Azospirillum Isolates Research Articles

Isolation and characterization of local Azospirillum and yeast strains to be used for wheat grains inoculation to improve plant growth and yield under field cultivation

Isolation and characterization of local Azospirillum and yeast strains to be used for wheat grains inoculation to improve plant growth and yield under field cultivation

Characterization of cellular, biochemical and genomic features of the diazotrophic plant growth-promoting bacterium Azospirillum sp. UENF-412522, a novel member of the Azospirillum genus

Given their remarkable beneficial effects on plant growth, several Azospirillum isolates currently integrate the formulations of various commercial inoculants. Our research group isolated a new strain, Azospirillum sp. UENF-412522, from passion fruit rhizoplane. This isolate uses carbon sources that are partially distinct from closely-related Azospirillum isolates. Scanning electron microscopy analysis and population counts demonstrate the ability of Azospirillum sp. UENF-412522 to colonize the surface of passion fruit roots. In vitro assays demonstrate the ability of Azospirillum sp. UENF-412522 to fix atmospheric nitrogen, to solubilize phosphate and to produce indole-acetic acid. Passion fruit plantlets inoculated with Azospirillum sp. UENF-41255 showed increased shoot and root fresh matter by 13,8% and 88,6% respectively, as well as root dry matter by 61,4%, further highlighting its biotechnological potential for agriculture. We sequenced the genome of Azospirillum sp. UENF-412522 to investigate the genetic basis of its plant-growth promotion properties. We identified the key nif genes for nitrogen fixation, the complete PQQ operon for phosphate solubilization, the acdS gene that alleviates ethylene effects on plant growth, and the napCAB operon, which produces nitrite under anoxic conditions. We also found several genes conferring resistance to common soil antibiotics, which are critical for Azospirillum sp. UENF-412522 survival in the rhizosphere. Finally, we also assessed the Azospirillum pangenome and highlighted key genes involved in plant growth promotion. A phylogenetic reconstruction of the genus was also conducted. Our results support Azospirillum sp. UENF-412522 as a good candidate for bioinoculant formulations focused on plant growth promotion in sustainable systems.

English

The objective of this study was to improve rice and maize yields using native Azospirillum-based biofertilizer. To reach this objective, samples of rhizosphere soil, non-rhizosphere soil and roots of maize plants were collected from the particular locations of Samanko and Bamako of the south Mali. Thirty-three different colonies of bacteria were isolated from the different samples. Based on their better growth in nitrogen free semi-solid medium, their morphological, biochemical and plant growth promotion characteristic, ten bacterial isolates were identified as Azospirillum isolates following the Bergey’s Manual of Determinative Bacteriology. Ten isolates were selected: Az1, Az2, Az3, Az4, Az5, Az6, Az7, Az8, Az9 and Az10. Strain Az6 showed great potential on both rice and maize production. Therefore, this strain is suggested for large scale rice and maize fields’ application. While the Azospirillum sp. Az5, Az6 and Az10 strains are suggested for large scale application in maize field, which may reduce production cost. Top dressing with 25% of the recommended nitrogen-fertilizer was found to decrease maize grain yield.   Key words: Azospirillum, Nitrogen-fixing bacteria, plant growth-promoting rhizobacteria (PGPR), biopesticide, maize, rice, Mali.

Assessment of effect of inoculation of efficient <em>Azospirillum </em>strains<em> </em>on growth and yield of foxtail millet [<em>Setaria italica </em>(L.) Beauv.

A pot culture experiment was conducted under greenhouse condition to study the effect of efficient isolates of Azospirillum strains on plant growth and yield parameters in foxtail millet. From foxtail millet growing areas of Raichur and Koppal districts, 40 samples were collected for isolation and enumeration of Azospirillum. In the investigation 21 isolates were found to be negative for denitrification and thus selected for further study. Based on in vitro studies, six efficient strains of Azospirillum were selected for pot culture experiment on foxtail millet growth and yield parameters at 30 and 60 days after sowing and at harvest. All the isolates including reference strain showed significant increase in plant height, No. of leaves per hill, No. of tillers/hill, Plant biomass, No. of panicles/hill, No. of filled seeds/panicle, No. of chaffy seeds/panicle, test weight, seed yield/plant, straw yield/hill of foxtail millet was observed over control. The strain MARV-18 was found to be efficient strain for foxtail millet crop.

