Rhizobacterial bioformulation for the effective management of Macrophomina root rot in mungbean

Abstract

Fluorescent pseudomonads based bioformulation was evaluated for their ability to control Macrophomina root rot disease in mungbean (Vigna mungo). P. fluorescens isolate Pf1 showed the maximum inhibition in mycelial growth of Macrophomina phaseolina under in vitro conditions. Bioformulation of Pf1 with chitin was effective in reducing the root rot incidence in green gram both under glasshouse and field conditions. The rhizosphere colonization of P. fluorescens was observed appreciable with the green gram plants. However, Pf1 amended with chitin colonized effectively. Furthermore, the induction of defence-related enzymes and chemicals in plants by Pf1 amended with or without chitin and neem were tested. Increased accumulation of defence enzymes viz., phenylalanine ammonia lyase (PAL), peroxidase (PO), polyphenol oxidase (PPO), chitinase, β-1,3-glucanse and phenolics were observed in Pf1 bioformulation amended with chitin, pre-treated plants challenge inoculated with M. phaseolina under glasshouse conditions. The present study reveals that in addition to direct antagonism and plant-growth promotion, PGPR strains amended with chitin bioformulation induced defence-related enzymes and pathogenesis related (PR) proteins which collectively enhance the resistance in green gram against the infection of M. phaseolina.     

Induction of defense-related proteins in tomato roots treated with Pseudomonas fluorescens Pf1 and Fusarium oxysporum f. sp. lycopersici

Pseudomonas fluorescens isolate Pf1 was found to protect tomato plants from wilt disease caused by Fusarium oxysporum f. sp. lycopersici. Induction of defense proteins and chemicals by P. fluorescens isolate Pf1 against challenge inoculation with F. oxysporum f. sp. lycopersici in tomato was studied. Phenolics were found to accumulate in bacterized tomato root tissues challenged with F. oxysporum f. sp. lycopersici at one day after pathogen challenge. The accumulation of phenolics reached maximum at the 5^th day after pathogen challenge. In pathogen-inoculated plants, the accumulation started at the 2^nd day and drastically decreased 4 days after the pathogen inoculation. Activities of phenylalanine ammonia-lyase (PAL), peroxidase (PO) and polyphenol oxidase (PPO) increased in bacterized tomato root tissues challenged with the pathogen at one day after pathogen challenge and activities of PAL and PO reached maximum at the 4^th day while activity of PPO reached maximum at the 5^th day after challenge inoculation. Isoform analysis revealed that a unique PPO1 isoform was induced and PO1 and PPO2 isoforms were expressed at higher levels in bacterized tomato root tissues challenge inoculated with the pathogen. Similarly, -1,3 glucanase, chitinase and thaumatin-like proteins (TLP) were induced to accumulate at higher levels at 3-5 days of challenge inoculation in bacterized plants. Western blot analysis showed that chitinase isoform Chi2 with a molecular weight of 46 kDa was newly induced due to P. fluorescens isolate Pf1 treatment challenged with the pathogen. TLP isoform with molecular weight of 33 kDa was induced not only in P. fluorescens isolate Pf1-treated root tissues challenged with the pathogen but also in roots treated with P. fluorescens isolate Pf1 alone and roots inoculated with the pathogen. These results suggest that induction of defense enzymes involved in phenylpropanoid pathway and accumulation of phenolics and PR-proteins might have contributed to restriction of invasion of F. oxysporum f. sp. lycopersici in tomato roots.     

Plant growth promoting potentials of Pseudomonas spp. strain OG isolated from marine water

Abstract

Bacterium Pseudomonas spp. olive green (OG) was isolated from marine water, yet, it was characterized as plant growth promoting bacterium (PGPB). Multiple plant growth promoting traits of OG isolate were determined in vitro. It was tested positive for Indole-3-acetic acid (IAA) production with 29 µg ml^?1 of IAA yield, phosphate solubilization with 34 µg ml^?1 solubilization of Tri-calcium-phosphate and it showed maximum of 32 µg ml^?1 of ammonia production. OG isolate was affirming siderophore production, hydrocyanic acid (HCN) production and catalase production. 16S rRNA gene sequence comparison was used to identify the isolate which showed its closest neighbor to be Pseudomonas fluoroscens strain BCPBMS-1. Efficacy of this PGPB was tested on the seedling growth of two test plants chickpea and green gram. Both the test plants treated with OG-based talc bioformulation showed significant growth promotion. Chickpea showed enhanced overall fresh biomass by 24%, overall dry biomass by 27% was observed after 15 days of seeded in pots. Green gram showed enhanced overall dry biomass by 28% was observed after 10 days of seeded in pots.     

