3. Origin of Bacteria and of Torulæ by Archebiosis.
The evidence on this part of the subject is, I think, sharply defined and conclusive. Simple experiments can be had recourse to, which are not admissible in the discussion of the question as to the origin of Bacteria and Torulæ by Heterogenesis. There, we wish to establish the fact that living matter is capable of undergoing a certain metamorphosis, and consequently, we must deal with living matter. Here, however, with the view of establishing the fact that living matter can arise de novo, if we are able, shortly after beginning our experiment, to arrive at a reasonable and well-based assurance that no living thing exists in the hermetically sealed experimental vessel—if the measures that we have adopted fully entitle us to believe that all living things which may have pre-existed therein have been killed—we may feel pretty sure that any living organisms which are subsequently found, when the vessel is broken, must have originated from some re-arrangements which had taken place amongst the not-living constituents of the experimental solutions, whereby life-initiating combinations had been formed.
The possibility of the de novo origination of Bacteria, Torulæ, and other such organisms, is one which is intimately associated with the doctrine as to the cause of fermentation and putrefaction. With regard to the almost invariable association of such organisms with some of these processes, almost all are agreed. There is, moreover, a very frequent association of particular kinds of organisms with particular kinds of fermentation. Hence the assumption is an easy and a natural one to many persons, that the organisms which are invariably met with in some cases are the causes of these fermentations,[9] although it is quite obvious that the facts on which this view is based, are equally explicable on the supposition that the organisms are concomitant results or products (due to new chemical combinations) of the fermentative changes. In the one case the fermentative changes are believed to be initiated by the influence of living organisms; and those who regard living things as the only true ferments, for the most part also believe that living things are incapable of arising de novo. They think that those organisms which serve to initiate the changes in question, have been derived from a multitudinous army of omnipresent atmospheric germs, which are always ready, in number and kind suitable for every emergency. This is the doctrine of M. Pasteur and others. On the other hand, fermentations and putrefactions may be regarded as sets of chemical changes, which are apt to occur in organic and other complex substances—these changes being due either to the intrinsic instability of the body which manifests them, or to molecular movements communicated to it by a still more unstable body. Baron Liebig says:—“Many organic compounds are known, which undergo, in presence of water, alteration and metamorphosis, having a certain duration, and ultimately terminating in putrefaction; while other organic substances that are not liable to such alteration by themselves, nevertheless, suffer a similar displacement or separation of their molecules, when brought into contact with the ferments.”
Each substance belonging to the first class, would be at the same time, therefore, both ferment and fermentable substance; whilst a small portion of such substance, when brought into contact with a less unstable substance, might induce such molecular movements as to make it undergo a process of fermentation. With regard to the cause of such induced fermentative changes, Gerhardt[10] says, in explaining Liebig’s views:—“Every substance which decomposes or enters into combination is in a state of movement, its molecules being agitated; but since friction, shock, mechanical agitation, suffice to provoke the decomposition of many substances (chlorous acid, chloride of nitrogen, fulminating silver), there is all the more reason why a chemical decomposition in which the molecular agitation is more complete, should produce similar effects upon certain substances. In addition, bodies are known which when alone are not decomposed by certain agents, but which are attacked, when they exist in contact with other bodies incapable of resisting the influence of these agents. Thus platinum alone does not dissolve in nitric acid, but when allied with silver, it is easily dissolved; pure copper is not dissolved by sulphuric acid, but it does dissolve in this when it is allied with zinc, &c. According to M. Liebig it is the same with ferments and fermentable substances; sugar, which does not change when it is quite alone, changes—that is to say ferments—when it is in contact with a nitrogenous substance undergoing change, that is, with a ferment.”
Thus, in accordance with this latter view, living ferments are not needed—mere dead, organic or nitrogenous matter suffices to initiate the processes in question.[11] Those who hold this opinion may or may not believe that organisms are capable of arising de novo;[12] though there can be little doubt that a belief in the truth of such a doctrine does, almost inevitably, entail a belief in the de novo origination of living things. No one who has looked into the evidence, doubts the fact of the association between some of these processes and the presence of organisms; the only question is, as to the relation in which they stand to one another. If organisms are not the causes of those fermentative changes with which they are invariably associated, then they are, in all probability, the results of such changes; and they must certainly have been produced de novo if it can be shown that fermentation or putrefaction may take place under the influence of conditions which make it certain that pre-existing living organisms could have had nothing to do with the process.
Now, in order to lend some air of probability to the former hypothesis, concerning the necessity for the existence of living ferments, it was incumbent upon its supporters to endeavour to show that the air did contain such a multitude of “germs,” or living things, as were demanded by the requirements of their theory. Spallanzani and Bonnet had, as far as the imagination was concerned, done all that was necessary. They had proclaimed the universal diffusion of “germs” of all kinds of organisms throughout the atmosphere—which were ready to develop, whenever suitable conditions presented themselves. So far, however, this was but another hypothesis. To establish the doctrine that fermentation cannot take place without the agency of living ferments, we cannot receive hypotheses in evidence: facts are needed. These, no one attempted to supply in an adequate manner[13] anterior to the investigations of M. Pasteur. Speaking of his researches, even M. Milne-Edwards says,[14] “Previous to this time, the existence of reproductive particles, or infusorial germs in the atmosphere was nothing more than a plausible hypothesis, put forward in order to explain the origin of such creatures in a manner conformable with the general laws of reproduction; but it was only a mere supposition, and no one had been able actually to see or to handle these reproductive corpuscles.”
We have to look, therefore, to M. Pasteur’s investigations, and to others which may have been since conducted, for all the scientific evidence in support of what has been called the “Panspermic hypothesis.”
