EVALUATION OF THE INTRADERMAL SENSITIVITY TEST IN PATIENTS SUBMITTED TO HETEROLOGOUS SEROTHERAPY

Review article

EVALUATION OF THE INTRADERMAL SENSITIVITY TEST IN PATIENTS SUBMITTED TO HETEROLOGOUS SEROTHERAPY.

J. A. AYRES

, B. BARRAVIERA

1 Course for a Degree in Nursing, School of Medicine of Botucatu, São Paulo State University, SP, Brazil; 2 Department of Tropical Diseases, School of Medicine of Botucatu and Center for the Study of Venoms and Venomous Animals – São Paulo State University, SP, Brazil.

ABSTRACT: This review deals with heterologous sera produced and used in Brazil. The authors studied 64 patients. Of these, 35 had been attacked by domestic and wild animals and received antirabies serum; 20 had been bitten by venomous animals (snakes and scorpions) and were treated with specific antivenoms; and 9 had traumas and received antitetanic serum All these patients were submitted to the intradermal sensitivity test before, and 30 days after reveiving heterologous serotherapy. The following results obtained by the authors agree with those in literature:

- The intradermal test using undiluted heterologous serum produced a more acute reaction than that using heterologous serum diluted 1:10;

- The larger the volume of serum, the larger was the wheal directly after inoculation;

- The antigenic concentration influenced the final local reaction;

- The reading 30 minutes after inoculation was always higher than that 15 min after;

- No systemic reaction was observed during the tests;

- The use of promethazine as a prophylactic medication did not protect against the early reactions;

- Tests performed 30 days after serotherapy showed a higher reactivity, probably due to sensitization;

- The intradermal sensitivity test did not allow the authors to predict early reactions.

After these observations, the authors do not recommend the intradermal sensitivity test for patients submitted to heterologous serotherapy. They, however, strongly recommend a careful observation during the infusion and clinical follow up in the first 24 hours.

KEY WORDS: sensitivity test, heterologous serotherapy.

INTRODUCTION

Serotherapy consists of the practical application of knowledge about artificially acquired passive immunity (20). This immunization is acquired by transferring antibodies from a sensitized individual, the donor, to another individual, the receptor. Serotherapy is homologous when both the donor and the receptor belong to the same animal species, and heterologous when they belong to different species (20).

The principles of serotherapy were based on the studies of two researchers, the French Albert Calmette and the Brazilian Vital Brazil. These principles are still used today (36,48,51).

The experiments related to serotherapy started in the 19th century (48,51). In 1884, Calmette (51) extracted venom from glands of venomous snakes of the genus Naja, and then injected repeated doses in experimental animals. This author observed that this procedure made the animals resistant to other venom inoculations. He also observed that the serum of the hyperimmunized animals neutralized the Naja venom. This finding led to the use of this serum for the treatment of human envenoming (51). However, Calmette's conclusions did not consider the specificity of the serum of immunized animals against the specific venom used to produce antivenom. Calmette's antivenom was distributed for use in the general treatment of snake envenoming in various parts of the world, such as Indochina, India, Australia, and Europe (51).

Later, other researchers observed that Calmette's serum antivenimeux only neutralized the effects of Naja venom, with no effective action on the venom of other snakes (51).

Vital Brazil was the first to demonstrate the specificity of the antivenom in 1901, a fact which was neither recognized nor admitted by Calmette. This researcher started then to prepare mono and polyvalent antivenoms in Brazil. The monovalent antivenoms included the anticrotalic and antibothropic sera. Snake antivenom is composed of two monovalent sera, the anticrotalic and antibothropic. Thus, Vital Brazil is considered the creator of heterologous serotherapy with the development of an effective therapy for snakebite victims (9,20,21,36,49).

This serotherapy has been used worldwide for over a hundred years and has also been indicated for the treatment of diphtheria, tetanus, rabies, as well as snake, spider, and scorpion envenoming (3,4,20,41,46,47,51).

Production of heterologous serum consists of hyperimmunization of animals using increasing doses of toxins, toxoids, and/or venoms. In Brazil, horses are the animal of choice due to their considerable capacity of production of plasma and appropriate immune response (40,41). This process lasts approximately 30 days. After this period, a blood sample is drawn to confirm antibody titration in the animals. If an immune response is obtained, the final bleeding is performed by drawing 15 liters of blood from an animal weighing around 500 kg. This procedure is conducted in 2 stages with an interval of at least 48 hours. Antibody purification and concentration are performed, resulting in the final product, the serum, which may or may not be lyophilized. The sera produced in Brazil are bottled and distributed in liquid form and must be kept under refrigeration at temperatures ranging from 2ºC and 8ºC (6,7,37,41).

The purification method most commonly used for the production of hyperimmune heterologous sera is that of serum gamma fraction purified by precipitation using ammonium sulfate. Pepsin then promotes enzyme digestion, causing the partial digestion of immunoglobulins, as well as the production of (Fab')2 fragments (3,20,41). This leads to the loss of the biological characteristics of crude heterologous IgG in the heterologous serum, which may cause adverse reactions to the receptor. This procedure allows the production of a relatively pure serum, which maintains its neutralizing properties. Despite these careful procedures, the heterologous sera produced in Brazil can still cause major side effects (10).

