IMMUNITY (see also BACTERIA AND DISEASE). The term "immunity" is used in science in the technical sense. An animal is described as naturally immune against the microbe of a disease if the microbe in question cannot establish itself in the organism and as artificially immune if it was naturally susceptible and has been rendered insusceptible. Similarly, an animal is described as naturally immune against a poison if its organism is naturally proof, and as artificially immune if its organism has been rendered proof against the poison.
Only a few species of microbes (the staphylococcus, streptococ cus, pneumococcus and perhaps the microbes of all genuinely septicaemic diseases) can proliferate in antitryptic blood fluids. Further, these microbes (serophytic microbes) produce when grown in plasma (generalizing from observations made with staph ylococcus and streptococcus) trypsin which quenches the anti tryptic power of the surrounding blood fluids and then digests vacuoles in the surrounding clot. Again, all serophytic microbes grow very much more vigorously (the streptococcus for example very many thousand times more vigorously) in the blood fluids when these microbes have, by an artificial addition of trypsin, been relieved from the task of themselves producing enough trypsin to quench the antitryptic power of the surrounding medium. And finally, all those kinds of microbes which are unable to grow in antitryptic blood fluids begin to pullulate there the moment tryp sin is added—and let it be noted here that a spontaneous addition of trypsin occurs regularly in wounds as soon as the emigrated leucocytes are broken down under the influence of bacterial growth and unfavourable external influences.
Anticipating, we may say that later it will be shown with regard to acquired immunity to infections that this depends upon the development of (a) increased antibacterial substances in the blood fluid, (b) increased efficiency in the leucocytes and (c) increased capacity for producing antibacterial substances in response to in fection. Similarly it will be shown with regard to acquired im munity to poisons that this depends upon the appearance of neutralizing elements (so-called antitoxins) in the blood coupled with the acquirement of increased capacity for elaborating these elements in response to an incorporation of the corresponding poisons.
Confining ourselves to the problem as to how the bacterial in fections are combated in the organism, we may begin by con sidering the two chief opposing doctrines which were promulgated, when, under the prompting of Pasteur's practical achievements in the field of prophylaxis, problems of immunity began to be first seriously considered. These theories included natural as well as acquired immunity.
What applies to natural, applies also to acquired immunity. There is nothing to show that increased resistance to serophytic microbes and septic infections generally is accompanied by the development of any bactericidal power in the blood fluids.
With regard to these free-living members of the protozoal colony Mechnikov divined that they function as defensive cells. And he showed in connection with sponges and other inverte brates that these wandering cells collect round invading microbes and other foreign intrusions and that they thereafter proceed to ingest and digest them or to aid otherwise in their elimination. Lastly Mechnikov pointed out that the leucocytes of the verte brate were homologous to those wandering cells and performed exactly the same defensive offices. In particular when leucocytes emigrate from the capillaries into a focus of bacterial infection, in the normal case, they follow this up by ingesting the intrud ing micro-organisms, and killing them intracellularly. Natural or native immunity was thus, in the conception of Mechnikov, due to efficient leucocytic functioning ; and acquired immunity to the leucocytes having by a process of training (as he called it) ac quired a power of more effectively confronting, ingesting and destroying microbes.
In connection with the first kind of microbe the fact that although they grow freely in the serum and plasma, these are killed in large numbers in the blood and in the serum, when liv ing leucocytes are added, shows that the cellular elements of the blood here do the work of destruction.
In connection with the successful killing of these microbes, the leucocytes must, however, in all cases have free mechanical access to the microbes. There would for example be default in this respect if, as would happen in dealing with infected defibrinated or infected centrifuged blood in vitro, the leucocytes settled to the bottom and the microbes were buoyed up out of their reach in the serum. The same would of course happen in vivo in all serous effusions.
In connection with those microbes which are not directly poisoned but fail to proliferate in the serum, the most important restraining influence is the native antitryptic power of the blood, but once the antitryptic inhibitory action of the blood fluids and serous effusions has been neutralized the leucocytes, even when everything else favours them, will be impotent to inhibit microbic growth. On the contrary as soon as the leucocytes degenerate they will furnish a further quantum of trypsin and in this way directly conduce to the pullulation of all manner of microbes.
