Being in the UK, I will start by quoting one of the most famous physicians in the history of this country, the surgeon Joseph Lister. Addressing Louis Pasteur, he expressed himself as follows:
It is my great privilege to convey to you, tributes, thanks and respect from all involved in medicine and surgery; it is true to say that, of all people in the world today, medical sciences owe you the most…For centuries, infectious diseases have been shrouded, as it were under a dark curtain. In discovering the microbial origin of disease you have raised that dark curtain! (Lister 1893).
Lister stated this at a ceremony held at the Sorbonne, in Paris, on the occasion of the 70th birthday anniversary of Louis Pasteur, on December 27, 1892. On that jubilee occasion, delegates from throughout the world gathered to pay homage to the scientist and to express their gratitude and admiration on behalf of their respective countries. Joseph Lister represented the Royal Societies of London and Edinburgh.
In 1892, the thoughts that were uppermost in the minds of Joseph Lister and the other participants at the jubilee ceremony no doubt centred on Pasteur’s most recent accomplishments in the field of infectious diseases. Today, more than a century later, we are far enough removed from those events to be able to more fully appreciate the impact of his body of work.
From among Pasteur’s early period, in the late 1840s, the image which comes most readily to mind is that of the young 26-year-old chemist, fresh out of the Ecole Normale, sorting out crystals of tartaric acid while the eminent crystallographer, Jean-Baptiste Biot, looked on in astonishment. After the experiment was over, and according to Pasteur himself, the illustrious old man was deeply moved. ‘My dear boy’, he said taking Louis by the arm, ‘I have loved science so much all my life that this touches my heart.’
What was this dramatic discovery?
What Pasteur had shown to Biot was that a rare form of tartrate, referred to as paratartrate, was in fact composed of equal quantities of two types of molecules whose crystals, although very similar, could be distinguished by the orientation of one tiny facet.
In addition to this minute difference in the shape of their crystals, these two molecular species, though identical in all of their physical and chemical properties, nonetheless had one other difference: their solutions rotated the plane of light polarization in opposite directions. Pasteur hypothesized that these two forms of tartrate corresponded to two different spatial configurations of the atoms within the molecule, two forms which were asymmetrical in themselves, but symmetrical with respect to one another, just as our two hands are. This was a most revolutionary hypothesis: two molecules, containing the same atoms, linked to one another by the same bonds, could nevertheless differ in the spatial arrangement of these atoms. This hypothesis, which would not be definitely confirmed until 30 years later with the establishment of the principle of carbon asymmetry, laid the foundation for stereochemistry. The far-reaching consequences of this concept can only fully be appreciated by recalling the fact that, as Pasteur himself sensed and as molecular biology would demonstrate many years later, all interactions between biological molecules, and hence all life processes result from the precise three-dimensional arrangement of the atoms within these molecules. Moreover, the emergence of the critical notions of symmetry and symmetry-breaking were of capital importance, for they laid the basis for a number of modern theories of physics such as that of elementary particles and phase changes.
Upon observing that all compounds whose solutions rotated the plane of polarized light (optically active) arose from plant and animal sources, Pasteur began to suspect that asymmetry was a sign of life. With this idea in mind, he, began studies on fermentation in 1854. Fermentation was known even in ancient times, for it was used in the preparation of bread, wine and many other types of food and drink. But when Pasteur began to examine it, the question of fermentation was the subject of utmost confusion. While in some cases, the participation of micro-organisms in such transformations of organic matter vas recognized, in general their role was completely misunderstood. Justus Liebig, the renowned German chemist, a contemporary and adversary of Pasteur, had stated:‘Yeast from the malt…(transfers) its own state of decomposition to that which is around it. The movement that disturbs the balance imprinted in its own elements also communicates with other elements of bodies in contact with it’ (Liebig 1839).
Needless to say, Pasteur could not accept such a confused explanation. The observation that optically active products appeared during the process of fermentation led him to propose an entirely different and much more accurate hypothesis. To Pasteur, ‘…fermentation, far from being a lifeless phenomenon, is a living process…all phenomena of fermentation correlate with the development of mycodermic cells and plants which I have prepared and studied in an isolated and pure state’. Indeed, he describes this method in his Note on lactic fermentation, published in 1857 and which can be considered as the birth certificate of Microbiology.
