N2-fixing Bacteria Research Articles

Un-culturable Microbial Community Analysis of Paddy Ecosystem at Different Depths using Nif H Gene DGGE Analysis Approach

Background: Biological N2 fixation is gaining importance in rice ecosystem, because of current concern on the environmental and soil health that are caused by the continuous use of nitrogenous fertilizers and the need for improved sustainable rice productivity. BNF is the conversion of atmospheric nitrogen by certain microorganism’s viz., N2 fixing microorganisms or diazotrophs into forms that plants can use. A substantial molecular diversity of N2 fixing bacteria has been detected in the rice field soil based on retrieval of nif H gene fragment from DNA by molecular approach, i.e., DGGE (Denaturant Gradient Gel Electrophoresis). The present investigation aimed to assess the diversity of N2 fixing bacteria by sequencing and phylogenetic analysis of nif H gene of the metagenomic DNA collected from different depth of flooded rice soil. Methods: In this laboratory investigation, soil samples were collected from the paddy field at different depths viz., rhizosphere region, 5 cm, 10 cm, 15 cm depth using the core sampler during active tillering stage. In the laboratory, the collected soil samples were analyzed by molecular methods involving the use of primer sets based on nif H sequences (nitrogenase reductase). Result: Our investigations in the genetic diversity of diazotrophs associated with rice field soil using nif H revealed the existence of seven discrete bands. The sequencing results showed all sequences were classified as uncultured bacterium clone nitrogenase iron protein (nif H) gene, partial cds. The present works based on the applications of molecular methods have facilitated the study of individual ecosystem and their specific activity.

Abundance and community succession of nitrogen-fixing bacteria in ferrihydrite enriched cultures of paddy soils is closely related to Fe(III)-reduction

In flooded paddy soils, some metal reducers are also capable of nitrogen (N) fixation, which is essential in ensuring a reliable N-supply for rice growth. Microbial iron [Fe(III)] reduction is an important biogeochemical process that can be stimulated by ferrihydrite amendment to paddy soil. Therefore, this study aimed to investigate the abundance and succession of the N2-fixing bacterial community in ferrihydrite enriched paddy soils collected from Hunan (HN) and Sichuan (SC) provinces, China. The relationship between the N2-fixing bacterial community and Fe(III) reduction was also assessed. When compared with the control treatment, ferrihydrite enrichment significantly enhanced nitrogenase (nifH) gene abundance by 8.05 × 105 to 4.45 × 106 copies g−1 soil during the 40-day flooding of HN soil, while nifH gene abundance in SC soil was remarkably increased by 5.90 × 107 to 9.56 × 107 copies g−1 soil during day 1 to 5 in response to ferrihydrite amendment. The relative abundance of N2-fixing bacteria peaked on day 5 (21.5% in HN soil and 5.4% in SC soil) and gradually decreased to a stable abundance after day 20. Remarkable increases in relative abundance of N2-fixing bacteria during the first 10 days of flooding were detected in both soils with ferrihydrite enrichment, whereas little difference was found after day 10 of flooding. During the early stage of flooding, the Shannon and Simpson indexes of N2-fixing bacteria with ferrihydrite enrichment were significantly decreased, and the community structure changed greatly. Most N2-fixing bacteria in ferrihydrite enriched paddy soils were phylogenetically related to the order Clostridiales, with some of those potentially capable of Fe(III) reduction. The community succession of N2-fixing bacteria closely correlated with Fe(III) reduction. Thus, improving N2-fixation via stimulation of Fe(III) reduction might aid in the reduction of N-fertilizer application to paddy field.

Enrichment of Nitrogen-Fixing Bacteria in a Nitrogen-Deficient Wastewater Treatment System.

Anthropogenic nitrogen fixation is essential to sustain a global population of 7.7 billion. However, there has been a long-standing desire to find cheaper and more environmentally friendly alternatives to the Haber-Bosch process. In this study, we developed a new strategy of nitrogen fixation by enriching free-living N2-fixing bacteria (NFB) in reactors fed with low nitrogen wastewater, analogous to those usually found in certain industrial effluents such as paper mills. Our reactors fixed appreciable quantities of nitrogen with a rate of 11.8 mg N L-1 day-1. This rate is comparable to recent "breakthrough" nitrogen-fixing technologies and far higher than observed in low C/N reactors (fed with organic matter and nitrogen). NFB were quantified using quantitative polymerase chain reaction (qPCR) of the nifH (marker gene used to identify biological nitrogen fixation) and 16S rRNA genes. The nifH gene was enriched by a factor of 10 in the nitrogen-fixing reactors (compared to controls) attaining 13% of the bacterial population (1:4.2 copies of nifH to 16S rRNA). The Illumina MiSeq 16S rRNA gene amplicon sequencing of reactors showed that the microbial community was dominated (19%) by Clostridium pasteurianum. We envisage that nitrogen-enriched biomass could potentially be used as a biofertilizer and that the treated wastewater could be released to the environment with very little post-treatment.