Lastly, the leucocytes do not seem to contribute to the defence of the organism against those microbes which, like the typhoid bacillus and the cholera vibrio, are directly poisoned by the serum. When we implant such microbes into defibrinated blood and then incubate and make microscopic preparations those microbes which are quickly ingested are found intact within the phagocytes while those which are left exposed to the action of the serum are dis torted and dissolved. And again, when living emigrated leucocytes are brought to bear upon typhoid bacilli it would seem that few of the microbes are killed. Precisely similar results are obtained with extracts made from leucocytes. It was shown by Schatten froh that such extracts exert bactericidal action upon the staphy lococcus and streptococcus while they exert no such action upon the typhoid bacillus and the cholera vibrio. And again it was shown by Colebrook that the products of inflammation derived from foci of inflammation set up by incorporating into rabbits lint soaked in typhoid vaccine are powerfully bactericidal for the staphylococcus and streptococcus while for the microbes of typhoid and cholera they are not more bactericidal than ordinary serum.
This review of the facts shows that only in the case of sero phytic microbes does the defence of the body depend upon the leucocytes and that when it is a question of defence against the other two classes of microbes (those whose growth is entirely inhibited and those which are directly poisoned by the serum) the leucocytes are either impotent or directly harmful.
Leucocytes Concerned in Immunization.—Mechnikov originally taught that the leucocytes were attracted to microbes and induced to phagocytose them by the toxin secreted by the microbes; and that there was here only an interaction between leucocytes and microbes and that the blood fluids could be left entirely out of the story. That that doctrine cannot be sustained can be demonstrated by a simple experiment.
We begin by receiving a sample of blood taken direct from the vessels into normal salt solution. We then centrifuge and recen trifuge in further volumes of this normal salt solution so as to wash the cellular elements free from all traces of the blood fluids. We then make two so-called phagocytic mixtures combining in the one case (a) one volume of a bacterial suspension with (b) one volume of washed leucocytes and (c) one volume of o.85% salt solution; and in the other case (a) one volume of washed leucocytes, (b) one volume of the same bacterial suspension and (c) one volume of normal serum. These mixtures are then placed in the incubator and after a suitable lapse of time a sample of each is examined under the microscope. In the first phagocytic mixture—that in which no serum was employed—the leucocytes have failed to ingest any microbes. In the specimen in which serum has been employed, the microbes have been plentifully in gested. This result depends not upon any stimulating effect ex erted upon the leucocytes, but upon the fact that the serum has effected a chemical change—a so-called opsonic change—in the microbes. That this opsonic change does not in any sense affect the vitality of the microbe is shown in the case of serophytic microbes by the fact that the serum that exerts an opsonic effect furnished culture medium in which the affected microbes will all grow out into colonies. The blood fluids intervene further in the destruction of microbes in the interior of the phagocyte by combining, with them in such a way as to favour their intracellular digestion. This action, which was first described by Douglas, is known as the protryptic action of the blood fluids.
The living leucocyte can also kill microbes apart from phago cytosis. The following experiment is instructive. A shallow re ceptacle is filled with a solid nutrient medium whose surface has been uniformly implanted with staphylococcus or streptococcus. The centre portions of the three cover glasses are thickly carpeted with living leucocytes obtained direct from the blood. We now, after different treatment, impose these cover glasses side by side upon the implanted nutrient surface. In the case of the first cover glass the adhering leucocytes are brought into application in con junction with the adhering serum; in the case of the second they have been washed free from every trace of serum ; and in the case of the third cover glass the adhering leucocytes, instead of being employed living, have been killed by drying.
