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Man swabs his cheek for a genetic test.

Genetic testing

Genetic testing looks for variants in genes that are known to cause primary immunodeficiency (PI) and may fast-track your diagnosis.

What is genetic testing?

Primary immunodeficiencies (PIs), also known as inborn errors of immunity (IEIs), are disorders caused by changes, or variants, in a person’s DNA. Four nucleotides make up DNA: adenine (A), thymidine (T), guanine (G), and cytosine (C). The order of these nucleotides, or the DNA "sequence," encodes instructions for making proteins, cells, and organs, similar to how specific sequences of letters form words, sentences, and paragraphs.

When patient DNA shows a difference in sequence relative to reference DNA, that difference is known as a variant.
When DNA from a person shows a difference in sequence relative to reference DNA, that difference is known as a variant.

Variants in the DNA sequence lead to genetic differences between people. Genetic testing looks for these variants. Some variants do not significantly change the overall genetic instructions, and those variants do not cause disease. Other variants do significantly change the instructions, and those types of variants can cause genetic conditions like PI.

Healthcare providers use genetic testing to confirm a diagnosis based on clinical symptoms or laboratory test results, or to reach a more precise diagnosis if symptoms don’t point to a particular disorder. Genetic testing can also determine if someone who does not yet have symptoms is at high risk for developing PI.

Results from genetic testing can not only provide a genetic diagnosis by pinpointing the gene variant responsible for PI but can also inform treatment and clinical management, family planning decisions, and whether relatives are at risk of PI. For example, several genetic variants with different patterns of inheritance cause chronic granulomatous disease (CGD). Knowing which variant an individual with CGD has is key for family planning and understanding who else in the family is at risk.

Genetic testing is a powerful tool. However, it’s important to understand that all of us have differences in our DNA compared to other people, and not all genetic variants are harmful or cause medical conditions. In addition, the genetic basis for some of the most common types of PI remains unknown, so not everyone who gets genetic testing will receive a genetic diagnosis. For some people, genetic testing will only rule out variants they do not have.

Genetic testing carries unique risks

Information from genetic testing is more sensitive than other types of medical information. In 2008, Congress passed the Genetic Information Non-discrimination Act (GINA). GINA bans employers and health insurance companies from using genetic information in employment or coverage decisions. GINA also clarified that genetic information is a type of medical information subject to Health Insurance Portability and Accountability Act (HIPAA) privacy rules.

However, GINA has limitations. For example, the military is exempt from its employer regulations and genetic information can legally be used to determine someone’s eligibility for life or long-term care insurance. In addition, law enforcement can subpoena genetic information from medical records, which could implicate an individual or their relatives in a crime.

In addition to being used in ways you may not agree with, your genetic testing results can reveal information you may not anticipate. This information can range from secondary findings that identify a genetic condition aside from PI to learning that your presumed parents are not your biological parents. Genetic counselors can help individuals understand and think through these scenarios before undergoing genetic testing.

Types of genetic tests

Generally, there are three types of genetic testing used to diagnose PI: panel testing, whole exome sequencing (WES), and whole genome sequencing (WGS). All three use the same underlying technology, which reads a person’s DNA sequence.

However, the different types of testing differ in scope.

Panel testing is the most widely available type of genetic testing but is also the most limited in scope. It looks for variants in a limited number of predetermined genes, typically several hundred, known to be related to a particular diagnosis or set of diagnoses.

Sometimes, individuals who have the genetic condition a panel tests for get false negative results—not because the test is faulty but because the individual has a variant that is not covered by the panel. Panels for the same diagnosis can differ between laboratories and the details of what the panel does and does not cover affect the results.

Some panels only include the most common genes associated with a diagnosis; others may not be updated with genes that have only recently been linked to the diagnosis. Often, panels only include the parts of a gene that code for its protein (also known as exons), which means that any variants in non-coding regions may be missed. However, variants in these non-coding regions can result in PI just as variants in exons can.

With panel testing, both the individual getting tested and the provider ordering the test must understand the limitations of the panel.

