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Groundnut (Arachis hypogaea) nodule Rhizobium and passenger endophytic bacterial cultivable diversity and their impact on plant growth promotion
Abstract
Root nodules offer habitat not only for Rhizobium also for a wide variety of endophytic bacteria that have a positive impact on growth and nutrient status of the plant. To evaluate this statement, the present study was aimed to reveal the cultivable Rhizobium (R) and passenger endophytes (PE) of groundnut nodule and explored their impact on plant growth parameters. Two groundnut phenotypes, bunch (cv., VRI 2) and semi-spreading (cv., VRI 7) varieties nodule were collected for isolation of endophytes. Molecular characterization of nodule endophytic bacterial isolates were done using 16S rDNA sequencing. Isolates were screened based on their early plant growth promotion and selected efficient R and PE were co-inoculated to assess the impact on plant growth. Ninety eight nodule endophytic bacterial isolates were distributed among the Firmicutes, Alphaproteobacteria, Betaproteobacteria and Gammaproteobacteria. In bunch type groundnut (VRI 2) co-inoculation of Rhizobium phaseoli S18 and Pantoea dispersa YBB19B registered significantly higher shoot length, root length, dry weight and nodule number. Similarly, in semi-spreading groundnut (VRI 7) co-inoculation of Rhizobium mayense S19 with Kosakonia oryzae ESB1 recorded maximum shoot length, root length, dry weight and nodule number. R and PE in nodules are visualized in SEM micrograph and changes in root exudates composition as a result of endophytic bacteria inoculation was assessed through GC-MS. This study showed that the nodule endophytic bacterial diversity of groundnut is vast and co-inoculation of native R and nodule PE significantly improved the plant growth than that of Rhizobium alone.
... While, the former traditionally belong to the genera Azorhizobium, Bradyrhizobium, Ensifer, Mesorhizobium and Rhizobium NRE include Aminobacter, Aerobacter, Bacillus, Enterobacter, Erwinia, Klebsiella, Paenibacillus, Pantoea, Pseudomonas, Staphylococcus etc. [10]. Despite the potential of endophytes in stress management and adequate knowledge on the nodule endophytes [11] studies on bio-control potential of nodule endophytic bacteria against stem rot disease in groundnut crop are limited. Therefore the present study aimed to exploit the nodule endophytic bacteria with antagonistic ability as potential biocontrol agents for the management of stem rot disease in groundnut. ...
... The endophytic bacteria from root nodules of groundnut were isolated based on the method given by [11]. The nodules were separated from the roots carefully avoiding any damage and were surface-sterilized using 3 per cent sodium hypochlorite for 30 s. ...
Groundnut is a major oilseed crop grown throughout the world. The crop is constrained by several biotic and abiotic stresses, among which stem rot disease caused by Sclerotium rolfsii poses significant threat to groundnut production. The management of stem rot disease using chemical methods has adverse health and ecological impacts. Eco-friendly alternatives such as the use of microbial antagonists including bacterial endophytes is an sustainable approach. In this context, the study explored the potential of nodule endophytic bacteria of groundnut for antagonism against S. rolfsii. A survey was conducted in major groundnut growing regions of Telangana during rabi, 2023. Eight different isolates of S. rolfsii and twenty nodule endophytic bacterial isolates were isolated from the infected and healthy groundnut plants, respectively. S. rolfsii isolate GNS1 was the most virulent of all with the lowest incubation period and days to permanent wilting. Among the 20 bacterial endophytes, isolates GNEB13 and GNEB6 were the most effective with highest mycelial inhibition of S. rolfsii under in vitro conditions. These isolates have the potential to be evaluated under field conditions for sustainable management of stem rot disease in groundnut.
... Rhizobial endophytes Rhizobium pusense S6R2, R. phaseoli S18, Rhizobium mayense S11R1, and R. phaseoli S10R2, passenger endophytes BreviBacillus brevis S8R1, Bacillus tequilensis NBB13, Pantoea dispersa YBB19B, Bacillus altitudinous TBB5A, Enterobacter cloacae S23, and Paenibacillus illionoisensis YBB20 were previously isolated from groundnut nodules (Preyanga et al., 2021) were selected for the present study. The pure cultures of rhizobial and passenger endophytes were routinely cultured in liquid yeast extract mannitol and nutrient agar medium at 28±2 • C for 24-48 h, respectively. ...
... Under moisture stress conditions, groundnut plants treated with rhizobial and passenger endophytes had their root exudates examined. A funnel flask experimental setup was used to collect groundnut variety VRI 2 root exudates (Preyanga et al., 2021;Oviya et al., 2023). Groundnut seeds (VRI 2) were obtained from Regional Research Station located at Vriddhachalam, Tamil Nadu Agricultural University, India. ...
Moisture stress poses a significant threat to global agriculture, compromising crop yields and food security. In the quest for sustainable solutions, endophytic microorganisms have emerged as promising candidates for enhancing plant resilience to drought. The study's primary goal was to analyse the significance of bacterial endophytes, both rhizobial and passenger endophytes, in alleviating the effects of moisture stress. Here, PEG 6000 was used to test the drought endurance of the ten identified rhizobial and passenger endophytes. Rhizobium pusense S6R2, Enterobacter cloacae S23 and Bacillus tequilensis NBB13 were selected as best performing endophytes as they showed high tolerance of poly ethylene glycol (PEG) and maximum plant growth promoting traits like Indole Acetic Acid, exopolysaccharide production, biofilm formation, 1-aminocyclopropane1-carboxylate (ACC) deaminase activity, siderophore, zinc and phosphorous solubilisation even in PEG induced moisture stress condition. Metabolite analysis revealed that twenty-four significant compounds mostly belong to fatty acyls, amino acids, peptides, polyketides, and benzenoids were found in the root exudates of groundnut treated with endophytes. The best-performing endophytes were used in a pot culture experiment, with groundnut as the test crop. The current study found that co-inoculation of Rhizobium pusense S6R2 and Enterobacter cloacae S23 significantly increased nodule number, growth, photosynthetic pigment, anti-oxidant enzymes, and osmolyte under moisture stressed conditions when compared to other treatments. As a result, co-inoculation of Rhizobium and entophytic bacteria may be recommended as a bio-inoculant for groundnut for moisture stress alleviation after confirming the results in field evaluation.
... B6, which was isolated in Algerian desert soil, had been demonstrated to have biopesticide properties against Locusta migratoria nymphs [6]. We isolated Kosakonia oryzae ESB1 and Enterobacter cloacae S23 from groundnut nodules that possessed multiple plant growth-promoting traits and promoted groundnut growth under pot culture conditions [7]. There are several Enterobacter species and related members of the Enterobacteriaceae family that exhibit PGP in response to abiotic stress have also been identified in recent years. ...
... Passenger endophytes (PE) are endophytes that enter a plant by accident and remain there in the absence of forces that would maintain their presence. In a previous study [7], we carefully characterized the rhizobial and passenger endophytes in groundnut nodules and recorded their PGP. The capacity of endophytes to confer plant stress resistance opens a new avenue for mitigating the detrimental effects of various stresses on agricultural output. ...
Aim
This study aims to reveal the passenger endophytic bacterium Enterobacter cloacae S23 isolated from groundnut nodules and to underpin the molecular mechanism and genes responsible for abiotic stress tolerance.
Background
A variety of microorganisms that contribute to nodulation and encourage plant development activity in addition to the nodulating Rhizobium. Passenger endophytes (PE) are endophytes that accidentally penetrate the plant without any selective pressure keeping them in the interior tissue of the plant. PE possesses characteristics that encourage plant development and boost output while reducing pathogen infection and improving biotic and abiotic stress tolerance. However, there is a lack of molecular evidence on the passenger endophyte-mediated alleviation of abiotic stresses.
Objective
This study was formulated to reveal the draft genome sequence of Enterobacter cloacae S23, as well as genes and characteristics involved in plant growth promotion and stress tolerance.
Method
The data were submitted to PATRIC and the TORMES-1.0 Unicyclker tools were used to conduct a complete genome study of Enterobacter cloacae S23. The TORMES-1.0 platform was used to process the reads. RAST tool kit (RASTtk) was used to annotate the S23 sequence. The plant growth-promoting traits such as indole acetic acid production, siderophore secretion, production of extracellular polysaccharides, biofilm formation, phosphate solubilization, and accumulation of osmolytes were examined under normal, 7% NaCl and 30% polyethylene glycol amended conditions to determine their ability to withstand salt and moisture stressed conditions, respectively.
Result
We report the size of Enterobacter cloacae S23 is 4.82Mb which contains 4511 proteincoding sequences, 71 transfer RNA genes, and 3 ribosomal RNA with a G+C content of DNA is 55.10%. Functional analysis revealed that most of the genes are involved in the metabolism of amino acids, cofactors, vitamins, stress response, nutrient solubilization (kdp, pho, pst), biofilm formation (pga) IAA production (trp), siderophore production (luc, fhu, fep, ent, ybd), defense, and virulence. The result revealed that E. cloacae S23 exhibited multiple plant growth-promoting traits under abiotic stress conditions.
Conclusion
Our research suggested that the discovery of anticipated genes and metabolic pathways might characterise this bacterium as an environmentally friendly bioresource to support groundnut growth through several mechanisms of action under multi-stresses.
... Legume-rhizobial symbiosis is known for the fixation of atmospheric nitrogen through a unique structure called root nodules. Root nodules are typical structures harbouring different types of bacteria (Kumar et al., 2017;Dekak et al., 2020), which are predominantly colonized with rhizobial species (Palaniappan et al., 2010), while other bacteria present inside the nodules are collectively known as non-rhizobial endophytes (NREs) (Preyanga et al., 2021). Divergent groups of NREs have been reported to assist the nodulation process and stimulate plant growth (Preyanga et al., 2021). ...
... Root nodules are typical structures harbouring different types of bacteria (Kumar et al., 2017;Dekak et al., 2020), which are predominantly colonized with rhizobial species (Palaniappan et al., 2010), while other bacteria present inside the nodules are collectively known as non-rhizobial endophytes (NREs) (Preyanga et al., 2021). Divergent groups of NREs have been reported to assist the nodulation process and stimulate plant growth (Preyanga et al., 2021). These NREs enter the plant through tissues and maintain themselves in the internal compartment of the plant (Hardoim et al., 2008). ...
Non-rhizobial endophytes (NREs) are active colonizers inhabiting the root nodules. Though their active role in the lentil agroecosystem is not well defined, here we observed that these NREs might promote the growth of lentils, modulate rhizospheric community structure and could be used as promising organisms for optimal use of rice fallow soil. NREs from root nodules of lentils were isolated and examined for plant growth-promoting traits, exopolysaccharide (EPS) and biofilm production, root metabolites, and the presence of nifH and nifK elements. The greenhouse experiment with the chosen NREs, i.e., Serratia plymuthica 33GS and Serratia sp. R6 significantly increased the germination rate, vigour index, development of nodules (in non-sterile soil) and fresh weight of nodules (33GS 94%, R6 61% growth) and length of the shoot (33GS 86%, R6 51.16%) as well as chlorophyll levels when compared to the uninoculated control. Scanning Electron Microscopy (SEM) revealed that both isolates could successfully colonize the roots and elicit root hair growth. The inoculation of the NREs resulted in specific changes in root exudation patterns. The plants with 33GS and R6 treatment significantly stimulated the exudation of triterpenes, fatty acids, and their methyl esters in comparison to the uninoculated plants, altering the rhizospheric microbial community structure. Proteobacteria dominated the rhizospheric microbiota in all the treatments. Treatment with 33GS or R6 also enhanced the relative abundance of other favourable microbes, including Rhizobium, Mesorhizobium, and Bradyrhizobium. The correlation network analysis of relative abundances resulted in numerous bacterial taxa, which were in cooperation with each other, having a possible role in plant growth promotion. The results indicate the significant role of NREs as plant growth promoters, which also includes their role in root exudation patterns, enhancement of soil nutrient status and modulation of rhizospheric microbiota, suggesting their prospects in sustainable, and bio-based agriculture.
... For more than a century, nitrogen-fixing nodules were thought to be uniquely inhabited by rhizobia, and culture-based studies have provided important information about the benefits of rhizobia, including differences in the efficiency of the BNF process among legumes and also between strains of the same host legume, e.g., [1][2][3][4][5]. It was only recently that the first genomic studies reported complex microbial communities inside the nodules [25][26][27][28], which was confirmed in our study in nodule microbiomes of both soybean and common bean. However, despite evolving as unique ecological niches for nitrogen fixation through the symbiotic relationship between the host legumes and rhizobia, the biological implications of sharing the nodule environment with other bacteria are not well understood yet. ...
... However, despite evolving as unique ecological niches for nitrogen fixation through the symbiotic relationship between the host legumes and rhizobia, the biological implications of sharing the nodule environment with other bacteria are not well understood yet. One hypothesis is that other bacteria living inside the nodules and that probably entered the nodule together with rhizobia are endophytes that can help in plant-growth promotion by other microbial processes, such as the synthesis of phytohormones, antimicrobial molecules, siderophores, mineral solubilization capacity, among others [27][28][29][30]. As an example of benefit, an interesting study of nodule endophytes performed with Lotus burtii pointed out that in healthy nodules of this legume, Pseudomonas species were the prevalent non-rhizobia, and when used as inoculum infected the plant together with a beneficial Mesorhizobium, but not with an ineffective Rhizobium, benefiting the symbiosis by decreasing the number of ineffective nodules [31]. ...
In this paper we set up to explore root an nodule microbiomes from soybean and common bean searching for effects after inoculation with nitrogen-fixing bacteria.
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Biological nitrogen fixation (BNF) is a key process for the N input in agriculture, with outstanding economic and environmental benefits from the replacement of chemical fertilizers. However, not all symbioses are equally effective in fixing N2, and a major example relies on the high contribution associated with the soybean (Glycine max), contrasting with the low rates reported with the common bean (Phaseolus vulgaris) crop worldwide. Understanding these differences represents a major challenge that can help to design strategies to increase the contribution of BNF, and next-generation sequencing (NGS) analyses of the nodule and root microbiomes may bring new insights to explain differential symbiotic performances. In this study, three treatments evaluated in non-sterile soil conditions were investigated in both legumes: (i) non-inoculated control; (ii) inoculated with host-compatible rhizobia; and (iii) co-inoculated with host-compatible rhizobia and Azospirillum brasilense. In the more efficient and specific symbiosis with soybean, Bradyrhizobium presented a high abundance in nodules, with further increases with inoculation. Contrarily, the abundance of the main Rhizobium symbiont was lower in common bean nodules and did not increase with inoculation, which may explain the often-reported lack of response of this legume to inoculation with elite strains. Co-inoculation with Azospirillum decreased the abundance of the host-compatible rhizobia in nodules, probably because of competitiveness among the species at the rhizosphere, but increased in root microbiomes. The results showed that several other bacteria compose the nodule microbiomes of both legumes, including nitrogen-fixing, growth-promoters, and biocontrol agents, whose contribution to plant growth deserves further investigation. Several genera of bacteria were detected in root microbiomes, and this microbial community might contribute to plant growth through a variety of microbial processes. However, massive inoculation with elite strains should be better investigated, as it may affect the root microbiome, verified by both relative abundance and diversity indices, that might impact the contribution of microbial processes to plant growth.