Testing the Efficiency of Different Isolates of Azotobacter, Azospirillum and Pseudomonas for Some Traits Related to Plant Growth Promoters on Varieties of Onion

Testing the Efficiency of Different Isolates of Azotobacter, Azospirillum and Pseudomonas for Some Traits Related to Plant Growth Promoters on Varieties of Onion

Isolation and identification of Azospirillum and Azotobacter species from Acacia spp. at Riyadh, Saudi Arabia

Populations of Azospirillum and Azotobacter isolated and identified from the soil rhizosphere of Acacia ehernbergiana, A. gerrardii and A. tortilis from three sites varied widely irrespective of tree species, size and site. Azospirillum strains were purified and characterized through morphological, microscopic and biochemical tests for motility, cell shape, pigmentation, colony color, gram reaction, optimal temperatures, biotin requirement, pH growth, acidification of peptone glucose medium, ability of nitrogen fixation, growth on 3% NaCl, and D-glucose and D-ribose utilization. Twenty Azospirillum isolates were divided into four matching characters. They were identified as A. lipoferum, A. brasilense, A. dobereinerae and A. zeae. The same above tests with omission of temperatures, biotin requirement, and acidification of peptone and addition of starch hydrolysis, H2S, urease, oxidase and sucrose, maltose and mannitol as carbon source were applied to Azotobacter strains. Azotobacter isolates were divided into three matching characters. They were identified as A. chroococcum, A. beijerinckii and A. salinestris.

Analysis of physico-chemical parameters of soil and population density of Azospirillum isolates from paddy fields of Theni district, Tamilnadu

The rhizosphere region of eight different paddy field areas of Theni district was studied for their physico-chemical analysis of soil and the population density of Azospirillum sp. The study results showed range of values of pH (7.4-7.9), bulk density (1.13g/cm3 -1.60g/cm3), water holding capacity (34.31% - 18.25%), electrical conductivity (1.31 – 1.11), organic carbon (0.93% - 0.71%) . The macronutrient values namely total nitrogen (1.72% - 0.78%), phosphorus content (0.177% - 0.122%) and potassium (1.364% - 1.273%) were observed. Also micronutrients of various paddy fields like Zn (0.9% - 0.5%), Cu (2.3% - 1.7%), Fe (10.9% -8.2%), Mn (6.7% - 5.2%) were recorded, whereas the values of available macronutrients namely nitrogen (295kg/ha - 223kg/ha), phosphorous (89Kg/acre - 49kg/acre), potassium (790kg/acre - 490kg/acre) were noted. The Azospirillum population density was highest in Chinnamanur (192 x105 CFU/g) and lowest in Royyanpatti (91 x105 CFU/g). Thus through this work we were able to isolate and identify novel high yielding Azosprillium sp from paddy fields of Theni district.

Evaluation of salt-tolerant Azospirillum spp and its role in improvement of wheat growth parameter

Eighteen isolates of Azospirillum were collected from different sites at Kafrelsheikh Governorate. The isolates were cultivated on nitrogen-free malate (NFM) medium supplemented with different NaCl gl-1 concentrations (Zero, 0.7 and 1.5%) and tryptophan to study the salt tolerant in vitro and indol acetic acid production. Pots were used to evaluate the most salt-tolerant and efficient species and soils of this experiment had artificially salinized with different salinity levels. The most salt-tolerant A10 and A11 species were genetically identified by 16S rRNA sequencing and the results revealed to be closest matched at 99% A. lipoferum strains and selected to inoculate wheat plants. The investigated A. lipoferum species had variable divergence growth at different salinity levels. Actually, increased salinity had deleterious effect on IAA production. In addition, salinity had deleterious effect on dry weights of plants, number and dry weight of branches, total chlorophyll, sodium and potassium% with increasing salinity levels. But, inoculation with associative nitrogen fixation A. lipoferum isolates enhanced the previous parameters. Thus, inoculation with the salt-tolerant A.lipoferum isolates decreased the deleterious effect of salt stress on wheat plants.

SCREENING OF DIFFERENT ISOLATES OF AZOSPIRILLUMFOR GROWTH AT DIFFERENT SALT (NACL) CONCENTRATIONS

Azospirillum was abundant in the rhizosphere of halophytes collected from El- Thaliga salt marsh near Benghazi, Libya. More than twenty five isolated of Azospirillum were isolated from rhizosphere by enrichement method using DN semisolid medium with or without salt. After several transfer for purification, 13 isolates were screened to select the most efficient strains in a microcosms experiment with wheat. According to the cultural, morphological and biochemical experiments Azospirillum isolated were belonging to A. brasilense and isolated was belonging to A. lipoferum, Azospirillum isolates were screened for growth at different NaCl concentration in liquid DN medium.

