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Fatty acids (FAs) are useful biomarkers in food web ecology because they are typically assimilated as a complete molecule and transferred into consumer tissue with minor or no modification, allowing the dietary routing between different trophic levels. However, the FA trophic marker approach is still hampered by the limited knowledge in lipid metabolism of the soil fauna. This study used entirely labelled palmitic acid ((13)C16:0, 99 atom%) as a tracer in fatty acid metabolism pathways of two widespread soil Collembola, Protaphorura fimata and Heteromurus nitidus. In order to investigate the fate and metabolic modifications of this precursor, a method of isotopologue profiling is presented, performed by mass spectrometry using single ion monitoring. Moreover, the upstream laboratory feeding experiment is described, as well as the extraction and methylation of dominant lipid fractions (neutral lipids, phospholipids) and the related formula and calculations. Isotopologue profiling does not only yield the overall (13)C enrichment in fatty acids derived from the (13)C labeled precursor but also produces the pattern of isotopologues exceeding the mass of the parent ion (i.e., the FA molecular ion M(+)) of each labeled FA by one or more mass units (M(+1), M(+2), M(+3), etc.). This knowledge allows conclusions on the ratio of dietary routing of an entirely consumed FA in comparison to de novo biosynthesis. The isotopologue profiling is suggested as a useful tool for evaluation of fatty acid metabolism in soil animals to disentangle trophic interactions.

Introduction Food-web models, which depict the trophic relationships between organisms within a community, form a powerful and versatile approach to study the relationships between community structure and ecosystem functioning. Although food-web models have recently been applied to a wide range of ecological studies (Memmott, 2009; Sanders et al., 2014), such approaches can be greatly improved by introducing high-resolution trophic information from empirical studies and experiments that realistically describe topological structure and energy flows (de Ruiter et al., 2005). Over the last decades major technological advances have been made in empirically characterizing trophic networks by describing, in detail, the connectedness and flows in food webs. Existing empirical techniques, such as stable isotope probing (SIP) (Layman et al., 2012), have been refined and new approaches have been created by combining methods, e.g., combining Raman spectroscopy or fatty acid analysis with SIP (Ruess et al., 2005a; Li et al., 2013). These empirical methods can provide insight into different aspects of food webs and together form an extensive toolbox to investigate trophic interactions. It is crucial to recognize the potential and limitations of a range of empirical approaches in order to choose the right method in the design of empirically based food-web studies. Empirically based food webs are generally classified according to the type of input information that is required. In the following lines we will provide an overview of four types of food-web model: connectedness webs, semi-quantitative webs, energy-flow webs, and functional webs. Paine (1980) introduced three of those webs, which are widely accepted and applied in food-web studies across ecosystems. We propose to add a fourth type of empirically based food web, the semi-quantitative web. All of these food webs have the same basic structure, but the conceptual webs differ in the type of trophic information they describe and represent (Figure 16.1). Connectedness webs (Figure 16.1a) define the basic structure of a food web by describing the food-web connections per se.

Subsurface microorganisms are essential constituents of the soil purification processes associated with groundwater quality. In particular, soil enzyme activity determines the biodegradation of organic compounds passing through the soil profile. Transects from surface soil to a depth of 3.5 m were investigated for microbial and chemical soil characteristics at two groundwater recharge sites and one control site. The functional diversity of the microbial community was analyzed via the activity of eight enzymes. Acid phosphomonoesterase was dominant across sites and depths, followed by l-leucine aminopeptidase and β-glucosidase. Structural [e.g. phospholipid fatty acid (PLFA) pattern] and functional microbial diversities were linked to each other at the nonwatered site, whereas amendment with nutrients (DOC, NO3−) by flooding uncoupled this relationship. Microbial biomass did not differ between sites, whereas microbial respiration was the highest at the watered sites. Hence, excess nutrients available due to artificial groundwater recharge could not compensate for the limitation by others (e.g. phosphorus as assigned by acid phosphomonoesterase activity). Instead, at a similar microbial biomass, waste respiration via overflow metabolism occurred. In summary, ample supply of carbon by flooding led to a separation of decomposition and microbial growth, which may play an important role in regulating purification processes during groundwater recharge

Subsurface microorganisms are crucial for contaminant degradation and maintenance of groundwater quality. This study investigates the microbial biomass and community composition [by phospholipid fatty acids (PLFAs)], as well as physical and chemical soil characteristics at woodland flooding sites of an artificial groundwater recharge system used for drinking water production. Vertical soil profiles to c. 4 m at two watered and one nonwatered site were analyzed. The microbial biomass was equal in watered and nonwatered sites, and considerable fractions (25-42%) were located in 40-340 cm depth. The microbial community structure differed significantly between watered and nonwatered sites, predominantly below 100 cm depth. Proportions of the bacterial PLFAs 16:1ω5, 16:1ω7, cy17:0 and 18:1ω9t, and the long-chained PLFAs 22:1ω9 and 24:1ω9 were more prominent at the watered sites, whereas branched, saturated PLFAs (iso/anteiso) dominated at the nonwatered site. PLFA community indices indicated stress response (trans/cis ratio), higher nutrient availability (unsaturation index) and changes in membrane fluidity (iso/anteiso ratio) due to flooding. In conclusion, water recharge processes led to nutrient input and altered environmental conditions, which resulted in a highly active and adapted microbial community residing in the vadose zone that effectively degraded organic compounds

Title: Dry weight of plant tissues. Description: Table of the dry weight of plant tissues at different growth stages (Root and Shoot flushes) for the respective treatments: control, P. penetrans, P. croceum and co-inoculation of P. penetrans and P. croceum. ANOVA with *, ** and *** with P

International audience; Food webs in agricultural systems are complex and trophic linkages are difficult to track using conventional methodologies. Here, we review three alternative approaches that allow empirical assessment of feeding interactions: DNA-based techniques, and stable isotope and fatty acid analyses. DNA-based methods, namely multiplex PCR and nextgeneration sequencing, allow identification of food types and host-parasitoid linkages, resulting in taxonomically highly resolved feeding networks. Stable isotopes and fatty acids reflect the assimilation of broader categories of resources, as metabolised into the consumers' tissue, together with the associated energy and nutrient fluxes in the food web. We discuss the strengths of the approaches but also highlight their limitations, providing practical advice on which technique is best suited to answer specific questions in examining food web interactions in agroecosystems. Future refinements of these techniques, especially when used in combination, could herald a new era in agricultural food web ecology, enabling management and environmental impact to be placed in the mechanistic context of trophic networks.

Food webs in agricultural systems are complex and trophic linkages are difficult to track using conventional methodologies. Here, we review three alternative approaches that allow empirical assessment of feeding interactions: DNA-based techniques, and stable isotope and fatty acid analyses. DNA-based methods, namely multiplex PCR and next-generation sequencing, allow identification of food types and host-parasitoid linkages, resulting in taxonomically highly resolved feeding networks. Stable isotopes and fatty acids reflect the assimilation of broader categories of resources, as metabolised into the consumers' tissue, together with the associated energy and nutrient fluxes in the food web. We discuss the strengths of the approaches but also highlight their limitations, providing practical advice on which technique is best suited to answer specific questions in examining food web interactions in agroecosystems. Future refinements of these techniques, especially when used in combination, could herald a new era in agricultural food web ecology, enabling management and environmental impact to be placed in the mechanistic context of trophic networks.