Plant-Growth-Promoting Rhizobacteria (PGPR) and its Effects
Author: Dharmendra Kumar
Microorganisms are both foes and friends of man. Though a layman understands microbes as evils due to their devastating effects on his crop plants, animals and as well as on himself, he had been taking advantage of their enormous beneficial contribution in this world. Some of the microbes that inhabit soils, rivers, lakes, oceans and air are beneficial to man.
From the microbial point of view soil is a nutritional desert. In contrast, soil adjacent to roots is relatively nutrient rich because as much as 40% of the photosynthates are lost to the soil through root exudation. Thus the area surrounding the roots is rich in nutrients and harbours great microbial activity and is called as “rhizosphere”. Population of bacteria in the rhizosphere is high ranging from 1010 to 10 12 cells per gram of soil. These bacteria in the rhizosphere are in continuous interaction with each other as well as with plants. Some of these interactions are beneficial, some are deleterious and some are neutral. The term plant growth promoting rhizobacteria (PGPR) has been introduced (Kloepper et al., 1980) for the bacteria that colonize the plant roots and stimulate plant growth and crop yield. Kloepper et al. (1989) observed increase in crop yield as high as 160% using PGPR. Many bacteria belonging to genera Pseudomonas.Bacillus, Azospirillum, Enterobacter, Serratia and Arthrobacter have a vast promise as PGPR and are now used in agriculture as bioinocuants. A subset of PGPR strains which enhance the nodulation of legumes by rhizobia have been designated as nodulation promoting rhizobacteria (NPR).
Successful crop production is highly dependent on the availability of various nutrients. Among these, nitrogen (N) is the most needed one, which is limiting in Indian soils. Nitrogenous fertilizers constitute the major components among the chemical fertilizers used in agriculture. Hellriegel and Wilforth (1888) showed that legumes could utilize atmospheric nitrogen and that their utilization was dependent upon bacteria present in the nodules. Subsequent studies led to the discoveries of Rhizobium, Clostridium, Azotobacter, Azospirillum, and blue green algae as very important N2 - fixing organisms.
Phosphorus (P) is one of the important and in some cases may be the first limiting nutrient for plant growth. Phosphorous nutrition benefits the plant by producing deeper and abundant roots. Hence, the supply of this element to plant is essential for achieving optimum crop yield. It is supplied through phosphatic fertilizers, animal manure, plant residues, domestic organic wastes and rock phosphate. The introduction of the most efficient phosphate solubilizing microorganisms (PSM) in the rhizosphere of crop increases the availability of phosphorus from insoluble sources of phosphate and also increases the efficiency of phosphate fertilizers such as superphosphate and rockphosphate. The phosphate uptake was enhanced and the yields were increased by 10 to 50% with different crops. A combination of rock phosphate and PSM was found to be more effective than using rock phosphate alone particularly in soils having neutral to alkaline pH. In general, 40% of the superphosphate could be saved by combined use of rock phosphate and PSM.
It is very clear that NPK are not only quite important but also limits crop production and consequently affect our livelihood. It is also obvious, that the modern agriculture is heavily dependent on chemical fertilizers to meet the food demands of ever increasing population. The production of chemical fertilizer is based on non-renewable and constantly depleting petroleum based feed stocks. It is estimated that the crops remove about 9 -10 million tonnes more N + P2O5 + K2O every year than the total additions through fertilizers in India. Progressive depletion of major plant nutrients in soil due to intensive cultivation has necessitated the use of higher dose of chemical fertilizers particularly in tropical soils where the organic matter content is very low. This huge drain on nutrients will continue to impoverish the soils unless these are replenished by natural or artificial means.
Application of high doses of chemical fertilizers may temporarily help to increase crop production and may relieve the problem of demand and supply. However, this may turn bitter and with highly regrettable consequences where soil fertility will be depleted or become acidic and devoid of macro and micro-nutrients for crops to grow and microorganisms to proliferate. Thus it is absolutely necessary to awake timely and be able to use ecofriendly inputs such as beneficial PGPR and save our ‘currency’, the soil, and its constituents.
