Ecology, 96(3), 2015, pp. 672–683 Ó 2015 by the Ecological Society of America Multi-stressor impacts on fungal diversity and ecosystem functions in streams: natural vs. anthropogenic stress M. TOLKKINEN, 1,5 H. MYKRA ¨ , 1,2 M. ANNALA, 3 A. M. MARKKOLA, 3 K. M. VUORI, 4 AND T. MUOTKA 3 1 Finnish Environment Institute (SYKE), P.O. Box 413, FI-90014, University of Oulu, Finland 2 Thule Institute, P.O. Box 7300, FIN-90014, University of Oulu, Finland 3 Department of Biology, P.O. Box 3000, 90014 University of Oulu, Finland 4 Finnish Environment Institute (SYKE), Technopolis, Jyva ¨skyla ¨ Office, Survontie 9, 40500 Jyva ¨skyla ¨, Finland Abstract. Biological assemblages are often subjected to multiple stressors emerging from both anthropogenic activities and naturally stressful conditions, and species’ responses to simultaneous stressors may differ from those predicted based on the individual effects of each stressor alone. We studied the influence of land-use disturbance (forest drainage) on fungal decomposer assemblages and leaf decomposition rates in naturally harsh (low pH caused by black-shale dominated geology) vs. circumneutral streams. We used pyrosequencing to determine fungal richness and assemblage structure. Decomposition rates did not differ between circumneutral and naturally acidic reference sites. However, the effect of forest drainage on microbial decomposition was more pronounced in the naturally acidic streams than in circumneutral streams. Single-effect responses of fungal assemblages were mainly related to geology. Community similarity was significantly higher in the naturally acidic disturbed sites than in corresponding reference sites, suggesting that land-use disturbance simplifies fungal assemblages in naturally stressful conditions. Naturally acidic streams supported distinct fungal assemblages with many OTUs (operational taxonomic unit) unique to these streams. Our results indicate that fungal assemblages in streams are sensitive to both structural and functional impairment in response to multiple stressors. Anthropogenic degradation of naturally acidic streams may decrease regional fungal diversity and impair ecosystem functions, and these globally occurring environments therefore deserve special attention in conservation planning. Key words: aquatic fungi; biodiversity; catchment geology; forest drainage; leaf decomposition; multiple stressors; natural acidity; next-generation sequencing; streams. INTRODUCTION Studies on the effects of human disturbance on biological assemblages have focused on single-stressor responses despite the fact that most ecosystems are subjected simultaneously to multiple stressors of both anthropogenic and natural origin. Biotic responses to multiple stressors are often nonadditive (Paine et al. 1998). For example, if species’ tolerances to natural and anthropogenic stressors are negatively correlated, diver- sity can be severely reduced as a result of synergistic interactions (Vinebrooke et al. 2004). Acclimating or adapting to one stressor may increase organisms’ sensitivity to novel stressors, suggesting a potential cost associated with greater tolerance (Kashian et al. 2007). Increased tolerance requires more energy to repair the damages caused by a stressor, potentially limiting the ability of organisms to withstand additional environ- mental perturbation (Congdon et al. 2001). It is equally possible, however, that naturally stressful environments act as a strong environmental filter that eliminates sensitive specialists, leaving a subset of generalists able to tolerate the harsh conditions (Chase and Myers 2011). Such tolerant species may be less sensitive to novel stressors, and assemblages in naturally harsh conditions may thus be more tolerant to additional stress than those in more benign environments (Vinebrooke et al. 2004). The loss of biodiversity has been related to impaired ecosystem functioning in both terrestrial (Tilman et al. 2001) and aquatic environments (Covich et al. 2004), including freshwater fungi (Ba¨ rlocher and Corkum 2003, Dang et al. 2005). The autumnal input of riparian leaf litter is the primary source of energy and carbon in small woodland streams, and microbes and shredding invertebrates are the key players in the decomposition of leaf material. Low water pH and increased metal concentrations may have detrimental effects on micro- bial diversity and stream ecosystem functioning. The diversity of aquatic fungi generally declines with decreasing pH (Wood-Eggenschwiler and Ba¨ rlocher 1983, Baudoin et al. 2008) and leaf decomposition is typically reduced in acidic streams (Chamier 1987, Dangles et al. 2004a, Simon et al. 2009). However, Manuscript received 18 April 2014; revised 11 August 2014; accepted 25 August 2014. Corresponding Editor: W. V. Sobczak. 5 E-mail: mikko.tolkkinen@ymparisto.fi 672