Molecular composition of straw and green manure at different stages of decomposition

The dominance of a single but shifting molecular signature in the wheat straw before and after incubation for both alkyl ratio and C/N is consistent with studies reporting decreasing proportions of straw carbon as carbohydrate and increasing proportions of aromatic compounds during straw decomposition (Cogle et al., 1989; Gao et al., 2016). The alkyl ratio increase indicates the decomposition of straw compounds, likely by a shift from substituted aliphatic alcohols and ethers to unsubstituted C in paraffinic structures (Wilson et al., 1983; Kögel-Knabner, 2002), whereas the decrease in C/N is consistent with a loss of C caused by microbial respiration (Geissen & Brümmer, 1999) and the preservation of N by decomposers in their tissues and by-products. However, the greater heterogeneity of both alkyl ratio and C/N of straw at the end of the incubation (Table 2) indicates that decomposition was spatially heterogeneous and that some regions of the straw amendment decomposed faster than others. Although this could be explained by preferential decomposition of specific plant-derived structural compounds by microorganisms during the first stages of decomposition (Golchin et al., 1997), this spatially heterogeneous degradation of the chemically homogeneous straw amendment is more likely attributable to abiotic factors, i.e. the microscale conditions of the microbial habitats surrounding POM that regulates soil moisture and microbial accessibility of the OM (Dungait et al., 2012).
In contrast to the clear shift in POM composition during incubation of straw, changes in the composition of green manure were less clear, a result of the greater heterogeneity of green manure before and after incubation. Indeed, the general increase in C/N and decrease in alkyl ratio after incubation are contrary to typical changes induced by decomposition of organic materials (Golchin et al., 1994; Baldock et al., 1997; Stone et al., 2001; Kögel-Knabner, 2002). We attribute this to a shift in the exposed surface of the material before and after incubation. The green manure material contained highly different types of plant tissues, such as leaves, woody parts and bark material and was composted before we used it. While pieces of bark and partially decomposed green manure were observed at the beginning of the experiment, these particles may have significantly peeled off and dispersed during the incubation, exposing internal woody structures of green manure material. To better explain the changes in green manure chemistry, further research into the molecular composition of the various types of organic material present in the mixture and their decomposition is required.

Straw amendments are more decomposed in topsoils than in subsoils

The lower C/N and larger alkyl ratio of straw particles (Table 2), and the stronger shifts in distribution in the topsoil at the end of the incubation indicate a more advanced degradation than in the subsoil (Figure 2), which we attribute to the differences in the initial conditions of the two soil matrices that drive decomposition processes and rates. Generally, topsoils accommodate more diverse microbial communities and more microbial biomass than subsoils (Taylor et al., 2002), and contain more SOC. The production of dissolved organic matter (DOM) is correlated with high SOC contents and abundance of microorganisms (Guigue et al., 2015) and larger quantities of DOM are measured in topsoils than in subsoils (Kalbitz et al., 2000; Kaiser & Kalbitz, 2012). DOM is is composed of labile and energy-rich compounds easily converted either by microbial resynthesis or respiration (Strauss & Lamberti, 2002; Guggenberger & Kaiser, 2003; Kaiser & Kalbitz, 2012). This fuels the production of extracellular hydrolytic enzymes that contribute to POM decomposition (Berg & McClaugherty, 2014), and enables the faster decomposition of the added fresh organic matter in topsoils than subsoils.

C/N spatial distribution and coupling of C and N cycling

The C/N of above 40 after incubation (Figure 2) are well above typical lower bounds for SOM in central European agricultural soils, which rarely exceed C/N of 15 (Matschullat et al., 2018). This indicates that the decomposition processes of the POM was not completed during the period of incubation and we assume that C/N of POM would continue to decrease if the incubation was prolonged. The initially lower C/N of green manure in the topsoil is likely the result of the prior composting of this material. It may also result from a greater proportion of N-rich compounds, such as bark, on the surface of the composted material when it was introduced to the topsoil for incubation.
The C/N of the POM fraction has been reported as being greater than 20 in soils under cropland, forest and grassland (Warren & Whitehead, 1988; Meijboom et al., 1995; Hassink, 1995; Baldock et al., 2003; Bimüller et al., 2014). In addition, C/N ranging from 20 to 25 are generally accepted to be the thresholds for the shift of microbial N immobilisation to N mineralisation (Nicolardot et al., 2001; Robertson & Groffman, 2015) that stabilises the C/N by balancing C and N losses and further leading to a rapid mineralisation of POM. Straw decomposition is reported to start rapidly before slowing down during the process (de Willingen et al., 2008), with decomposition mostly fuelled by hemicellulose while more recalcitrant ligneous materials are decomposed slower. Thus, the relative contribution of ligneous material increases during decomposition (Cogle et al., 1989; Gao et al., 2016), concurrent with decreases in the decomposition rate, corresponding to the preservation of remaining straw residues with a high C/N. Furthermore, lignin macromolecules decomposition is promoted when the macrostructures are firstly shredded by macro fauna (Scheu, 1992). The absence of such organisms in our experiment likely supported the preservation of large lignin-like moieties with high C/N.

Conclusion

The coupling of VNIR imaging with machine learning modelling was successful for the sub-millimetre scale mapping of molecular composition of various types of POM at distinct decomposition stages. Our novel approach based on model averaging of a random forest with an ensemble ANN overcomes issues relating to training of a single ANN and extrapolation of results beyond calibration ranges, presenting a new direction in machine learning and spectroscopy in soils. With this technique, we demonstrated the spatially heterogeneous changes in alkyl ratio and C/N of POM during decomposition with an overall increase in alkyl ratio and decrease in C/N of straw as a result of decomposition. The changes of the more heterogeneous green manure showed an opposite trend, likely associated with the preferential decomposition of N-rich bark tissues together with the preservation of less decomposable C-rich plant residues enriched in ligneous material. The decomposition of both straw and green manure was retarded in the subsoil compared to topsoil, as highlighted by smaller changes in the POM in the subsoil after the 6-month incubation in the subsoil. The visualisation approach presented has a great potential for applications aiming to investigate the spatial heterogeneity in molecular changes of organic particles during decomposition, and can help to disentangle the concurrent roles of accessibility and recalcitrance during the first steps of the decomposition cascade of organic matter in soils.

Acknowledgements

This work was financially supported by the German Federal Ministry for Education and Research, through the BonaRes initiative (BMBF, grant FKZ 031B0026B, Soil3 project and 031B0511C BonaRes centre). As a part of the BonaRes initiative, the authors would like to thank all contributors to the Soil3 project and the BonaRes centre.