Exploration, isolation and characterization of indigenous rhizobacteria from patchouli rhizosphere as PGPR candidates in producing IAA and solubilizing phosphate

Microorganisms that are active and aggressive colonizing the rhizosphere are known as rhizobacteria. They are able to act as biofertilizers, bioprotectants, biostimulants and bioremediation. This study aims to identify and characterize groups of rhizobacteria present in the patchouli rhizosphere that can produce IAA compounds and have the ability to solubilize phosphate in the soil. Soil samples were taken from the patchouli rhizosphere at Purwosari Village, Nagan Raya, Aceh Province, Indonesia. This study used quantitative and qualitative descriptive analysis through serial dilutions to obtain rhizobacterial strains. Parameters observed were macroscopic and microscopic characteristics, gram test, IAA production and phosphate solubilization. The study obtained 37 isolates of rhizobacteria from Purwosari (PS), comprising 25 isolates of gram positive and 12 isolates of gram negative. The rhizobacteria PS 5/1 produced the lowest IAA at 21.66 ppm, whereas isolate 5/6 C produced the highest IAA at 83.38 ppm. Twenty-five isolates of rhizobacteria could solubilize phosphate while the remaining 12 isolates did not have this ability. The rhizobacteria PS 7/1 resulted in the highest PSI at 2.55 and isolates PS 8/7 produced the lowest PSI at 1.33. The rhizobacteria isolates that can produce IAA and phosphate solubilizing have the potential to be used as PGPR candidates.

Plant growth promoting potential of multifarious endophytic Pseudomonas lini strain isolated from Cicer arietinum L.

Chickpea (Cicer arietinum), one of the major pulse crops in India, endured extreme reduction in production due to various abiotic and biotic stresses. Endophytic bacteria residing in the nodules and roots of chickpea plants enable host in combating these stresses. Twenty one endophytic bacteria isolated from nodules and roots of chickpea were screened for multiple plant growth promoting traits like ammonia, organic acid, siderophore, hydrogen cyanide (HCN) and phytohormone indole acetic acid (IAA) production. Out of these, 86% isolates produced ammonia, around 50% isolates produced organic acid, HCN and siderophore, 29% isolates produced ACC (1-aminocyclopropane-1-carboxylic acid) deaminase, while only 14% isolates solubilized phosphate. Interestingly, all the isolates were able to produce IAA ranging from 11.6–85.2 μg/ml, isolate CPJN 13 being the maximum IAA producer (85.5±2.33 μg/ml). Isolate CPJN13 was selected for IAA optimization studies. The yield of IAA increased up to 4 fold i.e. 331±4.96 μg/ml at optimized conditions. IAA production was also confirmed by TLC and HPLC analysis of crude IAA extract. The application of CPJN13 on chickpea seedlings resulted in significant increase in plant growth parameters. The 16S rDNA sequencing of CPJN13 revealed its similarity with Pseudomonas lini strain and submitted to NCBI with accession number MF574502. To best of our knowledge, this is the first report of the presence of P. lini as endophyte in chickpea nodules. The results of this study imply that the endophytic P. lini has a potential role to enhance the plant growth.

Isolation, Identification and Evaluation of Indigenous Plant Growth Promoting Bacterium Klebsiella pneumoniae PNE1