By an ingenious method of filtration, which is fully described in his memoir,[15] M. Pasteur separated from the air that passed through his apparatus the solid particles which it contained. This search convinced him that there were, as he says, “constantly in ordinary air a variable number of corpuscles whose form and structure declare them to be organized.” Some of these, he thinks, resemble the spores of fungi, and others the ova of ciliated infusoria, though he adds:—“But as to affirming that this is a spore, much less the spore of any definite species, and that one is an egg, and belonging to such an infusorium, I believe that this is not possible.” He limits himself, in fact, to the statements, that the corpuscles which he found, were (in his opinion) evidently organized; that they resembled in form and appearance the germs of the lower kinds of organisms; and that, from their variety in size, they probably belonged to many different sorts of living things. Even here, therefore, we have to do with the impressions of M. Pasteur, rather than with verified statements. All that has been established by his direct investigation as to the nature of the solid bodies contained in the atmosphere is this: that the air contains a number of round or ovoidal corpuscles, often quite structureless, which he could not distinguish from the spores of fungi[16]—some of which, being about the right size, were round or ovoidal, and structureless. In addition, however, it has been shown that the air contains other rounded corpuscles which are similarly structureless, though composed of silica or starch. It may therefore be asked, in the first place, whether the conclusion is a sufficiently safe one that many of the corpuscles found by M. Pasteur were spores of fungi; and in the next place, supposing this to have been established, whether such spores were living or dead. These questions would have been answered satisfactorily if M. Pasteur could state that he had actually watched the development of such corpuscles, in some suitable apparatus, into distinct organisms. But any such development, he distinctly states, he never witnessed. He says[17]:—“What would have been the better and more direct course would have been to follow the development of these germs with the microscope. Such was my intention; but the apparatus which I had devised for this purpose not having been delivered to me at a convenient time, I was diverted from this investigation by other work.” The evidence which he does adduce, in subsequent portions of his memoir, in order to prove that some of these corpuscles were really “fertile germs,” is almost valueless, because all the facts are open to another interpretation, which is just as much, nay, even more, in accordance with Baron Liebig’s than with his own doctrine of fermentation.
But another most important consideration presents itself. M. Pasteur’s researches as to the nature of the dust contained in the atmosphere enable him to say nothing concerning the presence of Bacteria, although he himself admits that these are generally the first organisms which display themselves in fermentations or putrefactions, and that in a very large majority of the cases in which fermentation occurs in closed vessels they are the only organisms which make their appearance.[18] And yet, notwithstanding these facts, M. Pasteur says, in reference to the common form of Bacterium:—“This infusorial animal is so small that one cannot distinguish its germ, and still less fix upon the presence of this germ, if it were known, amongst the organized corpuscles of the dust which is suspended in the air.”
Here, then, we have a confession from M. Pasteur himself, that all evidence fails, where it is most wanted, in support of his hypothesis.
If a large number of fermentations begin with the presence of Bacteria as the only living things, and if in a number of cases no other organisms ever occur, it is useless to adduce as evidence, in proof of the view that fermentations are always initiated by air-derived organisms, the fact that certain corpuscles (supposed to be spores of fungi) are recognizable in the atmosphere—capped by the distinct statement[19] that Bacteria or their germs are not recognizable. If Bacteria are not recognizable in the atmosphere, what scientific evidence is there that the fermentations in which these alone occur are initiated by Bacteria derived from the atmosphere, or from certain imaginary Bacteria germs,[20] which we are supposed to be unable to distinguish? M. Pasteur may, moreover, be reminded that when he resorts to the supposition of Bacteria possessing “germs” which are indistinguishable, he is again resorting to hypothesis rather than to fact, in order to prove the truth of the particular doctrine of fermentation which he advocates. Bacteria are known to reproduce and multiply only by a process of fission; each of the parts into which they divide being nothing more than a part of the original Bacterium, and therefore endowed with similar properties of resisting heat, desiccation, and other agencies. Any resort to invisible germs to account for the multiplication of Bacteria, which are known to reproduce freely in other ways, is obviously not permissible, unless such postulation be more or less necessitated by the occurrence of facts otherwise inexplicable.
Although, therefore, no direct evidence has been adduced tending to show that Bacteria are present in the atmosphere, even if this evidence had been forthcoming, it would have been necessary, in reference to M. Pasteur’s hypothesis, for it to be supplemented by further evidence to the effect that Bacteria were well capable of resisting such an amount of desiccation as must have been involved by their presence for an indefinite time in the atmosphere even of the hottest and driest regions of the earth. For, organic substances in solution do not only putrefy in moist weather or moist climates; they putrefy most rapidly and surely when the temperature is high, and quite irrespectively of the amount of moisture contained in the atmosphere. A property of resisting the effects of desiccation—the possession of which, by Bacteria, is so necessary for the truth of M. Pasteur’s argument—ought to have been shown by scientific evidence to be a real attribute of such organisms; though it seems, on the contrary, to have been assumed to exist, with almost equal readiness by both parties, in the controversies concerning the possibility of “spontaneous generation.” This error may be ascribed to the misguiding influence of a treacherous analogy. Whilst it may be true that certain seeds and spores, and also that Rotifers, “Sloths,” and some Nematoids are capable of resisting the influence of a prolonged exposure to desiccating influences, it may well be asked, whether the same fact necessarily holds good for organisms such as Bacteria, which have no chitinous or other envelopes to protect them, and which are merely minute fragments of naked protoplasm. Having elsewhere[21] shown how far presumptions had stolen a march upon established facts, in reference to the supposed possession of a similar property by the Free Nematoids, my eyes were opened to the reality of this uncertainty with regard to Bacteria. It is, however, no easy matter definitely to prove or to disprove the possession of this property by organisms so minute as Bacteria, and therefore so difficult to identify. If dried Bacteria are added to a drop of a suitable solution—similar to that in which they had been bred—it soon becomes quite impossible to distinguish those which have been added from those which arise in the fluid. Taking into consideration the fate of other simple organisms, however, it is by no means improbable that they should be killed even by a short desiccation. I have found, for instance, that desiccation for half-an-hour in a room at a temperature of 65° F. suffices to kill all the larger, naked, lower organisms with which I have experimented—including long Vibrios, Amœbæ, Monads, Chlamydomonads, Euglenæ, Desmids, Vorticellæ and all other Ciliated Infusoria.
