OsARF16, a transcription factor,is required for auxin and phosphate starvation response in rice (Oryza sativa L.)

Plant responses to auxin and phosphate (Pi) starvation are closely linked. However, the underlying mechanisms connecting auxin to phosphate starvation (?Pi) responses are largely unclear. Here, we show that OsARF16, an auxin response factor, functions in both auxin and ?Pi responses in rice (Oryza sativa L.). The knockout of OsARF16 led to primary roots (PR), lateral roots (LR) and root hair losing sensitivity to auxin and ?Pi response. OsARF16 expression and OsARF16::GUS staining in PR and LR of rice Nipponbare (NIP) were induced by indole acetic acid and ?Pi treatments. In ?Pi conditions, the shoot biomass of osarf16 was slightly reduced, and neither root growth nor iron content was induced, indicating that the knockout of OsARF16 led to loss of response to Pi deficiency in rice. Six phosphate starvation‐induced genes (PSIs) were less induced by ?Pi in osarf16 and these trends were similar to a knockdown mutant of OsPHR2 or AtPHR1, which was a key regulator under ?Pi. These data first reveal the biological function of OsARF16, provide novel evidence of a linkage between auxin and ?Pi responses and facilitate the development of new strategies for the efficient utilization of Pi in rice.     

Effect of brassinosteroids on salinity stress induced inhibition of seed germination and seedling growth of rice (Oryza sativa L.)

The effect of 24-epibrassinolide and 28-homobrassinolide on the inhibitionof germination and seedling growth of rice (Oryza sativa) induced bysalinity stress was studied. Brassinosteroids were found to reverse theinhibitory effect on germination and seedling growth. The activation ofseedling growth by brassinosteroids under salinity stress was associatedwith enhanced levels of nucleic acids and soluble proteins.     

Relative proportion of antioxidative enzyme activity in locally grown Indian rice cultivars (Oryza sativa L.) under submergence condition

Abstract

Breeders are trying to develop true submergence-tolerant genotypes for lowland irrigated and/or rainfed areas, where rice is often affected by short-term submergence. This study was conducted to evaluate the submergence tolerance, response of antioxidative defense systems to hypoxic and re-aerated conditions, and growth and survival percentage among 19 rice genotypes. The rice seedlings submerged into concrete tanks for 10 days exhibited decreased activities of antioxidant enzymes. The antioxidant enzymes, superoxide dismutase, catalase, and ascorbate peroxidase increased, whereas polyphenol oxidase and glutathione reductase decreased in subsequent re-aerated situation in tolerant genotypes. On the other hand, the activities of enzymes decreased greatly in susceptible genotypes under hypoxic and re-aerated conditions. Complete submergence of the seedling increased the plant height of 12 genotypes. However, it decreased the dry matter production. Genotypes having better root and shoot growth maintained higher dry weight and exhibited better survival than did others. The results indicated that genotypes tolerant of flash flood maintained higher activities of antioxidant enzymes and dry matter accumulation to protect against postanoxic injury.     

Responses of rice (Oryza sativa L.) genotypes to different levels of submergence

The study aims at identifying some submergence-tolerant rice genotypes through morphological and molecular characterization and their genetic variability analysis. Ten rice genotypes including two submergence-tolerant checks, two susceptible varieties and six advanced lines were evaluated for submergence tolerance in the laboratory and in the field during January–December 2015. The experiment was conducted in the field following randomized complete block design in a two-factor arrangement using five replications. Ten characters, viz. days to flowering, plant height, tiller number plant^?1, effective tiller plant^?1, and yield plant^?1 etc. were studied for four treatments. A significant genotype × environment interaction was observed for all traits studied in this experiment. The yield was reduced for all genotypes at a different level of submergence stress compared to control. Binadhan-11, Binadhan-12, RC 249 and RC 251 showed tolerance, whereas RC 192, RC 193 and RC 225 showed moderate tolerance in submerged condition. The phenotypic coefficient of variance (PCV) was higher than the genotypic coefficient of variance (GCV) in all the studies traits. High heritability (75–97%) was found for all traits. High heritability along with high genetic advance was found for days to flowering (45.55) and plant height (40.05). Molecular characterization of the used genotypes was done with three SSR markers viz. RM 24, and submergence specific SC3 and SUB1. SC3 was found reliable for detection of submergence tolerant genotypes due to the highest gene diversity (0.840) compared to others. The banding pattern of the submergence specific markers SC3 and SUB1 identified in Binadhan-11, Binadhan-12, RC 192, RC 193, RC 225, RC 227, RC 249, and RC 251, which possess the SUB1 gene. Finally, clustering also separates the tolerant genotypes from the susceptible by dividing them into different clusters. The identified genotypes might be useful for the breeding programme for the development of submergence tolerant as well as resistant rice variety in Bangladesh.     

