Loop-mediated Isothermal Amplification (LAMP) for Identification of Pythium insidiosum.
Int J Infect Dis
; 101: 149-159, 2020 Dec.
Article
en En
| MEDLINE
| ID: mdl-32987181
ABSTRACT
OBJECTIVE:
Pythium insidiosum causes a life-threatening condition called pythiosis. High morbidity and mortality of pythiosis are consequences of delayed diagnosis. We aimed to develop a loop-mediated isothermal amplification (LAMP) assay for the rapid detection of P. insidiosum for use in remote areas, where pythiosis is prevalent.METHODS:
We designed four LAMP primers to amplify the rDNA sequence. A side-by-side comparison evaluated performances of LAMP and the previously-established multiplex PCR (M-PCR), using gDNA samples extracted from colonies of P. insidiosum (n = 28) and other fungi (n = 54), and tissues of animals with (n = 16) or without (n = 13) pythiosis.RESULTS:
LAMP demonstrated a 50% shorter assay duration (1.5 h) and a 10-fold lower limit of detection (10-4 ng) than did M-PCR. Based on colony-extracted gDNAs, LAMP and M-PCR correctly reported P. insidiosum in all 28 samples, providing 100% sensitivity. While M-PCR did not amplify all fungal controls (100% specificity), LAMP falsely detected one organism (98% specificity). Based on the clinical samples, LAMP and M-PCR provided an equivalently-high specificity (100%). However, LAMP showed a markedly-higher sensitivity than that of M-PCR (88% vs. 56%).CONCLUSIONS:
LAMP is a simple, useful, efficient assay for the detection of P. insidiosum in clinical specimens and pure cultures in resource-limited laboratories.Palabras clave
Texto completo:
1
Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Pythium
/
Técnicas de Amplificación de Ácido Nucleico
/
Técnicas de Diagnóstico Molecular
/
Enfermedades de los Perros
/
Pitiosis
/
Enfermedades de los Caballos
Tipo de estudio:
Diagnostic_studies
/
Evaluation_studies
/
Prognostic_studies
Límite:
Animals
/
Humans
Idioma:
En
Revista:
Int J Infect Dis
Asunto de la revista:
DOENCAS TRANSMISSIVEIS
Año:
2020
Tipo del documento:
Article
País de afiliación:
Tailandia