But, certain indirect evidence seems to speak most authoritatively against the supposition that the air contains any notable quantity of living Bacteria, or Bacteria germs, whether visible or invisible. I have always found that a simple solution of ammonic tartrate, which has been placed—without previous boiling—in a corked bottle of greater capacity, will become turbid in two or three days, owing to the presence of myriads of Bacteria; whilst a similar solution, previously boiled, may remain for ten days, three weeks, or more, without showing the least trace of turbidity, although the open neck of the bottle or flask in which it is contained, may be covered only by a loose cap of paper. And yet, at any time, in order to make this fluid become turbid in from 24 to 48 hours, all that one has to do is to bring it into contact with a small glass rod which has just been dipped into a solution containing living Bacteria.[22]
If we find that an eminently inoculable fluid will remain for two or three weeks, or perhaps more, in contact with the air without becoming turbid, though it will always become turbid in two or three days if brought into contact with living Bacteria, what can we conclude, but that living Bacteria are not very common in the atmosphere? These most striking facts can be easily verified by other observers.
Thus we find ourselves, at present, in this position. After all that has been said and done to prove the wonderful prevalence of “germs” in the atmosphere, we are really still in the region of hypothesis—no further advanced than we were in the time of Bonnet and of Spallanzani, so far as it concerns the organisms which are all important—Bacteria. Neither these nor their germs have been shown to exist in any recognizable abundance in the atmosphere, and yet in most fermentations they are the first organisms which display themselves; whilst in many such fermentations Bacteria alone occur. Nay more, even were they present in any great abundance, there is some reason to believe that the majority of them would exist as mere dead, organic particles—because Bacteria are more than likely to be unable to resist anything like an extreme or prolonged exposure to desiccating influences.
The first and essential data in support of M. Pasteur’s hypothesis must, therefore, be regarded as entirely unproved in respect to Bacteria—which are the most important of all organisms, in relation to the cause of fermentation and putrefaction.
Without the aid of elaborate experiments, however, the evidence which the microscope can supply is capable of leading us to the conclusion that such search for atmospheric Bacteria germs, was comparatively useless. If it can be shown that Bacteria can arise in a fluid independently of visible germs, then, obviously, any inquiries as to the nature of the visible contents of the atmosphere, can have only a very indirect bearing upon the question as to the mode of origin of these organisms. And yet by the aid of the microscope, as I have elsewhere stated, one can watch the appearance of almost motionless specks, more or less uniformly diffused through a motionless film of fluid, and can see them gradually develop into moving Bacteria or into Torulæ. So that, where no visible germs previously existed, visible particles of living matter develop, and more or less rapidly grow into distinct Bacteria. This may be best seen in a drop of a fresh and very strong turnip infusion, which has been filtered several times through the finest paper. The drop, placed in a live-box, should be flattened into a thin film by the application of the cover.
Thus protected, evaporation takes place very slowly, and with the live-box resting on one of Stricker’s hot-water plates, at a temperature of 85° to 90° F., and the latter upon the stage of the microscope, one can easily select a portion of the field in which either no particles or only a countable number exist. If, therefore, around and between any mere granules which may pre-exist, or in a clear space, one gradually sees in the course of two or perhaps three hours, a multitude of almost motionless specks (at first about 1/100000″ in diameter) in positions where no such specks previously existed; and if these specks may be seen gradually to increase in size and develop into Bacteria and Torulæ, then, at all events, we are able to say that these organisms can be developed without pre-existing visible germs, and we have just the same amount of actual evidence for believing that they have been formed de novo, as we should have for believing that crystals had been formed de novo, if we had seen them appearing under our eyes in the same manner. Whether they really arise after the fashion of crystals, without the aid of pre-existing though invisible germs, is a matter which can only be settled inferentially, by a subsequent resort to strict methods of experimentation.
Seeing however, that we are able, with the aid of the microscope alone, to demonstrate that Bacteria and Torulæ can develop in situations where no visible germs had previously existed, it is useless, as I have said before—so far as the question of their mode of origin is concerned—to search the atmosphere to ascertain what visible germs it may contain. If some Bacteria and Torulæ arise from germs at all, it must be from germs which are invisible to us. The finding of visible germs in the atmosphere can, therefore, only have an indirect bearing upon the solution of the problem. Since it can be shown that some visible spores and ova exist in the atmosphere, this affords a certain amount of warrant for the supposition that invisible, living, reproductive particles may also exist—more especially if the existence of an amount of organic matter, which is ordinarily invisible, can be revealed in the air, by the agency of the electric beam, or by any other means.
Nothing can be more illegitimate, however, in the way of inference, than the assumption at once indulged in by Prof. Tyndall and others (who might have been expected, by their previous scientific work, to have learned more caution) that this impalpable organic dust was largely composed of impalpable germs. Yet, without a shadow of proof, without even an attempt to prove it, the air was for a time represented to be a mere stirabout, thick with invisible germs. The briefest reflection, however, upon the probabilities of the case, should have sufficed to suggest a totally different interpretation. The surface of the earth is clothed with living things of all kinds, animal and vegetal, which are not only continually throwing off organic particles and fragments during their life, but are constantly undergoing processes of decay and molecular disintegration after their death. The actual reproductive elements of these living things are extremely small in bulk, when compared with the other parts which are not reproductive, and although Bacteria and Torulæ do exist abundantly, and do materially help to bring about some of the decay in question, yet their bulk, also, is extremely small in comparison with the amount of organic matter itself that is continually undergoing disintegration of a dry kind, in which Bacteria and Torulæ take no part. When, moreover, it is considered that in the neighbourhood of populous cities (the air of which alone exhibits this very large quantity of impalpable, mixed with palpable, organic dust), there is constantly going on a wear and tear of the textile fabrics and of the organic products of various kinds which are daily subservient to the wants of man; and that the chimneys of manufactories and dwelling-houses are also continually emitting clouds of smoke thick with imperfectly consumed organic particles, some idea may be gained of the manifold sources whence the organic particles and fragments found in the atmosphere may emanate, and also as to what proportion of them is likely to be composed of living or dead reproductive elements, or “germs.”
Thus, then, so far as the two rival doctrines of fermentation are concerned, the investigation of the nature of the solid particles contained in the atmosphere has revealed facts which are thoroughly in harmony with all the requirements of Liebig’s physical theory, though it has almost utterly failed to give anything like a scientific basis to the vital theory of Pasteur. So far from being able to show that living Bacteria (which are the first and oftentimes the only organisms concerned in many processes of fermentation and putrefaction) are universally diffused through the air, Pasteur admits that these cannot be detected, and that their “germs” are not recognizable.
