Tuber Magnatum Research Articles

Tuber magnatum Picco: a new record for the Turkish mycobiota

Tuber magnatum Picco, the Italian white truffle mushroom recorded for the first time in Türkiye, is a rare and economically valuable mushroom. Samples were collected from Sakarya province during field studies in 2024. The samples were identified as T. magnatum according to their macro- and microscopic features. To make molecular confirmation of the species, the samples' ITS1 and ITS4 gene regions were analysed and registered in Genbank with the number PP239641. The sample showed 100% compatibility with other T. magnatum accessions in Genbank. The distributions of Tuber species in Türkiye were given, and their taxonomic features were compared.

In vitro interactions between Bradyrhizobium spp. and Tuber magnatum mycelium.

Tuber magnatum is the most expensive truffle, but its large-scale cultivation is still a challenge compared to other valuable Tuber species. T. magnatum mycelium has never been grown profitably until now, which has led to difficulties to studying it in vitro. This study describes beneficial interactions between T. magnatum mycelium and never before described bradyrhizobia, which allows the in vitro growth of T. magnatum mycelium. Three T. magnatum strains were co-isolated on modified Woody Plant Medium (mWPM) with aerobic bacteria and characterised through microscopic observations. The difficulties of growing alone both partners, bacteria and T. magnatum mycelium, on mWPM demonstrated the reciprocal dependency. Three bacterial isolates for each T. magnatum strain were obtained and molecularly characterised by sequencing the 16S rRNA, glnII, recA and nifH genes. Phylogenetic analyses showed that all nine bacterial strains were distributed among five subclades included in a new monophyletic lineage belonging to the Bradyrhizobium genus within the Bradyrhizobium jicamae supergroup. The nifH genes were detected in all bacterial isolates, suggesting nitrogen-fixing capacities. This is the first report of consistent T. magnatum mycelium growth in vitro conditions. It has important implications for the development of new technologies in white truffle cultivation and for further studies on T. magnatum biology and genetics.

Identification of Bis(methylsulfanyl)methane and Furan-2(5H)-one as Volatile Marker Compounds for the Differentiation of the White Truffle Species Tuber magnatum and Tuber borchii.

Some truffles are expensive and, therefore, are prone to food fraud. A particular problem is the differentiation of high-priced Tuber magnatum truffles from cheaper Tuber borchii truffles, both of which are white truffles with similar morphological characteristics. Using an untargeted approach, the volatiles isolated from samples of both species were screened for potential marker compounds by comprehensive two-dimensional gas chromatography-time-of-flight mass spectrometry (GC×GC-TOFMS) and statistical analysis of the obtained semiquantitative data. Results suggested bis(methylsulfanyl)methane and furan-2(5H)-one as compounds characterizing T. magnatum and T. borchii, respectively. Exact quantitation of both volatiles by conventional one-dimensional gas chromatography-mass spectrometry in combination with stable isotopologues of the target compounds as internal standards confirmed both as marker compounds. The method is suitable to be used in the routine analysis for the objective species differentiation of T. magnatum and T. borchii.

White Acqualagna truffle (Tuber magnatum Pico): Evaluation of volatile and non-volatile profiles by GC-MS, sensory analyses and elemental composition by ICP-MS

The White Truffle is the most expensive edible underground mushroom. In this study the first characterization of the Acqualagna white truffle was delivered, taking into consideration the soil of origin and the human perception. The volatile profile was identified by GC–MS and compared with the descriptors obtained by sensory analysis. The non-volatile characterization was done using elemental composition by ICP-MS analysis, elemental analysis, and spectrophotometric assays. The volatile profile consists mainly of bis(methylthio)methane (78.72%) and other minor constituents, linked to seven odorant descriptors: garlic-like, nutty-like, geosmine-like, floral, mushroom-like, pungent and green/herbal. ICP-MS revealed that truffle has a higher content of K, P, S, Ca and Mg (97% of the elements investigated) and that it assimilates the Rare Earth Elements (REE) from the soil without discriminating them. In conclusion, this project is the first step for the enhancement of local food, linked to the territory conditions in which it is produced.

From Nontargeted to Targeted Analysis: Feature Selection in the Differentiation of Truffle Species (Tuber spp.) Using 1H NMR Spectroscopy and Support Vector Machine.