Whole exome sequencing (WES) is broader than panel testing and picks up variants in all of the exons of all 20,000+ human genes. In addition to what panels find, WES can turn up variants in genes that:

  • Are linked to PI but are not included in available panel tests.
  • Have not previously been linked to PI but have a known function in the immune system.
  • Have not been previously linked to PI or the immune system.
Apple next to an apple slice to demonstrate the genome versus the exome.
Whole exome sequencing is like looking at a slice of an apple, while whole genome sequencing looks at the entire apple.

Since WES turns up many variants in many genes, how the data are analyzed is key to getting useful information. A study in the United Kingdom of more than 49,000 people found that each person, on average, had thousands of variants identified by WES. If WES only finds variants in genes that haven’t been linked to PI before, it can be difficult to make sense of the data.

In the past, clinicians generally used WES as a second-tier test when panel testing did not identify a PI-causing variant, but it’s being used as a first-tier test more and more. Like panels, WES returns only variants in exons (the parts of a gene that code for its protein), not variants in non-coding DNA. The analysis of WES data also prioritizes genes most relevant to the condition for which the person is being tested.

Whole genome sequencing (WGS) is the most comprehensive type of genetic testing available. In WGS, all of a person’s DNA is sequenced, including gene exons, non-coding bits in a gene called introns, and non-coding regions between genes. In addition to the types of variants WES can detect, WGS can also detect variants in:

  • Gene introns (non-coding spacers between the exons).
  • Stretches of DNA between genes, which may house sequences important for turning genes on and off.

Again, how WGS data is analyzed is critical to extracting useful information. WGS turns up even more variants than WES, which means even more potential leads to follow up.

Test results

Because the goal of genetic testing is to find variants causing a particular condition, computers analyze the raw sequence data to look for the variants most likely to cause PI. Genetic testing reports provide details on the variants that were found, if any.

Individuals undergoing genetic testing should talk to their healthcare provider or a genetic counselor about the implications of the testing, including:

VUS are variants that do not have strong evidence on whether they affect the immune system in a way that causes PI or not. Secondary findings are genetic variants linked to conditions other than PI.

In genetic testing, getting a positive test result means that the panel, WES, or WGS found a genetic variant that causes PI. On the report, this variant may be labeled pathogenic or likely pathogenic. This means that there is published scientific evidence that the variant found in your DNA disrupts the immune system in a way that is consistent with PI.

You can also get an indeterminant or negative result from genetic testing.

Indeterminant results mean that no pathogenic or likely pathogenic variants were found that are consistent with PI but that one or more VUS were found. While VUS may be detected in raw genetic data, not all laboratories report them unless the provider ordering the test opts for them to be included. In many cases, your physician will determine whether there is additional testing that can help to determine whether a VUS may be associated with PI, which often includes testing family members.

A negative result means that no pathogenic, likely pathogenic, or VUS variants were found. However, a negative result does not mean that there is no genetic variant underlying your PI. There are certain types of variants, such as large sections of DNA that are missing or duplicated, that next-generation sequencing methods do not detect well. In addition, specialized analysis of the raw data may be necessary to detect some genetic causes of PI, such as somatic variants

Because WES and WGS cover a large part of a person’s DNA, testing may find variants associated with conditions other than PI. These variants are called secondary findings.

The American College of Medical Genetics and Genomics (ACMG) recommends that all laboratories look for and return secondary findings of pathogenic variants within a limited list of genes. These variants cause preventable or treatable genetic conditions that tend to be asymptomatic for long periods of time before striking suddenly, such as familial hypercholesterolemia. The goal of reporting these findings is to alert the individual and their healthcare provider to ‘hidden’ genetic conditions that can be managed if they are caught in time. Increasingly, even panels include the ACMG genes.

The ACMG list is considered a minimum list of secondary findings that should be reported for WES and WGS, but some laboratories may offer additional secondary findings if the ordering provider opts in.