Relative toxicity of arsenite and arsenate on germination and early seedling growth of rice (Oryza sativa L.)

Elevated soil arsenic levels resulting from long-term use of arsenic contaminated ground for irrigation in Bangladesh may inhibit seed germination and seedling establishment of rice, the country's main food crop. A germination study on rice seeds and a short-term toxicity experiment with different concentrations of arsenite and arsenate on rice seedlings were conducted. Percent germination over control decreased significantly with increasing concentrations of arsenite and arsenate. Arsenite was found to be more toxic than arsenate for rice seed germination. There were varietal differences among the test varieties in response to arsenite and arsenate exposure. The performance of the dry season variety Purbachi was the best among the varieties. Germination of Purbachi was not inhibited at all up to 4 mg l^–1 arsenite and 8 mg l^–1 arsenate treatment. Root tolerance index (RTI) and relative shoot height (RSH) for rice seedlings decreased with increasing concentrations of arsenite and arsenate. Reduction of RTI caused by arsenate was higher than that of arsenite. In general, dry season varieties have more tolerance to arsenite or arsenate than the wet season varieties.     

Morphological and physiological characteristics of a root-hairless mutant in rice (Oryza sativa L.)

This paper reports morphological and physiological characteristics of a first root-hairless mutant (RH2) of rice (Oryza sativa L.), which can be useful in advancing knowledge on the role of root hairs in water and nutrient uptake, and genetics of root hairs. The mutant was selected among NaN₃ mutagenized progeny of the rice cultivar Oochikara. Microscopic observations showed absence of root hairs in RH2. At the seedling stage, RH2 showed shorter seedling height and shorter roots compared to the wild type variety Oochikara. Because of the differences in seedling growth, all comparisons between Oochikara and RH2 in uptake-related characters were made on the basis of values adjusted by the dry weight of either the shoot or the root. When grown at low water potential in soil, Oochikara and RH2 were similar in shoot water content and transpiration per unit shoot dry weight, and similarly, at low water potential in solution culture, there was no significant difference between Oochikara and RH2 in transpiration per unit shoot dry weight. These results suggest that at the seedling stage, root hairs do not significantly contribute to uptake of water. In solution culture, Oochikara and RH2 did not significantly differ in phosphate uptake per unit root dry weight. This result supports the previous work that root hairs do not contribute to phosphate uptake in solution culture. Regarding to response to plant hormones, RH2 showed a higher level of resistance to two synthetic auxins, 2,4-dichlorophenoxyacetic acid (2,4-D) and 1-naphthaleneacetic acid (NAA) than Oochikara. NAA treatment induced very short root hairs in RH2, suggesting that the absence of root hairs in RH2 may be due to a shortage of endogenous auxin. Genetic analysis showed that the root hairless character in RH2 is inherited as a single recessive gene.     

