Abstract
Zebra chip disease (ZC) of potato tubers, caused by the unculturable bacterium ‘Candidatus Liberibacter solanacearum’ (Lso) can be difficult to detect from potato tuber tissue. The polymerase chain reaction is an important tool for the detection of plant pathogens, but there are limits to its sensitivity. High-fidelity PCR incorporates a proofreading enzyme with 3′-5′ exonuclease activity and has been shown to increase detection of pathogens when the titer is low or when large amounts of competitor DNA or other host cellular contents inhibit PCR amplification. In this study, high-fidelity PCR and conventional PCR protocols were performed on tubers presenting ZC symptoms. Asymptomatic tubers were used as control. The high-fidelity PCR increased detection of Lso from symptomatic tubers by 30–40 % over conventional PCR. Slight modifications were made from previous high-fidelity protocols to decrease testing time and simplify the protocol.
Resumen
La enfermedad de la Zebra chip (ZC) o papa rayada en tubérculos de papa, causada por la bacteria no cultivable ‘Candidatus Liberibacter solanacearum’ (Lso), puede ser difícil de detectar de tejido de tubérculo. La reacción en cadena de la polimerasa (PCR) es una herramienta importante para la detección de fitopatógenos, pero hay límites en su sensibilidad. La PCR de alta fidelidad incorpora una enzima leíble en la prueba con actividad de exonucleasa 3′-5′ y se ha demostrado que incrementa la detección de patógenos cuando el título es bajo o cuando grandes cantidades del ADN competidor u otro contenido celular del hospedante inhibe la amplificación de PCR. En este estudio, se efectuaron protocolos de PCR de alta fidelidad y PCR convencional en tubérculos con síntomas de ZC. Se usaron tubérculos asintomáticos como testigos. La PCR de alta fidelidad aumentó la detección de Lso de tubérculos con síntomas en un 30–40 % sobre la PCR convencional. Se hicieron ligeras modificaciones de protocolos previos de alta fidelidad para disminuir el tiempo de prueba y simplificar el protocolo.
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Cating, R. A., Rondon, S. I., and Hamm, P. B. 2014. High-fidelity PCR improves the detection of ‘Candidatus Liberibacter solanacearum’ in potato tubers. American Journal of Potato Research ##:##-##.
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Cating, R.A., Rondon, S.I. & Hamm, P.B. High-Fidelity PCR Improves the Detection of ‘Candidatus Liberibacter solanacearum’ in Potato Tubers. Am. J. Potato Res. 92, 451–454 (2015). https://doi.org/10.1007/s12230-015-9434-z
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DOI: https://doi.org/10.1007/s12230-015-9434-z