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Rapid detection of Pecan Root-Knot Nematode, Meloidogyne partityla, in laboratory and field conditions using loop-mediated isothermal amplification

Sumyya Waliullah, Jessica Bell, Tammy Stackhouse, Ganpati Jagdale, Abolfazl Hajihassani, Timothy Brenneman, View ORCID ProfileMd Emran Ali
doi: https://doi.org/10.1101/2020.01.09.900076
Sumyya Waliullah
1University of Georgia, Department of Plant Pathology, Tifton, GA 31793, USA
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Jessica Bell
1University of Georgia, Department of Plant Pathology, Tifton, GA 31793, USA
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Tammy Stackhouse
1University of Georgia, Department of Plant Pathology, Tifton, GA 31793, USA
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Ganpati Jagdale
2University of Georgia, Department of Plant Pathology, Athens, GA 30602, USA
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Abolfazl Hajihassani
1University of Georgia, Department of Plant Pathology, Tifton, GA 31793, USA
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Timothy Brenneman
1University of Georgia, Department of Plant Pathology, Tifton, GA 31793, USA
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Md Emran Ali
1University of Georgia, Department of Plant Pathology, Tifton, GA 31793, USA
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  • ORCID record for Md Emran Ali
  • For correspondence: emran.ali@uga.edu
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Abstract

Meloidogyne partityla is the dominant root-knot nematode (RKN) species parasitizing pecan in Georgia. This species is known to cause a reduction in root growth and a decline in yields from mature pecan trees. Rapid and accurate diagnosis of this RKN is required to control this nematode disease and reduce losses in pecan production. In this study, a loop-mediated isothermal amplification (LAMP) method was developed for simple, rapid and on-site detection of M. partityla in infested plant roots and validated to detect the nematode in laboratory and field conditions. Specific primers were designed based on the sequence distinction of internal transcribed spacer (ITS)-18S/5.8S ribosomal RNA gene between M. partityla and other Meloidogyne spp. The LAMP detection technique could detect the presence of M. partityla genomic DNA at a concentration as low as 1 pg, and no cross reactivity was found with DNA from other major RKN species such as M. javanica, M. incognita and M. arenaria, and M. hapla. We also conducted a traditional morphology-based diagnostic assay and conventional polymerase chain reaction (PCR) assay to determine which of these techniques was less time consuming, more sensitive, and convenient to use in the field. The LAMP assay provided more rapid results, amplifying the target nematode species in less than 60 min at 65°C, with results 100 times more sensitive than conventional PCR (~2-3 hrs). Morphology-based, traditional diagnosis was highly time-consuming (2 days) and more laborious than conventional PCR and LAMP assays. These features greatly simplified the operating procedure and made the assay a powerful tool for rapid, on-site detection of pecan RKN, M. partityla. The LAMP assay will facilitate accurate pecan nematode diagnosis in the field and contribute to the management of the pathogen.

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Posted January 09, 2020.
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Rapid detection of Pecan Root-Knot Nematode, Meloidogyne partityla, in laboratory and field conditions using loop-mediated isothermal amplification
Sumyya Waliullah, Jessica Bell, Tammy Stackhouse, Ganpati Jagdale, Abolfazl Hajihassani, Timothy Brenneman, Md Emran Ali
bioRxiv 2020.01.09.900076; doi: https://doi.org/10.1101/2020.01.09.900076
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Rapid detection of Pecan Root-Knot Nematode, Meloidogyne partityla, in laboratory and field conditions using loop-mediated isothermal amplification
Sumyya Waliullah, Jessica Bell, Tammy Stackhouse, Ganpati Jagdale, Abolfazl Hajihassani, Timothy Brenneman, Md Emran Ali
bioRxiv 2020.01.09.900076; doi: https://doi.org/10.1101/2020.01.09.900076

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