Rhizosphere and Soil microorganisms
Soil is the natural growth media for living plants and microorganisms. The physical and chemical properties of soil directly affect the soil microorganisms. Bacteria are the most dominant group of microorganisms in the soil and probably equal to one half of the microbial biomass present there in. The most abundant bacterial forms or genera present in the soil are Pseudomonas, Arthrobacter, Clostridium, Bacillus, Micrococcus, Flavobacterium, Chromobacter, Sarcina, Enterobacter, Corynebacterium, Mycobacter etc. (Mishra 1996; Rangaswami and Bagyaraj, 1998; Subba Rao, 2000). Soil is also an environment in which the underground parts of the plant have their abode. As they grow through the soil, the condition of soil in the immediate vicinity of the root witness a drastic change in various ways. The microhabitat of the soil in close proximity of root changes and this has an impact on the microorganisms residing in the region. This area of enhanced activity surrounding living roots that is composed of soil particles and active communities of soil microorganisms is called the ‘rhizosphere’ (Hiltner, 1904). This environment is richer in nutrients, and its microbial communities differ from those present in area not influenced by the roots (Alexander, 1977). It is now recognized that the ‘rhizosphere effect’ is due to the root exudates which attract some soil microorganisms. The width of the zone of soil thus influenced by the root varies with the plant species its age and cultural conditions, soil conditions, environmental conditions etc. (Rangaswami and Bagyaraj, 1998).
Some bacterial species living in the rhizosphere can affect plant growth in either a positive or in a negative way. Rhizosphere bacteria having favourable effects on plant growth and yield of commercially important crops are denominated as Plant Growth Promoting Rhizobacteria (PGPR) (Kloepper and Schroth, 1978). Studies on beneficial effects of rhizospheric bacteria have most often been based on increased plant growth, faster seed germination, better seedling emergence, enhanced nodulation and nitrogen fixation in leguminous crops and suppression of diseases. As a consequence, PGPR have been further divided into subsets like emergence promoting rhizobacteria (EPR); nodulation promoting rhizobacteria (NPR) and disease suppressing rhizobacteria (DSR) (Kloepper et al., 1986b). However, there can be considerable overlap among these subsets. For instance, PGPR that act through biological control can also enhance germination or growth. In recent years, many reviews have appeared dealing with PGPR isolation, screening, ecology, physiology and their use as agrobiotechnological inoculants (Glick 1995; Lazarovits and Nowak, 1997; Saxena et al., 2000).
Mechanisms of plant growth promotion
The mechanisms by which PGPR promote plant growth are not fully understood. However, studies carried out by different workers suggest some of these as follows (i) the ability to produce or change the concentration of the plant hormones-indole acetic acid (IAA), gibberellic acid cytokinins and ethylene (ii) asymbiotic N2 fixation (iii) antagonism against phytopathogenic microorganisms by production of siderophores, b - 1,3 - Glucanases, chitinases, antibiotics and cyanide and (iv) solubilization of mineral phosphates and other nutrients
Effect of PGPR on growth and yield of legume plants
Rhizobium, a symbiotic nitrogen fixer, has been shown to act as a plant growth promoting rhizobacteria for non-leguminous crops. Reports indicate that non-legumes react to the presence of bradyrhizobia and rhizobia in the rhizosphere. Root hair curling induced by these symbiotic bacteria was observed on maize, rice and oat plants (Plazinsky et al., 1985; Terouchi and Syono 1990). Studies also show that Nod-factors produced by Bradyrhizobium and Rhizobium, can be perceived by tomato, as indicated by the induction of alkalization in tomato cell cultures. Nodulating and non-nodulating strains of Rhizobium leguminosarum produce IAA (Wang et al., 1982). Noel et al., (1996) observed under gnotobiotic conditions, a direct growth promotion of the early seedling root of canola and lettuce by R. leguminosorum. Chabot et al. (1996a) observed that phosphate solubilizing strains of R. leguminosarum bv phaseoli improved the growth of maize and lettuce. Hotlich et al. (1994) obtained significant shoot dry matter yield increases (7-8%) by inoculating maize, spring wheat and spring barley (Hordeum vulgare L.) with strain R 39 of R. leguminosarum bv. trifolii, in field experiments. Yanni et al. (1995) also observed that certain effective wild type strains of R. leguminosarum bv. trifolii establish natural plant-bacterial associations that have the potential to promote rice growth under both field and laboratory conditions. Antoun et al. (1998) also observed increase in the dry matter yield of radish due to inoculation of strain Tal 629 of B. japonicum.
