An Approach to Constructing Multispecies Biofilm Communities from Rhizosphere Soil.

The multispecies biofilm is a naturally occurring and dominant lifestyle of bacteria in nature, including in rhizosphere soil, although the current understanding of it is limited. Here, we provide an approach to rapidly establish synergistic multispecies biofilm communities. The first step is to extract cells from rhizosphere soil using the differential centrifugation method. Afterward, these soil cells are inoculated into the culture medium to form pellicle biofilm. After 36 h of incubation, the bacterial composition of the biofilm and the solution underneath are determined using the 16S rRNA gene amplicon sequencing method. Meanwhile, high-throughput bacterial isolation from pellicle biofilm is conducted using the limiting dilution method. Then, the top 5 bacterial taxa are selected with the highest abundance in the 16S rRNA gene amplicon sequencing data (pellicle biofilm samples) for further use in constructing multispecies biofilm communities. All combinations of the 5 bacterial taxa were quickly established using a 24-well plate, selected for the strongest biofilm formation ability by the crystal violet staining assay, and quantified by qPCR. Finally, the most robust synthetic bacterial multispecies biofilm communities were obtained through the methods above. This methodology provides informative guidance for conducting research on rhizosphere multispecies biofilm and identifying representative communities for studying the principles governing interactions among these species.

Enhanced surface colonisation and competition during bacterial adaptation to a fungus.

Bacterial-fungal interactions influence microbial community performance of most ecosystems and elicit specific microbial behaviours, including stimulating specialised metabolite production. Here, we use a co-culture experimental evolution approach to investigate bacterial adaptation to the presence of a fungus, using a simple model of bacterial-fungal interactions encompassing the bacterium Bacillus subtilis and the fungus Aspergillus niger. We find in one evolving population that B. subtilis was selected for enhanced production of the lipopeptide surfactin and accelerated surface spreading ability, leading to inhibition of fungal expansion and acidification of the environment. These phenotypes were explained by specific mutations in the DegS-DegU two-component system. In the presence of surfactin, fungal hyphae exhibited bulging cells with delocalised secretory vesicles possibly provoking an RlmA-dependent cell wall stress. Thus, our results indicate that the presence of the fungus selects for increased surfactin production, which inhibits fungal growth and facilitates the competitive success of the bacterium.

Metabolic interactions affect the biomass of synthetic bacterial biofilm communities.

IMPORTANCE: Co-occurrence network analysis is an effective tool for predicting complex networks of microbial interactions in the natural environment. Using isolates from a rhizosphere, we constructed multi-species biofilm communities and investigated co-occurrence patterns between microbial species in genome-scale metabolic models and in vitro experiments. According to our results, metabolic exchanges and resource competition may partially explain the co-occurrence network analysis results found in synthetic bacterial biofilm communities.

Annulment of Bacterial Antagonism Improves Plant Beneficial Activity of a Bacillus velezensis Consortium.

Bacillus sp. strains that are beneficial to plants are widely used in commercial biofertilizers and biocontrol agents for sustainable agriculture. Generally, functional Bacillus strains are applied as single-strain communities since the principles of synthetic microbial consortia constructed with Bacillus strains remain largely unclear. Here, we demonstrated that the mutual compatibility directly affects the survival and function of two-member consortia composed of Bacillus velezensis SQR9 and FZB42 in the rhizosphere. A mutation in the global regulator Spo0A of SQR9 markedly reduced the boundary phenotype (appearance of a visible boundary line at the meeting point of two swarms) with wild-type FZB42, and the combined use of the SQR9(△spo0A) mutant and FZB42 improved biofilm formation, root colonization, and the production of secondary metabolites that are beneficial to plants. Furthermore, alleviation of antagonistic interactions of two-member Bacillus consortia improved its beneficial effects to cucumber in a greenhouse experiment. Our results provide evidence that social interactions among bacteria could be an influencing factor for achieving a desired community-level function. IMPORTANCE Bacillus velezensis is one of the most widely applied bacteria in biofertilizers in China and Europe. Additionally, the molecular mechanisms of plant growth promotion and disease suppression by representative model strains are well established, such as B. velezensis SQR9 and FZB42. However, it remains extremely challenging to design efficient consortia based on these model strains. Here, we showed that swarm encounter phenotype is one of the major determinants that affects the performance of two-member Bacillus consortia in vitro and in the rhizosphere. Deletion in global regulatory gene spo0A of SQR9 reduced the strength of boundary formation with FZB42 and resulted in the improved plant growth promotion performance of the dual consortium. This knowledge provides new insights into efficient probiotics consortia design in Bacillus spp.