The whole preparation is now incubated at blood heat for I 2 hours or more. The microbes will then be found to have grown out forming an uninterrupted sheet of colonies over the whole surface of the nutrient medium except under those areas of cover glasses i and 2 which are carpeted with living leucocytes. Here the implanted microbes have not proliferated, and microscopic exam ination of the cover glasses shows that where the leucocytes came into action in conjunction with serum the microbes are all lying intracellularly; while where the leucocytes were washed free from serum, the microbes are all lying extracellularly. The experiment thus shows that microbes can be killed by leucocytes both intra and extra-cellularly, i.e., both by phagocytosis and also apart from phagocytosis. This holds true both under the conditions obtaining in this particular experiment, and also in numerous other con ditions. That destruction of microbes by leucocytes without the intervention of phagocytosis occurs also very frequently in vivo is practically certain.
The above deals only with the normal protective machinery of the body as distinguished from that which comes into play in artificial immunity. In connection with the latter Metchnikov, as will be remembered, taught that acquired resistance was due not to any changes in the blood fluids, but to the leucocytes hav ing been subjected to a process of training which gave to them a greater capacity for confronting and ingesting microbes. This doctrine had to go by the board when it was shown in numberless cases that increased phagocytosis goes hand in hand with increas ing opsonic power in the blood fluids.
From this it was incautiously assumed—though this tenet was never definitely formulated—that the leucocytes constitute in artificial immunity an invariable, and the blood fluids the only variable, factor. That the phagocytic efficiency of the leucocytes was also a variable factor was first shown by Shattock and Dudgeon, who observed that the phagocytic efficiency of the patient's leucocytes is in many cases of pyrexial infection greater than that of the normal man. The phagocytic efficiency of a patient's leucocytes may also, as was further shown by Shattock and Dudgeon, be less than normal.
These observations have a direct bearing upon Metchnikov's doctrine with respect to acquired immunity, since, in all infections associated with constitutional disturbances, antigens from the foci of infection are being brought into operation—in other words the organism is experiencing and is reacting to auto-inoculations. It follows that, conformably with the doctrine of Metclikov, the leucocytes should in every case of pyrexial infection be conducted by successive degrees to a condition of continually increased phagocytic efficiency. Instead of that these cellular elements are, sometimes in localised infections, and practically always in streptococcus septicaemia, reduced to a condition of diminished efficiency.
Pasteur's Work.—After Jenner came Pasteur and with Pasteur scientific methods are for the first time brought into application in connection with prophylactic inoculation. His initial achieve ment was to recognise that the essential in Jennerian vaccina tion was that for a virulent infective organism obtained from actual cases of smallpox there has been substituted an infective organism which by the operations of nature—to wit by transfer to the cow—had been attenuated in such a manner as to render it non-lethal for man. By the exploitation of that general prin ciple, by the employment of pure culture and by a technique of artificial attenuation adapted with infinite resource to each separate case, the whole series of Pasteurian successes in the field of artificial immunization were one after another achieved. Artificial immunization was not, however, purged from all its risks by the procedures of Pasteur.
It had not yet been transformed into a scientifically regulated procedure. The Pasteurian vaccines were in point of fact stand ardized only thus far that recourse to attenuation placed in each case a certain limit upon the proliferation of the vaccinating material in the organism of a normally resistant man or animal.
Standardization.—A great step in advance was taken when it was established in connection with anti-typhoid inoculation that the antigen required for the setting in motion of the machinery of immunization can be furnished by the incorporation of sterilized microbic cultures. And further, important steps to the achieve ment of a standardization of bacterial vaccines were made when a technique for the enumeration of the microbes in bacterial sus pensions was devised, and when it was recognized that weighed quanta of desiccated and powdered bacterial substance could be employed in cases where, owing to the felting together of the mi crobes, enumeration of the microbial suspension was impracti cable. The counting of the microbes or the weighing of the bac terial substance is, however, only a means to an end—the stand ardization of a vaccine, i.e., the determination of the doses which will give the best "curve of immunization." The Curve of Immunization.—The expression curve of immuni zation calls attention to certain fundamentally important points in connection with the reaction of the body to the incorporation of vaccines. In the pre-Pasteurian and Pasteurian periods, when ideas about the nature of immunizing response were still vague, it was assumed with regard to vaccines that they produced their effects only after a certain incubation period (ordinarily only after ten days) . And it was further taken for granted that the curve of immunization would from the beginning move always in the upward direction. When, however, immunization curves came to be constructed (and this was done first in connection with anti typhoid inoculation) unanticipated features revealed themselves and in connection with these also time-relations which did not conform with expectation.
