A novel loop-mediated isothermal amplification-based test for detecting Neospora caninum DNA

Andrea Estefanía Ramos, Marina Muñoz, Jesús Alfredo Cortés-Vecino, Paola Barato, Manuel Alfonso Patarroyo

Resultado de la investigación: Contribución a RevistaArtículo

2 Citas (Scopus)

Resumen

BACKGROUND: Neospora caninum is a cyst-forming, coccidian parasite which is known to cause neurological disorders in dogs and abortion and neonatal mortality in cows and other livestock. This study reports the development of a loop-mediated isothermal amplification (LAMP) assay based on the Neospora caninum Nc-5 gene and compares its efficacy for detecting DNA to that of a semi-nested PCR test.

RESULTS: Six primers were designed based on the Nc-5 repeat region of N. caninum. Specific LAMP primers led to successful amplification of N. caninum DNA at 63 °C in 30 min. The LAMP assay was highly specific (i.e. it did not reveal cross-reactivity with other parasite species) and had a low N. caninum plasmid DNA limit of detection (1 fg), which is ten times higher than that for the semi-nested PCR. LAMP applicability was evaluated using a set of naturally-infected samples (59 from canine faeces and five from bovine abortions). Thirty-nine percent (25/64) of the naturally-infected samples were positive for N. caninum DNA by LAMP and 36% (23/64) by semi-nested PCR. However, the LAMP assay is much faster to perform than semi-nested PCR and provides results in 30 min.

CONCLUSION: The optimized reaction conditions described in this study resulted in a sensitive, specific and rapid technique for detecting N. caninum DNA. Considering the advantages of LAMP for detecting N. caninum DNA, further assays aimed at testing its usefulness on a wider range of field samples are recommended.

Idioma originalEnglish (US)
Páginas (desde-hasta)1-9
Número de páginas9
PublicaciónParasites and Vectors
Volumen10
N.º1
DOI
EstadoPublished - nov 29 2017

Citar esto

Ramos, Andrea Estefanía ; Muñoz, Marina ; Cortés-Vecino, Jesús Alfredo ; Barato, Paola ; Patarroyo, Manuel Alfonso. / A novel loop-mediated isothermal amplification-based test for detecting Neospora caninum DNA. En: Parasites and Vectors. 2017 ; Vol. 10, N.º 1. pp. 1-9.
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abstract = "BACKGROUND: Neospora caninum is a cyst-forming, coccidian parasite which is known to cause neurological disorders in dogs and abortion and neonatal mortality in cows and other livestock. This study reports the development of a loop-mediated isothermal amplification (LAMP) assay based on the Neospora caninum Nc-5 gene and compares its efficacy for detecting DNA to that of a semi-nested PCR test.RESULTS: Six primers were designed based on the Nc-5 repeat region of N. caninum. Specific LAMP primers led to successful amplification of N. caninum DNA at 63 °C in 30 min. The LAMP assay was highly specific (i.e. it did not reveal cross-reactivity with other parasite species) and had a low N. caninum plasmid DNA limit of detection (1 fg), which is ten times higher than that for the semi-nested PCR. LAMP applicability was evaluated using a set of naturally-infected samples (59 from canine faeces and five from bovine abortions). Thirty-nine percent (25/64) of the naturally-infected samples were positive for N. caninum DNA by LAMP and 36{\%} (23/64) by semi-nested PCR. However, the LAMP assay is much faster to perform than semi-nested PCR and provides results in 30 min.CONCLUSION: The optimized reaction conditions described in this study resulted in a sensitive, specific and rapid technique for detecting N. caninum DNA. Considering the advantages of LAMP for detecting N. caninum DNA, further assays aimed at testing its usefulness on a wider range of field samples are recommended.",
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A novel loop-mediated isothermal amplification-based test for detecting Neospora caninum DNA. / Ramos, Andrea Estefanía; Muñoz, Marina; Cortés-Vecino, Jesús Alfredo; Barato, Paola; Patarroyo, Manuel Alfonso.

En: Parasites and Vectors, Vol. 10, N.º 1, 29.11.2017, p. 1-9.

Resultado de la investigación: Contribución a RevistaArtículo

TY - JOUR

T1 - A novel loop-mediated isothermal amplification-based test for detecting Neospora caninum DNA

AU - Ramos, Andrea Estefanía

AU - Muñoz, Marina

AU - Cortés-Vecino, Jesús Alfredo

AU - Barato, Paola

AU - Patarroyo, Manuel Alfonso

PY - 2017/11/29

Y1 - 2017/11/29

N2 - BACKGROUND: Neospora caninum is a cyst-forming, coccidian parasite which is known to cause neurological disorders in dogs and abortion and neonatal mortality in cows and other livestock. This study reports the development of a loop-mediated isothermal amplification (LAMP) assay based on the Neospora caninum Nc-5 gene and compares its efficacy for detecting DNA to that of a semi-nested PCR test.RESULTS: Six primers were designed based on the Nc-5 repeat region of N. caninum. Specific LAMP primers led to successful amplification of N. caninum DNA at 63 °C in 30 min. The LAMP assay was highly specific (i.e. it did not reveal cross-reactivity with other parasite species) and had a low N. caninum plasmid DNA limit of detection (1 fg), which is ten times higher than that for the semi-nested PCR. LAMP applicability was evaluated using a set of naturally-infected samples (59 from canine faeces and five from bovine abortions). Thirty-nine percent (25/64) of the naturally-infected samples were positive for N. caninum DNA by LAMP and 36% (23/64) by semi-nested PCR. However, the LAMP assay is much faster to perform than semi-nested PCR and provides results in 30 min.CONCLUSION: The optimized reaction conditions described in this study resulted in a sensitive, specific and rapid technique for detecting N. caninum DNA. Considering the advantages of LAMP for detecting N. caninum DNA, further assays aimed at testing its usefulness on a wider range of field samples are recommended.

AB - BACKGROUND: Neospora caninum is a cyst-forming, coccidian parasite which is known to cause neurological disorders in dogs and abortion and neonatal mortality in cows and other livestock. This study reports the development of a loop-mediated isothermal amplification (LAMP) assay based on the Neospora caninum Nc-5 gene and compares its efficacy for detecting DNA to that of a semi-nested PCR test.RESULTS: Six primers were designed based on the Nc-5 repeat region of N. caninum. Specific LAMP primers led to successful amplification of N. caninum DNA at 63 °C in 30 min. The LAMP assay was highly specific (i.e. it did not reveal cross-reactivity with other parasite species) and had a low N. caninum plasmid DNA limit of detection (1 fg), which is ten times higher than that for the semi-nested PCR. LAMP applicability was evaluated using a set of naturally-infected samples (59 from canine faeces and five from bovine abortions). Thirty-nine percent (25/64) of the naturally-infected samples were positive for N. caninum DNA by LAMP and 36% (23/64) by semi-nested PCR. However, the LAMP assay is much faster to perform than semi-nested PCR and provides results in 30 min.CONCLUSION: The optimized reaction conditions described in this study resulted in a sensitive, specific and rapid technique for detecting N. caninum DNA. Considering the advantages of LAMP for detecting N. caninum DNA, further assays aimed at testing its usefulness on a wider range of field samples are recommended.

U2 - 10.1186/s13071-017-2549-y

DO - 10.1186/s13071-017-2549-y

M3 - Article

C2 - 29187255

VL - 10

SP - 1

EP - 9

JO - Parasites and Vectors

JF - Parasites and Vectors

SN - 1756-3305

IS - 1

ER -