Defining the wheat microbiome: towards microbiome-facilitated crop production

Wheat is one of the world’s most important crops, but its production relies heavily on agrochemical inputs which are notoriously harmful to the environment. It is well known that a multitude of microbes interact with eukaryotic organisms, including plants, and the sum of microbes and their functions associated with a given host is termed the microbiome. Plant-microbe interactions can be beneficial, neutral or harmful to the host plant. Over the last decade, with the development of next generation DNA sequencing technology, our understanding of the plant microbiome structure has dramatically increased. Considering that defining the wheat microbiome is key to leverage crop production in a sustainable way, here we describe how different factors drive microbiome assembly in wheat, including crop management, edaphic-environmental conditions and host selection. In addition, we highlight the benefits to take a multidisciplinary approach to define and explore the wheat core microbiome to generate solutions based on microbial (synthetic) communities or single inoculants. Advances in plant microbiome research will facilitate the development of microbial strategies to guarantee a sustainable intensification of crop production

Development of a defined compost system for the study of plant-microbe interactions

Plant growth promoting rhizobacteria can improve plant health by providing enhanced nutrition, disease suppression and abiotic stress resistance, and have potential to contribute to sustainable agriculture. We have developed a sphagnum peat-based compost platform for investigating plant-microbe interactions. The chemical, physical and biological status of the system can be manipulated to understand the relative importance of these factors for plant health, demonstrated using three case studies: 1. Nutrient depleted compost retained its structure, but plants grown in this medium were severely stunted in growth due to removal of essential soluble nutrients - particularly, nitrogen, phosphorus and potassium. compost nutrient status was replenished with the addition of selected soluble nutrients, validated by plant biomass; 2. When comparing milled and unmilled compost, we found nutrient status to be more important than matrix structure for plant growth; 3. In compost deficient in soluble P, supplemented with an insoluble inorganic form of P (Ca3(PO4)2), application of a phosphate solubilising Pseudomonas strain to plant roots provides a significant growth boost when compared with a Pseudomonas strain incapable of solubilising ca3(PO4)2. Our findings show that the compost system can be manipulated to impose biotic and abiotic stresses for testing how microbial inoculants influence plant growth

Inorganic Chemical Fertilizer Application to Wheat Reduces the Abundance of Putative Plant Growth-Promoting Rhizobacteria

The profound negative effect of inorganic chemical fertilizer application on rhizobacterial diversity has been well documented using 16S rRNA gene amplicon sequencing and predictive metagenomics. We aimed to measure the function and relative abundance of readily culturable putative plant growth-promoting rhizobacterial (PGPR) isolates from wheat root soil samples under contrasting inorganic fertilization regimes. We hypothesized that putative PGPR abundance will be reduced in fertilized relative to unfertilized samples. Triticum aestivum cv. Cadenza seeds were sown in a nutrient depleted agricultural soil in pots treated with and without Osmocote⃝R fertilizer containing nitrogen-phosphorous-potassium (NPK). Rhizosphere and rhizoplane samples were collected at flowering stage (10 weeks) and analyzed by culture-independent (CI) amplicon sequence variant (ASV) analysis of rhizobacterial DNA as well as culture- dependent (CD) techniques. Rhizosphere and rhizoplane derived microbiota culture collections were tested for plant growth-promoting traits using functional bioassays. In general, fertilizer addition decreased the proportion of nutrient-solubilizing bacteria (nitrate, phosphate, potassium, iron, and zinc) isolated from rhizocompartments in wheat whereas salt tolerant bacteria were not affected. A “PGPR” database was created from isolate 16S rRNA gene sequences against which total amplified 16S rRNA soil DNA was searched, identifying 1.52% of total community ASVs as culturable PGPR isolates. Bioassays identified a higher proportion of PGPR in non-fertilized samples [rhizosphere (49%) and rhizoplane (91%)] compared to fertilized samples [rhizosphere (21%) and rhizoplane (19%)] which constituted approximately 1.95 and 1.25% in non-fertilized and fertilized total community DNA, respectively. The analyses of 16S rRNA genes and deduced functional profiles provide an in-depth understanding of the responses of bacterial communities to fertilizer; our study suggests that rhizobacteria that potentially benefit plants by mobilizing insoluble nutrients in soil are reduced by chemical fertilizer addition. This knowledge will benefit the development of more targeted biofertilization strategies