Coinoculation of PGPR strains with the root nodule bacteria ( Rhizobium and Bradyrhizobium) has been shown to influence the nodulation, nitrogen fixation and growth of leguminous plants. Grimes and Mount (1987) found that Pseudomonas putida strain M17 increased Rhizobium nodulation of bean in field soils. In Soybean, inoculation of a strain of P. fluorescens can enhance the nodulation ability of B. japonicum. Preincubation of B. japonicum with P. fluorescens before inoculation further increased the level of nodulation (Nishijima et al., 1988). Chanway et al. (1989) tested nine PGPR strains on a single cultivar of lentil and pea in the field. None of the strains stimulated the growth of pea, but in plots inoculated with one or more rhizobacterial strains, there were significant increase in emergence, vigour, nodulation; C2H2 reduction activity and root weight. Bolton et al. (1990) found that preinoculation of Bradyrhizobium japonicum and Pseudomonas fluorescens increased the level of nodulation in pea plant indicating a bacterial-bacterial interaction. Turner and Backman (1991) observed that treatment of peanut seeds with Bacillus subtilis was associated with improved germination and emergence, enhanced nodulation by Rhizobium spp., enhanced plant nutrition, reduced levels of root cankers caused by Rhizoctonia solani AG-4 and increased root growth. Singh (1994) suggested the role of phosphate solubilising microorganisms in stimulating the activity of native rhizobial strains resulting in significant increase in nodulation, plant growth and grain yield (0-41.38%) in soybean plants. Podile (1995) found that seed bacterization of pigeonpea with Bacillus subtilis AF1 enhanced the percentage emergence, growth and nodulation.
Singh and Gaur (1992) reported two strains of rhizospheric bacteria that further improved the nodulation as well as competitiveness of an effective strain of chickpea Rhizobium. They further suggested the role of flavonoid like substances produced by rhizobacterial strains in nod gene expression (Singh and Gaur, 1995). Similar results were reported for chickpea (Parmar and Dadarwal, 1999). Coinoculation of rhizobacteria with effective Rhizobium strains of chickpea resulted in a significant increase in nodule weight, root and shoot biomass and total plant nitrogen in chickpea. The nodule stimulating rhizobacteria enhanced levels of flavonoid-like compounds in roots on seed bacterization. Also, ethyl acetate extracts of culture supernatant fluids when applied to seeds resulted in enhancement of flavonoids in roots, suggesting that the rhizobaceria have a direct influence on root flavonoids which might be an additional factor in root promotion by these bacteria (Parmar and Dadarwal, 1999).
IAA producing Bacillus isolates promoted root growth and (or) nodulation when coinoculated with Rhizobium etli on Phaseolus vulgaris ‘contender’ in growth chambers (Srinivasan et al., 1996). Similarly coinoculation of soybean withB. japonicum and Serratia liquefaciens 2-68 or S. proteamaculans 1-102 increased soybean grain and protein yield compared to the non treated controls (Dashti et al., 1997, 1998). Gupta et al. (1998) isolated 16 strains of PGPR from rhizotic zone of green gram which were identified as Enterobacter, Pseudomonas and Bacillus spp. These PGPR were found to increase shoot and root length and fresh and dry biomass of green gram plants.Enterobacter isolate EG-ER-2 increased the nodule occupancy of Bradryhizobium spp. strain S24 on green gram and another isolate KG-ER-1 did the same for rhizobial strain COG 15 (Gupta et al., 1998).
Gupta et al. (1998) observed 60-80% increase in nodule occupancy and increased shoot biomass, N-contents and grain yield of green gram plant as a result of coinoculation of PGPR and Bradyrhizobium.
