Research Article |
Corresponding author: Gloria Georgieva ( gloriq_georgieva1@abv.bg ) Academic editor: Ventsislava Petrova
© 2023 Gloria Georgieva, Trayana Nedeva, Marina Badalova, Veronika Deleva, Valentin Savov.
This is an open access article distributed under the terms of the Creative Commons Attribution License (CC BY 4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Citation:
Georgieva G, Nedeva T, Badalova M, Deleva V, Savov V (2023) Study of the plant growth-promoting capacity of Pseudomonas putida 1046 in a model plant system. In: Chankova S, Danova K, Beltcheva M, Radeva G, Petrova V, Vassilev K (Eds) Actual problems of Ecology. BioRisk 20: 115-128. https://doi.org/10.3897/biorisk.20.97581
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Plant Growth Promoting Rhizobacteria (PGPR) represent a microbial community that exerts growth-promoting capabilities in plants by various mechanisms. Among the PGPR genera, Pseudomonas spp. deserves special attention. It is due to its characteristic traits, like the production of phytohormones and siderophores, solubilization of minerals and phosphates, and plant protection from biotic and abiotic stress. These PGPR properties depend on the microorganism and its plant counterpart. The use of microbial strains as bioinoculants must consider the physiological and economic aspects of the process, and the plant growth stimulating effect has to be checked and proved. This study aimed to explore the PGP capacity of Pseudomonas putida 1046 strain in a model plant system of the economically important corn culture (Zea mays). The effect of the strain’s metabolic status on the plant germination capacity was evaluated. Bacterial cultures, grown 16 h and 48 h, were explored for the treatment of the corn seeds at three experimental concentrations: 0.1, 0.2, and 0.4%, and monitoring of their germination capacity through the growth indicators length of the radicle, length of the coleoptile, and the number of lateral roots. The data obtained outline the positive effect of Pseudomonas putida 1046 on the germination capacity of corn when applied at 0.2% concentration. The in vitro treatment of the model plants with 0.2% suspension resulted in a 22.87%–28.33% increase in the length of the radicle, a 35.96%–49.56% increase in the length of the coleoptile, and a 5.41–16.67% increase in the number of the lateral roots. High values of the vigour index (2125 for 16 h and 2721 for 48 h culture) were also registered. The strain’s ability to produce siderophores of hydroximate type and exhibit phosphate solubilizing activity is proved. The optimal treatment parameters of the corn seeds comprise the application of 0.2% suspension of 16 h grown Pseudomonas putida 1046 strain for five days.
biofertilizer, PGPR, rhizosphere microorganisms, Zea mays
The soil microbial communities influence plant growth and development. The playground for this effect is the rhizosphere since the plant root system and the soil microorganisms interact there. This interaction is a complex process influenced by various environmental factors, such as temperature, humidity, pH, availability of nutrients, etc. (
The need for environmentally friendly fertilizers or biofertilizers is constantly increasing nowadays (
Many species belonging to g. Pseudomonas possess plant growth-promoting (PGP) characteristics that allow their application as bacterial fertilizers. They are abundantly present in the rhizosphere. Pseudomonas spp. are known for their well-established growth-promoting mechanisms. They encompass better root colonization, the production of enzymes, metabolites, phytohormones and siderophores, mineral and phosphate solubilization, and plant protection from biotic and abiotic stress (
The application of the microbial PGP properties depends on the microorganism and its plant counterpart. Corn (Zea mays) is one of the most important cereal crops in the world after wheat and rice. It is used as livestock feed, human food, and raw material for several industries (
This study aimed to explore the PGP capacity of bacteria belonging to Pseudomonas putida species in a model plant system of the economically important corn culture Zea mays.
Pseudomonas putida 1046 strain was used in this study. It possesses the biochemical capacity to catabolize aromatic hydrocarbons and their derivatives. Corn seeds (Dekalb – DKC 5830 HD (Hybrid 101)) were used as a model plant test system. They are characterized by a fast initial development, high quality of the grain, tolerance to high sowing norms, and a response to high fertilizers and sowing norms. In some of the experiments, soybean seeds (Glycine max L.) were used.
