1822 – 1895

	Louis Pasteur, the only son of a poorly educated tanner, Jean, was born on December 27th, 1822 in the small village of Dole, in the region of Jura, France. He was not a particularly bright scholar and showed no flair for the physical sciences, but he did have a fine artistic talent and preferred drawing and fishing to other subjects. His drawings were so professional that he could have been a skilled portrait artist, but his ambition was to matriculate at the Sorbonne and become a chemist. He enrolled in a preparatory school for later entrance into the Sorbonne, but during the first few months there he became so homesick that he returned to Dole and his family. Several months later he decided to try again and returned to the same preparatory school where his grades improved sufficiently to be acceptable and although the examiner for his entrance chemistry examination described him as 'mediocre', he was admitted to the Sorbonne in 1843.
Crystallography In 1847, at the age of 26, Pasteur was in his final year at the University, working for his doctorate in chemistry in the Antoine Balard laboratory. It was there that he did his first work on molecular asymmetry, bringing together the principles of crystallography, chemistry and optics. Crystallography was just emerging as a branch of chemistry and Pasteur’s project was to crystallise a number of different compounds. He started working with tartaric acid, crystals of which are found in large amounts in the sediments of fermenting wine. Also found in the sediments in the wine barrels a second acid called paratartaric acid or ‘racemic acid’. These acids were identical, but in solution there was a difference. Tartaric acid rotated a beam of polarised light passing through it to the right, whereas paratartaric acid was unable to do the same. Pasteur refused to accept the notion that two compounds having the same chemical composition, but acting so differently in respect to the rotation of light, could be identical. He decided to examine the crystals of the tartaric acid that had no power to change the direction of polarised light. His perceptive mind noticed that the form of dried tartaric acid comprised two slightly different sorts of crystals. With small tweezers he collected each form in separate tubes of pure water. When he placed the first container before the source of polarised light, the light rotated to the right, the second container containing the second variety of crystals turned the light to the left. Through this simple experiment, Pasteur had made two major discoveries. Firstly, he had isolated a previously unknown tartaric acid, one that turned polarised light to the left and secondly, more importantly he had determined why one of the two tartaric acids was unable to rotate polarised light at all: it was composed of crystals that turned polarised light in opposite directions, therefore neutralising each other. This significant finding by a 26 year-old chemist in a doctoral dissertation made Pasteur famous, his discovery was the beginning of the science of stereochemistry. To Pasteur his discovery had a deeper meaning; he proposed that asymmetrical molecules were indicative of living processes. Today, we know that all the proteins of higher animals are made up of only those amino acids that exist in the left-hand form; the mirror image right-hand amino acids are not used by human or animal cells. Likewise, our cells burn only the right-handed form of sugar, not the left-handed form that can be made in a test tube. Immediately after his discovery, Pasteur was appointed to the professorship of chemistry at Dijon, where he stayed for a brief time before being transferred to Strasbourg University where he continued his studies on molecular asymmetry. It was here that he met and married the University Rector’s daughter, Marie Laurent, who was a devoted wife and mother and scientific helpmate to him for the rest of his life.