Beneficial effects of coinoculation of Azospirillum withRhizobium have been reported by different workers (Tilaket al., 1981; Sarig et al., 1986; Yahalom et al., 1987; Saxena and Tilak, 1994; Galal, 1997; Tchebotaret al., 1998). Coinoculation of Rhizobium spp. and Azospirillum spp. enhances the nodule number and grain yield of various leguminous crops (Sarig et al., 1986; Yahalom et al., 1987). Coinoculation also increases the shoot length, dry weight, number of root hairs and root diameter of alfalfa (Itzigsohnet al., 1983). Dual inoculation with a mixture of Bradyrhizobium japonicum and A. brasilense strains improved growth and nitrogen fixation of soybean (Galal, 1997). The combined inoculation of white clover with Rhizobium leguminosarum bv. trifolii and A. lipoferum enhanced the number of nodules by 2-3 times and acetylene reduction activity by 2.3-2.7 times (Tchebotar et al., 1998).
Some bacterial species living in the rhizosphere can affect plant growth in either a positive or in a negative way. Rhizosphere bacteria having favourable effects on plant growth and yield of commercially important crops are denominated as Plant Growth Promoting Rhizobacteria (PGPR)
Anith, K.N. (1997). Molecular basis of antifungal activity of a fluorescent pseudomonad, Ph.D. Thesis, Indian Agricultural Research Institute, New Delhi.
Antoun, H., Besuchamp, C.J., Goussard, N., Chabot, R. and Lalande, R. (1998). Potential of Rhizobium and Bradyrhizobium species as plant growth promoting rhizobacteria on non-legumes : Effect on radishes (Raphanus sativus L.). Plant and Soil, 204 : 57-67.
Arshad, M. and Frankenberger Jr., W.T. (1984). Influence of ethylene produced by soil microorganisms on etiolated pea seedlings. Appl. Environ. Microbiol., 54 : 2728-2732.
Bakker, A.W. and Schippers, B. (1987). Microbial cyanide production in the rhizosphere in relation to potato yield reduction and Pseudomonas spp. mediated plant growth stimulation. Soil Biol. Biochem., 19(4) : 451-457.
Bansal, S. (1999). Studies on major storage proteins in developing seeds of Pisum sativum L. Ph. D. Thesis, Kurukshetra University, Kurukshetra.
Belimov, A.A. (2001). Characterization of plant growth promoting rhizobacteria isolated from polluted soil and containing 1 - aminocyclopropane - 1- carboxylate deaminase. Can. J. Microbiol., 47 : 642-652.
Beringer, J.E. (1974). R-factor transfer inRhizobium leguminosarum. J. Gen. Microbiol., 84 : 188-198.
*Berg, G. and Bahl, H. (1997). Characterization of beneficial rhizobacteria of oil seed rape for biological control of Verticillium wilt. Gesunde Pflanzen, 49: 76-82.
Bergey’s mannul of Determinative Bacteriology. (1994). Ed. J. G. Holte. Baltimore, Williams and Wilkins.
Bochow, H. and Fritzsche, S. (1990). Induction of phytoalexins biosynthesis by culture filtrate of bacterial antagonists. In Proceedings of International Workshop on PGPR in Switzerland. pp. 158-160.
Boddey, R.M. and Dobereiner, J. (1995). Nitrogen fixation associated with grasses and cereals : Recent progress and perspectives for the future. Fert. Res., 42 : 241-250.
Chandna, M. (1992). Structural and genetical studies on seed proteins of Lathyrus sativus L. Ph. D. Thesis, Kurukshetra University, Kurukshetra.
Chanway, C.P., Hynes, R.K. and Nelson, L.M. (1989). Plant growth promoting rhizobacteria : effect on the growth and nitrogen fixation of lentils (Lens esculenta Moench.) and pea (Pisum sativum L.). Soil Biol. Biochem., 21 : 511-512.