铜绿假单胞菌对镉胁迫苗期水稻根系活力及叶片生理特性的影响

为揭示耐镉铜绿假单胞菌缓解镉胁迫水稻的生理效应,以无镉处理为对照,通过添加菌液、空载体、菌剂及20μmol·L^-1 Cd进行水培试验,分析了菌株对苗期水稻根系活力及叶片生理特性的影响.结果表明:镉胁迫显著抑制了水稻的根系活力,降低了叶片光合效率、抗氧化酶活性及可溶性蛋白、类黄酮与总酚含量,提高了叶片丙二醛(MDA)和超氧阴离子(O2-)含量.与镉处理相比,添加菌液、菌剂处理的水稻根系活力分别提高了36.1%~42.5%、49.4%~53.0%;叶片净光合速率提高了118.5%~147.1%、137.6%~156.9%;可溶性蛋白含量提高了37.0%~49.3%、37.7%~72.6%.菌剂处理的水稻叶片超氧化物歧化酶(SOD)、过氧化物酶(POD)、过氧化氢酶(CAT)活性分别比Cd处理提高了36.9%~42.6%、82.7%~92.6%、43.3%~52.2%,菌液处理的SOD、POD、CAT则分别比Cd处理提高了25.8%~36.6%、40.9%~55.9%、24.0%~29.2%,菌剂对水稻叶片抗氧化酶的促进效应显著高于菌液;菌剂、菌液处理的水稻叶片MDA含量分别比Cd处理降低了44.8%~54.7%、29.4%~41.9%;O2-含量减少了9.9%~10.2%、3.0%~7.1%;菌剂处理后类黄酮、总酚含量分别比Cd处理提高了125.4%~135.7%、100.8%~119.4%;菌液处理后则分别提高了139.4%~146.7%、115.0%~134.7%.可见,铜绿假单胞菌及其菌剂通过提高苗期水稻根系活力、光合作用促进了苗期水稻的生长.铜绿假单胞菌通过增强水稻抗氧化酶活性、提高类黄酮和总酚等抗氧化物质含量,表现出显著的缓解镉胁迫效应.     

Mechanism associated with nonstructural carbohydrate accumulation in submergence tolerant rice (Oryza sativa L.) cultivars

Nonstructural carbohydrate (NSC) accumulation in submergence tolerant rice cultivars (cv) was studied in six Indica rice [Oryza sativa (L.)] cv under control and simulated submerged conditions. Tolerant cultivars accumulated greater contents of NSC compared to the susceptible cultivars. Starch and total NSC content showed significant positive association with survival percentage. On the other hand, elongation due to submergence was significantly a negative association with survival. The CO₂ photosynthetic rate, chlorophyll content, maximum photochemical efficiency of PS II (Fv/Fm), and activities of Rubisco were not significantly different between tolerant and susceptible cv under control condition. The ADP glucose pyrophosphorylase (AGPPase) activity was significantly higher in the tolerant cv and was a positive association with starch/NSC, whereas Fructose 1,6-diphosphatase (FDPase) activity was significantly higher in susceptible cv compared to tolerant cv and was a negative association with starch/NSC. Greater activities of AGPPase along with lower activities of FDPase might facilitate greater accumulation of NSC in tolerant rice cultivars.     

Heterosis breeding in rice (Oryza sativa L.)

Summary Studies conducted at the International Rice Research Institute (IRRI) during 1980 and 1981 have shown up to 73% heterosis, 59% heterobeltiosis and 34% standard heterosis for yield in rice. The latter was estimated in comparison to commercial varieties: IR36 and IR42 (yield 4–5 t/ha in wet season trials and 7–8 t/ha in dry season trials). Generally speaking, absolute yield was lower and extent of standard heterosis was higher in wet season than in dry season with some exception. Yields up to 5.9 t/ha (22% standard heterosis) in the wet season and 10.4 t/ha (34% standard heterosis) in the dry season were obtained. Most of the hybrids performed better in some season while some performed better in both seasons. Hybrids showed better lodging resistance although they were 5–10 cm taller. F₁ hybrids had significant positive correlations with the parental traits viz., yield (r = 0.446), tillering (r = 0.746), height (r = 0.810) and flowering (r = 0.843). Selection of parents among elite breeding lines on the basis of their per se yield performance, diverse origin and resistance to insects and diseases should give heterotic combination. Yield advantage of hybrids was due primarily to increase in number of spikelets per unit area even though tiller number was reduced. Grain weight was either the same or slightly higher. High yielding hybrids also showed significant heterosis and heterobeltiosis for total dry matter and harvest index. For commercial utilization of heterosis in rice, effective male sterility and fertility restoration systems are available and up to 45% natural outcrossing on male sterile lines has been observed. Consequently, F₁ rice hybrid have been successfully developed and used in China. Prospects of developing hybrid rice varieties elsewhere appear bright especially in countries that have organized seed production, certification and distribution programs and where hybrid seed can be produced at a reasonable cost.     

QTLs and epistasis for seminal root length under a different water supply in rice (Oryza sativa L.)

