Quantitative detection of Lactarius deliciosus extraradical soil mycelium by real-time PCR and its application in the study of fungal persistence and interspecific competition

Parladé, Javier; Hortal, Sara; Pera, Joan; Galipienso, Luis

doi:10.1016/j.jbiotec.2006.09.010

Cited by 57 publications

(35 citation statements)

References 31 publications

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“…Real-time PCR provides useful tools for studying and quantifying mycorrhizal fungus interactions and biomass (33,47,53,59). Unlike workers in previous studies, we did not use the ITS rRNA region to determine the number of Wilcoxina molecules.…”

Section: Discussionmentioning

confidence: 99%

Impact of Endochitinase-Transformed White Spruce on Soil Fungal Biomass and Ectendomycorrhizal Symbiosis

Stefani

Tanguay

Pelletier

et al. 2010

Appl Environ Microbiol

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The impact of transgenic white spruce [Picea glauca (Moench) Voss] containing the endochitinase gene (ech42) on soil fungal biomass and on the ectendomycorrhizal fungi Wilcoxina spp. was tested using a greenhouse trial. The measured level of endochitinase in roots of transgenic white spruce was up to 10 times higher than that in roots of nontransformed white spruce. The level of endochitinase in root exudates of three of four ech42-transformed lines was significantly greater than that in controls. Analysis soil ergosterol showed that the amount of fungal biomass in soil samples from control white spruce was slightly larger than that in soil samples from ech42-transformed white spruce. Nevertheless, the difference was not statistically significant. The rates of mycorrhizal colonization of transformed lines and controls were similar. Sequencing the internal transcribed spacer rRNA region revealed that the root tips were colonized by the ectendomycorrhizal fungi Wilcoxina spp. and the dark septate endophyte Phialocephala fortinii. Colonization of root tips by Wilcoxina spp. was monitored by real-time PCR to quantify the fungus present during the development of ectendomycorrhizal symbiosis in ech42-transformed and control lines. The numbers of Wilcoxina molecules in the transformed lines and the controls were not significantly different (P > 0.05, as determined by analysis of covariance), indicating that in spite of higher levels of endochitinase expression, mycorrhization was not inhibited. Our results indicate that the higher levels of chitinolytic activity in root exudates and root tissues from ech42-transformed lines did not alter the soil fungal biomass or the development of ectendomycorrhizal symbiosis involving Wilcoxina spp.

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Section: Discussionmentioning

confidence: 99%

Impact of Endochitinase-Transformed White Spruce on Soil Fungal Biomass and Ectendomycorrhizal Symbiosis

Stefani

Tanguay

Pelletier

et al. 2010

Appl Environ Microbiol

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“…Only amplification efficiencies ranging from 95% to 105% were accepted and considered for analysis. qPCRs, processing of data, and quantification of L. vinosus DNA were performed following the methods described in a previous study (11).…”

Section: Methodsmentioning

confidence: 99%

“…In this context, new techniques that do not require the collection of mushrooms can improve the estimation of mushroom yields. Except for a few cases (7), no correlations have been found between the belowground mycelial biomass of specific species and their mushroom production (8)(9)(10), even though positive correlations have been found between root mycorrhizal colonization and soil mycelia (11). In this regard, new monitoring approaches using fungal spores, such as spore trapping, may represent a new way of addressing this challenge.…”

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confidence: 99%

Mushroom Emergence Detected by Combining Spore Trapping with Molecular Techniques

