Figure - uploaded by Lily Pereg
Content may be subject to copyright.
The recovery rate of added plant-growth-promoting rhizobacteria (PGPR) inoculants on rice field soil using total plate count and colony immunoblotting (CIB).
Source publication
Inoculant plant-growth-promoting bacteria are emerging as an important component of sustainable agriculture. There is a need to develop inexpensive methods for enumerating these organisms after their application in the field, to better understand their survival and impacts on yields. Immunoblotting is one potential method to measure viable cells, b...
Context in source publication
Similar publications
Salt stress is one of the major constraints hampering agricultural production owing to its impact on ethylene production and nutritional imbalance. A check on the accelerated ethylene production in plants could be helpful in minimizing the negative effect of salt stress on plant growth and development. Four Pseudomonas, 1 Flavobacterium, and 1 Ente...
Abstract The recently developed transparent soil consists of particles of Nafion, a polymer with a low refractive index (RI), which is prepared by milling and chemical treatment for use as a soil analogue. After the addition of a RI-matched solution, confocal imaging can be carried out in vivo and without destructive sampling. In a previous study,...
Citations
... Their presence was assessed by the observation of color change in Petri dishes using serial dilutions, which allowed the determination of populations of up to 1 Â 10 3 CFU per gram of soil at various phenological stages of the crop. Krishnen et al. (2011) employed the immunoblot method to monitor the presence of Pseudomonas fluorescens, Azospirillum brasilense, and Rhizobium leguminosarum in the rice crop. This methodology allowed the quantification of the inoculum up to 147 days of cultivation, enabling the establishment of a correlation between changes in irrigation events and the population dynamics of the inoculum. ...
The microbial inoculant industry, encompassing formulation, commercialization, and use, has witnessed remarkable global growth in recent years. Legal regulations are necessary to ensure the quality of microbial inoculants, protect farmers' and consumers' rights, and guide the commercial promotion of microbial products. However, regulating microbial inoculants remains a worldwide challenge, as they are often classified under chemical fertilizer laws due to the lack of experience with microorganism-based products. The lack of a consensus on the definition of these products and insufficient communication between the scientific community and farmers have hindered their legal regularization. To establish a framework for the development of microbial inoculants, these issues must be addressed. This includes reporting the microbial quality of the products, such as identification, abundance, plant growth-promoting capabilities, and contamination levels. In addition, physical and chemical factors, mode of application, dosage, and other aspects impacting product quality should be considered. Numerous organizations in different countries are currently working towards regulating the production, commercialization, and use of organic inputs in agriculture. Therefore, efforts are needed to define microbial inoculants and standardize regulations across countries, emphasizing cooperation among scientists, producers, sellers, and farmers.
... Their presence was assessed by the observation of color change in Petri dishes using serial dilutions, which allowed the determination of populations of up to 1 Â 10 3 CFU per gram of soil at various phenological stages of the crop. Krishnen et al. (2011) employed the immunoblot method to monitor the presence of Pseudomonas fluorescens, Azospirillum brasilense, and Rhizobium leguminosarum in the rice crop. This methodology allowed the quantification of the inoculum up to 147 days of cultivation, enabling the establishment of a correlation between changes in irrigation events and the population dynamics of the inoculum. ...
In agriculture, the use of plant-associated microorganisms has emerged as an alternative to reduce the excessive use of chemicals and toxic fertilizers. The plant microbiome, composed of a diversity of microorganisms such as bacteria, fungi, algae, and nematodes, plays a fundamental role in plant growth and health, and the use of these microorganisms requires bioprospecting studies to isolate them from agroecosystems similar to those in which they are intended to be applied. These microorganisms can directly promote plant growth through various mechanisms, including the production of phytohormones, nitrogen fixation, phosphate solubilization, and the production of ACC deaminase enzymes. They can also exert indirect effects by combating plant pathogens through the production of antibiotics, lipopeptides, siderophores, and lytic enzymes. Advances in omics technologies have revolutionized the analysis of microbial inoculants' mechanisms of action from different perspectives. Genomics enables the determination of the genetic potential of microorganisms as plant growth promoters. The transcriptomic analysis provides insights into the physiological and metabolic status of these microorganisms. Proteomics is used to identify and quantify proteins related to plant growth-promoting traits. The metabolomic analysis examines the metabolites resulting from the interaction between the genome, transcriptome, and proteome of microorganisms. This comprehensive set of approaches allows for a deeper understanding of the metabolic mechanisms in plant-microorganism interactions which is of vital importance for the development of sustainable agricultural practices.
