Intracutaneous Inoculation Research Articles

Local Skin Reaction Obtained by Intravenous Injection of Agar Following Intracutaneous Inoculation of Meningococcus Toxin

Summary Intravenous injections of agar may be followed by reactions in skin areas prepared by injections of meningococcus toxic filtrate. Local reactions did not occur at the site of intracutaneous injections of agar when followed by intravenous injections of agar or of toxin. No reactions were obtained after the injection of other nonbacterial substances such as galactose, gelatin, serum and India ink, intravenously, following intracutaneous injection of meningococcus toxic filtrate.

REACTIONS OF RABBITS TO NON-HEMOLYTIC STREPTOCOCCI

Rabbits immunized intravenously with living culture or nucleoproteins of non-hemolytic streptococci react to subsequent intracutaneous inoculations with homologous streptococci with smaller and harder lesions than are shown by normal animals similarly inoculated; and they do not develop the general manifestations of hypersensitiveness such as are shown by animals previously inoculated into the tissues with the same cultures. A rabbit may react to intracutaneous inoculation with non-hemolytic streptococci in one of four ways, depending on whether it is normal, hypersensitive, immune or cachectic. Most normal animals show a secondary reaction about 10 days after inoculation with suitable strains of non-hemolytic streptococci; hypersensitive, allergic, or hyperergic animals show much larger lesions than do normals with the corresponding doses of the same streptococci, and practically never show secondary reactions; immune animals show smaller and harder early lesions and usually do not have secondary reactions if they are fairly well immunized. Cachectic animals show very soft and rapidly fading primary reactions and no secondary reactions.

TRANSMISSION OF RESPIRATORY ANAPHYLAXIS (ASTHMA) FROM MOTHER TO OFFSPRING

Rabbits immunized intravenously with living culture or nucleoproteins of non-hemolytic streptococci react to subsequent intracutaneous inoculations with homologous streptococci with smaller and harder lesions than are shown by normal animals similarly inoculated; and they do not develop the general manifestations of hypersensitiveness such as are shown by animals previously inoculated into the tissues with the same cultures. A rabbit may react to intracutaneous inoculation with non-hemolytic streptococci in one of four ways, depending on whether it is normal, hypersensitive, immune or cachectic. Most normal animals show a secondary reaction about 10 days after inoculation with suitable strains of non-hemolytic streptococci; hypersensitive, allergic, or hyperergic animals show much larger lesions than do normals with the corresponding doses of the same streptococci, and practically never show secondary reactions; immune animals show smaller and harder early lesions and usually do not have secondary reactions if they are fairly well immunized. Cachectic animals show very soft and rapidly fading primary reactions and no secondary reactions.

Intracutaneous Vaccination of Rabbits with Pneumococcus. I. Antibody Response.

Suspensions of heat killed pneumococci were injected into the skin of rabbits at intervals of 7 days during a period of 10 to 14 weeks. The total amount of bacterial substance injected was equivalent to and often greater than that ordinarily employed in the routine immunization of rabbits by the intravenous method. Pneumococci of Types I and III and a degraded “R” strain derived from Type II were used. The sera of the treated rabbits were tested for the presence of agglutinins, precipitins, and protective antibodies. The serum obtained from 85% of the animals immunized intracutaneously with Type I pneumococcus failed to show the presence of any demonstrable type specific antibodies. Virulent cultures of Type I were not agglutinated, nor were solutions of the specific soluble substance from organisms of the homologous type precipitated by these sera even when used in high concentrations. Only rarely did the serum confer any passive protection upon mice infected with a virulent strain of Type I, and in these instances the protective titre was low. In only 15% of animals studied was there any serological evidence of type specific response to repeated intracutaneous inoculation of Type I organisms; in these instances the presence in the serum of specific agglutinins was demonstrable only in low dilutions varying from 1:1 to 1:20. In terms of its capacity to stimulate the formation of type specific antibodies, Type III pneumococcus is at best a poor antigen. It was to be expected, therefore, that organisms of this type, when injected intracutaneously into rabbits, would fail to elicit the type specific response. Such proved to be the case. None of the rabbits treated by the skin method developed demonstrable serum antibodies against Type III.