When, instead of a quantum of vaccine which produces a con stitutional disturbance, a smaller dose is inoculated, the negative phase is elided and a positive phase is well developed already 24 hours after the incorporation of the vaccine. And, finally, when excessive doses of vaccine, such as produce very severe constitu tional disturbances, are incorporated, the negative phase may be correspondingly intense and may persist for many weeks. These facts have an important bearing upon prophylactic operations; they must also, as reflection will show, have an importance in connection with the immunization procedure to which horses are subjected with a view to their producing antibacterial and anti toxic sera Again substantially the same relations as between the quantum of vaccine inoculated and the type of response elicited, obtain in the case where vaccines are inoculated into patients who are the subjects of infection. But in that case the conditions are so far different that we have to consider in each case two quanta of antigen : that administered in the vaccine, and that already con tained in the patient's organism. Where a patient is the subject of only a minimal infection, we may employ doses of vaccine nearly as great as those employed for the prophylaxis of healthy men. Where a patient is heavily infected, we are restricted to the employment of minimal doses, and finally, when the patient is already labouring under an excessive infection, the injection of vaccines can only do harm.
The principle that the kind of response, and the amplitude of the reaction, and the time-relations of the phases are in each case a function of the quantum of antigen brought into applica tion is found to apply also to the case where the vaccine is added to the extravascular blood. We obtain in vitro every variety of effect according to the dose of vaccine brought into application and the time for which it operates upon the blood—the effects varying from an instantaneous increase of bactericidal power to a loss of most of that destructive power. And further different effects are obtained according as we select now one and now an other method of testing. We obtain for example one result when we measure the bactericidal power of the whole blood ; another when we measure the opsonic power of the serum ; a third when we measure the phagocytic efficiency of the leucocytes.
The lessons which can be drawn from immunization curves may now be summarised. The first of these is that excessive doses of vaccine may delay and possibly interfere with the prophylactic response, and further that such excessive doses of vaccine admin istered to patients suffering from an infection may definitely aggravate their condition. Other important lessons are that when appropriate doses of vaccine (i.e., doses which are not followed by a negative phase) are administered, prophylactic effects may be obtained almost immediately after inoculation. And further it should be possible to arrest a general infection by inoculating in the incubation period and then employing a reduced dose of vaccine.
Considerable evidence showing that immunization develops very rapidly after the inoculation of an antigen is furnished in the work of Pirquet. If a first implantation of vaccinia is fol lowed by similar implantations on successive days, the response to these latter differs from the response made to the first. This altered clinical response—which would appear to indicate a more rapid destruction of the later implanted infective material—was described under the name of allergy. It was further shown by Pirquet in connection with the inoculation of foreign serum that the supervening clinical reaction—which appears to indi cate the throwing out of that foreign serum from the blood— is accelerated when the injection of serum is made into an organ ism which has been before subjected to that procedure. The inter val between the injection of the foreign serum and the clinical manifestations associated with its elimination from the blood (normally I(3-14 days) may be reduced to a very few minutes.
A series of further questions in relation to artificial immunity have to be considered (I) Are there agencies other than vaccines proper—in other words, other than living or dead microbes or substances derived from the bodies of microbes—which will affect the bactericidal power of the blood and leucocytes? (2) Again, are the antibacterial substances in the serum specific in the sense of operating only upon one particular variety of microbe or are they non-specific? (3) Further, will leucocytes which have acquired increased phagocytic efficiency, ingest more actively only one particular species of microbe, or all microbes without distinc tion? (4) Finally, what are the cells in the body which elaborate the anti-bacterial substances? All these questions are intimately linked up.
