Assessment of genetic diversity in some of black and brown Iranian truffles by ISSR markers

Document Type : Original research paper


Department of Horticultural Sciences, Faculty of Agriculture and Natural Resources, University of Mohaghegh Ardabili, Ardabil, Iran


Tuber species are edible fungi and plant-symbiotic microorganisms that form a beneficial relationship with the roots of certain trees and plants (ectomycorrhizae). After interaction with a plant host, tuber species produced hypogeous fruit bodies of great economic value known as forest truffles. There are different species of truffles, but based on species and place of origin varied their quality and market price. Truffle identification is based on morphological analysis maybe fail to distinguish them due to highly susceptible to environmental conditions. But using molecular markers to identify truffles can be more accurate, less expensive and reliable monitoring. In this context, twelve inter-simple sequence repeats (ISSR) primers were chosen for amplifying the genetic materials of black and brown truffles. In this study, a total of 57 polymorphic bands were amplified (an average of 5.18 bands). The Polymorphism Information Content (PIC) value and gene diversity (H) was with an average 0.37 and 0.50, respectively. During the ISSR screening good amplification products were obtained from primers based on GA, (AG) G, (AG)T, and GAC repeats. The population analysis result revealed that there are three main clusters A, B and C. Four strains Ardabil, Khalkhal, Zanjan and Urmia were identified to be in the group A cluster.  The strains of at second and third groups were black and brown truffles respectively. The results indicated that truffles had two separate speciation events (DK = 2). According to DK = 2, the samples of Ardabil, Khalkhal, Zanjan and Urmia grouped in the same group and rest of truffles in other groups.


[1] Araujo, F., Pacheco. M., Vieira. F. 2016. ISSR molecular markers for the study of the genetic diversity of Mimosa caesalpiniaefolia Benth. IDESIA (Chile) Junio Volumen 34, Pg 47-52
[2] Belfiori. B., D'Angelo, V., Riccioni, C., Leonardi, M., Paolocci. F., Pacioni, G., Rubini. A. 2020. Genetic Structure and Phylogeography of Tuber magnatum Populations. Diversity 12(2), p.44.
[3] Cekic, C., Calis, O., Ozturk E. 2018. Genetic diversity of wild raspberry genotypes (Rubus idaeus L.) in North Anatolia based on ISSR markers. Appl Ecol Environ Res, 16(5):6835-6843.
[4] Cullere, L., Ferreira. V., Chevret B. Venturini ME. Sánchez-Gimeno AC. Blanco D. 2010. Characterisation of aroma active compounds in black truffles (Tuber melanosporum) and summer truffles (Tuber aestivum) by gas chromatography–olfactometry. Food Chem, 122(1), pp.300-306.
[5] Estoup, A., Presa, P., Krieg. F., Vaiman, D. 1993. (CT)n and (GT)n microsatellites: a new class of genetic markers for Salmo trutta L. (brown trout). Heredity 71: 488–496.
[6] Felix, F., Cardoso, F., Chagas, K., Ferrari, C., Vieira, F. 2020. Applications of ISSR markers in studies of genetic diversity of Pityrocarpa moniliformis. Revista Caatinga 33(4):1017-1024
[7] Gajera, BB., Kumar, N., Singh, A.S., Punvar, B.S., Ravikiran, R., Subhash. N., Jadeja, G.C. 2010. Assessment of genetic diversity in castor (Ricinus communis L.) using RAPD and ISSR markers. Ind Crops Prod, 32(3), pp.491-498.
[8] García-Cunchillos I. Sánchez S. Barriuso JJ. Pérez-Collazos E. 2014. Population genetics of the westernmost distribution of the glaciations-surviving black truffle Tuber melanosporum. Mycorrhiza 24(1), pp.89-100.
[9] Mello A. Murat C. Vizzini A. Gavazza V. Bonfante, P. 2005. Tuber magnatum Pico, a species of limited geographical distribution: its genetic diversity inside and outside a truffle ground. Environ. Microbiol, 7(1), pp.55-65.
[10] Molinier V, Murat C, Frochot H, Wipf D. Splivallo R. 2015. Fine‐scale spatial genetic structure analysis of the black truffle Tuber aestivum and its link to aroma variability. Environ. Microbiol, 17(8), pp.3039-3050.
[11] Molinier V. Murat C. Morin E. Gollotte A. Wipf D. Martin F. 2013. First identification of polymorphic microsatellite markers in the Burgundy truffle, Tuber aestivum (Tuberaceae). APPS, 1(2), p.120.
[12] Murat C. Díez J. Luis P. Delaruelle C. Dupré C. Chevalier G. Bonfante P. Martin F. 2004. Polymorphism at the ribosomal DNA ITS and its relation to postglacial re-colonization routes of the Périgord truffle Tuber melanosporum. New Phytologist 164:401-411
[13] Muthusamy   S.  Kanagarajan S. Ponnusamy S. 2008. Efficiency of RAPD and ISSR markers system in accessing genetic variation of rice bean (Vigna umbellata) landraces. Electronic Journal of Biotechnology 11(3):32-41.
[14] Paolocci F. Rubini A. Granetti B. Arcioni S. 1999. Rapid molecular approach for a reliable identi¢cation of Tuber spp. Ectomycorrhizae. FEMS Microbiology Ecology 28 (1): 23-30.
[15] Riccioni C. Belfiori B. Rubini A. Passeri V. Arcioni S. Paolocci, F. 2008. Tuber melanosporum outcrosses: analysis of the genetic diversity within and among its natural populations under this new scenario. New Phytologist 180(2), pp.466-478.
[16] Riccioni C. Rubini A. Belfiori B. Gregori G. Paolocci F. 2016. Tuber magnatum: the special one. What makes it so different from the other Tuber in True Truffle (Tuber spp.) in the World (pp. 87-103). Springer, Cham.
[17] Riccioni C. Rubini A. Türkoğlu A. Belfiori B. Paolocci F. 2019. Ribosomal DNA polymorphisms reveal genetic structure and a phylogeographic pattern in the Burgundy truffle Tuber aestivum Vittad. Mycologia 111(1), pp.26-39.
[18] Rubini A, Topini F, Riccioni C, Paolocci F, Arcioni S. 2004. Isolation and characterization of polymorphic microsatellite loci in white truffle (Tuber magnatum). Molecular Ecology Notes 4:116–118, doi:10.1111/j.1471-8286.2004.00587.x
[19] Splivallo R. Ottonello S. Mello A. Karlovsky P. 2011. Truffle volatiles: from chemical ecology to aroma biosynthesis. New Phytologist 189(3), pp.688-699.
[20] Tabouret P. 2011. Exclusive Description dun site franc ais producteur de truffe blanche d’Italie. Le Trufficulteur 78, p.18.
[21] Tanya P. Taeprayoon P. Hadkam Y. Srinives, P. 2011. Genetic diversity among Jatropha and Jatropha-related species based on ISSR markers. Plant Molecular Biology Reporter 29(1), pp.252-264.
[22] Weden C. Danell, E. Camacho FJ. Backlund A. 2004. The population of the hypogeous fungus Tuber aestivum syn. T. uncinatum on the island of Gotland. Mycorrhiza 14(1), pp.19-23.
Volume 7, Issue 2
December 2021
Pages 63-70
  • Receive Date: 05 June 2021
  • Revise Date: 02 July 2021
  • Accept Date: 04 September 2021
  • First Publish Date: 04 September 2021