Integration of Pseudomonas fluorescens and salicylic acid improves citrus canker disease management caused by Xanthomonas citri subsp citri-A*

The individual and combined effects of Pseudomonas fluorescens (Pf) and salicylic acid (SA) were investigated for control of citrus bacterial canker (CBC). Both treated plants with copper hydroxide and untreated ones were used as controls. Mexican lime (Citrus aurantifolia) seedlings were treated with SA at 10 mM, Pf and distilled water. Plants were initially inoculated with Xanthomonas citri subsp citri 72 h post treatments. Results indicated that the Pf and SA treatment controlled CBC more effectively compared to separately applying Pf or SA. The application of Pf in combination with SA significantly reduced lesion number per leaf (72%) and disease severity (84%). Significant changes in the activities of peroxidase and catalase were found. In conclusion, the integration of Pf with SA complements each other and can be applied to manage citrus canker disease in conjunction with other control programmes.     

PGPR mediates induction of pathogenesis-related (PR) proteins against the infection of blast pathogen in resistant and susceptible ragi [Eleusine coracana (L.) Gaertner] cultivars

The Pseudomonas fluorescens isolate 1 (Pf1) was found to protect the ragi [Eleusine coracana (L.) Gaertner] blast fungus, Pyricularia grisea. Induction of defense proteins viz. chitinase, β-1,3 glucanase, peroxidase (PO) and polyphenol oxidase (PPO) by the Pf1 isolate was studied against P. grisea. Chitinase in a resistant, susceptible and commonly used cultivar with and without challenge inoculation of P. grisea, revealed changes in the isoform pattern by UV illumination after staining the gel with fluorescent brightner 28. Native PAGE (polyacrylamide gel electrophoresis) of PO showed the single isoform in all the treatments including the control and a significant increase in the intensity of the band in the inoculated control and Pf1 treatment in all the varieties. Isoform analysis of PPO showed the induction of PPO in P. fluorescens treated plants challenged with P. grisea.     

Influence of phylloplane colonizing biocontrol agents on the black spot of rose caused by Diplocarpon rosae

Abstract

An attempt was made to study the biocontrol efficacy of antagonistic microorganisms in phylloplane of rose cv. Edward to manage the black spot (Diplocarpon rosae) disease. Eight antagonistic microorganisms were tested in vivo against the black spot pathogen. Among these, Trichoderma viride and Pseudomonas fluorescens pf1 reduces the mycelial growth significantly. These two biocontrol agents were evaluated for their ability to induce defense-related enzymes and chemicals in plants. Increased activity of phenylalanine ammonia lyase (PAL), peroxidase (PO), polyphenoloxidase (PPO) and total phenolics were recorded in all the biocontrol agents treated leaves. P. fluorescens Pf1 recorded early and increased synthesis of the entire defense-related enzymes and total phenol within 6 days. The application of biocontrol agents induced the defense-related enzymes involved in phenyl propanoid pathway in addition to direct antagonism, which collectively contribute for enhanced resistance against invasion of Diplocarpon rosae in rose.     

Integrated control of fruit rot and powdery mildew of chilli using the biocontrol agent Pseudomonas fluorescens and a chemical fungicide

A combined strategy of chilli fruit rot and powdery mildew control consisting of reduced fungicide dose and biological control with antagonistic Pseudomonas fluorescens (Pf1) was evaluated. The sensitivity of P. fluorescens to fungicide azoxystrobin at different concentrations (100, 150, 200, 250 and 300 ppm) was tested by turbidometric method. The grown bacterium (Pf1) was tolerant to all concentrations of azoxystrobin. In two field trials, Pf1 tested in combination with reduced concentration of azoxystrobin was highly efficient in management of both diseases of chilli. Biological control of Colletotrichum capsici and Leveillula taurica with P. fluorescens (Pf1) was effective but less so than fungicide alone at the standard dose. However, combination of the biological control agent with a 50% reduction of fungicide dose was as effective as the standard fungicide alone. Application of P. fluorescens along with azoxystrobin significantly increased the survival of Pf1 in the phylloplane of chilli. Further, there were ‘twofold’ increases in activities of peroxidase, polyphenol oxidase, phenylalanine ammonia-lyase, β-1,3-glucanase, chitinase and phenolics in plants treated with Pf1 plus azoxystrobin.     