Isolation and Characterization of Azospirillum Isolates from Soil and Their Effect on Growth and Yield of Wheat (Triticum aestivum L.) under Different Levels of Nitrogen Fertilizer

This study involved isolation of Azospirillum from soils that are different in texturefrom Aswan governorate. Also, study the response of wheat (Triticum aestivum L.) to inoculation with the isolated Azospirillum strain (M-1, M-2 and M-3) in presence of different levels of nitrogen fertilization (40, 60 and 80 kg N/fed.) was tested in field experiments in the two seasons 2014/2015 and 2015/2016.

In vitro PGPR properties and osmotic tolerance of different Azospirillum native strains and their effects on growth of maize under drought stress

Osmotic variations in the soil can affect bacterial growth diminishing the number of inoculated bacteria. In a scenario of water deficit having tolerant bacteria would be beneficial to achieve a better response of the plant to stress. Thus, selection of more resistant bacteria could be useful to design new inoculants to be used in arid zones. In this sense, a group of Azospirillum isolates deposited in INTA collection was characterized in order to select strains tolerant to osmotic stress. The results obtained demonstrated that Az19 strain has similar in vitro PGPR characteristics to Az39, the most used strain in Argentina for inoculants industries, with the advantage of a better tolerance to osmotic and salt stress. Inoculation of maize plants with this strain resulted in a better response against water deficit compared to Az39 strain, encouraging us to further study the behavior of this strain in greenhouse and field trials in view of developing new inoculants suitable for areas with water deficit.

Azospirillum spp. from native forage grasses in Brazilian Pantanal floodplain: biodiversity and plant growth promotion potential.

A sustainable alternative to improve yield and the nutritive value of forage is the use of plant growth-promoting bacteria (PGPB) that release nutrients, synthesize plant hormones and protect against phytopathogens (among other mechanisms). Azospirillum genus is considered an important PGPB, due to the beneficial effects observed when inoculated in several plants. The aim of this study was to evaluate the diversity of new Azospirillum isolates and select bacteria according to the plant growth promotion ability in three forage species from the Brazilian Pantanal floodplain: Axonopus purpusii, Hymenachne amplexicaulis and Mesosetum chaseae. The identification of bacterial isolates was performed using specific primers for Azospirillum in PCR reactions and partial sequencing of the 16S rRNA and nifH genes. The isolates were evaluated in vitro considering biological nitrogen fixation (BNF) and indole-3-acetic acid (IAA) production. Based on the results of BNF and IAA, selected isolates and two reference strains were tested by inoculation. At 31 days after planting the plant height, shoot dry matter, shoot protein content and root volume were evaluated. All isolates were able to fix nitrogen and produce IAA, with values ranging from 25.86 to 51.26mgNmL-1 and 107-1038µmolL-1, respectively. The inoculation of H. amplexicaulis and A. purpusii increased root volume and shoot dry matter. There were positive effects of Azospirillum inoculation on Mesosetum chaseae regarding plant height, shoot dry matter and root volume. Isolates MAY1, MAY3 and MAY12 were considered promising for subsequent inoculation studies in field conditions.

Nitrogen fixing efficiency and physiological characteristics of Azospirillum isolates from the paddy fields of North Bengal

The microbial activity of ten selected Azospirillum isolates was measured in terms of the amount of CO2 evolved by the isolates after incubation for 5 days which ranged from 6.88 to19.25 mg. Nitrogen fixing efficiency all of the isolates was determined by microkjeldhal method and the nitrogen fixing efficiency ranged from 10.03 to 13.11 mg N/g substrate. The growth of Azospirillum isolates was significant affected by different physiological factors such as pH, temperature and salinity. Most of the isolates showed optimum growth at pH 6.8, temperature 37°C and in the absence of NaCl. Azospirillum has the potential to be used as a substitute and or supplement of N-fertilizers. Further research is needed to estimate N-supplement potentials of biological nitrogen fixation (BNF) systems at the farm level.Jahangirnagar University J. Biol. Sci. 3(1): 47-53, 2014 (June)

Pertumbuhan Padi Varietas Ciherang Setelah Diinokulasi dengan Azospirillum Mutan Multifungsi