... Not applicable [156] Functional gene β-galactosidase (lacZ) -Plate and indigenous bacteria with β-D-galactosidase activity -Culture dependent quantification -In situ detection 10 3 -10 5 CFU/g [161,162] β-glucoranidase (gusA) -Culture dependent quantification -Substrate dependent -In situ detection -Semi-quantitative -Endophytic detection 10 3 -10 5 CFU/g [163] Bacterial luciferase (lux) -Culture dependent quantification -Substrate and metabolism dependent -In situ detection -Semi-quantitative -No endogenous and indigenous activity -Inexpensive substrate 10 3 -10 4 CFU/cm [164,165] DGGE, Denaturant gradient gel electrophoresis; T-RFLP, terminal restriction fragment length polymorphism; ARDRA, amplified ribosomal DNA restriction analys. ...
... Immunoblot (Western blot) 방법을 이용하여 벼농사 토양에 서 3종의 PGPB (P. fluorescens, A. brasilense 및 R.Leguminosarum)을 검출할 수 있었다[156]. 이와 같이 면역 법 기반 PGPB 검출방법은 in situ 적용이 가능하고 특정 세 ...
Remediating soils contaminated with heavy metals due to urbanization and industrialization is very important not only for human health but also for ecosystem sustainability. Of the available remediation technologies for heavy metal-contaminated soils, phytoremediation is a relatively low-cost environmentfriendly technology which preserves biodiversity and soil fertility. The application of plant growth-promoting bacteria (PGPB) during the phytoremediation of heavy metal-contaminated soils can enhance plant growth against heavy metal toxicity and increase heavy metal removal efficiency. In this study, the sources of heavy metals that have adverse effects on microorganisms, plants, and humans, and the plant growthpromoting traits of PGPB are addressed and the research trends of PGPB-assisted phytoremediation over the last 10 years are summarized. In addition, the effects of environmental factors and PGPB inoculation methods on the performance of PGPB-assisted phytoremediation are discussed. For the innovation of PGPB-assisted phytoremediation, it is necessary to understand the behavior of PGPB and the interactions among plant, PGPB, and indigenous microorganisms in the field.
... The use of DAS-ELISA determined both the colonization of the root tip (3 × 10 7 CFU g −1 rhizosphere soils) and basal root (6 × 10 7 CFU g −1 rhizosphere soils) sections and resulted in numbers of inoculated strains 80% higher than those detected by plate counts. In contrast, plate counts yielded higher numbers when Krishnen et al. (2011) tracked B. subtilis B9, Bacillus amyloliquefaciens E19, and P. fluorescens 1N in a peat carrier biofertilizer. However, DAS-ELISA does not allow the inoculated PGPB to be detected in situ because of soil particle interference (Quadt-Hallmann and Kloepper, 1996). ...
... Therefore, IGS must be combined with tracking methods (Gamalero et al., 2003). Krishnen et al. (2011) used immunoblot (also known as western blot) to track three PGPB (P. fluorescens 1N, A. brasilense Sp245 and R. Leguminosarum bv. ...
Since the 1980s, plant growth‒promoting bacteria (PGPB) have been studied as a sustainable alternative to the use of chemical fertilizers to increase crop yields, and effective PGPB have been isolated from diverse plant (e.g., endosphere and phyllosphere) and soil (e.g., rhizosphere) compartments. Despite the promising plant growth promotion results commonly observed under laboratory and greenhouse conditions, the successful application of PGPB under field conditions has been limited, partly by the lack of knowledge of the ecological/environmental factors affecting the colonization, prevalence and activity of beneficial bacteria on crops. It is generally accepted that the effectiveness of PGPB depends on their ability to colonize a niche and compete with the indigenous plant microbiome under agronomic conditions. However, most studies do not include tracking or monitoring of PGPB in the environment after their application, and the beneficial effects on plants are measured by determining biomass‒ and physiology‒related parameters without confirming bacterial colonization. To date, methods based on reporter genes, immunological reactions and nucleic acids have been applied to track or monitor PGPB in seeds, soils or in planta after inoculation. In this review, we describe, compare and discuss the methods used for tracking and monitoring PGPB, including challenges and perspectives on some novel methods.