Transmission of Respiratory Anaphylaxis (Asthma) From Mother to Offspring.

Rabbits immunized intravenously with living culture or nucleoproteins of non-hemolytic streptococci react to subsequent intracutaneous inoculations with homologous streptococci with smaller and harder lesions than are shown by normal animals similarly inoculated; and they do not develop the general manifestations of hypersensitiveness such as are shown by animals previously inoculated into the tissues with the same cultures. A rabbit may react to intracutaneous inoculation with non-hemolytic streptococci in one of four ways, depending on whether it is normal, hypersensitive, immune or cachectic. Most normal animals show a secondary reaction about 10 days after inoculation with suitable strains of non-hemolytic streptococci; hypersensitive, allergic, or hyperergic animals show much larger lesions than do normals with the corresponding doses of the same streptococci, and practically never show secondary reactions; immune animals show smaller and harder early lesions and usually do not have secondary reactions if they are fairly well immunized. Cachectic animals show very soft and rapidly fading primary reactions and no secondary reactions.

RECIPROCAL EFFECTS OF CONCOMITANT INFECTIONS

An experiment is reported in which was studied the effects of a concomitant vaccinal infection and of vaccinal immunity upon the reaction to syphilis in rabbits induced by intracutaneous inoculation. The results obtained showed that the reaction was modified by both conditions. A vaccinal infection initiated at the time of syphilitic inoculation was associated with a defensive reaction of lessened efficiency, the ensuing syphilis being more severe than in control animals. A state of vaccinal immunity present at the time of syphilitic inoculation was associated with a reaction of heightened efficiency, the ensuing syphilis being very mild. These results are in harmony with those obtained in other experiments in which the intratesticular route of syphilitic inoculation was employed.

ULCERATIVE TYPES AS DETERMINED BY INHERITANCE AND AS RELATED TO NATURAL RESISTANCE AGAINST TUBERCULOSIS: AN EXPERIMENTAL STUDY ON INBRED GUINEA PIGS

Five families of strictly inbred guinea pig whose general resistance to experimental tuberculosis had previously been determined by Wright and Lewis have now been studied with reference to the characteristics of the local lesion produced by intracutaneous inoculation with the tubercle bacillus. It is found that there are clearly recognizable familial types based on the size and quality of the nodular lesion, the ulcerative lesion consecutive to this and the general effectiveness of the healing process when in evidence. Family 39 which has the lowest general resistance forms an initial papule which does not differ appreciably from that formed by Family 35 which is the most resistant. In the ulcerative stage Family 39 shows an indolent excoriating process which exhibits none of the qualities which would be expected to make for healing. Family 35 exhibits a firm primary papule and nodule followed by an active healthy type of ulceration which is definitely restrained in comparison with that of the low resistance Family 13. Family 35 also presents the largest number of completely healed lesions in the later observation periods. Family 13 shows a larger, softer, primary papule and nodule than any of the others. The ulcer when formed is less restrained; that is, it is larger in proportion to the total size of the lesion. It is also more generally destructive. This family also shows a singular tendency to the formation of secondary ulcers along the lymphatic channels leading toward the adjacent lymph nodes. The adjacent lymph nodes are likewise more severely affected and frequently ulcerate through the skin giving rise to residual discharging sinuses. Families 2 and 32 are less definitely characterized. They are of the same order as 35 and 13 in that the ulcerations are active and healthy. Such similarities as are recognizable place Family 2 more nearly with 13 and Family 32 more nearly with 35. In general the qualities of lesion exhibited are such as to agree with the general resistance as previously determined and it is believed that the qualities underlying the tissue reaction may be safely considered to be among the influential factors in the make-up of the natural resistance against tuberculosis. The differences in reaction are in part manifested against agents causing simple inflammation also, and hence must be designated as non-specific with reference to the tubercle bacillus. There are definite indications of other "constitutional" differences in the character of the tissues and the general make-up of these families. The accumulated evidence from the study of the separate families and intercrosses between them is to the effect that the differences in question are transmitted by the blending type of inheritance and are therefore controlled by multiple unit characters. It is also suggested by the observations that certain of the significant characters are recessive in nature. It is of considerable interest that the observations agree very well with the older conception of an inherited, predisposing, constitutional diathesis as a significant factor in the incidence of tuberculosis.