Biological nitrogen fixation in field-grown sorghum under different edaphoclimatic conditions is confirmed by N isotopic signatures

The association between sorghum and N2-fixing bacteria has been assessed only under limited conditions. We investigated biological nitrogen fixation (BNF) in situ in fifteen sorghum genotypes with dry or succulent culm types under five edaphoclimatic conditions. One randomized block experiment was established in each of five locations, from the humid to the semiarid regions of Pernambuco state, Brazil. BNF was estimated using the 15N natural abundance method by comparing the average δ15N value of each sorghum genotype with those of the reference species. High levels of productivity, up to 22 Mg shoot biomass ha−1 in the 3-month cycle, were obtained where rainfall was high, and up to 5 Mg ha−1 was obtained under low rainfall. The nitrogen contents showed a similar pattern as biomass production, and the genotypes with the highest productivity accumulated from 200 to 300 kg N ha−1. BNF ranged from 55 to 78% of plant N in one location and from 36 to 56% in another location, but BNF did not occur in the other three locations. Although the factors that blocked effective symbiosis were not determined, symbiosis was not influenced by P or K availability. The proportion of N2 fixation was similar in the grain-producing, dry culm genotypes and in the sugar-rich, succulent culm genotypes. The sorghum genotypes fixed N2, reaching up to 218 kg ha−1 N, without inoculation with diazotrophs. Therefore, sorghum has a high potential to fix atmospheric N2, but the factors that block N2 fixation must be identified for crop management planning.

The Non-Legume Parasponia andersonii Mediates the Fitness of Nitrogen-Fixing Rhizobial Symbionts Under High Nitrogen Conditions.

Organisms rely on symbiotic associations for metabolism, protection, and energy. However, these intimate partnerships can be vulnerable to exploitation. What prevents microbial mutualists from parasitizing their hosts? In legumes, there is evidence that hosts have evolved sophisticated mechanisms to manage their symbiotic rhizobia, but the generality and evolutionary origins of these control mechanisms are under debate. Here, we focused on the symbiosis between Parasponia hosts and N2-fixing rhizobium bacteria. Parasponia is the only non-legume lineage to have evolved a rhizobial symbiosis and thus provides an evolutionary replicate to test how rhizobial exploitation is controlled. A key question is whether Parasponia hosts can prevent colonization of rhizobia under high nitrogen conditions, when the contribution of the symbiont becomes nonessential. We grew Parasponia andersonii inoculated with Bradyrhizobium elkanii under four ammonium nitrate concentrations in a controlled growth chamber. We measured shoot and root dry weight, nodule number, nodule fresh weight, nodule volume. To quantify viable rhizobial populations in planta, we crushed nodules and determined colony forming units (CFU), as a rhizobia fitness proxy. We show that, like legumes and actinorhizal plants, P. andersonii is able to control nodule symbiosis in response to exogenous nitrogen. While the relative host growth benefits of inoculation decreased with nitrogen fertilization, our highest ammonium nitrate concentration (3.75 mM) was sufficient to prevent nodule formation on inoculated roots. Rhizobial populations were highest in nitrogen free medium. While we do not yet know the mechanism, our results suggest that control mechanisms over rhizobia are not exclusive to the legume clade.

Isotopic constraints on plant nitrogen acquisition strategies during ecosystem retrogression.

