Usefulness of a multiplex PCR for the rapid identification of Candida glabrata species complex in Mexican clinical isolates
ABSTRACT Candida glabrata complex includes three species identified through molecular biology methods: C. glabrata sensu stricto , C. nivariensis and C. bracarensis . In Mexico, the phenotypic methods are still used in the diagnosis; therefore, the presence of C. nivariensis and C. bracarensis among...
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doaj-25506bd810344367b584b3319cb6af9d2020-11-25T02:04:08ZengUniversidade de São PauloRevista do Instituto de Medicina Tropical de São Paulo1678-99466110.1590/s1678-9946201961037S0036-46652019005000219Usefulness of a multiplex PCR for the rapid identification of Candida glabrata species complex in Mexican clinical isolatesMaría del Rocío Reyes-MontesGustavo Acosta-AltamiranoEsperanza Duarte-EscalanteEduardo García SalazarErick Martínez-HerreraRoberto ArenasGloria GonzálezMaría Guadalupe Frías-De-LeónABSTRACT Candida glabrata complex includes three species identified through molecular biology methods: C. glabrata sensu stricto , C. nivariensis and C. bracarensis . In Mexico, the phenotypic methods are still used in the diagnosis; therefore, the presence of C. nivariensis and C. bracarensis among clinical isolates is still unknown. The aim of this study was to evaluate the utility of a multiplex PCR for the identification of the C. glabrata species complex. DNA samples from 92 clinical isolates that were previously identified through phenotypic characteristics as C. glabrata were amplified by four oligonucleotides (UNI-5.8S, GLA-f, BRA-f, and NIV-f) that generate amplicons of 397, 293 and 223-bp corresponding to C. glabrata sensu stricto , C. nivariensis , and C. bracarensis , respectively. The amplicon sequences were used to perform a phylogenetic analysis through the Maximum Likelihood method (MEGA6), including strains and reference sequences of species belonging to C. glabrata complex. In addition, recombination and linkage disequilibrium were estimated (DnaSP version 5.0) for C. glabrata sensu stricto isolate s . Eighty-eight isolates generated a 397-bp fragment and only in one isolate a 223-bp amplicon was observed. In the phylogenetic tree, the sequences of 397-bp were grouped with C. glabrata reference sequences , and the sequence of 223-bp was grouped with C. bracarensis reference sequences, corroborating the PCR identification. The number of recombination events for the isolates of C. glabrata sensu stricto was zero, suggesting a clonal population structure. Three isolates that did not amplify any of the expected fragments were identified as Saccharomyces cerevisiae through the sequencing of the D1/D2 domain region within the 28S rDNA gene. The multiplex PCR is a fast, cost-effective and reliable tool that can be used in clinical laboratories to identify C. glabrata complex species.http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0036-46652019005000219&lng=en&tlng=enCandida glabrata complexCandida bracarensisCandida nivariensisMultiplex PCRCandidiasisMexico |
collection |
DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
María del Rocío Reyes-Montes Gustavo Acosta-Altamirano Esperanza Duarte-Escalante Eduardo García Salazar Erick Martínez-Herrera Roberto Arenas Gloria González María Guadalupe Frías-De-León |
spellingShingle |
María del Rocío Reyes-Montes Gustavo Acosta-Altamirano Esperanza Duarte-Escalante Eduardo García Salazar Erick Martínez-Herrera Roberto Arenas Gloria González María Guadalupe Frías-De-León Usefulness of a multiplex PCR for the rapid identification of Candida glabrata species complex in Mexican clinical isolates Revista do Instituto de Medicina Tropical de São Paulo Candida glabrata complex Candida bracarensis Candida nivariensis Multiplex PCR Candidiasis Mexico |
author_facet |
María del Rocío Reyes-Montes Gustavo Acosta-Altamirano Esperanza Duarte-Escalante Eduardo García Salazar Erick Martínez-Herrera Roberto Arenas Gloria González María Guadalupe Frías-De-León |
author_sort |
María del Rocío Reyes-Montes |
title |
Usefulness of a multiplex PCR for the rapid identification of Candida glabrata species complex in Mexican clinical isolates |
title_short |
Usefulness of a multiplex PCR for the rapid identification of Candida glabrata species complex in Mexican clinical isolates |
title_full |
Usefulness of a multiplex PCR for the rapid identification of Candida glabrata species complex in Mexican clinical isolates |
title_fullStr |
Usefulness of a multiplex PCR for the rapid identification of Candida glabrata species complex in Mexican clinical isolates |
title_full_unstemmed |
Usefulness of a multiplex PCR for the rapid identification of Candida glabrata species complex in Mexican clinical isolates |
title_sort |
usefulness of a multiplex pcr for the rapid identification of candida glabrata species complex in mexican clinical isolates |
publisher |
Universidade de São Paulo |
series |
Revista do Instituto de Medicina Tropical de São Paulo |
issn |
1678-9946 |
description |
ABSTRACT Candida glabrata complex includes three species identified through molecular biology methods: C. glabrata sensu stricto , C. nivariensis and C. bracarensis . In Mexico, the phenotypic methods are still used in the diagnosis; therefore, the presence of C. nivariensis and C. bracarensis among clinical isolates is still unknown. The aim of this study was to evaluate the utility of a multiplex PCR for the identification of the C. glabrata species complex. DNA samples from 92 clinical isolates that were previously identified through phenotypic characteristics as C. glabrata were amplified by four oligonucleotides (UNI-5.8S, GLA-f, BRA-f, and NIV-f) that generate amplicons of 397, 293 and 223-bp corresponding to C. glabrata sensu stricto , C. nivariensis , and C. bracarensis , respectively. The amplicon sequences were used to perform a phylogenetic analysis through the Maximum Likelihood method (MEGA6), including strains and reference sequences of species belonging to C. glabrata complex. In addition, recombination and linkage disequilibrium were estimated (DnaSP version 5.0) for C. glabrata sensu stricto isolate s . Eighty-eight isolates generated a 397-bp fragment and only in one isolate a 223-bp amplicon was observed. In the phylogenetic tree, the sequences of 397-bp were grouped with C. glabrata reference sequences , and the sequence of 223-bp was grouped with C. bracarensis reference sequences, corroborating the PCR identification. The number of recombination events for the isolates of C. glabrata sensu stricto was zero, suggesting a clonal population structure. Three isolates that did not amplify any of the expected fragments were identified as Saccharomyces cerevisiae through the sequencing of the D1/D2 domain region within the 28S rDNA gene. The multiplex PCR is a fast, cost-effective and reliable tool that can be used in clinical laboratories to identify C. glabrata complex species. |
topic |
Candida glabrata complex Candida bracarensis Candida nivariensis Multiplex PCR Candidiasis Mexico |
url |
http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0036-46652019005000219&lng=en&tlng=en |
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