsson T, Lycke E, Melen B, Olin G, Sahlenstedt R, et al. Potency of poliovirus vaccines. Correlation between antigenic extinction limit titers in guinea pigs and results obtained in man. Arch ges Virusforsch 1958; VIII: 423–9. 9. Norrby E. Nobel prizes and life sciences. New Jersey, World Scientific, 2010. 10. Butler D. Polio risk looms over Europe. Cases in Syria highlights vulnerability of nearby countries to the viral disease. Nature 2013; 502: 601–2. 11. ECDC. Wild type poliovirus in Israel – what is the risk to the EU/EEA? ECDC. 213. 601–2. € ttiger M, Lagercrantz R. First International 12. Gard S, Bo Conference on Live Polio Virus Vaccine. Vaccination with Attenuated Poliovirus Type 1, the CHAT Strain. Pan Am Sanitary Bur Scient Publ 1959; 44: 350–5. 13. Faxen N, Kallings LO, Lund E. Excretion period of attenuated polio vaccine virus in infants. Arch ges Virusforsch 1962; VIII: 1–6.
ª2014 Foundation Acta Pædiatrica. Published by John Wiley & Sons Ltd 2014 103, pp. 483–487
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MEDICAL ESSAY
The Swedish polio vaccine in memoriam Erik Lycke ([email protected]) €teborg, Sweden Sahlgren0 s Academy, University of Gothenburg, Go
Keywords Vaccine, Vaccination programmes, Vaccination Correspondence Erik Lycke MD, PhD, Aschebergsgatan 11A, 41127 €teborg, Sweden. Go Tel.: +46 31-121634 | Email: [email protected] Received 28 January 2014; accepted 29 January 2014. DOI:10.1111/apa.12584
INTRODUCTION This essay refers to personal memories of the 1950s and 1960s, when the Swedish polio vaccine was produced and we eradicated wild endemic polio virus. It is also a tribute to Sven Gard (1905–1998) (Figure 1), professor of virology, my mentor and scientific advisor, whose scientific contributions were crucial for the success of the vaccine and the vaccination campaigns.
THE EPIDEMICS It is difficult for the younger generations of today to image the fear in the past of contracting polio. The fear for polio had its roots in religious and superstitious believes, like the horror for plague in medieval times. Only a very select group of medical professionals knew anything about the nature of viruses and the epidemiology and the pathogenesis of poliomyelitis. One physician who was a patient during the 1953 epidemic said: The fear of polio was a reality, the treatment was a bed, nothing more, there were too many paralytic cases to take care of (1). The most severe epidemics, which resulted in thousands of cases of paralysis
Figure 1 Sven Gard (1905–1998), professor of virology at Karolinska Institutet in Stockholm.
and deaths hit Sweden in 1912, 1937, 1944, 1949 and 1953. It was not until 1963 that we had a polio free year. THE POLIO VACCINES For production of the inactivated polio vaccine (IPV) the virus suspensions are treated with formaldehyde, a process usually referred to as the inactivation of the virus, since after this treatment the vaccine shall no longer contain infectious virus. On the other hand, the viral proteins shall remain unaffected as much as possible during the inactivation process. The subtle balance between safety and immunogenicity is essential for the quality of the IPV, and consequently, the inactivation process must be elaborated with great care. The orally administered polio vaccine (OPV) contains attenuated, low-virulent but infectious and replicating virus. These fundamental differences between IPV and OPV are of importance when and how IPV or OPV are to be ideally used. I shall later in this essay discuss these matters in some detail. The first batches of the Swedish polio vaccine were produced already in 1954 but had to be discarded when safety tests revealed presence of infectious virus after the formaldehyde treatment. However, in 1957 and 1958, more than a million children were vaccinated with batches which had passed the many compulsory safety tests. And, in order to avoid further delays in the vaccination programme, vaccine imported from USA was used for the youngest age groups. From 1959 onwards, only Swedish vaccine was administered and in 1965 more than 4.5 million children and young adults had been vaccinated. The effect of the vaccinations was soon apparent. The epidemics disappeared and no more outbreaks were registered after 100 cases in Gothenburg in 1961. In the late 1960s, polio virus was no longer found in stool specimens or sewage samples, indicating that endemic wild polio virus was no longer circulating.
