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Notes Rec. R. Soc. (2010) 64, 59–65
doi:10.1098/rsnr.2009.0029
Published online 26 August 2009
JOSEPH LISTER: FIRST USE OF A BACTERIUM AS A ‘MODEL ORGANISM’
TO ILLUSTRATE THE CAUSE OF INFECTIOUS DISEASE OF HUMANS
by
MELVIN SANTER*
Department of Biology, Haverford College, 370 Lancaster Avenue,
Haverford, PA 19041, USA
Joseph Lister’s goal was to show that a pure culture of Bacterium lactis, normally present in
milk, uniquely caused the lactic acid fermentation of milk. To demonstrate this fact he
devised a procedure to obtain a pure clonal population of B. lactis, a result that had not
previously been achieved for any microorganism. Lister equated the process of
fermentation with infectious disease and used this bacterium as a model organism,
demonstrating its role in fermentation; from this result he made the inductive inference
that infectious diseases of humans are the result of the growth of specific, microscopic,
living organisms in the human host.
Keywords: fermentation; infectious disease; model organism;
clonal population; pure culture
In the Transactions of the Pathological Society of London for the session 1877–78 there
appeared a paper by Joseph Lister FRS entitled ‘On the lactic fermentation and its bearing on
pathology’.1 Lister’s intent ‘was to obtain, if possible, absolute proof’2 that a pure culture of a
unique bacterium, which he designated Bacterium lactis, was responsible for the production
of lactic acid and the consequent curdling of milk. By demonstrating that a microscopic living
entity smaller than a yeast cell could cause fermentation, he was able to argue ‘that other
organisms may exist . . . smaller than the B. lactis’, and not readily visible in diseased human
tissues, could be the cause of infectious disease in humans.3 This paper was a landmark for
two reasons. It was the first example of the use of a bacterium as a model organism and also
for the invention of a procedure, now characterized as the limiting dilution method, for
isolating a specific bacterium in a pure form, providing a first case of bacterial cloning.4
The opening paragraph of Lister’s paper contains his goals and procedures:
A few years ago it would have seemed very improbable that the souring of milk should
have any bearings upon human disease: but all will now be ready to admit that the study
of fermentative changes deservedly occupies a prominent place in the minds of
pathologists. In order that any sure steps may be taken to elucidate the real nature of
the various important diseases which may be presumed to be of a fermentative nature,
*msanter@haverford.edu
59
This journal is q 2009 The Royal Society
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M. Santer
such as the specific fevers or pyemia, the first essential, as it appears to me, is that we
should have clear ideas, based upon positive knowledge, with regard to the more
simple forms of fermentation, if I may so speak—more simple because they can be
conducted and investigated in our laboratories.5
Lister implicitly recognized that it was not methodologically possible to demonstrate that
bacteria would induce an infectious disease in humans, although he believed that this was
the case, but it was feasible to establish that a specific ‘disease’ of milk could be
demonstrated by introducing the proper bacterium. This work was the first example of the
use of a bacterium as a model organism.
There is now an extensive literature documenting the history of the use of particular
organisms in biological research, as well as a literature that deals with the philosophical
issues raised by the use of model organisms.6 In 1877, however, did Lister have reason to
believe that the study of the cause of the lactic fermentation, his ‘simple fermentation’,
using a bacterium, would yield knowledge that would illuminate the cause of infectious
disease in humans?
There were several reasons for Lister to take this position despite the fact that there were
serious contending, incompatible, views about the cause of fermentation and infectious
disease. Lister could rely on the fact that the concept of fermentation and the cause of
infectious disease had been part of the same theoretical discourse since the seventeenth
century.7 Willis also explained the transmission of disease. From every body ‘effluvia of
atoms constantly fly away . . . of remarkable virtue and energy; these little bodies . . . retain
the contagion of pestilence. . . . With its ferment it imbues the next little bodies, and so
acquires new forces.’8
The relationship between fermentation and infectious disease was revisited in the
nineteenth century. The most influential proponent of a purely chemical theory of
fermentation and infectious disease was J. Liebig9 (1803 – 73), who rejected a living agent
for both these processes:
There is no opinion so destitute of a scientific foundation as that which admits, that
miasms and contagions are living beings, parasites, fungi, or infusoria, which are
developed in the healthy body, are there propagated and multiplied, and thus increase
the diseased action, and ultimately cause death.10
A theory of the cause of fermentation and putrefaction, which is utterly fallacious in its
fundamental principles, has hitherto furnished the chief support of the parasitic theory of
contagion.11
The opinions concerning the cause of putrefaction, which the adherents of the parasitic
theory have formed, are founded chiefly on observations which have been made on the
fermentation of yeast in the fermentation of wine and beer. But the investigation into
the nature of yeast is not yet completed.12
The chemical–mechanical theories of disease inherited from the late seventeenth and early
eighteenth centuries had not adequately explained how the putative disease agent seemed to
multiply as it passed from one host to another, nor could they provide an answer to the
question of specificity of a particular disease agent. Liebig had provided a ‘solution’ to
the problem of transmissibility and the attendant increase in the causative agent by turning
the process of fermentation into a ‘contagiousness of chemical action’,13 not an entirely
new principle but now based on the more substantial chemistry of the nineteenth century.14
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Joseph Lister’s use of a bacterium as a ‘model organism’
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The strong support for a biological agent as the cause of fermentation and infectious
disease emerged in the writings of Jacob Henle15 (1809 – 85), who relied on the
experimental work on yeast alcoholic fermentation by Theodore Schwann (1810– 82) and
Charles Cagniard Latour (1777– 1859) in the 1830s. The role of yeast in alcoholic
fermentation gained support from the later work of Louis Pasteur16 (1822 – 95).
