Free-living Nitrogen-fixing Bacteria Research Articles

Isolation, Screening and Identification of Free-Living Diazotrophic Bacteria from Salinated Arid Soils

Investigation of free-living nitrogen-fixing bacteria from the salinated arid soil ecosystems is important for developing novel biological solutions for dryland agriculture. The present work reports isolation, screening and identification of two strains from salt-affected soils of Latakia (Syria) and Aral Sea basin (Uzbekistan): Gracilibacillus sp. Str01 and Halomonas sp. Str02. Nitrogenase activity of Gracilibacillus sp. Str01 and Halomonas sp. Str02 (as acetylene reduction) was 2.24 ± 0.61 and 0.63 ± 0.11 C2H4 nmol h–1, respectively and these strains may be considered free-living diazotrophic bacteria from arid saline soils.

Biodegradation of hexabromocyclododecane by Rhodopseudomonas palustris YSC3 strain: A free-living nitrogen-fixing bacterium isolated in Taiwan.

The persistent organic pollutant, brominated flame retardant, hexabromocyclododecane (HBCD), identified as an emerging contaminant has been detected in various environmental matrix. The increased level of this toxic organic compound in the environment has been associated with serious human health risks. The results obtained from this study revealed that various Rhodopseudomonas palustris strains isolated from paddy soil in Taiwan possessed good HBCD biodegradation capability when they were cultured aerobically. Among these strains, YSC3 was considered as one of the most potential isolates for HBCD degradation. The optimum HBCD biodegradation occurred at neutral pH and at 35°C in all our pH and temperature tests at an initial HBCD concentration of 1ppm. HBCD degradation kinetics generally decreased with the increase of initial HBCD concentration. The study also suggested that the cultivation temperature played a vital role on YSC3 for its initiation of cellular HBCD degradation. The relative-molar ratio of the released bromide ions during the biodegradation of HBCD was observed in the range between 1 and 3.5, suggesting that the debromination reactions occurred. Concomitant with the loss of HBCD, there was a concurrent production of two metabolites, pentabromocyclododecanol and pentabromocyclododecene, which were determined by liquid chromatography and mass spectrometry techniques. On the basis of the obtained results, the possible biodegradation pathways were also proposed in this study.

Study on the efficiency of free-living nitrogen fixing bacteria isolated from rice rhizospheresoil on auxin and giberellin production

Study on the efficiency of free-living nitrogen fixing bacteria isolated from rice rhizospheresoil on auxin and giberellin production

Isolation, Identification and Screening for Nitrogen Fixing Activities by <i>Azotobacter chroococcum</i> Isolated from Soil of Keffi, Nigeria as Agent for Bio-fertilizer Production

Free living nitrogen fixing bacteria are those that lives freely on rhizosphere of a young growing plant or those living in a close association with root of plants (Rhizobacteria) but not symbiotically. In most tropical soil, Nitrogen and other essential nutrients element are the most limiting nutrients which deficiencies could lead to slow growth and reduced crop production. Free living nitrogen fixing bacteria has the potential of secreting nitrogenase enzymes and produces organic acids such as glycolic, acetic, malic, succunic acids which fix atmospheric nitrogen directly into the soil for plants growth and development. To this end, the study on isolation, identification and screening of Azotobacter chroococcum from soil of Keffi, Nasarawa State, Nigeria was carried out from May to September 2018. Soil samples were collected from eight different locations and Azotobacter strains were isolated and identified using standard microbiological methods. The 16SrRNA gene sequence analysis of the strain showed maximum similarity of 96% with Azotobacter chroococcum of the reference type strain deposited in RDP Gen Bank database. Azotobacter strains isolated from four different locations showed coloured zone ranging between 16-10mm. Similarly, Percentage amount of nitrogen released by each Azotobacter strain in the culture broth ranging between 1.19% - 5.11% in an increasing order.

Structural characterization and applications of a novel polysaccharide produced by Azospirillum lipoferum MTCC 2306.