... Similarly, higher root growth in the inoculated treatments caused more P acquisition (Aloo et al. 2019;Araujo et al. 2021). The cultivation of inoculated legumes enhances K availability in aboveground plant parts by enhancing its solubility (Preyanga et al. 2021). ...
Low soil organic matter contents are among the most predominant reasons behind reduced soil health and decreased agricultural productivity. Sustainable agricultural approaches encourage minimal farmer’s dependence on chemical fertilizers and favor their supplementation with organic and biofertilizers to ensure smooth crop growth and sustained soil health. These organic amendments improve soil health by regulating various soil physicochemical and biological properties and thereby, control soil nutrient dynamics. Moreover, their integration with beneficial bacteria has also yielded significant outcomes in lieu of plant growth promotion and soil health improvement. Here, we performed a pot experiment to check the efficiency of different bacterial combinations alone and in combination with biochar and press mud (0, 0.25 and 0.50% w/w) for improving the yield and quality attributes of mash bean. Mash bean seeds were coated with freshly prepared inocula of three different combinations of rhizobia and PGPR strains. We found that inoculation with different bacterial consortia led to a significant increment in all the measured growth matrices. However, among them, combination (Bacillus subtilis + Bradyrhizobium japonicum) along with BC at 0.50% and PM at 0.25% caused maximum increase in SDW (124%), No. of pods per plant (360%), soil dehydrogenase (297%), alkaline phosphatase (149%), seed crude protein (119%), N (118%), P (127%), K (63%), niacin (68%), and thiamine (195%) contents as compared to their respective uninoculated controls. Pearson correlation heatmap revealed a strong positive correlation between various plant attributes and soil enzyme activities such as ALP (r = 0.69***) and ACP (r = 0.70***) levels. Furthermore, ALP was positively correlated with the plant yield (r = 0.66). We concluded that beneficial consortium (Bacillus subtilis + Bradyrhizobium japonicum) applied synergistically along with press mud and biochar can significantly improve the yield and seed quality of mash bean and at the same time, enhance the soil biological health as pragmatic by increased soil enzyme activities.
... In addition, it appears that the root nodule bacteria are not alone in the nodule microbiome as it has been recently recognized that there are entire communities of bacteria within these root nodules, known as nodule-associated bacteria, or NAB [14][15][16]. Many of these NAB have plant growth promoting (PGP) metabolic activities, such as N-fixation, pathogen suppression, production of phytohormones for growth, nutrient acquisition, and enhancement of root and shoot length, all of which aid in the primary root nodule development and plant growth [15][16][17][18][19][20][21][22]. However, little is known about the NAB communities in tropical forest leguminous tree nodules. ...
Leguminous tree root nodule nitrogen-fixing bacteria are critical for recuperation of soil C and N cycle processes after disturbance in tropical forests, while other nodule-associated bacteria (NAB) may enhance nodule development and activity, and plant growth. However, little is known of these root nodule microbiomes. Through DNA analysis, we evaluated the bacterial taxa associated with the root nodules of the 1-year-old, 2-year-old, 13-year-old, and old growth Inga punctata trees in a cloud forest. Bradyrhizobium diazoefficiens was the dominant taxon found in all nodules at 63.16% to 85.71% mean percent sequences (MPS) of the total nodule bacterial DNA and was found in the youngest nodules examined (1 year old), suggesting that it is the primary nodular bacteria. There were 26 other NAB genera with collective MPS levels between 7.4% to 12.2%, while 15 of these genera were found in the Bulk Forest soils at collective MPS levels of 4.6%. These bacterial community compositions were different between the NAB and Bulk Forest soils, suggesting the NAB became concentrated within the root nodules, resulting in communities with different compositions from the Bulk Forest soils. Twenty-three of the 26 NAB genera were previously identified with the potential to perform 9 plant growth promoting (PGP) activities, suggesting their importance in root nodule development and plant growth. These NAB communities appeared to successionally develop over time into more complex taxonomic communities, which is consistent with the outcome of advanced microbial communities following succession. The presence of both B. diazoefficiens and the NAB communities in the nodules across all ages of tree roots, and the potential for PGP activities linked with most of the NAB genera, suggest the importance of B. diazoefficiens and the NAB community for nodule development and enhanced development and growth of I. punctata throughout its lifespan, and most critically in the younger plants.
... Similarly, among the Gammaproteobacteria enriched in roots, nif H sequences associated with the genera Kosakonia and Enterobacter were found . Members of the Enterobacteriaceae with the ability to fix N 2 have been isolated as endophytes of rice (Prakamhang et al. 2009 ), and the genus Kosakonia has been reported as a PGPB in rice (Mosquito et al. 2020 ), sugarcane and groundnut (Taulé et al. 2019, Leite et al. 2021, Preyanga et al. 2021. ...
Aim
Bacteria that promote plant growth, such as diazotrophs, are valuable tools for achieving a more sustainable production of important non-legume crops like rice. Different strategies have been used to discover new bacteria capable of promoting plant growth. This work evaluated the contribution of soil diazotrophs to the endophytic communities established in the roots of rice seedlings cultivated on seven representative soils from Uruguay.
Methods and results
The soils were classified into two groups according to the C and clay content. qPCR, terminal restriction fragment length polymorphism (T-RFLP), and 454-pyrosequencing of the nifH gene were used for analyzing diazotrophs in soil and plantlets’ roots grown from seeds of the same genotype for 25 days under controlled conditions. A similar nifH abundance was found among the seven soils, roots, or leaves. The distribution of diazotrophs was more uneven in roots than in soils, with dominance indices significantly higher than in soils (nifH T-RFLP). Dominant soils’ diazotrophs were mainly affiliated to Alphaproteobacteria and Planctomycetota. Conversely, Alpha, Beta, Gammaproteobacteria, and Bacillota were predominant in different roots, though undetectable in soils. Almost no nifH sequences were shared between soils and roots.
Conclusions
Root endophytic diazotrophs comprised a broader taxonomic range of microorganisms than diazotrophs found in soils from which the plantlets were grown and showed strong colonization patterns.
... IAC Tatu other collaborative microorganisms, e.g., Azospirillum brasilense (Gericó et al., 2020), Bacillus spp. (Figueredo et al., 2014;Preyanga et al., 2021;Kaschuk et al., 2022), Serratia marcescens, and Trichoderma harzianum (Badawi et al., 2011). The benefits of coinoculation may be direct on nodule formation or indirect; for example, a meta-analysis revealed that coinoculation of bradyrhizobia and Bacillus spp. ...
Groundnut plants can obtain N from N 2 fixation via symbiosis with rhizobia, and inoculation with selected strains can improve grain yields. We report the results of four field experiments carried out under subtropical conditions to confirm whether microbial inoculants can improve groundnut performance through the effects of single inoculation with Bradyrhizobium arachidis (SEMIA6144), coinoculation with Arthrospira platensis (IPR7059) or Synechocystis sp. (IPR7061), or N fertilization with 100 kg ha ⁻¹ N on plant growth, nodulation, N accumulation in tissues, grain protein concentration (GPC), and grain yield. There were no effects of inoculation treatment or N fertilizer on shoot or root dry weight. In clayey soil, coinoculation with B. arachidis and cyanobacteria increased grain productivity by an average of 19% compared to that in the noninoculated control. In this clayey soil with a higher P content, regardless of whether coinoculated with B. arachidis or cyanobacteria or single inoculated, grain productivity was 16% greater on average than that resulting from N fertilizer addition. In conclusion, the success of rhizobial inoculation in groundnuts is dependent on the soil, probably due to P limitation and weather conditions.
... Tolerance of Isolates to High Temperature, NaCl, and Low pH All the isolates were tested for growth at high temperature, elevated NaCl, and low pH on YMA agar plates. Freshly grown isolates were spotted four times as replications for a single isolate on YMA agar plates and incubated at 28 • C for 48 h for low pH (4,5,7) and 72 h for various NaCl concentrations (0, 0.15, 0.3, and 0.6 M). For high temperature, isolates were incubated at 37 • C, 40 • C, and 45 • C for 72 h. ...
It is evident that legume root nodules can accommodate rhizobial and non-rhizobial bacterial endophytes. Our recent nodule microbiome study in peanuts described that small nodules can harbor diverse bacterial endophytes. To understand their functional role, we isolated 87 indigenous endophytes from small nodules of field-grown peanut roots and characterized them at molecular, biochemical, and physiological levels. The amplified 16S rRNA genes and phylogenetic analysis of these isolates revealed a wide variety of microorganisms related to the genera Bacillus, Burkholderia, Enterobacter, Herbaspirillum, Mistsuaria, Pantoea, Pseudomonas, and Rhizobia. It was observed that 37% (100% identity) and 56% (>99% identity) of the isolates matched with the amplified sequence variants (ASVs) from our previous microbiome study. All of these isolates were tested for stress tolerance (high temperature, salinity, acidic pH) and phosphate (P) solubilization along with ammonia (NH3), indole-3-acetic acid (IAA), 1-aminocyclopropane-1-carboxylate deaminase (ACCD), and siderophore production. The majority (78%) of the isolates were found to be halotolerant, thermotolerant, and acidophilic, and a few of them showed a significant positive response to the production of IAA, NH3, siderophore, ACCD, and P-solubilization. To evaluate the plant growth promotion (PGP) activity, plant and nodulation assays were performed in the growth chamber conditions for the selected isolates from both the non-rhizobial and rhizobial groups. However, these isolates appeared to be non-nodulating in the tested conditions. Nonetheless, the isolates 2 (Pantoea), 17 (Burkholderia), 21 (Herbaspirillum), 33o (Pseudomonas), and 77 (Rhizobium sp.) showed significant PGP activity in terms of biomass production. Our findings indicate that these isolates have potential for future biotechnological applications through the development of biologicals for sustainable crop improvement.
... The latter species and other Kosakonia spp. have been found as endophytes and PGP bacteria in rice (Mosquito et al. 2020), sugar cane, and groundnut (Taulé et al. 2019;Preyanga et al. 2021;Leite et al. 2021). Komagataeibacter spp., an acetobacteria closely related to the genus Gluconacetobacter, which includes well-known nitrogen-fixing species (Yamada et al. 2012) and is outstanding for its successful association with sugarcane (Cavalcante and Dobereiner 1988;Fischer et al. 2012), was also dominant inside rice roots, particularly enriched at FF stage (T-RF 107 bp). ...
Biological nitrogen fixation contributes greatly to the sustainability of agroecosystems. However, more information is needed about the impact that agricultural intensification, a frequent practice nowadays, would have on diazotrophic communities associated with plants. This work addresses the influence of intensifying rice cropping systems on the abundance, diversity, and structure of diazotrophic communities associated with rice roots (rhizospheric and endophytic bacteria) from a field experiment. Three different rice rotation systems (rice–pasture, rice–soybean, and continuous rice) at two crop growth stages (TBF: tillering before flooding and FF: flowering-flooded) were studied. The results showed that the rhizospheric soils had the greater nifH gene abundance, and the abundance and diversity of rhizospheric and endophytic diazotrophic communities significantly increased at the FF stage. Conversely, nifH abundance in bulk soils remained unaltered. Phylogenetically and metabolically diverse diazotrophic communities were found to be associated with rice roots from the different crop stages and rotations. According to multivariate, clustering and statistical analyses performed on results retrieved by nifH T-RFLP (primers F2/R6 and PolF/PolR and AluI), the interaction of root compartment and crop growth stage (PERMANOVA, p < 0.001) was the major driver of diazotrophic communities. Furthermore, a significant effect of the rice rotation systems on the structure of diazotrophic communities was found (PERMANOVA, p < 0.05), suggesting that crop intensification could impact diazotrophic communities associated with rice plants, which play a key role in plant growth promotion. The implications that this could have should be explored and considered when developing sustainable intensification strategies in rice production.
... Acinetobacter baumannii have been recognized as an environmental pollutant degrading bacterial agent (Zhang et al. 2021). Kosakonia oryzae have been recognized as a plant endophytic and growth-promoting strain (Preyanga et al. 2021) and have the biodegradation potential for pollutants (Dash and Osborne 2020). To the best of our knowledge, there has been no report on NH 4 + -N or S 2− removal capacity of Kosakonia sp. ...
In this study, two NH4⁺-N and S²⁻ removal strains, namely, Kosakonia oryzae (FB2-3) and Acinetobacter baumannii (L5-4), were isolated from the packing materials in a long-running biotrickling filter (BTF). The removal capacities of combined FB2-3 and L5-4 (FB2-3 + L5-4) toward 100 mg L⁻¹ of NH4⁺-N and 200 mg L⁻¹ of S²⁻ reached 97.31 ± 1.62% and 98.57 ± 1.12% under the optimal conditions (32.0 °C and initial pH = 7.0), which were higher than those of single strain. Then, FB2-3 and L5-4 liquid inoculums were prepared, and their concentrations respectively reached 1.56 × 10⁹ CFU mL⁻¹ and 1.05 × 10⁹ CFU mL⁻¹ by adding different resuspension solutions and protective agents after 12-week storage at 25 °C. Finally, pilot-scale BTF test showed that NH3 and H2S in the real exhaust gases from a pharmaceutical factory were effectively removed with removal rates > 87% and maximum elimination capacities were reached 136 g (NH3) m⁻³ h⁻¹ and 176 g (H2S) m⁻³ h⁻¹ at 18 °C-34 °C and pH 4.0–7.0 in the BTF loaded with bamboo charcoal packing materials co-immobilized with FB2-3 and L5-4. After co-immobilization of FB2-3 and L5-4, in the bamboo charcoal packing materials, the new microbial diversity composition contained the dominant genera of Acinetobacter, Mycobacterium, Kosakonia, and Sulfobacillus was formed, and the diversity of entire bacterial community was decreased, compared to the control. These results indicate that FB2-3 and L5-4 have potential to be developed into liquid ready-to-use inoculums for effectively removing NH3 and H2S from exhaust gases in BTF.
Graphical abstract
... They are called root nodule bacteria [8]. These days, several names have been proposed by different reports like non-rhizobia endophytes [9], nodule endophytes [10], nodule associated bacteria [11,12] and most recently Preyanga et al. [13] proposed passenger endophyte bacteria. Martinez-Hidalgo and Hirsch [4] have revealed that rhizobia and the passenger endophytes act together as a community within the root nodules to facilitate plant health and survival, particularly under conditions of environmental stress. ...
