constructing multispecies biofilms from rhizosphere samples for developing advanced biofertilizers

从根际土壤中开发协同多物种生物膜

Biofilms represent intricate microbial communities either affixed to surfaces or linked with interfaces. There is broad acknowledgment that the majority of bacteria encountered in various environments, such as natural habitats, clinical settings, and industrial contexts, endure within biofilm communities1. In soil, the rhizosphere&#8212;encompassing the root surface and its adjacent 2 mm thick region&#8212;forms an ecological niche with heightened nutrient availability. Specific bacteria have evolved strategies to exploit this niche, fostering microbial proliferation and fostering the formation of biofilm communities in the rhizosphere2.
Rhizosphere microbes are considered the &#39;second genome&#39; of plants3. Plant growth-promoting microorganisms (PGPM) engage in mutualistic symbiosis with plants, providing significant growth promotion and biocontrol functions4. On the one hand, PGPM can supply nutrients to plant roots, enhance nutrient uptake, defend against pathogens, and degrade pollutants in the rhizosphere soil5. On the other hand, PGPM utilizes plant root exudates as a nutrient source for growth and evolution, thereby influencing the rhizosphere soil and root system through their own metabolism. It is worth noting that the prerequisite for all these functions is the effective colonization of PGPM in the plant rhizosphere, forming stable biofilms6.
Rhizosphere biofilm affects the assembly process of rhizosphere microbes, and the temporal and spatial characteristics of rhizosphere microbe assembly are closely related to the interaction of multispecies biofilm7. It serves as the hub of plant-microbiome interactions, providing a stable and controllable niche for them&#160;to interact effectively. Therefore, studying rhizosphere biofilm is also an important direction to gain insight into the interaction between plants and microbes.
However, there is currently little research on rhizosphere multispecies biofilms. The high complexity of microbiome composition makes it challenging to answer fundamental ecological questions surrounding natural microbial communities8. In the process of experiments, many conditions, such as soil type, plant type, and the large number of microbial communities, need to be considered. Various factors limit the research of rhizosphere multispecies biofilms.
To further investigate multispecies biofilm communities from rhizosphere soil, such as interspecies synergistic interactions or metabolite cross-feeding8, it is desirable to artificially construct a simplified, representative, stable, and tractable synthetic microbial community under laboratory conditions9,10. Here, a standardized protocol combines several of the latest microbiological and bioinformatic techniques.

作者聚焦：开发合成微生物群落以生产第二代生物肥料

一种从根际土壤构建多物种生物膜群落的方法

从根际样品构建多物种生物膜以开发高级生物肥料

constructing-multispecies-biofilms-from-rhizosphere-samples-for

Research

JoVE Journal

Biology

Constructing Multispecies Biofilms from Rhizosphere Samples for Developing Advanced Biofertilizers

53 次观看. 首先，获取黄瓜种子并用 75% 乙醇消毒，然后用 2% 次氯酸钠溶液消毒。用无菌水冲洗种子后，在无菌条件下发芽。将持续发芽的两叶期幼苗移植到装有两公斤黑土的花盆中。当幼苗达到四叶阶段时，倒置花盆，使用消毒手套将根与两毫米厚的根际土壤隔离。将根部放入 50 毫升离心管中的 30 毫升 PBSS 缓冲液中。摇动试管 20 分钟，然后通过 100 微?...

视频: 从根际样品构建多物种生物膜以开发高级生物肥料

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.

A rapid and standardized procedure for establishing synergistic multispecies biofilm communities from various rhizosphere soils is presented here. It is a unique protocol designed to probe and simulate the complex rhizosphere soil microbiota.

The multispecies biofilm is a naturally occurring and dominant lifestyle of bacteria in nature, including in rhizosphere soil, although the current ...