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Primary ciliary dyskinesia (PCD) is a rare genetic disease characterized by dyskinetic cilia. Respiratory symptoms usually start at birth. The lack of diagnostic gold standard tests is challenging, as PCD diagnostics requires different methods with high expertise. We founded PCD-UNIBE as the first comprehensive PCD diagnostic center in Switzerland. Our diagnostic approach includes nasal brushing and cell culture with analysis of ciliary motility via high-speed-videomicroscopy (HSVM) and immunofluorescence labeling (IF) of structural proteins. Selected patients undergo electron microscopy (TEM) of ciliary ultrastructure and genetics. We report here on the first 100 patients assessed by PCD-UNIBE. All patients received HSVM fresh, IF, and cell culture (success rate of 90%). We repeated the HSVM with cell cultures and conducted TEM in 30 patients and genetics in 31 patients. Results from cell cultures were much clearer compared to fresh samples. For 80 patients, we found no evidence of PCD, 17 were diagnosed with PCD, two remained inconclusive, and one case is ongoing. HSVM was diagnostic in 12, IF in 14, TEM in five and genetics in 11 cases. None of the methods was able to diagnose all 17 PCD cases, highlighting that a comprehensive approach is essential for an accurate diagnosis of PCD.

Background Diagnosis of primary ciliary dyskinesia (PCD) is challenging since there is no gold standard test. The European Respiratory (ERS) and American Thoracic (ATS) Societies developed evidence-based diagnostic guidelines with considerable differences. Objective We aimed to compare the algorithms published by the ERS and the ATS with each other and with our own PCD-UNIBE algorithm in a clinical setting. Our algorithm is similar to the ERS algorithm with additional immunofluorescence staining. Agreement (Cohen's κ) and concordance between the three algorithms were assessed in patients with suspicion of PCD referred to our diagnostic centre. Results In 46 out of 54 patients (85%) the final diagnosis was concordant between all three algorithms (30 PCD negative, 16 PCD positive). In eight patients (15%) PCD diagnosis differed between the algorithms. Five patients (9%) were diagnosed as PCD only by the ATS, one (2%) only by the ERS and PCD-UNIBE, one (2%) only by the ATS and PCD-UNIBE, and one (2%) only by the PCD-UNIBE algorithm. Agreement was substantial between the ERS and the ATS (κ=0.72, 95% CI 0.53–0.92) and the ATS and the PCD-UNIBE (κ=0.73, 95% CI 0.53–0.92) and almost perfect between the ERS and the PCD-UNIBE algorithms (κ=0.92, 95% CI 0.80–1.00). Conclusion The different diagnostic algorithms lead to a contradictory diagnosis in a considerable proportion of patients. Thus, an updated, internationally harmonised and standardised PCD diagnostic algorithm is needed to improve diagnostics for these discordant cases., There is no gold standard test for diagnosing PCD. The use of existing diagnostic algorithms leads to contradicting results in many patients (15% in this study). Thus, an updated and internationally harmonised diagnostic guideline is needed. https://bit.ly/2U19Vvq

BackgroundPrimary ciliary dyskinesia (PCD) is a rare genetic disorder resulting in abnormal ciliary motility/structure, extremely heterogeneous at genetic and ultrastructural levels. We aimed, in light of extensive genotyping, to identify specific and quantitative ciliary beating anomalies, according to the ultrastructural phenotype.MethodsWe prospectively included 75 patients with PCD exhibiting the main five ultrastructural phenotypes (n=15/group), screened all corresponding PCD genes and measured quantitative beating parameters by high-speed video-microscopy (HSV).ResultsSixty-eight (91%) patients carried biallelic mutations. Combined outer/inner dynein arms (ODA/IDA) defect induces total ciliary immotility, regardless of the gene involved. ODA defect induces a residual beating with dramatically low ciliary beat frequency (CBF) related to increased recovery stroke and pause durations, especially in case of DNAI1 mutations. IDA defect with microtubular disorganisation induces a low percentage of beating cilia with decreased beating angle and, in case of CCDC39 mutations, a relatively conserved mean CBF with a high maximal CBF. Central complex defect induces nearly normal beating parameters, regardless of the gene involved, and a gyrating motion in a minority of ciliated edges, especially in case of RSPH1 mutations. PCD with normal ultrastructure exhibits heterogeneous HSV values, but mostly an increased CBF with an extremely high maximal CBF.ConclusionQuantitative HSV analysis in PCD objectives beating anomalies associated with specific ciliary ultrastructures and genotypes. It represents a promising approach to guide the molecular analyses towards the best candidate gene(s) to be analysed or to assess the pathogenicity of the numerous sequence variants identified by next-generation-sequencing.