The price of different truffle types varies according to their culinary value, sometimes by more than a factor of 10. Nonprofessionals can hardly distinguish visually the species within the white or black truffles, making the possibility of food fraud very easy. Therefore, the identification of different truffle species (Tuber spp.) is an analytical task that could be solved in this study. The polar extract from a total of 80 truffle samples was analyzed by 1H NMR spectroscopy in combination with chemometric methods covering five commercially relevant species. All classification models were validated applying a repeated nested cross-validation. In direct comparison, the two very similar looking and closely related black representatives Tuber melanosporum and Tuber indicum could be classified 100% correctly. The most expensive truffle Tuber magnatum could be distinguished 100% from the other relevant white truffle Tuber borchii. In addition, signals for a potential Tuber borchii and a potential Tuber melanosporum marker for targeted approaches could be detected, and the corresponding molecules were identified as betaine and ribonate. A model covering all five truffle species Tuber aestivum, Tuber borchii, Tuber indicum, Tuber magnatum, and Tuber melanosporum was able to correctly discriminate between each of the species.

Tuber magnatum Picco: the challenge to identify ascoma-associated bacteria as markers for geographic traceability.

OPINION article Front. Microbiol., 16 May 2023Sec. Microbial Symbioses Volume 14 - 2023 | https://doi.org/10.3389/fmicb.2023.1142214

Truffle flavored commercial products veracity and sensory analysis from truffle and non-truffle consumers

The price of the truffle species Tuber melanosporum and Tuber magnatum can be up to fifty times higher, or even more, than the cheapest edible truffle species due to their appreciated aroma and low production levels. This aroma is seriously affected after the application of treatments for the conservation of food products, usually thermal (freezing or sterilization). Hence, many of the truffled products that are retailed are characterized by the use of truffle species of low economic value and the addition of flavoring substances. Most of the time, the added flavorings do not mimic fresh truffle aroma and do not correspond to the truffle species appearing in the ingredients list and the statement of identity. These products sometimes include pictures of truffles or the term ‘white or black truffle’ in the label, which might confuse the consumer. To study this practice in the food industry, 51 products were evaluated through instrumental techniques determining truffle species presence by microscopy and molecular techniques, as well as the level of truffle flavorings added by HS-GC-MS analysis and by sensory perception scale. Finally, a sensory analysis of eight products was carried out by consumers distributed into two groups, those who had previously tasted fresh truffles and those who had not. Lower-value truffle species such as Tuber aestivum and Tuber indicum were frequently found in products in which the labeling did not indicate so. Also, 48% of the products contained high levels of added flavorings. In the sensory analysis, non-consumers of truffles rated flavored products more positively (up to 2 more points in some products) than truffle consumers. Also, this group associated negative attributes (weird and disappointing) to products elaborated with real black truffle, whereas truffle consumers associated positive attributes (truffle flavor, truffle smell and gourmet) to them. • 51 truffled products were tested to detect frauds. • Only 20% of truffled products were correctly labelled. • The combination of analysis allows detect which truffle specie is added. • Truffle consumers endowed truffle flavor, truffle smell and gourmet to products. • Negatives attributes were related with fresh-truffle products by some consumers.

The Mysteries of the White Truffle: Its Biology, Ecology and Cultivation

Tuber magnatum Picco is the most expensive of the truffles and a great deal of research has been carried out in an attempt to solve the mysteries of its ecology and biology. However, considerable work remains to be done particularly on those secrets of its life cycle that remain a mystery. It is known that T. magnatum is heterothallic, but it has yet to be determined how fertilization occurs between the two strains of different mating types. It is also known that the white truffle is an ectomycorrhizal fungus, and its mycorrhizas can be produced in greenhouses, but then they seem to disappear in the field. The role of other soil microorganisms, fungi and bacteria, on its soil mycelial development and fructification is intriguing but is far from being completely understood. All these uncertainties have made the cultivation of T. magnatum extremely difficult and only recently have we had the scientific proofs that it is possible. Even so, many questions remain unanswered and the management practices of T. magnatum plantations are still to be better defined to also enable the taming of this truffle.

Role and potentialities of bacteria associated with Tuber magnatum: A mini-review.