Background
In soils, phosphorous (P) mostly exists in fixed/insoluble form and unavailable for plants use in soil solution, hence it is in scarcity. P is fixed in the form of aluminium, iron and manganese phosphates in acidic soils and calcium phosphate in alkaline soils. Phosphate solubilizing bacteria, the ecological engineers play a pivotal role in the mobilization of fixed forms of P by using different mechanisms. The objectives of this study were to evaluate inorganic phosphate solubilizing efficiency and other multiple plant growth promoting traits of Erythrina brucei root nodule endophytic bacteria and to investigate effects of the selected endophytic bacteria on the growth of wheat plant under phosphorous deficient sand culture at greenhouse conditions.
Results
Among a total of 304 passenger endophytic bacteria, 119 (39%) exhibited tricalcium phosphate (TCP) solubilization; however, none of them were formed clear halos on solid medium supplemented with aluminum phosphate (Al-P) or iron phosphate (Fe–P). Among 119 isolates, 40% exhibited IAA production. The selected nine potential isolates also exhibited potentials of IAA, HCN, NH 3 and/or hydrolytic enzymes production. All the selected isolates were potential solubilizers of the three inorganic phosphates (Al-P, Fe–P and TCP) included in liquid medium. The highest values of solubilized TCP were recorded by isolates AU4 and RG6 ( A. soli ), 108.96 mg L ⁻¹ and 107.48 mg L ⁻¹ , respectively at sampling day3 and 120.36 mg L ⁻¹ and 112.82 mg L ⁻¹ , respectively at day 6. The highest values of solubilized Al-P and Fe–P were recorded by isolate RG6, 102.14 mg L ⁻¹ and 96.07 mg L ⁻¹ , respectively at sampling days 3 and 6, respectively. The highest IAA, 313.61 µg mL ⁻¹ was recorded by isolate DM17 ( Bacillus thuringiensis ). Inoculation of wheat with AU4, RG6 and RG5 ( Acinetobacter soli ) increased shoot length by 11, 17.4 and 14.6%, respectively compared to the negative control. Similarly, 76.9, 69.2 and 53.8% increment in shoot dry weight is recorded by inoculation with RG6, AU4 and RG5, respectively. These nine potential endophytic isolates are identified to Gluconobacter cerinus (4), Acinetobacter soli ( 3) , Achromobacter xylosoxidans (1) and Bacillus thuringiensis (1).
Conclusion
AU4, RG6 and RG5 can be potential bio-inoculants candidates as low cost agricultural inputs in acidic and/or alkaline soils for sustainable crop production.
... 42 S were detected inside the root nodules. Similarly, detection of R. mayense was detected earlier in Arachis hypogea nodule using SEM (Preyanga et al., 2021). Moreover, colonization of Bradyrhizobium sp. ...
Diazotrophic nodule isolates are acknowledged promoters of plant growth and rhizospheric community. Consequently, in the lentil agroecosystem, inoculation of atypical rhizobial isolates could be a viable alternative to chemical fertilizers for fallow land usage optimization. The aim of this study is to evaluate and select the rhizobial isolates of lentil nodules with plant-growth-promoting (PGP) attributes and to elucidate their application in rice-fallow soil for determining the growth of lentils and its impact on the rhizospheric bacterial community. Lentil’s nodule isolates were identified and screened for their PGP attributes, biofilm, exopolysaccharide (EPS) formation, and early plant growth promotion. The pot experiment with the selected atypical rhizobial isolates Pararhizobium giardinii (P1) and Ochrobactrum sp. (42 S) significantly enhanced germination, vigour index, nodule formation (P1 60%, 42 S 42% increase), nodule fresh weight, shoot length (65% P1 & 35% 42 S), and chlorophyll content as compared to the uninoculated control treatment. The genes for nitrogen fixation nifH and nifK were detected in both isolates. Scanning Electron Microscopy (SEM) revealed successful root and nodule colonization by both isolates, while Transmission Electron Microscopy (TEM) displayed nitrogen-fixing zones within root nodules. Proteobacteria predominated in the lentil rhizosphere of all the treatments. Whereas, application of either P1 or 42 S increased Rhizobium, Mesorhizobium, and Bradyrhizobium genra, thus positively modulating rhizospheric community structure. The correlation network analysis revealed an abundance of some interdependent bacterial genera with a possible role in overall plant growth. Functional genes for siderophore biosynthesis and ABC transporter were positively modulated by application of either P1 or 42 S. This study showed the significant effect of P. giardinii P1 and Ochrobactrum sp. 42 S of L. culinaris on lentil growth, improving fallowsoil health for optimum usage, and modulated rhizospheric community structure which strongly manifest prospects of low-cost, eco-friendly and sustainable biofertilizers.
... Similarly, detection of R. mayense was detected earlier in Arachis hypogea nodule using SEM (Preyanga et al., 2021). Moreover, colonization of Bradyrhizobium sp. ...
... [15,92,100,121]. Many reports which were used in this meta-analysis confirmed the capacity of Bacillus strains to solubilize P in growth media and in soil used for the cultivation of inoculated legumes [19,34,38,44,49,78,79,91]. In addition to the above results, shoot and root biomasses of chickpeas and soybeans were increased by the rhizobiabacilli coinoculation and suggested that root growth provides more resources for shoot growth (Table 3). ...
Coinoculation of symbiotic N2-fixing rhizobia and plant growth-promoting Bacillus on legume seeds can increase crop productivity. We collected highly resolved data on coinoculation of rhizobia and bacilli on 11 grain legume crops: chickpea, common bean, cowpea, faba bean, groundnut, lentil, mung bean, pea, pigeon pea, soybean, and urad bean to verify the magnitude of additive effects of coinoculation in relation to single inoculation of rhizobia on plant growth and yield of grain legumes. Coinoculation of rhizobia and bacilli on legume seeds and/or soil during sowing significantly increased nodulation, nitrogenase activity, plant N and P contents, and shoot and root biomass, as well as the grain yield of most grain legumes studied. There were however a few instances where coinoculation decreased plant growth parameters. Therefore, coinoculation of rhizobia and Bacillus has the potential to increase the growth and productivity of grain legumes, and can be recommended as an environmental-friendly agricultural practice for increased crop yields.
... The antibiosis, induced systemic resistance, hydrogen cyanide, siderophore, and enzyme production by endophytic microbes protect plants from pathogen attack, so indirectly enhance plant growth (Adeleke and Babalola, 2022). An increase in plant growth and crop yield upon inoculation with plant growth-promoting endophytic bacteria in the genera Bacillus, Enterobacter, Klebsiella, Pantoea, and Rhizobium has been documented (Nascimento et al., 2020;Mowafy et al., 2021;Preyanga et al., 2021). In addition, some endophytic bacteria isolated from oilseed crops have been reported to enhance plant growth due to their plant growthpromoting traits (Lally et al., 2017;Abdel-Latef et al., 2021). ...
Harnessing endophytic microbes as bioinoculants promises to solve agricultural problems and improve crop yield. Out of fifty endophytic bacteria of sunflowers, 20 were selected based on plant growth-promoting. These plant growth-promoting bacteria were identified as Bacillus, Pseudomonas, and Stenotrophomonas. The qualitative screening showed bacterial ability to produce hydrogen cyanide, ammonia, siderophore, indole-3-acetic acid (IAA), exopolysaccharide, and solubilize phosphate. The high quantity of siderophore produced by B. cereus T4S was 87.73%. No significant difference was observed in the Bacillus sp. CAL14 (33.83%), S. indicatrix BOVIS40 (32.81%), S. maltophilia JVB5 (32.20%), S. maltophilia PK60 (33.48%), B. subtilis VS52 (33.43%), and P. saponiphilia J4R (33.24%), exhibiting high phosphate-solubilizing potential. S. indicatrix BOVIS40, B. thuringiensis SFL02, B. cereus SFR35, B. cereus BLBS20, and B. albus TSN29 showed high potential for the screened enzymes. Varied IAA production was recorded under optimized conditions. The medium amended with yeast extract yielded high IAA production of 46.43 µg/ml by S. indicatrix BOVIS40. Optimum IAA production of 23.36 and 20.72 µg/ml at 5% sucrose and 3% glucose by S. maltophilia JVB5 and B. cereus T4S were recorded. At pH 7, maximum IAA production of 25.36 µg/ml was obtained by S. indicatrix BOVIS40. All the isolates exhibited high IAA production at temperatures 25, 30, and 37 • C. The in vitro seed inoculation enhanced sunflower seedlings compared to the control. Therefore, exploration of copious endophytic bacteria as bioinoculants can best be promising to boost sunflower cultivation.
... This species is occasionally isolated as an endophyte of banana (Andrade et al., 2014), tomato (Bhattacharya et al., 2019), prairie berry (Abdallah et al., 2016), citrus (Daungfu et al., 2019) and medicinal plants like Aloe vera (Akinsanya et al., 2019), Dahurian angelica (Li et al., 2018), Fagonia indica (Rahman et al., 2017). This bacterium was also reported as a passenger endophyte of groundnut nodules (Preyanga et al., 2021). The occurrence of Bacillus tequilensis was also reported in other ecological niches like 2000-years old shaft tomb (Gatson et al., 2006), mangrove soil (Sharifi & Kunchirman, 2021), saline water pond (Cortés-Camargo et al., 2021), geothermal hot spring (Fachrial et al., 2020), soil crust (Zhao et al., 2019), coastal soil (Pathak et al., 2014), fermented goat milk (Abid et al., 2019), fermented soybean (Chaurasia et al., 2020), Thai milk kefir (Vijitra et al., 2018) and chicken gastrointestinal tract (Rani et al., 2017). ...
Aim:
The plant-growth-promoting putative competitive endophytes offer significant benefits to sustainable agriculture. The unworthy opportunistic and passenger endophytes are inevitable during the isolation of putative competitive endophytes. This study aimed to discriminate the putative competitive endophytes undoubtedly from the opportunistic and passenger endophytes.
Methods and results:
The newly-isolated endophytes from field-grown rice were inoculated to 5-days old rice seedlings under gnotobiotic conditions. Re-isolation of the inoculated strains from the root surface, inner tissues of the whole plant, root, and shoot was performed after 5-days. All the re-isolated colonies were compared with native isolate for the homology by BOX-A1R-based repetitive extragenic palindromic-PCR (BOX-PCR) and enterobacterial repetitive intergenic consensus (ERIC-PCR) DNA fingerprints. The results revealed that the putative competitive endophyte (RE25 and RE10) showed positive for re-isolation and BOX and ERIC fingerprints for the whole plant, root, and shoot. The opportunistic (RE27 and RE8) and passenger endophytes (RE44 and RE18) failed in re-isolation either from root or shoot. The epiphytes (ZSB15 and Az204) showed negative for endophytic re-isolation and positive for surface colonization.
Conclusion:
This modified procedure can discriminate the putative competitive endophytes from others.
Significance and impact of the study:
Eliminating the opportunistic and passenger endophytes and epiphytes early by this method would help develop endophytic inoculants to enhance rice productivity.
... A single endophytic bacterial strain may have more than one of these plant-growthpromoting traits (e.g. [37, 41, 48, [49,55] in Table 1, and [56,57,63,65,66,68] in Table 2). ...
Plant-associated microbiomes confer fitness advantages to the plant host by growth promotion through different mechanisms including nutrient uptake, phytohormones production, resistance to pathogens, and stress tolerance. These effects of the potentially beneficial microbes have been used in a diversity of biotechnological approaches to improve crop performance applying individual bacterial cultures. However, healthy plants host a diversity of microorganisms (microbiota). Next-generation sequencing technologies have offered insights into the relative abundances of different phylogenetic groups in a community and the metabolic and physiological potential of its members. In the last decade, researchers have started to explore the possibilities to use temporal and functional combinations of those bacteria in the form of synthetic communities. In this chapter, we review the benefits of using endophytic bacteria in legumes, the available methodological approaches to study the effects of bacterial communities, and the most recent findings using synthetic communities to improve the performance of legume crops.
... However, the effect of passenger endophytes invasion on nodular activity and plant growth was only investigated before nodule formation. Indeed, many studies have revealed the beneficial effect of passenger endophytes co-inoculated with native rhizobium on nodule activity in terms of nodule leghemoglobin content, nitrogenase activity and plant growth Subramanian et al., 2015;Preyanga et al., 2021). Chihaoui et al. ...
Agrobacterium species are one of the most frequently isolated endophytes from root nodules of a wide range of wild and cultivated legumes. Many studies have investigated the role of endophytes in phytoprotection and phytoremediation enhancement. However, their biological significance and their implication in nodule functioning have been scarcely studied, especially, the effect of the inoculation time on nodule activity and plant growth which still unknown. Accordingly, in this study, nodule-endophytic Agrobacterium sp.10C2 was inoculated before and after nodule formation, to determine the effect of the inoculation time on the plant growth and nodule functioning in the reference line Jemalong A17 of the model legume Medicago truncatula in symbiotic association with Sinorhizobium meliloti TII7.
The results revealed that nodule functioning was strongly influenced by the inoculation time of the endophytic Agrobacterium sp.10C2. In fact, the early inoculation with Agrobacterium sp.10C2 induced an increase of the highest nodules nitrogenase activity, recorded at the day 66 after the inoculation, by 66%, the leghemoglobin concentration by 26% and the total sugar content by 67%, in addition to a higher photosynthesis activity and biomass plant production compared to the control. Interestingly, when inoculation occurred after nodule formation, nodules have induced a defense response against Agrobacterium sp. 10C2 evidenced by an overproduction of Guaicol peroxidase which increased from 1,04 μmol H2O2 min⁻¹ mg⁻¹ proteins in the control to 2,36 μmol H2O2 min⁻¹ mg⁻¹ proteins in the late inoculated plants, leading to a loss of the beneficial impact of the endophytic Agrobacterium on both biomass production and nodule functioning parameters.
The finding of this study showed for the first time that nodule responses to endophytic agrobacteria change according to the time of inoculation. When inoculated at early stage, 10C2 plays the role of a Plant Growth Promoting Rhizobacteria (PGPR), however, after nodule formation, 10C2 could act as a pathogen agent. Thus, the inoculation time could be considered as a crucial parameter for the nodule functioning and plant growth.
... That the internal environments of oleaginous plants provide unique ecological habitats for the growth and sustenance of PHA-accumulating bacteria [12] has been well justified from our laboratory [13,14]. The peanut plant (Arachis hypogaea L.), the fourth most important edible oil source of the world [15] has been investigated quite extensively and recognized as a treasured microbial repository [16,17]. The seed endophytes, in particular, have gained significant interest as they are transmitted vertically and ensure their presence in progeny plants [18,19]. ...
