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Simulated atmospheric N deposition alters fungal community composition and suppresses ligninolytic gene expression in a northern hardwood forest.

Edwards IP, Zak DR, Kellner H, Eisenlord SD, Pregitzer KS - PLoS ONE (2011)

Bottom Line: Our results indicate that across four Acer-dominated forest stands spanning a 500-km transect, community-scale expression of the cellulolytic gene cbhI under elevated N deposition did not differ significantly from that under ambient levels of N deposition.Simulated N deposition increased the proportion of basidiomycete sequences recovered from forest floor, whereas the proportion of ascomycetes in the community was significantly lower under elevated N deposition.Our results suggest that chronic atmospheric N deposition may lower decomposition rates through a combination of reduced expression of ligninolytic genes such as lcc, and compositional changes in the fungal community.

View Article: PubMed Central - PubMed

Affiliation: School of Natural Resources and Environment, University of Michigan, Ann Arbor, Michigan, United States of America. iedwards@umich.edu

ABSTRACT
High levels of atmospheric nitrogen (N) deposition may result in greater terrestrial carbon (C) storage. In a northern hardwood ecosystem, exposure to over a decade of simulated N deposition increased C storage in soil by slowing litter decay rates, rather than increasing detrital inputs. To understand the mechanisms underlying this response, we focused on the saprotrophic fungal community residing in the forest floor and employed molecular genetic approaches to determine if the slower decomposition rates resulted from down-regulation of the transcription of key lignocellulolytic genes, by a change in fungal community composition, or by a combination of the two mechanisms. Our results indicate that across four Acer-dominated forest stands spanning a 500-km transect, community-scale expression of the cellulolytic gene cbhI under elevated N deposition did not differ significantly from that under ambient levels of N deposition. In contrast, expression of the ligninolytic gene lcc was significantly down-regulated by a factor of 2-4 fold relative to its expression under ambient N deposition. Fungal community composition was examined at the most southerly of the four sites, in which consistently lower levels of cbhI and lcc gene expression were observed over a two-year period. We recovered 19 basidiomycete and 28 ascomycete rDNA 28S operational taxonomic units; Athelia, Sistotrema, Ceratobasidium and Ceratosebacina taxa dominated the basidiomycete assemblage, and Leotiomycetes dominated the ascomycetes. Simulated N deposition increased the proportion of basidiomycete sequences recovered from forest floor, whereas the proportion of ascomycetes in the community was significantly lower under elevated N deposition. Our results suggest that chronic atmospheric N deposition may lower decomposition rates through a combination of reduced expression of ligninolytic genes such as lcc, and compositional changes in the fungal community.

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Triplot based on a Canonical Correspondence Analysis showing the relationships between samples of a fungal community developing under ambient conditions of N deposition (open squares) and under conditions of simulated elevated N depostion (closed squares), the relative abundance of the 12 most widespread fungal Operational Taxonomic Units (OTUs, black circles), and total N depostion (vector) in the forest floor of a maple dominated hardwood ecosystem.The primary axis accounts for 22% of the variance in OTU relative abundances; the second axis a further 37%. The relationship between fungal relative abundances and total N is not significant (Monte Carlo P = 0.52). OTU codes correspond to the phylogenies (Fig. 2, Fig. 3): OceanaOTU_1, Typhula sp; OceanaOTU_4, Mycena sp; OceanaOTU_11, Entoloma sp; OceanaOTU_12, Athelia sp.; OceanaOTU_13, Gomphales sp.; OceanaOTU_15, Ceratosebacina sp.; OceanaOTU_16, Ceratobasidium sp.; OceanaOTU_18, Sistotrema sp.; OceanaOTU_23, unidentified Ascomycota; OceanaOTU_36, Leotiomycete sp.; OceanaOTU_41, Leotiomycete sp.; OceanaOTU_42, Leotiomycete sp.
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pone-0020421-g004: Triplot based on a Canonical Correspondence Analysis showing the relationships between samples of a fungal community developing under ambient conditions of N deposition (open squares) and under conditions of simulated elevated N depostion (closed squares), the relative abundance of the 12 most widespread fungal Operational Taxonomic Units (OTUs, black circles), and total N depostion (vector) in the forest floor of a maple dominated hardwood ecosystem.The primary axis accounts for 22% of the variance in OTU relative abundances; the second axis a further 37%. The relationship between fungal relative abundances and total N is not significant (Monte Carlo P = 0.52). OTU codes correspond to the phylogenies (Fig. 2, Fig. 3): OceanaOTU_1, Typhula sp; OceanaOTU_4, Mycena sp; OceanaOTU_11, Entoloma sp; OceanaOTU_12, Athelia sp.; OceanaOTU_13, Gomphales sp.; OceanaOTU_15, Ceratosebacina sp.; OceanaOTU_16, Ceratobasidium sp.; OceanaOTU_18, Sistotrema sp.; OceanaOTU_23, unidentified Ascomycota; OceanaOTU_36, Leotiomycete sp.; OceanaOTU_41, Leotiomycete sp.; OceanaOTU_42, Leotiomycete sp.