Schwann and Cagniard Latour demonstrated microscopically the presence and
multiplication of yeast in a medium containing a mixture of sugar and nitrogen with the
concomitant production of alcohol and carbon dioxide. Schwann concluded in his classic
paper on the cell theory17 that the yeast that caused fermentation was a prime example of
a cell’s doing what he stipulated was essential for the life of a cell; that is, performing
processes that lead to cell division. Yeast represented for Schwann the building blocks
from which all organized living bodies are constructed.18 Cells have the ability to make
more cells and consequently ‘we must ascribe to all cells an independent vitality.’19 This
vitality was based on ‘fundamental powers’20 that come from the combination of cell
components, and the ability of cells to perform ‘metabolic phenomena’.21
To present a concrete example of the metabolic power of cells, Schwann wrote about
‘vinous fermentation’:
I could not avoid bringing forward fermentation as an example, because it is the best
known illustration of the operation of the cells, and the simplest representation of the
process which is repeated in each cell of the living body.22
He continued:
We have every conceivable proof that the fermentation-granules are fungi. Their form is
that of fungi; . . . they grow, like fungi . . . . Now, that these fungi are the cause of
fermentation, follows, first from the constancy of their occurrence during the process;
secondly, from the cessation of fermentation under any influences by which they are
known to be destroyed, especially boiling heat, arseniate of potass, &c; and, thirdly,
because the principle which excites the process of fermentation must be a substance
which is again generated and increased by the process itself, a phenomenon which is
met with only in living organisms.23
Henle equated the ability of yeast to multiply to the apparent capacity of pox material to do
the same. ‘An atom of pox poison can produce rash over the entire body. The pus from each
of these pustules is again capable of infecting a new organism.’24
Henle recognized that a living agent theory of disease could not be demonstrated by
simple observation of the presence of an agent when the disease is present, because it
may be there accidentally. He suggested that it could be demonstrated by the deliberate
use of a pure sample of the agent administered to an appropriate host. However, such an
experiment was not possible because methods were not available to obtain a pure sample
of a living microscopic agent.
Lister had accepted the contested principle that biological agents cause both infectious
disease and fermentation. Pasteur and others, he wrote, had already provided the work,
tending to prove that all true fermentations of organic liquids are due to the development
of organisms within them. . . . But opinion is by no means universal in our profession.25
Consequently Lister proceeded to study a ‘model system’, a ‘simple’ system, which is the
lactic fermentation already studied by Pasteur in 1857.14
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M. Santer
The souring – curdling of milk fulfilled Lister’s requirements for a simple system. The
effect on milk is visible and is therefore easily determined. The milk solidifies quickly,
and souring can be tested easily. The event happens very rapidly. The process is unique
to the lactic acid bacterium and consequently it is unlikely that accidental contamination
will interfere with the experiment.26
The experiment depended on the ability of a pure culture of Bacterium lactis (so named by
Lister) to cause the lactic fermentation; Lister would therefore have to provide a method to
achieve this pure population. To do this he invented what is now called the ‘limiting
dilution method’.27 Lister devised this method to solve the problem of contamination. In his
original milk sample, which was the source of the bacterium producing lactic acid, the
predominant organisms were of the B. lactis type, but there was present a significantly
smaller number of another, morphologically different, microscopic organism. Lister
adopted the strategy that simply diluting the sample would eliminate the minority organism
and at the highest dilution could yield a single B. lactis bacterium in a specific volume.