Alcoholic Fermentation In 1854 Pasteur was appointed Dean and Professor of Chemistry at the Faculty of Sciences in Lille, France. Lille was an industrial town containing a number of distilleries and factories. Pasteur enjoyed taking his students on tours of the factories and advised the managers that he was always available to help solve any problems they might encounter. M. Bigot, father of one of his students in chemistry, contacted Pasteur to help him overcome difficulties he was having manufacturing alcohol by fermentation of beetroot. Often, instead of alcohol, M. Bigot's fermentations produced lactic acid. At that time, fermentation leading to the production of wine, beer and vinegar was believed to be a straightforward chemical breakdown of sugar to the desired molecules. The experts of the day stated that the breakdown of sugar into alcohol during fermentation of sugar to wine and beer was due to the presence of inherent unstabilising vibrations. These could be transferred from a vat of finished wine to new grape pressings to start the process of fermentation again. Yeast cells were found in the fermenting wine and although recognised as being live organisms, they were believed to be either a product of fermentation or catalytic agents that provided useful ingredients for fermentation to proceed. The few biologists, who believed that yeast was the cause of, and not the product of fermentation, were ridiculed by the scientific experts. The scientific establishment believed that chemistry had come too far to allow a vitalistic life force theory to challenge chemical explanations of molecular reaction. This attitude did not help the problems being experienced by the brewers of wine, beer and vinegar; yields of alcohol might suddenly drop, wine might become sour or turn to vinegar, and vinegar, when desired, might not be formed and lactic acid appear instead; the quality of beer could suddenly change making quality control impossible. Too often the producers would have to throw the lot out and start again, with no more success. Pasteur visited M. Bigot’s factory and quickly found three clues that enabled him to solve the problem of alcoholic fermentation. Firstly, when alcohol was produced normally, the yeast cells were plump and budding, but when lactic acid formed instead of alcohol, small rod like microbes were always mixed with the yeast cells. Secondly, analysis of the batches of alcohol showed that amyl alcohol and other complex organic compounds were being formed during fermentation. Thirdly, some of the compounds were asymmetric, i.e. rotating light. Pasteur concluded and was able to prove that living cells, the yeast, were responsible for forming alcohol from sugar and that contaminating micro-organisms turned the fermentation sour. Over the next few years, Pasteur identified and isolated the specific micro-organisms responsible for the problems and demonstrated that if he heated the wine, beer, milk to moderately high temperatures for a few minutes, he could kill living micro-organisms and so sterilise (pasteurise) the batches and prevent their contamination. If pure cultures of microbes and yeasts were added, predictable fermentation would follow.
Spontaneous Generation While the scientific world was full of excitement and controversy over Pasteur's research on fermentation there was also the bitterly debated obscure matter of spontaneous generation. In 1702, Leeuwenhoek dismissed the concept of living things arising from inanimate objects as ridiculous. The idea that beetles, eels, maggots and now microbes could arise from putrefying matter was speculated on as far back as Greek and Roman times, and in the 1860s the debate was still going on. Against the advice of his colleagues, Pasteur joined in the arguments. Based on his work on fermentation, it seemed obvious to him that the sources of yeasts and other micro-organisms that were found during fermentation and putrefaction entered from the outside, for example, from the air. He conducted various experiments, but the one that settled the arguments was his design and use of the swan-neck flask. The flask had a long 'swan-like' neck open to the air, but dust and airborne microbes were unable to reach the liquid. In the experiment, Pasteur placed some fermentable juice in the flask and after sterilisation; the neck was heated and drawn out as a thin tube taking a gentle downward then upward arc – similar to the neck of a swan. Providing it remained sealed the contents remained unchanged. If the flask was opened, by removing the end of the neck, air entered but dust was trapped on the wet walls of the neck. Under this condition, the fluid would remain forever sterile, showing that air alone cannot set off the growth of micro-organisms, however, tip the flask and allow the liquid to touch the contaminated walls and then return to the broth, micro-organisms immediately began to grow. To quote Pasteur:- "Never will the doctrine of spontaneous generation recover from the mortal blow of this simple experiment. No, there is now no circumstance known in which it can be affirmed that microscopic beings came into the world without germs, without parents similar to themselves."
Diseases of Silkworms Pasteur hoped he could return to his laboratory and continue his studies of chemicals and their reactions with one another, but he was not allowed to do so. The Department of Agriculture asked him to head a commission to investigate a disease of silkworms that was destroying the French silk industry. He knew nothing about silkworms and was not aware that they could suffer from disease, but as he had been insisting since 1863 that all putrefying processes were caused by micro-organisms, he maintained that putrefaction was destroying the small creatures. He spent five years investigating the sickness killing the silkworms. He considered his studies a milestone in his research into infection and infectious diseases. He found that healthy worms became infected when allowed to nest on leaves used by infectious works. He also noted that some worms died shortly after infection, whereas other died some weeks later and some not at all. He established that temperature, humidity, ventilation, quality of the food, sanitation and adequate separation of the broods of newly hatched worms each played a part in the vulnerability of newly hatched worms. He was never able to identify the germ that he was sure was responsible, but he was able to devise methods of quickly detecting the illness in the silkworms. By diligent isolation and hygienic methods, he prevented the spread of the disease and eventually eliminated the epidemic.
