1. Soil as a Biological System

Nannipieri Paolo

Soil is an unique biological system with an abundant microflora and a very high microbial diversity. The space occupied by microorganisms is very low because only few microsites have the right set of conditions suitable for microbial life. Surface-reactive particles can adsorb important biological molecules, such as DNA and enzymes, which become resistant to microbial degradation and thus genes are preserved and extracellular enzymes can be reactive when conditions are not suitable for microbial activity. Most soil functions mainly depend on microbial activity but soil fauna can accelerate microbial processes and complete food webs in soil. Omics techniques, such as metagenomics, metatranscriptomics and proteomics, have several problems when applied to soil. However, if used in a complementary way these techniques are promising for providing an integrated picture of the relationship between composition and activity of soil microflora.

2. Functional Genomics Analysis of Key Bacterial Traits Involved in Rhizosphere Competence during Microbial-Host Interactions

Matthieu Barret, John P. Morrissey and Fergal O'Gara

The rhizosphere is a nutrient rich environment, where numerous interactions between plant and microorganisms occur, ranging from mutualism to parasitism. The enrichment of specific microbial populations in the rhizosphere is dependent on the capability of these micro-organisms to utilize root exudates, to effectively colonize the root surface and to interact or compete with other micro-organisms. Analysis of the rhizospheric communities incorporating both established techniques, and recently developed "omic technologies" can now facilitate investigations into the molecular basis underpinning the establishment of plant-microbial interactomes in the rhizosphere. Therefore, the aim of this chapter is to present an overview of bacterial functions enriched in the rhizosphere of different plant species using data obtained from several functional genomics analyses.

3. Soil Metagenomics: Potential Applications and Methodological Problems

Jan Dirk van Elsas, Mariana Silvia Cretoiu, Anna Maria Kielak and Francisco Dini-Andreote

Metagenomics has been defined as the study of the collective genomes of the microbiota in given habitat. Soil offers a huge microbial diversity and the use of metagenomics approaches will allow a deeper understanding of soil microbial diversity and function. The two areas, phylogenetically-based diversity and functional gene based function, are complementary and may be used side-by-side in order to allow a better understanding of the living soil. Moreover, genes for relevant functions can be cloned into suitable vectors, after which they can be studied and possibly explored for biotechnological purposes. Thus, opportunities for novel product discovery via metagenomics are rapidly rising. However, there are caveats in what metagenomics techniques can tell us about the soil environment and its functioning, and also in the chances of successful exploration of soil. In this chapter, we review the developments in the metagenomics-based exploitation and exploration of soil and examine how soil metagenomics can enhance our vision about natural functioning and exploration for biotechnological novelty. One major issue, the need for advanced bioinformatics tools, is stressed. We conclude that the rich microbiota of soil offers an astonishing big playground for metagenomics, but that methodological and conceptual problems still hamper its full exploitation.

4. Screening Phylogenetic and Functional Marker Genes in Soil Microbial Ecology

Sotirios Vasileiadis, Edoardo Puglisi, PierSandro Cocconcelli and Marco Trevisan

Many of the ecosystem services are soil associated with microbes playing a predominant role. Nevertheless, our current knowledge of microbial contribution to ecosystem processes is still limited, partly because in the past centuries research was mostly based on culture-dependent methods, being oblivious of the vast un-cultivable microbial majority as proven during the last decades. Current molecular biology advances provide us with the ability to screen for microbial identities or functions by targeting marker genes in nucleic acid extracts of environmental samples, therefore partly bypassing previous methodological limitations. Topics addressed here aim at providing an overview of methodologies and concepts related to marker gene screening from environmental samples. Such are the description of marker gene categories, examples of their use in soil environments and the description of marker gene screening state-of-the-art methodologies and specifications. Finally we will exemplify the use of late methodologies for the case of the bacterial small ribosomal subunit screening in soil environments.

5. Soil Metatranscriptomics

Yongkyu Kim, Carl-Eric Wegner and Werner Liesack

Metatranscriptomics is defined as the analysis of microbial community gene expression in a particular environment, as opposed to metagenomics which is the study of the genomic content of entire microbial communities. Massively parallel sequencing of RNA is the key component of metatranscriptomics. The analysis of enriched mRNA has the potential to discover novel genes and to uncover functional adaptations of microbial communities to local environmental conditions. Alternatively, total RNA may be used for analysis. This approach provides insight into the taxonomic composition of microbial communities. The subject of this review is soil metatranscriptomics. We discuss the experimental and bioinformatic workflow that can be applied to the metatranscriptomic analysis of soil microbial communities. Since research in the field of soil metatranscriptomics is still in its infancy, we also review the recent advances in marine metatranscriptomics.

