Introduction
Primary ciliary dyskinesia (PCD) refers to a group of related hereditary motile ciliopathies. Cases of PCD are rare, affecting just 1 per 10,000—20,000 births [
1]. In childhood, abnormal motile cilia activity can result in progressive respiratory diseases that present in the form of neonatal respiratory distress (NRD), a persistent chronic wet cough often present from birth, and chronic rhinitis/sinusitis. The primary consequences of chronic airway infection and inflammation include bronchiectasis and the impairment of lung function [
2]. Prior to the onset of these serious events, patients generally exhibit nonspecific respiratory manifestations that overlap with other conditions such as recurrent respiratory infections, asthma, aspiration, and immunodeficiencies. If the genetic mutations underlying PCD also result in embryonic nodal cilia dysfunction, affected individuals also exhibit a spectrum of organ laterality defects, including situs inversus totalis and situs ambiguus (SA), in which organ arrangement falls somewhere between a normal and mirrored arrangement [
3,
4].
Confirmation of a PCD diagnosis remains challenging, as no single diagnostic test has been shown to offer 100% sensitivity and specificity. Historically, the diagnosis of PCD has been based on the presence of ultrastructural defects in the ciliary axoneme detected via transmission electron microscopy (TEM), but this approach is subject to significant limitations given that ~ 30% of PCD patients have normal ciliary ultrastructural characteristics [
5,
6], and nonspecific ciliary changes, which can be induced by infection, may appear similar in presentation to PCD under TEM [
7]. Other tests, including nasal nitric oxide (nNO) measurements, high-speed video analysis (HVSA) with ciliary beat pattern and beat frequency analyses, immunofluorescent staining of ciliary proteins, and genetic testing have emerged as alternative approaches to the diagnosis of PCD.
PCD is characterized by substantial genetic heterogeneity, with mutations present predominantly in autosomal recessive genes and less often in X-linked genes coding for axonemal, cytoplasmic, and regulatory proteins that have been implicated in this disease [
6,
8]. PCD patients are thought to exhibit striking genetic stratification based on their population of origin [
9‐
13]. In patients with mutations in specific genes associated with the pathogenesis of PCD, clinical symptoms and disease severity can vary significantly [
14‐
16]. China is a vast country, and two pediatric PCD case series from northern and eastern China have attested to the marked genetic heterogeneity and diverse distributions among PCD patients in this country [
10,
11]. The present survey of the clinical data and distributions of disease-causing mutations in a study population from Western China may thus offer valuable context for efforts to more comprehensively understand the characteristics of PCD in China.
Discussion
In the present study, we analyzed a small pediatric PCD patient cohort in western China (Sichuan and Guizhou Provinces or Chongqing). The vast majority of patients were of Han nationality, while only two were minorities. We focused on the clinical manifestations, ciliary phenotypes, and genetic characterization of these subjects. Furthermore, we integrated the genetic results from these individuals with those from other pediatric PCD cohorts in China and gained relatively comprehensive insights into the features of the PCD genetic spectrum in China.
Clinical features of PCD including chronic wet cough, recurrent sinusitis, otitis media, neonatal respiratory distress syndrome, laterality defects, and bronchiectasis [
3] were observed in this study. All of these patients had a chronic wet cough, and this high rate may stem from the bias of diseases treated in this specialty department. In our cohort, only 5% of patients presented with laterality defects, a much lower frequency than in other reports [
11,
12]. A low frequency of laterality defects increases the challenges associated with the early diagnosis of PCD [
21]. Laterality defects exhibit an overlapping genetic etiology with PCD and thus represent a relatively specific marker for this condition [
22,
23]. However, when comparing the incidence of situs inversus (SI) (1:6000–1:8000) [
24], and PCD, prior studies have concluded that most individuals affected by SI do not suffer from PCD. To date, there are 21 known PCD causative genes not associated with laterality defects [
6,
25,
26]. Four individuals in the present study exhibited mutations in genes not associated with laterality defects (
HYDIN, RSPH9, CCNO, RSPH4A). Another possible reason for the extremely low rate of laterality defects was that patients with severe organ malformation, particularly congenital heart disease, are usually referred to other specialists. Recurrent wheezing has not previously been reported as a diagnostic feature of PCD [
18,
27], yet was common in the present PCD patient cohort (7/16). Prior studies have also observed high rates of recurrent wheezing or asthma in PCD patients [
10]. Interestingly, ICS therapy is generally insufficient to relieve asthma symptoms in PCD patients, as shown in one study comparing airway inflammation between patients with asthma and PCD [
28]. Wheezing in PCD patients likely arises due to recurrent airway infection or inflammation, although further research on this topic is warranted. Children with PCD often develop persistent respiratory symptoms that start in the first year of life [
8,
27]. In previous reports, the median age of initial respiratory symptoms was 1 to 3 months [
10,
13]. However, in our cohort, only 44.7% of patients reported that their initial symptoms had been present since infancy age. This may be attributable to recall bias and differences in respiratory disease severity linked to specific genes. Bronchiectasis represents a severe pulmonary sequela that can affect PCD patients, but the timing of bronchiectasis development is unclear. Prior studies have detected bronchiectasis in infants with PCD [
29], while we observed it in children as young as 2 years old upon chest CT evaluation. Other reports have suggested that 70% of children with PCD present with bronchiectasis at a median age of 8 years [
30]. For pediatricians, patients with chronic wet cough or recurrent wheezing should be treated cautiously, even when they do not exhibit laterality defects or bronchiectasis. Awareness of PCD among medical practitioners and taking past history into account can help avoid a delayed diagnosis [
31].