If, therefore, M. Pasteur still maintains the truth of his theory, it should be distinctly understood that it rests originally, not upon established facts, but upon a mere hypothesis—the hypothesis that the air teems with multitudes of invisible Bacteria germs. He is driven to such a doctrine, not only by his own confessions concerning Bacteria, but also by the microscopical evidence to which I have referred.
So that in explaining the results of any experiments made with the view of throwing light upon the cause of fermentation or putrefaction, it is especially necessary to bear in mind two considerations:—
I. That dust filtered from the atmosphere cannot be proved to include living Bacteria; though it is known to contain a multitude of organic particles which may be capable in the presence of water, in accordance with Liebig’s hypothesis, of acting as ferments.
II. It must also be recollected that, in the opinion of many, Life represents a higher function which is displayed by certain kinds of organic matter; and that this higher function may be deteriorated or rendered non-existent by an amount of heat which might not be adequate to decompose the organic matter itself.
It is all the more necessary to call attention to these two considerations, because M. Pasteur invariably speaks as though it had been established that the air contains multitudes of living Bacteria, when, really, he had only proved that the air contains a number of corpuscles resembling spores of fungi, &c. And, as I have already intimated, the existence of spores of fungi in the atmosphere, however well established, is of little or no importance as an explanation of the cause of a very large number of fermentations. Their presence is even of still less importance, owing to the fact of the co-existence with these fungus-spores, of multitudes of organic fragments, which—in accordance with the views of Liebig, Gerhardt, and other chemists—are capable of acting as ferments. To this latter consideration M. Pasteur never even alludes when he speaks (loc. cit. p. 40) of his “ensemencements,” and of other experiments which are equally, or even more, capable of being interpreted in accordance with Liebig’s views than with his own.
Bearing these considerations in mind, we shall be in a better position to enquire into the real interpretation that may be given to many of M. Pasteur’s results, and into the question as to how far the facts which he records are favourable to his own, or to the adverse doctrine concerning the causes of fermentation.
In the memoir so often alluded to on “The Organized Corpuscles which exist in the Atmosphere,” M. Pasteur adduced various kinds of evidence, tending, as he thought, to show that the first Bacteria which make their appearance in putrefying or fermenting solutions, have been derived from living Bacteria or their “germs,” which pre-existed in the atmosphere.
Some of the experiments by which he endeavoured to substantiate this position were of a very simple nature. Their narration attracted much attention at the time, as it was supposed that by their means M. Pasteur had—as he professed—conclusively shown the erroneousness of the views of those who believed in what was called “spontaneous generation.” These experiments were soon repeated by other observers, who, using different fluids, obtained quite opposite results. Thus it became obvious to impartial critics, that whilst the means adopted by M. Pasteur might be adequate to check the processes of fermentation or putrefaction in certain fluids, they were quite powerless to effect this when many other fluids were employed.
These particular experiments, however, still seem to exercise a very great influence on the minds of many in this country, who are either unaware of, or disbelieve in, the possibility of obtaining opposite results.
The chapter in which M. Pasteur detailed these experiments is thus entitled:—“Another very simple method of demonstrating that all the organized products of Infusions (previously heated) owe their origin to the corpuscles which exist suspended in the Atmosphere.” Whilst claiming to have already rigorously established the validity of this conclusion by the experiments described in previous chapters, M. Pasteur adds:—“If there remained the least doubt on this subject, in the mind of the reader, it would be dissipated by the experiments of which I am now about to speak.” (p. 66.)
Sweetened yeast-water, urine, infusions of pear and of beetroot, were placed in flasks with long necks, variously drawn out and bent. The flasks were subsequently treated as follows. M. Pasteur says:—“I then raise the liquid to the boiling-point for several minutes until steam issues abundantly from the extremity of the drawn-out neck of the flask, which is permitted to remain open. I then allow the flask to cool. But, singular fact—and one well calculated to astonish every one acquainted with the delicacy of the experiments relating to what is called ‘spontaneous generation’—the liquid of this flask will remain indefinitely without alteration. The flask may be handled without any fear, it may be transported from place to place, allowed to experience all the seasonal variations of temperature, and its liquid does not undergo the slightest alteration, whilst it preserves its odour and its taste.” If, however, the neck of one of these flasks be broken off close to the flask itself, then, according to M. Pasteur, the previously unaltered fluid will, in a day or two, undergo the ordinary changes, and swarm with Bacteria and Mucedineæ.
“The great interest of this method is,” M. Pasteur adds, “that it completes, unanswerably, the proof that the origin of life in infusions which have been raised to the boiling point, is solely due to the solid particles[23] which are suspended in the air.” He believes that any living things pre-existing in the fluid itself would be destroyed by the high temperature to which it had been raised; and that those contained in the air of the flask would also be destroyed, if not expelled, by the process of ebullition. Believing that the air is the source of germs from which Life is first developed in infusions, he thinks that what rapidly enters at first, on the cessation of ebullition, has its germs destroyed by contact with the almost boiling liquid; whilst the air which enters subsequently, and more slowly, is supposed to deposit its germs in the various flexures of the tubes, so that none are able to reach the fluid itself. Infusions, thus protected, do not undergo putrefaction, says M. Pasteur, because the access of pre-existing living things is necessary for the initiation of this change, and such access is prevented by the tortuous and bent neck of the flask.
Others say that some fluids submitted to the conditions mentioned, will undergo putrefactive changes, and that, therefore, these experiments of M. Pasteur are utterly incapable of settling the general question as to the cause of fermentation and putrefaction, and also that concerning the origin of Life. Although acknowledging a certain difficulty in explaining the results which are sometimes attained by this method, some of us would rather confess this than confidently offer explanations—as M. Pasteur did—which may in a short time be stultified by the results of other experiments with different fluids.
