Presence Of Antitoxin Research Articles

Anomalous enhancement of botulinum toxin type A neurotoxicity in the presence of antitoxin

The neutralization of botulinum toxin serotype A with polyclonal equine antitoxin was studied in isolated mouse hemidiaphragms and compared to the same action in live mice. The biological activity of the toxin in the isolated muscle could be markedly reduced with excess antitoxin, estimated as 3:1 molar ratios of IgG Ab:toxin or better. Toxin neutralization in vivo required higher ratios of Ab:toxin, ranging from 30:1 at high toxin doses and increasing to 100:1 at 10×LD50 toxin. At equimolar Ab to toxin ratios in the isolated muscle, the biological activity of the toxin underwent a statistically significant increase. This paradoxical effect of the polyclonal antisera was serotype selective and independent of the presence or absence of hemagglutinin in the toxin. The enhancement of toxin activity was subsequently localized to occupancy of one of four epitopes on the toxin using monoclonal antibodies to mimic the effect of the antitoxin. The enhancement of toxin activity suggests that botulinum toxin may undergo a conformational change upon binding antibodies to certain domains. This phenomenon could contribute to the observed concentration dependent changes in neutralization efficacy with antitoxin in vivo.

Primum non nocere

The objective of a vaccine for pertussis is to prevent the paroxysmal cough with its complications. Prevention of the paroxysmal cough should prevent transmission of B. pertussis. Clinical studies indicate that a critical level of antitoxin confers protection against pertussis and that the long-lived protective immunity that follows pertussis is best explained by the presence of antitoxin. Vaccine-induced protective immunity can be mediated by a critical level of antitoxin alone. In addition to preventing pertussis with vaccines, we predict this level of antitoxin will also exert epidemiologic control (herd immunity) by inhibition of colonization with B. pertussis and by prevention of the paroxysmal cough which will reduce transmission of this pathogen. Lastly, a pertussis toxoid need not have detectable pharmacologic activity to exert its protective actions: residual activity could exert a deleterious effect on glucose metabolism and exert immunomodulating effects. Accordingly the new pertussis vaccine should contain inactivated pertussis toxoid alone and there is no need to include other components. Several candidates could serve as a satisfactory pertussis toxoid. The admonition to physicians given by Hippocrates, Primum non nocere (first of all do no harm), should be heeded by those responsible for the development and use of the new pertussis vaccine.

Tetanus antitoxin titration by haemagglutination.

The behaviour of sheep cells sensitized with tetanus toxins and toxoids in the presence of antitoxin has been investigated.Sensitized sheep cells were agglutinated in the presence of tetanus antitoxin. The agglutinin titres of such direct agglutinin systems were roughly proportional to the antitoxin content of the sera with some marked exceptions. The discrepancies were associated with the presence of non-specific agglutinins (antibodies other than specific antitoxin) and with the immunization history of the horses providing the sera.When sensitized cells were used as indicators of the presence of free antitoxin in toxin or toxoid-antitoxin mixtures, the correlation within vivotests was quite good at the L+ level of test, but not so reliable at the 0·01 L+ level of test.Sensitized cells preserved with formalin were found to be more reliable and the results with such materials were more reproducible than those obtained with freshly prepared sensitized cells.Cells sensitized with different concentrations of antigen gave different titres by direct agglutination with simple dilution of antitoxin; this difference was very much less in haemagglutination inhibition tests.The LAdose (the volume of toxin or toxoid which when mixed with one unit of antitoxin gave a + + ± end-point) varied according to the sensitivity of the suspension used and the purity of the components of the reacting mixtures.Prozones (zones of agglutination inhibition in the region of antibody excess) were found to be associated with the immunization history of the horses from which the sera were obtained. In general the more thorough the immunization the longer the prozones found. The length of prozones was proportionately increased with decrease in the concentration of sensitizing antigen. Possible reasons for the behaviour of certain antitoxins in these tests have been discussed.I am indebted to Prof. M. Raynaud of the Pasteur Institute, Garches, for the supply of tetanus toxoids of high purity.I am very grateful to Mr Charles Newman for considerable technical assistance.

Studies with H. Pertussis

Summary By intradermal testing in normal and immunized animals it has been shown that both the thermolabile and the thermostable toxins of H. pertussis are definitely although weakly antigenic in rabbits but apparently not in human beings. In normal animals, and in a few humans tested, the agglutinogen shows no primary toxicity but in immunized rabbits and human beings an immune response lasting a day or two is obtained. The toxins, on the other hand, produce a primary toxic reaction in normal animals which lasts for one week and longer; the thermolabile toxin (TLT) produces necrosis. In immunized animals the prolonged primary toxic reaction to the toxins is not shown; the reaction appears as a non-necrotic lesion of duration corresponding to the immune response produced by the agglutinogen. The above-mentioned effects were produced in animals immunized slowly and in a prolonged series of injections either by the subcutaneous or intravenous route with either live or formalin-killed whole organisms or sonic extracts. The presence of antitoxin in the sera of the immunized animals was determined by neutralization tests in the skin of normal rabbits. By this procedure no antitoxin could be detected in human hyperimmune or in convalescent sera containing agglutinins. Both toxins were obtained from both phase III and IV and para-pertussis organisms, but in much smaller amounts than from phase I strains. By neutralization tests the toxins from the several phases and in para-pertussis were shown to be related in antigenic specificity. The fallacy of using toxicity as a sole criterion in evaluation of strains suitable for the preparation of vaccines is discussed. The possible use of the various components of H. pertussis as reagents in dermal tests for susceptibility, in diagnosis and as immunizing agents is discussed.