Azospirillum lipoferum MTCC 2306, a free-living nitrogen fixing bacteria, has a doubling time of 1.7h in MPSS media. At the end of 28h at a pH of 7 and temperature of 30°C it produces 1.8 ± 0.013g/L biomass and 2.1 ± 0.018g/L of cyclic beta glucan (CβG) in MPSS medium with a yield coefficient (YP/S) of 2.1. This novel polysaccharide is a water-soluble cyclic biopolymer and is generally produced by Rhizobiaceae and predominantly made up of glucose. The CβG has a degree of polymerisation varying between 10 and 13 and has both α- and β-glycosidic linkages. It is not substituted with any functional groups such as acetates or succinates. Its ability to bind to aniline blue suggests that it can be a potential candidate for being used ascarrier in medical imaging as well as in reducing toxicity of textile effluents. It is able to encapsulate rifampicin, a hydrophobic drug and increase its aqueous solubility by 71%. So, CβG appears to have promising applications in the field of drug, food, cosmetic and nutraceutical industries.

Silvopastoral systems drive the nitrogen-cycling bacterial community in soil

ABSTRACT Intercropping tree legumes with forage grasses in a silvopastoral system can avoid pasture degradation benefiting the soil. In such a system, nitrogen (N) is supplied by symbiosis between legumes and bacteria. However, the pasture quality determines the action of free-living nitrogen-fixing bacteria, which possess nifH genes, which encode nitrogenase enzyme. Ammonium-oxidizing bacteria (AOB), involved in the nitrification step, can be evaluated by specific regions of the 16S rRNA corresponding to AOB. Thus, we investigated the influence of the introduction of tree legumes into a silvopastoral system on the community structure and abundance of total bacteria, diazotrophic bacteria and ammonium-oxidizing bacteria by DGGE (denaturing gradient gel electrophoresis) and real-time qPCR (quantitative PCR). The experiment involved nine plots of one hectare each, planted with sabia (Mimosa caesalpinifolia), a Gliricidia species (Gliricidia sepium), and a Brachiaria species (Brachiaria decumbens) in a randomized block design, forming three treatments: I-Brachiaria intercropped with sabia; II-Brachiaria intercropped with Gliricidia and III-Brachiaria only, with three replicates. The structures of the total bacterial and ammonium-oxidizing bacterial communities were influenced by tree legume introduction, possibly through modification of the soil chemical attributes. The copy numbers of total bacteria, ammonium-oxidizing bacteria and diazotrophic bacteria were higher in soils planted with legumes, which provided better conditions for microbial growth compared to planting with the Brachiaria species alone. Silvopastoral management with tree legumes improves the biological quality of soil, favouring the bacterial community linked to N-cycling.

Effects of Different Sources of Nitrogen on Endophytic Colonization of Rice Plants by Azospirillum sp. B510.

Azospirillum sp. B510, a free-living nitrogen-fixing bacterium isolated from the stems of rice (Oryza sativa cv. Nipponbare), was investigated to establish effective conditions for the colonization of rice plants. We analyzed the effects of the nitrogen sources KNO3, NH4Cl, urea (CO[NH2]2), and NH4NO3 at different concentrations (0.01–10 mM) on this colonization. Nitrogen promoted plant growth in a concentration-dependent manner, with minor differences being observed among the different nitrogen sources. Bacterial colonization was markedly suppressed on media containing NH4+ concentrations higher than 1 mM. Since concentrations of up to and including 10 mM NH4+ did not exhibit any antibacterial activity, we analyzed several factors affecting the NH4+-dependent inhibition of endophytic colonization, including the accumulation of the reactive oxygen species H2O2 and the secretion of the chemotactic substrate malic acid. The accumulation of H2O2 was increased in rice roots grown on 1 mM NH4Cl. The amounts of malic acid secreted from NH4-grown rice plants were lower than those secreted from plants grown without nitrogen or with KNO3. Although the bacterium exhibited chemotactic activity, moving towards root exudates from plants grown without nitrogen and KNO3-grown plants, this activity was not observed with root exudates from NH4+-grown plants. NH4+, but not NO3−, caused the acidification of growth media, which inhibited plant bacterial colonization. These NH4+-dependent phenomena were markedly suppressed by the stabilization of medium pH using a buffer. These results demonstrate that the type and concentration of nitrogen fertilizer affects the colonization of rice plants by Azospirillum sp. B510.