Having previously shown[24] that living things could appear and multiply in such a flask as M. Pasteur describes—in any flask, in fact,—which had been hermetically sealed during the ebullition of a suitable fluid within; this was deemed to be a result so contradictory to the explanations of M. Pasteur, that it appeared needless to add my testimony, as I could have done, to that of M. Victor Meunier and others, as to the different results obtainable by operating, in M. Pasteur’s fashion, with different fluids. It seemed to me that if organisms were to be procured in flasks from which air had been altogether expelled, it was useless still to urge the preservative virtues of any process of filtration of air—with the object of showing that living things in infusions derived their origin from atmospheric germs. Obviously, if there were no atmosphere, there could be no atmospheric germs present; and if living things were, nevertheless, developed under these exclusive circumstances, how could M. Pasteur or his disciples still expect to convince others that the first living things in infusions always proceeded from pre-existing atmospheric germs—even although it could be shown, that in many cases, when these were filtered off by flasks with narrow and tortuous necks, no living things were developed in such fluids. Granting to the full the truth of such facts, they could do nothing to establish the doctrine of the origin of infusorial life from pre-existing atmospheric germs, so long as it could also be shown that living things might be developed in boiled solutions to which air, instead of being filtered, was never allowed to enter at all.
It is not, therefore, because I think that some of the experiments which will subsequently be related afford any stronger or more direct support to my own conclusions, but because I think they may do this indirectly—by shaking the faith of many in some of the reasonings of M. Pasteur—that I am induced to give an account of them.[25]
What has been hitherto said, also applies to the more recent statements concerning the efficacy of cotton-wool as an agent for filtering germs from the atmosphere. Prof. Huxley says he has never seen putrefaction or fermentation occur after certain organic fluids have been boiled for ten or fifteen minutes, if a good plug of cotton-wool has been inserted into the neck of the flask in which they are contained whilst ebullition is going on, and has, subsequently, been allowed to remain in the same situation. Using other or perhaps stronger fluids, however, I have found that such a method of proceeding is by no means adequate to stop the growth and development of organisms. And, also, even if it had been always efficacious—the reason adduced could not hold good, in the face of my other experiments, which had shown that a development of life might go on in cases where the air, which had been similarly driven out, was subsequently, in place of being filtered, prevented from gaining access to the fluid.
If germs derived from the air are the sole causes of putrefaction, then, surely, deprivation from air ought to be just as efficacious as any process of filtration of air—more especially when the filtration or the deprivation have a common starting point. And the mode of procedure, in both cases, is precisely the same up to a certain point. A fluid is boiled for a short time in order to kill the germs which may be within the flask, and to expel its previously contained air. At a certain stage of the ebullition, this may be arrested, if we have to do with a bent-neck flask, or one whose neck is plugged with cotton-wool, and no change, it is said, will subsequently take place in the contained fluid, because the air which enters is, by either of these means, filtered from its germs. But if, whilst ebullition continued, the neck of the flask had been hermetically sealed—so as altogether to prevent the re-ingress of air—and if the fluid, thus contained in vacuo, would nevertheless undergo fermentation, obviously the former explanation must be altogether shelved.
In the face of M. Pasteur’s explanations, and those of Professor Huxley, these frequent positive results with fluids contained in vacuo are absolutely contradictory. There may naturally arise, therefore, a very grave doubt as to the validity of the explanation adduced by M. Pasteur, and adopted by Professor Huxley and others.
All these experiments to which I have been alluding are based upon the supposition (assented to by Pasteur and Huxley) that Bacteria which pre-existed in the solution would certainly be destroyed by its being raised for a few minutes to a temperature of 212° F. This conclusion is, I believe, perfectly correct,[26] and in support thereof I will adduce the following additional information.
Limits of ‘Vital Resistance’ to Heat displayed by Bacteria and Torulæ.
After stating elsewhere[27], that Vibriones are partly broken up or disintegrated by an exposure for a few minutes to a temperature of 212° F. in an infusion which is being boiled, and also that, in all probability, the life of Bacteria would be destroyed by such a treatment, I made the following remarks:—“With reference to these organisms, however, one caution is necessary to be borne in mind by the experimenter. The movements of monads and Bacteria may be, and frequently are, of two kinds. The one variety does not differ in the least from the mere molecular or Brownian movement, which may be witnessed in similarly minute, not-living particles immersed in fluids. Whilst the other seems to be purely vital—that is, dependent upon their properties as living things. These vital movements are altogether different from the mere dancing oscillations which not-living particles display, as may be seen when the monad or Bacterium darts about over comparatively large areas, so as frequently to disappear from the field. After an infusion has been exposed for a second or two to the boiling temperature, these vital movements no longer occur, though almost all the monads and Bacteria may be seen to display the Brownian movement in a well-marked degree. They seem to be reduced by the shortest exposure to a temperature of 100° C. to the condition of mere not-living particles, and then they become subjected to the unimpaired influence of the physical conditions which determine these movements.” I now have various facts to add in confirmation of these conclusions, and in extension of our knowledge concerning the vital resistance to heat of Bacteria and Torulæ.
It would be a most important step if we could ascertain some means by which these primary movements of living Bacteria might be distinguished from the secondary, or communicated, movements of not-living particles. In many cases, organisms that are truly living may only exhibit very languid movements, which, as movements, are quite indistinguishable from those that the same Bacteria may display when they are really dead. Because the movements, therefore, are of this doubtful character, persons are apt, unfairly, to argue that the Bacteria which present them, are no more living than are the minute particles of carbon obtained from the flame of a lamp, which may exhibit similar movements. This, however, is a point of view which becomes obviously misleading if too much stress is laid upon it; and it is more especially so in this case, when those Bacteria which display the most characteristic sign of vitality—viz., “spontaneous” division or reproduction—do, at the time, almost always exhibit only the same languid movements. Mobility is, in fact, not an essential characteristic of living Bacteria, whilst the occurrence of the act of reproduction is the most indubitable sign of their life. It should be remembered, therefore, that any Bacteria which are almost motionless, or which exhibit mere Brownian movements, may be living, whilst those which spontaneously divide and reproduce, are certainly alive—whatever may be the kind of movement they present.