Immune Tissue Response to Non-hemolytic Streptococci.

We1 originally reported that if a rabbit had been recently inoculated by any route with streptococci of any type it would fail to show a secondary reaction following intracutaneous inoculation with suitable strains of streptococcus viridans. This might be due to one of two conditions: first a hyperergic, second an immune state. First, if due to a hyperergic state there might be a combination of accelerated hyperergic-secondary-reaction with the primary reaction; hence the initial response to introduction of the same sized inoculum would constantly increase up to a certain point. Second, if an effective immune state had been induced, the local protective mechanism against the inoculum might be so efficient that the sum of the primary and accelerated secondary reactions would be smaller than the initial response of a normal animal.These two states were easily demonstrated in rabbits, according to the route used for inoculation. When repeated intracutaneous injections were made of 0.1, 0.01 and 0.0001 cc. at...

Cutaneous Reactions with Streptococcus Filtrates in Rabbits Rendered Allergic with Extracts of Guinea Pig Kidneys.

In a series of articles published in La Pediatria, Di Cristina, Sindoni, and Caronia1 describe a small diplococcus which they are able to obtain in culture from scarlet fever. These authors attribute certain specific reactions to this diplococcus which resemble the reactions obtained with the hemolytic streptococcus isolated from cases of scarlet fever in this country. The diplococcus has been obtained in blood cultures from scarlet fever and grown in media containing fragments of guinea pig tissue. When the filtrates of these cultures are injected intracutaneously, the response is apparently identical with that resulting from an intracutaneous inoculation with a filtrate of Streptococcus scarlatinae. These authors have prepared vaccines which they believe are capable of inducing an active immunity in persons susceptible to scarlatina.In 1925 Brokman2 and his associates undertook a comparison of the properties of scarlatinal hemolytic streptococcus and this diplococcus of Di Cristina and Caronia. Carboliz...

SKIN INFECTION OF RABBITS WITH HEMOLYTIC STREPTOCOCCI ISOLATED FROM A PATIENT WITH ERYSIPELAS

Cultures of a hemolytic streptococcus isolated from a human case of erysipelas were tested for ability to produce lesions by injecting various amounts intradermally in shaved rabbits. Severe lesions could be developed at will in this way despite the fact that intravenous or intraperitoneal inoculations of large doses of the cultures failed to kill. The lesions were of two sorts, phlegmonous and erysipelas-like. A serum capable of warding off the injurious action of the streptococcus on the skin was procured from animals receiving repeated intracutaneous inoculations of the organism. Normal serum by contrast had no protective effect. By intradermal injections at several points in the same normal rabbit of small quantities of mixtures of equal parts of serum and various dilutions of a suspension of streptococci it proved possible to test sera for the presence of substances protective against the streptococcus.

The production of an exotoxin by certain strains of staphylococcus aureus.

We have succeeded in demonstrating in sterile filtrates of broth cultures of four out of twenty-one strains of staphylococcus aureus isolated from various conditions, a powerful poison with a selective action for the skin. The poison is produced by the growth of these strains in any well buffered broth medium containing only a small amount of glucose. The medium from which our most powerful poisons were obtained was prepared by adding an equal volume of M/15 phosphate buffer solution pH 7.4 to ordinary sugar—free meat infusion broth containing 4 per cent Witte peptone, boiling, filtering and autoclaving. After inoculation with a suitable strain of staphylococcus, the poison can be demonstrated in the sterile Berkefeld filtrates of this broth after 24 hours growth, but our most toxic poisons were obtained after four to six days growth.The toxicity of the filtrates was tested by intracutaneous inoculation in rabbits. One-tenth cc. of filtrates was inoculated intradermally and at the same time a control of 0...