Plant root associations with microbes such as mycorrhizal fungi or N-fixing bacteria enable ecosystems to tap pools of nitrogen (N) that might otherwise be inaccessible, including atmospheric N or N in large soil organic molecules. Such microbially assisted N-foraging strategies may be particularly important in late-successional retrogressive ecosystems where productivity is low and soil nutrients are scarce. Here, we use natural N-stable isotopic composition to constrain pathways of N supplies to different plant functional groups across a well-studied natural soil fertility gradient that includes a highly retrogressive stage. We demonstrate that ectomycorrhizal fungi, ericoid mycorrhizal fungi, and N-fixing bacteria support forest N supplies at all stages of ecosystem succession, from relatively young, N-rich/phosphorus (P)-rich sites, to ancient sites (ca. 500 ky) where both N supplies and P supplies are exceedingly low. Microbially mediated N sources are most important in older ecosystems with very low soil nutrient availability, accounting for 75-96% of foliar N at the oldest, least fertile sites. These isotopically ground findings point to the key role of plant-microbe associations in shaping ecosystem processes and functioning, particularly in retrogressive-phase forest ecosystems.

Particle size and concentration dependent toxicity of copper oxide nanoparticles (CuONPs) on seed yield and antioxidant defense system in soil grown soybean (Glycinemax cv. Kowsar).

Increasing applications of engineered nanomaterials (ENMs) warrant lifecycle assessment of their potential toxicity. Herein, we investigated potential phytotoxicity of copper oxide nanoparticles (CuONPs) on seed yield, focusing on particle size- and concentration-dependent responses of multiple antioxidant defense biomarkers, in soil-grown Glycinemax (cv. Kowsar) during its lifecycle. To this end, we synthesized three distinct sizes CuONPs (25, 50 and 250nm): all with high purity, monoclinic crystal structure, and same surface charge. Each pot received two seeds, placed in soil inoculated with N-fixing bacteria (Rhizobium japonicum) and grown outdoor for 120days. Our results show lipid peroxidation (MDA) and several antioxidant biomarkers (SOD, CAT, POX, APX) were differentially altered by the copper compound type, concentrations, and their interactions (p<0.01). We show particle size- and concentration-dependent influence of CuONPs on lipid peroxidation, and such antioxidant biomarkers including SOD, CAT, POX, and APX, in soybean leaf at 120-day post-plantation. Particularly, the effects of CuONP-25 were consistently higher for most antioxidant biomarkers tested compared to the two larger size CuONPs (CuONP-50, CuONP-250) or Cu2+ ions treatments. We show that the concentration-response curves for CuONP-25 and Cu2+ ions were linear (R2>0.65), unlike for the larger size CuONPs (CuONP-50, CuONP-250) the relationships were nonlinear (R2<0.45), for most antioxidant biomarkers. The concentration-response curves for seed yield for all types of Cu compounds were linear (R2>0.65). Soybean seed yield also mirrored particle size- and concentration-dependent inhibition with CuONPs, and inhibition of CuONP-25 was significantly higher than the two larger size CuONPs or Cu2+ ions at all concentrations tested. All in all, our findings indicate differential nano-specific toxicity compared to ionic Cu2+ toxicity in soybean. These results may guide researchers and regulators on how best to tailor ENMs with specific particle characteristics rendering them more or less toxic, and better inform risk assessment of CuONPs in soil grown food crops such as soybean.

Interaction with Soil Bacteria Affects the Growth and Amino Acid Content of Piriformospora indica.

Exploration of the effect of soil bacteria on growth and metabolism of beneficial root endophytic fungi is relevant to promote favorable associations between microorganisms of the plant rhizosphere. Hence, the interaction between the plant-growth-promoting fungus Piriformospora indica and different soil bacteria was investigated. The parameters studied were fungal growth and its amino acid composition during the interaction. Fungus and bacteria were confronted in dual cultures in Petri dishes, either through agar or separated by a Perspex wall that only allowed the bacterial volatiles to be effective. Fungal growth was stimulated by Azotobacter chroococcum, whereas Streptomyces anulatus AcH 1003 inhibited it and Streptomyces sp. Nov AcH 505 had no effect. To analyze amino acid concentration data, targeted metabolomics was implemented under supervised analysis according to fungal-bacteria interaction and time. Orthogonal partial least squares-discriminant analysis (OPLS-DA) model clearly discriminated P. indica–A. chroococcum and P. indica–S. anulatus interactions, according to the respective score plot in comparison to the control. The most observable responses were in the glutamine and alanine size groups: While Streptomyces AcH 1003 increased the amount of glutamine, A. chroococcum decreased it. The fungal growth and the increase of alanine content might be associated with the assimilation of nitrogen in the presence of glucose as a carbon source. The N-fixing bacterium A. chroococcum should stimulate fungal amino acid metabolism via glutamine synthetase-glutamate synthase (GS-GOGAT). The data pointed to a stimulated glycolytic activity in the fungus observed by the accumulation of alanine, possibly via alanine aminotransferase. The responses toward the growth-inhibiting Streptomyces AcH 1003 suggest an (oxidative) stress response of the fungus.

