16S ribosomal RNA gene (rDNA) amplicon analysis remains the standard approach for the cultivation-independent investigation of microbial diversity. The accuracy of these analyses depends strongly on the choice of primers. The overall coverage and phylum spectrum of 175 primers and 512 primer pairs were evaluated in silico with respect to the SILVA 16S/18S rDNA non-redundant reference dataset (SSURef 108 NR). Based on this evaluation a selection of 'best available' primer pairs for Bacteria and Archaea for three amplicon size classes (100-400, 400-1000, ≥ 1000 bp) is provided. The most promising bacterial primer pair (S-D-Bact-0341-b-S-17/S-D-Bact-0785-a-A-21), with an amplicon size of 464 bp, was experimentally evaluated by comparing the taxonomic distribution of the 16S rDNA amplicons with 16S rDNA fragments from directly sequenced metagenomes. The results of this study may be used as a guideline for selecting primer pairs with the best overall coverage and phylum spectrum for specific applications, therefore reducing the bias in PCR-based microbial diversity studies.

Evaluation of general 16S ribosomal RNA gene PCR primers for classical and next-generation sequencing-based diversity studies

https://web.science.uu.nl/pmi/publications/PDF/2012/TiPS-Berendsen-2012.pdf

The rhizosphere microbiome and plant health

Since the 1900s, the link between soil biotic activity, soil organic matter (SOM) decomposition and stabilization, and soil aggregate dynamics has been recognized and intensively been studied. By 1950, many studies had, mostly qualitatively, investigated the influence of the five major factors (i.e. soil fauna, microorganisms, roots, inorganics and physical processes) on this link. After 1950, four theoretical mile-stones related to this subject were realized. The first one was when Emerson [Nature 183 (1959) 538] proposed a model of a soil crumb consisting of domains of oriented clay and quartz particles. Next, Edwards and Bremner [J. Soil Sci. 18 (1967) 64] formulated a theory in which the solid-phase reaction between clay minerals, polyvalent cations and SOM is the main process leading to microaggregate formation. Based on this concept, Tisdall and Oades [J. Soil Sci. 62 (1982) 141] coined the aggregate hierarchy concept describing a spatial scale dependence of mechanisms involved in micro- and macroaggregate formation. Oades [Plant Soil 76 (1984) 319] suggested a small, but very important, modification to the aggregate hierarchy concept by theorizing the formation of microaggregates within macroaggregates. Recent research on aggregate formation and SOM stabilization extensively corroborate this modification and use it as the base for furthering the understanding of SOM dynamics. The major outcomes of adopting this modification are: (1) microaggregates, rather than macroaggregates protect SOM in the long term; and (2) macroaggregate turnover is a crucial process influencing the stabilization of SOM. Reviewing the progress made over the last 50 years in this area of research reveals that still very few studies are quantitative and/or consider interactive effects between the five factors. The quantification of these relationships is clearly needed to improve our ability to predict changes in soil ecosystems due to management and global change. This quantification can greatly benefit from viewing aggregates as dynamic rather than static entities and relating aggregate measurements with 2D and 3D quantitative spatial information.

A history of research on the link between (micro)aggregates, soil biota, and soil organic matter dynamics

Biofilms are complex communities of microorganisms attached to surfaces or associated with interfaces. Despite the focus of modern microbiology research on pure culture, planktonic (free-swimming) bacteria, it is now widely recognized that most bacteria found in natural, clinical, and industrial settings persist in association with surfaces. Furthermore, these microbial communities are often composed of multiple species that interact with each other and their environment. The determination of biofilm architecture, particularly the spatial arrangement of microcolonies (clusters of cells) relative to one another, has profound implications for the function of these complex communities. Numerous new experimental approaches and methodologies have been developed in order to explore metabolic interactions, phylogenetic groupings, and competition among members of the biofilm. To complement this broad view of biofilm ecology, individual organisms have been studied using molecular genetics in order to identify the genes required for biofilm development and to dissect the regulatory pathways that control the plankton-to-biofilm transition. These molecular genetic studies have led to the emergence of the concept of biofilm formation as a novel system for the study of bacterial development. The recent explosion in the field of biofilm research has led to exciting progress in the development of new technologies for studying these communities, advanced our understanding of the ecological significance of surface-attached bacteria, and provided new insights into the molecular genetic basis of biofilm development.

Microbial Biofilms: from Ecology to Molecular Genetics

A global perspective on the use, sales, exposure pathways, occurrence, fate and effects of veterinary antibiotics (VAs) in the environment

A specific eucaryotic DNA marker from Solanum tuberosum cv Bintje (688 bp patatin cDNA fragment) was cloned into the unique HindIII-site of plasmid RP4. RP4:: pat was transferred from Escherichia coli to Pseudomonas fluorescens R2f by filter mating.

A specific marker, pat, for studying the fate of introduced bacteria and their DNA in soil using a combination of detection techniques

Survival of Pseudomonas fluorescens R2f containing either the conjugative plasmid RP4 or the nonconjugative plasmid pRK2501, and stability of the plasmids were studied in two soils, Ede loamy sand and Guelph loam, and in extracts prepared from these soils. In sterile soils, the introduced bacterial populations initially increased and then remained stable over a 47-day period. The presence of wheat roots did not significantly influence bacterial numbers in Guelph loam, whereas a slight increase occurred in Ede loamy sand. In Guelph loam, both plasmids were stably maintained in the introduced populations, but in Ede loamy sand substantial plasmid loss was observed. The presence of added phosphate in Ede loamy sand enhanced plasmid maintenance in the introduced R2f population. In nonsterile Guelph loam, a slow decline in the introduced populations was noted, regardless of plasmid type, whereas in Ede loamy sand the decline was more rapid. There was no detectable effect of plasmid type on host survival. Both ...

Survival of Pseudomonas fluorescens containing plasmids RP4 or pRK2501 and plasmid stability after introduction into two soils of different texture

Microbial community analysis by PCR-single-strand conformation polymorphism (PCR-SSCP).

Compte rendu et discussion (72 references) des differentes methodes permettant d'introduire et de decouvrir des microorganismes genetiquement recombines dans l'environnement. Les interets des nouvelles techniques genetiques et moleculaires pour la recherche sur l'environnement ont ete discutes

A review of selected methods in environmental microbial genetics

Influence of soil type on the transfer of RP4p from Pseudomonas fluorescens to indigenous bacteria.

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