International audience; Primary ciliary dyskinesia (PCD) is an autosomal-recessive disease due to functional or ultra-structural defects of motile cilia. Affected individuals display recurrent respiratory-tract infections; most males are infertile as a result of sperm flagellar dysfunction. The great majority of the PCD-associated genes identified so far encode either components of dynein arms (DAs), which are multiprotein-ATPase complexes essential for ciliary motility, or proteins involved in DA assembly. To identify the molecular basis of a PCD phenotype characterized by central complex (CC) defects but normal DA structure, a phenotype found in ∼15% of cases, we performed whole-exome sequencing in a male individual with PCD and unexplained CC defects. This analysis, combined with whole-genome SNP genotyping, identified a homozygous mutation in DNAJB13 (c.833T>G), a gene encoding a HSP40 co-chaperone whose ortholog in the flagellated alga Chlamydomonas localizes to the radial spokes. In vitro studies showed that this missense substitution (p.Met278Arg), which involves a highly conserved residue of several HSP40 family members, leads to protein instability and triggers proteasomal degradation, a result confirmed by the absence of endogenous DNAJB13 in cilia and sperm from this individual. Subsequent DNAJB13 analyses identified another homozygous mutation in a second family; the study of DNAJB13 transcripts obtained from airway cells showed that this mutation (c.68+1G>C) results in a splicing defect consistent with a loss-of-function mutation. Overall, this study, which establishes mutations in DNAJB13 as a cause of PCD, unveils the key role played by DNAJB13 in the proper formation and function of ciliary and flagellar axonemes in humans.

High speed videomicroscopy (HSV) becomes a key test in primary ciliary dyskinesia (PCD), providing a qualitative evaluation and recently a quantitative analysis of ciliary beating. We aimed to correlate ultrastructural phenotype and ciliary beating Method: 75 PCD patients were prospectively included to reach 15 patients per main ultrastructural phenotypes (combined outer/inner dynein arm defect, outer dynein arm defect, inner dynein arm defect with microtubular disorganization, central complex defect and PCD with normal ultrastructure). Ciliary beating was assessed by the proportion of beating ciliated edges, the ciliary beat frequency (CBF), a qualitative evaluation and a quantitative analysis allowing measuring objective parameters Result: Combined outer/inner dynein arm defect induced a total ciliary immotility, whatever the involved gene; (ii) Outer dynein arm defect induced a minimal residual beating with dramatically low CBF related to increased pause and recovery duration, especially in case of DNAI1 mutations; (iii) Inner dynein arm defect induced a weak proportion of ciliated edges still beating, a decreased beating angle and furthermore a relatively conserved CBF in case of CCDC39 mutations; (iv) Central complex defect induced nearly normal quantitative parameters with a circular gyrating motion in a minority of ciliated edges, whatever the involved gene; (v) PCD with normal ultrastructure induced a wide discrepancy in beating parameters but a frequent increased CBF Conclusion: Quantitative analysis by HSV allows objectively detecting abnormal values of ciliary beating parameters characteristic of each of the main ultrastructural phenotypes reported in PCD.