Chen, C., Bauske, E.M., Musson, G., Rodriquez-Kabana, R. and Kloepper, J.W. (1995). Biological control of Fusarium wilt on cotton by use of endophytic bacteria. Biol. Cont., 5 : 83-91.
Christensen,. (1946). Urea decomposition as a means of differentiating Proteus and Paravolon cultures from each other and from Salmonella ans Shigella types. J. Bacteriol., 25 : 461.
Cook, R.J. (1993). Making greater use of introduced microorganisms for biological control of plant pathogens. Annu. Rev. Phytopath., 31 : 53-80.
Costacurta, A. and Venderleyden, J. (1995). Synthesis of phytohormones by plant-associated bacteria. Crit. Rev. Microbiol., 21 : 1-18.
Davis, B.J. (1964). Electrophoresis III. Methods and Application to Human Serum Protein. Ann. New York Acad. Sci., 121 : 404-408.
Day, J.M., Harris, D., Dart, P.J. and Van Berkum, P. (1975). The broadbalk experiment : An investigation of nitrogen gains from non-symbiotic fixation. In : Nitrogen fixation by free-living microorganisms (ed. W.D.P. Stewart) vol. 6, pp. 72-84. International Biological Programme Series, Cambridge Univ. Press, Cambridge.
Dubeikovsky, A.N., Mordukhova, E.A., Kochetkov, V.V., Polikurpova, F.V. and Boranin, M.M. (1993). Growth promotion of black current soft wood cutting by recombinant strain Pseudomonas fluorescens. BSP 53a Synthesizing and increased amount of indole-3-acetic acid. Soil Biol. Biochem., 25 : 1277-1281.
Duffy, B.K., Simon, A. and Weller, D.M. (1996). Combination of Trichoderma koningii with fluorescent Pseudomonas for control of take-all on wheat. Phytopath., 86 : 188-194.
Dworkin, M. and Froster, J. (1958). Experiments with some microorganisms which utilize ethane and hydrogen. J. Bacteriol., 75 : 592-601.
Gupta, A., Saxena, A.K., Gopal, M. and Tilak, K.V.B.R. (1998a). Bacterization of green gram with rhizosphere bacteria for enhanced plant growth. J. Sci. Indust. Res., 57 : 726-736.
Gupta, A., Saxena, A.K., Gopal, M. and Tilak, K.V.B.R. (1998b). Enhanced nodulation of green gram by introduced Bradyrhizobium when co-inoculated with plant growth promoting rhizobacteria. J. Sci. Indust. Res., 57 : 720-725.
Handelsman, J., Raffel, S., Mester, E.H., Wunderlich, L. and Grau, C.R. (1990). Biological control of damping-off of alfalfa seedling byBacillus cereus UV 185. Appl. Environ. Microbiol., 56 : 713-718.
Hardy, R.W.F., Holsten, R.D., Jackson, E.K. and Burns, R.C. (1968). The acetylene - ethylene assay for N2 fixation : Laboratory and Field evaluation. Plant Physiol., 43 : 1185-1207.
Harrigan, W.F., McCance, M.E. (1961). Laboratory Methods in Microbiology. Academic Press, New York.
*Hiltrner, L. (1904). Liber neuere Erfahrungen und problems aufdem Gebiet der Bodenbackteriologie und unter besonderer Berucksichtingung der Grundringing und Brache. Arb. Dtsch. London Ges, 98 : 59-78.
Hirari, M.Y., Fujiwara, T., Komeda, Y., Chino, M., Naito, S. and Barrow, N.J. (1993). Temporal and nutritional regulation of soybean 7S seed stroage protein gene promoters in transgenic Petunia. Proceeding of Plant nutrition from genetic engineering to field practice: International Plant Nutrition Colloguium.pp, 143-146.
Holbrook, A.A., Edge, W.J.W. and Bailey, F. (1961). Spectrophotometric method for determination of gebberellic acid. In Gibberellin. A.C.S. Washington, D.C. pp. 159-167.
Hollwach, L.P., Thompson, J.F. and Medison, J.T. (1984). Effects of exogenous methionine on storage protein composition of soybean cotyledon cultured in vitro. Plant Physiol., 74 : 576-583.