... Detailed methods for ensuring quality control of the inoculant strains have been published in a widely distributed laboratory monograph Krishnen et al. 2011). This manual includes a range of molecular and physiological methods applicable to many other PGPR microbes ...
Since their original isolation from rice paddies near Hanoi, the set of microbial strains comprising the biofertilizer BioGro have been subjected to extensive and intensive experimentation in both laboratory and the field. Based on a hypothesis that such strains inoculated onto rice and other plants could significantly reduce the need for chemical fertilizers, this has been successfully tested using numerous procedures, documented in a series of peer-reviewed papers. The BioGro strains have been examined by a range of molecular and biochemical techniques, also providing means of quality control of inoculants. A positive response by rice plants to BioGro strains has been confirmed by proteomics. More than 20 randomized block design field experiments conducted in Vietnam or Australia have confirmed their effectiveness under a range of field conditions, reviewed here. Interactions with different rice cultivars have also been examined. While the response to inoculation is complex, the hypothesis of increased nutrient efficiency has been amply confirmed as consistent with observations. Finally, an extensive participatory research project over 3 years in the Mekong Delta showed reductions in fertilizer needs as high as 52 % as rice farmers learned to apply the technology. This result shows the importance of such adaptive practices for successful application of this biofertilizer technology in field condition.
It is known that members of the bacterial genus Azospirillum can promote the growth of a great variety of plants, an ability harnessed by the industry to create bioproducts aimed to enhance the yield of economically relevant crops. Its versatile metabolism allows this bacterium to adapt to numerous environments, from optimal to extreme or highly polluted. The fact of having been isolated from soil and rhizosphere samples collected worldwide and many other habitats proves its remarkable ubiquity. Azospirillum rhizospheric and endophytic lifestyles are governed by several mechanisms, leading to efficient niche colonization. These mechanisms include cell aggregation and biofilm formation, motility, chemotaxis, phytohormone and other signaling molecules production, and cell-to-cell communication, in turn, involved in regulating Azospirillum interactions with the surrounding microbial community. Despite being infrequently mentioned in metagenomics studies after its introduction as an inoculant, an increasing number of studies detected Azospirillum through molecular tools (mostly 16S rRNA sequencing) as part of diverse, even unexpected, microbiomes. This review focuses on Azospirillum traceability and the performance of the available methods, both classical and molecular. An overview of Azospirillum occurrence in diverse microbiomes and the less-known features explaining its notorious ability to colonize niches and prevail in multiple environments is provided.
It is critical to monitor plant growth-promoting rhizobacteria (PGPR) in soils after inoculation. Among common PGPR, the Azospirillum sp. strains TSA2s and TSH100 have the ability to mitigate nitrous oxide (N2O) emissions from agricultural soils; however, their mechanism by which they successfully colonize and achieve beneficial effects in plants remains poorly understood. Here, a simple and robust procedure was developed to design strain-specific primers based on the whole-genome sequences of these two strains. After evaluating their specificity and amplification efficiency, three primer pairs were screened for each Azospirillum sp. strain. Quantification of these two strains in the rhizosphere soils of red clover revealed distinct inoculant dynamics for each strain under greenhouse conditions. Specifically, 22, 34, and 41 days after inoculation with TSA2s, the population size of the inoculant was notably greater than that in the non-inoculated control, and reached a maximum at day 34. In contrast, the population size of the TSH100 inoculant was largest at day 22, then decreased dramatically from 34 days after inoculation. These results suggest that different PGPR may have different strategies for colonization and prevalence after inoculation. Of the 31 rhizosphere carbon sources added to the Biolog Eco Micro plate to simulate plant root exudates, TSA2s utilized 14 whereas TSH100 utilized only one. TSA2s utilized more diverse root exudates of red clover, indicating better colonization and prevalence than TSH100 after inoculation. Therefore, this study presents a method of monitoring PGPR after inoculation. Furthermore, this method can screen PGPR with a better ability to survive and colonize, enabling the development of efficient, stable, and standardized biofertilizers.
