A novel nematode diagnostic method using the direct quantification of major plant-parasitic nematodes in soil by real-time PCR

Nematology ◽  
2012 ◽  
Vol 14 (3) ◽  
pp. 265-276 ◽  
Author(s):  
Yu Yu Min ◽  
Koki Toyota ◽  
Erika Sato
Keyword(s):  
Real Time ◽  
Sweet Potato ◽  
Real Time Pcr ◽  
Extraction Method ◽  
Cyst Nematode ◽  
Parasitic Nematodes ◽  
Plant Parasitic Nematodes ◽  
Pcr Method ◽  
Plant Parasitic ◽  
Direct Quantification

We have developed a direct quantification method using real-time PCR for various plant-parasitic nematodes. Firstly, specific primers were designed for the root-knot nematode Meloidogyne incognita, the root-lesion nematode Pratylenchus penetrans, the potato cyst nematode Globodera rostochiensis and the soybean cyst nematode Heterodera glycines. A DNA extraction method was then developed beginning with 20 g of soil, a relatively large amount of soil but a necessary amount in the consideration of heterogeneous distribution of nematodes in soil. To estimate the density of the target nematode in soil, calibration curves for each plant-parasitic nematode were obtained by inoculating different numbers of the target nematode and then extracting DNA from the soils. The detection limit was 4-5 nematodes (20 g soil)−1. This method was applied to nematode diagnostics. Soil sampling was done when transplanting of radish and sweet potato in fields was taking place, and the density of plant-parasitic nematodes was measured using both the Baermann funnel extraction method and real-time PCR methods. In some soils, P. penetrans and M. incognita were not found with the Baermann method but were detected with the real-time PCR method. At harvest, damage to crops was evaluated and its relationship with initial densities was investigated. The real-time PCR method more precisely predicted damage to radish and sweet potato by nematodes and was considered to be a powerful tool in the diagnosis of nematode diseases.

The use of Pluronic F-127 to study the development of the potato cyst nematode, Globodera pallida

Nematology ◽  
2012 ◽  
Vol 14 (7) ◽  
pp. 869-873 ◽  
Author(s):  
Ayano Sasaki-Crawley ◽  
Rosane Curtis ◽  
Michael Birkett ◽  
Apostolos Papadopoulos ◽  
Rod Blackshaw ◽  
...  
Keyword(s):  
Aqueous Solution ◽  
Solanum Tuberosum ◽  
Transgenic Plants ◽  
Cyst Nematode ◽  
Potato Cyst Nematode ◽  
Globodera Pallida ◽  
Parasitic Nematodes ◽  
Plant Parasitic Nematodes ◽  
Plant Parasitic

This paper demonstrates a simple novel in vitro method using Pluronic F-127 aqueous solution to study the development of the potato cyst nematode, Globodera pallida, in Solanum spp. without any need for sterilisation of either the plants or the nematodes. In this study, this method was successfully applied to comparative studies on the development of G. pallida in Solanum tuberosum (potato) or S. sisymbriifolium (sticky nightshade). The protocol described here could be useful for screening transgenic plants or different plant cultivars/species for their ability to allow development not only of G. pallida but also any other plant-parasitic nematodes.

scholarly journals Studies on plant-parasitic nematodes associated with sweet potato (Ipomoea batatas L., Lam.) in Gombe State, Nigeria

2021 ◽  
Vol 6 (4) ◽  
pp. 477-482
Author(s):  
Jidere Caleb Iliya ◽  
Simon Lilian Dada ◽  
Sulaiman Ibrahim ◽  
Abraham Peter
Keyword(s):  
Sweet Potato ◽  
Ipomoea Batatas ◽  
Parasitic Nematodes ◽  
Soil Core ◽  
Biotic Factors ◽  
Plant Parasitic Nematodes ◽  
High Similarity ◽  
Soil Extraction ◽  
Meloidogyne Spp ◽  
Plant Parasitic

