scholarly journals Plasmodiophora brassicae in its environment-effects of temperature and light on resting spore survival in soil.

2021 ◽  
Author(s):  
Kher Zahr ◽  
Alian Sarkes ◽  
Yalong Yang ◽  
Hafiz Ahmed ◽  
Qixing Zhou ◽  
...  
Keyword(s):  
Plasmodiophora Brassicae ◽  
Uv Light ◽  
Quantitative Polymerase Chain Reaction ◽  
Soil Surface ◽  
Spore Viability ◽  
Resting Spore ◽  
Infected Root ◽  
Resting Spores ◽  
Negative Effect ◽  
C 30

Clubroot caused by Plasmodiophora brassicae is an important disease on cruciferous crops worldwide. Management of clubroot is challenging, largely due to the millions of resting spores produced within an infected root that can survive dormant in the soil for many years. This study was conducted to investigate some of the environmental conditions that may affect the survival of resting spores in the soil. Soil samples containing clubroot resting spores (1 × 107 spores/g soil) were stored at various temperatures for two years. Additionally, other samples were buried in soil, or kept on the soil surface in the field. The content of P. brassicae DNA and the numbers of viable spores in the samples were assessed by quantitative polymerase chain reaction (qPCR) and pathogenicity bioassays, respectively. The results indicated that 4°C, 20°C, and being buried in the soil were more conductive conditions for spore survival compared to -20°C, 30°C, and at the soil surface. 99.99% of the spores kept on the soil surface were non-viable, suggesting a negative effect of light on spore viability. Additional experiments confirmed the negative effect of UV light on spore viability as spores receiving 2- and 3-hour UV light exhibited lower disease potential and contained less DNA content compared to the untreated control. Finally, this work confirmed that DNA-based quantification methods such as qPCR can be poor predictors of P. brassicae disease potential due to the presence and persistence of DNA from dead spores.

scholarly journals Plasmodiophora brassicae in its environment-effects of temperature and light on resting spore survival in soil

2019 ◽  
Author(s):  
Kher Zahr ◽  
Alian Sarkes ◽  
Yalong Yang ◽  
Qixing Zhou ◽  
David Feindel ◽  
...  
Keyword(s):  
Plasmodiophora Brassicae ◽  
Uv Light ◽  
Quantitative Polymerase Chain Reaction ◽  
Soil Surface ◽  
Spore Viability ◽  
Resting Spore ◽  
Infected Root ◽  
Resting Spores ◽  
Negative Effect ◽  
C 30

AbstractClubroot caused by Plasmodiophora brassicae is an important disease on cruciferous crops worldwide. Management of clubroot has been challenging, due largely to the millions of resting spores produced within an infected root that can survive dormant in the soil for many years. This study was conducted to investigate some of the environmental conditions that may affect the survival of resting spores in the soil. Soil samples containing clubroot resting spores (1 × 107 spores g-1 soil) were stored at various temperatures for two years. Additionally, other samples were buried in soil, or kept on the soil surface in the field. The content of P. brassicae DNA and the numbers of viable spores in the samples were assessed by quantitative polymerase chain reaction (qPCR) and pathogenicity bioassays, respectively. The results indicated that 4°C, 20°C and being buried in the soil were better conditions for spore survival than were −20°C, 30°C and at the soil surface. Most of the spores kept on the soil surface were killed, suggesting the negative effect of light on spore viability. Additional experiments confirmed that ultraviolet (UV) light contributed a large negative effect on spore viability as lower pathogenicity and less DNA content were observed from the 2-and 3-hour UV light treated spores compared to the untreated control. Finally, this work demonstrated that DNA-based quantification methods such as qPCR can be poor predictors of P. brassicae disease potential due to the presence and persistence of DNA from dead spores.

scholarly journals Propidium Monoazide Improves Quantification of Resting Spores of Plasmodiophora brassicae with qPCR

