Multi-Target Strategy for Pan/Foot-and-Mouth Disease Virus (FMDV) Detection: A Combination of Sequences Analysis, in Silico Predictions and Laboratory Diagnostic Evaluation
Foot-and-mouth disease (FMD) is a highly contagious viral disease affecting cloven-hoofed animals that causes severe economic losses. The disease is characterized by a vesicular condition and it cannot be differentiated from other vesicular diseases. Therefore, laboratory confirmation of any suspect...
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doaj-c201f14bf15d492cba965aae916459ff2020-11-24T23:56:09ZengFrontiers Media S.A.Frontiers in Veterinary Science2297-17692018-07-01510.3389/fvets.2018.00160385721Multi-Target Strategy for Pan/Foot-and-Mouth Disease Virus (FMDV) Detection: A Combination of Sequences Analysis, in Silico Predictions and Laboratory Diagnostic EvaluationLiliam Rios0Carmen L. Perera1Liani Coronado2Damarys Relova3Ana M. Álvarez4Llilianne Ganges5Llilianne Ganges6Heidy Díaz de Arce7José I. Núñez8Lester J. Pérez9Reiman Cancer Research Laboratory, Faculty of Medicine, University of New Brunswick, Saint John, NB, CanadaCentro Nacional de Sanidad Agropecuaria, OIE Collaborating Centre for Diagnosis and Risk Analysis of the Caribean Region, San José de las Lajas, CubaCentro Nacional de Sanidad Agropecuaria, OIE Collaborating Centre for Diagnosis and Risk Analysis of the Caribean Region, San José de las Lajas, CubaCentro Nacional de Sanidad Agropecuaria, OIE Collaborating Centre for Diagnosis and Risk Analysis of the Caribean Region, San José de las Lajas, CubaInstituto Nacional de Investigaciones Agricolas, Maracay, VenezuelaOIE Reference Laboratory for Classical Swine Fever, IRTA-CReSA, Barcelona, SpainIRTA, Centre de Recerca en Sanitat Animal (CReSA, IRTA-UAB), Campus de la Universitat Autonoma de Barcelona, Barcelona, SpainHospital Italiano de Buenos Aires, Buenos Aires, ArgentinaIRTA, Centre de Recerca en Sanitat Animal (CReSA, IRTA-UAB), Campus de la Universitat Autonoma de Barcelona, Barcelona, SpainDalhousie Medicine New Brunswick, Dalhousie University, Saint John, NB, CanadaFoot-and-mouth disease (FMD) is a highly contagious viral disease affecting cloven-hoofed animals that causes severe economic losses. The disease is characterized by a vesicular condition and it cannot be differentiated from other vesicular diseases. Therefore, laboratory confirmation of any suspected FMD case is compulsory. Despite viral isolation in cell cultures has been considered for many years as the gold standard for FMD diagnosis, the advantages of real-time reverse transcription polymerase chain reaction (rRT-PCR) technology have motivated its use directly in clinical specimens for FMD diagnosis. The current work was aimed to develop and validate a molecular multi-check strategy using rRT-PCR (mMulti-rRT-PCR) based on SYBR-Green I for pan/foot-and-mouth disease virus (pan/FMDV) diagnosis. From in silico approaches, different primer pairs previously reported were selected and modified to reduce the likelihood of viral escape as well as potential failures in the pan/FMDV detection. The analytical parameters were evaluated using a high number of representative viral strains. The repeatability of the assay and its performance on field samples were also assessed. The mMulti-rRT-PCR was able to detect emergent FMDV strains that circulated in South America between the years 2006–2010 and on which the single rRT-PCRs failed when they were applied independently. The results obtained here showed that the proposed system is an accurate and rapid diagnosis method for sensitive and specific detection of FMDV. Thus, a validated mMulti-rRT-PCR assay based on SYBR-Green I detection coupled to melting curves resolution for pan/FMDV diagnosis on clinical samples is proposed. This study also highlights the need to incorporate the multi-target detection principle in the diagnosis of highly variable agents, specially, of those listed by OIE like FMDV.https://www.frontiersin.org/article/10.3389/fvets.2018.00160/fullfoot-and-mouth disease virusmulti-target strategysequences analysisprimer efficiency evaluationentropyamplicon stability |
collection |
DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