The purity of a ferment, its homogeneity, its unhindered development, with the help of a nutrient perfectly adapted to its individual nature, this is one of the essential conditions for obtaining high-quality fermentations (Pasteur 1857).
These are the basic principles of microbiology: isolate a micro-organism, and provide it with the adequate growth medium. With these simple rules, man had learned to master the microbe. As applied by Pasteur and his students to the preparation of wine, beer, vinegar and dairy products, these principles were to revolutionize those industries. They formed the basis for what we now refer to as biotechnology.
Each fermentation process results from the action of a specific micro-organism. To obtain a good fermentation, one has to introduce the right kind of micro-organism, or, at least, make sure that it is present, and avoid the presence of other types of micro-organisms, which could alter the process. Once the product is made, contaminating micro-organisms could still alter it. It is to prevent this alteration, in the case of wine, that Pasteur introduced a process now named after him, pasteurization.
Before Pasteur, when milk was getting sour, when meat or fish were decaying, nobody really knew why. Pasteur demonstrated that putrefaction, like fermentation, was due to the growth of micro-organisms. The empirical methods used in the past to delay food decay could then be rationalized and improved.
Pasteur understood that putrefaction, unpleasant as it may be, plays a major role in the recycling of elements between the living world and the mineral world. This ‘immense role of infinitely small bodies in the general economy of nature’, as Pasteur himself says (Pasteur 1862), warrants reflection in our present day era in which the protection of environment has become one of humanity’s major priorities.
Fermentation and putrefaction were often perceived as being spontaneous phenomena. But what was Pasteur to make of the ferments and yeasts which he felt were essential in the unfolding of those processes? Where did they come from? Did they appear spontaneously in the media, or did they come from elsewhere? Pasteur could not escape this debate, a debate stemming from ancient beliefs, concerning the spontaneous generation of bees, frogs, mice, etc., but which had taken on new strength toward the end of the 17th century following the discovery of ‘animalcules’ by Anton van Leeuwenhoek, the Dutch inventor of the microscope. On the subject of these microscopic organisms, the 18th century witnessed such supporters of spontaneous generation as the English scholar John Needham and France’s renowned naturalist Buffon taking part in heated debate with other scientists such as the Italian abbot Spallanzani.
Pasteur kept an open mind in his approach to this question. In 1859, in a letter to the man who was to become his most ardent opponent on the subject, Felix Archimede Pouchet, he wrote that the question of spontaneous generation was ‘entirely open and still awaiting proof ’ and that all of this was ‘unknown and warranted experimentation’ (Pasteur 1859).
Pasteur responded to his own invitation. By extremely painstaking experimental methodology, he demonstrated that the appearance of micro-organisms in a presterilized medium could always be explained by germs coming from the outside. He thus succeeded in discrediting any experimental basis for the theory of spontaneous generation.
On a philosophical level, the repercussions were resounding. The onset of life was decidedly not a predictable phenomenon, regularly occurring in any fermentable medium. The question of the origin of life was thereafter clearly set forth – and remains so today.
Pasteur’s research on fermentation and on so-called spontaneous generation inevitably led him toward the study of infectious diseases. He had only recently succeeded in demonstrating that if environmental yeasts are prevented from being deposited on grapes, the juices of these grapes will not ferment, when he wrote:
‘By analogy, is it unreasonable to hope that the day will come when easily administered preventive measures will be able to stop the scourges which terrify and decimate populations, such as yellow fever and the bubonic plague?’ (Pasteur 1879).
In other words, infectious diseases, like fermentations, are probably due to ‘germs’, and it may be possible to protect human beings against them, as one can ‘protect’ grapes against yeast.