To identify the genetic background of seminal root length under different water-supply conditions, a recombinant inbred (RI) population consisting of 150 lines, derived from a cross between an indica lowland rice, IR1552, and a tropical japonica upland rice, Azucena, was used in both solution culture (lowland condition) and paper culture (upland condition). Quantitative trait loci (QTLs) and epistatic loci for seminal root length were analyzed using 103 restriction fragment length polymorphism (RFLP) markers and 104 amplified fragment length polymorphism (AFLP) markers mapped on 12 chromosomes based on the RI population. One QTL for seminal root length in solution culture (SRLS) and one for seminal root length in paper culture (SRLP) were detected on chromosomes 8 and 1, and about 11% and 10% of total phenotypic variation were explained, respectively. The QTL for SRLP on chromosome 1 was very similar with the QTL for the longest nodal root referred to in a previous report; this QTL may be phenotypically selectable in a breeding program using paper culture. Five pairs of epistatic loci for SRLS were detected, but only one for SRLP, which accounted for about 60% and 20% of the total variation in SRLS and SRLP, respectively. The results indicate that epistasis is a major genetic basis for seminal root length, and there is a different genetic system responsible for seminal root growth under different water supply conditions. Received: 26 May 2000 / Accepted: 19 October 2000     

Effects of 24-epibrassinolide on seed germination, seedling growth, lipid peroxidation, proline content and antioxidative system of rice (Oryza sativa L.) under salinity stress

The effects of 24-epibrassinolide (24-epiBL) on seedling growth, antioxidative system, lipid peroxidation, proline and soluble protein content were investigated in seedlings of the salt-sensitive rice cultivar IR-28. Seedling growth of rice plants was improved by 24-epiBL treatment under salt stress conditions. When seedlings treated with 24-epiBL were subjected to 120 mM NaCl stress, the activities of superoxide dismutase (EC 1.15.1.1), catalase (EC 1.11.1.6) and glutathione reductase (EC 1.6.4.2) did not show significant difference, whereas the activity of ascorbate peroxidase (EC 1.11.1.11) significantly increased. Increased activity of peroxidase (EC 1.11.1.7) under NaCl stress showed remarkable decrease in the 24-epiBL+NaCl-applied group. Lipid peroxidation level significantly increased under salt stress but decreased with 24-epiBL application revealing that less oxidative damage occurred in this group (24-epiBL+NaCl). In addition, increased proline content in the NaCl-applied group was decreased by 24-epiBL application in the 24-epiBL+NaCl-applied group. Soluble protein content was increased by 24-epiBL application even under NaCl stress, being also higher than control conditions (no 24-epiBL or NaCl treatment). 24-epiBL treatment considerably alleviated oxidative damage that occurred under NaCl-stressed conditions and improved seedling growth in part under salt stress in sensitive IR-28 seedlings.     

Genetic diversity and association mapping of seed vigor in rice (Oryza sativa L.)

Key message

Twenty-seven QTLs were identified for rice seed vigor, in which 16 were novel QTLs. Fifteen elite parental combinations were designed for improving seed vigor in rice.

Abstract

Seed vigor is closely related to direct seeding in rice (Oryza sativa L.). Previous quantitative trait locus (QTL) studies for seed vigor were mainly derived from bi-parental segregating populations and no report from natural populations. In this study, association mapping for seed vigor was performed on a selected sample of 540 rice cultivars (419 from China and 121 from Vietnam). Population structure was estimated on the basis of 262 simple sequence repeat (SSR) markers. Seed vigor was evaluated by root length (RL), shoot length (SL) and shoot dry weight in 2011 and 2012. Abundant phenotypic and genetic diversities were found in the studied population. The population was divided into seven subpopulations, and the levels of linkage disequilibrium (LD) ranged from 10 to 80 cM. We identified 27 marker–trait associations involving 18 SSR markers for three traits. According to phenotypic effects for alleles of the detected QTLs, elite alleles were mined. These elite alleles could be used to design parental combinations and the expected results would be obtained by pyramiding or substituting the elite alleles per QTL (apart from possible epistatic effects). Our results demonstrate that association mapping can complement and enhance previous QTL information for marker-assisted selection and breeding by design.     