Diversity and characterization of Azotobacter isolates obtained from rice rhizosphere soils in Taiwan

Azotobacter species, free-living nitrogen-fixing bacteria, have been used as biofertilizers to improve the productivity of non-leguminous crops, including rice, due to their various plant growth-promoting traits. The purposes of this study were to characterize Azotobacter species isolated from rice rhizospheres in Taiwan and to determine the relationship between the species diversity of Azotobacter and soil properties. A total of 98 Azotobacter isolates were isolated from 27 paddy fields, and 16S rRNA gene sequences were used to identify Azotobacter species. The characteristics of these Azotobacter strains were analyzed including carbon source utilization and plant growth-promoting traits such as nitrogen fixation activity, indole acetic acid production, phosphate-solubilizing ability, and siderophore secretion. Of the 98 strains isolated in this study, 12 were selected to evaluate their effects on rice growth. Four species of Azotobacter were identified within these 98 strains, including A. beijerinckii, A. chroococcum, A. tropicalis, and A. vinelandii. Of these four species, A. chroococcum was predominant (51.0%) but A. beijerinckii had the highest level of nucleotide diversity. Strains within individual Azotobacter species showed diverse profiles in carbon source utilization. In addition, the species diversity of Azotobacter was significantly related to soil pH, Mn, and Zn. Members of the same Azotobacter species showed diverse plant growth-promoting traits, suggesting that the 98 strains isolated in this study may not equally effective in promoting rice growth. Of the 12 strains evaluated, A. beijerinckii CHB 461, A. chroococcum CHB 846, and A. chroococcum CHB 869 may be used to develop biofertilizers for rice cultivation because they significantly promoted rice growth. This study contributes to the selection of suitable Azotobacter strains for developing biofertilizer formulations and soil management strategies of Azotobacter for paddy fields.

Arsenic Contamination in Agricultural Soil Reduces Metabolic Activity of Total and Free-Living Nitrogen-Fixing Bacteria as Revealed by Real-Time qPCR

ABSTRACTLaboratory microcosms were used to assess the impact of arsenic (As) contamination on agricultural soil bacterial activity focusing free-living nitrogen fixers for three months. Periodically collected microcosm samples were analyzed by RT–qPCR following extraction of total RNA and cDNA preparation for assessing the metabolically active bacterial population. RT–qPCR data showed the gradual increase of 16S rRNA and nifH gene expression, and relative activity of diazotrophs in the enriched soil bacterial consortia under short time As exposure up to 20 ppm and 10 ppm, respectively. A similar trend of these variables was also noticed but up to 1 ppm As when incubating the bulk soil for the same duration. Reduced bacterial activity was noticed at higher concentration of As although in short time exposure. Extending the As exposure time, the bacterial activities in both enriched consortia and bulk soil were decreased. Although, the relative activity of diazotrophs in enriched consortia was increased in presence of 10 ppm As, the same was decreased in bulk soil when exposed to >1 ppm As for long time indicating susceptibility of nitrogen fixer to As contamination in soil. PCA of the data obtained also indicated a negative correlation between As concentrations and diazotrophic activity.

Co-Inoculation of Pleurotus ostreatus with Beneficial Bacteria Enhanced Substrate Utilization and Mushroom Production: An Implication for the Development of a Novel Bio-Fertilizer

The oyster mushroom (Pleurotus ostreatus) is one of the most widely cultivated edible mushrooms around the world. Mycelial colonization by beneficial bacteria has been shown to promote the growth of P. ostreatus and the other cultivated mushrooms. Therefore, present study was conducted to investigate the effect of beneficial bacteria which was isolated from soil, on growth, mushroom yield, primordia initiation of P. ostreatus, reducing sugar release, secretion of lignolytic enzymes such as laccase and manganese peroxidase aiming at developing a novel bacterial inoculum for P. ostreatus cultivation. Minimal salt medium (MSM) supplemented with cellulose and/or starch as a sole source of carbon and energy was used to isolate bacteria from soil whereas Ashby’s nitrogen free mannitol medium was used for the isolation of free-living nitrogen fixing bacteria. Pure cultures of bacterial isolates were obtained after several rounds of streaking on solid plates of the same medium, and co-cultured them with P. ostreatus to screen the isolates which did not cause inhibitory effects on mycelial growth. The beneficial isolates were identified based on Gram staining morphology and biochemical tests. While in vitro mycelial colonization studies were performed in liquid medium, the in vivo studies were conducted in pots filled with sterilized lignocellulosic solid substrate to evaluate beneficial effects of isolates on growth, mushroom yield, nitrogen content, primordia initiation of P. ostreatus and laccase and manganese peroxidase activities. Pots which did not receive any bacterial inoculum were treated as control. Heavy mycelial colonization by beneficial isolates was observed in liquid cultures. Mushroom yield was enhanced by two-fold in pots treated with microbial inoculum. While 30% and 50% increase in laccase and manganese peroxidase activities were observed in pots treated with bacterial inoculum than that of control pots, about 42.73% increment in nitrogen content of the dried mushroom samples was observed in treated pots. This could be possibly due to the presence of diazotrophs, cellulolytic and amylolytic isolates in the inoculum. Significant increase (p<0.05) in median values of primordia formed in pots treated with bacterial inoculum was also observed. The results of this study, therefore clearly indicate the importance of the application of such microbial inoculum as a bio-fertilizer in mushroom cultivation. Keywords: Pleurotus ostreatus, Laccase, Bio-fertilizer, Co-inoculation, Mycelial colonization