The experimental strain was maintained on Nutrient agar (BB-NCIPD Ltd., Bulgaria). Batch cultures in Nutrient broth were obtained after cultivation at 28 °C for 16 h or 48 h on a rotary shaker (220 rpm).
Pseudomonas putida 1046 biochemical profile was established applying ApiZYM rapid systems for the detection of bacterial enzymes and Api 20 NE standardized system for the identification of non-fastidious, non-enteric Gram-negative rods. The tests performance was according to the methods described by
The seed germination method was applied to study the PGP effect of Pseudomonas putida 1046 strain on corn (Zea mays) seeds. The corn seeds were subjected to surface sterilization before the germination test. Sodium hypochlorite (0.02%) solution was applied for 2 min., followed by intense rinsing with sterile distilled water. The seeds’ germination capacity was assessed through the growth parameters of the plant root system (
Correlation analysis was performed by calculating the correlation coefficients with MS Excel software of two variables data sets – Pseudomonas putida 1046 suspension concentration (0% (control), 0.1%, and 0.2%) and the growth parameters of the plant root system (the indices length of the radicle, length of the coleoptile, the number of the lateral roots, and the vigour index).
All presented data are mean values of at least 3 individual experiments. The data were analysed by MS EXCEL build-in function, and the results were presented as means with standard deviations (n=3).
Pseudomonas putida 1046 strain has been selected on the basis of preliminary biochemical analyses that indicate its potential plant growth-promoting capacity. The biochemical profile of the strain was established applying ApiZYM and Api 20 NE detection systems. The biochemical profile data presented in Table
ApiZYM | Api 20NE | ||
---|---|---|---|
Alkaline phosphatase | positive | NO3 Reduction of nitrates to nitrites | negative |
Esterase (C 4) | positive | Reduction of nitrates to nitrogen | positive |
Esterase Lipase (C 8) | positive | TRP Indole production (tryptophan) | negative |
Lipase (C 14) | negative | GLU Fermentation (glucose) | negative |
Leucine arylamidase | positive | ADH Arginine dihydrolase | positive |
Valine arylamidase | negative | URE Urease | negative |
Cystine arylamidase | negative | ESC Hydrolysis (βglucosidase) (esculin) | negative |
Trypsin | negative | GEL Hydrolysis (protease) (gelatin) | negative |
α-chimotrypsin | negative | PNGP β-Galactosidase (paranitropheny l-ßDgalactopyranosidase) | negative |
Acid phosphatase | positive | GLU Assimilation (glucose) | positive |
Naphthol-AS-BI-phosphohydrolase | positive | ARA Assimilation (arabinose) | positive |
α-galactosidase | negative | MNE Assimilation (mannose) | positive |
β- galactosidase | negative | MAN Assimilation (mannitol) | positive |
β-glucuronidase | negative | NAG Assimilation (N-acetylglucosamine) | positive |
α-glucuronidase | negative | MAL Assimilation (maltose) | negative |
β-glucosidase | negative | GNT Assimilation (potassium gluconate) | positive |
N-acetyl-β-glucosaminidase | negative | CAP Assimilation (capric acid) | positive |
α-mannosidase | negative | ADI Assimilation (adipic acid) | positive |
α-fucosidase | negative | MLT Assimilation (malate) | positive |
CIT Assimilation (trisodium citrate) | positive | ||
PAC Assimilation (phenylacetic acid) | positive | ||
OX Cytochrome oxidase | positive |
The effect of the strain metabolic status on the test plants’ germination was evaluated in order to check the PGP capacity of Pseudomonas putida 1046 strain. The PGP capacity of the bacterial strain was assessed in respect to its test plants. The seeds germination of the test plants was used as an assessment indicator through the quantitative measurement of three indices: length of the radicle, length of the coleoptile, and number of lateral roots. The corn seeds at three experimental concentrations (0.1, 0.2, and 0.4%) were treated with bacterial cultures, grown for 16 h and 48 h and monitored for their germination capacity. The length of the radicle, length of the coleoptile, and the number of lateral roots were used as growth indicators.