Staphylococcus antitoxin in the blood and milk of cows and other animals

Summary (1) It has been found that in cattle (male and female) and in goats, as in some other animals, there is a direct relationship between the age of the animal and the staphylococcus antitoxin titre of its blood. Evidence is brought forward to show that the presence of antitoxin in the blood of cows is determined in part by infection of the udder with Staph. aureus , and in the same connection it may be noted that the incidence of udder infection with Staph. aureus also increases with age. (2) As would be expected, in cows and sheep injections of staphylococcus toxoid lead to an increased concentration of antitoxin in the blood, and this is reflected at times by an increase of antitoxin in the milk. (3) Staphylococcus toxin can be demonstrated in the udder secretion of acute cases of staphylococcus mastitis. (4) In the milk whey from normal quarters the amount of antitoxin present is usually 1/80 to 1/40 of that in the blood serum, whereas in colostrum the quantity of antitoxin may equal or even exceed that present in the blood. When the quarter is inflamed, e.g. , as a result of streptococcus infection, the amount of staphylococcus antitoxin in the whey rises. (5) The facts brought forward in this paper have a bearing on the general question of the source of “natural” antitoxin in blood and of antibodies in milk.

Susceptibility of Eskimos to the Common Cold and a Study of Their Natural Immunity to Diphtheria, Scarlet Fever and Bacterial Filtrates

Summary Eskimos are very susceptible to upper respiratory infections on contact with the outside world. Ordinary bacterial infections rarely occur. Diphtheria and scarlet fever are unknown clinically and in a group of about fifty subjects all were negative to the Dick test and all the adults were also Schick negative. Children up to the age of twelve years were invariably Schick positive. Three serums were found to contain antitoxin both for diphtheria and scarlet fever. It is therefore concluded that the immunity to the disease and the negative skin tests depend on the presence of antitoxin. This is interpreted as being due to a natural hereditary immunity dependent upon some non-specific antitoxic mechanism. Skin reactions with filtrates of streptococci isolated from cases of rheumatic fever were mildly positive in a small percentage of cases. Neutralizing antitoxin was demonstrated in all three sera but it was not invariably present for all three toxins. The Eskimos showed a high percentage of positive reactions when tested with a staphylococcus aureus filtrate.

Scarlet Fever.???The Development of Toxin Sensitivity of the Skin in Infants and its Relation to the Presence of Antitoxin in the Blood

Scarlet Fever.???The Development of Toxin Sensitivity of the Skin in Infants and its Relation to the Presence of Antitoxin in the Blood

VIII. THE ANAPHYLACTIC FACTOR IN SCARLET FEVER

In a series of studies 1 on the sensitivity of the skin to scarlatinal streptococcus filtrate toxin and its neutralizing antibody, the following facts have been observed: In the new-born, the skin was usually not sensitive to considerable amounts of scarlatinal toxin, and this lack of sensitivity was not due to the presence of antitoxin in the blood. Relatively few infants developed sensitivity during the early months of life, but by the end of the first year the skin had become sensitive in a large proportion. Although antitoxin was not demonstrable in many young infants whose skin was insensitive to toxin, in older children the lack of sensitivity was practically always accompanied by antitoxin in the blood. Hemolytic streptococci isolated from young infants with infections of the upper respiratory tract in certain instances formed a toxin which was neutralized by convalescent scarlet fever serum, but not by serum

II. THE DEVELOPMENT OF TOXIN SENSITIVITY OF THE SKIN IN INFANTS AND ITS RELATION TO THE PRESENCE OF ANTITOXIN IN THE BLOOD

In a previous paper,1it was shown that skin sensitivity to scarlatinal streptococcus filtrate toxin was present at birth in only a small number of infants, and that in most infants up to 3 months of age, the skin did not react to a considerable amount of the toxin. This lack of skin sensitivity was not entirely due to the presence of antitoxin, since many such infants without a demonstrable amount of antitoxin in the blood had negative skin reactions between one and three months after birth. Those without antitoxin at birth, however, developed some skin sensitivity to the toxin within three months in a larger percentage than those in whom antitoxin was demonstrable. The present paper deals with a study of the skin sensitivity to toxin during later infancy and childhood. It was not possible to follow up the same infants tested at birth except in occasional instances,