The Effect of Pseudomonas fluorescens and Fusarium oxysporum f.sp. cubense on Induction of Defense Enzymes and Phenolics in Banana

The effect of Pseudomonas fluorescens treatment and Fusarium oxysporum f. sp. cubense inoculation on induction of phenylalanine ammonia-lyase (PAL), peroxidase (POX), chitinase, -1,3-glucanase and accumulation of phenolics in banana (Musa sp.) was studied. When banana roots were treated with P. fluorescens strain Pf10, a two-fold increase in phenolic content in leaf tissues was recorded 3 – 6 d after treatment. Challenge inoculation with F. oxysporum, the wilt pathogen, steeply increased the phenolic content in P. fluorescens-treated banana plants. Significant increase in POX activity was detected 6 – 9 d after P. fluorescens treatment. PAL, chitinase and -1,3-glucanase activities increased significantly from 3 d after P. fluorescens treatment and reached the maximum 6 d after treatment. Challenge inoculation with F. oxysporum further increased the enzyme activities. These results suggest that the enhanced activities of defense enzymes and elevated content of phenolics may contribute to bioprotection of banana plants against F. oxysporum.     

Biological control of rice root-knot nematode,Meloidogyne graminicola through mixture of Pseudomonas fluorescens strains

The plant growth promoting rhizobacterium, Pseudomonas fluorescens strains PF1, TDK1, and PY15 were evaluated individually and in combinations for their efficacy against root-knot nematode, Meloidogyne graminicola, in rice plants under in vitro, glass house and field conditions. Culture filtrates of these strains either individually or as mixture inhibited egg hatching and caused mortality of juveniles of M. graminicola in vitro. The efficacy was more pronounced when filtrates of the strain were used as mixtures than as individual strains. Mixtures of P. fluorescens strains signficantly reduced M. graminicola infestation when applied as bacterial suspensions through seed treatment. The higher activity of peroxidase and chitinase enzymes was observed in plants treated with P. fluorescens mixtures than the plants treated with individual strains, two strain mixtures and untreated control. In field trials on rice, talc formulations of the P. fluorescens strains individually as well as mixtures were evaluated as seed treatment, soil treatment and combination of both. A mixture of the three strains was the most effective when applied either as seed + soil treatment or as seed treatment alone. The introduced P. fluorescens strains survived endophytically on rice roots. The application of the P. fluorescens mixture PF1 + TDK1 + PY15 in seed + soil treatment resulted in higher grain yield which provided a 27.3% increase over the control followed by P. fluorescens mixture PF1 + TDK1 + PY15 in seed treatment alone, which increased the grain yield of rice by 24.7% compared to the control.     

Induced systemic resistance to tomato leaf curl virus and increased yield in tomato by plant growth promoting rhizobacteria under field conditions

The talc-based formulation of two Pseudomonas fluorescens strains (Pf1 and VPT10) and its mixture (with and without chitin) were tested against tomato leaf curl virus in tomato under greenhouse and field conditions. The mean percentage of tomato leaf curl virus infected plants were significantly lower (25%) with less symptom severity and delayed symptom expression up to nine additional days in Pseudomonas with chitin (VPT10 + chitin) treated tomato plants compared to non-bacterised control plants upon challenge inoculation with tomato leaf curl virus. Tomato leaf curl virus was partially purified and antiserum was developed. Using the antiserum the tomato leaf curl virus was detected in symptomatic leaves and in whitefly vector through direct antigen coating enzyme linked immunosorbent assay which revealed the low virus titre in Pseudomonas treated plants (VPT10 + chitin) and insect vector compared to untreated tomato plants. The results indicate the potentiality of plant growth promoting rhizobacteria strains and talc-powder formulations in the effective management of this tomato leaf curl virus in tomato under field conditions.     