Germ Theory The pinnacle of Pasteur’s achievements must be his development of the germ theory of disease and the use of vaccines to prevent them. His studies on the contamination of wine and beer by airborne yeast clearly showed that these ‘diseases’ were due to foreign micro-organisms. In England, Lister was so impressed by Pasteur’s work that he began to sterilise his equipment and sprayed phenol solutions in his wards to reduce infections following surgery. By 1875 many physicians recognised that some diseases were caused by specific micro-organisms, but the general medical opinion did not agree that important diseases like cholera, diphtheria, scarlet fever, childbirth fever, syphilis, smallpox, etc. could ever be caused by these agents. According to Pasteur’s son-in-law Vallery-Radot, , between April 1st and May 10th, 1856, there were 64 fatalities due to childbirth fever out of 347 confinements in the Paris Maternity Hospital. They closed the hospital and transferred the patients, but the contagion followed and nearly all the women died. Although he was not a physician, Pasteur managed to visit the morgues of hospitals, taking particular interest in women who had died of puerperal fever. He obtained samples of blood of the uterus and its secretion from the Parisian mothers who had died from the fever. When he examined the samples and took cultures, he always found a micro-organism that appeared to be composed of a series of tiny beads (which are now called streptococci). During his visits to hospital wards, Pasteur became ever more aware that infection was being spread by physicians and hospital attendants from sick to healthy patients. He stressed that avoidance of microbes meant avoidance of infection. In a famous speech to the Academy of Medicine in Paris, he said: "This water, this sponge, this lint with which you wash or cover a wound, may deposit germs which have the power of multiplying rapidly within the tissue….If I had the honour of being a surgeon….not only would I use none but perfectly clean instruments, but I would clean my hands with the greatest care…I would use only lint bandages and sponges previously exposed to a temperature of 1300 to 1500 degrees." Slowly, but surely, through the preaching’s of Pasteur, Lister and other physicians, antiseptic medicine and surgery became the rule.
Chicken Cholera In 1878 Pasteur turned his attention to the organisms causing chicken cholera. When chickens were injected with a culture of the cholera germs, they died within twenty-four hours, but one day Pasteur took a chance and injected two chickens with a culture that was not fresh, but several weeks old. The two chickens became ill, but then recovered. At that point in the experiments, the entire laboratory staff went on holiday and when they returned they continued injecting the chickens with the usual fresh cultures of cholera, including the two that had survived earlier with the old culture. All the chickens died the next day, with the exception of the two that had previously survived. When Pasteur saw the results, he realized that in a way, he was repeating the studies of Jenner 80 years earlier, who had discovered immunity to smallpox by vaccinating individuals with a mild form of cowpox. Pasteur then reproduced manufactured cultures of chicken cholera vaccines by growing the cholera bacillus at 42-43 degrees C. at which temperature the bacillus is not infectious. For a while Pasteur fantasized that the injection of an aged or weakened culture of cholera germs would not only protect an animal from a later infection of cholera, but would also offer protection from all other diseases. However, after months of enthusiastic but disappointing study of other diseases, he had to accept that there were limitations to his discovery. Injection of a weakened cholera germ would only give protection against cholera, no other diseases, but it might be worthwhile giving an injection of the weakened germs of other diseases to protect the individual against that particular disease.
Anthrax At this time, anthrax was a fatal disease of sheep and cattle, destroying the sheep industry and the economy of France. Pasteur was not the first scientist to examine the cause of anthrax, the German physician/scientist Robert Koch had already isolated the anthrax bacillus, previously identified by the French physician Davain, from infected spleens and showed that under resting conditions the bacillus formed long-lived spores. So Pasteur was aware of two things, anthrax was caused by a germ, and that as far as cholera was concerned, chickens could be protected against the disease. Could the same principle be used for protection against anthrax by inoculating an animal with an age-weakened culture of anthrax bacilli? Pasteur had to find a foolproof method of developing cultures of weakened anthrax bacilli which were too weak to kill or seriously incapacitate recipients of the injection but strong enough to provide immunity to a later injection of lethal anthrax bacilli. Although he found that usually he could give protection against a lethal injection of anthrax, occasionally the inoculation led to the death of the injected animal. Despite these occasional failures, in 1880, Pasteur prematurely announced that he had found a vaccine that could protect sheep and cattle from anthrax. This claim was so exciting to some and so unbelievable to others, that in 1881 he was challenged by the veterinarian Rossignol to provide a public demonstration of his