TY - JOUR
T1 - Enhancing Trypanosomatid Identification and Genotyping with Oxford Nanopore Sequencing
T2 - Development and Validation of an 18S rRNA Amplicon-Based Method
AU - Cruz-Saavedra, Lissa
AU - Ospina, Carlos
AU - Patiño, Luz H.
AU - Villar, Juan C.
AU - Sáenz Pérez, Luis D.
AU - Cantillo-Barraza, Omar
AU - Jaimes-Dueñez, Jeiczon
AU - Ballesteros, Nathalia
AU - Cáceres, Tatiana
AU - Vallejo, Gustavo
AU - Ramírez, Juan D.
N1 - Publisher Copyright:
© 2024 Association for Molecular Pathology and American Society for Investigative Pathology
PY - 2024
Y1 - 2024
N2 - Trypanosomatids, including Trypanosoma and Leishmania species, present significant medical and veterinary challenges, causing substantial economic losses, health complications, and even fatalities. Diagnosing and genotyping these species and their genotypes is often complex, involving multiple steps. This study aimed to develop an amplicon-based sequencing (ABS) method using Oxford Nanopore long-read sequencing to enhance Trypanosomatid detection and genotyping. The 18S rDNA gene was targeted for its inter-species conservation. The Trypanosomatid-ABS method effectively distinguished between 11 Trypanosoma species (including Trypanosoma evansi, Trypanosoma theileri, Trypanosoma vivax, and Trypanosoma rangeli) and 6 Trypanosoma cruzi discrete typing units (TcI to TcVI and TcBat), showing strong concordance with conventional methods (κ index of 0.729, P < 0.001). It detected co-infections between Trypanosomatid genera and T. cruzi, with a limit of detection of one parasite per mL. The method was successfully applied to human, animal, and triatomine samples. Notably, TcI predominated in chronic Chagas samples, whereas TcII and TcIV were found in the acute stage. Triatomine vectors exhibited diverse Trypanosomatid infections, with Triatoma dimidiata mainly infected with TcI and occasional TcBat co-infections, and Rhodnius prolixus showing TcI and TcII infections, along with T. rangeli co-infections and mixed TcII infections. Animals were infected with T. vivax, T. theileri, and T. evansi. The ABS method's high resolution, sensitivity, and accuracy make it a valuable tool for understanding Trypanosomatid dynamics, enhancing disease control strategies, and enabling targeted interventions.
AB - Trypanosomatids, including Trypanosoma and Leishmania species, present significant medical and veterinary challenges, causing substantial economic losses, health complications, and even fatalities. Diagnosing and genotyping these species and their genotypes is often complex, involving multiple steps. This study aimed to develop an amplicon-based sequencing (ABS) method using Oxford Nanopore long-read sequencing to enhance Trypanosomatid detection and genotyping. The 18S rDNA gene was targeted for its inter-species conservation. The Trypanosomatid-ABS method effectively distinguished between 11 Trypanosoma species (including Trypanosoma evansi, Trypanosoma theileri, Trypanosoma vivax, and Trypanosoma rangeli) and 6 Trypanosoma cruzi discrete typing units (TcI to TcVI and TcBat), showing strong concordance with conventional methods (κ index of 0.729, P < 0.001). It detected co-infections between Trypanosomatid genera and T. cruzi, with a limit of detection of one parasite per mL. The method was successfully applied to human, animal, and triatomine samples. Notably, TcI predominated in chronic Chagas samples, whereas TcII and TcIV were found in the acute stage. Triatomine vectors exhibited diverse Trypanosomatid infections, with Triatoma dimidiata mainly infected with TcI and occasional TcBat co-infections, and Rhodnius prolixus showing TcI and TcII infections, along with T. rangeli co-infections and mixed TcII infections. Animals were infected with T. vivax, T. theileri, and T. evansi. The ABS method's high resolution, sensitivity, and accuracy make it a valuable tool for understanding Trypanosomatid dynamics, enhancing disease control strategies, and enabling targeted interventions.
UR - http://www.scopus.com/inward/record.url?scp=85189094874&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=85189094874&partnerID=8YFLogxK
U2 - 10.1016/j.jmoldx.2024.01.012
DO - 10.1016/j.jmoldx.2024.01.012
M3 - Research Article
C2 - 38360211
AN - SCOPUS:85189094874
SN - 1525-1578
JO - Journal of Molecular Diagnostics
JF - Journal of Molecular Diagnostics
ER -