Among the hypogeous ectomycorrhizal fungi, the white truffle Tuber magnatum Picco is the species of greatest interest, both from an ecological and economic point of view. The increasing market demand of the precious white truffle along with the fall in its natural production led to a growing interest in cultivation techniques and encouraged truffle growers and researchers to deeper investigate factors that could affect and improve T. magnatum productivity. In this context, microbial communities play a central role. Indeed, in the last few years, the hypothesis of a potential link between microbial community composition and truffle orchard productivity is arousing a greater attention. Moreover, since the value of the prized T. magnatum can vary in relation to its provenience, the need to define a reliable tracking system is also emerging and bacteria appear to be a promising tool. Accordingly, the present mini-review summarises the knowledge currently available on T. magnatum microbial communities, focusing on the role of truffle-associated bacteria and highlighting similarities and differences between samples of different origin, to address the following issues: (i) Is there a correlation between microbial taxa and truffle ground productivity? (ii) Can bacteria actually be used as markers of T. magnatum geographic origin? The identification of microorganisms able to promote T. magnatum formation may represent an important advance in the field of truffle farming. Similarly, the detection of bacterial taxa that can be used as markers of T. magnatum origin could have a considerable impact on truffle industry and trade, even at local scale.

Bacterial Communities in the Fruiting Bodies and Background Soils of the White Truffle Tuber magnatum.

Tuber magnatum Picco is a greatly appreciated truffle species mainly distributed in Italy. Its price and characteristics mostly depend on its geographical origin. Truffles represent a fundamental step of the life cycle of Tuber species promoting spore dissemination. They consist of two main parts, gleba, the inner part, and peridium, which is in direct contact with ground soil. Within the truffle and around in the growing soil, both the occurrence and abundance of different microbial species seem to play an essential role in truffle production. The development of the next-generation sequencing (NGS) based technology has greatly improved to deepen the role of the composition of microbial communities, thus improving the knowledge of the existing relationships between microbial taxa in a specific condition. Here, we applied a metabarcoding approach to assess the differences in T. magnatum samples collected from three areas in Tuscany (Italy). Peridium and gleba were analyzed separately with the aim to distinguish them based on their microbial composition. Also, soil samples were collected and analyzed to compare productive and unproductive truffle grounds to confirm the presence of specific patterns linked to truffle production. Results indicate that differences occurred between truffle compartments (gleba and peridium) as well as between analyzed soils (productive and unproductive), with distinctive taxa associated. Furthermore, findings also demonstrated specific characteristics associated with truffle collection areas, thus indicating a degree of microbial selection related to different environments.

Fungal and Bacterial Diversity in the Tuber magnatum Ecosystem and Microbiome.

Fungi belonging to the genus Tuber produce edible ascocarps known as truffles. Tuber magnatum Picco may be the most appreciated truffle species given its peculiar aroma. While its life cycle is not yet fully elucidated, some studies demonstrated an active role of microorganisms. The main goal of this study was to determine how the T. magnatum microbiome varies across space and time. To address this, we characterized microbial communities associated with T. magnatum through high-throughput amplicon sequencing of internal transcribed spacer (ITS) and 16S rDNAs in three productive natural sites in Italy across 2years. At each site, four truffles were sampled as well as the soil underneath and at 40, 100, and 200cm from the harvesting points, to assess for microbial variation between substrates, years, and sites. A statistically significant site-related effect on microbial communities was identified, whereas only the prokaryotic community was significantly affected by the distance of soil from the truffle. Significant differences between sampling years were also found, demonstrating a possible relation among rainfall precipitation and Firmicutes and Actinobacteria. Thirty-six bacterial OTUs in truffles and 11 bacterial OTUs in soils beneath truffles were identified as indicator taxa. As shown for other truffle species, the dominance of Bradyrhizobium, Rhizobium, and Ensifer spp. within the truffle fruiting body suggests an evolutionary adaptation of this microorganism to the genus Tuber. The present work offers novel and relevant insights into the microbial ecology of T. magnatum ecosystems and fruiting bodies. The function and role of these bacteria in the truffle microbiome and life cycle are in need of further investigation.

Portable vs. Benchtop NIR-Sensor Technology for Classification and Quality Evaluation of Black Truffle.