Physiological and biochemical changes related to methyl jasmonate-induced chilling tolerance of rice (Oryza sativa L.) seedlings

Physiological and biochemical changes related to methyl jasmonate (MeJA)-induced chilling tolerance of rice (Oryza sativa L. cv. Taichung Native 1) seedlings were investigated. Treatment of whole plants with 10 mmol m^?3 MeJA for 48 h before chilling (5 °C) was optimal for the induction of chilling tolerance. MeJA greatly improved the survival ratio of chilled seedlings and ameliorated chilling injury such as demolition of membrane structure (estimated by electrolyte leakage). MeJA also prevented water loss in chilled seedlings by reducing the opening of stomata and decreasing the root bleeding rate. Putrescine and spermine levels in shoots increased but spermidine levels decreased on exposure to MeJA. In roots, putrescine levels also increased and spermidine levels increased transiently on exposure to MeJA. Activities of arginine decarboxylase (ADC; EC 4.1.1.19) and S-adenosylmethionine decarboxylase (SAMDC; EC 4.1.1.50) in both shoots and roots increased on exposure to MeJA, while the activity of ornithine decarboxylase (ODC; EC 4.1.1.17) remained unchanged. The MeJA-induced putrescine increase was inhibited by 50 mmol m^?3α-difluoromethylarginine (DFMA), an irreversible inhibitor of ADC, but not by 50 mmol m^?3α-difluoromethylornithine (DFMO), an irreversible inhibitor of ODC. The effect of MeJA on the induction of chilling tolerance was also reduced by 50 mmol m^?3 DFMA. The effects of DFMA were partly prevented by 1 mol m^?3 putrescine. This indicates that putrescine accumulation is required for the induction of chilling tolerance of rice seedlings by MeJA.     

Structural homology between semidwarfism-related proteins and glutelin seed protein in rice (Oryza sativa L.)

Summary Two semidwarfism-related proteins, SRP-1 and SRP-2, were detected as major spots in a long-culm rice cultivar, Norin 29 and its semidwarf near-isogenic line, SC-TN1, respectively, by two-dimensional gel electrophoresis (2D-PAGE). The testcross showed that SRP-1 and SRP-2 are controlled by codominant alleles, Srp-1 and Srp-2, respectively, at a single locus Srp. This locus was considered to be closely linked with the semidwarfing gene locus sd-1. SRP-1 and SRP-2 were separated by 2D-PAGE, electroblotted onto a polyvinylidene difluoride membrane, and sequenced by a gas-phase protein sequencer. The N-terminal amino acid sequences, however, could not be determined due to the blockage of the N-terminals of these proteins. After removal of the N-terminal residue with pyroglutamyl peptidase given to the membrane, the amino acid sequence in the N-terminal region was determined. The N-terminal and internal amino acid sequences of SRP-1 and SRP-2 were highly homologous with those of the glutelin -subunits of seed endosperm storage protein, which were deduced by the cDNA sequences. In the seed endosperms of Norin 29 and SC-TN1, a total of eight glutelin -subunits was identified by 2D-PAGE. The amino acid sequences in the N-terminal and internal regions of these proteins were determined. This experiment confirmed that SRP-1 and SRP-2 are almost identical in structure with the glutelin _5a- and _5b-subunits, respectively, which were identified in several organs such as endosperms, embryos, and leaves, unlike the other glutelin -subunits.     

OsMOGS is required for N‐glycan formation and auxin‐mediated root development in rice (Oryza sativa L.)

N‐glycosylation is a major modification of glycoproteins in eukaryotic cells. In Arabidopsis, great progress has been made in functional analysis of N‐glycan production, however there are few studies in monocotyledons. Here, we characterized a rice (Oryza sativa L.) osmogs mutant with shortened roots and isolated a gene that coded a putative mannosyl‐oligosaccharide glucosidase (OsMOGS), an ortholog of α‐glucosidase I in Arabidopsis, which trims the terminal glucosyl residue of the oligosaccharide chain of nascent peptides in the endoplasmic reticulum (ER). OsMOGS is strongly expressed in rapidly cell‐dividing tissues and OsMOGS protein is localized in the ER. Mutation of OsMOGS entirely blocked N‐glycan maturation and inhibited high‐mannose N‐glycan formation. The osmogs mutant exhibited severe defects in root cell division and elongation, resulting in a short‐root phenotype. In addition, osmogs plants had impaired root hair formation and elongation, and reduced root epidemic cell wall thickness due to decreased cellulose synthesis. Further analysis showed that auxin content and polar transport in osmogs roots were reduced due to incomplete N‐glycosylation of the B subfamily of ATP‐binding cassette transporter proteins (ABCBs). Our results demonstrate that involvement of OsMOGS in N‐glycan formation is required for auxin‐mediated root development in rice.     

qLTG-9, a stable quantitative trait locus for low-temperature germination in rice (Oryza sativa L.)