Modern agriculture is very closely related to the application of fertilizer to induce plants grow. The application of bio-fertilizers is expected to reduce the negative impact of chemical fertilizers. The purpose of this study was to determine the effect of multi-functional Azospirillum N 2 fixation, P solubility and IAA production on the growth of Ciherang rice in pot experiment in greenhouse BB Biogen. The experiment treatment were 3 types of inoculation (non-inoculation, inoculation using wildtype Aj Bandung 6.4.1.2 and the mutant isolate of AJM 3.7.1.14 ), and 4 levels of fertilizer application (non-fertilization, a quarter dose, a half dose, and the real dose of fertilization on rice in lowland). The Azospirillum isolates were used wildtype isolate Aj Bandung 6.4.1.2 and mutant isolate AJM 3.7.1.14 that was isolated and mutated using ethyl methanesulfonate (EMS) in BB Biogen. Seeds of Ciherang rice were inoculated using Azospirillum at cell density 10 6 cell/ml in different seedling tray. After 14 days, the seedlings were transferred to planting pots which consist of 3 plants per pot. Parameters observed were plant height, number of tillers, number of panicles per hill, wet and dry weight of panicles per hill, weight of 100 seeds, N and P content of the stover. The results showed that both wild- Azospirillum and mutant inoculum had no effect on the vegetative growth of Ciherang, but showed significant effect on the number of panicle per hill, grain weight per hill and dry weight of seeds per panicle. The use of Azospirillum and N fertilizer combination affected the growth and rice yields, also reduced chemical fertilizer application.

Characterization of Azospirillum and phosphosolubilizing bacterial isolate from salt-affected soil and their effect on rice (Oryza sativa) crop

The study was designed to isolate and characterize nutrient mobilizing soil microbes from salt affected soil of north Bihar. Out of 43 total 17 isolates of Azospirillum (12) and PSB (5) were selected in which, all the isolates produced one or the other different characteristics involved in plant growth promotion. They produced phytohormones like indole acetic acid, Phospho-solubilization, siderophore and N2 fixation. The observations were made with 21 treatments. The experiments on rice were carried in Randomized Block Design (RBD) with three replications. In the present investigation an attempt has been made to ascertain the effect of PGPR in different plant parameter such as tillers, effective tillers, plant height, panicle length, grain yield, straw yield and test weight.

Inoculación con Azospirillum spp y Enterobacter aglomerans en Pasto Guinea (Panicum maximum Jacq.) en el Departamento de Cesar (Colombia)

Se evaluó el efecto de los aislados SRGM2, SRGM3 y SRGM4 de Azospirillum spp. en el crecimiento y desarrollo de plantas de pasto Guinea (Panicum maximum Jacq cv. Tanzania) en la Estación Experimental Motilonia de Corpoica, del municipio Agustín Codazzi (Cesar, Colombia). Se realizó inoculación por imbibición de las semillas, y en el momento de la siembra, en las macetas en condiciones de casa malla. Se utilizó inoculación simple de los aislados de Azospirillum sp y co-inoculación con una cepa fosfato solubilizadora de Enterobacter aglomerans (UV1), aislada de suelos cultivados con algodón en el departamento de Cesar. Las variables estudiadas en las plantas fueron: germinación, peso foliar fresco y seco, peso radical fresco y seco, fósforo y proteína foliar. Los resultados mostraron que la inoculación simple y combinada promovió la germinación de semillas del pasto guinea (P≤0,05) con respecto al tratamiento sin inoculación (testigo absoluto); y aunque no se presentaron diferencias significativas (P≤0,05) en las demás variables estudiadas, se encontraron aumentos de hasta 26,80% en proteína cruda, y 45,67% en materia seca foliar comparada con las plantas tratadas con 100% de fertilización nitrogenada. Se observó que el aislado de Azospirillum SRGM2 co-inoculado con Enterobacter aglomerans UV1 registró los mayores valores en el crecimiento vegetal del pasto guinea, por lo que se consideró como una rizobacteria promisoria para la biofertilización de esta gramínea utilizada en la alimentación bovina en la costa caribe colombiana.

Phenotypic and molecular characterization of native Azospirillum strains from rice fields to improve crop productivity

Beneficial microorganisms have been considered as an important tool for crop improvement. Native isolates of Azospirillum spp. were obtained from the rhizospheres of different rice fields. Phenotypic, biochemical and molecular characterizations of these isolates led to the identification of six efficient strain of Azospirillum. PCR amplification of the nif genes (nifH, nifD and nifK) and protein profile of Azospirillum strains revealed inter-generic and inter-specific diversity among the strains. In vitro nitrogen fixation performance and the plant growth promotion activities, viz. siderophore, HCN, salicylic acid, IAA, GA, zeatin, ABA, NH3, phosphorus metabolism, ACC deaminase and iron tolerance were found to vary among the Azospirillum strains. The effect of Azospirillum formulations on growth of rice var. Khandagiri under field condition was evaluated, which revealed that the native formulation of Azospirillum of CRRI field (As6) was most effective to elevate endogenous nutrient content, and improved growth and better yield are the result. The 16S rRNA sequence revealed novelty of native Azospirillum lipoferum (As6) (JQ796078) in the NCBI database.

