Service Communautaire d'Information sur la Recherche et le Développement - CORDIS

Optimised protocol for growing AM fungal cultures tolerant to heavy metals

Two AM isolates of Glomus intraradices Schneck and Smith obtained from two heavy metal (HM) contaminated localities of anthropogenic origin in the Czech Republic were tested to optimise protocol for cultivation of AMF isolates from heavy metal contaminated soils:
- G. intraradices PH5 from Pb-contaminated waste in the proximity of the Pøíbram lead smelter and

- G. intraradices BEG140 from Mn-contaminated waste deposit Chvaletice.

Both isolates were obtained via trap cultures and subsequent multispore cultures. After successful sporulation in multi-spore culture, two lineages of each isolate were established either in the original HM-contaminated substrate or in inert metal-free substrate (clinoptilolite/sand). G. intraradices PH5 and BEG140 have been cultured for 5 years in these two lineages. Culture conditions for maintaining HM tolerance of AM fungi lineages were determined.

It was shown that the isolates of AM fungi originated from contaminated soils could potentially loose their tolerance to heavy metal when sub-cultured for a long term in inert media without contamination. This phenomena was highly significant for G. intraradices isolate BEG140 obtained from substrate contaminated with Mn, however, it was less pronounced for the isolate of the same AM fungal species from lead polluted substrate. In general, AM fungi were found to be more resistant to elevated concentration of cadmium than maize plants, but for Pb and Mn the results were opposite. Mycorrhizal inoculation exhibited potential to increase HM uptake to the roots and also to elevate P uptake into plant tissue. The isolate of G. intraradices from Pb contaminated soil has shown higher accumulation of heavy metals (including Pb) in the ERM as compared to reference isolate of G. intraradices from non-contaminated soil.

In conclusion, the cultivation protocol for AM fungi isolates with heavy metal tolerance requires continuation of heavy metal stress, i.e. either cultivation of fungi in original soils or in inert media with simulated HM stress, to ascertain maintenance of heavy metal tolerance during sub-culturing of the isolate. Similar behaviour might be expected for fungal tolerance to other kinds of environmental stress what should be further tested (salinity, pollution with xenobiotics etc.). Therefore, these results are potentially usable for industrial producers of mycorrhizal inocula targeted for bioremediation. Desired maintenance of heavy metal stress tolerance requires suggested optimisation of cultivation protocol for these AM fungal strains originated in contaminated soils.

It is therefore essential to cultivate mother cultures of AM strains from contaminated soils in their original soils to maintain stress (and in consequence to preserve the stress tolerance of the fungal isolate. Alternatively, the stress can be simulated in inert cultivation media e.g. by adding solution of relevant heavy metals at similar levels of available elements as in the original soil. This should ascertain maintenance of HM tolerance of AM strains isolated from contaminated soils or AM strains transformed by insertion of targeted methallothionein gene (later is being currently tested for transformed strain of Gigaspora rosea).

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Miroslav VOSÁTKA, (Head of scientific division)
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