Old meets new: most probable number validation of metagenomic and metatranscriptomic datasets in soil

Metagenomics and metatranscriptomics provide insights into biological processes in complex substrates such as soil, but linking the presence and expression of genes with functions can be difficult. Here, we obtain traditional most probable number estimates (MPN) of Rhizobium abundance in soil as a form of sample validation. Our work shows that in the Highfield experiment at Rothamsted, which has three contrasting conditions (>50 years continual bare fallow, wheat and grassland), MPN based on host plant nodulation assays corroborate metagenomic and metatranscriptomic estimates for Rhizobium leguminosarum sv. trifolii abundance. This validation is important to legitimize soil metagenomics and metatranscriptomics for the study of complex relationships between gene function and phylogen

The potato rhizosphere microbiota correlated to the yield of three different regions in Korea

We examined potato rhizosphere bacterial and fungal communities across three regions: Cheongju, Pyeongchang, and Gangneung. These regions have varying soil and climate conditions, resulting in different yields. We found that precipitation was the main limiting factor in our study while soil physiochemical factors affect bacterial and fungal microbiota in correlation with yield. Both bacterial and fungal microbiota showed distinct patterns according to the regions. ASVs positively correlated with yield were predominantly found in the Pyeongchang region which also produced the highest yields, while ASVs negatively correlated with yield were associated with Gangneung where the lowest yields were observed. The greatest bacterial and fungal diversity was detected in Pyeongchang consisting of Propionibacteriales, Burkholderiales, and Vicinamibacteriales. Gangneung, on the other hand primarily belong to Sordariales, Mortierellales, Cystofilobasidiales, and Tremellales. The putative yield-negative ASVs detected in Gangneung may have been influenced by drought stress. This work has highlighted key bacterial and fungal taxa as well as core taxa that may potentially be associated with high and low yields of potato in relation to metadata which includes soil chemical and physical parameters as well as weather data. Taken together we suggest that this information can be used to assess site suitability for potato production

Development of a defined compost system for the study of plant-microbe interactions

Plant growth promoting rhizobacteria can improve plant health by providing enhanced nutrition, disease suppression and abiotic stress resistance, and have potential to contribute to sustainable agriculture. We have developed a sphagnum peat-based compost platform for investigating plant-microbe interactions. The chemical, physical and biological status of the system can be manipulated to understand the relative importance of these factors for plant health, demonstrated using three case studies: 1. Nutrient depleted compost retained its structure, but plants grown in this medium were severely stunted in growth due to removal of essential soluble nutrients - particularly, nitrogen, phosphorus and potassium. Compost nutrient status was replenished with the addition of selected soluble nutrients, validated by plant biomass; 2. When comparing milled and unmilled compost, we found nutrient status to be more important than matrix structure for plant growth; 3. In compost deficient in soluble P, supplemented with an insoluble inorganic form of P (Ca3(PO4)2), application of a phosphate solubilising Pseudomonas strain to plant roots provides a significant growth boost when compared with a Pseudomonas strain incapable of solubilising Ca3(PO4)2. Our findings show that the compost system can be manipulated to impose biotic and abiotic stresses for testing how microbial inoculants influence plant growth

Assessment of core and accessory genetic variation in Rhizobium leguminosarum symbiovar trifolii strains from diverse locations and host plants using PCR-based methods