ANALYSIS OF GENETIC VARIABILITY IN SOURSOP (Annona muricata L.) POPULATIONS FROM CENTRAL JAVA AND EAST JAVA BASED ON RANDOM AMPLIFIED POLYMORPHIC DNA (RAPD) MARKER

Nitrogen fixing bacteria (Azospirillum and Azotobacter) were isolated from coastal mangrove in Pulau seribu. The aims of this experiment was to find out isolates of nitrogen fixing bacteria which were tolerant to high salinity. The isolates can be used as a biofertilizer to support coastal agriculture. A total of 28 isolates (14 isolates of Azospirillum and 14 isolates of Azotobacter) were tested their tolerance to salt by growing them in their respective media containing 1% 2% and 3 % NaCl,. Salt tolerant isolates obtained, then used as inoculants in paddy. The experiment laid out factorial based randomized complete block design which was comprised of 7 fertilizer treatments and 5 watering treatments with 5 replicates for each treatment at green house of Microbiology Division, Research Center of Biology, Indonesian Institute of Sciences. The first factor was fertilizer treatments 1. Compost + mixed bacteria (K1), 2. Control without inoculant, 3. Compost, 4. NPK, 5. Compost + NPK + mixed microbial, 6. Azotobacter isolates; 7. Azospirillum isolates. The second factor was watering treatments, plant was watered by : 1. freshwater, 2. freshwater + sea water at mixture ratio 1:1, 3. Sea water, 4. sea water + 2% NaCl (20 g NaCl / l), 5. freshwater + 5% NaCl (50 g NaCl / l). The result showed that there were 9 isolates of Azospirillum and 4 isolates of Azotobacter which were tolerant to grow at media with 3% NaCl. The green house experimental result revealed that the plants were treated with bacteria can survive up to the level of salinity 12.43 dS-1m.

THE EFFECT OF SALT TOLERANT NITROGEN FIXING BACTERIA ON THE GROWTH OF PADDY RICE (Oryza sativa. L)

Nitrogen fixing bacteria (Azospirillum and Azotobacter) were isolated from coastal mangrove in Pulau seribu. The aims of this experiment was to find out isolates of nitrogen fixing bacteria which were tolerant to high salinity. The isolates can be used as a biofertilizer to support coastal agriculture. A total of 28 isolates (14 isolates of Azospirillum and 14 isolates of Azotobacter) were tested their tolerance to salt by growing them in their respective media containing 1% 2% and 3 % NaCl,. Salt tolerant isolates obtained, then used as inoculants in paddy. The experiment laid out factorial based randomized complete block design which was comprised of 7 fertilizer treatments and 5 watering treatments with 5 replicates for each treatment at green house of Microbiology Division, Research Center of Biology, Indonesian Institute of Sciences. The first factor was fertilizer treatments 1. Compost + mixed bacteria (K1), 2. Control without inoculant, 3. Compost, 4. NPK, 5. Compost + NPK + mixed microbial, 6. Azotobacter isolates; 7. Azospirillum isolates. The second factor was watering treatments, plant was watered by : 1. freshwater, 2. freshwater + sea water at mixture ratio 1:1, 3. Sea water, 4. sea water + 2% NaCl (20 g NaCl / l), 5. freshwater + 5% NaCl (50 g NaCl / l). The result showed that there were 9 isolates of Azospirillum and 4 isolates of Azotobacter which were tolerant to grow at media with 3% NaCl. The green house experimental result revealed that the plants were treated with bacteria can survive up to the level of salinity 12.43 dS-1m.

Development and characterization of abiotic stress tolerant Azospirillum.

Abiotic stress is one of the most important environmental constraints that limit survival and productivity of staple crops like wheat and maize particularly in arid and semi-arid regions. Plant growth promoting bacteria ( Azospirillum ) are beneficial bacteria present in soil and forming associations with roots of plants. This investigation deals with the isolation and characterization of Azospirillum from maize roots. Azospirillum strain was identified by physiological, morphological and biochemical characters. These strains was identified based on morphological characters such as shape, and biochemical characters such as IMViC, urease were studied. Factors affecting growth of Azospirillum like temperature, salt stress, effect of pesticides and antibiotic tolerance were studied. The pot culture experiment was carried out to study the Azospirillum strain used on the growth of wheat plants. Salt stress has become an ever increasing threat to food production, irrigation being a major problem of agricultural fields due to gradual salinization.