Mycorrhiza (1996) 6:499–505 Q Springer-Verlag 1996 ORIGINAL PAPER Teresa E. Pawlowska 7 Janusz Błaszkowski Åke Rühling The mycorrhizal status of plants colonizing a calamine spoil mound in southern Poland Accepted: 29 November 1996 Abstract The arbuscular mycorrhizal (AM) status of Silene vulgaris, dominated the early succession commu- two plant communities on a calamine spoil mound (rich nity and contributed 64% to the relative cover. Spores in cadmium, lead and zinc) in southern Poland was sur- of G. fasciculatum and Entrophospora sp. averaged 20 veyed: an undisturbed grassland community and an per 100 g dry substrate at the disturbed site. early succession community that developed after com- plete removal of the surface layer of the calamine sub- Key words Mycorrhiza 7 Arbuscular mycorrhizal strate about 10 years earlier. The undisturbed site har- fungi 7 Calamine spoil mound 7 Heavy metals bored 40 herbaceous species making up 87% of the ab- solute cover. AM colonization was recorded in 25 spe- cies accounting for 77% of the relative cover. Species Introduction with 51–75% AM root colonization such as Festuca ovi- na and Leontodon hispidus dominated the undisturbed Heavy-metal tolerance in plants results from various turf, contributing 45% to the relative cover. Carex ssp. biochemical and physiological adaptations (Verkleij were the most abundant nonmycorrhizal plants and ac- and Schat 1990). Mycorrhiza formation may contribute counted for 9% of the relative cover. Spores of Glomus by providing a metal exclusion barrier and improving aggregatum, G. constrictum, G. fasciculatum, G. pansi- plant nutritional status (Turnau et al. 1993; Weissen- halos, Glomus sp. and Entrophospora sp. averaged 25 horn et al. 1995a). The ameliorating function of my- per 100 g dry substrate at the undisturbed site. The dis- corrhizal symbiosis has been documented in plants with turbed site was colonized by 25 species accounting for ericoid mycorrhiza (Bradley et al. 1981, 1982) as well as 17% of the absolute cover. Among the AM plants, in several species of ectomycorrhizal plants (Denny and most abundant were the species with up to 20% AM Wilkins 1987; Colpaert and Van Assche 1993). root colonization, such as Agrostis stolonifera and Thy- The role of arbuscular mycorrhiza (AM) in metal mus pulegioides, wich accounted for 24% of the relative uptake remains largely unclear (Galli et al. 1994; Hasel- cover. Nonmycorrhizal species, such as Biscutella laevi- wandter et al. 1994). There is ample laboratory evi- gata, Cardaminopsis arenosa, Gypsophila fastigiata and dence that AM plants are more efficient than nonmy- corrhizal in the acquisition of micronutrients such as Cu and Zn when available at low concentrations (Faber et T. E. Pawlowska (Y)1 al. 1990). However, field relationships between AM Institute of Botany, Jagiellonian University, Lubicz 46, colonization, shoot Cu or Zn concentrations and plant PL-31-512 Kraków, Poland biomass are not fully understood (Sanders and Fitter J. Błaszkowski 1992b). When these or other metals are present in ex- Department of Plant Pathology, Academy of Agriculture, cess, AM colonization may result in metal toxicity and Slowackiego 17, PL-71-434 Szczecin, Poland decrease in plant biomass (Killham and Firestone Å. Rühling 1983). Other evidence suggests that AM may be benefi- Department of Plant Ecology, University of Lund, cial to the host under metal stress, although, this de- Sölvegatan 37, P.O. Ecology Building, pends strongly on plant growth conditions, AM fungus S-223 62 Lund, Sweden and the metal (Weissenhorn et al. 1995a). Several AM Present address: fungal strains have been isolated from metal-enriched 1 Department of Plant Biology, University of Minnesota, 220 Biological Sciences Center, 1445 Gortner Ave., soils, indicating that AM fungi are able to develop hea- St. Paul, MN 55108-1095, USA vy-metal tolerance mechanisms (Gildon and Tinker fax: c1-612-625 1738; e-mail: pawl00146maroon.tc.umn.edu 1981; Weissenhorn et al. 1993; Griffioen 1994). 500 This paper presents results of a survey of the AM Table 1 Chemical properties of the calamine substrate in Boles- status of plants colonizing a calamine spoil mound that law, southern Poland, assessed in the uppermost 20 cm of the spoil material, which corresponds to the plant rooting zone in the originated from lead and zinc ore mining in Boleslaw in undisturbed turf southern Poland. The spoil mound contained Cd, Pb a and Zn at concentrations greatly exceeding natural soil pH (H2O) 6.8 levels (Godzik 1991; Kabata-Pendias and Pendias pH (KCl)b 6.4 Organic matter (%)c 19.5 1992). The history of ore mining in this region extends CEC (cmol kgP1)d 21.4 back to medieval times (Dobrzanska 1955). The vascu- Nitrate-N (mg kgP1)e 13.1 lar plant species occurring at the spoil mound constitute P (mg kgP1)f 9 an example of a highly specialized