Liliana Jazmín Cortés1, Sofía Duque1, Miryam Consuelo López2, Diego Moncada3, Diego Molina1, Jorge Enrique Gómez-Marín3, María Luz Gunturiz4. 1. Grupo de Parasitología, Subdirección de Investigación Científica y Tecnológica, Instituto Nacional de Salud. 2. Departamento de Parasitología, Facultad de Medicina, Universidad Nacional de Colombia, Bogotá, D.C, Colombia. 3. Grupo de Parasitología Molecular (GEPAMOL), Centro de Investigaciones Biomédicas, Universidad del Quindío, Armenia, Colombia. 4. Grupo de Fisiología Molecular, Subdirección de Investigación Científica y Tecnológica, Instituto Nacional de Salud, Bogotá, D.C, Colombia.
Abstract
INTRODUCTION: There are no reports describing polymorphisms in target genes of anti- Toxoplasma drugs in South American isolates. OBJECTIVE: This study sought to perform cloning and sequencing of the dihydrofolate reductase ( dhfr ) and dihydropteroate-synthase ( dhps ) genes of the reference Rh strain and two Colombian isolates of Toxoplasma gondii . MATERIALS AND METHODS: Two isolates were obtained from the cerebrospinal fluid of HIV-infected patients with cerebral toxoplasmosis. A DNA extraction technique and PCR assay for the dhfr and dhps genes were standardized, and the products of amplification were cloned into Escherichia coli and sequenced. RESULTS: One polymorphism (A « G) was found at position 235 of exon 2 in the dhps gene. In addition, two polymorphisms (G « C) at positions 259 and 260 and one polymorphism (T « G) at position 371 within exon 4 of the dhps gene were detected. In this last exon, a bioinformatic analysis revealed a non-synonymous polymorphism in the coding region that could lead to the substitution of Glu (CAA or CAG) for His (encoded by codons AAU or AAC). A structural model of the T. gondii DHPS protein was calculated, and the results revealed modifications in secondary structure due to mutations. CONCLUSIONS: The methods described in this study can be used as a tool to search for polymorphisms in samples from patients with different clinical manifestations of toxoplasmosis and to examine their relationship with the therapeutic response.
INTRODUCTION: There are no reports describing polymorphisms in target genes of anti- Toxoplasma drugs in South American isolates. OBJECTIVE: This study sought to perform cloning and sequencing of the dihydrofolate reductase ( dhfr ) and dihydropteroate-synthase ( dhps ) genes of the reference Rh strain and two Colombian isolates of Toxoplasma gondii . MATERIALS AND METHODS: Two isolates were obtained from the cerebrospinal fluid of HIV-infectedpatients with cerebral toxoplasmosis. A DNA extraction technique and PCR assay for the dhfr and dhps genes were standardized, and the products of amplification were cloned into Escherichia coli and sequenced. RESULTS: One polymorphism (A « G) was found at position 235 of exon 2 in the dhps gene. In addition, two polymorphisms (G « C) at positions 259 and 260 and one polymorphism (T « G) at position 371 within exon 4 of the dhps gene were detected. In this last exon, a bioinformatic analysis revealed a non-synonymous polymorphism in the coding region that could lead to the substitution of Glu (CAA or CAG) for His (encoded by codons AAU or AAC). A structural model of the T. gondiiDHPS protein was calculated, and the results revealed modifications in secondary structure due to mutations. CONCLUSIONS: The methods described in this study can be used as a tool to search for polymorphisms in samples from patients with different clinical manifestations of toxoplasmosis and to examine their relationship with the therapeutic response.
Authors: Gabriella de Lima Bessa; Ricardo Wagner de Almeida Vitor; Erica Dos Santos Martins-Duarte Journal: Parasitol Res Date: 2021-08-14 Impact factor: 2.289
Authors: Jorge Acosta-Reyes; Edgar Navarro; Maria José Herrera; Eloina Goenaga; Martha L Ospina; Edgar Parra; Marcela Mercado; Pablo Chaparro; Mauricio Beltran; Maria Luz Gunturiz; Lissethe Pardo; Catalina Valencia; Sandra Huertas; Jorge Rodríguez; Germán Ruiz; Diana Valencia; Lisa B Haddad; Sarah C Tinker; Cynthia A Moore; Hernando Baquero Journal: Emerg Infect Dis Date: 2017-06-15 Impact factor: 6.883