Mentions: We examined the effect of simulated N deposition on fungal community composition at the most southerly of our sites (site D, Table 1), in which consistently lower levels of fungal cbhI and lcc gene expression under simulated N deposition were observed in 2007 and 2009. We recovered a species-rich and diverse active fungal community from the forest floor of this site (Figs 2, 3). Although we recovered a higher diversity of ascomycete than basidiomycete sequences, basidiomycete sequences clearly dominated the rRNA gene library, and, moreover, represented 66% of the most widespread species at this site (Fig. 4). Lignin decomposition is principally associated with basidiomycete species, and Clitocybe, Collybia, Marasmius, and Mycena species are well known for their ligninolytic capability and are commonly recovered from forest floor environments [13], [30], [31]. We recovered taxa from Mycena, Clitocybe, Crepidotus and Clitopilus (Fig. 3), as well as Entoloma and Typhula that are also most likely saprotrophs, and these represented approximately half of the basidiomycete diversity. The most abundant and widespread basidiomycete taxa however, were members of the Cantharellales, Ceratobasidiales and Atheliales. The nutritional mode of these resupinate taxa is unclear; they are most likely saprotrophs, but mycorrhizal and parasitic life-strategies are also known from these groups [36]. A Gomphalian taxon recovered under both ambient and simulated N conditions is probably mycorrhizal, and Tremellomycete yeasts were also recovered under both treatments. Ascomycetes were predominantly recovered from the Dothideomycetes, Sordariomycetes and Leotiomycetes, although most remained unidentified at better than ordinal level (Fig. 2). This was especially true within the Leotiomycetes (Fig. 2). Although simulated N deposition had no significant effect on the relative abundance of the more widespread basidiomycete or ascomycete taxa, the proportion of ascomycete species in the active community was nevertheless significantly reduced under simulated N deposition. The consequences of the apparent reduction in ascomycete diversity during the later stages of litter decomposition are largely unknown; lcc is broadly distributed among the Basidiomycota, and less broadly among the Ascomycota and other fungi [35]. With the exception of some species of Sordariomycetes (e.g., Xylaria), saprotrophic ascomycetes isolated from forest soils appear to be primarily cellulolytic and chitinolytic organisms, rather than agents of lignin degradation [14], [15], [16]. Indeed, ascomycete-derived cellolulytic and chitinolytic genes have been recovered in a previous transcriptomic analysis of this ecosystem [37]. The reduced transcription of lcc that we observed may then be the result of basidiomycetes expressing less lcc as a physiological response to higher N availability; however, the extent of laccase distribution among ascomycetes is poorly understood [35]. Leotiomycete ecologies are generally considered to be plant-based, include pathogenic, endophytic, saprotrophic and mycorrhizal life strategies [38], and laccase positive species (e.g. Botryotinia fukeliana) are known [39]. Similarly, within the Dothideomycetes and Sordariomycetes, many of the taxa we recovered appear to place with plant pathogens such as Mycosphaerella, Neonectria and the Cryphonectriaceae. Species within these groups may use laccase as an “attack” enzyme during infection, and subsequently to break down senescent plant cells [39]. Although these taxa were sparsely distributed across the site, the possibility that their absence from the simulated N deposition treatment is in part responsible for the decline in lcc gene expression and laccase activity cannot be discounted. Although ligninolytic basidiomycetes are often considered the primary agents of late-stage litter decomposition [30], [35], [40], our results emphasize the need for further studies to connect functional genes recovered in the transcriptome to the species active in the community.