Endophytic bacteria, which thrive inside the plants, synthesize and accumulate intracellular biopolymer polyhydroxyalkanoates (PHAs) to strive against the hostile plant environment. The present study is focused on enhancing the production of poly(3-hydroxybutyrate) [P(3HB)], the most common PHA by the seed endophytic bacterium Bacillus pumilus AHSD 04 (GenBank accession number KY038573; MCC accession number 3573) isolated from the oleaginous plant Arachis hypogaea L. Interactions among the three most important influencing variables, glucose, tryptose, and initial pH affecting growth and P(3HB) production were studied using the central composite design (CCD) of response surface methodology (RSM). A 3.94-fold enhancement of P(3HB) production (5.36 g/L) was achieved over that of the 'one variable at a time' method. The isolate also produced co-polymers of 3-hydroxybutyrate (3HB) and 3-hydroxyvalerate (3HV) [P(3HB-co-3HV)] with the incorporation of 4.98 mol% 3HV during dual-step cultivation using glucose and valeric acid as co-substrate. The polymers, P(3HB) and P(3HB-co-3HV), so produced have been validated and characterized by Fourier transform infrared (FTIR) and 1H nuclear magnetic resonance (1H NMR) spectroscopic analysis. It is apparent that RSM has been successfully established as a promising tool to increase P(3HB) yield by the seed endophyte B. pumilus AHSD 04, which can further be utilized to scale up the production of these biodegradable polymers of industrial importance.
... In the field of marine agriculture, most research on growth promotion of mangrove has concentrated on PGPB from the rhizosphere (Bashan and Holguin, 2002;Kathiresan and Selvam, 2006;Youssef, 2010, 2011). Increasingly, endophytic bacteria have great attributes because they are less exposed to inhospitable environments of the soil and are located in living tissues where relevant activities occur (El-Tarabily et al., 2009, thus making them ideal for the next generation of PGP agents (Preyanga et al., 2021). Except of one report on endophytic PGPA to promote S. bigelovii growth under greenhouse conditions , the effect of applications of endophytic bacteria or fungi to promote growth of mangrove or other marine plants in the greenhouse/nursery/field is mostly negligible. ...
Mangrove forests in the Arabian Gulf are under continuous threat. To increase plantations of gray mangrove (Avicennia marina) in the United Arab Emirates (UAE), 27 endophytic actinobacterial isolates obtained from mangrove roots were in vitro pre-screened to detect the polyamines (PAs) [putrescine (Put), spermidine (Spd), and spermine (Spm)]. We also determined the abilities of the endophytic PA-producing actinobacterial isolates in enhancing the growth of A. marina under greenhouse conditions. Although three highly PA-producing isolates were recovered from inside mangrove root tissues, Streptomyces mutabilis UAE1 constantly colonized root and stem inner tissues for 12 weeks, suggesting an endophytic association between this actinobacterial isolate and mangrove seedlings. When roots were inoculated with S. mutabilis, mangrove growth was remarkably enhanced under gnotobiotic and greenhouse conditions. This was evident from the significant (P < 0.05) increases in dry weight and length of root (66.7 and 65.5%, respectively) and shoot (64.8 and 58.0%, respectively), number of branches (64.3%), total leaf area (40.2%), and photosynthetic pigments (54.5% chlorophyll a; 40.0% chlorophyll b; and 53.1% carotenoids) of mangrove compared to the PA-non-producing Streptomyces sp. or control treatment. Growth promotion in plants treated with S. mutabilis was also supported by significant (P < 0.05) elevations in the contents of mangrove in planta PAs, auxins, and cytokinins, accompanied by a decrease in abscisic acid levels. No difference was, however, detected in growth and amounts of PAs or any plant growth regulators (PGRs) in Frontiers in Marine Science | www.frontiersin.org 1 July 2021 | Volume 8 | Article 710200 El-Tarabily et al. Endophytic PGP Actinobacteria From Mangrove plants treated with or without the PA-non-producing isolate. Our findings indicate that plant growth parameters can be enhanced as a consequence of secretion of Put, Spd, and Spm by S. mutabilis resulting in regulation of endogenous PAs and other PGRs in mangrove tissues. This study is the first record that aims to improve sustainable agricultural management practices using plant growth promoting (PGP) actinobacteria, endophytic in mangrove tissues to promote mangrove growth under greenhouse conditions. Such research may allow this region to be a model to study the synergistic S. mutabilis-mangrove interactions and the future impacts on mangrove reforestation in the Arabian Gulf and elsewhere where mangrove forests are in threat.
EANUT plants can obtain N from N 2 fixation via symbiosis with rhizobia, and inoculation with selected strains can improve grain yields. The aim of this work is to conduct field trials to compare single inoculation with Bradyrhizobium sp., Anabaena circinalis, and A. variabilis with dual inoculation during the 2021 and 2022 seasons in order to verify whether microbial inoculants may enhance peanut performance, growth, and grain yield as a result of their advantageous effects. The findings demonstrated that T8 treatment (dual inoculation with Bradyrhizobium sp, A. circinalis and A. variabilis and (1:1:1) + 25% N + 100% PK), generally improved peanut plant growth, resulting in significantly higher chlorophylls (mg g-1 FW), carotenoids (µg g-1 FW), number of nodules and dry weight of nodules (mg plant-1). In addition, improved soil fertility by increasing dehydrogenase and CO 2 evolution, in the peanut rhizosphere during the two growing seasons. On the other hand, the percentages of N, P, and K in peanut plant leaves were affected by various inoculations in ways that were statistically significant (P ≤ 0.05), which were arranged by T8 > T6 > T7 > T5 for dual inoculation treatments and T2 > T4 > T3 > T1 for single inoculation treatments. The same pattern was observed for microelements (Fe, Cu, Mn and Zn). This was reflected in the yield (plant height, pod number, pod weight, yield, and 100-weight seeds), and the quality of the grain (percentage of oil, carbohydrates and protein). According to the current study, cyanobacteria and Bradyrhizobium are helpful in enhancing peanut plant development, physiological changes, biological performance, productivity, and seed quality.
Endophytes colonize interior regions of oilseeds depending on water, nutrient, space availability and microbial competition. Oilseeds are filled with oil globules making it difficult for microbes to obtain direct uptake of water and nutrient for growing into colonies. Recent research shows very less areas explored in endophytic study in oilseeds and as endophytes play a vital part in managing diseases in plants by releasing genes encoding jasmonate and salicylic acid which induces defense against pathogens and modify plant regions structurally adding to their fitness and increased productivity, their study is indispensable. This review focuses on the bacterial endophytes in oilseeds, the challenges faced in their isolation, their colonization and interactions with host and pathogens and their applications. Oilseed endophytic study is essential for understanding the benefits of oilseeds consumed as vegetable oil or used as fuel by people all over the world.
The regulation of legume‐rhizobia symbiosis by microorganisms has obtained considerable interest in recent research, particularly in the common rhizobacteria Bacillus . However, few studies have provided detailed explanations regarding the regulatory mechanisms involved. Here, we investigated the effects of Bacillus (Bac.B) on Bradyrhizobium– soybean ( Glycine max ) symbiosis and elucidated the underlying ecological mechanisms. We found that two Bradyrhizobium strains (i.e. Bra.Q2 and Bra.D) isolated from nodules significantly promoted nitrogen (N) efficiency of soybean via facilitating nodule formation, thereby enhanced plant growth and yield. However, the intrusion of Bac.B caused a reverse shift in the synergistic efficiency of N 2 fixation in the soybean –Bradyrhizobium symbiosis. Biofilm formation and naringenin may be importantin suppression of Bra.Q2 growth regulated by Bac.B. In addition, transcriptome and microbiome analyses revealed that Bra.Q2 and Bac.B might interact to regulateN transport and assimilation, thus influence the bacterial composition related to plant N nutrition in nodules. Also, the metabolisms of secondary metabolites and hormones associated with plant–microbe interaction and growth regulation were modulated by Bra.Q2 and Bac.B coinoculation. Collectively, we demonstrate that Bacillus negatively affects Bradyrhizobium– soybean symbiosis and modulate microbial interactions in the nodule. Our findings highlight a novel Bacillus ‐based regulation to improve N efficiency and sustainable agricultural development.
Legumes are renowned for their distinctive biological characteristic of forming symbiotic associations with soil bacteria, mostly belonging to the Rhizobiaceae familiy, leading to the establishment of symbiotic root nodules. Within these nodules, rhizobia play a pivotal role in converting atmospheric nitrogen into a plant-assimilable form. However, it has been discerned that root nodules of legumes are not exclusively inhabited by rhizobia; non-rhizobial endophytic bacteria also reside within them, yet their functions remain incompletely elucidated. This comprehensive review synthesizes available data, revealing that Bacillus and Pseudomonas are the most prevalent genera of nodule endophytic bacteria, succeeded by Paenibacillus, Enterobacter, Pantoea, Agrobacterium, and Microbacterium. To date, the bibliographic data available show that Glycine max followed by Vigna radiata, Phaseolus vulgaris and Lens culinaris are the main hosts for nodule endophytic bacteria. Clustering analysis consistently supports the prevalence of Bacillus and Pseudomonas as the most abundant nodule endophytic bacteria, alongside Paenibacillus, Agrobacterium, and Enterobacter. Although non-rhizobial populations within nodules do not induce nodule formation, their presence is associated with various plant growth-promoting properties (PGPs). These properties are known to mediate important mechanisms such as phytostimulation, biofertilization, biocontrol, and stress tolerance, emphasizing the multifaceted roles of nodule endophytes. Importantly, interactions between non-rhizobia and rhizobia within nodules may exert influence on their leguminous host plants. This is particularly shown by co-inoculation of legumes with both types of bacteria, in which synergistic effects on plant growth, yield, and nodulation are often measured. Moreover these effects are pronounced under both stress and non-stress conditions, surpassing the impact of single inoculations with rhizobia alone.
The present study was carried out to assess the growth-promoting ability of non-rhizobial endophytes in groundnut (Arachis hypogaea). Thirteen endophytic bacteria with different morphologies were isolated from the root and nodules of groundnut. These isolates significantly enhanced the growth of groundnut in sterilised vermiculite, though the isolates were unable to nodulate the host plant. The endophytic nature of these isolates was confirmed by their re-isolation from the sterilised and macerated roots of the plants. The isolates exhibited in vitro tricalcium phosphate and zinc solubilization, production of siderophores, auxins and ammonia as well as growth on different nitrogen-free media. The phosphate solubilization and auxin production varied from 50 to 196 and 17 to 71 µg/ml, respectively by the isolates. Based on phenotypic tests and 16S rRNA gene sequencing, four potential strains were identified as Klebsiella sp. R3, Pseudomonas putida R6, Klebsiella oxytoca GRE5 and Pseudomonas proteolytica GRE6. A significant increase in plant growth, chlorophyll content, nodule count and shoot nutrient content of groundnut was observed with these bacterial inoculations over the uninoculated control in greenhouse. The bacterial treatments resulted in increased N, P and K content in the shoot up to 87, 96 and 44%, respectively, over the control. Physico-chemical properties and available nutrient content of soil were also improved on bacterial inoculations. The results indicated that groundnut harbours beneficial non-rhizobial bacterial endophytes with the potential to be used as microbial inoculants in groundnut. Klebsiella oxytoca as a non-nodulating nodule endophyte of groundnut is reported for the first time.
Legumes improve soil fertility by interacting symbiotically with nitrogen-fixing rhizobia allocated in root nodules. Some bacterial endophytes can coexist with rhizobia in nodules and might help legumes by enhancing stress tolerance, producing hormones stimulating plant growth, and increasing plant nutrient intake. Twenty-six bacterial endophytes from Lens culinaris root nodules cultivated in intercropping with Triticum durum were identified and characterized molecularly and biochemically. Potential plant growth-promoting strains have been selected according to the indole acetic acid and 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase production, and for their inorganic phosphate solubilization ability. The presence of genes associated to ACC deaminase and nitrogenase was evaluated. Six selected strains were grown with varying NaCl and polyethylene glycol concentrations to test their salt and osmotic stress tolerance. Priestia megaterium 11NL3 and Priestia aryabhattai 19NL1, resulted to be tolerant to salinity and osmotic stress, were tested on four genotypes of T. durum seeds in different stress conditions. The effect of strain inoculation on seed germination, vigor, and root-to-shoot ratio varied depending on the type of stress and on the durum wheat genotypes. For future research, it will be necessary to test the selected bacterial strains at different plant phenological stages and to clarify the mechanisms involved in the different outcomes of plant-microbe interactions.
The ability of endophytes to confer plant stress resistance opens a new avenue for mitigating the negative impact of various stresses on crop production. Consequently, the purpose of this study was to investigate plant growth-promoting nodule endophytes for alleviating salt stress in groundnut. Under salt-stressed conditions, rhizobial isolates (Rhizobium phaseoli S18 and Rhizobium pusense S6R2) and passenger endophytes (Pantoea dispersa YBB19B and Bacillus tequilensis NBB13) exhibited multiple plant growth-promoting traits, including IAA, siderophore production, nutrient solubilisation, and ACC deaminase, and compared to the control, the co-inoculation of Rhizobium pusense S6R2 and Bacillus tequilensis NBB13 produced the highest number of nodules (18 per plant) in saline soil. Analysis of root exudates revealed the presence of acetic acid, phenol, and succinic acid compounds in groundnut treated with Rhizobium pusense S6R2 + Pantoea dispersa YBB19B might have resulted in an enhanced stress tolerance mechanism in groundnut. In a pot culture experiment, the combined application of Rhizobium pusense S6R2 and Pantoea dispersa YBB19B or Bacillus tequilensis NBB13 increased plant growth, pod yield, production of anti-oxidant enzymes such as catalase and ascorbate peroxidase, and resulted in accumulation of osmolytes. Similarly, in the field experiment, the co-inoculation of Rhizobium pusense S6R2 and Pantoea dispersa YBB19B resulted in the highest groundnut yield in comparison to the other treatments. This study demonstrated that plants co-inoculated with Rhizobium pusense S6R2 and Pantoea dispersa YBB19B alleviated salt stress; consequently, they may be recommended as a bio-inoculant for salt stress alleviation in groundnut following field evaluation at multi locations.
Groundnut can obtain N from the N 2 fixation in the symbiosis with rhizobia and inoculation with selected strains can improve grain yields. We report the results from four field experiments, aiming to verify if microbial inoculants may improve groundnut performance, through the effects of single inoculation with Bradyrhizobium sp. (SEMIA6144), of co-inoculation Arthrospira platensis IPR7059 or Synechocystis sp. IPR7061, and of the N fertilization with 100 kg ha − 1 of N on plant growth, nodulation, N accumulated in tissues, grain protein, and grain yield. There were no effects of inoculation treatments and N-fertilizer on shoot and root dry weights. In clayey soil, co-inoculation with Bradyrhizobium sp. and cyanobacteria increased grain productivity by an average of 19% compared to the non-inoculated control. In this clayey soil with higher P content, regardless of whether co-inoculated with Bradyrhizobium sp. and cyanobacteria or single inoculated, grain productivity was 16% higher on average compared to nitrogen fertilizer. In conclusion, it was clear that success of rhizobia inoculation in groundnut is dependent on the soil, probably due to P limitation, and weather conditions.