Plant Disease ◽  
2017 ◽  
Vol 101 (3) ◽  
pp. 442-447 ◽  
Author(s):  
Fadi Al-Daoud ◽  
Bruce D. Gossen ◽  
Justin Robson ◽  
Mary Ruth McDonald
Keyword(s):  
Heat Treatment ◽  
Plasmodiophora Brassicae ◽  
Quantitative Polymerase Chain Reaction ◽  
Infested Soil ◽  
Propidium Monoazide ◽  
Spore Viability ◽  
Heat Treated ◽  
Resting Spores ◽  
Polymerase Chain ◽  
The Impact

Plasmodiophora brassicae, which causes clubroot of Brassica crops, persists in soil as long-lived resting spores. Quantitative polymerase chain reaction (qPCR) analysis is often used to quantify resting spores but does not distinguish between DNA of viable and nonviable spores. The impact of pretreating spores with propidium monoazide (PMA), which inhibits amplification of DNA from nonviable microorganisms, was assessed in several experiments. Spore suspensions from immature and mature clubs were heat treated; then, PMA-PCR analyses and bioassays were performed to assess spore viability. Prior to heat treatment, assessments comparing PMA-PCR to qPCR for mature spores were similar, indicating that most of these spores were viable. However, only a small proportion (<26%) of immature spores were amplified in PMA-PCR. Bioassays demonstrated that clubroot severity was much higher in plants inoculated with mature spores than with immature spores. Heat treatment produced little or no change in estimates of mature spores from qPCR but spore estimates from PMA-PCR and clubroot severity in bioassays were both substantially reduced. Estimates of spore concentration with PMA-PCR were less consistent for immature spores. To facilitate use of PMA-PCR on infested soil, a protocol for extracting spores from soil was developed that provided higher extraction efficiency than the standard methods.

scholarly journals An Improved Evans Blue Staining Method for Consistent, Accurate Assessment of Plasmodiophora brassicae Resting Spore Viability

Plant Disease ◽  
2019 ◽  
Vol 103 (9) ◽  
pp. 2330-2336 ◽  
Author(s):  
M. W. Harding ◽  
T. B. Hill ◽  
Y. Yang ◽  
G. C. Daniels ◽  
S. F. Hwang ◽  
...  
Keyword(s):  
Evans Blue ◽  
Staining Method ◽  
Plasmodiophora Brassicae ◽  
Blue Staining ◽  
Inoculum Potential ◽  
Spore Viability ◽  
Modified Method ◽  
Resting Spore ◽  
Resting Spores ◽  
Vital Stains

Clubroot caused by Plasmodiophora brassicae is an important disease of brassica crops. The use of vital stains to determine the viability of P. brassicae resting spores can provide useful information regarding spore longevity, inoculum potential, or the efficacy of antimicrobial treatments. Evans blue is one example of a vital stain that has been reported to differentially stain viable and nonviable resting spores. Some previously published protocols using Evans blue to stain P. brassicae resting spores have not provided accurate or consistent results. In this study, we modified the Evans blue method by increasing the staining time to 8 h or more and evaluated P. brassicae resting spores after heat treatment at various combinations of temperature and time. Extending staining times significantly increased the numbers of stained resting spores up to 7 h, after which the numbers of stained spores did not change significantly (R2 = 96.88; P ≤ 0.001). The accuracy of the modified method to discriminate viable and nonviable spores was evaluated in repeated experiments and by comparing the staining data with those derived from inoculation assays and propidium monoazide quantitative PCR (qPCR). The results demonstrated that the modified Evans blue staining method improved the accuracy and consistency of measurement of P. brassicae resting spore viability. Additionally, it was equivalent to the qPCR method for differentiating viable and nonviable spores (R2 = 99.84; P ≤ 0.001) and confirmed in canola infection bioassays.

scholarly journals A Multiplex qPCR Assay for Detection and Quantification of Plasmodiophora brassicae in Soil

Plant Disease ◽  
2015 ◽  
Vol 99 (7) ◽  
pp. 1002-1009 ◽  
Author(s):  
Abhinandan Deora ◽  
Bruce D. Gossen ◽  
Sasan Amirsadeghi ◽  
Mary Ruth McDonald
Keyword(s):  
Plant Pathogens ◽  
Fluorescent Protein ◽  
Plasmodiophora Brassicae ◽  
Quantitative Polymerase Chain Reaction ◽  
Mineral Soil ◽  
Qpcr Assay ◽  
Spore Concentration ◽  
Resting Spore ◽  
Resting Spores ◽  
Detection And Quantification