The nitrogen-fixing symbiosis between Rhizobium leguminosarum and host legumes is recognized as a key part of sustainable agriculture. A culture collection containing rhizobia isolated from legumes of economic importance in the UK and worldwide, maintained at Rothamsted Research for many years, provided material for this study. We aimed to develop and validate efficient molecular diagnostics to investigate whether the host plant or geographical location had a greater influence on the genetic diversity of rhizobial isolates, and the extent to which the core bacterial genome and the accessory symbiosis genes located on plasmids were affected. To achieve this, core housekeeping genes and those involved in symbiosis interactions were sequenced and compared with genome-sequenced strains in the public domain. Results showed that some Rh. leguminosarum symbiovar trifolii strains nodulating clovers and Rh. leguminosarum sv. viciae strains nodulating peas and vicias shared identical housekeeping genes, clover nodule isolates from the same location could have divergent symbiosis genes, and others isolated on different continents could be very similar. This illustrates the likely co-migration of rhizobia and their legume hosts when crops are planted in new areas and indicates that selective pressure may arise from both local conditions and crop host genotypes

Insights into the rhizosphere microbiome: past and future perspectives

Microorganisms associated to plants can promote plant growth and development through direct and/or indirect mechanisms in addition to enabling them to tolerate biotic and abiotic stresses. Several factors are responsible for shaping the microbiome associated with plants and understanding how they influence the structure and functions of microbial communities is crucial for the development of sustainable agriculture. Using culture-independent methods and soils from Rothamsted Research, I will describe the importance of different factors in determining the root microbiome structure and diversity in wheat. In addition, I will describe a culturedependent approach that led to the development of a microbial product for maize. I also discuss gaps and benefits of taking a multidisciplinary approach to explore the plant microbiome to generate microorganism-based solutions for sustainable intensification of crop production

Effect of Plant Growth Promoting Bacteria on the Growth of Wheat Seedlings Subjected to Phosphate Starvation

Certain phosphorous solubilizing (PSB) and phosphorous mineralizing (PMB) bacteria may improve plant growth by improving nutrient availability. The aim of this work was to evaluate the effect of inoculation with two Bacillus spp. strains, 12A and 25A, on wheat seedlings growth. To this aim, a durum and a bread wheat genotype were grown under controlled conditions in a low P compost medium to evaluate: (i) the effect of the bacterial isolates on plant growth and root system architecture; (ii) the expression of two key genes indicative of the P-starvation response and phosphate (Pi) uptake, TaIPS1 and TaPHT1.6-B1. The results showed that 12A Bacillus sp. significantly increased root length, surface area and biomass. Furthermore, an enhanced shoot dry weight and P content were observed. This might be explained by the capacity of strain 12A to produce indole-3-acetic acid (IAA) in addition to P mineralizing and P solubilizing capability. No effect on plant growth was observed for 25A strain. The semi-quantitative gene expression analysis showed an overall lower expression of TaIPS1 in the inoculated plants and highest expression of TaPHT1.6-B1 in 12A inoculated plants. This suggests that Pi-responsive genes might be useful molecular indicators for the effectiveness of PSB and PM

Identification of novel aphid-killing bacteria to protect plants.

Aphids, including the peach-potato aphid, Myzus persicae, are major insect pests of agriculture and horticulture, and aphid control measures are limited. There is therefore an urgent need to develop alternative and more sustainable means of control. Recent studies have shown that environmental microbes have varying abilities to kill insects. We screened a range of environmental bacteria isolates for their abilities to kill target aphid species. Tests demonstrated the killing aptitude of these bacteria against six aphid genera (including Myzus persicae). No single bacterial strain was identified that was consistently toxic to insecticide-resistant aphid clones than susceptible clones, suggesting resistance to chemicals is not strongly correlated with bacterial challenge. Pseudomonas fluorescens PpR24 proved the most toxic to almost all aphid clones whilst exhibiting the ability to survive for over three weeks on three plant species at populations of 5–6 log CFU cm−2 leaf. Application of PpR24 to plants immediately prior to introducing aphids onto the plants led to a 68%, 57% and 69% reduction in aphid populations, after 21 days, on Capsicum annuum, Arabidopsis thaliana and Beta vulgaris respectively. Together, these findings provide new insights into aphid susceptibility to bacterial infection with the aim of utilizing bacteria as effective biocontrol agents