Isotopic evidences for microbiologically mediated and direct C input to soil compounds from three different leaf litters during their decomposition

M. Rubino 0 1 2 3 C. Lubritto 0 1 2 3 A. D'Onofrio 0 1 2 3 F. Terrasi 0 1 2 3 C. Kramer 0 1 2 3 G. Gleixner 0 1 2 3 M. F. Cotrufo 0 1 2 3 0 G. Gleixner Max Planck Institute for Biogeochemistry , Winzerlaer Strasse 10, 07745 Jena, Germany 1 C. Kramer Department of Analytical Chemistry and Reference Materials, Federal Institute for Materials Research and Testing (BAM) , Richard-Willstaetter-Strasse 11, 12489 Berlin, Germany 2 M. Rubino (&) C. Lubritto A. D'Onofrio F. Terrasi M. F. Cotrufo Department of Environmental Science, Second University of Naples , via Vivaldi, 81100 Caserta, Italy 3 Present Address: M. Rubino INRA, Unite UR1119 Unite Geochimie des Sols et des Eaux , F13545 Aix en Provence, France We show the potentiality of coupling together different compound-specific isotopic analyses in a laboratory experiment, where 13C-depleted leaf litter was incubated on a 13C-enriched soil. The aim of our study was to identify the soil compounds where the C derived from three different litter species is retained. Three 13C-depleted leaf litter (Liquidambar styraciflua L., Cercis canadensis L. and Pinus taeda L., d13CvsPDB & -43%), differing in their degradability, were incubated on a C4 soil (d13CvsPDB & -18%) under laboratory-controlled conditions for 8 months. At harvest, compound-specific isotope analyses were performed on different classes of soil compounds [i.e. phospholipids fatty acids (PLFAs), n-alkanes and soil pyrolysis products]. Linoleic acid (PLFA 18:2x6,9) was found to be very depleted in 13C (d13CvsPDB & from -38 to -42%) compared to all other PLFAs (d13CvsPDB & from -14 to -35%). Because of this, fungi were identified as the first among microbes to use the litter as source of C. Among n-alkanes, long-chain (C27-C31) n-alkanes were the only to have a depleted d13C. This is an indication that not all of the C derived from litter in the soil was transformed by microbes. The depletion in 13C was also found in different classes of pyrolysis products, suggesting that the litterderived C is incorporated in less or more chemically stable compounds, even only after 8 months decomposition. - The relentless increase of atmospheric CO2 concentration stimulated a large number of studies, in the past decades, devoted to the quantification of C fluxes between the atmosphere and the terrestrial biosphere. Terrestrial biosphere was proven to be an important sink for atmospheric CO2 (Janssens et al. 2003; Luo et al. 2007; Schimel et al. 2001), with soil organic matter (SOM) being the ultimate pool in which C can be stored for centuries or millennia (Schlesinger 1997; Yadav and Malanson 2007). Understanding C dynamics in soils is, thus, a main task for scientists working on the global C cycle. SOM is formed from the decomposition of dead plant and animal material (e.g. wood, roots, leaf litter, animal residues, etc.) due to the action of abiotic (e.g. wind, rain, solar radiation) and biotic (e.g. soil macrofauna and microbial community) factors (Aber et al. 1990; Spaccini et al. 2000). In particular, decomposition of leaf litter is one of the major processes determining SOM formation (Kuzyakov and Domanski 2000; Liski et al. 2002). Plant biomass is a complex mixture of several polymers of very different structure (Gleixner et al. 2001). After plant death, this mixture undergoes oxidative and hydrolytic degradation by micro-organisms, accompanied by secondary structural changes. Several different steps are used in describing SOM formation, including loss of labile compounds and CO2 and numerous reactions of biotransformation of recalcitrant compounds. On the other side, undecomposed litter fragments may directly enter the soil and may prime aggregate formation (Six and Jastrow 2002). This suggests that part of litter-derived C could enter SOM without any participation by microbes. The relative importance of factors governing these processes is poorly understood. In particular, a key question is how substrate quality affects the transformations of plant residues into stable SOM (Corbeels 2001). Mechanisms of SOM formation and stabilization have mainly been studied investigating the turnover and stability of SOM as a bulk (Zech et al. 1997). This just reveals mean pattern of soil C transformation, but it does not give any information about the phenomenon acting at the molecular level. Indeed, physical fractionation coupled with SOM labelling allowed progress in understanding changes in soil C stores, especially, the stabilization of SOM due to the interaction with the mineral part of soil (Del Galdo et al. 2003). However, to fully clarify soil C stabilization processes at the molecular level, the chemical nature of soil organic C has to be studied as well as the flow rates of each different group of compounds forming SOM need to be quantified (Gleixner et al. 2001; Lichtfouse 2000). This study focuses on three different groups of compounds: Phospholipids fatty acids Since micro-organisms play a fundamental role in the transformation of organic residues in SOM, the study of microbial community in soil is necessary to fully understand the dynamics of C during SOM formation. To characterize soil microbial communities, the fatty acids of phospholipids belonging to the cell membranes of microorganisms are routinely used (Allison et al. 2005; Zelles 1999). Phospholipids are essential membrane components of all living cells: because of their fast metabolization rate on cell death (Tollefson and McKercher 1983), phospholipids fatty acids (PLFAs) reflect the structure of viable soil microbial community. The PLFAs in soil are derived from a wide range of bacteria, fungi and invertebrates and present a complex mixture that is a challenge both to analyze and interpret (Zelles 1999). In many cases, specific types of fatty acids predominate in a given taxon so that they can be used to distinguish between the different groups of micro-organisms. Yet, not all the C derived from plant material is processed by micro-organisms. Recalcitrant compounds present in plants (i.e. lipids, waxes, etc.) can be transported into the soil without being involved in microbial metabolism. For example, n-alkanes are found in leaves for a number of reasons; for instance, to protect the exposed part of outer cells against drought, plant leaves are covered with nonpermeable, hydrophobic compounds (Gleixner et al. 2001). Most of studies on n-alkanes abundance are conducted on leaves or sediments samples: n-alkanes are used as biomarkers for the paleoenvironment (Lockheart et al. 1997; Naraoka and Ishiwatari 1999). In SOM studies, the extraction of n-alkanes from SOM gives the opportunity to test the presence of compounds recalcitrant to microbial attack, derived from wax lipids of decomposing leaf litter (Cayet and Lichtfouse 2001; Lichtfouse et al. 1995a, 1994). Pyrolysis products The use of Curie-point pyrolysis for the thermal degradation of organic substances gives a reliable and reproducible way to obtai (...truncated)