Endophytic nitrogen fixation – a possible ‘hidden’ source of nitrogen for lodgepole pine trees growing at unreclaimed gravel mining sites

2019 ◽  
Vol 95 (11) ◽  
Author(s):  
Kiran Preet Padda ◽  
Akshit Puri ◽  
Chris Chanway
Keyword(s):  
Nitrogen Fixation ◽  
Lodgepole Pine ◽  
Nifh Gene ◽  
Control Treatment ◽  
Nitrogen Fixing ◽  
Nitrogen Fixing Bacteria ◽  
Mining Sites ◽  
Greenhouse Study ◽  
Gravel Mining ◽  
Pine Trees

ABSTRACT Lodgepole pine (Pinus contorta var. latifolia) trees have been thriving on unreclaimed gravel mining sites in British Columbia, Canada, with tissue nitrogen-content and growth-rate unaffected by extremely low soil nitrogen-levels. This indicates that pine trees could be accessing a hidden nitrogen source to fulfill their nitrogen requirements – possibly via endophytic nitrogen-fixation. Endophytic bacteria originally isolated from native pine trees growing at gravel sites were selected (n = 14) for in vitro nitrogen-fixation assays and a year long greenhouse study to test the overall hypothesis that naturally occurring endophytic nitrogen-fixing bacteria sustain pine tree growth under nitrogen-limited conditions. Each of the 14 bacteria colonized the internal tissues of pine trees in the greenhouse study and fixed significant amounts of nitrogen from atmosphere (23%–53%) after one year as estimated through 15N isotope dilution assay. Bacterial inoculation also significantly enhanced the length (31%–64%) and biomass (100%–311%) of pine seedlings as compared to the non-inoculated control treatment. In addition, presence of the nifH gene was confirmed in all 14 bacteria. Our results support the possibility that pine trees associate with nitrogen-fixing bacteria, capable of endophytic colonization, to survive at unreclaimed gravel mining pits and this association could potentially be utilized for effective reclamation of highly disturbed sites in a sustainable manner.

scholarly journals Genomic characterization and computational phenotyping of nitrogen-fixing bacteria isolated from Colombian sugarcane fields

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Luz K. Medina-Cordoba ◽  
Aroon T. Chande ◽  
Lavanya Rishishwar ◽  
Leonard W. Mayer ◽  
Lina C. Valderrama-Aguirre ◽  
...  
Keyword(s):  
Antibiotic Resistance ◽  
Nitrogen Fixation ◽  
Plant Growth ◽  
Phosphate Solubilization ◽  
Nitrogen Fixing ◽  
Nitrogen Fixing Bacteria ◽  
Promising Tool ◽  
A Genome ◽  
Plant Growth Promoting ◽  
Growth Promoting

AbstractPrevious studies have shown the sugarcane microbiome harbors diverse plant growth promoting microorganisms, including nitrogen-fixing bacteria (diazotrophs), which can serve as biofertilizers. The genomes of 22 diazotrophs from Colombian sugarcane fields were sequenced to investigate potential biofertilizers. A genome-enabled computational phenotyping approach was developed to prioritize sugarcane associated diazotrophs according to their potential as biofertilizers. This method selects isolates that have potential for nitrogen fixation and other plant growth promoting (PGP) phenotypes while showing low risk for virulence and antibiotic resistance. Intact nitrogenase (nif) genes and operons were found in 18 of the isolates. Isolates also encode phosphate solubilization and siderophore production operons, and other PGP genes. The majority of sugarcane isolates showed uniformly low predicted virulence and antibiotic resistance compared to clinical isolates. Six strains with the highest overall genotype scores were experimentally evaluated for nitrogen fixation, phosphate solubilization, and the production of siderophores, gibberellic acid, and indole acetic acid. Results from the biochemical assays were consistent and validated computational phenotype predictions. A genotypic and phenotypic threshold was observed that separated strains by their potential for PGP versus predicted pathogenicity. Our results indicate that computational phenotyping is a promising tool for the assessment of bacteria detected in agricultural ecosystems.

scholarly journals High diversity of nitrogen-fixing bacteria in the upper reaches of the Heihe River, northwestern China