Application of chemical fertilizer is an integral practice to optimize crop productivity, but the dominant use of chemical fertilizers contributes largely to the deterioration of the environment, leads to loss of soil fertility, increases pollution, and causes hazardous diseases. Hence, the chemical fertilizers, pesticides and other supplements are being replaced by the plant growth promoting bacteria (PGPB) due to their improved potency and environment friendly nature. Plant growth-promoting bacteria (PGPB) can enhance plant growth by a wide variety of mechanisms like Phosphate (P) solubilization, Potassium solubilisation, siderophore production, biological nitrogen fixation and Indole acetic acid (IAA) production. The Klebsiella species is also known to exhibit important PGP traits like solubilization of phosphate, phytohormone production and good germination potential. In present study the Klebsiella pneumoniae PNE1 was selected from the isolates obtained from vegetable waste collected from Kadi market. The isolate was selected on the basis of its ability for Nitogen fixation, Phosphate solubilization, Potassium solubilization, IAA production, EPS production and biopolymer degradation. The molecular identification through 16S rRNA gene sequence, confirmed the isolate as Klebsiella pneumonia PNE1. Quantitative analysis of ammonia production revealed that isolate Klebsiella pneumonia PNE1 produced 0.5 µg/ml of ammonia (NH3) on 6th day of incubation and produced 0.09 µg/ml Nitrite after 8th day of incubation. The Phosphate solubilisation Index (SI) of the isolate was 4.16 and the isolate released 177.50 μg/ml Phosphate. The qualitative estimation of Potassium solubilisation by the isolate Klebsiella pneumoniae PNE1 in terms of Potassium solubilisation zone was found to increase gradually from day 1 to 7 days and was maximum at 2nd day with a KSI of 3.6. The isolate Klebsiella pneumoniae PNE1 released 29.94 mg/l Potassium on 21th day of incubation respectively. The IAA production was found to be 94.96 µg/ml. The maximum the EPS yield was 11.3 mg/ml. The Klebsiella pneumonia PNE1 had capacity to degrade Cellulose, Pectin and Xylan i.e. all biopolymers tested. The antibiotic susceptibility test indicated that isolate was sensitive to all 22 antibiotics tested. The Klebsiella pneumonia PNE1 thus, shows important plant growth promoting traits and can be used in a bio-fertilizer formulation for sustainable agriculture.

Fermentation of Washed Rice Water Increases Beneficial Plant Bacterial Population and Nutrient Concentrations

Washed rice water (WRW) is said to be a beneficial plant fertilizer because of its nutrient content. However, rigorous scientific studies to ascertain its efficiency are lacking. The purpose of this study was to determine the effect of fermenting WRW on the bacterial population and identification, and to measure how fermentation affects the nutrient composition of WRW. Rice grains were washed in a volumetric water-to-rice ratio of 3:1 and at a constant speed of 80 rpm for all treatments. The treatments were WRW fermented at 0 (unfermented), 3, 6, and 9 days. Bacterial N fixation and P and K solubilization abilities in the fermented WRW were assessed both qualitatively and quantitatively. The isolated bacterial strains and the WRW samples were also tested for catalase and indole acetic acid (IAA) production ability. Significantly greater N fixation, P and K solubilization, and IAA production were recorded after 3 days of fermentation compared with other fermentation periods, with increases of 46.9–83.3%, 48.2–84.1%, 73.7–83.6%, and 13.3–85.5%, respectively, in addition to the highest (2.12 × 108 CFU mL−1) total bacterial population. Twelve bacteria strains were isolated from the fermented WRW, and the gene identification showed the presence of beneficial bacteria Bacillus velezensis, Enterobacter spp., Pantoea agglomerans, Klebsiella pneumoniae and Stenotrophomonas maltophilia at the different fermentation periods. All the identified microbes (except Enterobacter sp. Strain WRW-7) were positive for catalase production. Similarly, all the microbes could produce IAA, with Enterobacter spp. strain WRW-10 recording the highest IAA of up to 73.7% higher than other strains. Generally, with increasing fermentation periods, the nutrients N, S, P, K, Mg, NH4+, and NO3− increased, while pH, C, and Cu decreased. Therefore, fermentation of WRW can potentially increase plant growth and enhance soil health because of WRW’s nutrients and microbial promotional effect, particularly after 3 days of fermentation.