Due to the immunological properties of heterologous serotherapy and the reactions it may cause, a careful investigation is advised of subjects with a history of reactions to previous serum administration. Administration of heterologous serotherapy must take place at an emergency service or a hospital, where appropriate test materials and specialized nursing and medical personnel are available for the management of possible allergic reactions. However, sensitivity tests are still, at least theoretically, the only procedure available to evaluate the patient’s reactivity to a certain allergen.

The authors studied patients submitted to heterologous serotherapy at the University Hospital of the School of Medicine of Botucatu between 1994 and 1996. Their objective was to evaluate the intradermal sensitivity tests using the heterologus serum as the antigen. This testing was performed before and 30 days after serotherapy.

Sixty-four adults of both sexes, 48 males (75%) and 16 females (25%), aged between 13 and 77 years old were evaluated. All these patients came from Botucatu and the surrounding area. Thirty-five individuals had been attacked by domestic animals (dog, cat, horse, and pig) (80%), and wild animals (monkey, bat, fox, and rat) (20%); 20 individuals had been bitten by venomous animals: 15 by Bothrops snakes (75%), 4 by Crotalus snakes (20%), and 1 by the scorpion Tityus bahiensis (5%); and 9 had traumas, 5 caused by sharp objects (56%) and 4 by car accidents (44%). All of these patients received specific heterologous serotherapy depending on the case and medical recommendation.

Diluted and undiluted sera were used in this test. Readings were made 15 min and 30 min after the test.

The diameter of the wheal resulting from the inoculation of undiluted serum was larger than that of diluted serum after both readings. This observation suggests that undiluted serum contain a higher antigenic concentration, therefore causing a more acute reaction. Another observation was that the larger the amount of serum inoculated, the larger the wheal.

In this study, 0.1 ml of the diluted and undiluted sera solution was inoculated resulting in a wheal of approximately 7mm in diameter. Other studies which used 0.02 ml and 0.125 ml of antigen, the wheals were approximately 3 mm and 7.2 mm in diameter, respectively (24,34).

The local reaction was influenced by the antigenic concentration. The test using undiluted serum showed a larger wheal diameter of 1 to 2 mm or more than that using diluted serum. On the other hand, the test using diluted serum showed a wheal 2 to 2.5 mm larger in diameter than that in the control test. These data suggest a clear association between the antigen concentration with the wheal size.

Lima et al. (28) recommend that readings are made 15 and 30 min after the test. Reaction is considered positive when an erythematous wheal develops with or without pseudopodia. If no reaction develops after 15 min, another reading at 30 min is recommended (34).

In this study, the authors observed that the reading at 30 min was more reliable for it showed the signs of a more acute reaction at the inoculation site. These observations agree with those of Lima et al. (28).

Most of the patients in this study were reevaluated 30 days after treatment with heterologous serum. The intradermal tests were repeated utilizing the same serum at the same concentration previously used.

Analyses of the results demonstrate that the reactions caused by undiluted serum were more acute than those caused by diluted serum. Comparing the reactions before specific treatment with heterologous serum with reactions that developed 30 days later, an evident increase in reactivity to the tests was seen. Based on these observations, the authors suggest that the use of heterologous serum contributes to an increase in the size of intradermal reactions.

Horse serum, used as a source of antitoxin, is known to contain a large amount of antigenic proteins. These are capable of inducing the formation of IgE antibodies through sensitization (1,5,22,26,33).

Sensitization begins when an individual receives sensitizing antigens. These promote the formation of IgE antibodies which firmly bind to basophils and mast cells by the Fc fragment (11,22,31). The second stage of the reaction occurs after a period of undefined length when the antigen binds to two specific IgE molecules in the Fab region. Then, the antigen causes, by a chemical process, the degranulation of mast cells with the resultant release of histamines (3,14,22,31).

It is known that the reactions, which can be triggered after treatment with heterologous sera, are related to complex immunological processes. These reactions usually occur in individuals with a history of previous administration of specific antigen (23). Anaphylaxis occurs when basophils and mast cells release their chemical mediators.

Approximately 40% of the individuals submitted to heterologous serotherapy may develop type I hypersensitivity (anaphylaxis), depending on several factors, such as, type of serum, dose, route and method of administration, population characteristics, and previous exposure to sensitizing substances (38).

Barraviera et al. (3) and Cupo et al. (12) report that type I hypersensitivity reactions, also called anaphylactic by Gell and Coombs, are mediated by IgE immunoglobulins. These authors also report that these reactions can, at least theoretically, be detected by skin sensitivity tests.

The intradermal sensitivity test is no longer efficient in the detection of sensitive individuals to horse serum because it has a low predictive value, delays treatment, and does not contraindicate serotherapy. (12,13,15,24,30,37). In addition, the intradermal sensitivity test is not free of risks, as it may trigger hypersensitivity (10,15,18,24,31) (32,34,42,45,50). Spaiti (42) reports that skin testing may sensitize a patient to horse serum. Faria et al. (16) report that anaphylactic reactions caused by intradermal tests are infrequent. The author, however, advises that tests should be performed at emergency services or hospitals, where possible severe systemic reactions are properly treated (16).