Low-temperature germination (LTG) is an important agronomic trait for direct seeding of rice in temperate regions of East Asia. To dissect the genetic control of LTG, we constructed a recombinant inbred line (RIL) population derived from a cross of japonica variety USSR5 and indica variety N22. Three putative QTL involved in LTG were detected and named qLTG-7, qLTG-9 and qLTG-12. They explained 9.5, 12.12 and 7.08 % of the phenotypic variation, respectively, and the alleles from USSR5 enhanced LTG. A set of advanced backcross lines selected for the presence of qLTG-9 (with the biggest contribution of the three QTL), by both linked markers and phenotype, was used to validate qLTG-9 in different generations, years and locations. A near-isogenic line in USSR5 background with a qLTG-9 insertion from N22 had retarded germination under low-temperature conditions. Finally, qLTG-9 was fine mapped between markers L9-25D and ID-1, to a 72.3-kb region in chromosome 9, which in the Nipponbare genome contains five predicted genes. This result provides a springboard for map-based cloning of qLTG-9 and is helpful in understanding the mechanism of seed germination under low-temperature conditions.     

Root aquaporins contribute to whole plant water fluxes under drought stress in rice (Oryza sativa L.)

Aquaporin activity and root anatomy may affect root hydraulic properties under drought stress. To better understand the function of aquaporins in rice root water fluxes under drought, we studied the root hydraulic conductivity (Lpr) and root sap exudation rate (Sr) in the presence or absence of an aquaporin inhibitor (azide) under well‐watered conditions and following drought stress in six diverse rice varieties. Varieties varied in Lpr and Sr under both conditions. The contribution of aquaporins to Lpr was generally high (up to 79% under well‐watered conditions and 85% under drought stress) and differentially regulated under drought. Aquaporin contribution to Sr increased in most varieties after drought, suggesting a crucial role for aquaporins in osmotic water fluxes during drought and recovery. Furthermore, root plasma membrane aquaporin (PIP) expression and root anatomical properties were correlated with hydraulic traits. Three chromosome regions highly correlated with hydraulic traits of the OryzaSNP panel were identified, but did not co‐locate with known aquaporins. These results therefore highlight the importance of aquaporins in the rice root radial water pathway, but emphasize the complex range of additional mechanisms related to root water fluxes and drought response.     

Azospirillum amazonense inoculation: effects on growth, yield and N2 fixation of rice (Oryza sativa L.)

Bacteria of the genus Azospirillum are considered to be plant-growth promoting bacteria (PGPR) and stimulate plant growth directly either by synthesising phyto-hormones or by promoting nutrition by the process of biological nitrogen fixation (BNF). Although this genus extensively studied, the effects of inoculation and the possible BNF contribution of the Azospirillum amazonense specie are very scarce. The aim of this study was to isolate, characterise and evaluate auxin production and nitrogenase activity of this species and to select, by inoculation of rice plants, promising isolates based on their ability to improve plant growth, yield and the BNF contribution. One hundred and ten isolates obtained from rice were characterised and grouped according to colony features. Forty-two isolates, confirmed as A. amazonense by the fluorescent in situ hybridization (FISH) technique, were tested for auxin production and nitrogenase activity in vitro. Subsequently plant growth promotion related to plant nutrition effect was evaluated, in vitro and in greenhouse experiments. The BNF contribution to plant growth was evaluated using the ¹⁵N isotope dilution technique. All A. amazonense strains tested produced indoles, but only 10% of them showed high production, above 1.33 μM mg protein⁻¹. The nitrogenase activity also was variable and only 9% of isolates showed high nitrogenase activity and the majority (54%) exhibited a low potential. The inoculation of selected strains in rice under gnotobiotic conditions reduced the growth of root and aerial part when compared to the control, showing the negative effects of excess of phytohormone accumulation in the medium. However, in the greenhouse experiment, inoculation of strains of A. amazonense increased grain dry matter accumulation (7 to 11.6%), the number of panicles (3 to 18.6%) and nitrogen accumulation at grain maturation (3.5 to 18.5%). BNF contributions up to 27% were observed for A. amazonense Y2 (wild type strain). The data presented here is the first report that the PGPR effect of A. amazonense for rice plants grown under greenhouse conditions is mainly due the BNF contribution as measured by ¹⁵N isotope dilution technique, in contrast to the hormonal effect observed with other Azospirillum species studied.