scholarly journals Quantitative Assessment of Ciliary Ultrastructure with the Use of Automatic Analysis: PCD Quant

Diagnostics ◽  
2021 ◽  
Vol 11 (8) ◽  
pp. 1363
Author(s):  
Andrea Felšöová ◽  
Tibor Sloboda ◽  
Lukáš Hudec ◽  
Miroslav Koblížek ◽  
Petr Pohunek ◽  
...  
Keyword(s):  
Quantitative Analysis ◽  
Automatic Analysis ◽  
Diagnostic Methods ◽  
Mutual Orientation ◽  
Transmission Electron ◽  
Complex Process ◽  
Golden Standard ◽  
Secondary Defect ◽  
Ciliary Dyskinesia ◽  
Ciliary Ultrastructure

The ciliary ultrastructure can be damaged in various situations. Such changes include primary defects found in primary ciliary dyskinesia (PCD) and secondary defects developing in secondary ciliary dyskinesia (SCD). PCD is a genetic disease resulting from impaired ciliary motility causing chronic disease of the respiratory tract. SCD is an acquired condition that can be caused, for example, by respiratory infection or exposure to tobacco smoke. The diagnosis of these diseases is a complex process with many diagnostic methods, including the evaluation of ciliary ultrastructure using transmission electron microscopy (the golden standard of examination). Our goal was to create a program capable of automatic quantitative analysis of the ciliary ultrastructure, determining the ratio of primary and secondary defects, as well as analysis of the mutual orientation of cilia in the ciliary border. PCD Quant, a program developed for the automatic quantitative analysis of cilia, cannot yet be used as a stand-alone method for evaluation and provides limited assistance in classifying primary and secondary defect classes and evaluating central pair angle deviations. Nevertheless, we see great potential for the future in automatic analysis of the ciliary ultrastructure.

scholarly journals PCD Detect: enhancing ciliary features through image averaging and classification

2020 ◽  
Vol 319 (6) ◽  
pp. L1048-L1060
Author(s):  
Amelia Shoemark ◽  
Andreia L. Pinto ◽  
Mitali P. Patel ◽  
Farheen Daudvohra ◽  
Claire Hogg ◽  
...  
Keyword(s):  
Genetic Diagnosis ◽  
Lung Damage ◽  
Chain Gene ◽  
Heavy Chain Gene ◽  
Transmission Electron ◽  
Improve Accuracy ◽  
Image Averaging ◽  
Biallelic Mutations ◽  
Ciliary Dyskinesia ◽  
Ciliary Ultrastructure

Primary ciliary dyskinesia (PCD) is an inherited disorder of the motile cilia. Early accurate diagnosis is important to help prevent lung damage in childhood and to preserve lung function. Confirmation of a diagnosis traditionally relied on assessment of ciliary ultrastructure by transmission electron microscopy (TEM); however, >50 known PCD genes have made the identification of biallelic mutations a viable alternative to confirm diagnosis. TEM and genotyping lack sensitivity, and research to improve accuracy of both is required. TEM can be challenging when a subtle or partial ciliary defect is present or affected cilia structures are difficult to identify due to poor contrast. Here, we demonstrate software to enhance TEM ciliary images and reduce background by averaging ciliary features. This includes an option to classify features into groups based on their appearance, to generate multiple averages when a nonhomogeneous abnormality is present. We validated this software on images taken from subjects with well-characterized PCD caused by variants in the outer dynein arm (ODA) heavy chain gene DNAH5. Examining more difficult to diagnose cases, we detected 1) regionally restricted absence of the ODAs away from the ciliary base, in a subject carrying mutations in DNAH9; 2) loss of the typically poorly contrasted inner dynein arms; and 3) sporadic absence of part of the central pair complex in subjects carrying mutations in HYDIN, including one case with an unverified genetic diagnosis. We show that this easy-to-use software can assist in detailing relationships between genotype and ultrastructural phenotype, improving diagnosis of PCD.

scholarly journals Ciliary Feature Counter: A program for the Quantitative Assessment of Cilia to Diagnose Primary Ciliary Dyskinesia