Virulence and indole-3-acetic acid (IAA) biosynthesis ability of Turkish Pseudomonas savastanoi pv. savastanoi isolates and susceptibility of some native olive genotypes

Aim of study: To evaluate the virulence and indole-3-acetic acid (IAA) biosynthesis ability of several Turkish P. savastanoi pv. savastanoi isolates and the susceptibility of some native genotypes to olive knot. Area of study: The Aegean, Marmara, and Mediterranean Regions of Turkey. Material and methods: 101 isolated bacteria were identified on the basis of biochemical, PCR for amplification of the bacterial iaaL gene, and pathogenicity tests. The virulence of the isolates was determined in a randomized experimental trial carried out by stem inoculation of pot-grown seedlings of olive (cv. ‘Manzanilla’) in the growing chamber. The amounts of IAA produced by the isolates were determined colorimetrically. The susceptibility of native olive genotypes was evaluated on 2-yr old plants inoculated with two distinct strains. Main results: Tested P. savastanoi pv. savastanoi isolates showed significant differences in virulence found to be associated with their geographical origin. The isolates produced IAA amounts varied from 148.67 to 0.3 μg mL-1. The geographical variation in IAA biosynthesis ability of the isolates was observed. No correlation (R=0.0225) was determined between virulence and IAA amounts of the isolates. Native olive genotypes indicated different susceptibility levels to the olive knot pathogen. No genotype tested had complete resistance. However, low susceptible genotypes (‘Memecik’, ‘Ayvalık’ and ‘Uslu’) were identified. Some genotypes had variable reactions depending on the isolate used. Research highlights: The results undergird the differences in the virulence and IAA production of the isolates within the area and also between geographical locations. Genotypes with low susceptibility can be used as genitors in further breeding studies.

Detection and Identification of Plant Growth Promoting Bacteria from Sorghum (Sorghum bicolor L. Moench) Rhizosphere Soil in Northern Ethiopia

Plant growth promoting rhizobacteria are the bacteria which subsist inside and outside of the plant tissue and promote plant growth through direct or indirect mechanisms. To increase sorghum production and productivity we utilize herbicides and chemical fertilizers to overcome sorghum production constraints, but those chemicals have negative side effects. The current study was conducted with the objective of isolation of PGPR from sorghum rhizosphere and screening for primary growth related trait, evaluation of potential PGPR at greenhouse for sorghum growth performance and identify through biochemical characterization. So that, in this study a total of 117 plant growth promoting rhizobacteria were isolated from the rhizosphere of 12 sorghum (Sorghum bicolor L. Moench) genotype by cultivating using 3 collected soil samples from the northern part of Ethiopia (Amhara and Tigray regional states) in greenhouse. Isolated bacteria were screened for primary growth promoting traits such as phosphate solubilization test, IAA production test at different concentration of L-tryptophan and ammonia production test. From the isolated bacteria 28% solubilized Phosphorous, 78% produced IAA at different concentration of tryptophan. The greatest IAA production was scored at 100 mg/L of tryptophan and the lowest production of IAA was scored at 150 mg/L of tryptophan, 69% of isolated bacteria produced ammonia. Hence, 15% of isolated bacteria fulfilled the above primary screening test and used for further greenhouse evaluation. Accordingly, eighteen bacteria were tested for greenhouse experiment using completely randomized design and all 18 isolates were significantly increased all the agronomic parameter as compared to the control such as plant shoot height, plant shoot fresh and dry weight, root length, root fresh and dry weight at p < 0.01 and P ≤ 0.001. Two isolates G6E29 and G4E19 had significantly increased all the parameter but two isolates (G12E19 and G3E40) were statistically non-significant for root fresh weight compared to the control. These 18 potential isolates were characterized morphologically and biochemically. Eight isolates were grouped at Pseudomonas genera. Six isolates were grouped at Azotobacter and the rest four isolates were grouped at Bacillus genera. Thus, the use of plant growth promoting rhizosphere bacteria could be useful to improve sorghum production and productivity. However, further molecular identification and evaluation of the isolates exhibiting multiple plant growths promoting traits on plant-microbe interaction for economic crop of Ethiopia is needed to uncover their efficacy as effective plant growth promoting rhizosphere bacteria.