2013 ◽  
Vol 10 (8) ◽  
pp. 5589-5600 ◽  
Author(s):  
X. S. Tai ◽  
W. L. Mao ◽  
G. X. Liu ◽  
T. Chen ◽  
W. Zhang ◽  
...  
Keyword(s):  
Water Conservation ◽  
Bacterial Communities ◽  
Nifh Gene ◽  
Qilian Mountains ◽  
Northwestern China ◽  
Heihe River ◽  
Meadow Soil ◽  
Nitrogen Fixing ◽  
Nitrogen Fixing Bacteria ◽  
Gene Copies

Abstract. Vegetation plays a key role in water conservation in the southern Qilian Mountains (northwestern China), located in the upper reaches of the Heihe River. Nitrogen-fixing bacteria are crucial for the protection of the nitrogen supply for vegetation in the region. In the present study, nifH gene clone libraries were established to determine differences between the nitrogen-fixing bacterial communities of the Potentilla parvifolia shrubland and the Carex alrofusca meadow in the southern Qilian Mountains. All of the identified nitrogen-fixing bacterial clones belonged to the Proteobacteria. At the genus level, Azospirillum was only detected in the shrubland soil, while Thiocapsa, Derxia, Ectothiorhodospira, Mesorhizobium, Klebsiella, Ensifer, Methylocella and Pseudomonas were only detected in the meadow soil. The phylogenetic tree was divided into five lineages: lineages I, II and III mainly contained nifH sequences obtained from the meadow soils, while lineage IV was mainly composed of nifH sequences obtained from the shrubland soils. The Shannon–Wiener index of the nifH genes ranged from 1.5 to 2.8 and was higher in the meadow soils than in the shrubland soils. Based on these analyses of diversity and phylogeny, the plant species were hypothesised to influence N cycling by enhancing the fitness of certain nitrogen-fixing taxa. The number of nifH gene copies and colony-forming units (CFUs) of the cultured nitrogen-fixing bacteria were lower in the meadow soils than in the shrubland soils, ranging from 0.4 × 107 to 6.9 × 107 copies g−1 soil and 0.97 × 106 to 12.78 × 106 g−1 soil, respectively. Redundancy analysis (RDA) revealed that the diversity and number of the nifH gene copies were primarily correlated with aboveground biomass in the shrubland soil. In the meadow soil, nifH gene diversity was most affected by altitude, while copy number was most impacted by soil-available K. These results suggest that the nitrogen-fixing bacterial communities beneath Potentilla were different from those beneath Carex.

scholarly journals Nitrogen-Fixing Bacteria Associated with Peltigera Cyanolichens and Cladonia Chlorolichens

Molecules ◽  
2018 ◽  
Vol 23 (12) ◽  
pp. 3077 ◽  
Author(s):  
Katerin Almendras ◽  
Jaime García ◽  
Margarita Carú ◽  
Julieta Orlando
Keyword(s):  
Bacterial Communities ◽  
Fragment Length Polymorphism ◽  
Multivariate Analyses ◽  
Nifh Gene ◽  
Shannon Index ◽  
Nitrogen Fixing ◽  
Nitrogen Fixing Bacteria ◽  
Symbiotic Interaction ◽  
Different Types ◽  
Terricolous Lichens

Lichens have been extensively studied and described; however, recent evidence suggests that members of the bacterial community associated with them could contribute new functions to the symbiotic interaction. In this work, we compare the nitrogen-fixing guild associated with bipartite terricolous lichens with different types of photobiont: Peltigera cyanolichens and Cladonia chlorolichens. Since cyanobacteria contribute nitrogen to the symbiosis, we propose that chlorolichens have more diverse bacteria with the ability to fix nitrogen compared to cyanolichens. In addition, since part of these bacteria could be recruited from the substrate where lichens grow, we propose that thalli and substrates share some bacteria in common. The structure of the nitrogen-fixing guild in the lichen and substrate bacterial communities of both lichens was determined by terminal restriction fragment length polymorphism (TRFLP) of the nifH gene. Multivariate analyses showed that the nitrogen-fixing bacteria associated with both types of lichen were distinguishable from those present in their substrates. Likewise, the structure of the nitrogen-fixing bacteria present in the cyanolichens was different from that of chlorolichens. Finally, the diversity of this bacterial guild calculated using the Shannon index confirms the hypothesis that chlorolichens have a higher diversity of nitrogen-fixing bacteria than cyanolichens.

scholarly journals Differing Courses of Genetic Evolution of Bradyrhizobium Inoculants as Revealed by Long-Term Molecular Tracing in Acacia mangium Plantations