These laboratory experiments are in consonance with the sta tistical results obtained in connection with anti-pneumonococcus inoculations at the Premier Mine in the Transvaal. Here in 1912, in addition to a striking reduction in the incidence and death-rate of pneumonia, there was achieved a striking reduction in the incidence and death-rate from "other diseases." Derivation of Immunization discussion of the deeper problems as to where antibacterial substances are elaborated in the body, and as to how their production is to be explained, may be deferred for a moment. For the facts relating to immunization against bacterial toxins must first be taken into consideration by bringing out the following points: (I) The machinery of immunization is in reality a machinery for neutral izing or otherwise disposing of poisonous substances—poisonous substances being by definition those which enter into crippling or lethal chemical combination with the blood fluids and tissues.
(2) The machinery of immunization achieves its ends by furnish ing substances which enter into neutralizing or precipitating or destructive union with the poisonous substances above spoken of.
(3) The machinery of immunization is brought into operation only by a particular class of poisons—to wit, by those which enter into crippling but not immediately lethal chemical combi nation with the cellular protoplasm—those which, to use the expressions of Ehrlich, intrude themselves into the "side-chains" and not into the "vital ring" of that protoplasm. Of such poisons four kinds specially invite attention. These are : (a) the poison ous constituents of the bacterial protoplasm, (b) the albuminous substances contained in foreign sera and certain other foreign al buminous substances, (c) bacterial toxins such as those which can be filtered off from cultures of diphtheria and tetanus and (d) vegetable and animal toxalbumens such as abrin, ricin and the various snake venoms. With respect to the first the body responds to their inoculation by a production of bacterio-tropic substances, i.e., substances which enter into detrimental or lethal chemical combination with bacteria. The inoculation of sera is followed by the elaboration and delivery into the blood of sero-tropic substances which neutralize and precipitate these sera. The in oculation of bacterial toxins in like manner—this discovery was made by Behring—leads to the production and delivery into the blood of toxitropic substances. These, known as bacterial anti toxins, neutralize and precipitate the corresponding toxins. And, finally, the inoculation of toxalbumens is followed by an elabo ration and delivery into the blood of the appropriate neutralizing substances.
Further, by the study of the antitoxin content of the blood in its relation to diphtheria toxin, it has been shown that the pres ence of even a small quantum of antitoxin in the blood protects against infection, and that the insusceptibility of the majority of adults to diphtheritic infection is correlated with the possession of a minute quantum of antitoxin derived, as it would seem, from repeated minimal diphtheritic infections contracted in their earlier life. The same would appear to hold also of the scarlet fever streptococcus. We have here, as reflection will show, facts which illuminate the epidemiology of diphtheria and scarlatina and show that it is possible and may under circumstances be advisable to substitute for a prophylactic inoculation of a bacterial vaccine, an injection of antitoxin, or alternatively an injection of such quantum of toxin as will evoke an antitoxic response.
Two further points about antitoxins have important bearings upon the problem as to where and how products of immunization are produced in the organism. (I) Antitoxins are quite rigidly specific—each antitoxin neutralizing only the particular kind of toxin in response to which it was engendered. (2) After a first inoculation of toxins antitoxins are only very slowly produced. Ordinarily an interval of ten to 20 days elapses before they make their appearance in the blood.
These properties are not, let it be noted, differential properties of antitoxins; they characterize also certain kinds of bacterio tropic substances. Specificity and comparatively tardy appear ance in the blood characterize for example agglutinins, and the so-called "thermostable immune bodies." Since there are two kinds of products of immunization : one kind that are eminently non-specific and are produced imme diately (and can, as we have seen, be produced in the blood in vitro) ; and another kind which are rigidly specific and are elab orated only after a considerable lapse of time and are so far as appears produced only in vivo; we may now seek for an answer to the problem as to how and where these various products of immunization are engendered. Since it may be taken as certain that these two kinds cannot well originate in the same cells and be engendered by the same kind of metabolic operation, we may divide up the problem and consider first by what cells and by what kind of metabolic operation the non-specific anti-bacterial substances are produced and then take up the question as to where and how antitoxins and such antibacterial substances as are specific are generated.
Further the facts comport with the idea that leucocytes which have elaborated antibacterial secretions but have not as yet ex creted these into the environing blood fluids will, by virtue of their increased content in antibacterial substances, possess in creased antibacterial efficiency, while the blood fluids will not have gained anything in antibacterial power.
