Establishment of a system for bacterial colonization experiments in wheat

Eukaryotes, including plants, are hosts to microbes. These microbes can be recruited from the environment as well as transferred between the generations horizontally or vertically. We hypothesize that in wheat, due to the priority effects, vertically transferred microbiota has priority advantage over bacteria recruited from the environment. Isolation efforts on Triticum aestivum seeds and seedlings have yielded few cultivable bacterial strains present both on and in the seed and seedling. Pantoea agglomerans was the species found in most of the samples, independent of the geographical origin (Germany, Turkey). Here we show that the GFP-tagged strain of Pantoea agglomerans, originally isolated from wheat, maintains its ability to colonize germ-free wheat roots and to proliferate within the root. Independently of the inoculum size used, log-phase-like population growth within the root was observed. Afterward, while the population growth was still ongoing, any further increase in the population size was tied to the growth of the root habitat. Once the population count was standardized based on root fresh weight, the plateau was observed within 8 days post-inoculation, showing that the carrying capacity of the host is limited. In planta competitions between GFP and mCherry tagged Pantoea strains were performed to check for numerical priority effects and adaptive priority effects. Competing strains were introduced with a time lag between them, or one of the strains was pre-cultivated in a plant, while the second was cultivated in liquid media. Our results show a strong impact of the priority effect. The numerical advantage of primary colonizer led to its high relative abundance. No adaptive advantage was observed. Compared to our single colonizer experiments, total carrying capacity of the host was unchanged. Persistence of Pantoea in the plant host was tested up to 46 days post-inoculation. Density of the population varies between the plant compartment, reaching highest CFU per gram of tissue in the roots, lowest in the leaves. My observations support the hypothesis of vertical transfer of Pantoea agglomerans and its association with host lifecycle (ability to colonize the seedling from the seed while outcompeting soil-borne Pantoea). This data provides the fundament for future experiments on mechanistic aspects of wheat colonization by Pantoea agglomerans, which can be performed by analysis of the bacterial genome and preparation of knock-outs. The established gnotobiotic system enables further competition and exclusion experiments

Advancing Our Functional Understanding of Host–Microbiota Interactions: A Need for New Types of Studies

Multicellular life evolved in the presence of microorganisms and formed complex associations with their microbiota, the sum of all associated archaea, bacteria, fungi, and viruses. These associations greatly affect the health and life history of the host, which led to a new understanding of “self” and establishment of the “metaorganism” concept.1 The Collaborative Research Centre (CRC) 1182 aims at elucidating the evolution and function of metaorganisms. Its annual conference, the Young Investigator Research Day (YIRD), serves as a platform for scientists of various disciplines to share novel findings on host–microbiota interactions, thereby providing a comprehensive overview of recent developments and new directions in metaorganism research. Even though we have gained tremendous insights into the composition and dynamics of host‐associated microbial communities and their correlations with host health and disease, it also became evident that moving from correlative toward functional studies is needed to examine the underlying mechanisms of interactions within the metaorganism. Non‐classical model organisms in particular possess significant potential to functionally address many open questions in metaorganism research. Here, we suggest and introduce a roadmap moving from correlation toward a functional understanding of host–microbiota interactions and highlight its potential in emerging ecological, agricultural, and translational medical applications

Comparative analysis of amplicon and metagenomic sequencing methods reveals key features in the evolution of animal metaorganisms

Background The interplay between hosts and their associated microbiome is now recognized as a fundamental basis of the ecology, evolution, and development of both players. These interdependencies inspired a new view of multicellular organisms as “metaorganisms.” The goal of the Collaborative Research Center “Origin and Function of Metaorganisms” is to understand why and how microbial communities form long-term associations with hosts from diverse taxonomic groups, ranging from sponges to humans in addition to plants. Methods In order to optimize the choice of analysis procedures, which may differ according to the host organism and question at hand, we systematically compared the two main technical approaches for profiling microbial communities, 16S rRNA gene amplicon and metagenomic shotgun sequencing across our panel of ten host taxa. This includes two commonly used 16S rRNA gene regions and two amplification procedures, thus totaling five different microbial profiles per host sample. Conclusion While 16S rRNA gene-based analyses are subject to much skepticism, we demonstrate that many aspects of bacterial community characterization are consistent across methods. The resulting insight facilitates the selection of appropriate methods across a wide range of host taxa. Overall, we recommend single- over multi-step amplification procedures, and although exceptions and trade-offs exist, the V3 V4 over the V1 V2 region of the 16S rRNA gene. Finally, by contrasting taxonomic and functional profiles and performing phylogenetic analysis, we provide important and novel insight into broad evolutionary patterns among metaorganisms, whereby the transition of animals from an aquatic to a terrestrial habitat marks a major event in the evolution of host-associated microbial composition

T4 Phage Tail Adhesin Gp12 Counteracts LPS-induced Inflammation In Vivo

Bacteriophages that infect Gram-negative bacteria often bind to the bacterial surface by interaction of specific proteins with lipopolysaccharide (LPS). Short tail fiber proteins (tail adhesin, gp12) mediate adsorption of T4-like bacteriophages to Escherichia coli, binding surface proteins or LPS. Produced as a recombined protein, gp12 retains its ability to bind LPS. Since LPS is able to exert a major impact on the immune response in animals and in humans, we have tested LPS-binding phage protein gp12 as a potential modulator of the LPS-induced immune response. We have produced tail adhesin gp12 in a bacterial expression system and confirmed its ability to form trimers and to bind lipopolysaccharide in vitro by dynamic light scattering. This product had no negative effect on mammalian cell proliferation in vitro. Further, no harmful effects of this protein were observed in mice. Thus, gp12 was used in combination with LPS in a murine model, and it decreased the inflammatory response to LPS in vivo, as assessed by serum levels of cytokines IL-1 alpha and IL-6 and by histopathological analysis of spleen, liver, kidney and lungs. Thus, in future studies gp12 may be considered as a potential tool for modulation and specifically for counteracting LPS-related physiological effects in vivo