Diagnostics ◽  
2020 ◽  
Vol 10 (8) ◽  
pp. 524
Author(s):  
Andreia L. Pinto ◽  
Ranjit K. Rai ◽  
Claire Hogg ◽  
Thomas Burgoyne
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Primary Ciliary Dyskinesia ◽  
Quantitative Assessment ◽  
Consensus Guideline ◽  
International Consensus ◽  
Transmission Electron ◽  
Speed Up ◽  
Ciliary Dyskinesia ◽  
Ciliary Ultrastructure

Primary ciliary dyskinesia (PCD) is a disorder that affects motile cilia in the airway that are required for the removal of mucus, debris, and pathogens. It is important to diagnose PCD in early childhood to preserve lung function. The confirmation of a diagnosis relies on the assessment of ciliary ultrastructure by transmission electron microscopy (TEM). TEM involves the quantitative assessment of the ciliary ultrastructure to identify PCD defects as well as abnormalities resulting from infection. Many specialist diagnostic centres still rely on physical counters to tally results and paper notes to summarise findings before transferring the results to computer databases/records. To speed up the diagnostic data collection and increase the protection of patient information, we have developed digital ciliary feature counters that conform to the PCD reporting international consensus guideline. These counters can be used on a computer or tablet, and automatically generate notes regarding sample observations. We show that the digital counters are easy to use and can generate TEM diagnostic reports that will be useful for many PCD diagnostic centres.

Quantitative analysis of ciliary ultrastructure in patients with primary ciliary dyskinesia

2008 ◽  
Vol 110 (1) ◽  
pp. 34-41 ◽  
Author(s):  
Serap Sirvanci ◽  
Z. Seda Uyan ◽  
Feriha Ercan ◽  
Bulent Karadag ◽  
Refika Ersu ◽  
...  
Keyword(s):  
Quantitative Analysis ◽  
Primary Ciliary Dyskinesia ◽  
Ciliary Dyskinesia ◽  
Ciliary Ultrastructure

Ultrastructural Abnormalities of Respiratory Cilia: A 25-Year Experience

2008 ◽  
Vol 132 (11) ◽  
pp. 1786-1791
Author(s):  
Thomas P. Plesec ◽  
Angela Ruiz ◽  
James T. McMahon ◽  
Richard A. Prayson
Keyword(s):  
Basal Body ◽  
Respiratory Infections ◽  
Clinical History ◽  
Radial Spoke ◽  
Transmission Electron ◽  
Respiratory Cilia ◽  
Tract Infections ◽  
Multiple Abnormalities ◽  
Ciliary Dyskinesia ◽  
Ciliary Ultrastructure

Abstract Context.—Ciliary dyskinesia is a rare, but significant, cause of chronic respiratory infections, and transmission electron microscopy is a critical adjunct to making the diagnosis. Objective.—To investigate a single institution's experience with patients demonstrating abnormal ciliary ultrastructure. Design.—Retrospective clinicopathologic review of 278 bronchial or nasal turbinate brushings or biopsies from 1983 through 2007. Results.—There were 12 women and 9 men (mean age, 19.6 years; range, 1–54 years) with abnormal ciliary ultrastructure. Clinical history was unavailable in 3 patients, 15 (83%) of 18 patients presented with chronic or recurrent upper respiratory infections, and 3 (17%) presented with infertility. Seven (39%) of 18 patients had findings of Kartagener syndrome with situs inversus, dextrocardia, and bronchiectasis. Truncation or absence of inner or outer dynein arms occurred in 15 (71%) of 21 cases, and 5 (24%) revealed transposition defects with displacement of the central microtubules and peripheral doublets in 9 + 0 and 8 + 1 patterns. Radial spoke defects with microtubular disarray occurred in 4 (19%) of 21 cases. Compound cilia with multiple axonemes within a single outer sheath and supernumerary microtubules each occurred in 2 (10%) of the cases. Random ciliary orientation was also found in 2 (10%) of the cases, and dense granular basal body inclusions occurred in 1 case (5%). Multiple abnormalities occurred in 6 (29%) of the 21 cases. Conclusions.—Most patients presented with chronic respiratory tract infections or infertility. Dynein arm defects, transposition defects, and radial spoke defects were the most commonly encountered abnormal findings. Less-frequent abnormal findings included compound cilia, supernumerary microtubules, and dense granular basal body inclusions.

