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Genetic and Bioinformatic Strategies to Improve Diagnosis in Three Inherited Bleeding Disorders in Bogotá, Colombia
Juliana Lago1, Helena Groot1, Diego Navas1
1Laboratorio de Genética Humana, Universidad de Los Andes, Bogotá 111711, Colombia.
Insights
This study identified novel genetic variations in inherited bleeding disorders like hemophilia A, hemophilia B, and von Willebrand Disease in Colombia. A low-cost, high-resolution melting technique with open-source analysis was validated for improved genetic diagnosis.
Area of Science:
- Medical Genetics
- Molecular Biology
- Congenital Diseases
Background:
- Inherited bleeding disorders (IBDs), including hemophilia A (HA), hemophilia B (HB), and von Willebrand Disease (VWD), are common congenital diseases in Colombia.
- Current diagnostic methods rely on laboratory assays, but limited accessibility can lead to incomplete diagnoses, necessitating genetic confirmation.
Purpose of the Study:
- To characterize the molecular genetic basis of IBDs in Colombian patients.
- To validate the high-resolution melting (HRM) technique as a low-cost genetic diagnostic tool for IBDs.
- To develop an open-source Python code for improved HRM data analysis.
Main Methods:
- Sequencing of the F8, F9, and VWF genes in 11 HA, 3 HB, and 5 VWD patients.
- Validation of the HRM variant-scanning technique using identified disease-causing variations.
- Development and application of an open-source Python code for HRM data analysis, assessing sensitivity and bias.
Main Results:
- Identified 12 variations in HA, 4 in HB, and 19 in VWD patients, with 25 novel variations discovered.
- The HRM technique demonstrated high sensitivity (95%) and unbiased results across different equipment and software.
- The developed Python code enhanced HRM data analysis, enabling analysis of amplicons over 300 bp.
Conclusions:
- Genetic characterization revealed novel variations in Colombian IBD patients.
- The validated HRM technique offers a cost-effective and sensitive method for genetic diagnosis of IBDs, suitable for developing countries.
- The open-source analysis tool improves the accessibility and reliability of HRM-based genetic diagnostics.
Abstract:
Inherited bleeding disorders (IBDs) are the most frequent congenital diseases in the Colombian population; three of them are hemophilia A (HA), hemophilia B (HB), and von Willebrand Disease (VWD). Currently, diagnosis relies on multiple clinical laboratory assays to assign a phenotype. Due to the lack of accessibility to these tests, patients can receive an incomplete diagnosis. In these cases, genetic studies reinforce the clinical diagnosis. The present study characterized the molecular genetic basis of 11 HA, three HB, and five VWD patients by sequencing the F8, F9, or the VWF gene. Twelve variations were found in HA patients, four in HB patients, and 19 in WVD patients. From these variations a total of 25 novel variations were found. Disease-causing variations were used as positive controls for validation of the high-resolution melting (HRM) variant-scanning technique. This approach is a low-cost genetic diagnostic method proposed to be incorporated in developing countries. For the data analysis, we developed an accessible open-source code in Python that improves HRM data analysis with better sensitivity of 95% and without bias when using different HRM equipment and software. Analysis of amplicons with a length greater than 300 bp can be performed by implementing an analysis by denaturation domains.
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