Various physical and chemical factors in soil can inhibit the detection and quantification of soilborne plant pathogens using quantitative polymerase chain reaction (qPCR) assays. A multiplexed TaqMan qPCR assay, including a competitive internal positive control (CIPC), was developed to identify and (where necessary) compensate for inhibition in the quantification of resting spores of Plasmodiophora brassicae from soil. The CIPC amplicon was developed by modifying a sequence coding for green fluorescent protein so that it could be amplified with P. brassicae-specific primers. Addition of CIPC at 5 fg/μl to the singleplex qPCR assay designed to quantify P. brassicae genomic DNA did not reduce the sensitivity, specificity, or reproducibility of the assay. Each of the soil samples, either artificially inoculated or naturally infested with P. brassicae, exhibited no amplification of the CIPC. When the samples were diluted and reassessed, the quantification cycle of the CIPC relative to the control (water only) was delayed in each sample. The magnitude of the delay was used to adjust the estimate of resting spore concentration. The corrected concentration estimates were significantly higher than the unadjusted estimate, which indicated the presence of DNA inhibitors in samples even after dilution. The only exception was a mineral soil sample inoculated with a low concentration (103 spores/g) of resting spores. The assay was optimized for use on a range of soil types. A sample of 0.25 g for mineral soil and 0.10 g for high-organic-matter soil was optimum for recovery of DNA of P. brassicae. The assay represents an improvement over existing assays for estimating resting spore concentration in infested fields.

The life history of Plasmodiophora brassicae Woron

1972 ◽  
Vol 180 (1058) ◽  
pp. 103-112 ◽  
Keyword(s):  
Life History ◽  
Root Hair ◽  
Plasmodiophora Brassicae ◽  
Root Hairs ◽  
Secondary Phase ◽  
Tissue Cultures ◽  
Resting Spore ◽  
Resting Spores ◽  
Seedling Roots ◽  
History Of

Resting spore germination and the root hair stages of the life history of Plasmodiophora brassicae were studied in stained preparations of infected Brassica rapa seedling roots. Naked protoplasts, usually possessing two unequal flagella, were released from resting spores through a small circular pore. They penetrated the root hairs of B. rapa and there developed into plasmodia which, after becoming multinucleate, cleaved to form zoosporangia con­taining incipient zoospores. Biflagellate zoospores were released from root hair zoosporangia and fused in pairs, although karyogamy did not occur. The resulting binucleate zoospores infected the cortical dells of B. rapa to form binucleate plasmodia, the earliest stages of the secondary phase of the life history. These findings are discussed in relation to previous studies on the life history of P. brassicae in Brassica plants and in Brassica tissue cultures, and a new complete life history, including nuclear fusion in the secondary plasmodium, is suggested for the organism.

Detecting the titer in forest soils of spores of the gypsy moth (Lepidoptera: Lymantriidae) fungal pathogen, Entomophaga maimaiga (Zygomycetes: Entomophthorales)

2002 ◽  
Vol 134 (2) ◽  
pp. 269-279 ◽  
Author(s):  
Ronald M. Weseloh ◽  
Theodore G. Andreadis
Keyword(s):  
Forest Soils ◽  
Gypsy Moth ◽  
Fungal Pathogen ◽  
Spore Viability ◽  
Resting Spore ◽  
Entomophaga Maimaiga ◽  
Resting Spores ◽  
A Site ◽  
Spore Load

AbstractBioassays and direct counts were used to assess the abundance of resting spores of the gypsy moth, Lymantria dispar (L.), fungal pathogen, Entomophaga maimaiga Humber, Shimazu and Soper in forest soils. Resting spores in soil collected in October, January, and March and held under refrigeration germinated as readily as spores collected in April, but those collected in April germinated faster. Bioassays of resting spores in soils from different sites in Connecticut were directly related to results obtained from physically counting spores in the soil, and weakly correlated with a previously developed forest-based bioassay. The number of resting spores in a site was inversely related to the number of years since the site had last been defoliated by the gypsy moth, resulting in an implied maximum viability of resting spores of about 10 years. This maximum longevity was similar to a direct measure of long-term resting-spore viability. The study implies that resting-spore load in the soil may be an important determinant of the ability of the pathogen to control the gypsy moth.