ª2014 Foundation Acta Pædiatrica. Published by John Wiley & Sons Ltd 2014 103, pp. 483–487
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SVEN GARD The Jubilee Fund, honoring King Gustav V’s 80th birthday, was established in 1938. And it was his wish that it should be used to finance the research and treatment of crippling diseases, in particular poliomyelitis and rheumatism. The Svedberg, head of the Department of Physical Chemistry at Uppsala University and a Nobel Prize laureate (1926) and another Nobel Prize recipient of the department, Arne Tiselius (1948) were asked to suggest the most urgent research project to overcome the poliomyelitis epidemics. They suggested studies on the chemical and physical structure of polio virus and approached Sven Gard, physician and scientist working with Svedberg0 s group in Uppsala. In addition, Gard received a grant to work at the Rockefeller Institute in New York. Staying there for almost 2 years Gard tried to purify mouse polio virus. And Albert Sabin – who later developed the OPV was to follow Gard to Uppsala for further studies. ‘We were supposed to solve all the riddles of polio’, as Gard later said. However, in 1939 the war made Sabin’s visit to Sweden impossible. Gard graduated in Uppsala but failed to purify the mouse polio virus. He later teamed up with H akan Leyon to present relevant data on the structure of polio virus, including electron microscopy images. But by then the findings of Schwerdt and Shaffer (2) were already published and confirmed. In 1948, Gard was appointed professor of virology at Karolinska Institutet (KI) in Stockholm, the first chair of virology in Sweden and probably also internationally. At Gard0 s own request the Department for Virus Research of KI was located at the National Bacteriological Laboratory (SBL). Gard had, for some years, collaborated with Carl Kling from SBL, and Kling had launched the hypothesis that polio was spread by fecal excretions. Kling and Gard gathered much of the information that would ultimately lead to important insights into the polio epidemiology. Both KI and SBL benefitted from the collaboration and in the years that followed Gard became an appreciated advisor to many of the physicians and chemists at SBL. The probably most impressive of Sven Gard0 s many talents as scientist was his analytical ability. He worked systematically according to a fixed protocol, was an advanced statistician and, fostered by the physical chemistry regime of the Uppsala department, he was not content until he could describe his findings with a valid formula. When I first met him he was already a living legend, dedicated to science, loved and feared. He was considered to be demanding while in reality he was generous and helpful. Just outside the door to his office there were three chairs. Once a week these chairs were occupied by doctors in spe, who were waiting with their dissertation manuscripts in trembling hands. Sven Gard was unpretentious, and always wore a grey suit and a black trench coat regardless of whether it was summer or winter. He walked from his home to the laboratory. His personal office in the department was small, nine square meters, with a desk, two chairs and bookshelves.
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He was admired by many but also criticised. His exceptional personal integrity made him sensitive about everything that he felt harmed scientific honesty. If he saw that someone was not presenting scientific data honestly and correctly he could become an unforgiving and dangerous enemy. In his view, scientific observations should not be patented, they belonged to all mankind. And a true scientist should be personally responsible for the proper use of his discoveries. Sven Gard was well aware of the considerable challenge of producing a polio vaccine. During the first decades of the 20th century some attempts to produce a polio vaccine had been made in USA but all had failed and some had even been detrimental. However, the January 1949 issue of Science published results from the Children0 s Hospital in Boston. John Enders, Thomas Weller and Frederick Robbins reported that they had observed multiplication of polio virus in human fetal tissue fragments suspended in a nutrient medium (3). For years it had been considered as an axiom that polio virus only replicated in neural tissues. Now Enders and coworkers reported, most sensationally, that polio virus seemed to infect and replicate also in nonneuronal cells. Suddenly a vaccine against polio was no longer just wishful thinking, and a production of a vaccine seemed a real possibility. In 1952 Gard visited Enders in Boston, saw polio virus infected cultures, and was convinced. He nominated Enders for a Nobel Prize in medicine and physiology. However, in his evaluations in the Nobel committee he later suggested that the prize should be given jointly to Enders, Weller and Robbins since ‘no one of the group seemed to have contributed more than any of the others’. In 1954 they received the prestigious prize.* In 1951, I attended a seminar by Sven Gard who was an excellent lecturer, and became irresistibly interested in virology. For 4 months I practiced bacteriological techniques at a large diagnostic laboratory in Stockholm and with this ‘meritorious’ background I approached Gard, was interviewed and offered a post as his assistant. His working hours were from 10 am to 10 pm and somehow I managed to adapt myself to his routines. Generously, Sven allowed me to set my own schedules and was free to include the times when I had to be absent due to my clinical courses. I was still a student but also the assistant of the Department for Virus Research. In 1953 I published three papers, had learned to cultivate cells and tissue fragments and was fostered by Sven as if I was a son of the house.