Background and Objectives Functional and structural lung evaluations are part of the follow-up of patients with primary ciliary dyskinesia (PCD). We aimed to evaluate transversal and longitudinal relationships between lung function test (LFT) and chest computed tomography (CT) in children with PCD, in stable clinical condition. Materials and Methods Data from children followed in the French National Center were retrospectively collected. Inclusion criteria were (i) definitive diagnosis of PCD, (ii) age less than 15 years at the beginning of follow-up, (iii) at least 8 years of follow-up, (iv) at least two couples of concurrent CT and LFT available in a phase of clinical stability of the lung disease without modification of the treatment regimen in the last 4 weeks. Twenty children (median age at entry 4.6 years, median follow-up 15.4 years) were included. Concurrent LFT (blood gas and spirometry) and CT (score) results were recorded. Results LFT indices (PaO2 (n = 210), FVC, FEV1, FEF2575% (n = 195)) significantly decreased with age, and the mean annual decrease (z-score (% predicted)) was −0.17 (−0.49%), −0.09 (−0.50%), −0.10 (−0.89%), and −0.07 (−1.73%), respectively. First CT (median age 8.7 years) revealed bronchiectasis (70%), mucous plugging (70%), peribronchial thickening (90%), parenchymal abnormalities (65%), and hyperinflation (50%). CT scores (n = 74) significantly increased with age, and was negatively correlated to PaO2, FVC, FEV1, and FEF2575% longitudinal changes. Conclusion In stable clinical condition, functional, and structural progressive impairments significantly correlated in children with PCD. Further prospective studies, including large populations of patients with various levels of disease severity, are needed to confirm whether lung function follow-up can be used to adjust CT frequency and help at minimizing the radiation burden in children with a good life expectancy. Pediatr Pulmonol. 2012. 47:816–825. © 2012 Wiley Periodicals, Inc.

International audience; Primary ciliary dyskinesia (PCD) is an inherited disease related to ciliary dysfunction, with heterogeneity in clinical presentation and in ciliary ultrastructural defect. Our study intended to determine if there are phenotypic differences in patients with PCD based on ciliary ultrastructural abnormality. In this retrospective study carried out among 60 children with a definitive diagnosis of PCD, we analyzed clinical, radiological, and functional features at diagnosis and at last recorded visit, according to cilia defect (absence of dynein arms: DAD group, n = 36; abnormalities of the central complex: CCA group, n = 24). Onset of respiratory symptoms occurred later in the CCA than in the DAD group (9.5 versus 0.5 months, p = 0.03). Situs inversus was only observed in the DAD group, while respiratory disease in siblings were more frequent in the CCA group (p = 0.003). At diagnosis, clinical presentation was more severe in the CCA group: frequency of respiratory tract infections (p = 0.008), rhinosinusitis (p = 0.02), otitis complications (p = 0.0001), bilateral bronchiectasis (p = 0.04), and number of hypoxemic patients (p = 0.03). Pulmonary function remained stable in both groups, but outcome was better in the CCA than in the DAD group: less antibiotic therapy and hypoxemic patients (p = 0.004). In conclusion, our results underlined the relationship between the severity of clinical presentation and the ultrastructural ciliary defect.

Background Primary Ciliary Dyskinesia (PCD) is a inherited disorder characterized by abnormal ciliary structure/function, responsible for impaired muco-ciliary transport and infertility. The diagnosis is often delayed for several years though PCD is a rare (~1/20,000) and heterogeneous disease. We elaborated recommendations aiming at improving the quality and efficacy of PCD diagnosis. Methods A multicentric working group, created in 2009 within the French National Centre for Rare Respiratory Diseases, elaborated precise and practical algorithm for PCD diagnosis, based on their experience and the recent literature. It includes adults and paediatric physicians, biologists and technicians involved in PCD, dealing since many years with the difficulties to confirm this diagnosis.

Background Primary Ciliary Dyskinesia (PCD) is a rare inherited disease (~1/20,000), characterized by ciliary structure/function abnormalities, responsible for impaired muco-ciliary transport, leading to recurrent upper and lower airways infections early in life and infertility. The diagnosis is confirmed on ciliate cell samples, collected by nasal and/or bronchial endoscopy. Patients usually need several samples, due to difficulties to get reliable results, especially during respiratory tract infections which are frequent in PCD. We created national guidelines aimed at obtaining the more efficient quality of ciliate cell samples for children and adults.