anthrax vaccine. Pasteur readily agreed and the event was arranged to take place at Poilly-le-Fort on May 31st – June 2nd. On May 5th, 1881, Pasteur and his assistants inoculated twenty-four of forty-eight sheep, three of six cows and one of two goats with the cultures of weakened anthrax bacilli. The same animals were reinjected on May 17th. On May 31st, the pre-injected animals together with the uninjected animals were inoculated with a lethal culture of anthrax bacilli. To prove his point, all of the control animals must die and the vaccinated animals must live. Pasteur's colleagues were most concerned that he had agreed to the test, the challenge was critical and there was no room for error. On June 1st Pasteur received a message stating that some of the vaccinated sheep given the deadly culture the day before were already becoming ill. In a fury, he blamed his colleague Pierre Roux for performing the vaccinations in a sloppy, careless way and said he would not attend at Poilly-le-Fort the following day; Roux could go instead and accept the blame. He eventually calmed down and later that evening a telegram arrived announcing that all the vaccinated sheep were doing well, so Pasteur caught the train to Poilly-le-Fort. There was intense publicity, a reporter from the London Times sent back daily despatches and newspapers in France followed events with daily bulletins. There were crowds of onlookers, farmers, engineers, veterinarians, physicians, scientists and everyone greeted him with cheers when he arrived at the demonstration field. The trial was a complete success, twenty-two of the sheep that had not received the vaccine were dead, and the remaining two were dying, the cows and goat who had not received the vaccine lay dying or dead, but the vaccine-protected sheep, cows and goat were completely well. It was a magnificent triumph for Pasteur and the fame of his experiments spread through France, Europe and beyond. He became an international hero and in days thousands of sheep and cattle growers were clamouring for his magical anthrax vaccine and hundreds of vials of the vaccine were sent from his laboratory. (Within ten years a total of 3.5 million sheep and half a million cattle had been vaccinated with a mortality rate of less than 1%, saving the French economy at least 7 million francs). Note: Because of the frequent failures, Robert Koch, annoyed because Pasteur had never referred to his own seminal anthrax studies, publicly criticised both the cholera and anthrax studies. This lead to a bitter confrontation between the two, but despite Koch’s antagonism, Pasteur was able to repeat his Poilly-le-Fort demonstrations in Germany in 1882 with complete success. Following his successes with anthrax and fowl cholera diseases, over the next few years Pasteur identified and isolated the microbes for many other diseases, including swine erysipelas, childbirth fever and pneumonia.
Rabies In 1882, at the age of 60 and partially crippled by an earlier stroke that had weakened his left leg, Pasteur could have rested on his laurels and become an administrator. Instead he turned his attention to study the vicious and fatal disease of rabies, or hydrophobia as it is known. He had never forgotten seeing a child who had been attacked by a mad dog having heated flesh-burning irons applied to the wounds. Experimenting with rabid dogs presented a terrible danger to Pasteur and his assistants and initially, the only way of studying the disease was to put a normal dog with a rabid one so that the sick animal could transmit rabies to the fit animal by repeatedly biting it. Pasteur and Roux attempted to transfer infection by injecting healthy dogs with the saliva from rabid ones. The results were variable and unpredictable. Later, noticing that there was a time delay between a human or an animal being bitten, and the onset of the symptoms of rabies, and noting that the symptoms arose chiefly from a disturbed brain and spinal cord, Pasteur concluded that the infectious agent must travel via the peripheral nerves, concentrating eventually in the brain and spinal chord. Pasteur and his assistants had been working for some years on rabies, when by chance they acquired a dog that had become rabid after being bitten by another dog with rabies. For the first time, the dog recovered and when injected with fresh brain tissue from a rabid animal, which previously had led to rabies, the dog remained totally healthy. This was the opportunity Pasteur needed, he began to inject spinal-chord tissue from rabbits infected with rabies, but he allowed the extracts to weaken for a number of days in the now famous Roux Bottle before they were injected. A strip of spinal chord was suspended from a hangar in the center of the bottle containing a hole at the top of the bottle and one on the side. Air entered from the bottom opening, passed over a drying agent, and exited over the top. The longer the chord was dried, the less potent was the tissue in producing rabies. He inoculated dogs each day with spinal-chord extracts obtained from rabid rabbits, starting with an extract that had been dried and set aside for fourteen days, and then injecting on each succeeding day an extract which had been left for one day less. After many trials, he found that by the fourteenth day the dogs could receive fresh chord extract – which initially would have given them rabies - and they showed no sign of the disease. Forty dogs were successfully treated in this way. Unlike his impatient