III. MODIFICATION OF SKIN SENSITIVITY TO TOXIN AS A RESULT OF NONSCARLATINAL INFECTIONS

In the course of a study1of skin sensitivity to scarlatinal streptococcus filtrate toxin in infants and younger children, it was noted that those in close contact with one another in an asylum showed striking differences from children in isolated homes. These differences, together with their relation to the presence of hemolytic streptococci in the pharynx, and to the presence of antitoxin in the blood, form the subject of this report. In addition, the variation in skin sensitivity which accompanies measles and possibly some other nonscarlatinal infections has been studied. METHOD The skin tests were made as previously described,1with 2, 20 and 50 standard skin test doses (STD) of a potent scarlatinal streptococcus filtrate that contained 100,000 STD per cubic centimeter. Antitoxin determinations were made by toxin neutralization tests. Pharyngeal cultures for hemolytic streptococci were made by rubbing a sterile swab, moistened with salt solution, on the

Skin Reactions to Scarlatinal Streptococcus Filtrate in New-Born Infants and Their Mothers.

Observations on over 150 infants during the first 2 weeks of life, and their mothers, were made by noting (1) the reaction of the skin to 2, 20 and 50 standard skin test doses of scarlatinal toxin, (2) the occurrence of antitoxin in the blood of a number of these infants and their mothers, and (3) later skin tests on a number of the same infants between the ages of 6 weeks and 2 months. It was found that the skin of infants in the first 2 weeks of life does not react to small amounts (2 S. T. D.) of toxin, only one instance of a positive reaction having been observed. With larger amounts (20 and 50 S. T. D.) only a small proportion (about one-eighth) show positive reactions, and the mothers of these babies in practically every instance have positive reactions to two skin test doses. It was quite exceptional for an infant to give a positive reaction to 20 or 50 S. T. D. if the mother was negative to 2 S. T. D. The presence of antitoxin in infant's serum had no relation to the infant's skin test, but is associated with the presence of antitoxin in the mother's blood. No antitoxin was demonstrated in infants of mothers who reacted to 2 S. T. D., but it was rather constantly present in infants whose mothers gave negative reactions to 2 S. T. D., even though skin tests with larger amounts were positive. On later examination of the infants at the age of 6 weeks to 2 months, the skin reaction had become positive in certain babies, to the larger amounts (20 to 50 S. T. D.) of toxin and occasionally to 2 S. T. D.

Relation of Skin Reactions to Scarlatinal Streptococcus Filtrate in Children, to Antitoxin in Blood.

A large number of children over 3 years of age have been given skin tests with varying amounts of a standard scarlatina streptococcus filtrate toxin. In a group giving a positive Dick test with 1 or 2 S. T. D., no antitoxin was demonstrable in the blood. In a few children who were negative to 2 S. T. D., but positive to 5 S. T. D., the antitoxin determinations were somewhat irregular, although when 10 or 20 S. T. D. of toxin or more was necessary to obtain a skin reaction, antitoxin was readily demonstrable, and in those children who gave negative skin tests to 100 or 400 S. T. D., antitoxin was present in the blood in rather large amount.In tests on younger children a similar association of antitoxin with negative skin tests was found. A certain number of infants, more commonly in the first year, but occasionally later, who gave negative skin reactions to 20 or 50 S. T. D. of toxin, had no antitoxin demonstrable in the blood.The association of the presence of antitoxin in the blood with negative skin tes...

Occurrence of Schick Negative Reactions in Absence of Free Antitoxin in Blood Stream.

The object of this communication is to present some observations concerning the relationship of antitoxic immunity to the Schick test.The Schick test, consisting of the intra-dermal injection of 1/50 M. L. D. of diphtheria toxin, shows under proper conditions, a reaction at the site of injection in the non-immune (positive), and a complete absence of reaction in those who are immune (negative). The generally accepted explanation of a negative Schick test is that antitoxin is present in the blood serum, the circulating antitoxin reaching and neutralizing the toxin before tissue damage results.This explanation is of course plausible, especially since we know of no product of the tissues capable of neutralizing toxin, except antitoxin. There was no ground for dispute of this hypothesis, so long as the presence of antitoxin in the blood was predicated, but it now develops as a result of a series of comparative observations with the Schick and the Kellogg1 tests that there are individuals who give absolutely c...

Experiments on the Sensitivity of the Human Skin to the Toxin of the Bacillus of Shiga-Kruse

Summary Experiments are presented showing that the negative skin reaction to the products of the Shiga-Kruse bacillus in human beings depends upon the presence of antitoxin in the blood. The antitoxin is specific, and, as in the case of “natural” immunity to diphtheria, it appears spontaneously in the individual, without a previous attack of the disease or artificial immunization. If this presence of dysenteric antitoxin in human beings provides an actual immunity against infection with dysentery, then the way must be open to attempt an active immunity in the same way as is done in respect to diphtheria and scarlet fever. We have actually begun to make this attempt with dysentery anatoxin. These experiments will be the suject of our next communication.