Bacillus velezensis stimulates resident rhizosphere Pseudomonas stutzeri for plant health through metabolic interactions.

Trophic interactions play a central role in driving microbial community assembly and function. In gut or soil ecosystems, successful inoculants are always facilitated by efficient colonization; however, the metabolite exchanges between inoculants and resident bacteria are rarely studied, particularly in the rhizosphere. Here, we used bioinformatic, genetic, transcriptomic, and metabonomic analyses to uncover syntrophic cooperation between inoculant (Bacillus velezensis SQR9) and plant-beneficial indigenous Pseudomonas stutzeri in the cucumber rhizosphere. We found that the synergistic interaction of these two species is highly environmental dependent, the emergence of syntrophic cooperation was only evident in a static nutrient-rich niche, such as pellicle biofilm in addition to the rhizosphere. Our results identified branched-chain amino acids (BCAAs) biosynthesis pathways are involved in syntrophic cooperation. Genome-scale metabolic modeling and metabolic profiling also demonstrated metabolic facilitation among the bacterial strains. In addition, biofilm matrix components from Bacillus were essential for the interaction. Importantly, the two-species consortium promoted plant growth and helped plants alleviate salt stress. In summary, we propose a mechanism in which synergic interactions between a biocontrol bacterium and a partner species promote plant health.

Advances in neuroimaging studies of alcohol use disorder (AUD).

Alcohol use disorder (AUD) is a worldwide problem and the most common substance use disorder. Chronic alcohol consumption may have negative effects on the body, the mind, the family, and even society. With the progress of current neuroimaging methods, an increasing number of imaging techniques are being used to objectively detect brain impairment induced by alcoholism and serve a vital role in the diagnosis, prognosis, and treatment assessment of AUD. This article organizes and analyzes the research on alcohol dependence concerning the main noninvasive neuroimaging methods, structural magnetic resonance imaging, functional magnetic resonance imaging, and electroencephalography, as well as the most common noninvasive brain stimulation - transcranial magnetic stimulation, and intersperses the article with joint intra- and intergroup studies, providing an outlook on future research directions.

Enhanced Control of Plant Wilt Disease by a Xylose-Inducible degQ Gene Engineered into Bacillus velezensis Strain SQR9XYQ.

Bacillus velezensis SQR9 (former B. amyloliquefaciens SQR9) is a plant-growth-promoting rhizobacterium (PGPR) that promotes plant growth and health. The colonization of PGPR strains along plant roots is a prerequisite for them to execute their specific functions. However, one problem of microbial introduction in practice is that the applied PGPR strains do not always successfully colonize the rhizosphere. In Bacillus spp., two-component signal transduction system (TCS) DegS/U regulates flagellar motility, biofilm formation and antibiotic production. Phosphorylation of DegU by DegS is positively affected by DegQ protein. In this study, we constructed a xylose-inducible degQ genetically engineered strain SQR9XYQ to improve the biocontrol activity. The results from in vitro, root in situ, greenhouse experiments and RT-qPCR studies demonstrate that (i) the phosphorylation of DegU in SQR9XYQ can be gradually activated by xylose, which is a component of both cucumber and tomato root exudates, and (ii) biofilm formation, antibiotic expression, colonization activity, and biocontrol efficiency were improved in SQR9XYQ compared with the wild-type strain SQR9. These results suggest that colonization trait is important to biocontrol strains for maintenance of plant health.