Liliam Rios Carmen L. Perera Liani Coronado Damarys Relova Ana M. Álvarez Llilianne Ganges Llilianne Ganges Heidy Díaz de Arce José I. Núñez Lester J. Pérez |
spellingShingle |
Liliam Rios Carmen L. Perera Liani Coronado Damarys Relova Ana M. Álvarez Llilianne Ganges Llilianne Ganges Heidy Díaz de Arce José I. Núñez Lester J. Pérez Multi-Target Strategy for Pan/Foot-and-Mouth Disease Virus (FMDV) Detection: A Combination of Sequences Analysis, in Silico Predictions and Laboratory Diagnostic Evaluation Frontiers in Veterinary Science foot-and-mouth disease virus multi-target strategy sequences analysis primer efficiency evaluation entropy amplicon stability |
author_facet |
Liliam Rios Carmen L. Perera Liani Coronado Damarys Relova Ana M. Álvarez Llilianne Ganges Llilianne Ganges Heidy Díaz de Arce José I. Núñez Lester J. Pérez |
author_sort |
Liliam Rios |
title |
Multi-Target Strategy for Pan/Foot-and-Mouth Disease Virus (FMDV) Detection: A Combination of Sequences Analysis, in Silico Predictions and Laboratory Diagnostic Evaluation |
title_short |
Multi-Target Strategy for Pan/Foot-and-Mouth Disease Virus (FMDV) Detection: A Combination of Sequences Analysis, in Silico Predictions and Laboratory Diagnostic Evaluation |
title_full |
Multi-Target Strategy for Pan/Foot-and-Mouth Disease Virus (FMDV) Detection: A Combination of Sequences Analysis, in Silico Predictions and Laboratory Diagnostic Evaluation |
title_fullStr |
Multi-Target Strategy for Pan/Foot-and-Mouth Disease Virus (FMDV) Detection: A Combination of Sequences Analysis, in Silico Predictions and Laboratory Diagnostic Evaluation |
title_full_unstemmed |
Multi-Target Strategy for Pan/Foot-and-Mouth Disease Virus (FMDV) Detection: A Combination of Sequences Analysis, in Silico Predictions and Laboratory Diagnostic Evaluation |
title_sort |
multi-target strategy for pan/foot-and-mouth disease virus (fmdv) detection: a combination of sequences analysis, in silico predictions and laboratory diagnostic evaluation |
publisher |
Frontiers Media S.A. |
series |
Frontiers in Veterinary Science |
issn |
2297-1769 |
publishDate |
2018-07-01 |
description |
Foot-and-mouth disease (FMD) is a highly contagious viral disease affecting cloven-hoofed animals that causes severe economic losses. The disease is characterized by a vesicular condition and it cannot be differentiated from other vesicular diseases. Therefore, laboratory confirmation of any suspected FMD case is compulsory. Despite viral isolation in cell cultures has been considered for many years as the gold standard for FMD diagnosis, the advantages of real-time reverse transcription polymerase chain reaction (rRT-PCR) technology have motivated its use directly in clinical specimens for FMD diagnosis. The current work was aimed to develop and validate a molecular multi-check strategy using rRT-PCR (mMulti-rRT-PCR) based on SYBR-Green I for pan/foot-and-mouth disease virus (pan/FMDV) diagnosis. From in silico approaches, different primer pairs previously reported were selected and modified to reduce the likelihood of viral escape as well as potential failures in the pan/FMDV detection. The analytical parameters were evaluated using a high number of representative viral strains. The repeatability of the assay and its performance on field samples were also assessed. The mMulti-rRT-PCR was able to detect emergent FMDV strains that circulated in South America between the years 2006–2010 and on which the single rRT-PCRs failed when they were applied independently. The results obtained here showed that the proposed system is an accurate and rapid diagnosis method for sensitive and specific detection of FMDV. Thus, a validated mMulti-rRT-PCR assay based on SYBR-Green I detection coupled to melting curves resolution for pan/FMDV diagnosis on clinical samples is proposed. This study also highlights the need to incorporate the multi-target detection principle in the diagnosis of highly variable agents, specially, of those listed by OIE like FMDV. |
topic |
foot-and-mouth disease virus multi-target strategy sequences analysis primer efficiency evaluation entropy amplicon stability |
url |
https://www.frontiersin.org/article/10.3389/fvets.2018.00160/full |
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