This review evaluates oilseed crop soybean endophytic bacteria, their prospects, and challenges for sustainable agriculture. Soybean is one of the most important oilseed crops with about 20–25% protein content and 20% edible oil production. The ability of soybean root-associated microbes to restore soil nutrients enhances crop yield. Naturally, the soybean root endosphere harbors root nodule bacteria, and endophytic bacteria, which help increase the nitrogen pool and reclamation of another nutrient loss in the soil for plant nutrition. Endophytic bacteria can sustain plant growth and health by exhibiting antibiosis against phytopathogens, production of enzymes, phytohormone biosynthesis, organic acids, and secondary metabolite secretions. Considerable effort in the agricultural industry is focused on multifunctional concepts and bioprospecting on the use of bioinput from endophytic microbes to ensure a stable ecosystem. Bioprospecting in the case of this review is a systemic overview of the biorational approach to harness beneficial plant-associated microbes to ensure food security in the future. Progress in this endeavor is limited by available techniques. The use of molecular techniques in unraveling the functions of soybean endophytic bacteria can explore their use in integrated organic farming. Our review brings to light the endophytic microbial dynamics of soybeans and current status of plant microbiome research for sustainable agriculture.
Low soil phosphorus (P) availability is a notable limitation for legume nodulation, nitrogen fixation and yield, and it is therefore important to improve legume growth under P-deficient conditions. This study aimed to determine the effects of coinoculation with Rhizobium and plant growth promoting rhizobacter (PGPR) on the nitrogen fixation and nutrient uptake of white clover (Trifolium repens) in a P-deficient soil. White clover plants were inoculated with one of the two Rhizobium strains CHB1120 and CHB1121 alone or coinoculated with either of the Rhizobium strains and one of the two PGPR strains, Bacillus aryabhattai strain Sb and Azotobacter vinelandii strain G31. White clover inoculated with either CHB1120 or CHB1121 alone and coinoculated with one of the two Rhizobium strains and the PGPR significantly enhanced nodule numbers compared with the non-inoculated control. The coinoculation of either CHB1120 or CHB1121 and Sb showed a significantly greater level of nitrogenase than the single inoculation of CHB1120 or CHB1121. In addition, the single inoculation of CHB1120 or CHB1121 and their coinoculation with PGPR significantly increased nitrogen contents in both shoots and roots under P-deficient conditions. The coinoculation of CHB1121 and G31 showed the greatest N content in shoots. Moreover, the single inoculation of the two Rhizobium strains and their coinoculation with G31 and Sb significantly increased the K, Ca, and Mg contents in shoots compared with the non-inoculated control. In summary, the PGPR used in this study can enhance the symbiotic potential of rhizobia under low P conditions. ARTICLE HISTORY
Common bean is a poor symbiotic N-fixer, with a low response to inoculation owing to its promiscuous nodulation with competitive but inefficient resident rhizobia. Consequently, farmers prefer to fertilize them rather than rely on their capacity for Biological Nitrogen Fixation (BNF). However, when rhizobial inoculants are based on autochthonous strains, they often have superior BNF performance in the field due to their genetic adaptations to the local environment. Nevertheless, there is scant information at the genomic level explaining their superiority or on how their genomes may influence the inoculant performance. This information is especially important in technologically advanced agri-systems like Europe, where environmental concerns and increasingly stringent fertilizer regulations are encouraging a return to the use of rhizobial inoculants, but based upon strains that have been thoroughly characterized in terms of their symbiotic performance and their genetics. The aim of this study was to design an inoculant formulation based on a superior autochthonous strain, Rhizobium leguminosarum bv. phaseoli LCS0306, to assess its performance in the field, and to determine the genomic features contributing to the high effectiveness of its symbiosis with common bean. Plants inoculated with the autochthonous strain LCS0306 fixed significantly more nitrogen than those with the allochthonous strains R. phaseoli ATCC 14482T and R. etli CFN42T, and had grain yield similar to the nitrogen-fertilized controls. Inoculation with LCS0306 was particularly efficacious when formulated with a carrier based upon a mixture of perlite and biochar. Whole genome comparisons revealed no differences in the classical symbiotic genes of strain LCS0306 within the symbiovar phaseoli. However, its symbiotic superior performance might be due to its genomic versatility, as it harbors a large assortment of genes contributing to fitness and competitiveness. It is concluded that inoculation with elite rhizobia formulated with perlite-biochar carriers might constitute a step-change in the sustainable cultivation of common bean in Spanish soils.
Endophytes are rich in plant tissues and play important roles in plant-microbial interactions and plant-growth regulation. Here, endophytic bacteria from two closely related tea cultivars of Zijuan and Yunkang-10 were isolated, and the diversities were compared. Plant-growth promoting (PGP) activities were determined on the dominant groups or nitrogen-fixing genera from the two cultivars. Endophytic bacteria were isolated by using of different selective media and methods, and the PGP activities were investigated by analytical and molecular technologies. A total of 110 isolates of 18 genera belonging to three phylums (Proteobacteria, Firmicutes, and Bacteroidetes) were obtained from Zijuan, while 164 isolates of 22 genera belonging to two phylums (Proteobacteria and Firmicutes) were obtained from Yunkang-10. PGP screening indicated that Herbaspirillum spp., Methylobacterium spp., and Brevundimonas spp. showed different PGP abilities. The PGP ability decreased in order of Herbaspirillum spp., Brevundimonas spp. and Methylobacterium spp., and the majority of Methylobacterium spp. did not showed PGP activity of nitrogen-fixation, P-solubilization, siderophore, indole-3-acetic acid (IAA) production or 1-aminocyclopropane-1-carboxylate (ACC) deaminase. The study of bacterial community and PGP activities confirmed that endophytes in tea plants are constantly changing in different seasons and tea cultivars, and the PGP bacteria in Zijuan are much abundant than those of Yunkang-10.
This study aimed at documenting the culturable methylotrophic bacterial diversity across different groundnut genotypes and evaluating their effect on the growth of groundnut. 80 methylotrophic bacterial isolates were obtained from the phyllosphere of 15 groundnut genotypes collected from Tamil Nadu, India. The bacterial isolates were identified through sequencing of the 16S rDNA and were tested for their plant growth-promoting properties. Groundnut seeds were inoculated with methylotrophic bacteria and their effect on growth was evaluated via in vitro and pot experiments. Molecular identification revealed that the isolates belonged to 30 different species. A higher diversity of methylotrophic bacteria at genus and species level was found in groundnut genotype TMV2. Shannon diversity index was the highest in genotype TMV7, followed by VRI2 and TMV2. Similarly, geographical location also influenced the diversity of methylotrophic bacteria. In vitro seed germination assay revealed that methylotrophic isolates enhanced root growth and improved formation of root hair. The radicle length of treated seeds ranged from 2.7 to 8.4 cm. A higher shoot length was observed in the plants from seeds treated with Methylobacterium radiotolerans VRI8-A4 (27.3 cm), followed by Pseudomonas psychrotolerans TMV13-A1 (26.3 cm) and Bacillus aryabhattai K-CO3-3 (23 cm). The findings of this study strongly suggest that beneficial methylotrophic bacteria associated with the phyllosphere of groundnut play a major role in regulating plant growth.
Aspalathus linearis (burm f.), commonly known as rooibos, grows in nutrient and organic matter poor sandy soils that limit its growth. In this study, samples of nodules from both declined and healthy rooibos plants were collected to determine the frequency of nodule nitrogen-fixing and endophytic bacteria. Standard microbiological procedures as well as sequence analysis of the 16S rRNA revealed that more than 75% of the bacterial isolates from the healthy plants contained microsymbionts belonging to the Rhizobium group and the remaining 25% were characterized as Pseudomonas and Burkholderia spp. The nodule from the declined plants lacks a sufficient number of rhizobia and was mostly white in color, small and contains the free-living endospore-forming Bacillus and other endophytic Burkholderia and Pseudomonas spp. The results provide a baseline data on the microsymbionts of rooibos nodules in Citrusdal and highlighted the need for further investigation using additional techniques.
Rhizobia induce root nodules and fix atmospheric N2 for most legume species in exchange for carbon. However, the diverse endophytic non-rhizobial bacteria in legume nodules that co-exist with rhizobia are often ignored because they are difficult to cultivate using routine cultivation approaches. To enhance our understanding of the incidence and diversity of legume–bacteria associations, a high-throughput sequencing analysis of bacterial 16S rRNA genes was used to examine the bacterial community in the rhizospheres and root nodules of Dalbergia odorifera seedlings that were uninoculated or inoculated with Bradyrhizobium elkanii H255, Rhizobium multihospitium–like HT221, or Burkholderia pyrrocinia–like H022238, in two growth media (nitrogen [N]-supplied soil or N-omitted potting mix). Seedlings inoculated with Bradyrhizobium had significantly more nodules than seedlings in the other inoculation conditions, regardless of growth media. Using the ¹⁵N natural abundance method, it was shown that the inoculated plants had significantly higher N2 fixation efficiency (48–57%) and specific nodule activity [269–313 μg N mg⁻¹ of dry weight (dwt) nodule] compared to the uninoculated plants (203 μg N mg⁻¹ dwt nodule). The 16S rRNA gene analysis showed that there was generally a higher bacterial diversity in the rhizosphere than in the nodules in the corresponding condition. Both rhizobial inoculation and media status significantly altered the bacterial communities in the rhizospheres and nodules (P < 0.05), with the exception of the inoculated soil rhizospheres. Regarding non-rhizobial bacteria, three genera, i.e., Lactococcus, Bacillus, and Pseudomonas, were consistently enriched in the rhizosphere and Bradyrhizobium, Chloroplast norank (which belongs to Cyanobacteria), and Lactococcus were commonly found in the nodules. In contrast, common rhizobial genera (including Rhizobium, Mesorhizobium, and Burkholderia) were only present in the nodules at low relative abundances (0.01–3.41%). Regarding non-rhizobial bacteria, 32 genera were found in the nodules, with non-rhizobial bacteria being predominant in the N omitted potting mix (with a relative abundance of 56–87%). This study suggests that legume nodules are inhabited by a high diversity of non-rhizobial bacteria, which may play a vital role in nodulation and N2 fixation in the host plants.
The discovery that the actinobacterium Micromonospora inhabits nitrogen-fixing nodules raised questions as to its potential ecological role. The capacity of two Micromonospora strains to infect legumes other than their original host, Lupinus angustifolius, was investigated using Medicago and Trifolium as test plants. Compatible rhizobial strains were used for coinoculation of the plants because Micromonospora itself does not induce nodulation. Over 50% of nodules from each legume housed Micromonospora, and using 16S rRNA gene sequence identification, we verified that the reisolated
strains corresponded to the microorganisms inoculated. Entry of the bacteria and colonization of the plant hosts were monitored using a GFP-tagged Lupac 08 mutant together with rhizobia, and by using
immunogold labeling. Strain Lupac 08 was localized in plant tissues, confirming its capacity to enter and colonize all hosts. Based on studying three different plants, our results support a non-specific relationship between Micromonospora and legumes. Micromonospora Lupac 08, originally isolated from Lupinus re-enters root tissue, but only when coinoculated with the corresponding rhizobia. The ability of Micromonospora to infect and colonize different legume species and function as a potential plant-growth promoting bacterium is relevant because this microbe enhances the symbiosis without interfering with the host and its nodulating and nitrogen-fixing microbes.
Nitrogen (N) fixation through legume-Rhizobium symbiosis is important for enhancing agricultural productivity and is therefore of great economic interest. Growing evidence indicates that other soil beneficial bacteria can positively affect symbiotic performance of rhizobia. Nodule endophytic plant growth promoting rhizobacteria (PGPR) were isolated from common bean nodules from Nakuru County in Kenya and characterized 16S rDNA partial gene sequencing. The effect of co-inoculation of rhizobium and PGPR, on nodulation and growth of common bean (Phaseolus vulgaris L.) was also investigated using a low phosphorous soil under greenhouse conditions. Gram-positive nodule endophytic PGPR belonging to the genus Bacillus were successfully isolated and characterized. Two PGPR strains (Paenibacillus polymyxa and Bacillus megaterium), two rhizobia strains (IITA-PAU 987 and IITA-PAU 983) and one reference rhizobia strain (CIAT 899) were used in the co-inoculation study. Two common bean varieties were inoculated with Rhizobium strains singly or in a combination with PGPR to evaluate the effect on nodulation and growth parameters. Co-inoculation of IITA-PAU 987 + B. megaterium recorded the highest nodule weight (405.2 mg) compared to IITA-PAU 987 alone (324.8 mg), while CIAT 899 + B. megaterium (401.2 mg) compared to CIAT 899 alone (337.2 mg). CIAT 899 + B. megaterium recorded a significantly higher shoot dry weight (7.23 g) compared to CIAT 899 alone (5.80 g). However, there was no significant difference between CIAT 899 + P. polymyxa and CIAT 899 alone. Combination of IITA-PAU 987 and B. megaterium led to significantly higher shoot dry weight (6.84 g) compared to IITA-PAU 987 alone (5.32 g) but no significant difference was observed when co-inoculated with P. polymyxa. IITA-PAU 983 in combination with P. polymyxa led to significantly higher shoot dry weight (7.15 g) compared to IITA-PAU 983 alone (5.14 g). Plants inoculated with IITA-PAU 987 and B. megaterium received 24.0 % of their nitrogen demand from atmosphere, which showed a 31.1% increase compared to rhizobium alone. Contrast analysis confirmed the difference between the co-inoculation of rhizobia strains and PGPR compared to single rhizobia inoculation on the root dry weight. These results show that co-inoculation of PGPR and Rhizobia has a synergistic effect on bean growth. Use of PGPR may improve effectiveness of Rhizobium biofertilizers for common bean production. Testing of PGPR under field conditions will further elucidate their effectiveness on grain yields of common bean.
The recent advent of DNA sequencing technologies facilitates the use of genome sequencing data that provide means for more informative and precise classification and identification of Bacteria and Archaea. Because the current species definition is based on the comparison of genome sequences between type and other strains in a given species, building a genome database with correct taxonomic information is a premium need to enhance our efforts in exploring prokaryotic diversity and discovering new species as well as for routine identifications. Here we introduce an integrated database, called EzBioCloud, that holds the taxonomic hierarchy of Bacteria and Archaea that are represented by quality-controlled 16S rRNA gene and genome sequences. Whole genome assemblies in the NCBI Assembly Database were screened for low quality and subjected to a composite identification bioinformatics pipeline that employs gene-based searches followed by the calculation of average nucleotide identity. As a result, the database is made of 61,700 species/phylotypes including 13,132 with validly published names, and 62,362 whole genome assemblies that were taxonomically identified at the genus, species and subspecies levels. Genomic properties, such as genome size and GC content, and the occurrence in human microbiome data were calculated for each genus or higher taxa. This united database of taxonomy, 16S rRNA gene and genome sequences, with accompanied bioinformatics tools, should accelerate genome-based classification and identification of Bacteria and Archaea. The database and related search tools are available at http://www.ezbiocloud.net/.