Prospection of Plant Growth-promoting Rhizobacterium Enterobacter cloacae and N2-fixing Rhizobium leguminosarum bv. viciae Effect on Faba Bean Evolution in Sustainable Agriculture

Biofertilizers are relying on ecofriendly approaches as sustainability and environmental safety of agricultural for crop production. The multiplicity of beneficial effects of microbial inoculants, particularly plant growth promoters (PGP) and N2-fixing bacteria strengthened to use biofertilizers in modern agriculture. Prospection study of Enterobacter cloacae KX034162 as PGPR was carried out in this study. Based on in vivo assays for PGPRs characterization; Exopolysaccharides production (EPS), biofilm formation, Phosphate, Zinc carbonate and zinc oxide solubilization and productivity of indole acetic acid (IAA) E. cloacae KX034162 was used. The synergistic inoculation effect of two microbial on faba bean (Vicia faba L.) growth parameters and productivity were evaluated in two successful winter seasons. The results showed that the dual inoculation of both microbes have a significant positive effect on the estimated parameters compared with the control. E. cloacae KX034162 produced IAA (47.0 mgl-1), polysaccharides (6.4 gl-1) and solubilized Zn. The dual inoculation enhanced nutrients absorption, nodulation, growth parameters, N% (it was 1.42 and 1.44 in the first and second season compared with 0.29 and 0.30 as un-inoculated treatments, respectively) and at the end faba bean production increased.

Rhizobium Inoculation Drives the Shifting of Rhizosphere Fungal Community in a Host Genotype Dependent Manner.

Rhizosphere microorganisms play important roles in plant health and nutrition, and interactions among plants and microorganisms are important for establishment of root microbiomes. As yet, plant-microbe and microbe-microbe interactions in the rhizosphere remain largely mysterious. In this study, rhizosphere fungal community structure was first studied in a field experiment with two soybean cultivars contrasting in nodulation grown in two rhizobium inoculation treatments. Following this, recombinant inbred lines (RILs) contrasting in markers across three QTLs for biological nitrogen fixation (BNF) were evaluated for effects of genotype and rhizobium inoculation to the rhizosphere fungal community as assessed using ITS1 amplicon sequencing. The soybean plants tested herein not only hosted rhizosphere fungal communities that were distinct from bulk soils, but also specifically recruited and enriched Cladosporium from bulk soils. The resulting rhizosphere fungal communities varied among soybean genotypes, as well as, between rhizobium inoculation treatments. Besides, Cladosporium were mostly enriched in the rhizospheres of soybean genotypes carrying two or three favorable BNF QTLs, suggesting a close association between soybean traits associated with nodulation and those affecting the rhizosphere fungal community. This inference was bolstered by the observation that introduction of exogenous rhizobia significantly altered rhizosphere fungal communities to the point that these communities could be distinguished based on the combination of soybean genotype and whether exogenous rhizobia was applied. Interestingly, grouping of host plants by BNF QTLs also distinguished fungal community responses to rhizobium inoculation. Taken together, these results reveal that complex cross-kingdom interactions exist among host plants, symbiotic N2 fixing bacteria and fungal communities in the soybean rhizosphere.

Endophytic Microbes from Diverse Wheat Genotypes and Their Potential Biotechnological Applications in Plant Growth Promotion and Nutrient Uptake