scholarly journals Greenhouse Evaluation of Clubroot Resistant-Brassica napus cv. Mendel and Its Efficacy Concerning Virulence and Soil Inoculum Levels of Plasmodiophora brassicae

Pathogens ◽  
2021 ◽  
Vol 10 (2) ◽  
pp. 151 ◽  
Author(s):  
Nazanin Zamani-Noor ◽  
Imke Krohne ◽  
Birger Koopmann
Keyword(s):  
Brassica Napus ◽  
Plasmodiophora Brassicae ◽  
Root Weight ◽  
Susceptible Cultivar ◽  
Clubroot Resistance ◽  
Soil Inoculum ◽  
Resting Spore ◽  
Greenhouse Evaluation ◽  
Major Resistance Gene ◽  
Resting Spores

Clubroot resistance of oilseed rape (OSR) cultivars frequently relies on a major resistance gene originating from cv. Mendel. The efficacy of this resistance was studied in greenhouse experiments using two Plasmodiophora brassicae isolates, which were either virulent (P1(+)) or avirulent (P1) on Mendel. Seeds of clubroot-susceptible cultivar Visby and clubroot-resistant cultivar Mendel were sown in soil mixtures inoculated with different concentrations of resting spores (101, 103, 105, and 107 resting spores/g soil). Clubroot severity, plant height, shoot and root weight as well as resting spore propagation were assessed for each isolate and cultivar separately at four dates after sowing. The OSR cultivars behaved significantly different in the measured parameters. The threshold of inoculum density to cause disease depended strongly on the virulence of the pathogen and susceptibility of the host plant. In Visby grown in soil infested with P1, clubroot symptoms and increases in root weight and the number of propagated resting spores occurred at inoculum levels of 101 resting spores and higher, whereas Mendel was not affected in soils under the three lowest inoculum densities. In contrast, the P1(+) isolate led to earlier and more severe symptoms, heavier galls, and a significantly higher number of new resting spores in both cultivars.

Hard to kill: Inactivation of Plasmodiophora brassicae resting spores using chemical disinfectants

Plant Disease ◽  
2021 ◽  
Author(s):  
T. Blake Hill ◽  
Greg C Daniels ◽  
Jie Feng ◽  
Michael Wayne Harding
Keyword(s):  
Sodium Hypochlorite ◽  
Time Course ◽  
Plasmodiophora Brassicae ◽  
Blue Staining ◽  
Resting Spore ◽  
Resting Spores ◽  
Time Course Experiment ◽  
High Concentrations ◽  
Management Recommendations ◽  
Very High

Biosafety practices, such as bioexclusion via sanitization, can prevent the spread of infectious soilborne threats such as the clubroot pathogen, Plasmodiophora brassicae. Twenty three chemical disinfectants were evaluated for efficacy against P. brassicae resting spores. Evans blue staining was used to directly measure the viability of P. brassicae resting spores after 20 min exposures to ten concentrations of each of the 23 chemical disinfectants. Only nine disinfectants were capable of greater than 95% inactivation, and only five were capable of inactivating >99% of resting spores. Bleach (sodium hypochlorite) and Spray Nine® were the most effective disinfectants for inactivation of clubroot resting spores. AES 2500, SaniDate® and ethanol also inactivated >99% of resting spores, but only at very high concentrations. A time course experiment showed that 10 to 12 min contact time was sufficient for ≥ 95% resting spore inactivation with Spray Nine® and sodium hypochlorite, but ≥ 30 min contact was required for other disinfectants evaluated. These results will assist in guiding management recommendations for sanitization aimed at bioexclusion and biocontainment of P. brassicae.

Sign in / Sign up

Export Citation Format

Share Document