Truffles represent the best known and most expensive edible mushroom. Known as Ascomycetes, they belong to the genus Tuber and live in symbiosis with plant host roots. Due to their extraordinary taste and smell, truffles are sold worldwide for high prices of up to 3000–5000 euros per kilogram (Tuber magnatum PICO). Amongst black truffles, the species Tuber melanosporum VITTAD. is highly regarded for its organoleptic properties. Nonetheless, numerous different sorts of black truffle are offered at lower prices, including Tuber aestivum VITTAD., Tuber indicum and Tuber uncinatum, which represent the most frequently consumed types. Because truffles do not differ visually for inexperienced consumers, food fraud is likely to occur. In particular, for the highly prized Tuber melanosporum, which morphologically forms very similar fruiting bodies to those of Tuber indicum, there is a risk of fraud via imported truffles from Asia. In this study, 126 truffle samples belonging to the four mentioned species were investigated by four different NIR instruments, including three miniaturized devices—the Tellspec Enterprise Sensor, the VIAVI solutions MicroNIR 1700 and the Consumer Physics SCiO—working on different technical principles. Three different types of measurement techniques were applied for all instruments (outer shell, rotational device and fruiting body) in order to identify the best results for classification and quality assurance in a non-destructive manner. Results provided differentiation with an accuracy up to 100% for the expensive Tuber melanosporum from Tuber indicum. Classification between Tuber melanosporum, Tuber indicum, Tuber aestivum and Tuber uncinatum could also be achieved with success of 100%. In addition, quality monitoring including discrimination between fresh and frozen/thawed, and prediction of the approximate date of harvesting, was performed. Furthermore, feasibility studies according to the geographical origin of the truffle were attempted. The presented work compares the performance for prediction and quality monitoring of portable vs. benchtop NIR devices and applied measurement techniques in order to be able to present a suitable, accurate, fast, non-destructive and reliable method for consumers.

Antioxidant Profile and Biosafety of White Truffle Mycelial Products Obtained by Solid-State Fermentation.

Solid-state fermentation may produce therapeutic compounds with higher biomass or better product characteristics than those generated by submerged fermentation. The objectives of this study were to analyze the antioxidant activities and biosafety of products obtained by white truffle (Tuber magnatum) solid-state fermentation in media with different ratios of soybean and red adlay. High levels of antioxidant components and high antioxidant activities such as DPPH radical scavenging, ferrous ion chelation, and reducing power were measured in 20 mg/mL water and ethanol extracts of the white truffle fermentation products. When the solid-state fermentation medium contained soybean and red adlay in a 1:3 ratio (S1A3), the fermentation product had more uniform antioxidant compositions and activities by principal component analysis (PCA). In addition, a 200 ppm water extract of the mycelial fermentation product was able to protect zebrafish embryos from oxidative stress induced by 5 mM hydrogen peroxide. Sprague–Dawley rats were fed the mycelial fermentation product for 90 consecutive days, revealing a no-observed-adverse-effect level (NOAEL) of 3000 mg/kg BW/day. Therefore, mycelial products obtained by white truffle solid-state fermentation can be used instead of expensive fruiting bodies as a good source of antioxidant ingredients.

Investigation of the Proteomes of the Truffles Tuber albidum pico, T. aestivum, T. indicum, T. magnatum, and T. melanosporum.

Truffles of the Tuber species are known as expensive foods, mainly for their distinct aroma and taste. This high price makes them a profitable target of food fraud, e.g., the misdeclaration of cheaper truffle species as expensive ones. While many studies investigated truffles on the metabolomic level or the volatile organic compounds extruded by them, research at the proteome level as a phenotype determining basis is limited. In this study, a bottom-up proteomic approach based on LC-MS/MS measurements in data-independent acquisition mode was performed to analyze the truffle species Tuber aestivum, Tuber albidum pico, Tuber indicum, Tuber magnatum, and Tuber melanosporum, and a protein atlas of the investigated species was obtained. The yielded proteomic fingerprints are unique for each of the of the five truffle species and can now be used in case of suspected food fraud. First, a comprehensive spectral library containing 9000 proteins and 50,000 peptides was generated by two-dimensional liquid chromatography coupled to tandem mass spectrometry (2D-LC-MS/MS). Then, samples of the truffle species were analyzed in data-independent acquisition (DIA) proteomics mode yielding 2715 quantified proteins present in all truffle samples. Individual species were clearly distinguishable by principal component analysis (PCA). Quantitative proteome fingerprints were generated from 2066 ANOVA significant proteins, and side-by-side comparisons of truffles were done by T-tests. A further aim of this study was the annotation of functions for the identified proteins. For Tuber magnatum and Tuber melanosporum conclusive links to their superior aroma were found by enrichment of proteins responsible for sulfur-metabolic processes in comparison with other truffles. The obtained data in this study may serve as a reference library for food analysis laboratories in the future to tackle food fraud by misdeclaration of truffles. Further identified proteins with their corresponding abundance values in the different truffle species may serve as potential protein markers in the establishment of targeted analysis methods. Lastly, the obtained data may serve in the future as a basis for deciphering the biochemistry of truffles more deeply as well, when protein databases of the different truffle species will be more complete.