THE
EXPERIMENT
Lister had a culture of B. lactis and had determined microscopically that the cells are present
in greater number than those of another morphologically different organism. He diluted the
sample to a point ‘calculated . . . to contain on the average a single Bacterium lactis’28 in the
volume he would use to initiate his experiment. Lister prepared 16 tubes of milk that had
been pre-heated to 2108F, thus eliminating all existent bacteria. He added to tubes 1 –10 a
volume containing one bacterium. To tubes 11 – 15 he added twice the volume, or two
bacteria per tube. For the last tube, number 16, he used double the volume again, to add
four bacteria to this tube.
The milk of tube number 16 was curdled in 3½ days, and as it did in tubes 11– 15. In all of
tubes 1 – 10 at 3.5 days the milk was still fluid, but during the next day five tubes became
curdled at different times. Five tubes remained permanently fluid even four months later
and did not contain any bacteria. Thus it seemed that whenever at least one bacterium
was introduced into a milk sample a lactic fermentation occurred.
It is obvious that some tubes of milk did not show fermentation; according to Lister, this
proved ‘the important truth’ that the agent responsible for the fermentation was particulate
and was a bacterium, for if the ferment had been soluble every sample of inoculum
would have contained the ferment.29 In addition it was ‘utterly inconceivable’ that there
was an inanimate factor, a ‘so-called chemical ferment’30 responsible for the souring of
milk, because they would have had to be of the same number as the bacteria and would
have had to be introduced with the bacteria in all tubes in which souring occurred and
excluded from all tubes in which fermentation did not take place.
The general applicability of this procedure to isolate and characterize microbial agents
was demonstrated by Lister in his experiments using highly diluted tap water to inoculate
boiled milk. Here again, some tubes remained free of fermentation but others showed
‘different kinds of fermentation’ that demonstrated again the particulate nature of the
ferments and, importantly, showed that water
contains several different kinds of ferments, which though generally confused through
being mixed up together, declare their individual peculiarities when isolated by this
method of separation.31
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Joseph Lister’s use of a bacterium as a ‘model organism’
63
CONCLUSION
In a re-evaluation of Lister’s contribution to theories of infectious disease, it is
acknowledged that he had a germ theory of disease but ‘not a theory of specific
organisms causing specific diseases’.32 This characterization was based on Lister’s paper
in 1873 that bacteria may be derived from fungi.33 If this had been the case—that he
believed that microorganisms did not have a fixed identity—then it would have followed
that there was no one-to-one causal relationship between a microorganism and a disease.
Nevertheless, the lactic acid fermentation paper in 1877 – 78 offers another perspective,
because it demonstrated that a specific organism caused a specific phenomenon. The
evidence that Lister provided is unambiguous. There are no bacteria in the air, outside of
stalls where milk is obtained from cows, that are lactic-acid-producing bacteria. There are
no microorganisms in the air that are so plastic that they can transform into B. lactis,
although there are airborne organisms that can grow in milk.
If bottled milk is exposed to air under various conditions, Lister wrote,
you will be sure to have organisms develop . . . of the nature of filamentous fungi and
bacteria and you will see fermentative changes ensue, . . . you will not see the
coagulation and souring of the lactic fermentation, nor will you find under the
microscope the peculiar organism to which I have given the name of Bacterium lactis.
. . . The filamentous fungi most frequently found in milk . . . include Penicillium
glaucum, Aspergillus glaucus, and two forms of Mucor.34
It is reasonable to conclude that for Lister the cause of infectious diseases and fermentations
was living, specific agents that found their particular ecological niche in bodies or liquids
and in the course of their growth performed the chemical processes leading to
fermentation or disease. The study of the lactic fermentation by B. lactis led him to this
position.
ACKNOWLEDGEMENTS
I thank Dora Wong of the Haverford College Science Library for her constant help, and Karl
Johnson of the Haverford Biology Department for his willingness to discuss history and for
his perceptive comments.
NOTES
1
2
3
4
5
6
J. Lister, ‘On the lactic fermentation and its bearing on pathology’, Trans. Pathol. Soc. Lond. 29,
425 –467 (1877–78).
Ibid., p. 451.
Ibid., p. 463.
R. A. Weiss, ‘Robert Koch: the grandfather of cloning?’, Cell 123, 539–542 (2005). Weiss
points out ( p. 540): ‘In 1878, Joseph Lister described the limiting dilution method in liquid
culture for isolating bacteria in pure form’. This procedure and the plating method of Koch
were the first examples of bacterial cloning.
Lister, op. cit. (note 1), p. 425.
