Pattern of Tuber melanosporum extramatrical mycelium expansion over a 20-year chronosequence in Quercus ilex-truffle orchards

Abstract

Successful cultivation of black truffle (Tuber melanosporum) requires a long-term investment and the maintenance of the symbiosis throughout its preproductive and productive years. Monitoring the symbiosis over time is challenging, as it requires methods that can detect the belowground proliferation of the fungus associated with its host tree. In this study, we used a chronosequence design to study the expansion pattern of this fungus as the host tree grows. We hypothesize that this expansion can be estimated by monitoring T. melanosporum DNA from soil beneath host trees of different ages (3, 5, 7, 10, 14, and 20 years old) and at different distances from the trunk of the trees (40, 100, and 200 cm). We also wished to evaluate the presences of Tuber brumale and Tuber indicum, potentially problematic truffle species, in these plantations. To detect the mycelium of T. melanosporum in these soils, we extracted DNA and performed polymerase chain reaction (PCR) with Tuber species-specific primers, and to estimate DNA amount, we measured relative band intensities from the amplicons in agarose gels. Both age and distance were related to T. melanosporum DNA quantity, which was more abundant in the oldest age classes, reaching a plateau in 5-7 years. At 40 cm from the tree, there were no differences in T. melanosporum DNA amounts in orchards of different ages, but at 100 and 200 cm, younger orchards had less T. melanosporum DNA. We did not detect DNA from T. brumale or T. indicum in any of our samples.

Shifts in soil fungal communities in Tuber melanosporum plantations over a 20-year transition from agriculture fields to oak woodlands

“…belonging to T. melanosporum and, with a fluctuation in the 10-year-old plantation, continued to increase through the 20-year plantation with 22 of 31 (71% of colonies) belonging to T. melanosporum, confirming its dominating capacity. This domination was also reflected in the relative quantities of T. melanosporum mycelium detected in soils from these plantations in a companion work by Liu et al (2014).…”

“…in collaboration with truffle farmers who have established successful black truffle plantations in the province of Teruel, (central eastern Spain). As reported in Liu et al (2014): Plantations are located in the valley between the ranges of Gudar and Javalambre, on relatively homogeneous calcareous soils with Tertiary sediments. The latitudes of the plots range from N 40º 20' to N 40º 2' and the longitudes from W 1º 10' to W 0º 41', with altitudes from 843 to 1124 m a.s.l.…”

“…The 6 trees selected are a subset of the 18 trees sampled to quantify T. melanosporum mycelium over 17 years by Liu et al (2014) and soil samples were collected as reported: From each tree we cored three soil subsamples (30 cm deep and 7 cm diameter) 40 cm from the trunk, at the vertices of a randomly-oriented equilateral triangle, and mixed them thoroughly into one pooled sample, which was passed through a 4 mm mesh to remove debris, stones and roots, and then placed on ice and taken to the laboratory where they were stored at -20ºC in a sealed plastic bag.…”

Black truffle cultivation: a global reality

“…The methods for quantifying mycelium of T. melanosporum in the soil have allowed Liu et al (2014) to monitor its spread and increase of density in young plantations. Suz et al (2008) showed the relation between the abundance of mycelium and the formation of the brûlé.…”

Tuber 2013: scientific advances in sustainable truffle culture

“…Sánchez et al (2014) present an easy and reliable method to quickly check for the presence of T. melanosporum ectomycorrhiza on Q. ilex plantation trees and conclude that only three trees need sampling to provide a reasonable estimate of black truffle ectomycorrhiza presence. Liu et al (2014) examine the rate of black truffle mycelial spread on Q. ilex plantation trees of different ages and report that mycelium extension reaches a maximum from the base of trees after 7 to 10 years. These findings address important questions regarding tree density in plantations and the potential for different dominant truffle mating types on individual trees to contact one another and initiate sporocarp formation.…”