scholarly journals New Diagnostic Methods and Treatment Recommendations in Primary Ciliary Dyskinesia

2020 ◽  
pp. 42-49
Author(s):  
Nagihan Emiralioğlu
Keyword(s):  
Diagnostic Tests ◽  
Primary Ciliary Dyskinesia ◽  
High Speed ◽  
Electron Tomography ◽  
Diagnostic Methods ◽  
European Respiratory Society ◽  
Transmission Electron ◽  
Nutritional Advice ◽  
Organ Laterality ◽  
Ciliary Dyskinesia

Primary ciliary dyskinesia (PCD) is a rare and genetically heterogeneous disease and clinically characterized by neonatal respiratory distress, organ laterality defects, persistent rhinosinusitis, chronic bronchitis, and eventually bronchiectasis. Currently, there is no single “gold standard” diagnostic test for PCD. PICADAR (Primary Ciliary Dyskinesia Rule) score is a guide to decide for further evaluation of diagnostic tests in PCD. European Respiratory Society (ERS) and American Thoracic Society (ATS) recommend diagnostic tests, including nasal nitric oxide (nNO), high-speed video analysis (HSVMA), transmission electron microscopy (TEM) and genetic testing. Cryo-electron tomography and immunofluorescence methods are new techniques recently performed by specialized centers and needs to be improved. Age at diagnosis for PCD changes according to awareness of disease and available diagnostic tests in different centers. Regular follow-up and multidisciplinary approach is important in the management of PCD. The main aim of the treatment is to prevent pulmonary exacerbations and slow the progression of the disease since there are no treatment approaches to correct the underlying cilia structure and its functions in PCD. Although, there are not enough randomized controlled trials for the treatment of PCD, recent treatments are usually based on to improve the mucociliary clearance. Early diagnosis with multidisciplinary management and nutritional advice could improve growth and delay disease progression leading to bronchiectasis and lung function impairment in PCD

scholarly journals Clinical and Genetic Analysis of Children with Kartagener Syndrome

Cells ◽  
2019 ◽  
Vol 8 (8) ◽  
pp. 900 ◽  
Author(s):  
Rute Pereira ◽  
Telma Barbosa ◽  
Luís Gales ◽  
Elsa Oliveira ◽  
Rosário Santos ◽  
...  
Keyword(s):  
Autosomal Recessive Disorder ◽  
Kartagener Syndrome ◽  
Mucus Clearance ◽  
Pathogenic Variants ◽  
Transmission Electron ◽  
Whole Exome ◽  
Dynein Arms ◽  
Organ Laterality ◽  
Ciliary Dyskinesia ◽  
Laterality Defects

Primary ciliary dyskinesia (PCD) is a rare autosomal recessive disorder characterized by dysfunction of motile cilia causing ineffective mucus clearance and organ laterality defects. In this study, two unrelated Portuguese children with strong PCD suspicion underwent extensive clinical and genetic assessments by whole-exome sequencing (WES), as well as ultrastructural analysis of cilia by transmission electron microscopy (TEM) to identify their genetic etiology. These analyses confirmed the diagnostic of Kartagener syndrome (KS) (PCD with situs inversus). Patient-1 showed a predominance of the absence of the inner dynein arms with two disease-causing variants in the CCDC40 gene. Patient-2 showed the absence of both dynein arms and WES disclosed two novel high impact variants in the DNAH5 gene and two missense variants in the DNAH7 gene, all possibly deleterious. Moreover, in Patient-2, functional data revealed a reduction of gene expression and protein mislocalization in both genes’ products. Our work calls the researcher’s attention to the complexity of the PCD and to the possibility of gene interactions modelling the PCD phenotype. Further, it is demonstrated that even for well-known PCD genes, novel pathogenic variants could have importance for a PCD/KS diagnosis, reinforcing the difficulty of providing genetic counselling and prenatal diagnosis to families.