But it was neither yellow fever nor bubonic plague that Pasteur would attempt to cure; that task would be left up to his followers. Instead, upon the request of his former mentor who was now senator in the department of the Gard, Jean-Baptiste Dumas, he next turned his attention toward silkworm disease. Pasteur knew virtually nothing about silkworms, but he accepted the challenge, seizing the opportunity to learn more about infectious diseases. After 5 years of painstaking research, from 1865 to 1870, he succeeded in saving the silk industry through a method called ‘graining’ which enabled the preservation of healthy eggs while eliminating those from contaminated females. This method is still used today in silk-producing countries. Indeed, in Japan, a law actually exists which controls its use!
As we mentioned earlier, according to Joseph Lister, it was Pasteur who demonstrated the microbial nature of infectious diseases. However, this does not imply that Pasteur alone was responsible for identifying the microbes which caused all major diseases. He did, of course, contribute to definitively identifying the agent of anthrax, which was decimating cattle and sheep herds; he identified staphylococcus, streptococcus and the septic vibrion. But Robert Koch, of the German school, provided an equally important contribution.
In reality, what is so uniquely important about the entire body of Pasteur’s work is that it laid the groundwork upon which the microbial theory of disease was built.
He demonstrated how to cultivate bacteria and later, for rabies, he set forth the premizes for the culture of viruses on animal tissues.
He had always been intrigued by the mechanisms through which pathogenic microbes caused profound disturbances in the physiology of the infected organism, what we would call today the mechanisms of pathogenicity. Not surprisingly, it is one of his closest collaborators, Emile Roux, who made the first major contribution to this field, by identifying diphtheria toxin.
Pasteur also demonstrated how pathogens are able to spread through animal and human populations, thereby laying the foundation for infectious epidemiology and defining the basic rules of hygiene.
Concerning epidemiology, I would like to recall two facts, one well known and one which is less, concerning anthrax.
The well known story is that of the so-called ‘cursed fields’, where it seemed that the cattle would systematically catch anthrax, and die. One day, while Pasteur was having a walk close to one such field, he noticed that the ground had a different colour in one place. He was told that, the year before, sheep which had died of anthrax had been buried at this spot. Getting closer, Pasteur noticed the presence, at the surface, of a multitude of those twisted tubes of earth which are excreted by worms. The idea then came to him that the worms, travelling up and down in the ground, could bring to the surface anthrax spores present in the dead bodies. He verified that, indeed, the excrements of worms taken at this place contained spores of the anthrax bacillus. The cattle could then contaminate itself by grazing in such spots. The cursed fields were thereafter eliminated, once farmers ceased to bury the dead animals in them.
Less well known, perhaps, are observations made, in 1881 by Ed. Nocard a veterinarian and a collaborator of Pasteur. At that time the battle was still raging between the ‘contagionists’ and the ‘spontaneists’. The former, with Pasteur, contended that infectious diseases could only be caught through contagion, whereas the latter claimed that they could appear spontaneously. In that context, Nocard was informed of the appearance of anthrax in farms where it had never been seen before, and where it thus must have appeared ‘spontaneously’. I will only relate his observations concerning one of these farms.
There, a young farmer full of dynamism, had decided to improve the yield of his fields. With this in mind he bought a large quantity of fertilizer, something unheard of in that area. He then got beautiful crops but, the year after, when he let his cattle graze on the same fields a large number immediately caught anthrax. That year he lost about a quarter of his herd. Nocard then discovered that the fertilizer was commercialized by a company that collected the carcasses of dead animals over a large area and did not take special precautions in transforming them into fertilizers. Among the carcasses some were bound to have been from animals which had died of anthrax, and the spores they contained had thus been distributed in the field of the unlucky young farmer. I am sure you will have already drawn a parallel with what happened at a time much closer to us in the dissemination of BSE.
Going back to Louis Pasteur himself, the last part of his life was no less productive than the rest. It is then, indeed, that he outlined the overall principles of vaccination and contributed to the foundation of immunology.
The concept of acquired immunity dates from ancient times. Indeed, Thucydides reported that those who were cured of the plague no longer ran the risk of falling victim to the disease. The first vaccine was developed by the English physician, Edward Jenner, who, at the end of the 18th century discovered that human beings could be protected against smallpox by inoculation of the similar but benign disease, called ‘cowpox’ in English, and ‘vaccine’ in French. Although of crucial importance, since it lead to complete eradication of smallpox in 1979, this empirical discovery could not be generalized to other diseases.
