Validation of a PCR Assay for Chlamydophila abortus rRNA gene detection in a murine model
Chlamydophila abortus (C. abortus) is associated with reproductive problems in cattle, sheep, and goats. Diagnosis of C. abortus using embryonated chicken eggs or immortalized cell lines has a very low sensitivity. Polymerase chain reaction (PCR) assays have been used to detect C. abortus infection in clinical specimens and organ fragments, such as placenta, fetal organs, vaginal secretions, and semen. The aim of this study was to develop a PCR assay for the amplification of an 856-bp fragment of the rRNA gene of the Chlamydiaceae family. The PCR assay was evaluated using organs from 15 mice experimentally infected with the S26/3 reference strain of C. abortus. The results of the rRNA PCR were compared to the results from another PCR system (Omp2 PCR) that has been previously described for the Omp2 (outer major protein) gene from the Chlamydiaceae family. From the 15 C. abortus-inoculated mice, 13 (K=0.84, standard error =0.20) tested positive using the rRNA PCR assay and 9 (K=0.55, standard error=0.18) tested positive using the Omp2 PCR assay. The detection limit, measured using inclusion-forming units (IFU), for C. abortus with the rRNA PCR (1.05 IFU) was 100-fold lower than for the Omp2 PCR (105 IFU). The higher sensitivity of the rRNA PCR, as compared to the previously described PCR assay, and the specificity of the assay, demonstrated using different pathogenic microorganisms of the bovine reproductive system, suggest that the new PCR assay developed in this study can be used for the molecular diagnosis of C. abortus in abortion and other reproductive failures in bovines, caprines, and ovines.
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Instituto de Tecnologia do Paraná - Tecpar
2009
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oai:scielo:S1516-891320090007000142010-02-11Validation of a PCR Assay for Chlamydophila abortus rRNA gene detection in a murine modelSilva-Zacarias,Francielle Gibson daAlfieri,Amauri AlcindoSpohr,Kledir Anderson HofstaetterLima,Bruna Azevedo de CarvalhoNegrão,Fábio JulianoLunardi,MicheleFreitas,Julio Cesar de bovine ovine caprine reproductive failures Chlamydophila abortus PCR Chlamydophila abortus (C. abortus) is associated with reproductive problems in cattle, sheep, and goats. Diagnosis of C. abortus using embryonated chicken eggs or immortalized cell lines has a very low sensitivity. Polymerase chain reaction (PCR) assays have been used to detect C. abortus infection in clinical specimens and organ fragments, such as placenta, fetal organs, vaginal secretions, and semen. The aim of this study was to develop a PCR assay for the amplification of an 856-bp fragment of the rRNA gene of the Chlamydiaceae family. The PCR assay was evaluated using organs from 15 mice experimentally infected with the S26/3 reference strain of C. abortus. The results of the rRNA PCR were compared to the results from another PCR system (Omp2 PCR) that has been previously described for the Omp2 (outer major protein) gene from the Chlamydiaceae family. From the 15 C. abortus-inoculated mice, 13 (K=0.84, standard error =0.20) tested positive using the rRNA PCR assay and 9 (K=0.55, standard error=0.18) tested positive using the Omp2 PCR assay. The detection limit, measured using inclusion-forming units (IFU), for C. abortus with the rRNA PCR (1.05 IFU) was 100-fold lower than for the Omp2 PCR (105 IFU). The higher sensitivity of the rRNA PCR, as compared to the previously described PCR assay, and the specificity of the assay, demonstrated using different pathogenic microorganisms of the bovine reproductive system, suggest that the new PCR assay developed in this study can be used for the molecular diagnosis of C. abortus in abortion and other reproductive failures in bovines, caprines, and ovines.info:eu-repo/semantics/openAccessInstituto de Tecnologia do Paraná - TecparBrazilian Archives of Biology and Technology v.52 n.spe 20092009-11-01info:eu-repo/semantics/articletext/htmlhttp://old.scielo.br/scielo.php?script=sci_arttext&pid=S1516-89132009000700014en10.1590/S1516-89132009000700014 |
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Silva-Zacarias,Francielle Gibson da Alfieri,Amauri Alcindo Spohr,Kledir Anderson Hofstaetter Lima,Bruna Azevedo de Carvalho Negrão,Fábio Juliano Lunardi,Michele Freitas,Julio Cesar de |
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Silva-Zacarias,Francielle Gibson da Alfieri,Amauri Alcindo Spohr,Kledir Anderson Hofstaetter Lima,Bruna Azevedo de Carvalho Negrão,Fábio Juliano Lunardi,Michele Freitas,Julio Cesar de Validation of a PCR Assay for Chlamydophila abortus rRNA gene detection in a murine model |
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Silva-Zacarias,Francielle Gibson da Alfieri,Amauri Alcindo Spohr,Kledir Anderson Hofstaetter Lima,Bruna Azevedo de Carvalho Negrão,Fábio Juliano Lunardi,Michele Freitas,Julio Cesar de |
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Silva-Zacarias,Francielle Gibson da |
title |
Validation of a PCR Assay for Chlamydophila abortus rRNA gene detection in a murine model |
title_short |
Validation of a PCR Assay for Chlamydophila abortus rRNA gene detection in a murine model |
title_full |
Validation of a PCR Assay for Chlamydophila abortus rRNA gene detection in a murine model |
title_fullStr |
Validation of a PCR Assay for Chlamydophila abortus rRNA gene detection in a murine model |
title_full_unstemmed |
Validation of a PCR Assay for Chlamydophila abortus rRNA gene detection in a murine model |
title_sort |
validation of a pcr assay for chlamydophila abortus rrna gene detection in a murine model |
description |
Chlamydophila abortus (C. abortus) is associated with reproductive problems in cattle, sheep, and goats. Diagnosis of C. abortus using embryonated chicken eggs or immortalized cell lines has a very low sensitivity. Polymerase chain reaction (PCR) assays have been used to detect C. abortus infection in clinical specimens and organ fragments, such as placenta, fetal organs, vaginal secretions, and semen. The aim of this study was to develop a PCR assay for the amplification of an 856-bp fragment of the rRNA gene of the Chlamydiaceae family. The PCR assay was evaluated using organs from 15 mice experimentally infected with the S26/3 reference strain of C. abortus. The results of the rRNA PCR were compared to the results from another PCR system (Omp2 PCR) that has been previously described for the Omp2 (outer major protein) gene from the Chlamydiaceae family. From the 15 C. abortus-inoculated mice, 13 (K=0.84, standard error =0.20) tested positive using the rRNA PCR assay and 9 (K=0.55, standard error=0.18) tested positive using the Omp2 PCR assay. The detection limit, measured using inclusion-forming units (IFU), for C. abortus with the rRNA PCR (1.05 IFU) was 100-fold lower than for the Omp2 PCR (105 IFU). The higher sensitivity of the rRNA PCR, as compared to the previously described PCR assay, and the specificity of the assay, demonstrated using different pathogenic microorganisms of the bovine reproductive system, suggest that the new PCR assay developed in this study can be used for the molecular diagnosis of C. abortus in abortion and other reproductive failures in bovines, caprines, and ovines. |
publisher |
Instituto de Tecnologia do Paraná - Tecpar |
publishDate |
2009 |
url |
http://old.scielo.br/scielo.php?script=sci_arttext&pid=S1516-89132009000700014 |
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