GUINEA-PIG INOCULATION FOR TUBERCLE TEST

Several methods of facilitating the rapid recovery of tubercle bacilli from inoculated guinea-pigs have been suggested. Intraperitoneal inoculation requires at least six weeks for the satisfactory development of lesions, subcutaneous inoculation at least four weeks. Exposure to roentgen rays previous to inoculation has been suggested to increase the susceptibility and make possible a recovery of the tubercle bacillus in two weeks. With intrahepatic inoculation, a search for tuberculous abscess is made after two weeks. All these methods require killing the animal, the test is interrupted, and further observation is impossible, particularly in apparently negative or doubtful cases. Besides, it may be necessary in some of these tests to make tissue examinations in order to arrive at a diagnosis. With intracutaneous inoculation, a result can be obtained in less than two weeks; the development and location of the lesion are always under observation, and the animal may be kept alive for

STUDIES BASED ON A MALIGNANT TUMOR OF THE RABBIT

The results of seven methods of transplantation of a malignant tumor of the rabbit are reported and contrasted with results obtained by intratesticular inoculation. It was found that inoculations made into the brain, the anterior chamber of the eye, and the muscles gave an almost uniform series of takes with the production of comparatively large and vigorously growing tumors. Intracutaneous inoculations were less favorable, while with subcutaneous, intravenous, and intraperitoneal inoculations, a growth was obtained in only 20 to 25 per cent of the animals. In contrast with the malignant course of the disease produced by testicular transplantation, the tumors produced by methods other than intracerebral inoculation were relatively benign. In all cases, the growth was of a purely local character; in some instances, there was invasion of the surrounding tissues with local or regional extension of the growth, but no metastases developed in distant organs. Tumors in the brain soon caused death as a result of pressure, and the ultimate fate of tumors in the eye was not determined, but the growth produced by other methods of local inoculation was of relatively short duration and terminated with spontaneous resolution. It was found that while a good growth could be obtained with material which had been subjected to repeated freezing, thawing, and grinding, the use of cell-free filtrates or desiccated tumor tissue gave negative results. In discussing the experiments reported, it was pointed out that there were three groups of factors which had played important parts in determining the results of transplantation. These were the capacity for growth of the tumor cells, animal resistance, and the advantage given the tumor or the animal by the use of a particular method of inoculation. From an analysis of the results obtained with reference to the operation of these factors, the conclusion was reached that while the quality of malignancy displayed by the tumor was conditioned upon the capacity for growth of the tumor cells, the degree of malignancy exhibited in any given instance was a function of animal resistance which was not entirely specific but corresponded closely with the ability of the animal to meet excess demands arising from other causes and, to this extent, represented an expression of the functional capacity of a mechanism which is concerned primarily with the maintenance of the general animal economy.

STUDIES BASED ON A MALIGNANT TUMOR OF THE RABBIT

Several attempts were made to transplant a spontaneous malignant tumor in a syphilitic rabbit before a method was devised which proved successful. Subcutaneous and intracutaneous inoculations were unsuccessful on account of the vigorous granulomatous reaction which was aroused by the introduction of tumor fragments into these tissues. This difficulty was overcome by the use of intratesticular inoculations. With this method, a good growth was obtained in practically all animals. It appeared possible, however, that a syphilitic infection had been transmitted along with the tumor, and hence treatment with arsphenamine was instituted as a means of eliminating this infection. A study of the transplanted tumor in the first and second generations showed that it possessed a high degree of malignancy. Metastases occurred at an early period in the majority of animals, and while some of them appeared to recover, the indications were that more than 50 per cent might be expected to succumb to the effects of the tumor growth within a period of 6 to 7 months. The method of intratesticular inoculation now in use is described in detail.