2014 ◽  
Vol 80 (18) ◽  
pp. 5709-5716 ◽  
Author(s):  
M. M. Perrineau ◽  
C. Le Roux ◽  
A. Galiana ◽  
A. Faye ◽  
R. Duponnois ◽  
...  
Keyword(s):  
Nitrogen Fixation ◽  
Growth Promotion ◽  
Acacia Mangium ◽  
Housekeeping Genes ◽  
Nitrogen Fixing ◽  
Microbial Inoculants ◽  
Nitrogen Fixing Bacteria ◽  
Content Type ◽  
Significant Difference

ABSTRACTIntroducing nitrogen-fixing bacteria as an inoculum in association with legume crops is a common practice in agriculture. However, the question of the evolution of these introduced microorganisms remains crucial, both in terms of microbial ecology and agronomy. We explored this question by analyzing the genetic and symbiotic evolution of twoBradyrhizobiumstrains inoculated onAcacia mangiumin Malaysia and Senegal 15 and 5 years, respectively, after their introduction. Based on typing of several loci, we showed that these two strains, although closely related and originally sampled in Australia, evolved differently. One strain was recovered in soil with the same five loci as the original isolate, whereas the symbiotic cluster of the other strain was detected with no trace of the three housekeeping genes of the original inoculum. Moreover, the nitrogen fixation efficiency was variable among these isolates (either recombinant or not), with significantly high, low, or similar efficiencies compared to the two original strains and no significant difference between recombinant and nonrecombinant isolates. These data suggested that 15 years after their introduction, nitrogen-fixing bacteria remain in the soil but that closely related inoculant strains may not evolve in the same way, either genetically or symbiotically. In a context of increasing agronomical use of microbial inoculants (for biological control, nitrogen fixation, or plant growth promotion), this result feeds the debate on the consequences associated with such practices.

Asymbiotic N2-fixing bacteria associated with three boreal conifers

1984 ◽  
Vol 14 (4) ◽  
pp. 595-597 ◽  
Author(s):  
L. Z. Florence ◽  
F. D. Cook
Keyword(s):  
Nitrogen Fixation ◽  
Acetylene Reduction ◽  
Nitrogen Fixing ◽  
Nitrogen Fixing Bacteria ◽  
Seedling Root ◽  
Natural Stands

Azotobacter spp., Azospirillum spp., and Desulfovibrio spp., were identified as the predominant nitrogen-fixing bacteria associated with seedling root sections collected from natural stands of Pinusbanksiana Lamb., Piceamariana (Mill.) B.S.P., and Larixlaricina (Du Roi) K. Koch growing in Alberta. Samples from the sandy upland habitat of P. banksiana exhibited lower average rates of nitrogen fixation relative to the wet lowland occupied by P. mariana and L. laricina. Average nitrogen-fixing capacity (by acetylene reduction) was greater among bacteria isolates from L. laricina than those from P. mariana. Azospirillum spp. were strongly associated with P. mariana, while Azotobacter spp. were isolated more frequently from L. laricina.

scholarly journals mRNA Extraction and Reverse Transcription-PCR Protocol for Detection of nifH Gene Expression by Azotobacter vinelandii in Soil

2003 ◽  
Vol 69 (4) ◽  
pp. 1928-1935 ◽  
Author(s):  
Helmut Bürgmann ◽  
Franco Widmer ◽  
William V. Sigler ◽  
Josef Zeyer
Keyword(s):  
Gene Expression ◽  
Nitrogen Fixation ◽  
Reverse Transcription ◽  
Liquid Culture ◽  
Azotobacter Vinelandii ◽  
Nifh Gene ◽  
Rna Extraction ◽  
Soil Microbial Communities ◽  
Nitrogen Fixing ◽  
Reverse Transcription Pcr

ABSTRACT The study of free-living nitrogen-fixing organisms in bulk soil is hampered by the great diversity of soil microbial communities and the difficulty of relating nitrogen fixation activities to individual members of the diazotroph populations. We developed a molecular method that allows analysis of nifH mRNA expression in soil in parallel with determinations of nitrogen-fixing activity and bacterial growth. In this study, Azotobacter vinelandii growing in sterile soil and liquid culture served as a model system for nifH expression, in which sucrose served as the carbon source and provided nitrogen-limited conditions, while amendments of NH4NO3 were used to suppress nitrogen fixation. Soil RNA extraction was performed with a new optimized direct extraction protocol that yielded nondegraded total RNA. The RNA extracts were of high purity, free of DNA contamination, and allowed highly sensitive and specific detection of nifH mRNA by a reverse transcription-PCR. The level of nifH gene expression was estimated by PCR amplification of reverse-transcribed nifH mRNA fragments with A. vinelandii-specific nifH primers. This new approach revealed that nifH gene expression was positively correlated with bulk nitrogen fixation activity in soil (r 2 = 0.72) and in liquid culture (r 2 = 0.84) and therefore is a powerful tool for studying specific regulation of gene expression directly in the soil environment.