Bacterial antagonists and hexanal-induced systemic resistance of mango fruits against Lasiodiplodia theobromae causing stem-end rot

Induction of defense-related enzymes, such as phenylalanine ammonia lyase (PAL), peroxidase (PO), polyphenol oxidase (PPO), superoxide dismutase (SOD) and catalase (CAT) due to bacterial antagonists viz., Pseudomonas fluorescens (Pf1) and Bacillus subtilis (EPCO16) and plant-derived lipoxygenase volatile compound hexanal, were studied in mango fruits against Lasiodiplodia theobromae causing stem-end rot disease. The results showed increased induction of all the defense-related enzymes in mango fruits 3–5 days after dipping treatment with combination of bacterial antagonists and hexanal when compared to untreated control treatment and treatment with fungicide carbendazim in storage condition. The increased activity was observed up to 3 days after treatment and thereafter declined. Further, increased expression of specific isoforms of PO, PPO, SOD and CAT were also observed in the treatment effect of P. fluorescens (0.5%)?+?hexanal (0.02%) treated fruits against L. theobromae. From the results obtained, it is inferred that due to the enhancement of defense-related enzymes via the phenylpropanoid pathway and due to secretion of secondary metabolites that would play significant role in hindering the pathogen quiescence and further invasion in mango fruits and thereby prevent the fruit rot.     

Plant Growth-Promoting Bacteria Facilitate the Growth of Barley and Oats in Salt-Impacted Soil: Implications for Phytoremediation of Saline Soils

Plant growth-promoting bacteria (PGPB) strains that contain the enzyme 1-amino- cyclopropane-1-carboxylate (ACC) deaminase can lower stress ethylene levels and improve plant growth. In this study, ACC deaminase-producing bacteria were isolated from a salt-impacted (～50 dS/m) farm field, and their ability to promote plant growth of barley and oats in saline soil was investigated in pouch assays (1% NaCl), greenhouse trials (9.4 dS/m), and field trials (6–24 dS/m). A mix of previously isolated PGPB strains UW3 (Pseudomonas sp.) and UW4 (P. sp.) was also tested for comparison. Rhizobacterial isolate CMH3 (P. corrugata) and UW3+UW4 partially alleviated plant salt stress in growth pouch assays. In greenhouse trials, CMH3 enhanced root biomass of barley and oats by 200% and 50%, respectively. UW3+UW4, CMH3 and isolate CMH2 also enhanced barley and oat shoot growth by 100%–150%. In field tests, shoot biomass of oats tripled when treated with UW3+UW4 and doubled with CHM3 compared with that of untreated plants. PGPB treatment did not affect salt uptake on a per mass basis; higher plant biomass led to greater salt uptake, resulting in decreased soil salinity. This study demonstrates a method for improving plant growth in marginal saline soils. Associated implications for salt remediation are discussed.     

Induction of defense-related proteins by mixtures of plant growth promoting endophytic bacteria against Banana bunchy top virus

Rhizosphere and endophytic bacterial isolates from the roots and corms of banana were tested for their biocontrol efficiency against Banana bunchy top virus (BBTV). Molecular characterization using RAPD and microsatellite markers revealed genomic variability in the endophytic Pseudomonas and Bacillus isolates. Bio-formulations of mixtures of the rhizobacterial isolate Pseudomonas fluorescens (Pf1) and endophytic Bacillus spp. (EPB22) were effective in reducing the incidence of BBTV under green-house (80%) and field conditions (52%). Reduction in virus titer (0.64) was noticed in the plants treated with compatible mixtures of rhizobacterial and endophytic bacterial isolates as evidenced by ELISA, in comparison to control plants (1.69). In addition to disease control, a significant increase in the yield (53.33%) was noticed in the bacterized plants when compared to the control plants. Pathogenesis-related (PR) proteins, chitinase and β-1,3-glucanase and defense-related proteins, peroxidase, polyphenol oxidase, phenylalanine ammonia-lyase and phenolic compounds were significantly activated in the bacterized plants, thus inducing resistance against bunchy top virus. Populations of endophytic bacteria also remained high and stable throughout the growing period. Thus, application of mixtures of rhizosphere and endophytic bacteria increases yield and has a potential role in inducing resistance against Banana bunchy top virus.     