Reanalyzing genetic testing data

Unlike other types of medical tests, your genetic sequence does not change over time. That means that your raw genetic data can be reanalyzed periodically as labs update their lists of PI-related or immune system-related variants and reclassify VUS. Be sure to get and retain a copy of your raw genetic testing data for this purpose.

Carrier testing for autosomal or X-linked recessive PIs

Panel testing, WES, and WGS can identify gene variants not only in individuals with PI but in their unaffected family members. In cases of a family history of autosomal recessive or X-linked recessive PI, family members, such as the parents or siblings of the person with PI, can undergo carrier testing. Carrier testing determines if someone ‘carries’ one copy of a gene variant linked to PI. In most cases, carriers do not themselves have PI symptoms, although there are exceptions, such as women who carry the variant that causes X-linked chronic granulomatous disease (CGD).

Prospective parents should seek information from healthcare professionals such as pediatricians, genetic counselors, immunologists, and obstetricians on current medical advances relating to the PI of concern. Genetic counseling is highly recommended in families with a known history of PI, as it allows the option for early detection, diagnosis, and treatment.

Prenatal testing

It is undeniable that raising a child with PI places a physical, emotional, and financial burden on families, and many families might wish to maintain their current family size to avoid the possibility of having another affected child. These choices are very personal.

In families with a PI, parents may wish to know whether future babies are affected by the disorder. Testing to determine whether unborn babies are affected is possible in most situations where the immunodeficiency has a known genetic cause, such as X-linked agammaglobulinemia (XLA) or RAG1 severe combined immunodeficiency (SCID). However, prenatal testing is only possible if the genetic cause for a PI has been defined. There are several methods of prenatal diagnosis for PI.

Chorionic villus sampling (CVS) or amniocentesis can obtain a fetal sample for chromosome, genetic, or biochemical testing. CVS is usually scheduled at 10-13 weeks of pregnancy and involves retrieving a tiny sample of the developing placenta from the womb. Amniocentesis is typically performed at 16-17 weeks of pregnancy and involves the withdrawal of amniotic fluid containing fetal cells. Both procedures have a small risk of miscarriage that should be balanced against the benefits of the testing.

Several types of analysis can be done with CVS or amniocentesis samples:

  • Gene variant analysis: This method is 100% reliable if the PI-causing variant is known.
  • Linkage analysis: If the PI-causing variant is unknown, but the diagnosis is definite, prenatal diagnosis may be possible using linkage analysis. This will be slightly less reliable than gene variant analysis.
  • Enzyme analysis: The concentration and/or function of the enzyme affected in a certain PI can be measured. Examples are adenosine deaminase (ADA) and purine nucleoside phosphorylase (PNP), deficiencies which both cause forms of SCID. Both enzymes can be analyzed in a CVS sample taken at 11-12 weeks of gestation.

Noninvasive prenatal testing (NIPT), sometimes called noninvasive prenatal screening (NIPS), analyzes small fragments of DNA circulating in a pregnant woman’s blood. NIPT involves taking a blood sample from the mother and is not associated with any risk of miscarriage.

Unlike most DNA, which lives inside cells, these small DNA fragments float freely outside cells and are called cell-free DNA (cfDNA). cfDNA comes from cells that die off and are broken down so that their contents, including DNA, are released into the bloodstream. During pregnancy, a mother’s blood contains a mix of cfDNA that comes from her cells and cells from the placenta. The placenta is tissue in the uterus that links the fetus and the mother’s blood supply. The DNA in placental cells is usually identical to the DNA of the fetus. 

Because of the technical challenges of determining whether a particular DNA fragment is from the placenta or the mother, NIPT is largely clinically available only for disorders that involve large insertions or deletions of DNA or extra chromosomes. Some clinical labs are working on techniques that allow single-gene variants to be detected through NIPT, but NIPT for single-gene disorders, which includes most genetically-defined PIs, is currently limited.