Sweet potato (Ipomoea batatas L., Lam.) is one of the most frequently eaten food crops. Its production is affected by plant-parasitic nematodes as well as biotic factors. This study was conducted to document the different plant-parasitic nematodes (PPN) that limit the gainful production of sweet potato in Gombe State. Thirty soil core samples per hectare were collected at random from sweet potato farms in the three local government areas (Nafada, Kaltungo, and Yamaltu Deba) of Gombe state. The Whitehead and Hemming method and identification keys were used for the soil extraction and genera identification of the plant-parasitic nematodes respectively. A total of 15 plant-parasitic nematodes were recovered throughout the surveyed areas among which 7 are considered major nematode pests of global importance. Irrespective of the surveyed locations, Meloidogyne spp., was found to record the highest population density and prevalence value. The frequency of occurrence in Y/Deba and Nafada LGAs shows that Meloidogyne spp., wasthe most occurring (32 %) genera. In Kaltungo LGA however, Scutellonema spp., and Rotylenchusspp., were the most occurring (17 %) genera. There was a high similarity percentage (≥ 68 %) of PPN genera where 8 genera (Scutellonema spp., Nacobbus spp., Pratylenchus spp., Meloidogyne spp., Heterodera spp., Xiphinema spp., Trichodorus spp., and Rotylenchus spp.) were found to be common amongst the surveyed locations. This is the first report of plant-parasitic nematodes associated with sweet potato in Gombe state, Nigeria. Hence, it is critical to educate farmers in the regions about their effects on the crop and how to successfully manage them.

scholarly journals Identification and characterisation of serotonin signalling in the potato cyst nematode Globodera pallida reveals new targets for crop protection

2018 ◽  
Author(s):  
Anna Crisford ◽  
Fernando Calahorro ◽  
Elizabeth Ludlow ◽  
Jessica M.C. Marvin ◽  
Jennifer K. Hibbard ◽  
...  
Keyword(s):  
Biogenic Amines ◽  
Tryptophan Hydroxylase ◽  
Plant Root ◽  
Crop Protection ◽  
Cyst Nematode ◽  
Potato Cyst Nematode ◽  
Globodera Pallida ◽  
Parasitic Nematodes ◽  
Plant Parasitic Nematodes ◽  
Plant Parasitic

AbstractPlant parasitic nematodes are microscopic pests that invade plant roots and cause extensive damage to crops worldwide. To investigate mechanisms underpinning their parasitic behaviour we used a chemical biology approach: We discovered that reserpine, a plant alkaloid known for its antagonism of the mammalian vesicular monoamine transporter VMAT and ability to impart a global depletion of synaptic biogenic amines in the nervous system, potently impairs the ability of the potato cyst nematode Globodera pallida to enter the host plant root. We show that this effect of reserpine is mediated by an inhibition of serotonergic signalling that is essential for activation of the stylet, a lance-like organ that protrudes from the mouth of the worm and which is used to pierce the host root to gain access. Prompted by this we identified core molecular components of G. pallida serotonin signalling encompassing the target of reserpine, VMAT; the synthetic enzyme for serotonin, tryptophan hydroxylase; the G protein coupled receptor SER-7 and the serotonin-gated chloride channel MOD-1. We found that inhibitors of tryptophan hydroxylase, SER-7 and MOD-1 phenocopy the plant protecting action of reserpine. Thus targeting the serotonin signalling pathway presents a promising new route to control plant parasitic nematodes.SummaryIndian snakeroot, an herbal medicine prepared from the roots of the shrub Rauwolfia serpentina, has been used for centuries for its calming action. The major active constituent is reserpine which works by depleting a specific class of mood regulating chemical in the brain, the biogenic amines. We have discovered a remarkable effect of reserpine on a pest of global concern, the plant parasitic nematodes. These microscopic worms invade the roots of crops presenting a severe threat to food production. We show that reserpine disables serotonin signalling in the worm’s ‘brain’ that regulates the rhythmic thrusting of the stylet: a lance-like structure that protrudes from its mouth to pierce the plant root and which is essential to its parasitic lifecycle. Thus, reserpine joins nicotine as another intriguing example of Nature evolving its own protection against pests. We have identified key components of the serotonin signalling pathway in the potato cyst nematode Globodera pallida and show that chemicals that target these sites inhibit the ability of the nematode to invade its host plant. We conclude that biogenic amine transmitters are intimately involved in the worm’s parasitic behaviour and provide a new discrete route to crop protection.