Plant growth-promoting rhizobacteria-mediated induction of phenolics in pea ( Pisum sativum) after infection with Erysiphe pisi

Qualitative and quantitative estimation of phenolic compounds was done through high performance liquid chromatography (HPLC) in different parts of pea (Pisum sativum) after treatment with two plant growth-promoting rhizobacteria (PGPR), viz., Pseudomonas fluorescens (strain Pf4) and Pseudomonas aeruginosa (referred to here as Pag) and infection by Erysiphe pisi. The phenolic compounds detected were tannic, gallic, ferulic, and cinnamic acids on the basis of their retention time in HPLC. In all the treated plants, synthesis of phenolic compounds was enhanced. The induction of gallic, ferulic, and cinnamic acids was manyfold more than those in the control. Maximum accumulation of phenolic compounds was observed in plants raised from PGPR-treated seeds and infection with E. pisi. Under pathogenic stress, Pag performed better because a relatively higher amount of phenolics was induced compared with plants treated with Pf4. Received: 20 August 2001 / Accepted: 20 September 2001     

Shoot endophytic plant growth-promoting bacteria reduce cadmium toxicity and enhance switchgrass (<Emphasis Type="Italic">Panicum virgatum</Emphasis> L.) biomass

The aims of the study were to increase the biomass and to alleviate the deleterious effects of cadmium (Cd) in the switchgrass cultivars (Panicum virgatum L.) Alamo and Cave-in-Rock (CIR) under cadmium (Cd) stress using Cd-tolerant shoot endophytic plant growth-promoting bacteria (PGPB). Four shoot endophytic bacterial strains, viz. Bc09, So23, E02, and Oj24, were isolated from the above-ground parts of plants grown in a Cd-polluted soil and were successfully identified by 16S rRNA gene sequencing as Pseudomonas grimontii, Pantoea vagans, Pseudomonas veronii, and Pseudomonas fluorescens, respectively. These four strains were adapted to high CdCl₂ concentrations as they had higher Cd uptake capacities. In addition, they possessed a huge amount of growth regulatory activities e.g., indole acetic acid production, 1-aminocyclopropane-1-carboxylic acid deaminase (ACCD) activity, and phosphate solubilization. Growth particularly the height and biomass of both cultivars increased significantly in response to PGPB inoculation in the 20 µM CdCl₂ stress. The shoot biomass of the PGPB-inoculated Alamo was higher than the CIR under Cd stress. Interestingly, the level of Cd inside PGPB-inoculated plant tissues and the translocation factors were lower compared with the noninoculated Cd control plants. CIR plants exhibited higher Cd content than Alamo plants. Through confocal microscopy, green fluorescence was observed in roots and leaf tissues 2 days after the inoculation of green fluorescent protein (GFP)-labeled bacteria in Alamo, which confirmed the successful colonization of bacteria inside the plant tissues. These shoot endophytic PGPB and switchgrass interactions are useful for the sustainable biomass production of bioenergy crop in a Cd-contaminated environment.     

Metabolic profile and antioxidant responses during drought stress recovery in sugarcane treated with humic acids and endophytic diazotrophic bacteria

Water deficit is the major yield‐limiting factor for sugarcane crop production that can be enhanced by inoculating with plant growth promoting bacteria (PGPB) combined with humic substances. The aim of this work was to examine changes to the metabolic profile and antioxidant enzyme activity of sugarcane treated with PGPB and humic acid (HA) after drought and then rehydration. The drought was imposed by withholding irrigation for 21 days thereby measuring enzyme activity, metabolic profile and photosynthetic rate 1 week after rehydratation. Growth of plants treated with HA, PGPB and with both treatments combined (PGPB + HA) was higher than control plants. The antioxidant enzymes superoxide dismutase (SOD), catalase (CAT) and ascorbate peroxidase (APX) activities remained higher after rehydration only in plants treated with HA. Plants treated with HA and PGPB + HA exhibited increased transpiration, stomatal conductance and net photosynthesis than plants treated with PGPB. The PGPB‐treated plants exhibited drought resistance that resembled ‘delayed stress onset’, which is a new term for preserving water in the plants tissues. Water preservation in plants treated with PGPB was corroborated by higher relative water content (RWC) than control plants at the end of the drought period. Plants treated with HA + PGPB exhibited the highest water potential after rehydration and high RWC. Osmotic adjustment in the other treatments (control, HA and PGPB) was indicated by a new pattern of metabolic response after rehydration, including generally enhanced carbohydrates and proteins and specific changes induced by HA‐enhancing aromatic compounds, whereas PGPB exhibited enhanced fatty acids and other aliphatic H species. Humic acids assist with drought stress recovery by inducing antioxidant enzyme activity whereas PGPB induced preservation of leaf water potential and RWC by closing stomata efficiently, resulting in plant water preservation.     

