Key signs and steps for diagnosing primary immunodeficiency in children with ASD.

A primary immunodeficiency (PID) is a group of more than 400 genetic disorders that affect the immune system, making it less effective at protecting the body against infections and other threats. In children with neurodevelopmental problems, identifying a PID can be crucial, as a poor immune response can aggravate existing conditions or contribute to the development of additional symptoms.

A timely diagnosis allows for the initiation of specific treatments. As parents of children with neurodevelopmental problems such as autism, it is important to be alert to signs such as recurrent infections, persistent fatigue or unexplained symptoms. This article aims to guide families in identifying whether they are facing a case of PID. 

It is important to remember that the immune system is like the body's army, protecting us from invaders that can cause disease, such as viruses, bacteria and fungi. To do this, the immune system uses two main types of responses: innate immunity and adaptive immunity. The innate immune response (first line of defense) is carried out by different types of cells capable of reacting immediately to microorganisms, these cells include: 

1. Phagocytic cells: neutrophils and macrophages.
2. Blood cells: eosinophils, basophils and mast cells; which release inflammatory proteins and are toxic to invading microorganisms.
3. Natural killer (NK) cells: one of whose functions is to kill infected cells. 

Instead adaptive immune response, which is our second line of defense, is more specific and “sophisticated” since it is activated after the innate response. It adapts and improves with each encounter with a specific invader. The main cells involved in this process are T and B lymphocytes. T lymphocytes attack microorganisms that are inside the body's cells and synthesize chemical substances called cytokines, which help activate other cells of the immune response. B lymphocytes, after maturing into plasma cells, synthesize immunoglobulins (or antibodies) whose mission is to neutralize specific microorganisms and help the function of phagocytic cells. 

Considering and ruling out primary immunodeficiency disorders (PIDs) in autistic children is crucial because alterations in the immune system can exacerbate the health and behavioral challenges associated with autism. Although autism spectrum disorder (ASD) is primarily associated with difficulties in communication, behavior, and social interaction, there is growing evidence that some children with ASD have immunological alterations that could influence their symptoms and quality of life.

It is important for parents to be alert to the symptoms presented by the child and the occurrence of recurrent infections so that the diagnosis is made and treatment is started as soon as possible, in order to avoid serious complications, such as severe respiratory impairment and septicemia, for example, which can be fatal.

This is a painstaking process that involves a series of detailed laboratory tests that are necessary to assess everything from immune cell counts to antibody functionality and other key components. 

The main signs of primary immunodeficiency are:

• 4 or more ear infections in less than 1 year.
• 2 or more respiratory tract infections in less than 1 year.
• Use of antibiotics for more than 2 months without effect.
• More than two cases of pneumonia in less than 1 year.
• Delay in child development.
• Recurrent intestinal infections.
• Emergence of complications from vaccines.
• Frequent appearance of skin abscesses.
• In addition, if the family has a history of primary immunodeficiency or the child is the child of a consanguineous marriage, there is a greater chance of developing primary immunodeficiency.

Symptoms of primary immunodeficiency usually appear in the first months of life; however, in some cases, symptoms may only appear in adulthood, as they depend on the type and severity of the genetic alteration. These may be noticed in any part of the body, as they may affect any organ or system. However, symptoms related to the respiratory system are the most noticeable, which may lead to primary immunodeficiency being underdiagnosed and managed as respiratory and infectious diseases of childhood.

Thus, in case of suspected primary immunodeficiency, the initial study should include:

• Complete Blood Count: This basic analysis evaluates the white blood cell (leukocyte) count, red blood cell (RBC) count, and platelet count. In the context of PID, special attention is paid to leukocyte levels and the study of their subtypes (lymphocytes, neutrophils, monocytes, eosinophils, and basophils).

Abnormalities in white blood cells may suggest certain types of PID. For example, a low neutrophil count may be related to immunodeficiencies that affect the production of these cells, while a low lymphocyte count may indicate defects in B or T cells.

• Serum Immunoglobulins (IgG, IgA, IgM, IgE): This test measures the levels of the main antibodies in the blood: IgG, IgA, IgM and IgE.

Immunoglobulins play a crucial role in defending against infections. A deficiency in IgG, for example, is common in many types of primary immunodeficiencies, as this is the most abundant immunoglobulin in the body. Absence or low levels of IgA or IgM can indicate specific types of PID. Each type of immunoglobulin has specific functions, so a particular deficiency offers clues to the nature of the immune defect.