6. Soil Proteomics

Giancarlo Renella, Laura Giagnoni, Mariarita Arenella and Paolo Nannipieri

Proteomics is a potent post genomic approach with the potential to interrogate natural complex systems such as soils. However, the great potentials of soil proteomics are currently limited by either the complexity of the soil matrix which is reactive, structured, teeming with microbial communities which are at the same time extremely diverse, in heterogeneous physiological state and normally poorly characterized. Taken together, these soil features pose problems of protein sampling, extraction and purification. This chapter, though not exhaustive, aims at illustrate the main approaches and achievements in soil proteomics and indicate some future directions for further developments soil proteomics.

7. Soil Volatile Organic Compounds as Tracers for Microbial Activities in Soils

Heribert Insam

Soil volatilomics is a scientific discipline that deals with the multitude of volatile organic compounds produced, stored or degraded in soils. These compounds may be of plant or microbial origin, or they may enter or leave the atmosphere through diffusion and convection processes. The present article focuses on soil volatile organic compounds of which many are produced by microorganisms. The challenges and chances of using such VOCs for characterizing soil microbial communities are addressed, and several examples on microbe-to microbe and microbe-plant interactions involving VOCs are given. Also, some problems concerning the measurement of these VOCs are addressed.

8. Proteogenomics: a New Integrative Approach for a Better Description of Protein Diversity Found in Soil Microflora

Céline Bland and Jean Armengaud

Proteogenomics is a relatively recent field at the junction of genomics and proteomics which consists of refining the annotation of the genome of model organisms with the help of high-throughput proteomic data. To get a comprehensive view on how a given microorganism functions, elucidating its genome is a prerequisite. Since the first complete genome of a cellular organism was sequenced, that of Haemophilus influenza in 1995, an impressive catalogue of genomes has been reported. Because automatic annotation software are not yet sufficiently confident, the annotation process should be complemented with experimental data. Alongside the development of high-throughput sequencing techniques, important innovations in tandem mass spectrometry and proteomic approaches have led to the possibility of analyzing thousands of proteins from a given sample. Proteogenomics has proved to be helpful in discovering new genes that were forgotten by automatic annotation software, identifying the true translational initiation codon of coding domain sequences and characterizing maturation events at the protein level, such as signal peptide processing. Consequently, proteogenomics is now proposed at the earliest stage of a genome sequencing project as exemplified by the Deinococcus deserti genome, for which unexpected results, such as the reversal of gene sequences in different bacteria or the use of non-canonical start codons for translation in Deinococcus species, are only some of the numerous corrections obtained by proteogenomics. Because an important issue is the identification of the correct translational start codons, we have pointed out the need for developing N-terminal-oriented strategies to reveal experimentally the precise sites of translation initiation. Today, a better description of the protein universe found in soil microflora can be achieved if proteogenomics is performed on a given set of representative models from this environment.

9. Analysis of Soil Metagenomes using the MEtaGenome ANalyzer (MEGAN)

Daniel H. Huson and Nico Weber

Soil is one of the most dense and diverse habitats of microorganisms, and metagenomics is a key technique for studying this environment. Given a dataset of DNA sequencing reads obtained from an environmental sample, the program MEGAN (Metagenome Analyzer) can be used to interactively perform an analysis and comparison of the data, both on a taxonomic and functional level. The program attempts to place all reads into the NCBI taxonomy and to map them to SEED functional roles or KEGG orthology groups. To compare various datasets, MEGAN allows the user to load multiple samples at once and combine them into a single comparison document. Multiple views makes it easy to inspect assignments to the different nodes. An alignment viewer is supplied to explore reference-guided multiple alignments of reads. The program supports various input formats for loading data and can export analysis results in different text-based and graphical formats. The program is designed to work with very large datasets. It is written in Java and installers for the three major computer operating systems are available from https://www-ab.informatik.uni-tuebingen.de.

10. Classical Techniques versus Omics Approaches

David D. Myrold and Paolo Nannipieri

This volume presents the state-of-the-art of omics in soil science, a field that is advancing rapidly on many fronts. The various omics approaches hold much promise but also await further refinement before they are ready for widespread adaptation. One way to judge their readiness is to compare them to methods that have become standards for soil microbiology research. Methods become standards because they provide useful information quickly and inexpensively. There is no question that omics can provide useful information, some of which cannot be obtained with traditional techniques, and integration of omics methods may provide insights into ecosystem functioning. In particular, the potential for omics to provide comprehensive coverage of genes and genes products make them well-suited for the study of general soil microbiological phenomena, such as decomposition, response to water stress, etc.