Background CCDC39 and CCDC40 genes have recently been implicated in primary ciliary dyskinesia (PCD) with inner dynein arm (IDA) defects and axonemal disorganisation; their contribution to the disease is, however, unknown. Aiming to delineate the CCDC39/CCDC40 mutation spectrum and associated phenotypes, this study screened a large cohort of patients with IDA defects, in whom clinical and ciliary phenotypes were accurately described. Methods All CCDC39 and CCDC40 exons and intronic boundaries were sequenced in 43 patients from 40 unrelated families. The study recorded and compared clinical features (sex, origin, consanguinity, laterality defects, ages at first symptoms and at phenotype evaluation, neonatal respiratory distress, airway infections, nasal polyposis, otitis media, bronchiectasis, infertility), ciliary beat frequency, and quantitative ultrastructural analyses of cilia and sperm flagella. Results Biallelic CCDC39 or CCDC40 mutations were identified in 30/34 (88.2%) unrelated families with IDA defects associated with axonemal disorganisation (22 and eight families, respectively). Fourteen of the 28 identified mutations are novel. No mutation was found in the six families with isolated IDA defects. Patients with identified mutations shared a similar phenotype, in terms of both clinical features and ciliary structure and function. The sperm flagellar ultrastructure, analysed in 4/7 infertile males, showed evidence of abnormalities similar to the ciliary ones. Conclusions CCDC39 and CCDC40 mutations represent the major cause of PCD with IDA defects and axonemal disorganisation. Patients carrying CCDC39 or CCDC40 mutations are phenotypically indistinguishable. CCDC39 and CCDC40 analyses in selected patients ensure mutations are found with high probability, even if clinical or ciliary phenotypes cannot prioritise one analysis over the other.

Methods Ciliated cells issued from nasal brushing (controls and primary ciliary dyskinesia patients) were recorded by high-speed video-microscopy (350 frames s). We have performed an original quantitative analysis of ciliary beat dynamics (CBD) by following cilium tips. It allows to describe different parameters including ciliary beat frequency also measured by Fast-Fourier-Transform and Video-Kymography. We have also developed the microbeads tracking method (MBT) to get an index of the global efficiency of ciliary beat. Here, micro-beads (4.5 μm) have been used as markers of the flow generated by beating cilia.

Background Primary ciliary dyskinesia (PCD) is a rare congenital respiratory disorder characterized by abnormal ciliary motility leading to chronic airway infections. Qualitative evaluation of ciliary beat pattern based on digital high-speed videomicroscopy analysis has been proposed in the diagnosis process of PCD. Although this evaluation is easy in typical cases, it becomes difficult when ciliary beating is partially maintained. We postulated that a quantitative analysis of beat pattern would improve PCD diagnosis. We compared quantitative parameters with the qualitative evaluation of ciliary beat pattern in patients in whom the diagnosis of PCD was confirmed or excluded. Methods Nasal nitric oxide measurement, nasal brushings and biopsies were performed prospectively in 34 patients with suspected PCD. In combination with qualitative analysis, 12 quantitative parameters of ciliary beat pattern were determined on high-speed videomicroscopy recordings of beating ciliated edges. The combination of ciliary ultrastructural abnormalities on transmission electron microscopy analysis with low nasal nitric oxide levels was the “gold standard” used to establish the diagnosis of PCD. Results This “gold standard” excluded PCD in 15 patients (non-PCD patients), confirmed PCD in 10 patients (PCD patients) and was inconclusive in 9 patients. Among the 12 parameters, the distance traveled by the cilium tip weighted by the percentage of beating ciliated edges presented 96% sensitivity and 95% specificity. Qualitative evaluation and quantitative analysis were concordant in non-PCD patients. In 9/10 PCD patients, quantitative analysis was concordant with the “gold standard”, while the qualitative evaluation was discordant with the “gold standard” in 3/10 cases. Among the patients with an inconclusive “gold standard”, the use of quantitative parameters supported PCD diagnosis in 4/9 patients (confirmed by the identification of disease-causing mutations in one patient) and PCD exclusion in 2/9 patients. Conclusions When the beat pattern is normal or virtually immotile, the qualitative evaluation is adequate to study ciliary beating in patients suspected for PCD. However, when cilia are still beating but with moderate alterations (more than 40% of patients suspected for PCD), quantitative analysis is required to precise the diagnosis and can be proposed to select patients eligible for TEM.

International audience