Endophytic bacteria mediate plant resistance against cotton bollworm

Abstract

The efficacy of endophytic bacterial strains was evaluated in cotton against American bollworm infestation under greenhouse conditions. Among the 103 endophytic bacterial strains, the Bacillus strains (EPCO 102 and EPCO 16) and Pseudomonas fluorescens strain Pf1 significantly reduced the bollworm incidence. Talc-based bioformulation of EPCO 102, EPCO 16 and P. fluorescens Pf1 with and without chitin in inducing systemic resistance was tested against bollworm. The application of the bioformulation through seed, soil and foliar spray significantly reduced the bollworm incidence. The amendment of chitin in the formulation further reduced the pest incidence. Maximum bollworm reduction by endophytic bacterial strains EPCO 102, EPCO 16 and Pf1 strain with chitin was recorded. In addition, endophytic bacterial bioformulation with chitin induced more and timely activities of chitinase, β-1, 3-glucanase, peroxidase, polyphenol oxidase, phenylalanine ammonia-lyase and phenol in cotton plants infested with Helicoverpa armigera.     

Growth promotion and disease suppression ability of Pseudomonas fluorescens in acid lime

Biochemical analysis was carried out for 10 isolates of Pseudomonas fluorescens. All isolates were found positive for siderophore and indole acetic acid production (except Pf IX) and phosphate solubilisation (except Pf VII). Biochemically efficient strains (Pf I, Pf IV, Pf VII and Pf IX) were selected for management of root rot, collar rot and damping off caused by Phytophthora parasitica, Pythium sp. and Fusarium solani. The activity of defence-related enzymes like esterase, peroxidase and polyphenol oxidase was also detected by polyacrylamide gel electrophoresis (PAGE). Consistent appearance of esterase isozyme bands was visualised, in the range of 0.193–0.349. The highest Rm value 0.349 was observed on Pf IV. Whereas, for peroxidase, Rm value ranged between 0.302 and 0.373 and for polyphenol oxidase, it was in the range of 0.211–0.800. P. fluorescens Pf IV was found significantly effective to arrest the per cent mycelial growth of P. parasitica (55.20%), Pythium sp. (65.33%) and F. solani (64.67%). Among bioagents, seed treatment and soil drenching with Pf IV at 15 and 30 days after sowing were found effective to reduce per cent disease incidence (30.55%) at 120 days after emergence. Seed treatment with copper oxychloride at 3g/kg seed and metalaxyl at 2 g/kg seed were also found effective.     

Pseudomonas fluorescens enhances resistance and natural enemy population in rice plants against leaffolder pest

Plant growth promoting fluorescent pseudomonad strains Pf1, TDK1 and PY15 were evaluated for their efficacy against leaffolder ( Cnaphalocrocis medinalis ) pest in rice plants under field conditions individually and in combinations. Application of mixtures of Pseudomonas fluorescens strains Pf1, TDK1 and PY15 significantly reduced the leaffolder damage in rice plants compared with untreated control. Interestingly, natural enemy population in plots treated with P. fluorescens was greater than the chemical and untreated controls. Further, support for these results was gathered by assaying activities of polyphenol oxidase (PPO) and lipoxygenase (LOX) under glasshouse conditions. The results showed the higher activity of PPO and LOX in plants treated with P. fluorescens mixtures (Pf1 + TDK1 + PY15) than the plants treated with individual strains, chemical and untreated controls. Further, fluorescent pseudomonad mixtures increased the rice yield compared with individual strain and non-bacterized treatments. The present study reveals that in addition to plant growth promotion, plant growth-promoting rhizobacterial (PGPR) strains-mediated induction of PPO and LOX in rice plants could be involved in enhanced natural enemy populations and resistance mechanisms against leaffolder attack.