Tuber borchii Vitt. mycorrhiza protects Cistus creticus L. from heavy metal toxicity

• We report the protective effects of mycorrhization with the fungus Tuber borchii in seedlings treated with heavy metals.
• Leaves’ chlorosis induced by heavy metals is remarkably reduced in mycorrhized seedlings compared with non mycorrhized seedlings.
• In mycorrhized seedlings, treatment with heavy metals does not reduce expression of genes involved in chlorophyll biosynthesis.
• DNA damage induced by HM stress is lower in mycorrhized than in non mycorrhized seedlings.

Heavy metals (HMs), such as copper, zinc, lead, mercury and cadmium, are the most abundant and dangerous inorganic environmental pollutants. Growing pieces of evidence suggest that mycorrhizal fungi can alleviate metal toxicity in plants. In this study, we focused attention on the ectomycorrhizal (ECM) fungus Tuber borchii Vitt., which is widespread in Italy and is of great ecological interest because of the mutualistic associations and the advantages it provides to host plants. Seedlings of the Mediterranean shrub Cistus creticus L., mycorrhized and non mycorrhized with the ECM fungus T. borchii, were treated with HMs (zinc, lead and chromium). HMs induced leaves’ chlorosis in non mycorrhized seedlings; while no significant difference was observed in pigmentation of mycorrhized seedlings’ leaves. This observation was confirmed by Euclidean Distance of color measurements in L*a*b* units from RGB digital images of leaves. The decrease in leaves pigmentation observed in HM treated non mycorrhized seedlings strongly correlated with a reduced expression of key genes associated with chlorophyll biosynthesis; instead, no significant variation of gene expression was detected in mycorrhized seedlings treated with HMs.Finally, a comet test on nuclei isolated from leaves of mycorrhized and non mycorrhized C. creticus seedlings, treated or not with HMs, confirmed that the DNA damage induced by HMs stress was lower in mycorrhized than in non mycorrhized seedlings. Taken together, the data suggest the involvement of the mycorrhizal fungus T. borchii in the improvement of HM stress tolerance in C. creticus host.