Species Typing of Trypanosomatids Using Amplicon-Based Sequencing

Area of Science:

• Molecular Biology
• Parasitology
• Genomics

Background:

• Amplicon-based sequencing offers high sensitivity and specificity for microorganism identification.
• Accurate identification of parasitic species like Leishmania and Trypanosoma is crucial for diagnostics.

Purpose of the Study:

• To detail amplification and sequencing protocols for HSP70 and 18S rRNA genes.
• To establish a comprehensive workflow for pathogen detection and genetic diversity assessment.

Main Methods:

• Optimization of Polymerase Chain Reaction (PCR) conditions for robust gene amplification.
• Sequencing using Oxford Nanopore Technologies for long-read, real-time data generation.
• Bioinformatics pipeline including quality control and phylogenetic analysis for data processing.

Main Results:

• Successful amplification and sequencing of target genetic markers (HSP70 and 18S rRNA).
• High-resolution taxonomic assignments achieved through long-read sequencing.
• Accurate pathogen detection and detailed assessment of genetic diversity.

Conclusions:

• The described workflow enhances accuracy in identifying Leishmania and Trypanosoma species.
• This approach facilitates in-depth analysis of genetic diversity within trypanosomatids.
• Amplicon sequencing provides a powerful tool for molecular diagnostics and microbial characterization.