Isolation and genetic characterization of Toxoplasma gondii in Spanish sheep flocks

Background: Toxoplasma gondii is a major cause of abortion in small ruminants and presents a zoonotic risk when undercooked meat containing cysts is consumed. The aim of the present study was to investigate the genetic diversity among the T. gondii strains circulating in ovine livestock in Spain. Me...

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Authors: Benavides, Julio, Fernández Escobar, Mercedes, Calero Bernal, Rafael, Regidor-Cerrillo, Javier, Guerrero-Molina, María Cristina, Gutiérrez-Expósito, Daniel, Collantes Fernández, Esther, Ortega Mora, Luis Miguel
Format: article
Publication Date:2020
Country:España
Institution:Universidad Complutense de Madrid (UCM)
Repository:Docta Complutense
Language:English
OAI Identifier:oai:docta.ucm.es:20.500.14352/96793
Online Access:https://hdl.handle.net/20.500.14352/96793
Access Level:Open access
Keyword:Toxoplasma gondii
Sheep
Abortion
Isolates
Genotyping
Sequencing,
Population structure
Spain
Ciencias Biomédicas
24 Ciencias de la Vida
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spelling Isolation and genetic characterization of Toxoplasma gondii in Spanish sheep flocksBenavides, JulioFernández Escobar, MercedesCalero Bernal, RafaelRegidor-Cerrillo, JavierGuerrero-Molina, María CristinaGutiérrez-Expósito, DanielCollantes Fernández, EstherOrtega Mora, Luis MiguelToxoplasma gondiiSheepAbortionIsolatesGenotypingSequencing,Population structureSpainCiencias Biomédicas24 Ciencias de la VidaBackground: Toxoplasma gondii is a major cause of abortion in small ruminants and presents a zoonotic risk when undercooked meat containing cysts is consumed. The aim of the present study was to investigate the genetic diversity among the T. gondii strains circulating in ovine livestock in Spain. Methods: Selected samples collected from abortion outbreaks due to toxoplasmosis (n = 31) and from chronically infected adult sheep at slaughterhouses (n = 50) in different Spanish regions were bioassayed in mice, aiming at parasite isolation. In addition, all original clinical samples and the resulting isolates were genotyped by multi-nested PCR-RFLP analysis of 11 molecular markers and by PCR-DNA sequencing of portions of the SAG3, GRA6 and GRA7 genes. Results: As a result, 30 isolates were obtained from 9 Spanish regions: 10 isolates from abortion-derived samples and 20 isolates from adult myocardial tissues. Overall, 3 genotypes were found: ToxoDB#3 (type II PRU variant) in 90% (27/30) of isolates, ToxoDB#2 (clonal type III) in 6.7% (2/30), and ToxoDB#1 (clonal type II) in 3.3% (1/30). When T. gondii-positive tissue samples (n = 151) were directly subjected to RFLP genotyping, complete restriction profiles were obtained for 33% of samples, and up to 98% of the specimens belonged to the type II PRU variant. A foetal brain showed a clonal type II pattern, and four specimens showed unexpected type I alleles at the SAG3 marker, including two foetal brains that showed I + II alleles as co-infection events. Amplicons of SAG3, GRA6 and GRA7 obtained from isolates and clinical samples were subjected to sequencing, allowing us to confirm RFLP results and to detect different single-nucleotide polymorphisms. Conclusions: The present study informed the existence of a predominant type II PRU variant genotype (ToxoDB#3) infecting domestic sheep in Spain, in both abortion cases and chronic infections in adults, coexisting with other clonal (ToxoDB#1 and ToxoDB#2), much less frequent genotypes, as well as polymorphic strains as revealed by clinical sample genotyping. The use of multilocus sequence typing aided in accurately estimating T. gondii intragenotype diversity.Universidad Complutense de Madrid20202020-01-0120202020-01-01journal articlehttp://purl.org/coar/resource_type/c_6501VoRhttp://purl.org/coar/version/c_970fb48d4fbd8a85info:eu-repo/semantics/articleapplication/pdfhttps://hdl.handle.net/20.500.14352/96793reponame:Docta Complutenseinstname:Universidad Complutense de Madrid (UCM)Inglésengopen accesshttp://purl.org/coar/access_right/c_abf2Attribution 4.0 Internationalhttp://creativecommons.org/licenses/by/4.0/info:eu-repo/semantics/openAccessoai:docta.ucm.es:20.500.14352/967932026-06-02T12:44:21Z
dc.title.none.fl_str_mv Isolation and genetic characterization of Toxoplasma gondii in Spanish sheep flocks
title Isolation and genetic characterization of Toxoplasma gondii in Spanish sheep flocks
spellingShingle Isolation and genetic characterization of Toxoplasma gondii in Spanish sheep flocks
Benavides, Julio
Toxoplasma gondii
Sheep
Abortion
Isolates
Genotyping
Sequencing,
Population structure
Spain
Ciencias Biomédicas
24 Ciencias de la Vida
title_short Isolation and genetic characterization of Toxoplasma gondii in Spanish sheep flocks
title_full Isolation and genetic characterization of Toxoplasma gondii in Spanish sheep flocks
