Carrier screening and prenatal detection of chronic granulomatous disease in Iran

OBJECTIVE: To develop an application that is simple and reliable using the nitroblue tetrazolium (NBT) method that clearly differentiates chronic granulomatous disease (CGD) patients with heterozygous carriers in groups suspected with CGD.

METHODS: This study was carried out in Shiraz University of Medical Sciences from October 2002 and March 2004. The study included 260 samples consisting of 123 children (2-24 months) and 106 neonates (<2 months), either suspected with bacterial infection or are immunodeficient, and 31 cord blood samples. Fifty healthy adult individuals were also diagnosed as normal control. Neutrophil reduction of NBT can be stimulated in vitro by protein kinase agonists such as phorbol myristate acetate (PMA), resulting to superoxide anion release.

RESULTS: The PMA is an exceptionally powerful stimulant and when we used it in conjunction with adherence of glass slides, it causes transformation of nearly 100% of all normal neutrophils, and reduces NBT to formazan deposits. Of 260 blood samples, 12 unrelated CGD patients and 16 carriers of X-linked or autosomal recessive CGD patients were diagnosed. The carriers had a range of 15-75% stimulated neutrophils.

CONCLUSION: We have established a PMA-stimulated NBT test for the detection of CGD patients, which clearly differentiate the CGD patients from heterozygote carriers. The results in the cord fetal blood samples indicate that this test may be used for antenatal diagnosis of affected boys, carrier females and autosomal recessive variants of CGD. The technique is simple, fast, inexpensive, and requires only a few microliters of blood.