Some of the flasks are still on a shelf at the Institut Pasteur in Paris. They contain nutrient broth prepared in the early 1860s, exposed to the open air ever since through a long curved glass neck, and after more than a century and a half nothing has grown in them.
They are the surviving apparatus from the experiment that ended a debate running back to Aristotle, and they are a better argument than anything Louis Pasteur wrote about it.
The Question
Spontaneous generation was the doctrine that living organisms can arise directly from non living matter. Maggots appear in meat, mice in stored grain, eels in mud. To an observer without a microscope, this is what the evidence looks like.
The doctrine had been steadily pushed back. In 1668 Francesco Redi covered jars of meat with gauze and showed that maggots appear only where flies can lay eggs. In the following century Lazzaro Spallanzani boiled sealed flasks of broth and found that nothing grew in them.
Spallanzani’s opponents had a serious counterargument. Sealing the flask excludes air, and if air contains some vital principle necessary for spontaneous generation, then his experiment does not test the doctrine, it merely suffocates it. Boiling might also destroy that principle. The dispute stalled on this point for a century.
By the middle of the nineteenth century the debate had a sharper edge because of what Leeuwenhoek’s microscopes had revealed: a world of organisms too small to see, present nearly everywhere, and appearing in any infusion left standing. Whether those organisms arose from the material itself was a question with consequences for medicine, brewing, food preservation, and the understanding of disease.
Pasteur Before Microbes
Pasteur began as a crystallographer. In 1848, aged twenty five, he examined crystals of a salt of tartaric acid and noticed that they came in two forms, mirror images of each other. He separated them by hand under a lens with a pair of tweezers and found that solutions of one rotated polarised light to the right and the other to the left, while a mixture was optically inactive.
This is the discovery of molecular chirality, and it remains one of the foundations of structural chemistry. It also shaped his later thinking, because the tartrate produced by fermentation contained only one form while the synthetic product contained both. Something about living processes generated asymmetry.
Appointed to Lille in 1854, in a region built on distilling, he was asked to investigate why some batches of beet alcohol spoiled. Examining the failed vats under the microscope, he found rod shaped organisms rather than the round yeast cells of the successful ones. Fermentation, he concluded, is not a purely chemical decomposition as Justus von Liebig maintained. It is the work of living microorganisms, and different organisms produce different results.
If fermentation and putrefaction are caused by living things, the obvious question is where those things come from.
The Swan Neck Flask
The Académie des Sciences offered a prize in 1860 for work settling the spontaneous generation question, largely because Félix Pouchet, a respected naturalist and director of the Rouen natural history museum, had published experiments he believed demonstrated it.
Pasteur’s solution was a piece of glassblowing. He prepared flasks of nutrient broth and drew their necks out into long tubes, bent downwards and then up again in an S shape, leaving the end wide open.
He boiled the broth to sterilise it and left the flasks open to the room.
Air passed freely in and out. Whatever vital principle the air might contain had unimpeded access. But dust and airborne particles, being heavier than air, settled in the bend of the neck and never reached the liquid.
Nothing grew. The broth stayed clear for months.
Then came the decisive step. Tilting the flask so that the broth ran into the curve of the neck and back, or simply snapping the neck off, produced growth within a day or two. The same broth, the same air, the same temperature. The only variable was whether the liquid had come into contact with the dust trapped in the bend.
The Mountain Flasks
Pasteur pressed the point with a second set of experiments designed to show that contamination varies with the dustiness of the air.
He prepared sealed flasks of sterile broth and carried them to different locations, breaking the seal to admit local air and resealing them. In the cellars of the Paris Observatory, where the air is still and clean, few flasks spoiled. In ordinary rooms, most did. On the Mer de Glace glacier on Mont Blanc, at around two thousand metres, only a small fraction of the twenty flasks he opened showed any growth.
The result is quantitative and it undercuts Pouchet’s position directly. Life does not appear at a rate determined by the nutritive richness of the medium. It appears at a rate determined by how much airborne material the medium is exposed to.
Pouchet, to be fair to him, had obtained growth in flasks opened on a Pyrenean glacier, and the discrepancy was probably caused by his use of hay infusions, which contain heat resistant bacterial spores that survive boiling. Neither man knew about spores in 1860. The commission of the Académie found for Pasteur, and Pouchet withdrew rather than repeat the trials under the commission’s conditions.
At a public lecture at the Sorbonne in 1864, Pasteur stated that the doctrine of spontaneous generation would never recover from the mortal blow of this simple experiment. He was right, and the flasks on the shelf continue to make the point.
What Followed
Establishing that microorganisms come from other microorganisms turned a philosophical dispute into a practical programme.
Preservation. If spoilage is caused by organisms arriving from outside, heating a product sufficiently and then protecting it from contamination will preserve it. Pasteur developed the process for wine in 1864, heating it briefly to around fifty five degrees, and it was later applied to beer and milk. Pasteurisation is his name attached to a public health measure that has saved an enormous number of lives.
Disease. If organisms cause fermentation and putrefaction, they may cause disease. Pasteur worked on the diseases of silkworms that were destroying the French silk industry, identifying the parasites responsible and devising a screening method. Joseph Lister in Glasgow read his work on putrefaction and reasoned that surgical wounds are infected the same way, introducing carbolic acid antisepsis in 1867 with dramatic effects on mortality. The connection between contamination and disease had already been established statistically, with no microscope involved, by John Snow’s map of a cholera outbreak in 1854.
Vaccination. In 1879, an assistant inoculated chickens with an old culture of the bacterium causing fowl cholera. The birds fell only mildly ill and then proved resistant to fresh, virulent cultures. Pasteur recognised that weakened organisms could produce immunity, generalising what Edward Jenner had achieved empirically with cowpox. He demonstrated an anthrax vaccine publicly at Pouilly le Fort in 1881, and in 1885 treated Joseph Meister, a boy bitten by a rabid dog, with an attenuated rabies preparation. The boy lived.
The Notebooks
Pasteur instructed his family not to release his laboratory notebooks, and they remained closed until 1971, when a descendant deposited them in the Bibliothèque Nationale.
Historians working through them, notably Gerald Geison, found that the public accounts were sometimes tidier than the laboratory record. In the anthrax demonstration, the vaccine actually used appears to have been prepared by a chemical method developed by a rival, Jean Joseph Toussaint, rather than the oxygen attenuation method Pasteur described. In the rabies case, the treatment given to Meister had been tested less extensively on animals than the published account implied.
This does not touch the validity of the results, which were confirmed by others many times over. It does complicate the heroic version, and it is a reasonably common pattern in nineteenth century science, where the published paper was understood as a demonstration of a conclusion rather than a diary of how it was reached.
Life Comes from Life
The principle Pasteur established, omne vivum ex vivo, is one of the load bearing statements of biology. It closed off a whole category of explanation and forced questions about origin, transmission and contamination to be answered concretely.
It also left one problem conspicuously open, as Pasteur himself acknowledged. If life only comes from life, the first appearance of life on Earth still requires an account, and that question moved out of everyday biology and into a separate field where it remains.
The experiment itself is worth remembering as a piece of design. Pasteur did not out argue his opponents. He identified the one objection that had kept the debate alive for a century, that sealing a flask changes the conditions, and built a piece of glass that removed it. The flask stays open. The air gets in. Nothing grows.
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