calamine flora asso- K (mg kgP1)g 141 P1 g ciated with extensive deposits of zinc ores (Dobrzanska Ca (mg kg ) 2690 Mg (mg kgP1)g 434 1955; Ernst 1974). The metal-tolerant vegetation of the P1 h calamine site offers an opportunity to investigate the Total metals (mg kg ) Cd 180 role of AM in plant interactions with heavy metals at a Cu 18 functional community level. Fe 86000 Pb 4560 Zn 49000 Materials and methods DTPA extractable metals (mg kgP1) Cd 37.4 Cu 1.1 Site description Fe 13.2 Pb 323 The investigated spoil mound is located in Boleslaw (50717b52nN, Zn 1270 19728b35nE) west of Kraków in southern Poland. It is composed mainly of the Triassic oolitic limestone and metalliferous dolom- a With H O (1:2.5 w/v) ite (calamine) from the mined ore (Dobrzanska 1955). The sub- 2 b With KCl (1:2.5 w/v) strate is stony and except for a thin humus layer associated with c Wet oxidation in K Cr O followed by FeSO (NH ) SO titra- densely vegetated turfs is not differentiated into horizons. 2 2 7 4 4 2 4 tion The spoil mound harbors grassland vegetation with scattered d Ammonium saturation/KCl displacement dwarfed pines Pinus sylvestris and birches Betula pendula. Study e Cadmium reduction of the mycorrhiza in herbaceous plants was carried out in 1989– f Olsen’s NaHCO 1990 at two sites of about 100 m2 each. One site was an undis- 3 g Extractable with neutral NH OAc turbed, densely vegetated turf. Table 1 summarizes some of the 4 h Extractable with hot NHO chemical properties of the calamine substrate at this site. The sec- 3 ond site was a disturbed area where the top layer of substrate had been completely stripped around 1980 and the site left to revege- tate naturally. The calamine substrate at the disturbed site con- AM fungi tained 0.8% (w/w) organic matter and did not differ significantly in total metal content from the undisturbed site (data not present- The AM fungi were assessed in samples of the uppermost 10 cm ed). of the spoil substrate. Three 1-kg samples were collected at ran- dom from each site in September 1989. Spores were extracted from the substrate by wet sieving and decanting (Gerdemann and Herbaceous cover Nicolson 1963). The AM fungi were identified based on their ori- ginal descriptions, and specimens were deposited in the collection Absolute cover was defined as a vertical projection of shoot area of J. Błaszkowski (Academy of Agriculture, Szczecin, Poland). to the ground surface expressed as a percent of the reference area Two species remain unidentified and are referred to by their ref- (Mueller-Dombois and Ellenberg 1974). The absolute cover of erence numbers. herbaceous plant species was estimated in May 1990 by a point intercept method (Mueller-Dombois and Ellenberg 1974). A 10- point frame was placed at random over the investigated sites; the number of sample points was 1000 per site. The relative cover was Results calculated for each species as its percent contribution to the total plant cover. The AM status of the undisturbed turf community The herbaceous cover in the undisturbed turf was Root sampling and preparation about 87% and comprised 40 species (Tables 2, 3). Intact roots of the herbaceous species were sampled in May, June, There were 15 species of nonmycorrhizal plants which July and September, as flowering occurred. The stony substrate constituted 37.5% of species recorded at this site and impeded retrieval of root systems in their entirety. Therefore, aft- contributed about 22% to the relative cover. The non- er washing, the fine lateral roots were cut off and roots of at least 5 specimens with a total fresh weight of about 1.5 g were pooled, mycorrhizal plants belonged to families Brassicaceae fixed and stored in 50% ethanol. After clearing in 2.5% KOH, (Alyssum montanum, Biscutella laevigata, Cardaminop- roots were stained in 0.05% trypan blue according to Koske and sis arenosa), Caryophyllaceae (Cerastium arvense, C. Gemma (1989). Roots were examined with a compound micro- fontanum subsp. triviale, Gypsophila fastigiata, Silene scope for the presence of structures characteristic of AM such as arbuscules, coils and vesicles. The AM percent root colonization vulgaris), Cyperaceae (Carex caryophyllea, C. ericeto- was estimated by a grid intersect method using a dissecting micro- rum), Polygonaceae (Rumex acetosella), Santalaceae scope (Giovannetti and Mosse 1980). (Thesium alpinum) and Scrophulariaceae (Euphrasia 501 Table 2 Mycorrhizal status of herbaceous plant species, ordered by families, found at the undisturbed site of the calamine spoil mound in Boleslaw, southern Poland. Values listed are means BSEM (np3) (A arbuscules, C coils, V vesicles) Family Species Mycorrhizal Percentage of structures roots colonized Polygonaceae Rumex acetosa L. V 4.0B 4.0 Rumex acetosella L. Absent Caryophyllaceae Cerastium arvense L. V 4.3B 2.2 Cerastium fontanum Baumg. subsp.