Biofertilizer effect of free living nitrogen fixing bacterium on the growth and water quality properties of Penaeid prawn culture system

The role of phytoplankton is significant in improving the yield of prawn culture. The growth of phytoplankton will be effected various factors viz. dissolved oxygen, production of natural food to shrimp larvae in control of ammonia. The growth of phytoplankton can be enhanced by application of nitrogeneous fertilizers to improve water quality and growth performance of Penaeus indicus. The present research work is focussed on a free-living nitrogen fixing bacterium Azotobacter chroococcum supplemented by nitrogeneous fertilizers to increase the content of proteins, carbohydrates and wet weight of P. indicus. Ten animals of juveniles of P. indicus were collected and grown in controlled conditions for fifteen days. The liquid suspension of free living nitrogen fixing bacteria A. chroococcum has been inoculated for experimental treatment purpose. The inoculum has been introduced to four treatments and for findings of water quality analysis andphytoplankton growth analysis by Strickland and Parsons method and biochemical analysis by Raymont et al., method. The research findings revealed that A. chroococcum supplemented with fertilizers will affect the growth of phytoplankton which is vital in prawn culture and enhance the growth of hormones and dry weight for improvement of the yield.

BIOREMEDIATION OF SOILS FROM OIL SPILL IMPACTED SITES USING BIOAUGMENTATION WITH BIOSURFACTANTS PRODUCING, NATIVE, FREE-LIVING NITROGEN FIXING BACTERIA

Oil spills are the most common source of environmental damage in areas with oil extraction activities. Bioaugmentation is a bioremediation technology that involves increasing microbiotas that can remove contaminant compounds from soils or water. This can be achieved by supplying exogenous microorganisms or by increasing the abundance of native microorganisms able to remove the contaminants. As biosurfactants enhance the solubility and bioavailability of hydrophobic compounds, the addition of native bacteria that produce biosurfactants may improve the rates of hydrocarbons biodegradation at oil spill impacted sites. The objective of this study was to investigate the effectiveness of using the bioaugmentation of native, free-living nitrogen fixing bacteria (NFB) that are able to remove hydrocarbons and produce biosurfactant compounds, to bioremediate soils at long term oil spill impacted sites containing total hydrocarbon (TH) levels of 120 000 ppm. For this study, two NFB that produce biosurfactants were selected from a group of 24 previously isolated from a long-term oil spill impacted site. After 16 months of bioaugmentation, the hydrocarbons removal reached 80 %. The NFB count increased from 13×10 4 colony-forming units (CFU) to 2×10 9 CFU, and the total bacterial population increased from 5×10 4 CFU to 22×10 11 CFU. According to literature, this is the first bioaugmentation study using native NFB biosurfactant producers to bioremediate soils from aged oil spill impacted sites. As such, these results demonstrate the potential of this bioaugmentation strategy for the bioremediation of oil spill impacted sites, especially those low in nitrogen.

Biofilm formation enables free-living nitrogen-fixing rhizobacteria to fix nitrogen under aerobic conditions.