THE CONTROVERSY WITH SALK Gunnar Olin, the director of SBL was also the administrative head for the polio vaccine project and Sven Gard was its scientific advisor. Tore Wessl en, a histologist with substantial knowledge in cell and tissue culturing, was recruited, and he set up a new vaccine production laboratory called Polio One. At Gard0 s request I moved to Polio One and we prepared with assistance of four technicians suspended tissue fragment cultures of human fetal tissues for production of virus suspensions. A mixture of formalin
ª2014 Foundation Acta Pædiatrica. Published by John Wiley & Sons Ltd 2014 103, pp. 483–487
Polio vaccine
Lycke
and glycine was added to the virus suspensions giving a final concentration of 0.006 M of formaldehyde and 0.02 M of glycine. The glycine-formaldehyde mixture was thought to function as a formaldehyde ‘buffer’ and probably this assumption was correct. The temperature was kept at 25°C to minimise the loss of immunogenicity of the viral proteins and samples were collected daily to assay remaining virus infectivity. Tore Wesslen, who had introduced the formaldehydeglycine mixture was keen to see this technical detail patented, Gard refused, and in 1955 Wesslen left for Uppsala. Jonas Salk (4) had repeatedly stated that the inactivation process of polio virus in presence of formaldehyde followed the kinetics of a first order chemical reaction, i.e. that for each day of treatment with formaldehyde the same amount of virus in log units lost its infectivity, or in other words that the log of remaining virus activity was a linear function of the time of treatment. Thus he claimed that it was possible to find the time of treatment when the vaccine was safe to use by simply extrapolate along a linear regression line. Our observations demonstrated, on the other hand, that Salk0 s statement must be wrong. In fact, the inactivation reaction graph deviated from a linear course, Figure 2 (5). At the Third International Poliomyelitis Conference in Rome in 1954, Gard pointed out that Salk0 s recommendations could mislead the US vaccine producers, but Salk was unwilling to discuss the matter. He simply meant that our observations were due to differences between American and Swedish inactivation techniques. Gard now asked me to test all possibly influential parameters such as temperature during the inactivation process, pH, formaldehyde concentration etc. In addition, I tested filtration, mechanical treatment and finally even purified virus, all to exclude that virus hidden in cellular debris or clusters possibly could escape exposure to the formaldehyde. However, in none of all our experiments the results supported the view that the deviation from a strict first order chemical reaction was a technical problem. In April of 1955, newspapers reported the so-called Cutter incident, 204 children in USA were infected by a vaccine produced by the Cutter factory following Salk0 s recommendations, and 11 children died. Gard0 s suspicions had been sadly verified.
GARD0 S FORMULA We wanted a better understanding of the kinetics of the inactivation process and Gard went to Besken, the first Swedish-made computer at KTH, the technical university of Stockholm. Besken was a huge monster, occupying alone a whole room, and Gard spent a week feeding the computer all the data we possessed about the inactivation process. However, Besken was reluctant to reveal the mystery of the kinetics. Nevertheless, Gard returned with a formula of his own which described the course of the inactivation remarkably well (6). Gard’s formula read:
Figure 2 Kinetics of the inactivation of polio virus infectivity by treatment with 0.006 M formaldehyde at 25°C. Shaded area: mean values of 15 assays and statistical confidence intervals. Heavy line: best fitting linear regression line. Ordinate: log virus activity. Abscissa: time of treatment in days.
log y0 log y ¼ a log (1+bt) log y refers to the remaining infectivity at time t according to the assays of viral activity and deducted from the initial virus activity, log y0, the amount of inactivated virus is obtained. Of the two parameters, a and b, a determines the shape of the curvature of the regression line, i.e. the deviation from linearity, and b its position in relation to origin. The two parameters can be statistically estimated from a series of inactivation data. As shown by Figure 3 there was an extraordinary good fit when German and Swedish results were compared giving us an indication that Gard0 s formula was a reliable instrument. Gard0 s formula provided us with the necessary information to produce a safe vaccine but one more quality guarantee was essential. Earlier in this essay I have touched upon the crucial balance between inactivation of viral infectivity and maintenance of the immunising capacity of the vaccine. A method was required to ascertain this balance so that the relative immunogenicity of the different
ª2014 Foundation Acta Pædiatrica. Published by John Wiley & Sons Ltd 2014 103, pp. 483–487
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at the dinner table he suddenly said: ‘do you hate me as much as Sven Gard does?’ We all laughed and I did not have to answer.
