Distribution and diversity of root-lession nematode (Pratylenchus spp.) associated with Miscanthus × giganteus and Panicum virgatum used for biofuels, and species identification in a multiplex polymerase chain reaction

Tesfamariam Mekete, Kimberly Reynolds, Horacio D. Lopez-Nicora, Michael E. Gray, Terry L. Niblack

Research output: Contribution to journalArticlepeer-review

Abstract

The distribution of Pratylenchus spp. from bioenergy field plots in six states (Illinois, Iowa, South Dakota, Kentucky, Tennessee, Georgia) of the USA were surveyed. The species were identified based on morphology and morphometrics and further characterised based on fragment sequences of the 28S rRNA of the D2-D3 region. The region revealed variations in sequencing information that supports the morphological identification. In this work, six Pratylenchus spp. were detected: Pratylenchus brachyurus, P. crenatus, P. hexincisus, P. neglectus, P. penetrans and P. scribneri. Pratylenchus scribneri, P. crenatus, and P. penetrans were distributed most widely, with detection of 34, 29 and 15%, respectively. Pratylenchus hexincisus, P. brachyurus and P. neglectus were distributed sporadically, with detection rates of 10.0, 2.6 and 2.0%, respectively. A one-step multiplex PCR was developed for the simultaneous detection of P. scribneri, P. crenatus and P. penetrans. Sequence data from this research and NCBI were used to generate different primer sets that are species-specific. We have therefore designed three sets of primers that discriminate P. scribneri, P. crenatus and P. penetrans in multiplex PCR. All the tested primers have shown specificity and have no cross-reaction with the non-target species. When used in a uniplex, duplex and triplex PCR, the selected three primers gave a unique electrophoretic DNA banding pattern characterised by a single DNA fragment for P. scribneri (ca 750), P. crenatus (ca 690), and P. penetrans (ca 520). The method could be used for routine diagnostic programmes.

Original languageEnglish (US)
Pages (from-to)673-686
Number of pages14
JournalNematology
Volume13
Issue number6
DOIs
StatePublished - 2011

Keywords

  • bioenergy
  • diagnostics
  • duplex
  • multiplex
  • rDNA D2-D3
  • triplex
  • uniplex

ASJC Scopus subject areas

  • Ecology, Evolution, Behavior and Systematics
  • Agronomy and Crop Science

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