title_fullStr Isolation and genetic characterization of Toxoplasma gondii in Spanish sheep flocks
title_full_unstemmed Isolation and genetic characterization of Toxoplasma gondii in Spanish sheep flocks
title_sort Isolation and genetic characterization of Toxoplasma gondii in Spanish sheep flocks
dc.creator.none.fl_str_mv Benavides, Julio
Fernández Escobar, Mercedes
Calero Bernal, Rafael
Regidor-Cerrillo, Javier
Guerrero-Molina, María Cristina
Gutiérrez-Expósito, Daniel
Collantes Fernández, Esther
Ortega Mora, Luis Miguel
author Benavides, Julio
author_facet Benavides, Julio
Fernández Escobar, Mercedes
Calero Bernal, Rafael
Regidor-Cerrillo, Javier
Guerrero-Molina, María Cristina
Gutiérrez-Expósito, Daniel
Collantes Fernández, Esther
Ortega Mora, Luis Miguel
author_role author
author2 Fernández Escobar, Mercedes
Calero Bernal, Rafael
Regidor-Cerrillo, Javier
Guerrero-Molina, María Cristina
Gutiérrez-Expósito, Daniel
Collantes Fernández, Esther
Ortega Mora, Luis Miguel
author2_role author
author
author
author
author
author
author
dc.contributor.none.fl_str_mv Universidad Complutense de Madrid
dc.subject.none.fl_str_mv Toxoplasma gondii
Sheep
Abortion
Isolates
Genotyping
Sequencing,
Population structure
Spain
Ciencias Biomédicas
24 Ciencias de la Vida
topic Toxoplasma gondii
Sheep
Abortion
Isolates
Genotyping
Sequencing,
Population structure
Spain
Ciencias Biomédicas
24 Ciencias de la Vida
description Background: Toxoplasma gondii is a major cause of abortion in small ruminants and presents a zoonotic risk when undercooked meat containing cysts is consumed. The aim of the present study was to investigate the genetic diversity among the T. gondii strains circulating in ovine livestock in Spain. Methods: Selected samples collected from abortion outbreaks due to toxoplasmosis (n = 31) and from chronically infected adult sheep at slaughterhouses (n = 50) in different Spanish regions were bioassayed in mice, aiming at parasite isolation. In addition, all original clinical samples and the resulting isolates were genotyped by multi-nested PCR-RFLP analysis of 11 molecular markers and by PCR-DNA sequencing of portions of the SAG3, GRA6 and GRA7 genes. Results: As a result, 30 isolates were obtained from 9 Spanish regions: 10 isolates from abortion-derived samples and 20 isolates from adult myocardial tissues. Overall, 3 genotypes were found: ToxoDB#3 (type II PRU variant) in 90% (27/30) of isolates, ToxoDB#2 (clonal type III) in 6.7% (2/30), and ToxoDB#1 (clonal type II) in 3.3% (1/30). When T. gondii-positive tissue samples (n = 151) were directly subjected to RFLP genotyping, complete restriction profiles were obtained for 33% of samples, and up to 98% of the specimens belonged to the type II PRU variant. A foetal brain showed a clonal type II pattern, and four specimens showed unexpected type I alleles at the SAG3 marker, including two foetal brains that showed I + II alleles as co-infection events. Amplicons of SAG3, GRA6 and GRA7 obtained from isolates and clinical samples were subjected to sequencing, allowing us to confirm RFLP results and to detect different single-nucleotide polymorphisms. Conclusions: The present study informed the existence of a predominant type II PRU variant genotype (ToxoDB#3) infecting domestic sheep in Spain, in both abortion cases and chronic infections in adults, coexisting with other clonal (ToxoDB#1 and ToxoDB#2), much less frequent genotypes, as well as polymorphic strains as revealed by clinical sample genotyping. The use of multilocus sequence typing aided in accurately estimating T. gondii intragenotype diversity.
publishDate 2020
dc.date.none.fl_str_mv 2020
2020-01-01
2020
2020-01-01
dc.type.none.fl_str_mv journal article
http://purl.org/coar/resource_type/c_6501
VoR
http://purl.org/coar/version/c_970fb48d4fbd8a85
dc.type.openaire.fl_str_mv info:eu-repo/semantics/article
format article
dc.identifier.none.fl_str_mv https://hdl.handle.net/20.500.14352/96793
url https://hdl.handle.net/20.500.14352/96793
dc.language.none.fl_str_mv Inglés
eng
language_invalid_str_mv Inglés
language eng
dc.rights.none.fl_str_mv open access
http://purl.org/coar/access_right/c_abf2
Attribution 4.0 International
http://creativecommons.org/licenses/by/4.0/
dc.rights.openaire.fl_str_mv info:eu-repo/semantics/openAccess
rights_invalid_str_mv open access
http://purl.org/coar/access_right/c_abf2
Attribution 4.0 International
http://creativecommons.org/licenses/by/4.0/
eu_rights_str_mv openAccess
dc.format.none.fl_str_mv application/pdf
dc.source.none.fl_str_mv reponame:Docta Complutense
instname:Universidad Complutense de Madrid (UCM)
instname_str Universidad Complutense de Madrid (UCM)
reponame_str Docta Complutense
collection Docta Complutense
repository.name.fl_str_mv
repository.mail.fl_str_mv
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