Phosphorus is one of the three macronutrients that are essential for plant growth and development. Inorganic phosphorus (P), which can make up to 70% of the total P content in soils, can exist in calcium-, aluminum-, or iron-complexed forms that are unavailable for plant use. As a result, mineral phosphorus, P2O5, is often used as a fertilizer to supplement the nutrient for crop growth. To reduce the addition of mineral phosphorus to agricultural soils, research in naturally occurring phosphate-solubilizing microorganisms has been conducted for decades. This study found bacteria that solubilized phosphate at very high rates. The most efficient of the bacteria presented in this paper, Pantoea sp. Pot1, can solubilize tricalcium phosphate (Ca3(PO4)2) at a rate of 956 mgL-1. This bacteria produces a variety of organic acids, including acetic, gluconic, formic, and propionic acids. Greenhouse experiments demonstrated that tomato plants with soil systems inoculated with Pantoea sp. Pot1 incorporated more P and produced much higher biomass weights than those plants without any added bacteria.
Five species of Acacia (Acacia ehrenbergiana Hayne, A. nilotica (L.) Delile, A. seyal Delile, A. tortilis(Forssk.) Hayne and A. laeta Delile) indigenous to Tamanrasset (Algeria) were investigated for theirnodulation status and nodular endophytic diversity. A. ehrenbergiana showed the highest nodulationability across the different sites in this region, indicating the widespread occurrence of compatiblerhizobia in the soils. Altogether 81 strains were purified. Among this endophytic strain collection, onlyfour bacterial endophytes nodulated their respective host plants. On the basis of partial 16S rDNAsequencing, they were affiliated to Ensifer sp., Ensifer teranga, Mesorhizobium sp. and Rhizobium sp.Among the 79 non- symbiotic endophytes, 24 representative strains on the basis of PCR-RFLP profileobtained with MSPI enzyme digestion were characterized. They belonged to nine genera, namely: Paenibacillus, Ochrobactrum, Stenotrophomonas, Pseudomonas, Microbacterium, Rhizobium,Agrobacterium, Brevibacillus and Advenella. The isolated nodular endophytes in this study revealed astrong tolerance profile to salinity and high temperature. Principal component analysis confirmed thatno correlation was found between bacterial tolerance to a maximum temperature of growth and soildepth of sampling. This tolerance profile was distributed over the three levels of soil depth sampling:20, 40 and 60 cm. On the other hand, there was no relationship between in vitro tolerances of rhizobialstrains to NaCl and high temperature and corresponding edaphoclimatic characteristics of the samplingsites. This study is a contribution to nodular bacterial diversity knowledge of desert African Acaciaspecies growing in preserved ecosystems.
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. Inoculation of ornamentals, forest trees, vegetables, and agricultural crops with PGPR may result in multiple effects on early-season plant growth, as seen in the enhancement of seedling germination, stand health, plant vigor, plant height, shoot weight, nutrient content of shoot tissues, early bloom, chlorophyll content, and increased nodulation in legumes. PGPR 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, sulphur, iron and copper, produce plant hormones, enhance other beneficial bacteria or fungi, control fungal and bacterial diseases and help in controlling insect pests. There has been much research interest in PGPR and there is now an increasing number of PGPR being commercialized for various crops. Several reviews have discussed specific aspects of growth promotion by PGPR. In this review, we have discussed various bacteria which act as PGPR, mechanisms and the desirable properties exhibited by them.
All plants are inhabited internally by diverse microbial communities comprising bacterial, archaeal, fungal, and protistic taxa. These microorganisms showing endophytic lifestyles play crucial roles in plant development, growth, fitness, and diversification. The increasing awareness of and information on endophytes provide insight into the complexity of the plant microbiome. The nature of plant-endophyte interactions ranges from mutualism to pathogenicity. This depends on a set of abiotic and biotic factors, including the genotypes of plants and microbes, environmental conditions, and the dynamic network of interactions within the plant biome. In this review, we address the concept of endophytism, considering the latest insights into evolution, plant ecosystem functioning, and multipartite interactions.
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The effects of rhizobial symbiosis on the growth, metal uptake, and antioxidant responses of Medicago lupulina in the presence of 200 mg kg(-1) Cu(2+) throughout different stages of symbiosis development were studied. The symbiosis with Sinorhizobium meliloti CCNWSX0020 induced an increase in plant growth and nitrogen content irrespective of the presence of Cu(2+). The total amount of Cu uptake of inoculated plants significantly increased by 34.0 and 120.4 % in shoots and roots, respectively, compared with non-inoculated plants. However, although the rhizobial symbiosis promoted Cu accumulation both in shoots and roots, the increase in roots was much higher than in shoots, thus decreasing the translocation factor and helping Cu phytostabilization. The rate of lipid peroxidation was significantly decreased in both shoots and roots of inoculated vs. non-inoculated plants when measured either 8, 13, or 18 days post-inoculation. In comparison with non-inoculated plants, the activities of superoxide dismutase and ascorbate peroxidase of shoots of inoculated plants exposed to excess Cu were significantly elevated at different stages of symbiosis development; similar increases occurred in the activities of superoxide dismutase, catalase, and glutathione reductase of inoculated roots. The symbiosis with S. meliloti CCNWSX0020 also upregulated the corresponding genes involved in antioxidant responses in the plants treated with excess Cu. The results indicated that the rhizobial symbiosis with S. meliloti CCNWSX0020 not only enhanced plant growth and metal uptake but also improved the responses of plant antioxidant defense to excess Cu stress.
In this study, we compare the variations of Margalef K and Shannon H diversity indices obtained for a fish community that suffered changes as a consequence of the impoundment of the upper Tocantins River in Goiás, Brazil. The analysis of different sites along the study area showed that the Shannon index was not sensitive to the environmental changes, whereas the Margalef K index varied significantly following the impoundment. Therefore, the Margalef K index appears to be a good diversity indicator as well as a valuable parameter for the temporal data series analysis, from changing environments and for the conservation of natural environments. Although the Margalef K index can be used as a diversity index, it can also be used to consistently indicate the speed of ecosystem evolution.
Endophytic bacteria from root and nodules of field pea being grown in CCS Haryana Agricultural University farm Hisar were isolated. A total of 15 endophytic bacteria from surface sterilized roots and 60 from the nodules of field pea were isolated. Out of 75 isolates, 67% in roots and 68% in nodules were gram positive. In roots 70% and in nodules 90% of the gram positive isolates was spore formers. Selected 9 endophytes from roots and 48 from nodules were used to determine molecular diversity by amplication of 16S rDNA by PCR, followed by RFLP by three restriction endonucleases Hae III, RsaI & Hinf I. Wide diversity among field pea bacterial endophytes was observed and considering each cluster as one genotype at 80% level of similarity coefficient, thereby in CCS Haryana Agricultural University soils, 3 genotypes were present in field pea roots while 17 genotypes were present in the nodules. A combined dandrogram showed that entirely a separate cluster was formed by the root endophytes as compared to nodules endophytes, which indicates that the number and type of bacterial genera enters depending upon the plant tissue and environmental conditions in field pea, besides the microbial population present in the soil.
Background and aims:
The legume clade Lotononis sensu lato (s.l.; tribe Crotalarieae) comprises three genera: Listia, Leobordea and Lotononis sensu stricto (s.s.). Listia species are symbiotically specific and form lupinoid nodules with rhizobial species of Methylobacterium and Microvirga. This work investigated whether these symbiotic traits were confined to Listia by determining the ability of rhizobial strains isolated from species of Lotononis s.l. to nodulate Listia, Leobordea and Lotononis s.s. hosts and by examining the morphology and structure of the resulting nodules.
Methods:
Rhizobia were characterized by sequencing their 16S rRNA and nodA genes. Nodulation and N2 fixation on eight taxonomically diverse Lotononis s.l. species were determined in glasshouse trials. Nodules of all hosts, and the process of infection and nodule initiation in Listia angolensis and Listia bainesii, were examined by light microscopy.
Key results:
Rhizobia associated with Lotononis s.l. were phylogenetically diverse. Leobordea and Lotononis s.s. isolates were most closely related to Bradyrhizobium spp., Ensifer meliloti, Mesorhizobium tianshanense and Methylobacterium nodulans. Listia angolensis formed effective nodules only with species of Microvirga. Listia bainesii nodulated only with pigmented Methylobacterium. Five lineages of nodA were found. Listia angolensis and L. bainesii formed lupinoid nodules, whereas nodules of Leobordea and Lotononis s.s. species were indeterminate. All effective nodules contained uniformly infected central tissue. Listia angolensis and L. bainesii nodule initials occurred on the border of the hypocotyl and along the tap root, and nodule primordia developed in the outer cortical layer. Neither root hair curling nor infection threads were seen.
Conclusions:
Two specificity groups occur within Lotononis s.l.: Listia species are symbiotically specific, while species of Leobordea and Lotononis s.s. are generally promiscuous and interact with rhizobia of diverse chromosomal and symbiotic lineages. The seasonally waterlogged habitat of Listia species may favour the development of symbiotic specificity.
Nine fast-growing bacteria were isolated from mung bean nodules and characterized with plant growth promoting properties. All the isolated bacteria were able to colonize mung bean root at varying level of 3 × 10 3 to 3 × 10 7 cfu (0.1 mg)-1 root, whereas, bacterial isolates M2, M4, M5 and M6 showed high biofilm formation ability on abiotic surface. None of them was able to nodulate mung bean plant when reinoculated. Bacterial isolate M2 was found to be an efficient indole acetic acid producer (28.3 µg mL-1), whereas M6 was an excellent phosphate solubilizer (21.8 µg mL-1). Two isolates, M1 and M3, were able to fix nitrogen. 16S rRNA gene sequence analysis of potential isolates revealed that bacterial isolates M2 and M6 showed maximum similarity with Bacillus subtilis, M4 with Bacillus simplex and M5 with Agrobacterium tumefaciens. Further, the impact of co-inoculation of these non-rhizobial bacteria with Bradyrhizobium sp. MN-S on nodulation, plant growth and grain yield of mung bean was also assessed. Generally, co-inoculation significantly improved nodulation and grain yield compared with Bradyrhizobium sp. MN-S alone inoculation. The enhancement due to co-inoculation in nodule number and nodule dry weight was 78 and 127%, respectively when compared with the Bradyrhizobium sp. MN-S alone. Co-inoculation combination of Bradyrhizobium sp. MN-S with B. subtilis M6 performed best by increasing 22% grain yield, while the rest combinations also benefited plants non-significantly. The results show that non-rhizobial plant growth promoting bacteria improve nodulation and grain yield of the legumes upon co-inoculation with crop specific rhizobia.
In the greenhouse experiment, two Bacillus and two Pseudomonas strains were examined for their ability to promote common bean growth (Phaseolus vulgaris L.) when co-inoculated with Rhizobium phaseoli. Co-inoculation with Rhizobium and Pseudomonas sp. LG or Bacillus sp. Bx improved shoot dry weight, nitrogen and phosphorus contents in bean plants, compared to inoculation with Rhizobium alone. Pseudomonas sp. LG promoted bean growth and particularly P uptake more efficiently than Bacillus sp. Bx. In vitro screening for plant growth-promoting traits showed phosphate solubilization, IAA, ammonia and siderophore production by Pseudomonas sp. LG, as opposed to Bacillus sp. Bx, which showed only ammonia production. Multiple plant growth-promoting traits of Pseudomonas sp. LG may be associated with its ability to improve common bean growth and nutrient uptake efficiently.
Aims
Low numbers of rhizobia in soil or inoculants delay nodulation and decrease symbiotic legume productivity. This study investigated the effect of co-inoculation with a helper bacterium, Pseudomonas fluorescens WSM3457 on the Medicago truncatula - Ensifer (Sinorhizobium) medicae WSM419 symbiosis challenged by a low inoculum dose.
Methods
In a glasshouse experiment the effect of co-inoculation with WSM3457 on the kinetics of nodule initiation and development was assessed 5, 7, 10, 14, 17, 21, and 42 days after inoculation of M. truncatula cv. Caliph with 103 cells/plant of E. medicae WSM419.
Results
Co-inoculated plants had enhanced rate of nodule initiation and development, greater numbers of larger crown nodules, and by day 42 accumulated more N than plants inoculated with E. medicae WSM419 alone. Nodule development was altered by co-inoculation. Approximately 25% of nodule initials on co-inoculated plants formed in closely associated pairs, young nodules were larger with multiple meristems and developed into cluster-like multi-lobed nodules compared to those on WSM419 inoculated plants. Molecular typing showed WSM3457 occupied a significant proportion of root nodules on co-inoculated plants.
Conclusion
Co-inoculation with P. fluorescens WSM3457 enhanced symbiotic effectiveness of M. truncatula when inoculated with a low inoculum dose of E. medicae WSM419.
Strains of Gram-negative, rod-shaped, non-spore-forming bacteria were isolated from nitrogen-fixing nodules of the native legumes Listia angolensis (from Zambia) and Lupinus texensis (from Texas, USA). Phylogenetic analysis of the 16S rRNA gene showed that the novel strains belong to the genus Microvirga, with 96.1 % or greater sequence similarity with type strains of this genus. The closest relative of the representative strains Lut6T and WSM3557T was M. flocculans TFBT, with 97.6-98.0 % similarity, while WSM3693T was most closely related to M. aerilata 5420S-16T, with 98.8 % similarity. Analysis of the concatenated sequences of four housekeeping gene loci (dnaK, gyrB, recA, rpoB) and cellular fatty acid profiles confirmed the placement of Lut6T, WSM3557T and WSM3693T within Microvirga. DNA:DNA relatedness values and physiological and biochemical tests allowed genotypic and phenotypic differentiation of Lut6T, WSM3557T and WSM3693T from each other and from other validly published Microvirga species. The nodA sequence of Lut6T was placed in a clade that contained strains of Rhizobium, Mesorhizobium and Sinorhizobium, while the 100 % identical nodA sequences of WSM3557T and WSM3693T clustered with Bradyrhizobium, Burkholderia and Methylobacterium strains. Concatenated sequences for nifD and nifH show that Lut6T, WSM3557T and WSM3693T were most closely related to Rhizobium etli CFN42T nifDH. On the basis of genotypic, phenotypic and DNA relatedness data, three novel species of Microvirga are proposed: Microvirga lupini (Lut6T = LMG26460T, = HAMBI 3236) Microvirga lotononidis (WSM3557T = LMG26455T, = HAMBI 3237) and Microvirga zambiensis (WSM3693T = LMG26454T, = HAMBI 3238).