Figure 3 Fitness of observations to Gard’s formula. Comparison of German (filled) circles and Swedish (open circles). German data: means of 60 batches. Swedish data: means of 21 batches. Parameters: a = 12.5, b = 0.779 and 0.339. Ordinate: log y0 – log y. Abscissa: log (1 + bt).
batches could be estimated. Using the so-called guinea pig test (7) we were able to estimate the antigenic extinction limit for each batch of vaccine. Together with our in-house references the guinea pig test soon provided us with a reliable control of the immunisation capacity of individual vaccine batches, and the test became an appreciated quality marker (8). The final protocol for the vaccine production was gradually developed. Because monkey kidney cell cultures produced ten times more virus than human fetal cultures a regular import of monkeys was organised and a second vaccine producing unit was established, Polio Two. More people was recruited, and the evaluation of vaccination results and surveillance of immunity became important issues. I graduated at KI in 1958 and in my thesis I discussed the results of my systematic studies on the inactivation of polio virus by formaldehyde. I received a Rockefeller scholarship in 1959 and spent a year at the University of Michigan. Before returning home I visited, among others, Jonas Salk and Julius Youngner, in Pittsburgh. Youngner, who was Salk0 s closest collaborator was interested in discussing various aspects of the inactivation process. While talking to him I became fully aware of that also he had noted the deviation from linearity of the inactivation kinetics and that it had worried him. Salk had invited me and my wife to his home and when we sat down
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NO NOBEL PRIZE FOR THE POLIO VACCINE The polio vaccines and the masterful accomplishment of the mass vaccination campaigns represent important achievements in the history of medicine. In fact they have been described as ‘milestones of progress’. Yet none of the scientists who were prominently involved in the development of the vaccines has been honored with a Nobel Prize. Salk was nominated for the first time in 1955, but in the preliminary evaluation Gard delivered to the Nobel committee he said that he was unable to reach a conclusion and wanted an ‘exhaustive analysis.’ (9). The following year when Salk was nominated for a second time Gard0 s thorough analysis for the Nobel committee commented on Salk’s misinterpretation of the inactivation kinetics. Gard argued that Salk0 s incorrect recommendations to the manufacturers were the cause to the Cutter incident. He ended his evaluation as follows: ‘According to mine opinion Salk has not demonstrated the cautiousness that one would expect to be applied in this context. It is my view, based on these conclusions, that Salk0 s publications on the poliomyelitis vaccine cannot be considered as prize worthy.’ Salk was nominated in 10 proposals in 1958 and seven times in 1959 but he was never considered worthy of a Nobel Prize, probably due to his misinterpretation of the inactivation kinetics and the Cutter incident. Koprowski and Sabin were nominated in 1958 for their attenuation of the polio strains used in OPV. Again Gard felt it premature to take a firm stand. In the late 1960s, I was approached by Rune Grubb, professor of bacteriology at Lund University. He wanted to nominate Salk and Gard for their achievements with the IPV and Koprowski and Sabin for the OPV. The draft nomination proposal was signed by many Swedish virologists, bacteriologists and immunologists and by their colleagues in the other Scandinavian countries. But the nomination never reached the Nobel Committee. When Gard heard about the proposal he vetoed it and said that he would not accept the nomination. His explanation was that the primary scientific achievement, the demonstration of the multiplication of polio virus in non-neuronal cells, already had been acknowledged, when Enders, Weller and Robbins received the prize. According to the Nobel statutes, secondary applications of already awarded accomplishments are not to be awarded. Sven Gard was a man of exceptional integrity.