• Lymphocyte Typing by Flow Cytometry: This study allows the counting and classification of different types of lymphocytes (T cells, B cells and NK cells) in the blood.

A low T, B, or NK cell count suggests specific failures. For example, low T cell counts may point to defects in cell-mediated immunity, characteristic of severe combined immunodeficiencies. B cells, responsible for antibody production, may be absent in certain types of humoral immunodeficiencies. Flow cytometry can identify problems not only in number, but also in the distribution of specific subtypes, which is critical in diagnosing PIDs.

• T Cell Function Analysis: It is a functional test to evaluate whether T lymphocytes respond appropriately to certain stimuli in vitro.

The response of T cells to an antigen or mitogen is measured in this test. If T cells are not activated properly, this suggests a deficiency in cellular immunity. These tests are especially useful for detecting combined immunodeficiencies, where T cells are defective.

• Antibody Response Assessment: This test analyzes the ability of the immune system to produce antibodies in response to a vaccine (such as tetanus or pneumococcal vaccine) given in the past or at the time of diagnosis.

People with PID often have a poor or absent response to certain vaccines. An inadequate response suggests dysfunction in B cells or antibody production, which is characteristic of humoral immunodeficiencies.

• Complement Analysis (C3, C4 and CH50): measures the levels and functionality of proteins of the complement system, a key component of innate immunity.

Low levels of C3 or C4 may indicate a deficiency in the complement system. In addition, CH50 assesses the overall ability of the complement to destroy bacteria and other pathogens. Complement deficiencies are often associated with recurrent bacterial infections and autoimmune diseases.

• Genetic Analysis: Genetic studies look for mutations in specific genes that have been associated with different types of PID.

Identification of mutations in specific genes, such as the BTK gene (associated with X-linked agammaglobulinemia) or the ADA gene (adenosine deaminase deficiency in severe combined immunodeficiency), confirms the diagnosis and allows for precise classification of the type of immunodeficiency. With the advancement of sequencing techniques, such as whole-exome sequencing, this analysis is an essential tool for definitive diagnosis.

• Neutrophil Function Test: assesses the ability of neutrophils to kill pathogens, particularly through tests such as nitroblue tetrazolium (NBT) reduction or the oxidative burst test.

Reduced neutrophil activity may indicate a dysfunction in their ability to eliminate pathogens, characteristic of diseases such as chronic granulomatous disease (CGD). This test is important to confirm specific defects in the antimicrobial defense system.

• IgG Subclass Levels: Assesses specific IgG subclasses (IgG1, IgG2, IgG3, IgG4). Some primary immunodeficiencies show a reduction in specific IgG subclasses, such as in selective IgG2 deficiency. This may be associated with recurrent infections, and its identification helps direct treatment towards replacement therapies.

The integration of these analyses allows the immunologist to establish a complete profile of the patient's immune system, helping to identify whether the problem is in antibody production, in the cellular response or in other components of the immune system. Accurate diagnosis is essential to determine the appropriate treatment, whether through intravenous immunoglobulins, prophylactic antibiotics or gene therapies for specific genetic defects.

It should be noted that the vast majority of primary immunodeficiencies are diagnosed with a correct clinical history and physical examination, a complete blood count and a determination of plasma immunoglobulins (Ig), and these tests are available in most laboratories and in primary care. The complete blood count with differential blood cell count helps us to recognize lymphopenia or neutropenia, the serum Ig count (IgG, IgM and IgA) mainly evaluates hypogammaglobulinemia. We must also consider special situations of differential diagnosis or directed suspicion where IgE can be very useful, ruling out each of the cell lines in their form, count and production helps us to have an early diagnosis.

A second diagnostic step would include the determination of functional antibodies against protein antigens, such as tetanus, or polysaccharides such as pneumococcus or Salmonella typhi. These could be absent even when total Ig levels were previously in the normal range. 

Lymphocyte immunophenotype helps us evaluate the surface characteristics of lymphocytes that can be identified using immunotyping techniques, thereby classifying lymphocytes according to their different subpopulations:

1. T lymphocytes (CD3+):
   • CD4+ (Helper T cells): They help activate other types of immune cells.
   • CD8+ (Cytotoxic T lymphocytes): Destroy infected or tumor cells.
2. B lymphocytes (CD19+): They are responsible for the production of antibodies.
3. NK (Natural Killer) cells: Part of the innate immune system, capable of recognizing and destroying infected or tumor cells without the need for prior sensitization.