The multicellular communities of microorganisms known as biofilms are of high significance in agricultural setting, yet it is largely unknown about the biofilm formed by nitrogen-fixing bacteria. Here we report the biofilm formation by Pseudomonas stutzeri A1501, a free-living rhizospheric bacterium, capable of fixing nitrogen under microaerobic and nitrogen-limiting conditions. P. stutzeri A1501 tended to form biofilm in minimal media, especially under nitrogen depletion condition. Under such growth condition, the biofilms formed at the air-liquid interface (termed as pellicles) and the colony biofilms on agar plates exhibited nitrogenase activity in air. The two kinds of biofilms both contained large ovoid shape 'cells' that were multiple living bacteria embedded in a sac of extracellular polymeric substances (EPSs). We proposed to name such large 'cells' as A1501 cyst. Our results suggest that the EPS, especially exopolysaccharides enabled the encased bacteria to fix nitrogen while grown under aerobic condition. The formation of A1501 cysts was reversible in response to the changes of carbon or nitrogen source status. A1501 cyst formation depended on nitrogen-limiting signaling and the presence of sufficient carbon sources, yet was independent of an active nitrogenase. The pellicles formed by Azospirillum brasilense, another free-living nitrogen-fixing rhizobacterium, which also exhibited nitrogenase activity and contained the large EPS-encapsuled A1501 cyst-like 'cells'. Our data imply that free-living nitrogen-fixing bacteria could convert the easy-used carbon sources to exopolysaccharides in order to enable nitrogen fixation in a natural aerobic environment.

Effects of Rhamnolipid and Microbial Inoculants on the Vermicomposting of Green Waste with Eisenia fetida.

The effects of adding the biosurfactant rhamnolipid, the lignolytic and cellulolytic fungus Phanerochete chrysosporium, and the free-living nitrogen-fixing bacterium Azotobacter chrococcum on vermicomposting of green waste with Eisenia fetida was investigated. The addition of rhamnolipid and/or either microorganism alone or in all combinations significantly increased E. fetida growth rate, the number of E. fetida juveniles and cocoons, the population densities of cellulolytic fungi and Azotobacter bacteria, and cellulase and urease activities in the vermicomposts. The quality of the final vermicompost (in terms of electrical conductivity, nutrient content, C/N ratio, humic acid content, lignin and cellulose contents, and phytotoxicity to germinating seeds) was enhanced by addition of rhamnolipid and/or microorganisms. The physical characteristics of vermicomposts produced with rhamnolipid and/or microorganisms were acceptable for agricultural application. The best quality vermicompost was obtained with the combined addition of P. chrysosporium, A. chrococcum, and rhamnolipid.

Isolation and characterization of free-living nitrogen fixing bacteria from alkaline soils

Free living nitrogen fixing bacteria were isolated from soil on Jensen agar plates and were characterized phylogenetically by 16S rDNA sequence analysis. All the isolates (VS1, VS2, VS3, VS4) were Gram –ve, rod shaped. Antibiotic test revealed VS2 to be resistant to ampicillin and VS4 was resistant to both ampicillin and kanamycin; otherwise all the isolates were sensitive to chloramphenicol. Nitrogen fixation was studied by the estimation of total nitrogen and available nitrogen fixed by cultures in the medium and compared with the control culture of Azotobacter CBD15 (Azo) procured from IARI. VS2 and VS3 fixed 12.02 ppm/ml and 10.635 ppm/ml as available nitrogen content and 14.44 ppm/ml and 18.73 ppm/ml as total nitrogen content. 16S rDNA studies revealed identification of the isolates- Pseudomonas sp. (VS2) and Paenibacillus sp. (VS3 and VS4). VS3 and VS4 showed 98% similarity with P. borealis. Soils from which these microbes isolated were also characterized to understand the environment of these microbes. The pH and chemical characterization (Organic Carbon, Phosphorus, Sulphur, heavy metal analysis of different metals e.g. Zn, Cu, Cr, Pb, Ni and water holding capacity) of the soils showed them to be slightly alkaline and clayey loamy. Cloning of VS2 was done successfully with plasmid pMMB277 isolated from E. coli 2842.The significance of this study lies in the isolation of those bacteria which are comparable in their nitrogen fixing potential to Azotobacter.