The values for the growth indicator length of the radicle are shown in Figs
PGP effect of Pseudomonas putida 1046 on the germination of corn seeds evaluated through length of the radicle (A), length of the coleoptile (B), and the number of lateral roots (C).
The data for the length of the coleoptile show that all three tested concentrations exhibit a positive effect on its growth (Figs
The third index (number of the lateral roots), used for assessment of the corn germination capacity, showed a relatively lower stimulatory effect (2.78%–18.60%) as compared to the previous two indicators. However, it is evident that the tendency for the highest positive results obtained by the 0.2% suspension of the bacterial strain is kept (Figs
Among the three tested bacterial concentrations, the 0.2% suspension was the right choice for the seeds’ treatment. The 0.1% expressed no or mild positive effect on the number of lateral roots and the length of radicles, as shown in Figs
The values for the vigour index are presented in Fig.
Based on the experimental data for the plant growth indices evaluated, correlation analysis for the bacterial and plant data sets was performed as described in the section Materials and Methods. The results are presented in Table
In order to confirm the PGP effect of Pseudomonas putida 1046 strain on the corn test plant, two different approaches were used: testing another plant model system and extended research on PGP characteristics of the bacterial strain.
Soybean (Glycine max L.) was exploited as a second model system. The crop has a global position as one of the most significant agri-cultures, a source of food, protein, and oil. The PGP effect of Pseudomonas putida 1046 was evaluated using two of the indices applied for the corn: length of the radicle and the epicotile (coleoptile). The tendency for enhanced germination due to the increased number of lateral roots and enlarged epicotile length, promoted by Pseudomonas putida 1046, is confirmed by the data depicted in Fig.
PGP effect of Pseudomonas putida 1046 on the germination of soybean seeds presented through the growth parameters length of radicle (A) and length of epicotile (B).
The ability of the model bacterial strain to produce siderophores and solubilize phosphates was examined by qualitative analyses. These activities were tested due to their promotional effect on plant growth: enhanced metal accumulation in plants and putative application for phytoremediation purposes of siderophores, and the growth-promoting ability attributed to phosphate solubilization. The data about these PGP characteristics are presented in Figs
Nowadays, extensive research is focused on PGPR as a versatile replacement of fertilizers, pesticides, and other agrochemicals for the promotion of plant growth. PGPR influence in a positive way (directly or indirectly) the soil structure and fertility, decomposition of organic matter and organic pollutants, solubilization of nutrients of mineral nature, production of plant growth regulators, stimulation of the root growth, and execution of biocontrol against soil and seed-borne pathogens. (
The data presented in the Results section outline the positive effect of Pseudomonas putida 1046 strain on the germination capacity of the technical crop Zea mays. Its biochemical profile speculated PGP potential, and the determined PGP traits confirmed it. The in vitro treatment of the model plants with bacterial strain improved their germination capacity.
The physiological status of the culture is a parameter correlating with its biosynthetic capacity. The metabolically active 16 h culture expressed high values for the three tested indices and strong correlation coefficients of the bacterial and model plant data sets resulting in a better germination capacity. Pseudomonas putida strains and plants are in commensal relationships. The root exudates of the plant feed the bacteria, and the bacteria stimulate plant growth through the production of hormone precursors and antibiotics that suppress the pathogens’ growth. It also supports nutrients immobilization (
The results demonstrate that the Pseudomonas putida 1046 strain possesses defined plant growth-promoting capacity. It enhances the process of corn germination due to the increased number of lateral roots and enlarged length of the coleoptile and the root. The optimal treatment parameters of the corn seeds, considering both the physiological and economic aspects of the process, comprise the application of 0.2% suspension of 16 h grown Pseudomonas putida 1046 strain for five days.
The positive germination capacity effect exerted by Pseudomonas putida 1046 strain contributes to improve our knowledge of the variety of approaches and mechanisms that are implicated in the plant growth promotion by these bacteria. However, greater understanding of the interaction between bacteria and host plants requires further study. From an economic point of view, in light of the possible application of Pseudomonas putida 1046 strain as a bioinoculant, the use of less concentrated suspension is more feasible and cost-effective.