Our current knowledge of plant-microbe interactions indicate that populations inhabiting a host plant are not restricted to a single microbial species but comprise several genera and species. No one knows if communities inside plants interact, and it has been speculated that beneficial effects are the result of their combined activities. During an ecological study of nitrogen-fixing bacterial communities from Lupinus angustifolius collected in Spain, significant numbers of orange-pigmented actinomycete colonies were isolated from surface-sterilized root nodules. The isolates were analysed by BOX-PCR fingerprinting revealing an unexpectedly high genetic variation. Selected strains were chosen for 16S rRNA gene sequencing and phylogenetic analyses confirmed that all strains isolated belonged to the genus Micromonospora and that some of them may represent new species. To determine the possibility that the isolates fixed atmospheric nitrogen, chosen strains were grown in nitrogen-free media, obtaining in some cases, significant growth when compared with the controls. These strains were further screened for the presence of the nifH gene encoding dinitrogenase reductase, a key enzyme in nitrogen fixation. The partial nifH-like gene sequences obtained showed a 99% similarity with the sequence of the nifH gene from Frankia alni ACN14a, an actinobacterium that induces nodulation and fixes nitrogen in symbiosis with Alnus. In addition, in situ hybridization was performed to determine if these microorganisms inhabit the inside of the nodules. This study strongly suggests that Micromonospora populations are natural inhabitants of nitrogen-fixing root nodules.
A universal method to detect and determine siderophores was developed by using their high affinity for iron(III). The ternary complex chrome azurol S/iron(III)/hexadecyltrimethylammonium bromide, with an extinction coefficient of approximately 100,000 M-1 cm-1 at 630 nm, serves as an indicator. When a strong chelator removes the iron from the dye, its color turns from blue to orange. Because of the high sensitivity, determination of siderophores in solution and their characterization by paper electrophoresis chromatography can be performed directly on supernatants of culture fluids. The method is also applicable to agar plates. Orange halos around the colonies on blue agar are indicative of siderophore excretion. It was demonstrated with Escherichia coli strains that biosynthetic, transport, and regulatory mutations in the enterobactin system are clearly distinguishable. The method was successfully used to screen mutants in the iron uptake system of two Rhizobium meliloti strains, DM5 and 1021.
Endophytic bacteria reside within plant tissues and have often been found to promote plant growth. Fourteen strains of putative endophytic bacteria, not including endosymbiotic Bradyrhizobium strains, were isolated from surface-sterilized soybean (Glycine max. (L.) Merr.) root nodules. These isolates were designated as non-Bradyrhizobium endophytic bacteria (NEB). Three isolates (NEB4, NEB5, and NEB17) were found to increase soybean weight when plants were co-inoculated with one of the isolates and Bradyrhizobium japonicum under nitrogen-free conditions, compared with plants inoculated with B. japonicum alone. In the absence of B. japonicum, these isolates neither nodulated soybean, nor did they affect soybean growth. All three isolates were Gram-positive spore-forming rods. While Biolog tests indicated that the three isolates belonged to the genus Bacillus, it was not possible to determine the species. Phylogenetic analysis of 16S rRNA gene hypervariant region sequences demonstrated that both NEB4 and NEB5 are Bacillus subtilis strains, and that NEB17 is a Bacillus thuringiensis strain.
One of the major mechanisms utilized by plant growth-promoting rhizobacteria (PGPR) to facilitate plant growth and development is the lowering of ethylene levels by deamination of 1-aminocyclopropane-1-carboxylic acid (ACC) the immediate precursor of ethylene in plants. The enzyme catalysing this reaction, ACC deaminase, hydrolyses ACC to alpha-ketobutyrate and ammonia. Several bacterial strains that can utilize ACC as a sole source of nitrogen have been isolated from rhizosphere soil samples. All of these strains are considered to be PGPR based on the ability to promote canola seedling root elongation under gnotobiotic conditions. The treatment of plant seeds or roots with these bacteria reduces the amount of ACC in plants, thereby lowering the concentration of ethylene. Here, a rapid procedure for the isolation of ACC deaminase-containing bacteria, a root elongation assay for evaluating the effects of selected bacteria on root growth, and a method of assessing bacterial ACC deaminase activity are described in detail. This should allow researchers to readily isolate new PGPR strains adapted to specific environments.
The use of Rhizobium inoculant for groundnut is a common practice in India. Also, co-inoculation of Rhizobium with other plant growth-promoting bacteria received considerable attention in legume growth promotion. Hence, in the present study we investigated effects of co-inoculating the sulfur (S)-oxidizing bacterial strains with Rhizobium, a strain that had no S-oxidizing potential in groundnut. Chemolithotrophic S-oxidizing bacterial isolates from different sources by enrichment isolation technique included three autotrophic (LCH, SWA5 and SWA4) and one heterotrophic (SGA6) strains. All the four isolates decreased the pH of the growth medium through oxidation of elemental S to sulfuric acid. Characterization revealed that these isolates tentatively placed into the genus Thiobacillus. Clay-based pellet formulation (2.5 x 10(7) cf ug(-1) pellet) of the Thiobacillus strains were developed and their efficiency to promote plant growth was tested in groundnut under pot culture and field conditions with S-deficit soil. Experiments in pot culture yielded promising results on groundnut increasing the plant biomass, nodule number and dry weight, and pod yield. Co-inoculation of Thiobacillus sp. strain LCH (applied at 60 kg ha(-1)) with Rhizobium under field condition recorded significantly higher nodule number, nodule dry weight and plant biomass 136.9 plant(-1), 740.0mg plant(-1) and 15.0 g plant(-1), respectively, on 80 days after sowing and enhanced the pod yield by 18%. Also inoculation of S-oxidizing bacteria increased the soil available S from 7.4 to 8.43 kg ha(-1). These results suggest that inoculation of S-oxidizing bacteria along with rhizobia results in synergistic interactions promoting the yield and oil content of groundnut, in S-deficit soils.
The effects of the pathogenic fungus Fusarium oxysporum f. sp. radicis-lycopersici and of the bacterial biocontrol strain Pseudomonas fluorescens WCS365, and of both microbes, on the amounts and composition of root exudate components of tomato plants grown in a gnotobiotic stonewool substrate system were studied. Conditions were selected under which introduction of F. oxysporum f. sp. radicis-lycopersici caused severe foot and root rot, whereas inoculation of the seed with P. fluorescens WCS365 decreased the percentage of diseased plants from 96 to 7%. This is a much better disease control level than was observed in potting soil. Analysis of root exudate revealed that the presence of F. oxysporum f. sp. radicis-lycopersici did not alter the total amount of organic acids, but that the amount of citric acid decreased and that of succinic acid increased compared with the nontreated control. In contrast, in the presence of the P. fluorescens biocontrol strain WCS365, the total amount of organic acid increased, mainly due to a strong increase of the amount of citric acid, whereas the amount of succinic acid decreased dramatically. Under biocontrol conditions, when both microbes are present, the content of succinic acid decreased and the level of citric acid was similar to that in the nontreated control. The amount of sugar was approximately half that of the control sample when either one of the microbes was present alone or when both were present. Analysis of the interactions between the two microbes grown together in sterile tomato root exudate showed that WCS365 inhibited multiplication of F. oxysporum f. sp. radicis-lycopersici, whereas the fungus did not affect the number of CFU of the bacterium.
We evaluated phytohormone and polyamine biosynthesis, siderophore production, and phosphate solubilization in two strains (Cd and Az39) of Azospirillum brasilense used for inoculant formulation in Argentina during the last 20 years. Siderophore production and phosphate solubilization were evaluated in a chemically defined medium, with negative results. Indole 3-acetic acid (IAA), gibberellic acid (GA(3)), and abscisic acid (ABA) production were analyzed by gas chromatography-mass spectrometry. Ethylene, polyamine, and zeatin (Z) biosynthesis were determined by gas chromatography-flame ionization detector and high performance liquid chromatography (HPLC-fluorescence and -UV), respectively. Phytohormones IAA, Z, GA(3), ABA, ethylene, and growth regulators putrescine, spermine, spermidine, and cadaverine (CAD) were found in culture supernatant of both strains. IAA, Z, and GA(3) were found in all two strains; however, their levels were significantly higher (p < 0.01) in Cd (10.8, 2.32, 0.66 microg ml(-1)). ABA biosynthesis was significantly higher (p < 0.01) in Az39 (0.077 microg ml(-1)). Ethylene and polyamine CAD were found in all two strains, with highest production in Cd cultured in NFb plus L-methionine (3.94 ng ml(-1) h(-1)) and Az39 cultured in NFb plus L-lysine (36.55 ng ml(-1) h(-1)). This is the first report on the evaluation of important bioactive molecules in strains of A. brasilense as potentially capable of direct plant growth promotion or agronomic yield increase. Az39 and Cd showed differential capability to produce the five major phytohormones and CAD in chemically defined medium. This fact has important technological implications for inoculant formulation as different concentrations of growth regulators are produced by different strains or culture conditions.
This study aimed to characterize endophytic bacteria associated with root-nodules of a wild legume (Genista cinerea: Fabaceae) growing in arid soils of Algeria. A total of ten non-symbiotic endophytic bacterial strains were isolated and identified using a combination of conventional and molecular approaches based on 16S rRNA gene sequencing. Moreover, growth variations of the isolates under different environmental conditions were examined using advanced statistical modeling techniques (Markov Chain Monte Carlo sampler for multivariate generalized linear mixed models-MCMCglmm). The results revealed the existence of a large physiological diversity among the endophyte isolates, which formed two distinct groups. Group 1, included fast-growing and salt-tolerant isolates and Group 2 covered acid-sensitive isolates. The bacterial isolates showed capabilities to assimilate different carbonaceous and nitrogenous substances, with consistent and large tolerances to pH [4–10], temperature [4–55°C], and salinity [NaCl=2–10%]. In addition, The MCMCglmm indicated that scored growth rate increased with pH and decreases at high level of salinity. The endophytic strains were classified as fast-growing bacteria belonged to the β and γ-Proteobacteria including Achromobacter, Klebsiella, Luteibacter, Pantoea and Pseudomonas. The present findings may support the idea that these isolates are passenger bacteria able to colonize nodules of Genista cinerea. Furthermore, the salt and temperature tolerant patterns found among the isolates reflect the environmental stresses pressure and the importance of using efficient indigenous endophytic strains for successful plant inoculation in arid agriculture.
Microbial communities associated with roots confer specific functions to their hosts, thereby modulating plant growth, health, and productivity. Yet, seminal questions remain largely unaddressed including whether and how the rhizosphere microbiome modulates root metabolism and exudation and, consequently, how plants fine tune this complex belowground web of interactions. Here we show that, through a process termed systemically induced root exudation of metabolites (SIREM), different microbial communities induce specific systemic changes in tomato root exudation. For instance, systemic exudation of acylsugars secondary metabolites is triggered by local colonization of bacteria affiliated with the genus Bacillus . Moreover, both leaf and systemic root metabolomes and transcriptomes change according to the rhizosphere microbial community structure. Analysis of the systemic root metabolome points to glycosylated azelaic acid as a potential microbiome-induced signaling molecule that is subsequently exuded as free azelaic acid. Our results demonstrate that rhizosphere microbiome assembly drives the SIREM process at the molecular and chemical levels. It highlights a thus-far unexplored long-distance signaling phenomenon that may regulate soil conditioning.
Plant growth-promoting rhizobacteria (PGPR) and rhizobia are potentially advantageous in improving plant growth in heavy metal contaminated soils. However, only limited information is available in literature on the manner through which the co-inoculation of PGPR and rhizobia can potentially supply nutrients to benefit plant growth in heavy metal contaminated soil. Accordingly, this study investigated the effects of Paenibacillus mucilaginosus (PGPR) and Sinorhizobium meliloti (rhizobia) co-inoculation on soil nutrients, enzyme activities, and microbial biomass in copper (Cu) contaminated soil planted with alfalfa (Medicago sativa). Moreover, we assessed soil bacterial community structure using high-throughput Illumina sequencing of 16S rRNA genes. Results showed that PGPR and/or rhizobia inoculation improved alfalfa growth. In particular, we found that this co-inoculation approach decreased Cu accumulation (48.6%) in shoots compared to the control (uninoculated). Both partial least squares path modeling (PLS-PM) and the relative importance of regressors in the linear models identified that enzyme activities, microbial biomass, and microbial community structure in Cu contaminated soil were major controlling variables of soil nutrient availability. The co-inoculation treatment significantly increased soil carbon (C) and nitrogen (N) concentrations by increasing urease (55.6%), saccharase (29.5%), and β-glucosidase (31.4%) activities compared to the control. Furthermore, the rhizosphere microbial community structure in the co-inoculation treatment was mainly regulated by soil N concentrations (i.e., both total N and available N) while altering alpha diversity (α-diversity). The relative abundances of Firmicutes (including biomarkers of the Bacillus genus) and Acidobacteria were enriched in the co-inoculated treatment, which can potentially improve soil nutrient availability and subsequently benefit plant growth. These findings indicated that the co-inoculation of PGPR and rhizobia plays an important role in promoting plant growth in Cu contaminated soil. This is because this approach can increase soil nutrient availability by enhancing soil enzyme activities and regulating rhizosphere microbial community structure.
Inoculation of the carrier-based mixed bioinoculants of N-fixer (Azospirillum lipoferum strain Az204/Rhizobium strain BMBS P47) and phosphate-solubilizing bacterium (Bacillus megaterium var phosphaticum strain Pb1) promoted growth and yield of pearl millet and blackgrain under pot-culture conditions. The mixed inoculant of Az204 and Pb1 enhanced germination, seedling vigor, plant height, and seed weight, and resulted in 6% increase in grain yield of pearl millet. Likewise, the mixed inoculant of BMBS P47 and Pb1 increased growth, nodulation, and yield in blackgram. The rhizosphere soil enzyme activities, including nitrogenase, urease, and phosphatase, in both pearl millet and blackgram were significantly increased by the inoculation of the mixed inoculant, compared to that of the individual inoculants. The results clearly indicate the beneficial effect of co-culturing the N-fixer and P-solubilizer in inoculants production.
Most healthy naturally propagated plants grown in field or potting soils are colonized by communities of endophytic bacteria, embracing a wide variety of species and genera. These bacteria form nonpathogenic relationships with their hosts: some beneficial, some neutral, and some detrimental. Such associations can increase plant growth and hasten development or improve resistance to environmental stress. Endophytic bacteria have been implicated in supplying biologically fixed nitrogen in non-legumes, and these associations can increase the nitrogen economy of a crop, reducing the requirement for N fertilizers. Bacterial endophytes have also been shown to prevent disease development through endophyte-mediated de novo synthesis of structural compounds and fungitoxic metabolites. Such induced protection responses have been linked to certain forms of systemic acquired (disease) resistance. Certain crop sequences have been shown to favor the build-up of specific plant growth-promoting bacterial endophyte populations. These can lead to the creation of beneficial host-endophyte allelopa-thies, with implications for the formation and maintenance of fertile, disease-suppressive soils. Manipulating bacterial populations in soils and within crops will be crucial if endophytes are to be utilized in crop production systems, and special techniques will be required to do so. This review surveys the natural associations between bacterial endophytes and their hosts, and discusses how such relationships can be employed most productively in sustainable systems of agricultural crop production.