When analyzing the immunophenotype, it is a guide to identify specific markers on the surface of these lymphocytes, such as CD45, CD19, CD20, CD3, CD4, CD8, among others. This information is essential to understand the state of the immune system and help in the diagnosis and monitoring of various diseases.

Another important point to evaluate is the proliferative function of lymphocytes, which is essential for the adaptive immune response. When lymphocytes are activated by antigens, receiving signals from other immune cells (such as antigen-presenting cells) or cytokines, they initiate a proliferation process that results in the creation of a clone of cells specific to the antigen. It evaluates the capacity of lymphocytes to react in vitro to different stimuli in order to draw a parallel to what could occur in vivo. It is used to detect cases of combined immunodeficiencies.

• B-cell activation:
  • Antigen presentation, B cells are activated when they recognize a specific antigen through their B cell receptors (BCR). 
  • Interaction with T Cells: Helper T cells (CD4+) can help B cells by releasing cytokines, promoting B cell proliferation and differentiation into antibody-producing plasma cells.

• T-cell activation:
  • Antigen Recognition: Cytotoxic T cells (CD8+) are activated when they recognize antigen fragments presented by major histocompatibility complex (MHC) molecules on the surface of infected cells. 
  • Costimulatory Signals: Interaction with costimuli (e.g. CD28 with B7) is crucial for their activation and subsequent proliferation. 
• Growth and Differentiation:
  • Cytokines: Cytokines such as IL-2 play a key role in lymphocyte proliferation and survival. IL-2, in particular, is essential for clonal expansion of T cells. 
  • Formation of Memory Cells: After proliferation, some lymphocytes differentiate into memory cells, which allow a faster and more effective response if they encounter the same antigen in the future.
• Proliferation Control:
  • Lymphocyte proliferation is tightly regulated to avoid inappropriate immune responses, which can lead to autoimmune diseases. Negative feedback mechanisms limit proliferation once the antigen has been eliminated.

In the case of leukocytosis with normal or high Ig, and the presence of abscesses in the skin or different organs, a test of the oxidative capacity of granulocytes using flow cytometry should be added (to rule out chronic granulomatous disease).

Another important study to take into account is that of leukocyte adhesion deficiency, although it is an extremely rare entity, it is key to rule it out since it includes a group of primary immunodeficiencies that are characterized by the inability of leukocytes to migrate correctly to infection sites. This is due to alterations in the adhesion molecules that are crucial for the diapedesis process, which is where leukocytes leave the bloodstream and head to inflamed tissues or tissues where the injury exists. The two most common types are:

• Leukocyte Adhesion Deficiency Type 1 (LAD-1): Characterized by a deficiency in integrins (particularly CD18), which are necessary for the adhesion of leukocytes to endothelial cells.
• Leukocyte Adhesion Deficiency Type 2 (LAD-2): Less common, it is due to the alteration of selectin adhesion molecules.

A diagnosis of PID opens the door to targeted interventions, such as intravenous immunoglobulins or prophylactic antibiotics, which can significantly reduce infections and improve the child's overall well-being. This, in turn, allows the child to be better able to benefit from behavioral and educational therapies.

At Enevia we offer specialized consulting services and different tests that can guide you in different areas such as neurology, genetics, nutrition and general medicine, as well as help you make the right decisions and analyze medical tests to achieve effective treatment for the pathologies that you may suffer from.

Enter our website through www.eneviacare.com and www.eneviahealth.com and you will be able to find the services that we can offer you.

At Enevia, we are your ally in health!

Article written by Enevia Health Advisor and Collaborator: Dr. Julianny Albarran 

Medical surgeon, general medicine with more than 5 years of experience in the field.

In our blog we also have more interesting articles about immunology and primary immunodeficiencies in neurodevelopmental pathologies that could be of interest to you or your loved ones. Below are some of them:

https://test.eneviahealth.com/blog/errores-innatos-sistema-inmune-infancia/

https://test.eneviahealth.com/blog/el-vinculo-entre-inmunidad-y-el-tea/

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