Endophytic microbes residing inside the tissues of plants play a significant role to enhance the growth and health of plants by different plant growth-promoting mechanisms. In the present investigation, N2-fixing endophytic bacteria were isolated and characterized by plant growth. A total of one hundred fifty-nine endophytic bacteria were isolated from surface-sterilized roots and stem of different genotypes of wheat growing in the Divine Valley of Baru Sahib, Himachal Pradesh. The isolated bacterial endophytes were screened in vitro for plant growth-promoting attributes. Out of one hundred fifty-nine, thirteen endophytic bacteria were selected based on multifarious plant growth-promoting attributes. Among plant growth-promoting activities, hydrogen cyanide producers (19%) were higher when compared to siderophores producers (16%) and P-solubilizers (16%), ammonia producers (14%), K-solubilizers (14%), IAA producers (12%), Zn-solubilizers (5%), N2-fixers (2%) and biocontrol (2%). One of the isolates EU-B2RT.R1 demonstrated that a significant level of nitrogenase activity, P-solubilization and IAA production was identified as Acinetobacter guillouiae EU-B2RT.R1 based on 16S rRNA gene sequencing and BLAST analysis. Acinetobacter guillouiae EU-B2RT.R1, exhibiting multifarious beneficial traits, is further evaluated for plant growth promotion of wheat cultivar PBW 343+Lr24+GPC in pot experiment under greenhouse conditions. The Acinetobacter guillouiae EU-B2RT.R1 with multifarious plant growth-promoting activity has emerged as one of the efficient biofertilizers that need to be explored for sustainable agriculture.

Nodule Inception Is Not Required for Arbuscular Mycorrhizal Colonization of Medicago truncatula.

Most legumes can engage in symbiosis with N-fixing bacteria called rhizobia. This symbiosis, called nodulation, evolved from the more widespread symbiosis that most land plants form with arbuscular mycorrhiza, which is reflected in a common requirement of certain genes for both these symbioses. One key nodulation gene, Nodule Inception (NIN), has been intensively studied. Mutants in NIN are unable to form nodules, which has made it difficult to identify downstream genes under the control of NIN. The analysis of data from our recent transcriptomics study revealed that some genes with an altered expression of nin during nodulation are upregulated in mycorrhizal roots. In addition, another study reported the decreased colonization of nin roots by arbuscular mycorrhiza. We therefore investigated a role for NIN in mycorrhiza formation. Our time course study, using two nin alleles with differing genetic backgrounds, suggests that that loss of NIN does not affect colonization of Medicago truncatula roots, either in the presence or absence of rhizobia. This, and recent phylogenetic analyses showing that the loss of NIN is correlated with loss of nodulation in the FaFaCuRo clade, but not with the ability to form mycorrhiza, argue against NIN being required for arbuscular mycorrhization in legumes.

Occurrence of heavy metal resistance in Sinorhizobium sp. isolated from root nodules of fenugreek, treated with tannery effluent

The aim of present study was to initiate preliminary work on heavy metal contamination in farm soil (contaminated with processed tannery effluent) and its potential influence on the development of metal resistance among N2fixing bacterium, Sinorhizobium sp. Contaminated plant samples were analyzed for various metals and minimum inhibitory concentration (MIC) of metals was determined. Fenugreek (Trigonella sp.) plants revealed accumulation of these metals in root and leaves. Sinorhizobium sp. were isolated (25) from the root nodules of fenugreek-treated with processed tannery wastewater and characterized morpho-biochemically. All isolates were evaluated for their resistance against Cr3+, Cr6+, Cd2+, Cu2+, Zn2+ and Ni2+. The maximum MIC of 1600 μg mL−1 was noticed against Cr3+ in 64% isolates. Among all the isolates, the lowest MIC of 25 μg mL−1 was detected against Ni2+. Some metal resistant isolates were evaluated for their resistance against frequently used antibiotics viz., tetracycline, ampicillin, gentamycin, kanamycin, chloramphenicol and nalidixic acid. About 46% Sinorhizobium isolates were resistant to nalidixic acid whereas 26.6% showed resistance to ampicillin and kanamycin. The isolates demonstrated confluent growth upto a salt concentration of 3%; whereas isolates (SM17, SM24) tolerated 10% NaCl. Acidic pH eliminated almost all the test population and neutral pH had no suppressive effect on growth while majority were tolerant to pH 9.

STRAINS OF Paraburkholderia ORIGINATED FROM RUPESTRIAN FIELDS PROMOTE THE GROWTH OF Mimosa foliolosa

ABSTRACT Mimosa foliolosa is a promising native species of rupestrian fields for revegetation of degraded areas in this ecosystem. The symbiosis between leguminous plants and N2-fixing bacteria may play an important role in the recovery of these areas, since these plants have better development and are more resistant to the attack of pathogens. In addition to the biological nitrogen fixation (BNF), these bacteria can promote plant growth through other processes, such as phosphate solubilization and siderophore production. We studied the cultural and genetic characteristics of 11 bacterial strains, isolated from rupestrian field soils using Mimosa tenuiflora as “trap” plant. We evaluated these strains considering their symbiotic characteristics such as nodulation, and growth-promotion of Mimosa foliolosa, as well as their ability for siderophore production and phosphate solubilization. Native N2-fixing bacterial strains belonging to the Paraburkholderia genus (UFLA01-750, UFLA01-728, UFLA01-725, and UFLA01-757), showed high symbiotic efficiency with M. foliolosa. These strains also solubilized calcium phosphate and produced siderophores, exhibiting high functional diversity and potential for use in revegetation projects.