Phylogenetic diversity of nitrogen-fixing bacteria and thenifHgene from mangrove rhizosphere soil

2012 ◽  
Vol 58 (4) ◽  
pp. 531-539 ◽  
Author(s):  
Jianyin Liu ◽  
Mengjun Peng ◽  
Youguo Li
Keyword(s):  
Phylogenetic Trees ◽  
Rhizosphere Soil ◽  
Nifh Gene ◽  
Soil Samples ◽  
Community Diversity ◽  
Nitrogen Fixing ◽  
Pcr Cloning ◽  
Nitrogen Fixing Bacteria ◽  
Soil Dna ◽  
Mangrove Ecosystems

Nine types of nitrogen-fixing bacterial strains were isolated from 3 rhizosphere soil samples taken from mangrove plants in the Dongzhaigang National Mangrove Nature Reserve of China. Most isolates belonged to Gammaproteobacteria Pseudomonas , showing that these environments constituted favorable niches for such abundant nitrogen-fixing bacteria. New members of the diazotrophs were also found. Using a soil DNA extraction and PCR-cloning-sequencing approach, 135 clones were analyzed by restriction fragment length polymorphism (RFLP) analysis, and 27 unique nifH sequence phylotypes were identified, most of which were closely related to sequences from uncultured bacteria. The diversity of nitrogen-fixing bacteria was assessed by constructing nifH phylogenetic trees from sequences of all isolates and clones in this work, together with related nifH sequences from other mangrove ecosystems in GenBank. The nifH diversity varied among soil samples, with distinct biogeochemical properties within a mangrove ecosystem. When comparing different mangrove ecosystems, the nifH gene sequences from a specific site tended to cluster as individual groups. The results provided interesting data and novel information on our understanding of diazotroph community diversity in the mangrove ecosystems.

Long-term effect of inoculating lodgepole pine seedlings with plant growth-promoting bacteria originating from disturbed gravel mining ecosystem

2020 ◽  
Author(s):  
Kiran Preet Padda ◽  
Akshit Puri ◽  
Christopher Chanway
Keyword(s):  
Plant Growth ◽  
Lodgepole Pine ◽  
Tree Regeneration ◽  
Lytic Enzyme ◽  
Plant Growth Promoting Bacteria ◽  
Bacterial Strains ◽  
Acetic Acid Production ◽  
Gravel Mining ◽  
Pine Seedlings ◽  
Pine Trees

Gravel mining is prevalent in forest landscapes of Canada, typically resulting in complete loss of vegetation and topsoil. Despite such extreme disturbance, lodgepole pine (<i>Pinus contorta</i> var. <i>latifolia</i>) trees are thriving at unreclaimed gravel pits located in central-interior British Columbia, possibly due, at least in part, to the association of pine trees with their endophytic bacteria. Testing this possibility, several bacterial strains were previously isolated from pine trees growing at these pits, of which 14 were identified as effective nitrogen-fixers. In this study, we evaluated the inoculation effect of these 14 strains on lodgepole pine growth under nitrogen-poor conditions. Each strain colonized the rhizosphere and internal tissues of pine seedlings and significantly enhanced their length (24–65%) and biomass (100–300%), 18 months after sowing and inoculation. Notably, three <i>Pseudomonas</i> strains increased pine seedling length by 1.6-fold and biomass by 4-fold. Most strains also demonstrated substantial potential to promote plant growth via phosphorus solubilization, siderophore production, 1-aminocyclopropane-1-carboxylic acid deaminase activity, indole-3-acetic acid production, lytic enzyme activity and catalase activity. Our results suggest that such effective bacteria could be sustaining pine growth on bare gravel, indicating a possible ecological association that may explain natural tree regeneration in such a disturbed ecosystem.

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