IPV OR OPV The vaccination campaigns administrated by the World Health Organisation (WHO) have successfully annihilated epidemic polio globally. However, the eradication of wild polio virus which was the ultimate goal has not been achieved despite costly mass vaccinations since 1988. Polio
ª2014 Foundation Acta Pædiatrica. Published by John Wiley & Sons Ltd 2014 103, pp. 483–487
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is still endemic in Afghanistan, Nigeria and Pakistan and minor outbreaks have been reported also from Syria to Israel. In his article ‘Polio risk looms over Europe’ (10), senior reporter Declan Butler quotes the concerns of Marc Sprenger, director of the European Center for Disease Prevention and Control (ECDC) that surveillance systems and vaccination rates are suboptimal in many European countries. There is a risk, Sprenger says, that wild polio virus will be transmitted to Europe. Wild polio virus seems to circulate in both Syria and Israel. OPV vaccinations have already been administered in extensive campaigns further campaigns are also planned for the neighboring countries. It is likely that OPV vaccinations will be recommended also for Europe. If so, the recommendation might be a cause for concern. The OPV stimulates protective intestinal immune responses but does not eliminate fecal excretion and transmission. And circulation of the vaccine virus complicates surveillance systems. Moreover, there is always the risk that OPV virus reverts to virulence. In fact true polio eradication demands the elimination of both wild and vaccine- derived virus. The WHO OPV vaccinations have faced these and many other problems in attempts to eradicate polio by OPV vaccination. Most European countries have been primarily vaccinated with IPV, and Finland, Holland and Sweden have exclusively used IPV. Europe has been reportedly free of polio since 2002. The IPV vaccination provides good individual protection against polio and if the vaccination rate is sufficiently high the unvaccinated individuals of a population are also protected by herd immunity. But individuals protected against polio might if exposed to infection excrete wild endemic virus, as was recently observed in Israel (11). They obviously represent a risk for the spread of the infection. However, the duration of the virus excretion period depends on the level of the systemic immunity induced by the IPV, thus the higher the titers of virus neutralising serum antibodies, the shorter is the excretion period (12,13). The experiences from the 1950s and 1960s
should not be neglected. IPV vaccinations and surveillance of subsequent immunity seems the preferable choice. And as Sven Gard said: ‘the only good virus is a dead virus’.
References 1. Olsson TH. Polioepidemin 1953 – ett outpl anligt patientminne. €kartidningen 2006; 103: 30–1. La 2. Schwerdt CE, Schaffer FL. Some physical and chemical properties of purified poliomyelitis virus preparations. Ann N Y Acad Sci 1955; 61: 740–50. 3. Enders J, Weller TH, Robbins FC. Cultivation of the Lansing strain of poliomyelitis virus in cultures of various human embryonic tissues. Science 1949; 109: 85–7. 4. Salk J. Poliomyelitis vaccine in the fall of 1955. Amer Health 1955; 46: 1–14. 5. Wesslen T, Lycke E, Gard S, Olin G. Inactivation of poliomyelitis virus by formaldehyde. Arch ges Virusforsch 1957; VII: 125–35. 6. Gard S, Lycke E. Inactivation of poliomyelitis virus by formaldehyde. Analysis of inactivation curves. Arch ges Virusforsch 1957; VII: 471–82. 7. Gard S, Wesslen T, Fagraeus A, Svedmyr A, Olin G. Arch ges Virusforsch 1956; 6: 401. 8. Gard S, Johnsson T, Lycke E, Melen B, Olin G, Sahlenstedt R, et al. Potency of poliovirus vaccines. Correlation between antigenic extinction limit titers in guinea pigs and results obtained in man. Arch ges Virusforsch 1958; VIII: 423–9. 9. Norrby E. Nobel prizes and life sciences. New Jersey, World Scientific, 2010. 10. Butler D. Polio risk looms over Europe. Cases in Syria highlights vulnerability of nearby countries to the viral disease. Nature 2013; 502: 601–2. 11. ECDC. Wild type poliovirus in Israel – what is the risk to the EU/EEA? ECDC. 213. 601–2. € ttiger M, Lagercrantz R. First International 12. Gard S, Bo Conference on Live Polio Virus Vaccine. Vaccination with Attenuated Poliovirus Type 1, the CHAT Strain. Pan Am Sanitary Bur Scient Publ 1959; 44: 350–5. 13. Faxen N, Kallings LO, Lund E. Excretion period of attenuated polio vaccine virus in infants. Arch ges Virusforsch 1962; VIII: 1–6.
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