In any particular case, however, can we decide whether Bacteria, that have been submitted to a given temperature, and which exhibit movements resembling those known as Brownian, are really dead or living? If the movements are primary, or dependent upon the inherent molecular activity of the organism itself, they ought, it might be argued, to continue when the molecules of the fluid are at rest; if, on the other hand, they are mere secondary or communicated movements, impressed upon the organisms as they would be upon any other similarly minute particles, by the molecular oscillations of the fluid in which they are contained, then the movements ought to grow less, and gradually cease, as the fluid approaches a state of molecular rest—if this be attainable. Following out this idea, some months ago, I first tested the correctness of the assumption by experimenting with fluids containing various kinds of not-living particles; such as carbon-particles from the flame of a lamp, or freshly precipitated baric sulphate. However perfect may have been the Brownian movements when portions of these fluids were first examined beneath a covering-glass, they always gradually diminished, after the specimen had been mounted by surrounding the covering-glass with some cement or varnish. Thus prepared, no evaporation could take place from the thin film of fluid, and after one, three, four, or more hours—the slide remaining undisturbed—most of the particles had subsided, and were found to have come to a state of rest. In order still further to test these views, I took an infusion of turnip, containing a multitude of Bacteria whose movements were of the languid description, and divided it into two portions. One of these portions was boiled for about a minute, whilst the other was not interfered with. Then, after the boiled solution had been cooled, a drop was taken from each and placed at some little distance from one another on the same glass slip; covering-glasses half an inch in diameter were laid on, and the superfluous fluid beneath each was removed by a piece of blotting-paper. When only the thinnest film of fluid was left, the covering-glasses were surrounded by a thick, quickly-drying cement.[28] Examined with the microscope immediately afterwards, it was generally found that the Bacteria which had been boiled presented a shrunken and shrivelled aspect—whilst some of them were more or less disintegrated—though, as far as movement was concerned, there was little to distinguish that which they manifested, from that of their plumper-looking relatives which had not been boiled.
If the specimens were examined again after twenty-four or more hours, there was still very little difference perceptible between them, as regards their movements. And the same was the case when the specimens were examined after a lapse of some days or weeks. One important difference does, however, soon become obvious. The Bacteria which have not been boiled, undergo a most unmistakeable increase within their imprisoned habitat; whilst those which have been boiled, do not increase. The two films may be almost colourless at first (if the Bacteria are not very abundant), but after a few days, that composed of unboiled fluid begins to show an obvious and increasing cloudiness, which is never manifested by the other. Microscopical examination shows that this cloudiness is due to a proportionate increase in the number of Bacteria.
Is the continuance of the movements of the organisms which had been boiled attributable to their extreme lightness, and to the slight difference between their specific gravity and that of the fluid in which they are immersed? I soon became convinced that this was one, if not the chief reason, when I found that Bacteria which had been submitted to very much higher temperatures, behaved in precisely the same manner as those which had been merely boiled, and also that other particles which—though obviously dead—had a similar specific lightness, also continued to exhibit their Brownian movements for days and weeks. This was the case more especially with the minute fat particles in a mounted specimen of boiled milk,[29] and also with very minute particles which were gradually precipitated[30] from a hay infusion that had been heated to 302° F. for four hours. Trials with many different substances, indeed, after a time convinced me that the most rapid cessation of Brownian movements in stationary films,[31] occurred where the particles were heavy or large; and that the duration of the movement was more and more prolonged, as the particles experimented with, were lighter or more minute. So that, when we have to do with Bacteria, the minute oil globules of milk, or with other similarly light particles, the movements continue for an indefinite time, and are, in part, mere exponents of the molecular unrest of the fluid. They are always capable of being increased or renewed by the incidence of heat or other disturbing agencies.
In respect of the movements which they may exhibit, therefore, really living, though languid, Bacteria, cannot always be discriminated from dead Bacteria. Both may only display mere Brownian movements.
It becomes obvious, then, that in doubtful cases we ought not to rely very strongly upon the character of their movements, as evidence of the death of Bacteria—although these may frequently be of so extensive a nature as to render it not at all doubtful whether the Bacteria which display them are living. In the experiments which I am about to relate, we shall be able to pronounce that the Bacteria are living or dead, by reference to the continuance or cessation of their most essentially vital characteristic. If Bacteria fail to multiply in a suitable fluid, and under suitable conditions, we have the best proof that can be obtained of their death.
Having made many experiments with solutions of ammonic tartrate and sodic phosphate, I have almost invariably observed that such solutions—when exposed to the air without having been boiled—become turbid in the course of a few days owing to the presence of myriads of Bacteria and Vibriones, with some Torulæ. These organisms seem to appear and multiply in such a solution almost as readily as they do in an organic infusion. On the other hand, having frequently boiled such solutions, and closed the flasks during ebullition, I have invariably found, on subsequent examination of these fluids, that whatever else may have been met with, Bacteria and Vibriones were always absent. The difference was most notable, and it seemed only intelligible on the supposition that any living Bacteria or dead ferments which may have pre-existed in the solution, were deprived of their virtues by the preliminary boiling. These experiments also seemed to show that such solutions, after having been boiled, and shut up in hermetically-sealed flasks, from which all air had been expelled, were quite incapable of giving birth to Bacteria. The unboiled fluid, exposed to the air, might have become turbid, because it was able to nourish any living Bacteria which it may have contained, or because it was capable of evolving these de novo, under the influence of dead ferments whose activity had not been destroyed by heat. Hence we have a fluid which is eminently suitable for testing the vital resistance of Bacteria,—one which, although quite capable of nourishing and favouring their reproduction, does not appear capable of evolving them, when, after previous ebullition, it is enclosed in a hermetically sealed flask from which all air has been expelled. Three flasks were half-filled with this solution.[32] The neck of the first (a) was allowed to remain open, and no addition was made to the fluid. To the second (b), after it had been boiled and had become cool, was added half a minim of a similar saline solution, which had been previously exposed to the air, and which was quite turbid with Bacteria, Vibriones and Torulæ. From this flask—after its inoculation with the living organisms—the air was exhausted by means of an air-pump, and its neck was hermetically sealed during the ebullition of the fluid, without the flask and its contents having been exposed to a heat of more than 90° F. The third flask (c) was similarly inoculated with living Bacteria, although its contents were boiled for ten minutes (at 212° F.), and its neck was hermetically sealed during ebullition. The results were as follows:—the solution in the first flask (a), became turbid in four or five days; the solution in the second (b), became turbid after thirty-six hours; whilst that in the third flask (c), remained perfectly clear. This latter flask was opened on the twelfth day, whilst its contents were still clear, and on microscopical examination of the fluid no living Bacteria were to be found. This particular experiment was repeated three times with similarly negative results, although on two occasions the fluid was only boiled for one minute instead of ten.
