Evaluation of genomic DNA extraction methods for the identification of Leptospira spp. in bovine urine samples by PCR

Authors

  • Alexandra Paola Revelo Ruales Facultad de Medicina y Zootecnia, Universidad Central del Ecuador, Quito, Ecuador
  • Euclides De La Torre M. Agencia de Regulación y Control Fito y Zoosanitario AGROCALIDAD, Tumbaco, Ecuador
  • Galo Martínez C. Carrera de Medicina Veterinaria y Zootecnia, Facultad de Medicina Veterinaria y Zootecnia, Universidad de Guayaquil, Ecuador
  • María Baquero C. Facultad de Medicina y Zootecnia, Universidad Central del Ecuador, Quito, Ecuador
  • Yveth Casart Q. Proyecto Prometeo, SENESCYT, Ecuador

DOI:

https://doi.org/10.15381/rivep.v31i2.15522

Keywords:

leptospirosis, zoonosis, diagnosis, DNA extraction, Chelex, PCR

Abstract

The purpose of this study was to select for the extraction of DNA from Leptospira spp from urine samples for the diagnosis of bovine leptospirosis by PCR. Three methods of DNA extraction methods were used: Ethanol-Sodium Hydroxide (EtNa), Chelex® 100 chelating resin and the PureLink® Genomic DNA Mini Kit commercial extraction case. The extracted DNA served for the standardization of three PCR protocols for the identification of the rrl, hap1 and rrs genes, respectively, in the AGROCALIDAD, Ecuador laboratory. A total of 72 bovine urine samples were collected from livestock farms in the province of Manabí, Ecuador. Ten positive samples were obtained by amplifying the rrl gene, which identifies the genus Leptospira. Of the genus-positive samples, eight amplified for the hap1 gene, which codes for the main outer membrane protein of pathogenic species. The agreement between the DNA extraction methods was evaluated with Chelex-100, and the PureLink® Genomic DNA Mini Kit, determining a agreement of 0.74 using the Kappa index.

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Author Biography

  • Alexandra Paola Revelo Ruales, Facultad de Medicina y Zootecnia, Universidad Central del Ecuador, Quito, Ecuador

    AGROCALIDAD-Dirección de Diagnóstico Animal de la Coordinación General de Laboratorios, Analista de Laboratorio de Microbiología.

    Instituto Nacional de Investigación en Salud Pública-INSPI-Dr. Leopoldo Izquieta Pérez, Analista del Centro de Referencia Nacional de Zoonosis (Trabajo actual)

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Published

2020-06-20

Issue

Vol. 31 No. 2 (2020)

Section

Artículos Primarios

License

Copyright (c) 2020 Alexandra Paola Revelo Ruales, Euclides De La Torre M., Galo Martínez C., María Baquero C., Yveth Casart Q.

Creative Commons License

This work is licensed under a Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International License.

AUTHORS RETAIN THEIR RIGHTS:

a. Authors retain their trade mark rights and patent, and also on any process or procedure described in the article.

b. Authors retain their right to share, copy, distribute, perform and publicly communicate their article (eg, to place their article in an institutional repository or publish it in a book), with an acknowledgment of its initial publication in the Revista de Investigaciones Veterinarias del Perú (RIVEP).

c. Authors retain theirs right to make a subsequent publication of their work, to use the article or any part thereof (eg a compilation of his papers, lecture notes, thesis, or a book), always indicating the source of publication (the originator of the work, journal, volume, number and date).

How to Cite

Revelo Ruales, A. P., De La Torre M., E., Martínez C., G., Baquero C., M., & Casart Q., Y. (2020). Evaluation of genomic DNA extraction methods for the identification of Leptospira spp. in bovine urine samples by PCR. Revista De Investigaciones Veterinarias Del Perú, 31(2), e15522. https://doi.org/10.15381/rivep.v31i2.15522