Historically, TEM was a traditional test for the diagnosis of PCD, with a reported overall 86% success rate in acquiring specimens adequate for formal interpretation from nasal scrape biopsies in children [
32]. In this study, this proportion was 92.3% (24/26), suggesting that mucosal biopsy via bronchoscopy was likely to be more efficient due to associated visualization and relatively integrated tissue. Three patients with definitive PCD exhibited normal ciliary ultrastructural characteristics together with mutations in
HYDIN or
DNH11, both of which are known to result in PCD with normal ciliary ultrastructural morphology [
5,
6]. Notably, one patient found to be aciliary under TEM was ultimately determined to harbor compound heterogeneous mutations in
CCNO, a gene related to the generation of multiple cilia [
6]. As such, in the absence of genetic testing, it was sometimes hard to determine whether ultrastructural defects were primarily due to genetic mutations or were secondary to infection and/or inflammation. Moreover, improper specimen handling/processing or inexperience with the interpretation of electron microscopy results can also impact the diagnostic utility of TEM [
3]. Therefore, TEM alone is not well suited to PCD diagnosis [
33].
To date, more than 50 genes have been associated with PCD, and over 70% of tested patients exhibit biallelic mutations in one of these genes [
8]. There is a striking genetic stratification among PCD patients with respect to their population of origin [
9,
13]. Mutations in the
DNAH5 and
DNAH11 genes are thought to be the most common cause of PCD in Europeans, whereas mutations in
LRRC6 and
CCDC103 are more common in South Asian populations [
9]. Genetic sequencing interpretation is not straightforward in PCD patients owing to high levels of genetic heterogeneity and allele rarity, with very little past sequencing data being available to date for Chinese patient populations that can provide important information used to exclude non-pathogenic variants. Here, we have provided clinical and genetic data for a small number of pediatric PCD patients in a single center in western China. By integrating these results with other data, we identified
DNAH11 and
DNAH5 mutations as the dominant PCD-related pathogenic variants among affected children in China. The diagnosis of patients with
DNAH11 mutations remains challenging, as mutations in this gene exhibit considerable variability in the resultant clinical phenotype [
34]. In addition, PCD cases associated with
DNAH11 mutations do not exhibited any specific ultrastructural defects and thus cannot be detected via TEM [
5,
35]. Therefore, the high prevalence of mutations in
DNAH11 in Chinese PCD patients may limit the utility of TEM as a first-line diagnostic approach. Moreover, locus heterogeneity and the high frequency of VUS present a substantial challenge to the translation of genomic variation to clinical practice [
36], and new methods must be developed to assist in diagnosis [
37,
38].
This study had a number of limitations, including a small clinical sample size and the potential for recall bias in family members when evaluating past medical history. ERS guidelines recommend nNO and HSVA testing for the initial diagnostic work-up of patients suspected of having PCD [
18]. Owing to a lack of testing facilities, however, we were unable to perform any HSVA testing and nNO measurements were not taken for the majority of patients despite the importance of this screening test. In addition, some parents refused to undergo bronchial biopsy for TEM evaluation, and future studies may thus benefit from including more comprehensive diagnostic and follow-up strategies for PCD patients.
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