Divulging diazotrophic bacterial community structure in Kuwait desert ecosystems and their N2-fixation potential.

Kuwait is a semi-arid region with soils that are relatively nitrogen-poor. Thus, biological nitrogen fixation is an important natural process in which N2-fixing bacteria (diazotrophs) convert atmospheric nitrogen into plant-usable forms such as ammonium and nitrate. Currently, there is limited information on free-living and root-associated nitrogen-fixing bacteria and their potential to fix nitrogen and aid natural plant communities in the Kuwait desert. In this study, free living N2-fixing diazotrophs were enriched and isolated from the rhizosphere soil associated with three native keystone plant species; Rhanterium epapposum, Farsetia aegyptia, and Haloxylon salicornicum. Root-associated bacteria were isolated from the root nodules of Vachellia pachyceras. The result showed that the strains were clustered in five groups represented by class: γ-proteobacteria, and α-proteobacteria; phyla: Actinobacteria being the most dominant, followed by phyla: Firmicutes, and class: β-proteobacteria. This study initially identified 50 nitrogen-fixers by16S rRNA gene sequencing, of which 78% were confirmed to be nitrogen-fixers using the acetylene reduction assay. Among the nitrogen fixers identified, the genus Rhizobium was predominant in the rhizosphere soil of R. epapposum and H. salicornicum, whereas Pseudomonas was predominant in the rhizosphere soil of F. aegyptia, The species Agrobacterium tumefaciens was mainly found to be dominant among the root nodules of V. pachyceras and followed by Cellulomonas, Bacillus, and Pseudomonas genera as root-associated bacteria. The variety of diazotrophs revealed in this study, signifying the enormous importance of free-living and root-associated bacteria in extreme conditions and suggesting potential ecological importance of diazotrophs in arid ecosystem. To our knowledge, this study is the first to use culture-based isolation, molecular identification, and evaluation of N2-fixing ability to detail diazotroph diversity in Kuwaiti desert soils.

Larger root nodules increased Fe, Mo, Mg, P, Ca, Mn, K in the roots and higher yield in chickpea grown from nano FeS2 pre-treated seeds: emulating nitrogenase

N2-fixing bacteria symbiotically dwell inside the root nodules of legume and converts atmospheric N2 to NH3. Unlike high temperature–pressure Haber’s process, this conversion in the nodule is orchestrated by nitrogenase: an enzyme with Fe-S, Fe-Mo, Fe-V cofactor, at its heart. Strategies to increase the nodule population could reduce nitrogen fertilizer use. Here we discovered that pre-treating the chickpea seeds with nano FeS2 (iron pyrite) resulted in a denser root network with larger root nodules. It improved the shoot system and resulted in a higher yield. We observed a higher concentration of Fe, Mo, Mg, P, Ca, Mn, K in the roots of chickpea: possibly emulating nitrogenase.

Root microbiomes as indicators of seagrass health.

The development of early warning indicators that identify ecosystem stress is a priority for improving ecosystem management. As microbial communities respond rapidly to environmental disturbance, monitoring their composition could prove one such early indicator of environmental stress. We combined 16S rRNA gene sequencing of the seagrass root microbiome of Halophila ovalis with seagrass health metrics (biomass, productivity and Fsulphide) to develop microbial indicators for seagrass condition across the Swan-Canning Estuary and the Leschenault Estuary (south-west Western Australia); the former had experienced an unseasonal rainfall event leading to declines in seagrass health. Microbial indicators detected sites of potential stress that other seagrass health metrics failed to detect. Genera that were more abundant in 'healthy' seagrasses included putative methylotrophic bacteria (e.g. Methylotenera and Methylophaga), iron cycling bacteria (e.g. Deferrisoma and Geothermobacter) and N2 fixing bacteria (e.g. Rhizobium). Conversely, genera that were more abundant in 'stressed' seagrasses were dominated by putative sulphur-cycling bacteria, both sulphide-oxidising (e.g. Candidatus Thiodiazotropha and Candidatus Electrothrix) and sulphate-reducing (e.g. SEEP-SRB1, Desulfomonile and Desulfonema). The sensitivity of the microbial indicators developed here highlights their potential to be further developed for use in adaptive seagrass management, and emphasises their capacity to be effective early warning indicators of stress.