Plant Growth Promoting Rhizobacteria-an Efficient Tool for Agriculture Promotion

Current soil management strategies are mainly dependent on inorganic chemical-based fertilizers, which caused a serious threat to human health and environment. Plant growth-promoting rhizobacteria (PGPR) are naturally occurring soil bacteria that aggressively colonize plant roots and benefit plants by providing growth promotion. Inoculation of crop plants with certain strains of PGPR at an early stage of development improves biomass production through direct effects on root and shoots growth. The major groups of PGPR can be found along with the phyla actinobacteria, bacteroidetes, firmicutes, and proteobacteria. Inoculation of agricultural crops with PGPR may result in multiple effects on early-season plant growth, as seen in the enhancement of seedling germination, plant health, vigor, height, shoot weight, nutrient content of shoot tissues, early bloom, chlorophyll content, and increased nodulation in legumes. PGPRs are reported to influence the growth, yield, and nutrient uptake by an array of mechanisms. They help in increasing nitrogen fixation in legumes, help in promoting free-living nitrogen-fixing bacteria, increase supply of other nutrients, such as phosphorus, iron and produce plant hormones that enhance other beneficial bacteria or fungi. Now a day’s an increasing number of PGPR being commercialized for various crops. Subsequently, there has been much research interest in PGPRs. Several reviews have discussed specific aspects of growth promotion by PGPRs. Therefore, PGPRs can help to generate wealth cooperatively in local communities, reducing the need for more expensive manufactured products, such as nitrogenous fertilizers and use of PGPR in world has the potential to provide valuable insight.

Isolation and Characterization of Phosphate Solubilizing Diazotrophic Bacteria from Rhizosphere of Rice Plant of Indo-Nepal Border

Free-living nitrogen fixing bacteria were isolated from the rhizosphere of rice plant of different regions of Indo-Nepal border. 33.78% isolates among them showed mineral phosphate solubilization. On the basis of bacterial characterization and identification with Bergey’s manual of Determinative Bacteriology (Holt et al, 1994), 38% isolates showed maximum similarity with Pseudomonas and Azotobacter, 30% with members of Enterobacteriaceae, 4% with Gram’s positive Microbacterium sp. and 28% was categorized unknown. ECI 12A isolate showed maximum solubilization index (S.I.), 0.809 on solid medium whereas 188.65 mg P/mg dry weight phosphate solubilization in liquid medium. The highest phosphate solubilizing isolate ECI 12A was sequenced after amplification of partial 16S rRNA gene, blast with BLASTn 2.2.17 program of NCBI showed 99 % identity having 99 % query coverage with Microbacterium sp. This isolate has been named as Microbacterium sp. Strain ECI-12A.The sequence of this strain has been deposited in NCBI Gene Bank under accession number EU155122 and this strain has promising potential for developing as a plant growth promoting rhizobacteria (PGPR) as well as biofertilizer.Academic Voices Vol.5 2015: 79-87

Uso de Leersia hexandra (Poaceae) en la fitorremediación de suelos contaminados con petróleo fresco e intemperizado

The oil industry has generated chronic oil spills and their accumulation in wetlands of the state of Tabasco, in Southeastern Mexico. Waterlogging is a factor that limits the use of remediation technologies because of its high cost and low levels of oil degradation. However, Leersia hexandra is a grass that grows in these contaminated areas with weathered oil. The aim of the study was to evaluate the bacteria density, plant biomass production and phytoremediation of L. hexandra in contaminated soil. For this, two experiments in plastic tunnel were performed with fresh (E1) and weathered petroleum (E2) under waterlogging experimental conditions. The E1 was based on eight doses: 6 000, 10 000, 30 000, 60 000, 90 000, 120 000, 150 000 and 180 000 mg.kg-1 dry basis (d. b.) of total petroleum hydrocarbons fresh (TPH-F), and the E2, that evaluated five doses: 14 173, 28 400, 50 598, 75 492 and 112 142 mg. kg-1 d. b. of total petroleum hydrocarbons weathered (TPH-W); a control treatment with 2 607 mg.kg-1 d. b. was used. Each experiment, with eight replicates per treatment, evaluated after three and six months: a) microbial density of total free-living nitrogen-fixing bacteria (NFB) of Azospirillum (AZP) and Azotobacter group (AZT), for viable count in serial plate; b) dry matter production (DMP), quantified gravimetrically as dry weight of L. hexandra; and c) the decontamination percentage of hydrocarbons (PDH) by Soxhlet extraction. In soil with TPH-F, the NFB, AZP y AZT populations were stimulated five times more than the control both at the three and six months; however, concentrations of 150 000 and 180 000 mg.kg-1 d. b. inhibited the bacterial density between 70 and 89 %. Likewise, in soil with TPH-W, the FNB, AZP and AZT inhibitions were 90 %, with the exception of the 14 173 mg.kg-1 d. b. treatment, which stimulated the NFB and AZT in 2 and 0.10 times more than the control, respectively. The DMP was continued at the six months in the experiments, with values of 63 and 89 g in fresh and weathered petroleum, respectively; had no significant differences with the control (p≤0.05). The PDH reached values of 66 to 87 % both TPH-F and TPH-W at six months, respectively. These results demonstrated the ability the L. hexandra rhizosphere to stimulate the high NFB density, vegetal biomass production and phytoremediation of contaminated soils (with fresh and weathered petroleum), in a tropical waterlogging environment.