It seemed, moreover, that by having recourse to experiments of the same kind, the exact degree of heat, which is fatal to Bacteria and Torulæ might be ascertained. I accordingly endeavoured to determine this point. Portions of the same saline solution, after having been boiled[33] and allowed to cool, were similarly inoculated with a drop[34] of very turbid fluid, containing hundreds of living Bacteria, Vibriones, and Torulæ. A drying apparatus was fixed to an air-pump, and the flask containing the inoculated fluid was securely connected with the former by means of a piece of tight india-rubber tubing,[35] after its neck had been drawn out and narrowed, at about two inches from the extremity. The flask containing the inoculated fluid was then allowed to dip into a beaker holding water at 122° F., in which a thermometer was immersed. The temperature of the fluid was maintained at this point for fifteen minutes,[36] by means of a spirit lamp beneath the beaker. The air was then exhausted from the flask by means of the pump, till the fluid began to boil; ebullition was allowed to continue for a minute or two, so as to expel as much air as possible from the flask, and then, during its continuance, the narrowed neck of the flask was hermetically sealed by means of a spirit-lamp flame and a blowpipe. Other flasks were similarly prepared, except that they were exposed to successively higher degrees of heat—the fluid being boiled off, in different cases, at temperatures of 131°, 140°, 149°, 158°, and 167° F. All the flasks being similarly inoculated with living Bacteria, Vibriones, and Torulæ, and similarly sealed during ebullition, they differed from one another only in respect to the degree of heat to which they had been submitted. Their bulbs were subsequently placed in a water bath, which during both day and night was maintained at a temperature of from 85° to 95° F. The results have been as follows:—The flasks whose contents had been heated to 122° and 131° F. respectively, began to exhibit a bluish tinge in the contained fluid after the first or second day; and after two or three more days, the fluid in each became quite turbid and opaque, owing to the presence and multiplication of myriads of Bacteria, Vibriones and Torulæ; the fluids in the flasks, however, which had been exposed to the higher temperature of 140°, 149°, 158°, and 167° F., showed not the slightest trace of turbidity, and no diminution in the clearness of the fluid while they were kept under observation—that is, for a period of twelve or fourteen days. One kind of conclusion only is to be drawn from these experiments, the conditions of which were in every way similar, except as regards the degree of heat to which the inoculated fluids were subjected—seeing that the organisms were contained in a fluid, which had been proved to be eminently suitable for their growth and multiplication.[37] If inoculated fluids which have been raised to 122° and 131° F. for ten minutes, are found in the course of a few days to become turbid, then, obviously, the organisms cannot have been killed by such exposure; whilst, if similar fluids, similarly inoculated, which have been raised to temperatures of 140°, 149°, 158°, and 167° F. remain sterile, such sterility can only be explained by the supposition that the organisms have been killed by exposure to these temperatures.
Some of these experiments have been repeated several times with the same results. On three occasions, I have found the fluid speedily become turbid, which had only been exposed to 131° F. for ten minutes, whilst on three other occasions I have found the inoculated fluid remain clear, after it had been exposed to a heat of 140° F. for ten minutes.[38]
In experimenting upon rather higher organisms, with which there is little difficulty in ascertaining, by microscopical examination, whether they are living or dead, I have found that an exposure even to the lower temperature of 131° F. for five minutes, always suffices to destroy all signs of life in Vibrios, Amœbæ, Monads, Chlamydomonads, Euglenæ, Desmids, Vorticellæ, and all other Ciliated Infusoria which were observed, as well as in free Nematoids, Rotifers, and other organisms contained in the fluids which had been heated.
These results are quite in harmony with the observations and experiments of M. Pouchet and of Professor Wyman, as to the capability of resisting heat displayed by Vibriones and all kinds of ciliated infusoria. According to the former,[39] the majority of ciliated infusoria are killed at, or even below, the temperature of 122° F., whilst large Vibriones are all killed at a temperature of 131° F.[40] According to the observations of Professor Wyman, the motions of all ciliated infusoria are stopped at less than 130° F., whilst Vibriones, taken from the most various sources, also seemed to be killed at temperatures between 130°–136.4° F. Similarly, we find Baron Liebig quite recently making the following remarks concerning a species of Torula:—“A temperature of 60° C. [140° F.] kills the yeast cells; after exposure to this temperature in water, they no longer undergo fermentation, and do not cause fermentation in a sugar solution. . . . In like manner, active fermentation in a saccharine liquid is stopped when the liquid is heated to 60° C., and it does not recommence again on cooling the liquid.”
That the organisms in question—being minute naked portions of living matter—should be killed by exposure to the influence of a fluid at these temperatures will perhaps not seem very improbable to those who have attempted to keep their fingers for any length of time in water heated to a similar extent. With watch in hand I immersed my fingers in one of the experimental beakers containing water at 131° F., and found that, in spite of my desires, they were hastily withdrawn, after an exposure of less than five-and-twenty seconds.
Wishing to ascertain what difference there would be if the inoculated fluids were exposed for a very long time, instead of for ten minutes only, to certain temperatures, I prepared three flasks in the same manner—each containing some of the previously boiled solution, which, when cold, had been inoculated with living Bacteria, Vibriones, and Torulæ. These flasks and their contents were then submitted to the influence of the following conditions:—One of them was heated for a few minutes in a beaker containing water at 113° F., and then by means of the air-pump a partial vacuum was procured, till the fluid began to boil. After the remainder of the air had been expelled by the ebullition of the fluid, the neck of the flask was hermetically sealed, and the flask itself was subsequently immersed in the water of the beaker, which was kept for four hours at a temperature between 113° and 118 1/2° F.[41] The two other flasks similarly prepared were kept at a temperature of 118 1/2°–127 1/2° F. for four hours. In two days, the fluid in the first flask became slightly turbid, whilst in two days more the turbidity was most marked. The fluid in the two other flasks which had been exposed to the temperature of 118 1/2°–127 1/2° F. for four hours, remained quite clear and unaltered during the twelve days in which they were kept in the warm bath under observation. These experiments seem to show, therefore, that the prolongation of the period of exposure to four hours, suffices to lower the vital resistance to heat of Bacteria and Torulæ by 14 1/2°–18° F.