Effect of Plant-Growth-Promoting Fungi on Eggplant (Solanum melongena L.) in New Reclamation Land

Land reclamation may expand the supply of usable land for food security. Immature soil is not suitable for plant growth and needs to be amended by the addition of organic matter and plant growth-promoting (PGP) microorganisms. However, the effects of different PGP fungi on plant growth in immature soil are largely unexplored. In order to obtain beneficial soil microorganisms with a good PGP ability in new reclamation land, 162 fungal isolates were isolated from different abandoned wastelands, four isolates of which were obtained in this study by the screening of P solubilization, siderophore production, and indole acetic acid (IAA) production. The result of this study revealed that isolate HZ123 had the highest ability to solubilize P and produce siderophores and IAA, followed by HZ23, HZ10, and HZ06. Based on the results of morphological and molecular analyses, isolate HZ06 was identified as Penicillium oxalicum, isolates HZ23 and HZ10 were identified as Aspergillus brunneoviolaceus, and isolate HZ123 was identified as Aspergillus tubingensis. Furthermore, the results of in vivo PGP assays demonstrated that isolate HZ123 has a minimal negative effect on the growth of eggplant; however, the other three isolates, particularly isolate HZ06, caused the greatest increase in eggplant biomasses. Overall, these results indicate that isolate HZ06 has great potential as a PGP fungus to develop biofertilizer for application in eggplant production in immature soil from new reclamation land.

Multifarious functional traits of free-living rhizospheric fungi, with special reference to Aspergillus spp. isolated from North Indian soil, and their inoculation effect on plant growth

Purpose Rhizospheric soil fungi are critical for plant and soil health. However, their multiple functional traits and impact on plant growth have not been systematically explored. Methods During this study, biochemical traits of 73 indigenous soil fungal isolates and 15 unidentified isolates related to plant growth promotion and production of extracellular enzymes were studied. Results Forty four (65.67%) of the total isolates produced indole acetic acid (IAA) followed by siderophore (52.23%), phosphate solubilization (37.31%), and antibiotic (11.93%). 91.04% of the studied isolates produced ammonia whereas 28.35% produced organic acid. Extracellular enzyme activities of lipase, amylase, chitinase, and cellulase were detected among 95.52%, 61.11%, 35.82%, and 41.79% isolates, respectively. Based on these activities, 73 fungal isolates were categorized into different biotypes. Quantitative analysis of IAA production and phosphate solubilization was carried out for Aspergillus, Penicillium, and Rhizopus isolates. Aspergillus isolates exhibited varying activities of IAA production and phosphate solubilization. Most of the Aspergillus isolates and some other fungi demonstrated multiple activities. Based on the multiple traits of selected fungal isolates, Aspergillus sp-07, Penicillium sp-03, and Rhizopus sp-02 were further evaluated in different combinations for their inoculation effect on the growth and yield of wheat under field conditions. Conclusions The results indicated that these isolates could be developed into bio-inoculants to enhance plant growth. The consortium of these three isolates was also found to be compatible and beneficial for plant growth.

Indole-3-acetic acid production by rhizobacteria Bacillus spp. to various abiotic stress factors

Indole-3-acetic acid (IAA) phytohormone plays an essential role in forming and initiating main, lateral, and adventitious roots in vegetative propagation. Plants are receiving IAA naturally from a diverse group of soil-plant associated rhizobacteria. However, IAA synthesis by rhizobacteria is influenced by abiotic growth conditions. Three indigenous Bacillus isolates were subject to in vitro assay for the effects of abiotic factors (temperature, salinity and pH) on growth and IAA production. All isolates grew well between 25 - 40°C, and only B. megaterium UPMLH3 was capable of synthesising IAA (21.18 µg/ml) at 40°C. All three bacterial growth under saline stress were slightly dropped over control (0% NaCl), but still producing IAA up to 1% NaCl condition. B. cereus UPMLH24 revealed high resistance to salinity up to 5% NaCl. The optimum growth of all three Bacillus spp. was at pH 7. B. cereus UPMLH1 and UPMLH24 discovered higher IAA production in slightly alkaline conditions (pH 8). Each rhizobacterium shows different physiology trait against each abiotic factor. However, the multiple tolerance ability of PGPR against abiotic factors is an indication that its ability to survive under harsh soil and plant environments while delivering benefits to the plant. Thus, B. cereus UPMLH1, B. megaterium UPMLH3 and B. cereus UPMLH24 might serve as potential biofertiliser, enhancing the growth performance of test plants at various environmental conditions.