production of the anthrax vaccine, Pasteur proceeded cautiously with these experiments and three years after beginning his work he was reasonably sure that his discovery would provide complete protection against rabies. In 1885, following confirmation of his success in protecting animals from rabies and after much enthusiastic publicity, it was suggested that the discovery should be used on humans. Pasteur was not a doctor and was afraid that the treatment might go wrong. He insisted that many years additional research was necessary before the vaccine could be tried on humans. However, events made him act sooner. Two initial trials in humans, which he had conducted with the help of his physician colleagues, were indecisive. One patient was discharged from the hospital after only one injection and no-one knows what happened to him, the second patient was a girl suffering from an advanced stage of rabies and she died shortly after the procedure had started. Then, on July 6th, 1885, a nine year-old boy, Joseph Meister and his mother appeared at Pasteur's laboratory. The mother was distraught and begged Pasteur to save her son. Two days earlier the child had been bitten fourteen times on his arms, leg and thigh by a rabid dog and was so badly mauled that he could hardly walk and without treatment would certainly have died. Pasteur sent the boy to two physician friends who, when they saw the festering bite wounds, urged him to administer his therapy to the child. He reluctantly agreed to give Joseph daily injections of chord extract for thirteen days of continually increasing strength. It demanded great courage on Pasteur’s part, knowing as he did, that the extract given on the thirteenth day would have been lethal, if given on the first. Despite all his misgivings, he saved the child and news of his success spread throughout the civilized world. (When the Pasteur Institute was founded, Joe Meister became its custodian for the rest of his life). A few weeks later, nineteen Russian peasants who had been bitten two weeks earlier by a rabid wolf came to Pasteur asking for his therapy. He injected them twice a day for seven days and when they all survived, the Russian Czar was so appreciative that he sent Pasteur the diamond cross of Sainte Anne, together with 100,000 francs to be used for the construction of a Pasteur Institute. Following reports of his successful treatments, victims of dog and wolf bites from France, Russia and the United States rushed into his laboratory for treatment. The newspapers and the public followed the therapy and cures with great interest. Pasteur became a hero and a legend. The Pasteur Institute, funded by public and government subscriptions, was built in Paris, initially to treat victims of rabies who were coming to Pasteur’s laboratory. Later Pasteur Institutes were built, including three in the United States, to deal with human rabies and other diseases. Within several years, extracts of the vaccine were prepared by laboratories in many countries and death due to rabies almost disappeared. It is interesting to note that although in 1885 Koch had derided this method of prevention, within a year he also started to use Pasteur’s method of preparing the extract. Rabies was the last great research carried out by Pasteur. His health was failing and the paralysis of his left side, caused by the stroke he suffered at forty-six, made working in the laboratory increasingly difficult. In 1892, for his seventieth birthday, he was honoured with a special medal before a distinguished audience, including the President of the French Republic. He was too weak to give his own speech, so his son presented it for him. Pasteur died in 1895 after a series of additional strokes, holding a crucifix in one hand and his wife’s hand in the other. He was buried, a national hero, by the French Government and his funeral was attended by thousands of people. His remains were initially interred in the Cathedral of Notre Dame, but were transferred to a permanent crypt in the Pasteur Institute in Paris. (In a tragic footnote to history, Joseph Meister, the first person publicly to receive the rabies vaccine, served at the Pasteur Institute as Gatekeeper for many years. Forty-five years after his treatment for rabies he was ordered by the German occupiers of Paris to open Pasteur’s crypt. Rather than comply, he committed suicide.) Without doubt, Louis Pasteur was France’s most distinguished scientist, his work served as a springboard for branches of science and medicine such as stereochemistry, microbiology, bacteriology, virology, immunology and molecular biology. He was short-tempered, egotistical, and reluctant to give credit to his predecessors or his contemporaries, sometimes dishonest, a great showman and a bitter foe of physicians, but his discoveries have protected millions of people from disease through vaccination and pasteurization. His work was revolutionary in discovering the link between germs and disease and he led the way for Robert Koch to discover later how each type of germ caused a specific disease, and who established a complete germ theory of disease. Pasteur’s work is not just the sum of his discoveries; it also represents the revolution of scientific methodology. He placed, above all, two irrefutable rules of modern research; the freedom of creative imagination necessarily subjected to thorough experimentation, as he taught his disciples: "Do not put forward anything that you cannot prove by experimentation."

Louis Pasteur