The impact of management strategies on the development and status of potato cyst nematode populations in Switzerland: An overview from 1958 to present

Plant Disease ◽  
2021 ◽  
Author(s):  
Andrea Caroline Ruthes ◽  
Paul Dahlin
Keyword(s):  
Integrated Management ◽  
Abiotic Factors ◽  
Management Strategies ◽  
Monitoring Program ◽  
Cyst Nematode ◽  
Globodera Pallida ◽  
Parasitic Nematodes ◽  
Plant Parasitic Nematodes ◽  
The Impact ◽  
Plant Parasitic

Globodera rostochiensis and Globodera pallida are some of the most successful and highly specialized plant-parasitic nematodes, and appear among the most regulated quarantine pests globally. In Switzerland, they have been monitored by annual surveys since their first detection in Swiss soil, in 1958. The dataset created was reviewed to give an overview of the development and actual status of PCN in Switzerland. Positive fields represent 0.2% of all the samples analyzed, and currently their distribution is limited to central-west and western Switzerland, suggesting that new introduction of PCN and the spread of the initial introduced PCN populations did not occur. In this way, the integrated management used in Switzerland appears to be effective. However, the increasing availability of potato varieties with resistance to G. rostochiensis and the limited availability of varieties with resistance to G. pallida, together with other biotic and abiotic factors promoted changes in the dominance of either species. Consequently, an extended monitoring program would be of interest to Swiss farmers, to avoid favoring virulent traits that could be present within Swiss Globodera populations.

scholarly journals Functional Analysis of Pathogenicity Proteins of the Potato Cyst Nematode Globodera rostochiensis Using RNAi

2005 ◽  
Vol 18 (7) ◽  
pp. 621-625 ◽  
Author(s):  
Qing Chen ◽  
S. Rehman ◽  
G. Smant ◽  
John T. Jones
Keyword(s):  
Functional Analysis ◽  
Host Location ◽  
Globodera Rostochiensis ◽  
Cyst Nematode ◽  
Potato Cyst Nematode ◽  
Parasitic Nematodes ◽  
Plant Parasitic Nematodes ◽  
Modified Method ◽  
Plant Parasitic

RNA interference (RNAi) has been used widely as a tool for examining gene function and a method that allows its use with plant-parasitic nematodes recently has been described. Here, we use a modified method to analyze the function of secreted β-1,4, endoglucanases of the potato cyst nematode Globodera rostochiensis, the first in vivo functional analysis of a pathogenicity protein of a plant-parasitic nematode. Knockout of the β-1,4, endoglucanases reduced the ability of the nematodes to invade roots. We also use RNAi to show that gr-ams-1, a secreted protein of the main sense organs (the amphids), is essential for host location.

The added value of proficiency tests: choosing the proper method for extracting Meloidogyne second-stage juveniles from soil

Nematology ◽  
2013 ◽  
Vol 15 (2) ◽  
pp. 143-151 ◽  
Author(s):  
Loes den Nijs ◽  
Wim van den Berg
Keyword(s):  
Extraction Method ◽  
Proficiency Test ◽  
Performance Test ◽  
Ring Test ◽  
Parasitic Nematodes ◽  
Plant Parasitic Nematodes ◽  
Method Performance ◽  
Second Stage ◽  
Baermann Method ◽  
Plant Parasitic

To gain insight into the different extraction processes used in laboratories in various European countries and the effect these methods might have on the number of Meloidogyne second-stage juveniles (J2) extracted from soil samples, a ring test with 2 elements was conducted. The first element used the Baermann funnel as a method performance test. The second element was a proficiency test, in which the laboratory’s standard technique was compared with the Baermann method. Chilled samples of 100 cm3 (10 per extraction method) were sent to participants in insulated containers. In the nematode counts distinction was made between Meloidogyne J2, other plant-parasitic nematodes and/or saprophytes. A total number of 18 participants took part in the method performance test and 13 institutes took part in the proficiency test using 11 different extraction techniques. The initial population density in the soil was 2025 J2 (100 cm3 soil)−1, determined by Oostenbrink elutriator and 4 weeks incubation. In the method performance test the institutes found Meloidogyne J2 numbers varied between 0 and 705 J2 (100 cm3)−1. Using the Baermann extraction method the median number per institute ranged from 0 to 377 J2 (100 cm3)−1. In the proficiency test comparison amongst the different methods showed an increase of more than 100-fold, with the highest median of 3733 J2 (100 cm3)−1 and the lowest of 34 J2 (100 cm3)−1; the difference was caused mainly by the incubation effect. For the other plant-parasitic nematodes and the saprophytes, the differences between the most and least efficient extraction methods were much smaller. The variance of the Meloidogyne counts was the highest for the standard Baermann extraction technique; other similar techniques also had high variances. The automated zonal centrifuge had the lowest variance. The Baermann method is not advisable for survey purposes.

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