W. C. Summers, ‘How bacteriophage came to be used by the phage group’, J. Hist. Biol. 26,
255 –267 (1993): M. Weber, ‘Model organisms: of flies and elephants’, in Philosophy
of experimental biology (Cambridge University Press, 2004), pp. 154–187; P. Meneely,
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7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
M. Santer
‘Model organisms’, in Advanced genetic analysis: genes, genomes, and networks in eukaryotes
(Oxford University Press, 2009), ch. 2, pp. 29– 90.
T. Willis, A medical –philosophical discourse of fermentation (H. Clark, London, 1684).
Ibid., p. 105. It is both interesting and fascinating that Willis equates the alteration of the
consistency of milk when it undergoes souring to the character of blood under conditions of
infection ( p. 56): ‘the blood is subject to alteration, which happens . . . most often to Milk . . .
from a Morbific cause.’ Two hundred and nineteen years later Lister provided a solution to
the cause of the souring of milk, comparing it to an infectious disease.
J. Liebig, Animal chemistry in its application to physiology and pathology, 3rd edn
(ed. W. Gregory) (John Wiley, New York, 1852).
Ibid., p. 127.
Ibid., p. 143.
Ibid., p. 145.
Ibid. Liebig suggested that the fermentation associated with the ‘yeast plant’ by its growth
produces a material capable of acting on sugar in the same manner as emulsin acts on
amygdalin, or diastase on starch, a catalytic phenomenon described in detail on pages 131–137.
J. R. Partington, A history of chemistry (4 volumes, 1961–70), vol. 4 (Macmillan, London,
1964), pp. 261 –264 and 302.
J. Henle, ‘On miasmata and contagia’ (transl. G. Rosen), Bull. Inst. Hist. Med. 6, 907–983
(1938).
L. Pasteur, ‘Mémoire sur la fermentation appelée lactique’, C. R. Acad. Sci. 45, 913– 916 (1857);
‘Mémoire sur la fermentation alcoolique’. Ann. Chim. Phys. (3) 58, 323– 426 (1860).
T. Schwann, Microscopical researches into the accordance in the structure and growth of
animals and plants (tr. H. Smith) (Sydenham Society, London, 1857).
Ibid., p. 191.
Ibid., p. 192.
Ibid., p. 193.
Ibid., p. 193.
Ibid., p. 197.
Ibid., p. 197.
Henle, op. cit. (note 5), p. 921.
Lister, op. cit. (note 1), p. 426. The contentious nature concerning the composition and origin of
the agent that caused infectious disease and fermentation is extensively covered by M. Pelling in
Cholera, fever and English medicine 1825–1865 (Oxford University Press, 1976); J. K. Crellin,
‘The dawn of the germ theory: particles, infection and biology’, in Medicine and Science in the
1860s (ed. F. N. L. Poynter), pp. 57 –76 (Wellcome Institute, London, 1968). The period of the
1870s in England is also discussed by J. E. Strick, Sparks of life: Darwinism and the Victorian
debates over spontaneous generation (Harvard University Press, Cambridge, MA, 2000), chs 5
and 6.
Lister, op. cit. (note 1), p. 437; Lister had previously written: ‘milk supplied for domestic use
will turn sour in summer weather within twenty-four hours, yet of all the many instances in
which I have observed alterations in milk caused by organisms introduced through
atmospheric exposure, in no single case did the true lactic acid fermentation occur’: ‘A
further contribution to the natural history of bacteria and the germ theory of fermentative
changes’, Q. J. Microsc. Sci. 13, 397 (1873). In Pasteur’s paper several products were
produced in addition to lactic acid, because there was more than one microorganism growing
in the milk. In short, there was not a pure culture.
C. Henry, J. Marbrook, D. C. Vann, D. Kodlin and C. Wofsy, ‘Limiting dilution analysis’, in
Selected methods in cellular immunology (ed. B. B. Mishell and S. M. Shiigi), pp. 138–152
(W. H. Freeman, San Francisco, 1980).
Lister, op. cit. (note 1), p. 451.
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Joseph Lister’s use of a bacterium as a ‘model organism’
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30
31
32
33
34
65
Lister, op. cit. (note 1), p. 442.
Lister, op. cit. (note 1), pp. 454 –455.
Lister, op. cit. (note 1), p. 442.
C. Lawrence and R. Dixey, ‘Practising on principle: Joseph Lister and the germ theories
of disease’, in Medical theory, surgical practice: studies in the history of surgery
(ed. C. Lawrence), pp. 153 –215 (Routledge, London, 1992), at p. 179.
J. Lister, ‘A further contribution to the natural history of bacteria and the germ theory of
fermentative changes’, Q. J. Microsc. Sci. 13, 380–408 (1873), at p. 381.
Lister, op. cit. (note 1), p. 439.