In vitro test for compatibility of biofertilizers containing phosphate solubilizers and nitrogen-fixing bacteria

The quality of biofertilizers is determined by the content of isolates and synergism between isolates. The experiment was aimed to investigate the compatibillty of biofertilizers containing P-solubilizers and N-fixing bacteria. In vitro test using isolates of PSB and N-fixing bacteria was done to study the synergystic character by using streak method on nutrient agar for the bacteria and potato dextrose agar for the fungi. Species of P-solubilizers were Pseudomonas mallei, P. cepaceae, Aspergillus niger, and Penicillium sp., and species of N-fixing bacteria were Azotobacter chroococum and Azospirillum sp. The results revealed that the P-soluilizers microbes and N-fixing bacteria were synergistic and did not antagonistic to each other. Biofertilizers which contain P-solubilizers and N-fixing bacteria can be used for agricultural purposes.

STUDIES ON PEANUT BRADYRHIZOBIUM BROTH INOCULA AS AFFECTED BY IRRIGATION WATER QUALITY IN NEWLY RECLAIMED DESERT SOILS.

Peanut being a leguminous crop is capable of fixing atmospheric nitrogen. The present investigation had been carried out in order to study and evaluate the efficiency of both liquid and solid inoculum of Bradyrhizobium ssp (N2-fixing bacteria) alone or with plant growth promoting rhizobacteria (PGPR); Azotobacter chroococcum DSM 2286, as co-inoculation to reduce the negative impact of salinity of irrigation water (Nile Water (563 mg L-1) (W1), Groundwater artesian well, about 1000 mgL-1 (W2) and Groundwater artesian well about 2000 mgL-1 (W3), were collected from cultivated regions of Sadat city.) on peanut plants grown in cultivated sandy soils. Nodulation, fresh and dry weights of the growing plants and their elemental composition, mainly contents of nitrogen, phosphorus, potassium, manganese, zinc and Iron, yield, yield components as well as quality traits of the seed and oil of two peanut cultivars grown in sandy soils were determined. The biofertilizers inoculation treatments as follows: 1-Control treatments (without any biofertilizers inoculation). 2- Inoculation by solid Bradyrhizobium alone. 3-Inoculation by liquid Bradyrhizobium alone. 4- Co-inoculation: by solid Bradyrhizobium + PGPR (Azotobacter chrococoum), and 5- Co-inoculation: by liquid Bradyrhizobium fertilization + PGPR (Azotobacter chrococoum). It could be concluded that inoculation by liquid Bradyrhizobium + Azotobacter chroococcum seems to be the recommended treatment for producing optimum seed and straw yield and other parameters studied. Likewise it achieved the least production costs, also reducing the harmful effects of salinity of soil and/or irrigation water. These in turn reflect an increase in dry matter yield and after that pod, seed and straw yield. However, this approach could be explored as an effective strategy with least production costs to improve salt tolerance index in peanut plants under drip irrigation system in newly reclaimed soils in Egypt.

Potential of Boehmia nivea as phytoremediator for petroleum-contaminated soil following nitrogen-fixing bacteria inoculation

Phytoremediation is one of an alternative technology to overcome a petroleum-contaminated soil. Boehmia nivea or Ramie has the potential to be used as a phytoremediatoragent to degrade petroleum hydrocarbons. The effort to improve the performance of Ramie in degrading petroleum hydrocarbons is through the concept of phytostimulant inoculation in biological fertilizer. The objective of pot experiment was to investigate the performance of Ramie as a phytoremediator of petrolleum waste. The research was designed in a randomized block design and consisted of Ramie treatment without inoulation, Ramie inoculated with 2% Azotobacter vinelandii, 2% Azospirillum sp., and 2% consortia of both inoculants. Experimental result showed that Ramie plants removed petroleum hydrocarbons up to 83.22% after 13 week of inoculation. The efficacy of hydrocarbon degradation increased significantly if Ramie plants was inoculated by N-fixing bacteria, but the biodegradation efficiency in biofertilizer treatments was not significantly different.