The increasing number of laboratories offering molecular genetic analysis of the CFTR gene and the growing use of commercial kits strengthen the need for an update of previous best practice guidelines (published in 2000). The importance of organizing regional or national laboratory networks, to provide both primary and comprehensive CFTR mutation screening, is stressed. Current guidelines focus on strategies for dealing with increasingly complex situations of CFTR testing. Diagnostic flow charts now include testing in CFTR-related disorders and in fetal bowel anomalies. Emphasis is also placed on the need to consider ethnic or geographic origins of patients and individuals, on basic principles of risk calculation and on the importance of providing accurate laboratory reports. Finally, classification of CFTR mutations is reviewed, with regard to their relevance to pathogenicity and to genetic counselling.

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Primary ciliary dyskinesia (PCD) is a genetically heterogeneous disorder characterized by chronic destructive airway disease and randomization of left/right body asymmetry. Males often have reduced fertility due to impaired sperm tail function. The complex PCD phenotype results from dysfunction of cilia of the airways and the embryonic node and the structurally related motile sperm flagella. This is associated with underlying ultrastructural defects that frequently involve the outer dynein arm (ODA) complexes that generate cilia and flagella movement. Applying a positional and functional candidate-gene approach, we identified homozygous loss-of-function DNAI2 mutations (IVS11+1G> A) in four individuals from a family with PCD and ODA defects. Further mutational screening of 105 unrelated PCD families detected two distinct homozygous mutations, including a nonsense (c.787C > T) and a splicing mutation (IVS3-3T > G) resulting in out-of-frame transcripts. Analysis of protein expression of the ODA intermediate chain DNAI2 showed sublocalization throughout respiratory cilia. Electron microscopy showed that mutant respiratory cells from these patients lacked DNAI2 protein expression and exhibited ODA defects. High-resolution immunofluorescence imaging demonstrated absence of the ODA heavy chains DNAH5 and DNAH9 from all DNAI2 mutant ciliary axonemes. In addition, we demonstrated complete or distal absence of DNAI2 from ciliary axonemes in respiratory cells of patients with mutations in genes encoding the ODA chains DNAH5 and DNAI1, respectively. Thus, DNAI2 and DNAH5 mutations affect assembly of proximal and distal ODA complexes, whereas DNAI1 mutations mainly disrupt assembly of proximal ODA complexes.