Nutrient Enrichment of Organic Manure Through Biotechnological Means

The residual slurry obtained after anaerobic digestion is termed as digestate which contain partially degraded organic material along with sludge and inorganic matter. Recycling of this digestate by applying back to the soil as an organic fertilizer could be the best option for its safe disposal. However, the residual slurry is usually characterized by high COD, BOD and low nutrient (N, P and K) content which makes it difficult for direct application as biomanure. In the present study, an attempt has been made to enrich the residual slurry obtained from codigestion through biotechnological means. Seven treatment sets were placed for nutrient enrichment studies which include 4 cyanobacteria, 2 free living nitrogen fixing bacteria and mixture of all. Further, the enriched bioslurry was applied to soil as biomanure at an application rate of 10 t/ha. Bhendi crop was cultivated in three different soils (biomanure, urea and control). After incubation period of 6 weeks it has been observed that there was an enhancement of N, P, K content by 2.76, 3.7 and 2.2 fold higher as compared to control with maximum NPK ratio of 25:1:3.5. Aulosira, Azosprillum and Azotobacter were found to enrich the nutrients more efficiently as compared to other strains. The soil supplemented with biomanure was found to have superior quality than the control and almost comparable to that chemical (urea) amended soil. The plant growth from biomanure treated plot was at par with the urea supplemented soil which advocates that synthetic fertilizer can be potentially replaced with this enriched residual slurry.

Rice (Oryza sativa L.) nutrient management using mycorrhizal fungi and endophytic Herbaspirillum seropedicae

Integrated nutrient management with biological and chemical fertilizers can improve rice (Oryza sativa L.) productivity, bio-fortification, soil health and fertility. Accordingly, this study was planned to evaluate the combined effects of biological fertilizers including arbuscular mycorrhizal (AM) fungi (Glomus mosseae) and free-living nitrogen-fixing bacteria (Herbaspirillum seropedicae), as well as chemical fertilizers on the yield and nutrient contents of wetland rice under field conditions. Seedlings were inoculated with AM fungi and the bacteria in the nursery and were then transplanted to the field. The experiment was carried out as a split factorial design with three replicates. Treatments included three rates of nitrogen (N1, N2 and N3) and phosphorous (P1, P2 and P3) fertilizers (100, 75 and 50% of the optimum level) in the main plots and mycorrhizal and bacterial treatments in the sub plots. The total of urea (g) used per plot was equal to N1=200, N2=150 and N3=100 at three different growth stages (seeding, tillering and heading) and the total of P (g) per plot used once at seeding using triple super phosphate including P1=16, P2=13 and P3=10. Plant growth and yield as well as the concentration of nitrogen (N), phosphorous (P), potassium (K), iron (Fe), and zinc (Zn) were measured in the soil, straw and grains. N-fertilizer and biological fertilizers had significant effects on root, shoot and grain yield of rice, however, P-fertilizer just significantly affected root and shoot dry weights. Interestingly, analyses of variance indicated that biological fertilization significantly affected all the experimental treatments except straw N. AM fungi, N1 and P1 resulted in the highest rate of rice growth and yield. The interactions of chemical and biological fertilization resulted in significant effects on grain Zn, Fe, P, and N as well as soil Fe, K and N. The highest rate of grain nutrient uptake was resulted by the combined use of biological fertilization and the medium level of chemical fertilization. Interestingly, with decreasing the rate of chemical N fertilization, rice nutrient use efficiency increased indicating how biological fertilization can be efficient in providing plants with its essential nutrients such as N. However, the highest rate of soil and straw nutrient concentration was related to the combined use of biological fertilization and the highest rate of chemical fertilization. We conclude that biological fertilizer, (mycorrhizal fungi and H. seropedicae) can significantly improve wetland rice growth and yield (resulting in the decreased rate of chemical fertilizer), especially if combined with appropriate rate of chemical fertilization, by enhancing nutrient uptake (fortification) and root growth.