LOK, E. H., O'HARA, G. & DELL, B. 2006. Nodulation of the legume Pterocarpus indicus by diverse strains of rhizobia. Pterocarpus indicus (Leguminosae: Papilionoideae) is a tropical legume with potential for commercial forest plantation. A glasshouse experiment was undertaken to investigate the effects of inoculation of 18 strains of diverse rhizobia comprising species from four genera on P. indicus seedlings. Pterocarpus indicus was effectively nodulated by Bradyrhizobium elkani WSM 2096. These seedlings had a significantly greater plant yield compared with seedlings with other strains and the controls. Strain Bradyrhizobium sp. WSM 3712, originally isolated from P. indicus from Malaysia, formed many partially effective nodules. However, three other slow- growing strains, B. liaoningense WSM 2098, Bradyrhizobium sp. TAL 643 and Bradyrhizobium sp. TAL 651, ineffectively nodulated P. indicus. The moderately fast-growing strain, Mesorhizobium ciceri WSM 2100, and four fast-growing strains, Sinorhizobium meliloti WSM 2114, Rhizobium hainanense WSM 2106, Rhizobium gallicum R 602 and Rhizobium tropici WSM 2110, also formed ineffective nodules on P. indicus. Pterocarpus indicus appears to be a diverse and promiscuous host for nodulation but with a narrow range for effective N2-fixing symbiosis. This information is useful for future inoculation programmes for P. indicus in containerized nurseries.
Plant-associated bacteria that live inside plant tissues without causing any harm to plants are defined as endophytic bacteria. The present investigation was carried out to analyse the phenotypic and genotypic diversity in the bacterial endophytes of two species of soybean viz. Glycine max and G. soja . A total of 65 bacterial endophytes were isolated from three tissues: stem, root and nodule. All the isolates were screened for Gram reaction, secretion of hydrolytic enzymes (pectinase and cellulase), fluorescent pigment production, and motility, resistance to streptomycin @ 100 )g/ml, capsule formation and IAA production. Genotypic variation was studied using PCR-based 16S rDNA-RFLP. Preliminary characterization of the 65 endophytes showed that approximately equal percentages of gram positive (49%) and gram negative (51%) bacteria were present. Approximately 80% were motile, 33% and 70% secreted pectinase and cellulase, respectively and 17% did not produce IAA in vitro . Phenotypically the 65 isolates were found to show less closeness among themselves for the characters studied. Molecular characterization of selected 35 endophytic bacteria was carried out by PCR amplification of 16S rDNA gene, and its restriction analysis using three tetra cutters, HaeIII , MboI and MspI . Two main clusters were observed at 48% and 43% similarity coefficients in which most of the endophytes belonged. Six of the total isolates (I-8, I-15, I-25, I- 68, I-121 and I-137) did not come into these clusters, showing their divergence from the rest. The genetic variation was more among endophytes isolated from G. max tissues than G. soja . Keyword: characterization, endophytic bacteria, phyllosphere, rhizosphere, soybean.
The aim of this study was to identifiy the endophytic bacteria recovered from the foliage, tap roots and nodules of red clover
plants (Trifolium pratense L.); and to assess the effects of the nodule bacteria, alone and in combination with Rhizobium spp., on the growth and development of red clover seedlings. Thirty-one bacteria species from 14 different genera were recovered
from within the foliage, roots and nodules of red clover plants cv. AC Charlie. Genera diversity and species number were greatest
in foliage tissues. Pantoea agglomerans (59.6%) was the most frequent species recovered in foliage tissues, Agrobacterium rhizogenes A in the tap root (49.2%) and Rhizobium leguminosarum BV phaseoli and R. loti B in the nodules (27.2% each). Recovery of Rhizobium species was not restricted to the nodules, and species of this genus were systemic throughout the plant. Clover root nodules
were host to 12 bacteria species other than rhizobia, of which 8 were specific to this tissue. Using non-selective media,
R. leguminosarum BV trifolii constituted only 8.8% of all the root nodule bacteria recovered. In root bacterization experiments, species of nodule bacteria
promoted growth of red clover more often when applied in combination with R. leguminosarum BV trifolii than when applied singly. However, Bacillus megaterium, Bordetella avium and Curtobacterium luteum consistently promoted growth either individually or in combination with R. leguminosarum BV trifolii. Nodulation was promoted when R. leguminosarum BV trifolii was coinoculated with Bacillus insolitus, B. brevis or A. rhizogenes A. Single isolate applications of Rhizobium species to roots always led to the depression of clover growth, but mixtures of R. leguminosarum BV trifolii and R. leguminosarum BV phaseoli resulted in growth promotion. The latter is considered further evidence of the beneficial allelopathic side effect of strain
competition for the same ecological niche.
Understanding the interaction mechanisms between plant growth-promoting rhizobacteria (PGPR), leguminous crops, and rhizobia is necessary to effectively use PGPR in increasing the biological nitrogen fixation of legumes. We determined the coinoculation effects of Bradyrhizobium japonicum A1017 and a gusA-marked strain of Pseudomonas fluorescens 2137, P. fluorescens WCS365, Azomonas agilis 125, and Azospirillum lipoferum 137 on soybean [Glycine max (L.) Merr] cv. Enrei grown under axenic conditions. The gusA-marked rhizobacteria effectively colonized the root tips and surfaces near the roots tips with a colonization rate ranging from 7.50 to 8.62 log colony forming units (cfu) gfw-1. P. fluorescens 2137 had the highest colonization activity on soybean roots whether inoculated alone or coinoculated with B. japonicum A1017. Coinoculation of P. fluorescens 2137 and B. japonicum A1017 increased the colonization of B. japonicum A1017 on soybean roots, nodule number, and acetylene reduction activity (ARA) at 10 and 20 days after inoculation. Moreover, the addition of sterile spent medium of P. fluorescens 2137 increased the growth of B. japonicum A1017 in yeast mannitol broth (YMB), indicating that P. fluorescens 2137 may have released substances that increased the rhizobial population. The results of this study suggest that the enhanced nodulation and ARA of soybean due to the high colonization of P. fluorescens on soybean roots could depend on the production of growth-promoting substances that stimulate the growth of B. japonicum. However, coinoculation with P. fluorescens WCS365 decreased the nodule number and ARA, despite its slight stimulation of the growth of B. japonicum on the roots, indicating that coinoculation effects are strain dependent.
Numerous species of soil bacteria which flourish in the rhizosphere of plants, but which may grow in, on, or around plant tissues, stimulate plant growth by a plethora of mechanisms. These bacteria are collectively known as PGPR (plant growth promoting rhizobacteria). The search for PGPR and investigation of their modes of action are increasing at a rapid pace as efforts are made to exploit them commercially as biofertilizers. After an initial clarification of the term biofertilizers and the nature of associations between PGPR and plants (i.e., endophytic versus rhizospheric), this review focuses on the known, the putative, and the speculative modes-of-action of PGPR. These modes of action include fixing N2, increasing the availability of nutrients in the rhizosphere, positively influencing root growth and morphology, and promoting other beneficial plant–microbe symbioses. The combination of these modes of actions in PGPR is also addressed, as well as the challenges facing the more widespread utilization of PGPR as biofertilizers.
A detailed study (involving in vitro bioassays, greenhouse and field trials) was carried out to develop an integrated approach to control fusarium wilt of banana. Eight biocontrol agents were screened against Fusarium oxysporum f. sp. cubense in vitro. Among these, the Pfm strain of Pseudomonas fluorescens showed maximum growth inhibition of the pathogen and a compatible reaction with extracts of neem cake. Similar effects were shown by T. viride and T. harzianum, P.fluorescens; these agents were used further to verify efficacy under greenhouse conditions using various methods of application. Among the methods tested, soil application of biocontrol agents proved to be most effective against the wilt disease and associated vascular discolouration. Soil application of talc formulation of P. fluorescens at 10 g/plant had the least effect when measured by wilt index and vascular discolouration index. Other methods, viz., sucker dipping corm injection and capsule application showed a significant reduction in wilt index and vascular discolouration. In field trials, there were 15 treatments including soil application, sucker dipping in spore suspension, basal application of neem cake, in all possible combinations, together with chemical and uninoculated control. Two field trials in two locations each were conducted during 2000–2001 and 2001–2002 to test its efficacy. The treatment: basal application of neem cake at 0.5 kg/plant+sucker dipping in spore suspension of P. fluorescens for 15 min+soil application of P. fluorescens at 10 g/plant at 3.5 and 7 months after planting showed the greatest suppression of wilt disease and this was on par with basal application of neem cake at 0.5 kg/plant+soil application of P. fluorescens at 10 g/plant at 3.5 and 7 months after planting. Other treatments also showed significant effects in reducing wilt incidence.
Thirty-nine endophytic bacterial strains were isolated from the nodule of Lespedeza sp. grown in two different locations of South Korea. All strains were checked for their plant growth promoting (PGP) abilities under in vitro conditions. Most of the isolates showed multiple PGP activity, i.e., indole acetic acid production, ACC deaminase activity, siderophore production, and phosphate solubilization. The strains were identified by using 16S rRNA gene sequence analysis as belonging to Alphaproteobacteria, Betaproteobacteria, Actinobacteria, and Firmicutes phylum with nine different genera Arthrobacter, Bacillus, Bradyrhizobium, Burkholderia, Dyella, Methylobacterium, Microbacterium, Rhizobium, and Staphylococcus. Gene nodA amplification showed positive results only for strains from Bradyrhizobium and Rhizobium genera. The strains from Bradyrhizobium and Rhizobium genera enhanced plant growth, nodulation, and acetylene reduction activity when inoculated on Vigna unguiculata L. (cowpea), whereas other strains did not induce nodule formation but enhanced plant growth. Herbaceous legume Lespedeza sp. formed root nodules with diverse bacterial group, and probably, these bacteria can be used for stimulating plant growth.
Bacterial endophytes live inside plants for at least part of their life cycle. Studies of the interaction of endophytes with their host plants and their function within their hosts are important to address the ecological relevance of endophytes. The modulation of ethylene levels in plants by bacterially produced 1-aminocyclopropane-1-carboxylate deaminase is a key trait that enables interference with the physiology of the host plant. Endophytes with this capacity might profit from association with the plant, because colonization is enhanced. In turn, host plants benefit by stress reduction and increased root growth. This mechanism leads to the concept of competent endophytes, defined as endophytes that are equipped with genes important for maintenance of plantendophyte associations. The ecological role of these endophytes and their relevance for plant growth are discussed here
Successful competition for iron by potential pathogens is essential to establish infection. The roles of the various types of microbial iron acquisition systems in host-pathogen interactions depend on the nature of the infection and the location of the pathogen within the host. Microbes infecting the extracellular spaces of the host employ different strategies for iron acquisition than those that invade and multiply within host cells.
The methodology, characteristics and application of the sensitive C2H2-C2H4 assay for N2 fixation by nitrogenase preparations and bacterial cultures in the laboratory and by legumes and free-living bacteria in situ is presented in this comprehensive report. This assay is based on the N2ase-catalyzed reduction of C2H2 to C2H4, gas chromatographic isolation of C2H2 and C2H4, and quantitative measurement with a H2-flame analyzer. As little as 1 μμmole C2H4 can be detected, providing a sensitivity 10³-fold greater than is possible with ¹⁵N analysis.
A simple, rapid and effective procedure utilizing syringe-type assay chambers is described for the analysis of C2H2-reducing activity in the field. Applications to field samples included an evaluation of N2 fixation by commercially grown soybeans based on over 2000 analyses made during the course of the growing season. Assay values reflected the degree of nodulation of soybean plants and indicated a calculated seasonal N2 fixation rate of 30 to 33 kg N2 fixed per acre, in good agreement with literature estimates based on Kjeldahl analyses. The assay was successfully applied to measurements of N2 fixation by other symbionts and by free living soil microorganisms, and was also used to assess the effects of light and temperature on the N2 fixing activity of soybeans. The validity of measuring N2 fixation in terms of C2H2 reduction was established through extensive comparisons of these activities using defined systems, including purified N2ase preparations and pure cultures of N2-fixing bacteria.
With this assay it now becomes possible and practicable to conduct comprehensive surveys of N2 fixation, to make detailed comparisons among different N2-fixing symbionts, and to rapidly evaluate the effects of cultural practices and environmental factors on N2 fixation. The knowledge obtained through extensive application of this assay should provide the basis for efforts leading to the maximum agricultural exploitation of the N2 fixation reaction.
Recent molecular studies on endophytic bacterial diversity have revealed a large richness of species. Endophytes promote plant growth and yield, suppress pathogens, may help to remove contaminants, solubilize phosphate, or contribute assimilable nitrogen to plants. Some endophytes are seedborne, but others have mechanisms to colonize the plants that are being studied. Bacterial mutants unable to produce secreted proteins are impaired in the colonization process. Plant genes expressed in the presence of endophytes provide clues as to the effects of endophytes in plants. Molecular analysis showed that plant defense responses limit bacterial populations inside plants. Some human pathogens, such as Salmonella spp., have been found as endophytes, and these bacteria are not removed by disinfection procedures that eliminate superficially occurring bacteria. Delivery of endophytes to the environment or agricultural fields should be carefully evaluated to avoid introducing pathogens.
The colonization pattern of Vitis vinifera L. by Burkholderia phytofirmans strain PsJN was determined using grapevine fruiting cuttings with emphasis on putative inflorescence colonization under nonsterile conditions. Two-week-old rooted plants harbouring flower bud initials, grown in nonsterile soil, were inoculated with PsJN:gfp2x. Plant colonization was subsequently monitored at various times after inoculation with plate counts and epifluorescence and/or confocal microscopy. Strain PsJN was chronologically detected on the root surfaces, in the endorhiza, inside grape inflorescence stalks, not inside preflower buds and flowers but rather as an endophyte inside young berries. Data demonstrated low endophytic populations of strain PsJN in inflorescence organs, i.e. grape stalks and immature berries with inconsistency among plants for bacterial colonization of inflorescences. Nevertheless, endophytic colonization of inflorescences by strain PsJN was substantial for some plants. Microscopic analysis revealed PsJN as a thriving endophyte in inflorescence organs after the colonization process. Strain PsJN was visualized colonizing the root surface, entering the endorhiza and spreading to grape inflorescence stalks, pedicels and then to immature berries through xylem vessels. In parallel to these observations, a natural microbial communities was also detected on and inside plants, demonstrating the colonization of grapevine by strain PsJN in the presence of other microorganisms.
Bacterial endophytes and their effects on plants and uses in agriculture
- Kobayashi
Kobayashi, D.Y., Palumbo, J.D., 2000. Bacterial endophytes and their effects on plants
and uses in agriculture. In: Bacon, C.W., White, J.F. (Eds.), Microbial Endophytes.
Marcel Dekker, New York, pp. 99-233.
Effects of co-cultures, containing N-fixer and P-solubilizer, on the growth and yield of pearl millet (Pennisetum glaucum (L.) R. Br.) and blackgram (Vigna mungo L.)
- Poonguzhali