It is often said that Jenner discovered vaccination, and Pasteur invented vaccines.
Pasteur’s fundamental discovery in this field dates back to 1879 and concerned the disease known as fowl cholera, which was rampant in chicken coops at that time.
The disease was due to bacteria which today bear the name ‘Pasteurella’. When inoculated into a chicken, several drops of a culture of these bacteria were sufficient to kill the animal. But Pasteur noted, partly by chance it would seem, that chicken inoculated with an old culture not only did not die but were protected against a later inoculation with a virulent culture. The principle of vaccination with attenuated pathogens was thus discovered.
From then on, Pasteur repeatedly applied this principle to other diseases. His first great success both at a scientific 1evel and in terms of public opinion – today we would refer to it as a ‘mass media happening’– was the vaccination against anthrax. The famous public experiment held in Pouilly le-Fort in 1881, during the course of which 24 vaccinated sheep survived an injection of the anthrax bacillus, while 24 nonvaccinated sheep died, had extraordinary repercussions, convincing a large portion of public opinion of the validity of Pasteur’s work.
The few remaining sceptics rallied around Pasteur for his final victory, that of human vaccination against rabies. The problem was complicated from the very outset, for the rabies microbe was invisible – we now know that it is a virus rather than a bacterium – and could not multiply in any culture medium. But the stakes were high, for although the disease was relatively rare in France, it had always fascinated the popular imagination, conjuring up fear and mystery. For Pasteur, conquering rabies would consolidate the final victory of his theories.
Even though he could not see or cultivate the microbe, Pasteur knew that it had to be there. It had to be in the nervous system, recognized as its target. As a replacement for in vitro cultivation, Pasteur transmitted the infectious agent from animal to animal, by intracerebral inoculation. He adapted the disease to the rabbit, and then undertook to attenuate the invisible microbe, which he did by dessication of the spinal cord of infected animals. In doing so, he thought, at first, that he was doing the same as with chicken cholera and anthrax, i.e. create an attenuated form of the microbe. In fact, as he was to realize later, most of the virus was presumably killed in his preparations. Thus, rather unknowingly, he opened the way for the second class of vaccines, besides attenuated live micro-organisms, and consisting in inactivated micro-organisms, which would themselves later lead to subunit vaccines.
The story of the vaccination against rabies is well known. So well known, in fact, that for many this was the sole accomplishment of Pasteur!
The success of the vaccination of Joseph Meister on July 6, 1885 and of the shepherd boy Jean-Baptiste Jupille in October of that same year, followed by hundreds of other bite-victims from throughout the world, brought glory to Pasteur and opened up the era of preventive medicine. On the heels of this success, the ‘Academie des Sciences’ launched an international fund-raising campaign to build the Institut Pasteur. It was there that Louis Pasteur lived out his last years, it is there that he lies in his final resting place, and it is there that, for over a century now, the work that he began is being pursued by his followers.
Louis Pasteur has left us many messages, which I have tried to briefly summarize in this presentation. In ending, I would like to quote him once more, and leave you with a last message, which seemed to me of particular relevance at a time when we are confronted with such terrifying emerging infectious diseases, as are AIDS and BSE. This quote is from an article published in 1881, the year of the vaccine against anthrax, when Pasteur had just found that virulence is not a fixed trait of micro-organisms. Virulence could be weakened to create attenuated strains, why could it not be increased under other conditions? Here is the quotation:
And so it is that virulence appears to us in a new light, rather disquieting for humanity, unless nature in its evolution during centuries of the past already encountered all possible occasions of creating virulent or contagious diseases, something which is very unlikely.
What makes a micro-organism harmless for a human being or any given animal? It is a micro-organism which cannot grow in our body or in the body of this animal; but nothing proves that provided this micro-organism were to penetrate one of the thousands of species of Creation, it might not invade it and make it ill. Its virulence, then reinforced by successive passages through members of this species, could become able to infect some animal of large size, man or certain domestic animals. In this way new virulences or contagions could be created. (Pasteur 1881)