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Paediatric Laboratory Medicine
Paediatric Laboratory Medicine
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Cystic Fibrosis (CF) Molecular Analysis

Background
Who should be tested?
Testing Methodology
Potential Outcomes & Interpretation of Test Results
Cautions
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Background

Cystic fibrosis is the most common autosomal recessive disorder among Caucasians of Northern European descent. Symptoms are highly variable between patients, but those with ‘classical’ CF produce thick, sticky mucous that clogs their respiratory airways and lungs. This mucous provides a breeding ground for infections, and subsequent damage of lung tissue, which can result in respiratory and heart failure. The mucous may cause gastrointestinal pseudo-obstructions, block pancreatic and bile ducts and interfere with exocrine function. Cystic fibrosis mutations are associated with male infertility in about 10 per cent of infertility cases.

CF is caused by mutations in the CFTR gene located on chromosome 7. Over 1,300 different mutations in the CFTR gene have been identified. The most common of these, deltaF508, is associated with about 70-72 per cent of the CF chromosomes in Caucasians of Northern European descent. The other CFTR mutations are rare although some of these have a high frequency in certain ethnic groups. To be affected with CF, an individual must have two mutations in the CF gene (one inherited from each parent); these may or may not be identical mutations.

The disease is present when a child receives two copies of a defective gene, one from each parent. Any person with one copy of the defective CFTR gene is a cystic fibrosis carrier. Carriers do not have, and will never develop, cystic fibrosis. However, if two carriers wish to have children, there is a one in four chance (25 per cent) that their baby will be born with cystic fibrosis. There is a three in four chance (75 per cent) that their baby will not have cystic fibrosis.

Within the CFTR gene lies a region, the Intron 8 T-tract, which affects the efficiency of the normal CFTR gene. There are three versions of this tract, which vary between individuals: 9T, 7T and 5T. The low-efficiency 5T variant can be considered a very mild mutation by itself and can increase the severity of other mutations within the same gene. For example, one CFTR mutation coupled with the 5T variant may result in mild symptoms of CF or male infertility.

Who should be tested

  • individuals clinically suspected of being affected with Cystic Fibrosis
  • individuals with a family history of Cystic Fibrosis and their spouses, to determine the carrier status of unaffected individuals
  • pregnancies at risk due to a family history of CF
  • parents of a fetus with echogenic bowel detected during ultrasound
  • gamete donors

Testing Methodology

Direct Mutation Analysis: This assay consists of several possible stages:

  1. Recurrent mutation analysis by multiplex-PCR for the common mutations listed below
  2. Direct sequence analysis to identify any point mutations or small insertions/deletions
  3. Dosage analysis to detect deletions/duplications if the CFTR gene
deltaF508N1303KS549NR347P621+1G>T3659delC3849+10kbC>T711+1G>T
deltaF508R553XV520FR1162XdeltaI507394delTT1898+1G>A2184delA
G542XG85ER334WY122XR560T3905insT3876delA2789+5G>A
G551DR117HA455ES549RR347H1078delT1717-1G>A3120+1G>A
S1255XA559TY1092XM1101K2307insA2183AA>G1898+5G>T5/7/9T

Known polymorphisms F508C, I507V, I506V.

Sensitivity of the Test: Detection rates vary significantly between ethnic groups. The recurrent mutations indicated above account for greater than 90 per cent of the mutations in North American Caucasians, and approximately 95 per cent in the Ashkenazi Jewish population.

    • These mutations are the most common cause of cystic fibrosis, however other mutations in the CFTR gene may also cause CF. A negative result does not rule out the possibility that the individual has a rare CF mutation not included in the assay.
      • For example, of 10 people with cystic fibrosis, this test is able to diagnose about 9. If the 10 people are all Jewish, virtually all of them would be detected by this test. If the 10 people are all Caucasian, about eight people would be detected by this test.

Potential Outcomes & Interpretation of Test Results

Reason for referral

CFTR Gene Mutations
allele 1 / allele 2

Explanation

carrier testingnone detected / none detected
  • This individual is unlikely to be a carrier of Cystic Fibrosis.
carrier testingmutation detected / none detected
  • This individual is a carrier of Cystic Fibrosis, and may transmit a mutation to offspring.
  

 

diagnosisnone detected / none detected
  • This result does not support a diagnosis of Cystic Fibrosis.
diagnosismutation detected / none detected
  • This result is unable to confirm a diagnosis of Cystic Fibrosis.
diagnosismutation detected / none detected
  • This result confirms a diagnosis of Cystic Fibrosis.

Cautions:

  • Current molecular testing will not detect all possible mutations in this gene. A negative result does not rule out the possibility that the individual carries a rare CF mutation not included in the assay.
  • Linkage analysis may be used to assess carrier risk when mutations are not detected in families with a clear clinical history of CF. Linked markers are used to calculate the likelihood that an individual has inherited a family mutation. It is not an absolute diagnosis, due to the possibility of recombination between the marker loci and the disease locus.
  • Test results should be interpreted in the context of clinical findings, family history, ethnic background and other laboratory data.
  • This test was developed and its performance characteristics validated by the Molecular Genetics Laboratory at the Hospital for Sick Children. It has not been cleared or approved by the U.S. Food and Drug Administration. The FDA has determined that such clearance or approval is not necessary. This test is used for clinical purposes.

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