Advancing Primary Ciliary Dyskinesia Diagnosis through High-Speed Video Microscopy Analysis

Abstract

Primary ciliary dyskinesia (PCD) is an inherited disorder that impairs motile cilia, essential for respiratory health, with a reported prevalence of 1 in 16,309 within Hispanic populations. Despite 70% of Puerto Rican patients having the RSPH4A [c.921+3_921+6del (intronic)] founder mutation, the characterization of the ciliary dysfunction remains unidentified due to the unavailability of advanced diagnostic modalities like High-Speed Video Microscopy Analysis (HSVA). Our study implemented HSVA for the first time on the island as a tool to better diagnose and characterize the RSPH4A [c.921+3_921+6del (intronic)] founder mutation in Puerto Rican patients. By applying HSVA, we analyzed the ciliary beat frequency (CBF) and pattern (CBP) in native Puerto Rican patients with PCD. Our results showed decreased CBF and a rotational CBP linked to the RSPH4A founder mutation in Puerto Ricans, presenting a novel diagnostic marker that could be implemented as an axillary test into the PCD diagnosis algorithm in Puerto Rico. The integration of HSVA technology in Puerto Rico substantially enhances the PCD evaluation and diagnosis framework, facilitating prompt detection and early intervention for improved disease management. This initiative, demonstrating the potential of HSVA as an adjunctive test within the PCD diagnostic algorithm, could serve as a blueprint for analogous developments throughout Latin America.

Keywords: Puerto Rico; ciliary dysfunction; diagnostic technology; high-speed video microscopy analysis; primary ciliary dyskinesia.

Figure 1

Ciliary beat frequency (CBF) measurement in patients with PCD with the RSPH4A [c.921+3_921+6del (intronic)] founder mutation (a) and healthy controls (b). This figure illustrates the CBF obtained from a series of samples measured using the manual method. The y-axis represents the CBF in Hertz (Hz), and the x-axis enumerates the individual patients with median measurements across each sample set. (c) Comparison of median CBF (Hz) between patients with PCD with the founder mutation versus healthy controls. Mann–Whitney test between cohorts showed statistical significance (p < 0.001, ****) among Median CBF. The graph’s dotted area and solid line denote the normal CBF range and median typically observed in healthy individuals, as in previous publications [17]. Patient with PCD #12 has a compound heterozygous for the RSPH4A [c.921+3_921+6del (intronic)] plus RSPH4A c.1103T>G (p.Val368Gly).

Figure 2

Representative illustration of nasal ciliary biopsy dynamics and ciliary beat pattern (CBP). Panel (a) presents a top view of the nasal ciliated epithelium from a patient homozygous for the RSPH4A [c.921+3_921+6del (intronic)] founder mutation, illustrating the cilia’s rotational motion. As quantified by tracker software, this pattern is characterized by wide-ranging oscillations at the cilia tips along the x and y axes. Panel (b) displays a top view from a healthy control subject, where cilia demonstrate the expected bidirectional movement, with pronounced and distinct oscillations observable on the x and y axes, also captured and analyzed with tracker software. An accompanying video that provides a dynamic visualization of the RSPH4A [c.921+3_921+6del (intronic)] founder mutation patterns (Video S1) as compared with healthy control (Video S2) is available in the Supplemental Material.

Figure 3

Population-targeted diagnostic algorithm for PCD in Puerto Rico. This algorithm presents a population-targeted diagnostic algorithm for PCD specifically designed for the Puerto Rican demographic, considering the region’s high prevalence of the RSPH4A founder mutation. (*): Threshold values below 77 nL/min are considered positive for PCD at baseline status in two separate visits.