Biochemical and Health Properties of Truffles

Truffles are the most expensive edible mushrooms refer to genus tuber which grows symbiotically in plant roots such as oaks and hazels. Truffles are underground mushrooms also known for their characteristic earthy flavor which is the major reason for their special place in the culinary. Their characteristic intense aroma helps them in reproduction by attracting small animals. Truffles can survive in a wide range of environments such as deep forests as well as deserts. The most expensive varieties of truffles include Tuber melanosporum (Black truffle), Tuber magnatum (White truffle), Tuber aestivum (Burgundy truffle), Tirmania nivea, and Terfezia chlaveryi (Dessert truffles). Truffles vary in their composition and flavor profile from species to species. The major volatile components which are responsible for truffle aroma are aldehydes, ketones, sulfur compounds, alcohols, and esters. Truffles are highly nutritious, rich in antioxidants, and have therapeutic properties such as antimicrobial activity, antiviral activity, antimutagenic activity, anti-inflammatory activity, hepatoprotective activity, etc. The major active components present in truffle are tuberoside, phenolics, anandamide, and ergosterol.

Aroma and bacterial communities dramatically change with storage of fresh white truffle Tuber magnatum

White truffles are highly valuable, but the aroma is susceptible to change with storage. How volatile composition and microbiome of Tuber magnatum evolve by storage time is poorly understood. Changes in bacterial community and volatile compounds of T. magnatum fruiting bodies with storage time at 23 °C were investigated. Truffles (21 fruiting bodies) were collected from different sites in Italy, Hungary, and Croatia. Single fruiting bodies were subsampled at four time points (day (D)0, 3, 6, and 9). Gas chromatography-mass spectrometry and microbiome composition using PCR-high throughput sequencing were used to analyse the volatiles and bacterial communities, respectively. Spoilage compounds atypical of truffles increased with storage time. Likewise, bacterial communities changed with storage time; families belonging to Xanthobacteraceae dominated at D0 and gradually evolved to more Rhizobiaceae and Pseudomonadaceae by D3 and D6, and finally with more prominent Xanthomonadaceae and Burkholderiaceae at D9. Overall, storage time had increased the diversity of bacterial communities. The microbiome and volatile data were correlated using regularised canonical correlation analysis and determined correlations between 2-methylisoborneol and five bacterial OTUs. Potential candidate bacteria and their volatile metabolites can serve as markers for truffle spoilage. The results highlighted the mechanisms by which the degradation of expensive truffles occur.

Aroma‐volatile profile and its changes in Australian grown black Périgord truffle (Tuber melanosporum) during storage

SummaryBlack Périgord truffle (Tuber melanosporum) is one of the most expensive fungi in the world that appreciated by gourmets. Studies have indicated the impact of growing location and soil microorganisms on the aroma profile of truffle. The aroma profile of West Australian black Périgord truffle (Tuber melanosporum) has not been previously reported, which was studied over a 14 day storage period. Sixty‐four compounds were identified in all truffle samples. Significant changes (P > 0.05) were observed in 11 key volatiles (carbon dioxide, acetaldehyde, 2‐butanone, 3‐methyl‐1‐butanal, toluene, 2‐butenal, formic acid 2‐methyl butyl ester, 3‐methyl‐1‐butanol, 6‐methyl‐2‐heptanol, 3‐octanol and dimethyl sulfoxide) over time. Comparison of these results against published aroma profile of European grown black Périgord truffle identified number of significant similarities and differences were also detected. Dimethyl sulfoxide, a compound previously identified in European grown white truffle (Tuber magnatum), was detected. Principle component analysis (PCA) showed that the major changes in the truffle aroma profile took place in the first 7 days of storage.