scholarly journals A recent update of the diagnostic methods for tuberculosis and their applicability in Indonesia: a narrative review

2019 ◽  
Vol 28 (3) ◽  
pp. 284-91
Author(s):  
Tri Nugraha Susilawati ◽  
Riska Larasati
Keyword(s):  
Delayed Diagnosis ◽  
Diagnostic Methods ◽  
World Health ◽  
Current State ◽  
Interferon Gamma Release Assays ◽  
High Prevalence ◽  
The World ◽  
Golden Standard ◽  
Tertiary Hospitals ◽  
Health Organization

Tuberculosis (TB) is a major global problem, especially with the high prevalence of HIV-TB co-infection. Delayed diagnosis and continual transmission contribute to high mortality in Indonesia, which has the third highest incidence of TB in the world, after China and India. Therefore, early diagnosis is needed to reduce the number of cases and to administer therapy to prevent the transmission of bacteria. The diagnosis of TB remains a challenge in clinical practice due to poor sensitivity and the requirement of skilled staff in microscopic tests, the slow growth Mycobacterium in culture, and the low number of bacilli present in extrapulmonary TB. Despite being the golden standard for TB diagnosis, cultures require 2–8 weeks to grow. Other methods for diagnosing TB include interferon-gamma release assays and serologic tests such as the tuberculin skin test. Recently, the World Health Organization recommended the GeneXpert MTB/RIF assay for diagnosing TB. This review presents the current state of TB epidemiology and various methods for TB diagnosis. In particular, the paper provides an in-depth discussion about the GeneXpert MTB/RIF assay that has been made available recently in selected tertiary hospitals in Indonesia.

scholarly journals Primary ciliary dyskinesia with normal ultrastructure: three-dimensional tomography detects absence of DNAH11

2018 ◽  
Vol 51 (2) ◽  
pp. 1701809 ◽  
Author(s):  
Amelia Shoemark ◽  
Thomas Burgoyne ◽  
Robert Kwan ◽  
Mellisa Dixon ◽  
Mitali P. Patel ◽  
...  
Keyword(s):  
Primary Ciliary Dyskinesia ◽  
Electron Tomography ◽  
Three Dimensional ◽  
Structural Abnormality ◽  
Healthy Controls ◽  
Ultrastructural Studies ◽  
Transmission Electron ◽  
Causal Variants ◽  
Diagnostic Samples ◽  
Ciliary Dyskinesia

In primary ciliary dyskinesia (PCD), motile ciliary dysfunction arises from ciliary defects usually confirmed by transmission electron microscopy (TEM). In 30% of patients, such as those with DNAH11 mutations, apparently normal ultrastructure makes diagnosis difficult. Genetic analysis supports diagnosis, but may not identify definitive causal variants. Electron tomography, an extension of TEM, produces three-dimensional ultrastructural ciliary models with superior resolution to TEM. Our hypothesis is that tomography using existing patient samples will enable visualisation of DNAH11-associated ultrastructural defects. Dual axis tomograms from araldite-embedded nasal cilia were collected in 13 PCD patients with normal ultrastructure (DNAH11 n=7, HYDIN n=2, CCDC65 n=3 and DRC1 n=1) and six healthy controls, then analysed using IMOD and Chimera software.DNAH11 protein is localised to the proximal ciliary region. Within this region, electron tomography indicated a deficiency of >25% of proximal outer dynein arm volume in all patients with DNAH11 mutations (n=7) compared to other patients with PCD and normal ultrastructure (n=6) and healthy controls (n=6). DNAH11 mutations cause a shared abnormality in ciliary ultrastructure previously undetectable by TEM. Advantageously, electron tomography can be used on existing diagnostic samples and establishes a structural abnormality where ultrastructural studies were previously normal.

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