Paulitz, T.C., Anas, O. and Frenando, D.G. (1992). Biocontrol ofPythium damping-off of seed treatment with Pseudomonas putida relationship with ethanol production by pea and soybean seeds. Bio. Sci. Tech., 2 : 193-201.
Pelczar, M.J. (1957). In : Manual of Microbiological Methods. McGraw Hill Book Company Inc., New York.
Saxena, A.K. and Tilak, K.V.B.R. (1998). Free living nitrogen fixers : their role in crop productivity. In : Microbes for health, wealth and Sustainable Environment (ed Ajit Varma) Malhotra Publishing House, New Delhi.
Saxena, A.K., Pal, K.K. and Tilak, K.V.B.R. (2000). Bacterial biocontrol agents and their role in plant disease management. In : Biocontrol Potential and its Exploitation in Sustainable Agriculture (Eds. R.K. Upadhyay, K.G. Mukerji and B.P. Chamola), pp. 25-37. Kluwer Academic/Plenum Publisher, New York.
Schroeder, H. E. (1984). Effects of applied growth regulators on pod growth and seed protein composition in Pisum sativum L. J. Exp. Bot. 35 : 813-821.
Subba Rao, N.S. (2000). Soil Microbiology. 4th ed. Oxford & IBH Publishing Co. Pvt. Ltd., New Delhi, Calcultta. p. 407.
Sundara Rao, W.V.B. and Sinha, M.K. (1963). Phosphate dissolving organisms in the soil and rhizosphere. Ind. J. Agric. Sci., 53 : 272-278.
Tilak, K.V.B.R., Singh, C.S. and Rana, J.P.S. (1981). Effect of combined inoculation of Azospirillum brasilense with Rhizobium trifolii, Rhizobium meliloti and Rhizobium sp. (cowpea miscellany) on nodulation and yield of clover ( Trifolium repens), lucerne (Medicago sativa) and chickpea (Cicer arietinum). Zbl. Bakt. II. Abt., 136 : 117-120.
Walker, R., Emslie, K.A. and Allan, E.J. (1996). Bioassay methods for the detection of antifungal activity by Pseudomonas antimicrobien against the grey mould pathogenBotrytis cinesea. J. Appl. Biotechnol., 81 : 531-537.
Wang, C., Knill, E., Glick, B. R. and Defago, G. (2000). Effect of transferring 1- aminocyclopropanr-1-carboxylic acid (ACC) deaminase genes into Pseudomonas fluorescens strain CHAO and its gac A derivative CHA96 on their growth promoting and disease suppressive capacities. Can. J. Microbiol., 46 : 898-907..
Yahalom, E., Okon, Y. and Devart, A. (1987). Azospirillum effects on susceptibility to Rhizobium nodulation and on nitrogen fixation of several forage legumes. Can. J. Microbiol., 33 : 51-514.
Yamagata, H., Sugimoto, T., Tanaka, K. and Kasai, Z. (1982). Biosynthesis of storage proteins in developing rice seeds. Plant Physiol. 70 : 1094-1100.
Yanni, Y.C., Rizk, R.Y., Corich, V., Squartini, A. and Dazzo, E.G. (1995). Endorhizosphere colonization and growth promotion of indica and japonica rice varieties by Rhizobium leguminosarum bv. trifolii. Abstr. 017. In : Proceedings of the 15th symbiotic nitrogen fixation conference. North Carrolina State University, Raleigh, N.C.
Zafar, Y., Malik, K.A. and niemann, E.G. (1987). Studies on N2-fixing bacteria associated with salt tolerant grass Leptochloa fusca (KL.) Kunth. Mircen. J. Appl. Microbiol., 3 : 45-56.
Zhang, Jin Xu, Howell, C.R. and Starr, J.L. (1996). Suppression of Fusarium colonization of cotton roots and Fusarium wilt by seed treatments with Gliocladium virens andBacillus subtilis. Biocont. Sci. Technol., (UK) 6 : 175-187.
About Author / Additional Info:
Currently doing Ph.D from IARI, New Delhi