Success and failures in the inoculation of five introduced trees in Chile with Tuber magnatum Pico: First advances for the domestication of the white truffle in South America

Truffles are highly demanded edible fungi. They belong to the genus Tuber and have very high economic value, with Tuber magnatum being one of the most expensive and gastronomically used. This study evaluates the potential of five introduce tree species as mycorrhizal hosts for T. magnatum in Chile. Seeds from Quercus cerris, Q. robur and Corylus avellana were harvested from adult trees. Cuttings from Populus nigra and Salix caprea were harvested in the field and asexually propagated under greenhouse conditions. After two months, they were transplanted to 260 cc pots containing sterilized composted pine bark. A spore suspension (106 spores/plant) of T. magnatum was injected directly into plant roots. Three months after inoculation we observed spore germination and the presence of mycelium around the roots in some seedlings. After seven months, we observed mycorrhizae from T. magnatum only in Q. cerris, Q. robur and C. avellana, characterized by their epidermoid mantle and awl-shaped, bristle-like cystidia. The identification the mycorrhizal structures was confirmed by sequencing of the nuclear ITS- rDNA region. This study provide the first advances for the domestication of this highly valuable truffle in Chile and South America and the successful mycorrhized plants could be used in further field assays. Some Quercus, Populus and Salix species have been used as ornamental plants and are naturalized in Chile for over 100 years. Additionally, hazelnut (Corylus avellana) is currently grown in the country under intensive silviculture. These tree species could act as a non-intentional host for truffles, dispersed from production sites.

Heterogeneity of the white truffle Tuber magnatum in a limited geographic area of Central-Southern Italy.

Molise region (Central-Southern Italy) is one of the Italian richest areas of truffles and contributes significantly to the national production of the precious Tuber magnatum. Nevertheless, Molise truffle has received little scientific attention. Accordingly, in the present study, two T. magnatum populations collected in two different sites of Molise region were characterised from a morphological, genetic and microbiological point of view. A considerable variability between and within the two analysed groups emerged, suggesting an interesting heterogeneity of Molise white truffle populations. Ascocarps of the two groups significantly differed in size and maturation degree, although no linear correlation between weight and maturity was found. Genetic investigations focused on the Sequence-Characterised Amplified Region SCAR A21-inf. Three haplotypes, randomly distributed within the two truffle groups regardless of their collection sites, were detected. The 16S rRNA gene amplicon high-throughput sequencing provided an overview of the composition of the ascocarp-associated bacterial communities. A predominance of α-Proteobacteria was observed, with Bradyrhizobium among the main genera. However, some truffles showed unusual microbial profiles, with Pedobacter, Polaromonas and other bacterial genera as dominant taxa.

Co-occurrence of true truffle mycelia in Tuber magnatum fruiting sites.

Tuber magnatum (the white truffle) is the most precious species of the genus Tuber which comprises the hypogeous ectomycorrhizal species called "true truffle." Despite its high economic value, the knowledge on its ecology is scant, principally due to the difficulty to find its mycorrhizas in the soil. The possibility to detect its mycelium by DNA extracted from soil has given a new chance for studying this truffle species. In this work, the co-occurrence of other Tuber species with T. magnatum mycelium was investigated by using species-specific primers in several productive areas located in central and northern Italy. Most (82%) of the examined soil samples showed at least one other Tuber species in addition to T. magnatum. The most common was T. maculatum (72% of soil samples) followed by T. borchii, T. rufum, T. brumale, T. dryophilum, T. macrosporum, and T. melanosporum (40%, 37%, 22%, 19%, 12%, and 1% of soil samples, respectively). Tuber aestivum was never detected in T. magnatum productive patches. Analysis of species co-occurrence showed that the pairwise associations between T. dryophilum-T. brumale, T. brumale-T. borchii, and T. borchii-T. dryophilum was significant. The results suggest that Tuber mycelial network in white truffle grounds is much more extensive than the distribution of their ectomycorrhizas and competitive exclusion between different Tuber species seems to take place only for root colonization.