Conversely leucocytes that have excreted their antibacterial substances will exhibit diminished antibacterial power and the blood fluids which have received these secretions will have re ceived an accretion of antibacterial power.
At the same time these counterpart substances differ funda mentally from the non-specific antibacterial substances which were considered above. First of all they are not, as are the sub stances last mentioned, available in the form of secretory prod ucts produced only with a view to their being ejected from the cell. Instead of that, specific counterpart substances are integral elements of the cellular protoplasm, and elements of which it may be assumed with certainty that they subserve special func tions in the internal economy of the cells of which they are con stituents. Further the specific counterpart substances we are here considering differ from the non-specific antibacterial sub stances in the respect that while these latter are elaborated only in one particular variety of cell (to wit, in the leucocyte) the for mer are widely distributed in the organism, being located in each case in a different assortment of cells. Thus, for example, the counterpart substances to which the diphtheritic toxin would anchor itself would be located in a different assortment of cells than the counterpart substances with which the tetanus toxin would combine. And again the counterpart substances which would combine with abrin would be different. This would hold true also of the counterpart substances which would unite with the poisons derived from each particular variety of bacterial protoplasm.
So far it has been shown only that susceptible animals must by the very nature of things contain in their cell-protoplasm con stituent elements which are the exact chemical counterparts of poisons. We are still very far from the solution of the mystery (a) of the organism furnishing antitoxins in the circulating blood; (b) of its furnishing antitoxins only to special classes of poisons; and (c) of its furnishing these in quantities far in excess of the quantum of counterpart substances originally contained in the organism. Ehrlich, in exploring for some way of exit from the labyrinth constituted by these questions oriented himself by the aid of a ground-plan, in which there was set out his general con ception of the stereo-chemistry of protoplasm. In the plan in question the protoplasm is a structure made up of side-chains assembled round a central ring—the continued life of the proto plasm depending upon the integrity of the central (or as we may call it vital) ring; while the side-chains consist of elements which are integrated into the protoplasm for its nourishing and vital functioning. To this original ground plan there was now added by Ehrlich a new feature. The conception which he now added was that the side-chains of the protoplasm would, as soon as they became redundant, be cast forth from the cell into the circulating blood, forming there what he called free receptors.
Interpreted in the light of this so-called side-chain theory, the incorporation of poisons which make a chemical attack upon the vital ring of the cell protoplasm would abrogate the life of the cell and would therefore be incompatible with an elaboration of antitoxins. The situation is entirely different when the poison, instead of directing its attack to the vital ring, anchors itself on to one of the side-chains. Af ter a temporary putting out of action of those functions which are discharged by the particular side chains in question, this would lead to the replacement of the crippled side-chains, and thereafter to a hyper-replacement and to such redundancy of these in the protoplasm as would involve casting forth these side-chain receptors into the circulating blood. This theory, which is quite as applicable to the production of specific antibacterial substances as to the production of antitoxins —would seem to lie open to critical assault in that the hyper-re placement of side-chains does not necessarily conduct to an excretion of these into the circulating blood. In point of fact in the case of muscle, on which Ehrlich here relies, it leads to some thing very different, to a hypertrophy of this tissue.
Finally, the side-chain theory has met with hostile criticism more especially on the ground that it would compel us to believe that the noble tissues, such as those of the central nervous tissue which are poisoned by the toxins of diphtheria and tetanus, can be converted into secretory organs so prolific as to furnish in the blood tens and hundreds of thousands of units of the correspond ing antitoxins. The side-chain theory does not in any way re quire us to believe this. It would do so only if it had been estab lished that poisons such as diphtheria and tetanus toxins attack only the central nervous system. But in point of fact Ehrlich assumed that the poisons which are responded to by a production of antitoxins are all polytropic—in other words they turn towards and combine chemically with a number of different tissues. So far therefore as the side-chain theory is concerned, we are thus authorised to assume that, not the cells of the central nervous system, but all or any of the other and less noble tissues which are affected by the toxins, are those which produce the harvest of antitoxins.