Analyzing cfDNA from the placenta provides an opportunity for early detection of certain genetic abnormalities without harming the fetus. However, NIPT is a screening instead of a diagnostic test, so it does not give a definitive answer about whether or not a fetus has a genetic condition. The test can only estimate whether the risk of having certain conditions is increased or decreased. If NIPT indicates an increased risk of a genetic condition, more definitive diagnostic testing, such as CVS or amniocentesis can confirm the results. In addition, the test may detect a genetic condition in the mother.

Pre-implantation genetic diagnosis (PGD) is a useful option for couples who are at high risk of having a child with a genetic disorder. It involves in vitro fertilization (IVF) of a number of eggs, followed by genetic testing of a single cell from each embryo, and subsequent implantation of embryos that do not have the PI-causing gene variant.

Implantation of an unaffected embryo via IVF in individuals with SCID, leukocyte adhesion deficiency (LAD), CGD, and XLA has been previously reported. Evidence also supports the efficacy of PGD in the detection of other immunodeficiencies such as Wiskott-Aldrich syndrome, X-linked hyper IgM syndrome (HIGM), NEMO deficiency, and ataxia-telangiectasia.

Note that the use of donor egg or sperm to conceive a child via in vitro fertilization (IVF) can be carry a risk of PI if the health or carrier status of the donor has not been established.

Talk with your provider

A few questions to ask your provider:

  • Have I had a genetic test? 
  • If I have previously had a genetic test, should I consider retesting since genetic tests have expanded and include over 400+ genes (i.e., a PI panel)?
  • There are no-charge genetic tests available with a doctor’s referral; do I qualify for one? 
  • Could the results of a genetic test alter my treatment or management program? 
  • While I understand testing is not for everyone, can you advise why you would not recommend me for a genetic test?
doctor with patients

No-charge genetic testing

There are currently two sponsored, no-charge genetic testing programs available for people who are suspected of having either APDS (activated PI3K delta syndrome) or chronic neutropenia/WHIM syndrome. Both programs include pre- and/or post-testing genetic counseling services that provide support to help your family better understand the testing process, what to anticipate in terms of results, and what information is needed by your physician. 

Talk to your provider to determine if you meet the criteria to qualify for either genetic testing program.

navigateapds

navigateAPDS

This program is available to patients in the U.S. and Canada who meet any two or more of the following bulleted criteria below:

Clinical features:

  • Bronchiectasis.
  • Lymphadenopathy for greater than one month.
  • Chronic hepatomegaly or chronic splenomegaly.
  • Severe, persistent, or recurrent Herpesviridae infections (e.g., EBV, cytomegalovirus).
  • Enteropathy.
  • Lymphoma at 0-25 years - meets the 2 eligibility criteria.
  • Lymphoma at ≥ 26 years of age - requires second eligibility criteria.

Labs:

  • Elevated levels of immunoglobulin M (IgM).
  • Increased number of follicular helper T cells.
  • Reduced number of naïve B cells.

History:

  • Common variable immune deficiency (CVID) phenotype or direct family member with CVID phenotype.
  • Relative with PIK3CD or PIK3R1 genotype (first or second degree) - meets the 2 eligibility criteria. 
path 4 ward logo

PATH4WARD

This program is available to patients in the U.S. and Canada who meet all three of the following criteria:

  1. You suspect you have congenital neutropenia OR a primary immunodeficiency.
  2. You have any prior history or a current absolute neutrophil count (ANC) of less than or equal to 1,000 on multiple occasions (not related to drugs, chemotherapy, or a viral infection).
  3. You have one or more of the following:
    1. Repeat and/or severe infections for you or a family member.
    2. Lymphopenia, or not having enough white blood cells or lymphocytes to help fight infection.
    3. Hypogammaglobulinemia, or low antibody levels that make it hard to fight infections.
    4. Warts that are hard to treat or won’t go away.
    5. A family history of neutropenia.

Download information about these programs to take to your healthcare provider.

This page contains general medical and/or legal information that cannot be applied safely to any individual case. Medical and/or legal knowledge and practice can change rapidly. Therefore, this page should not be used as a substitute for professional medical and/or legal advice.