Castaño

Oliva

Aragón

et al. 2017

Appl Environ Microbiol

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Obtaining reliable and representative mushroom production data requires time-consuming sampling schemes. In this paper, we assessed a simple methodology to detect mushroom emergence by trapping the fungal spores of the fruiting body community in plots where mushroom production was determined weekly. We compared the performance of filter paper traps with that of funnel traps and combined these spore trapping methods with species-specific quantitative real-time PCR and Illumina MiSeq to determine the spore abundance. Significantly more MiSeq proportional reads were generated for both ectomycorrhizal and saprotrophic fungal species using filter traps than were obtained using funnel traps. The spores of 37 fungal species that produced fruiting bodies in the study plots were identified. Spore community composition changed considerably over time due to the emergence of ephemeral fruiting bodies and rapid spore deposition (lasting from 1 to 2 weeks), which occurred in the absence of rainfall events. For many species, the emergence of epigeous fruiting bodies was followed by a peak in the relative abundance of their airborne spores. There were significant positive relationships between fruiting body yields and spore abundance in time for five of seven fungal species. There was no relationship between fruiting body yields and their spore abundance at plot level, indicating that some of the spores captured in each plot were arriving from the surrounding areas. Differences in fungal detection capacity by spore trapping may indicate different dispersal ability between fungal species. Further research can help to identify the spore rain patterns for most common fungal species.IMPORTANCE Mushroom monitoring represents a serious challenge in economic and logistical terms because sampling approaches demand extensive field work at both the spatial and temporal scales. In addition, the identification of fungal taxa depends on the expertise of experienced fungal taxonomists. Similarly, the study of fungal dispersal has been constrained by technological limitations, especially because the morphological identification of spores is a challenging and time-consuming task. Here, we demonstrate that spores from ectomycorrhizal and saprotrophic fungal species can be identified using simple spore traps together with either MiSeq fungus-specific amplicon sequencing or species-specific quantitative real-time PCR. In addition, the proposed methodology can be used to characterize the airborne fungal community and to detect mushroom emergence in forest ecosystems.

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“…The quantification of the target amount in unknown samples can be calculated by generating a standard curve (Song et al, 2002). Over the past ten years, at least five papers have reported the detection, distribution and abundance of a specific ectomycorrhizal fungus through qPCR in the soil (Guidot et al, 2002, Landeweert et al, 2003, Parladé et al, 2007, Suz et al, 2008and De la Varga et al, 2012. Suz et al (2008) attempted to correlate mycelia abundance and other factors related to truffle productivity in Tuber melanosporum-Quercus ilex orchards.…”

Section: Introductionmentioning

confidence: 99%

The detection of mating type genes of Tuber melanosporum in productive and non productive soils

Zampieri

Rizzello

Bonfante

et al. 2012

Applied Soil Ecology

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Truffles are hypogeous ectomycorrhizal fungi. Of all the different species, Tuber melanosporum is one of the most popular on the truffle market. The aim of this work was to set up a protocol in order to check the fertility of a T. melanosporum ground. The correlation between its abundance in soil, the presence of mating type genes and productivity was investigated. Soil sampling was conducted in a truffle-ground over two periods of the T. melanosporum life cycle, and under two different host species, to verify whether the time and plant species can affect the quantity of mycelium in the soil. An effective quantitative PCR protocol was set up and employed to the investigated truffle-orchard.We found a statistically significant difference in T. melanosporum abundance between the productive and unproductive soils collected in April. Mating type genes for T. melanosporum were detected under productive and formally productive trees and generally not under unproductive trees even though T. melanosporum was detected. In all the three situations the mating type genes were detected when more than 0.3 ng of T. melanosporum DNA was present. Our results suggest combining these approaches to increase knowledge on the fertility of truffle orchards. Highlights► We present a protocol in order to check the possibility of a ground to produce T. melanosporum. ► Soil samples from a truffle-ground were investigated. ► A quantitative real time PCR was set up to quantify mycelium in the soil. ► The mating type genes were identified when more than 0.3 ng of T. melanosporum was present. ► The correlation between T. melanosporum abundance and productivity is also discussed.

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Quantitative detection of Lactarius deliciosus extraradical soil mycelium by real-time PCR and its application in the study of fungal persistence and interspecific competition

Cited by 57 publications

References 31 publications

Impact of Endochitinase-Transformed White Spruce on Soil Fungal Biomass and Ectendomycorrhizal Symbiosis

Impact of Endochitinase-Transformed White Spruce on Soil Fungal Biomass and Ectendomycorrhizal Symbiosis

Mushroom Emergence Detected by Combining Spore Trapping with Molecular Techniques

The detection of mating type genes of Tuber melanosporum in productive and non productive soils

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