Such experiments would seem to be most important and crucial in their nature. They may be considered to settle the question as to the vital resistance of these particular Bacteria, whilst other evidence points conclusively in the direction that all Bacteria, whencesoever they have been derived, possess essentially similar vital endowments[42]. Seeing also that the solutions have been inoculated with a drop of a fluid in which Bacteria, Vibriones, and Torulæ are multiplying rapidly, we must suppose that they are multiplying in their accustomed manner, as much by the known method of fission, as by any unknown and assumed method of reproduction. In such a fluid, at all events, there would be all the kinds of reproductive elements common to Bacteria, whether visible or invisible, and these would have been alike subjected to the influence of the same temperature. These experiments seem to show, therefore, that even if Bacteria do multiply by means of invisible gemmules as well as by the known process of fission, such invisible particles possess no higher power of resisting the destructive influence of heat than the parent Bacteria themselves possess. This result is, moreover, as I venture to think, in accordance with what might have been anticipated à priori. Bacteria seem to be composed of homogeneous living matter, and any gemmule, however minute, could only be a portion of such living matter, endowed with similar properties.
Extent to which boiled Fermentable Fluids may be preserved in Vessels with Bent Necks, or in those whose Necks are guarded by a Plug of Cotton-Wool.
Having thus satisfied ourselves as to the truth of the conclusion that Bacteria are killed when the fluid containing them is boiled (at 212° F.), we are in a position to proceed with the inquiry as to the evidence which exists in respect to the statements made by M. Pasteur, Professor Huxley, and others, that fermentable fluids which have been boiled, will not undergo fermentation, either in vessels whose necks have been many times bent, or in those into whose necks a plug of cotton-wool has been inserted during the ebullition of their contained fluid. Organisms are not found in such cases, they say, because the “germs” from which the low organisms of infusions are usually produced, are arrested either in the flexures of the tube or in the cotton-wool. As I have before stated, however, it is obvious that if this explanation be the correct one, the preservation should be equally well marked in all cases—quite irrespectively of the amount of albumenoid or other nitrogenous material which may be contained in the fluid. Any exceptions to the rule should at once suggest doubts as to the validity of the explanation.
It was shown[43] in 1865 by M. Victor Meunier that some fluids were preserved after having been boiled in a vessel of this kind, whilst others, submitted to the same treatment, speedily became turbid from the presence of Bacteria and other organisms.[44] By these experiments he ascertained that strong infusions did frequently change, whilst weak ones might be preserved; and that even a strong infusion might be prevented from undergoing change if the period of ebullition were sufficiently prolonged.
The fluids most frequently employed by M. Pasteur were yeast-water, the same sweetened by sugar, urine, infusion of beetroot, and infusion of pear.
Taking urine as a fair example of such a fluid, I have found that the statements of M. Pasteur and of Professor Lister are perfectly correct. This fluid may generally remain for an indefinite period in such vessels[45] without becoming turbid, or undergoing any apparent change. The same is generally found to be the case with an infusion of turnip, and occasionally an infusion of hay may be similarly prevented from undergoing fermentation. On the other hand, if the turnip-solution be neutralized by the addition of a little ammonic carbonate, or liquor potassæ; or, better still, if even half a grain of new cheese be added to the infusion before it is boiled, then I have found that the fluid speedily becomes turbid, owing to the appearance of multitudes of Bacteria. In an infusion to which a fragment of cheese had been added, I have seen a pellicle form in three days, which, on microscopical examination, proved to be composed of an aggregation of Bacteria, Vibriones, and Leptothrix filaments. A mixture of albuminous urine and turnip-infusion has also rapidly become turbid in a vessel of this kind owing to the appearance of multitudes of Bacteria, and so has a mixture containing one-third of healthy urine with two-thirds of infusion of turnip.
Other infusions have been boiled for ten minutes in a vessel with a horizontal neck two feet long, into which, during ebullition, a good plug of cotton-wool had been carefully pushed down for a depth of twelve or fourteen inches, and cautiously increased in quantity during the continuance of the ebullition; whilst immediately after the withdrawal of the heat, the plug was pressed closer, and all the outer unoccupied portion of the tube was rapidly filled up in the same manner.
Preserved in such a vessel, a specimen of urine remained unchanged; a hay-infusion also underwent no apparent alteration; whilst a very strong infusion of turnip became turbid in five days, and ultimately showed a large quantity of deposit.[46]
Thus the rules laid down by Pasteur and others are not universal, and therefore, as I have previously pointed out, the explanation which he adduced of the preservation of those particular fluids which remained unchanged is at once rendered doubtful. More especially is there room for doubt on this subject when, as I have found, the result of the experiment can be, within certain limits, predicated beforehand, according to the nature of the fluid employed. If all organisms proceed from pre-existing germs, and these can be filtered from the air by a certain mechanical contrivance, then, if it be alleged that it is on account of such filtration that certain boiled fluids do not change, all fluids placed under these conditions ought, on this theory, to be similarly preserved. Exceptional cases cannot be accounted for on this hypothesis. To others, however, who say that organisms are capable of arising de novo, and that fermentation can be initiated without the agency of living things, the above facts appear quite natural. The more complex the nitrogenous or protein materials contained in a solution, the more is it fitted to undergo fermentative changes, which may be accompanied by the de novo origination of living things. Therefore the above results are just as compatible with the notions of M. Liebig and his school, as they are antagonistic to those of M. Pasteur. Certain fluids, it is found, do not undergo change; whilst other fluids, of a more complex description, will ferment under the influence of similar conditions. Prolonged ebullition also, by breaking up some of the more unstable compounds of a solution (those which most easily initiate these changes) will retard or prevent its fermentation.
The complete untenability of M. Pasteur’s explanations are, however, best revealed by having recourse to a series of comparative experiments, in which portions of the same fluid are boiled for an equal length of time in vessels of different kinds, and are then subsequently submitted, in a water-bath, to the influence of the same temperature.
I have made many experiments of this kind with different solutions, some of which I will now record. Owing to the different behaviour of the same fluids under different conditions, we are enabled to draw some most important conclusions; and owing to the different behaviour of different fluids under these respective conditions, our attention is strongly drawn to other facts which ought considerably to influence our judgment as to the relative merits of the two doctrines concerning the cause of fermentation and putrefaction.