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.

Truffles, such as Tuber magnatum and Tuber melanosporum, are greatly appreciated throughout the world, both as fresh fruiting bodies and as ingredients in processed products. Diagnostic methods are therefore required to check the identity of truffles in this kind of food. The present paper reports the application of microscopic and molecular techniques to authenticate truffle species in commercial products. Through the application of microscopic techniques, it has been possible to distinguish spores in a cream that could be ascribed to the truffle species (T. magnatum, the highest priced truffle) declared on the label and also spores of a less prized and aromatic truffle that was not indicated on the label. Good quality DNA was quickly obtained in a few hours using a kit generally employed for DNA extraction from soil. A new primer pair was developed to authenticate T. magnatum in commercial products and it was employed in a quantitative PCR assay (qPCR). T. melanosporum, which was neither indicated on the label nor recognized in the processed products containing truffles, was also detected in a cream and its amount was quantified by qPCR. This method can therefore be used to detect fraudulent practices and to protect the consumer of truffle delicacies.