In this study, no systemic reaction that might be attributable to the intradermal test was observed. During the test, the patients were properly monitored, and the reactions were restricted only to the inoculation site. Some systemic reactions that occurred were mild and were associated with the main dose of the serum, especially in the case of serotherapy for snake envenoming.

It must be emphasized that the intradermal sensitivity test does not have a predictive value, and therefore, several clinicians have ceased to recommend it because it fails in a high proportion of cases (27). It has also been shown that some patients have reacted locally to the preliminary test without causing systemic reactions. On the other hand, patients who showed no local reaction have occasionally developed an intense general reaction after injection of the total serum dose. This phenomenon has already been demonstrated by several investigators (12,15,25,30,35,43).

In view of possible early reactions to the treatment with heterologous serum, some authors propose premedication with the administration of antihistamines, such as H1 and H2 blocking drugs, vasoconstrictors, and hormonal anti-inflammatories (2,6,7,37).

Wen (52) reported that to date there is no therapeutic protocol to completely prevent or reduce the frequency of adverse reactions.

In our study, antihistamines showed to be inefficient in the prevention of type I hypersensitivity. Of the patients in group 1, who received 50 mg of promethazine before treatment, 7.4% showed early reactions, while of the patients in group 2, who did not receive antihistamines, 2.7% showed early reactions.

A study on the effect of promethazine for the prevention of immediate reactions to heterologous serum demonstrated that it did not protect against immediate reactions (52). In addition, we may suggest that immediate reactions were related to the amount of inoculated antigen and that the route of administration may be involved in the development of the reactions, as these affected the patients who received intravenous therapy.

With regard to the treatment of early reactions, subcutaneous adrenaline is the drug of choice, since it controls blood pressure and other adverse effects of anaphylatic mediators, and reduces the further release of mediators from mast cells and basophils (1,39). Antihistamines, bronchodilators, and corticosteroids may also be used to treat these reactions.

Bucaretch et al. (8) evaluated the protection of the H1 and H2 blocking drugs in children less than 15 years old, victims of bothropic and/or crotalic envenoming and observed that these drugs did not provide a safe protection against early reactions.

Sutherland (44) reported that the use of antihistamines is associated with possible side effects, such as sedation, tachycardia, and hypotension. These effects may make the treatment of anaphylaxis more difficult. On the other hand, Fatovich (17) suggests that there is not evidence supporting the use of antihistamines as prophylaxis of immediate reactions.

The use of corticosteroids in envenoming remains controversial. Corticosteroids are administered to prevent and reduce hypersentivity reactions. However, as the onset of the pharmacological activity of corticosteroids is four to six hours following administration, these are not effective in preventing early reactions (42).

Griffen (19) reported that the use of a single dose of hydrocortisone prior to antivenom therapy was an effort to prevent or ameliorate immediate reactions. The author also reported that although there was no evidence to support the use of steroids prophylactically to prevent immediate reactions, their use has been advocated.

Laprinzi (29) and Patterson (35), however, contraindicated the use of corticosteroids as antihistaminic agents due to their slow pharmacological action.

In this study, the authors observed that the patients' early reactions were mild. The procedures adopted were the interruption of serum infusion, hemodynamic control, and observation over 15 minutes. After this, the vital parameters returned to normal and the signs and symptoms disappeared. Antivenom therapy was completed with no clinical alterations shown by the patients. It should be highlighted that in all cases of early reactions there was no need for the use of drugs to block the effects observed or previous desensitization.

Based on the data in literature and on the findings of this study, the authors came to the following conclusions:

- The intradermal test using undiluted heterologous serum produced a more acute reaction than that using heterologous serum diluted 1:10;

- The larger the volume of serum, the larger was the wheal directly after inoculation;

- The antigenic concentration influenced the final local reaction;

- The reading 30 minutes after inoculation was always higher than that 15 minutes after;

- No systemic reaction was observed during the tests;

- The use of promethazine as a prophylactic medication did not protect against early reactions;

- Tests performed 30 days after serotherapy showed a higher reactivity, probably due to sensitization;

- The intradermal sensitivity tests did not allow the authors to predict early reactions.

Finally, the authors do not recommend the intradermal sensitivity tests for patients submitted to heterologous serotherapy. They, however, recommend a careful observation during the infusion and clinical follow up in the first 24 hours.

REFERENCES

Accepted 18 March 1998

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CORRESPONDENCE TO:

J. A. AYRES – Curso de Graduação em Enfermagem da Faculdade de Medicina de Botucatu, UNESP, Distrito de Rubião Junior, s/n, CEP 18618-000, Botucatu, São Paulo, Brazil.

Publication Dates

• Publication in this collection
  17 Sept 1999
• Date of issue
  1999

History

• Accepted
  18 Mar 1998
• Received
  18 Mar 1998