Simulated atmospheric N deposition alters fungal community composition and suppresses ligninolytic gene expression in a northern hardwood forest.

Edwards IP, Zak DR, Kellner H, Eisenlord SD, Pregitzer KS - PLoS ONE (2011)

Triplot based on a Canonical Correspondence Analysis showing the relationships between samples of a fungal community developing under ambient conditions of N deposition (open squares) and under conditions of simulated elevated N depostion (closed squares), the relative abundance of the 12 most widespread fungal Operational Taxonomic Units (OTUs, black circles), and total N depostion (vector) in the forest floor of a maple dominated hardwood ecosystem.The primary axis accounts for 22% of the variance in OTU relative abundances; the second axis a further 37%. The relationship between fungal relative abundances and total N is not significant (Monte Carlo P = 0.52). OTU codes correspond to the phylogenies (Fig. 2, Fig. 3): OceanaOTU_1, Typhula sp; OceanaOTU_4, Mycena sp; OceanaOTU_11, Entoloma sp; OceanaOTU_12, Athelia sp.; OceanaOTU_13, Gomphales sp.; OceanaOTU_15, Ceratosebacina sp.; OceanaOTU_16, Ceratobasidium sp.; OceanaOTU_18, Sistotrema sp.; OceanaOTU_23, unidentified Ascomycota; OceanaOTU_36, Leotiomycete sp.; OceanaOTU_41, Leotiomycete sp.; OceanaOTU_42, Leotiomycete sp.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3119081&req=5

pone-0020421-g004: Triplot based on a Canonical Correspondence Analysis showing the relationships between samples of a fungal community developing under ambient conditions of N deposition (open squares) and under conditions of simulated elevated N depostion (closed squares), the relative abundance of the 12 most widespread fungal Operational Taxonomic Units (OTUs, black circles), and total N depostion (vector) in the forest floor of a maple dominated hardwood ecosystem.The primary axis accounts for 22% of the variance in OTU relative abundances; the second axis a further 37%. The relationship between fungal relative abundances and total N is not significant (Monte Carlo P = 0.52). OTU codes correspond to the phylogenies (Fig. 2, Fig. 3): OceanaOTU_1, Typhula sp; OceanaOTU_4, Mycena sp; OceanaOTU_11, Entoloma sp; OceanaOTU_12, Athelia sp.; OceanaOTU_13, Gomphales sp.; OceanaOTU_15, Ceratosebacina sp.; OceanaOTU_16, Ceratobasidium sp.; OceanaOTU_18, Sistotrema sp.; OceanaOTU_23, unidentified Ascomycota; OceanaOTU_36, Leotiomycete sp.; OceanaOTU_41, Leotiomycete sp.; OceanaOTU_42, Leotiomycete sp.
Mentions: We examined the effect of simulated N deposition on fungal community composition at the most southerly of our sites (site D, Table 1), in which consistently lower levels of fungal cbhI and lcc gene expression under simulated N deposition were observed in 2007 and 2009. We recovered a species-rich and diverse active fungal community from the forest floor of this site (Figs 2, 3). Although we recovered a higher diversity of ascomycete than basidiomycete sequences, basidiomycete sequences clearly dominated the rRNA gene library, and, moreover, represented 66% of the most widespread species at this site (Fig. 4). Lignin decomposition is principally associated with basidiomycete species, and Clitocybe, Collybia, Marasmius, and Mycena species are well known for their ligninolytic capability and are commonly recovered from forest floor environments [13], [30], [31]. We recovered taxa from Mycena, Clitocybe, Crepidotus and Clitopilus (Fig. 3), as well as Entoloma and Typhula that are also most likely saprotrophs, and these represented approximately half of the basidiomycete diversity. The most abundant and widespread basidiomycete taxa however, were members of the Cantharellales, Ceratobasidiales and Atheliales. The nutritional mode of these resupinate taxa is unclear; they are most likely saprotrophs, but mycorrhizal and parasitic life-strategies are also known from these groups [36]. A Gomphalian taxon recovered under both ambient and simulated N conditions is probably mycorrhizal, and Tremellomycete yeasts were also recovered under both treatments. Ascomycetes were predominantly recovered from the Dothideomycetes, Sordariomycetes and Leotiomycetes, although most remained unidentified at better than ordinal level (Fig. 2). This was especially true within the Leotiomycetes (Fig. 2). Although simulated N deposition had no significant effect on the relative abundance of the more widespread basidiomycete or ascomycete taxa, the proportion of ascomycete species in the active community was nevertheless significantly reduced under simulated N deposition. The consequences of the apparent reduction in ascomycete diversity during the later stages of litter decomposition are largely unknown; lcc is broadly distributed among the Basidiomycota, and less broadly among the Ascomycota and other fungi [35]. With the exception of some species of Sordariomycetes (e.g., Xylaria), saprotrophic ascomycetes isolated from forest soils appear to be primarily cellulolytic and chitinolytic organisms, rather than agents of lignin degradation [14], [15], [16]. Indeed, ascomycete-derived cellolulytic and chitinolytic genes have been recovered in a previous transcriptomic analysis of this ecosystem [37]. The reduced transcription of lcc that we observed may then be the result of basidiomycetes expressing less lcc as a physiological response to higher N availability; however, the extent of laccase distribution among ascomycetes is poorly understood [35]. Leotiomycete ecologies are generally considered to be plant-based, include pathogenic, endophytic, saprotrophic and mycorrhizal life strategies [38], and laccase positive species (e.g. Botryotinia fukeliana) are known [39]. Similarly, within the Dothideomycetes and Sordariomycetes, many of the taxa we recovered appear to place with plant pathogens such as Mycosphaerella, Neonectria and the Cryphonectriaceae. Species within these groups may use laccase as an “attack” enzyme during infection, and subsequently to break down senescent plant cells [39]. Although these taxa were sparsely distributed across the site, the possibility that their absence from the simulated N deposition treatment is in part responsible for the decline in lcc gene expression and laccase activity cannot be discounted. Although ligninolytic basidiomycetes are often considered the primary agents of late-stage litter decomposition [30], [35], [40], our results emphasize the need for further studies to connect functional genes recovered in the transcriptome to the species active in the community.

