The MeloTuber Test: a real-time TaqMan® PCR-based assay to detect the root-knot nematodes Meloidogyne chitwoodi and M. fallax directly in potato tubers

EPPO Bulletin ◽  
2014 ◽  
Vol 44 (2) ◽  
pp. 166-175 ◽  
Author(s):  
E. G. de Haan ◽  
C. C. E. M. Dekker ◽  
W. I. L. Tameling ◽  
L. J. M. F. den Nijs ◽  
G. W. van den Bovenkamp ◽  
...  
Keyword(s):  
Real Time ◽  
Potato Tubers ◽  
Meloidogyne Chitwoodi ◽  
Root Knot Nematodes

Population fluctuations of root-knot nematodes Meloidogyne chitwoodi and M. hapla under field conditions

2021 ◽  
Author(s):  
Emre Evlice ◽  
Halil Toktay ◽  
Gökhan Yatkın ◽  
Fatma Dolunay Erdoğuş ◽  
Mustafa İmren
Keyword(s):  
Field Conditions ◽  
Population Fluctuations ◽  
Meloidogyne Chitwoodi ◽  
Root Knot Nematodes

scholarly journals Using the TxtAB Operon to Quantify Pathogenic Streptomyces in Potato Tubers and Soil

2008 ◽  
Vol 98 (4) ◽  
pp. 405-412 ◽  
Author(s):  
Xinshun Qu ◽  
Leslie A. Wanner ◽  
Barbara J. Christ
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Pathogenic Bacteria ◽  
Common Scab ◽  
Cost Effective ◽  
Potato Tubers ◽  
Pcr Assay ◽  
Streptomyces Species ◽  
Standard Curve ◽  
Target Dna

The phytotoxin thaxtomin, produced by plant pathogenic Streptomyces species, is the only known pathogenicity determinant for common scab diseases of potato and other root and tuber crops. Genes encoding thaxtomin synthetase (txtAB) are found on a pathogenicity island characteristic of genetically diverse plant pathogenic Streptomyces species. In this study, an SYBR Green quantitative real-time polymerase chain reaction (PCR) assay using primers designed to anneal to the txtAB operon of Streptomyces was developed to quantify pathogenic bacterial populations in potatoes and soil. The real-time PCR assay was specific for pathogenic Streptomyces strains. The detection limit of the assay was 10 fg of the target DNA, or one genome equivalent. Cycle threshold (Ct) values were linearly correlated with the concentration of the target DNA (correlation coefficient R2 = 0.99) and were not affected by the presence of plant DNA extracts, indicating the usefulness of the assay for quantitative analyses of the pathogenic bacteria in plant tissues. The amount of pathogenic Streptomyces DNA in total DNA extracts from 1 g asymptomatic and symptomatic tubers was quantified using the assay and ranged from 101 to 106 pg. A standard curve was established to quantify pathogenic Streptomyces in soil. Using the standard curve, numbers of pathogenic Streptomyces colony forming units were extrapolated to range from 103 to 106 per gram of soil from potato fields where common scab was found. This real-time PCR assay using primers designed from the txtAB operon allows rapid, accurate, and cost effective quantification of pathogenic Streptomyces strains in potato tubers and in soil.

scholarly journals A Multiplex Real-Time Polymerase Chain Reaction (TaqMan) Assay for the Simultaneous Detection of Meloidogyne chitwoodi and M. fallax

2006 ◽  
Vol 96 (11) ◽  
pp. 1255-1262 ◽  
Author(s):  
C. Zijlstra ◽  
R. A. Van Hoof
Keyword(s):  
Polymerase Chain Reaction ◽  
Real Time ◽  
Real Time Pcr ◽  
False Negative ◽  
Simultaneous Detection ◽  
Chain Reaction ◽  
Meloidogyne Chitwoodi ◽  
Negative Results ◽  
False Negative Results ◽  
Polymerase Chain

This study describes a multiplex real-time polymerase chain reaction (PCR) approach for the simultaneous detection of Meloidogyne chitwoodi and M. fallax in a single assay. The approach uses three fluorogenic minor groove binding (MGB) TaqMan probes: one FAM-labeled to detect M. chitwoodi, one VIC-labeled to detect M. fallax, and one NED-labeled to detect the internal amplification control (IAC) to monitor false negative results. One common primer set is used for the amplification of part of the internal transcribed spacer (ITS) region of M. chitwoodi and M. fallax and one primer set for the amplification of the IAC. The test enabled detection of M. chitwoodi and/or M. fallax in DNA samples extracted from batches of juveniles, from single juveniles, and from infected plant material. Compared with current assays to detect M. chitwoodi and M. fallax, the multiplex real-time PCR offers the following advantages: it is faster because the test can simultaneously detect both quarantine species without the need for post-PCR processing; and it is at least 10 times more sensitive than a comparable regular PCR also targeting the ITS sequence. Inclusion of the IAC facilitates the interpretation of the FAM and VIC cycle threshold (Ct) values and can prevent the scoring of false negative results when FAM, VIC, and NED Ct values are high. The test allows precise quantification when only one of the two species is present in the sample. However, experiments with mixtures of genomic DNA of M. chitwoodi and M. fallax revealed that the ability of the multiplex real-time PCR assay to detect small quantities of DNA of one species is reduced when large quantities of DNA of the other species are present.

scholarly journals Detection of Root-Knot Nematode Meloidogyne luci Infestation of Potato Tubers Using Hyperspectral Remote Sensing and Real-Time PCR Molecular Methods

2021 ◽  
Vol 13 (10) ◽  
pp. 1996
Author(s):  
Uroš Žibrat ◽  
Barbara Gerič Stare ◽  
Matej Knapič ◽  
Nik Susič ◽  
Janez Lapajne ◽  
...  
Keyword(s):  
Real Time ◽  
Hyperspectral Imaging ◽  
Real Time Pcr ◽  
Detection Methods ◽  
Parasitic Nematodes ◽  
Tuber Quality ◽  
Limiting Factor ◽  
Potato Tubers ◽  
Root Knot Nematode ◽  
Efficient Detection

Root-knot nematodes (Meloidogyne spp.) are considered the most aggressive, damaging, and economically important group of plant-parasitic nematodes and represent a significant limiting factor for potato (Solanum tuberosum) production and tuber quality. Meloidogyne luci has previously been shown to be a potato pest having significant reproductive potential on the potato. In this study we showed that M. luci may develop a latent infestation without visible symptoms on the tubers. This latent infestation may pose a high risk for uncontrolled spread of the pest, especially via seed potato. We developed efficient detection methods to prevent uncontrolled spread of M. luci via infested potato tubers. Using hyperspectral imaging and a molecular approach to detection of nematode DNA with real-time PCR, it was possible to detect M. luci in both heavily infested potato tubers and tubers without visible symptoms. Detection of infested tubers with hyperspectral imaging achieved a 100% success rate, regardless of tuber preparation. The real-time PCR approach detected M. luci with high sensitivity.

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