Two affected brothers and a negative carrier test
Two brothers, Luka (7) and Ivan (4), have haemophilia A, an X-linked recessive bleeding disorder. Their mother, Petra, is healthy. Petra's brother is also affected, but her father and mother are healthy. Petra's sister, Maja, has one affected son and one healthy son. Petra's husband, Tomo, is healthy and has no family history of bleeding disorders. Clotting tests show Petra's factor VIII activity is 48% (reference 50–150%), while Maja's is 62%. Genetic testing finds a known pathogenic variant in the F8 gene in both affected boys. Petra is tested for that same variant on a blood sample, and the result is negative. The counsellor explains that about 10% of carrier women have normal factor VIII activity, that skewed X-inactivation can lower factor VIII, and that a negative blood DNA test can miss a variant present in only some tissues. Petra asks about the risk to a future son and a future daughter.
1. Explain the most likely mode of inheritance and identify who in the family must be carriers.
Read the worked answer
Inheritance
The pattern is X-linked recessive. Only males are affected, affected males have unaffected parents, and the variant passes through unaffected females. Petra's father is healthy, so her affected brother must have inherited the variant from their mother. Her mother is therefore an obligate carrier, and Petra's prior probability of being a carrier is 1/2.
Maja's affected son makes Maja a carrier, which is consistent with the mother being a carrier. Petra's two affected sons are very strong evidence that Petra is a carrier too: if she were not, two independent new mutations in the same gene would be required, which is extremely unlikely. Tomo's healthy status means he cannot pass the variant to a son, because a son receives his Y chromosome from his father.
2. Petra's blood DNA test is negative, her factor VIII is 48%, and she has two affected sons. Explain the most probable explanation and estimate the risk to a future son.
Read the worked answer
Most probable explanation
Petra's blood DNA test is probably a false negative, or she has germline mosaicism. In germline mosaicism the pathogenic variant is present in a proportion of her egg cells but not in her blood cells, so a blood test can be negative while she still has affected sons. Her factor VIII activity of 48% is just below the reference range and is consistent with carrier status and skewed X-inactivation; it supports carrier status but does not prove it. A second tissue (buccal cells) or a more sensitive assay for the familial variant should be tested.
Risk to a future son
If she is a carrier or germline mosaic, each son has a 50% chance of inheriting the variant and being affected. Because two affected sons already make carrier status overwhelmingly likely, the practical risk is close to 1/2. If a second, more sensitive test is also negative, the residual risk is lower but not zero.
3. What is the risk to a future daughter, and what advice would you give the family?
Read the worked answer
Risk to a daughter
A daughter receives a normal X chromosome from her healthy father, so she cannot be affected by this X-linked recessive condition even if she inherits the variant from Petra. Her risk of being a carrier is 1/2 if Petra is a carrier or germline mosaic. Carrier girls can have mildly reduced factor VIII and a small bleeding risk, so carrier testing is still worthwhile.
Advice
Genetic counselling, testing the daughter for the familial variant after birth (or prenatally if the family wishes), and cascade testing of female relatives. The family should be told that a negative blood test in Petra does not remove the risk to sons, and that prenatal or pre-implantation testing is available for the known familial variant.