Bottom Line: Our results indicate that across four Acer-dominated forest stands spanning a 500-km transect, community-scale expression of the cellulolytic gene cbhI under elevated N deposition did not differ significantly from that under ambient levels of N deposition.Simulated N deposition increased the proportion of basidiomycete sequences recovered from forest floor, whereas the proportion of ascomycetes in the community was significantly lower under elevated N deposition.Our results suggest that chronic atmospheric N deposition may lower decomposition rates through a combination of reduced expression of ligninolytic genes such as lcc, and compositional changes in the fungal community.

View Article: PubMed Central - PubMed

Affiliation: School of Natural Resources and Environment, University of Michigan, Ann Arbor, Michigan, United States of America. iedwards@umich.edu

ABSTRACT
High levels of atmospheric nitrogen (N) deposition may result in greater terrestrial carbon (C) storage. In a northern hardwood ecosystem, exposure to over a decade of simulated N deposition increased C storage in soil by slowing litter decay rates, rather than increasing detrital inputs. To understand the mechanisms underlying this response, we focused on the saprotrophic fungal community residing in the forest floor and employed molecular genetic approaches to determine if the slower decomposition rates resulted from down-regulation of the transcription of key lignocellulolytic genes, by a change in fungal community composition, or by a combination of the two mechanisms. Our results indicate that across four Acer-dominated forest stands spanning a 500-km transect, community-scale expression of the cellulolytic gene cbhI under elevated N deposition did not differ significantly from that under ambient levels of N deposition. In contrast, expression of the ligninolytic gene lcc was significantly down-regulated by a factor of 2-4 fold relative to its expression under ambient N deposition. Fungal community composition was examined at the most southerly of the four sites, in which consistently lower levels of cbhI and lcc gene expression were observed over a two-year period. We recovered 19 basidiomycete and 28 ascomycete rDNA 28S operational taxonomic units; Athelia, Sistotrema, Ceratobasidium and Ceratosebacina taxa dominated the basidiomycete assemblage, and Leotiomycetes dominated the ascomycetes. Simulated N deposition increased the proportion of basidiomycete sequences recovered from forest floor, whereas the proportion of ascomycetes in the community was significantly lower under elevated N deposition. Our results suggest that chronic atmospheric N deposition may lower decomposition rates through a combination of reduced expression of ligninolytic genes such as lcc, and compositional changes in the fungal community.

Show MeSH
Related in: MedlinePlus