Reliable multiplication of seed for NERICA varieties of rice, <Emphasis Type="Italic">Oryza sativa</Emphasis> L.

A comparison of breeder seed from different seed sources produced guidelines for multiplication of NERICA (New Rice for Africa) varieties. Lower percentage detection of offtypes was noted with seed from a breeder than with seed from the Genetic Resources Unit (GRU) of the Africa Rice Center. The GRU grows breeder seed in the field as bulk populations of varieties and this seed is multiplied for the foundation seed. Breeders, however, always cultivate the variety in each family with several lines. In the comparison, each NERICA variety was cultivated as a single plant per hill for easy offtype detection in the field. Any offtypes or doubtful plants were recorded and immediately discarded. The breeder seed was not always genetically uniform because it was not yet fixed genetically and included mechanical seed mixture as well as natural mutants. Offtypes were classified into categories such as chlorophyll mutants, aberrations of plant height, plant types and panicle types, semi- or high sterility, awnness, apiculous color, leaf sheath color and excessively early plants. The causes of these offtypes were mainly (1) mechanical seed mixture, (2) segregation in genetically-unfixed material, (3) out-crossing with another variety, or (4) natural mutation. In 2006, we multiplied foundation seed using used home-saved seed from the 2005 crop harvested after removal of offtypes. The detection percentage of offtypes decreased compared with the previous year, but the number of semi- or high sterility plants increased significantly. This review confirms that breeder seed must be maintained with line culture within a family.     

Effect of mineral fertilizer,pig manure,and <Emphasis Type="Italic">Azospirillum rugosum</Emphasis> on growth and nutrient contents of <Emphasis Type="Italic">Lactuca sativa</Emphasis> L.

Benefits from the application of plant growth-promoting bacteria in agriculture largely depend on the complex interactions between several factors including the nature of fertilizers selected. This study was designed to determine the fine tuning between the inoculated bacteria and different fertilizers and their effect on the growth of lettuce plants (Lactuca sativa L.). Plant growth promotion by a novel species of the genus Azospirillum, namely A. rugosum IMMIB AFH-6, was tested by biochemical, bioassay, and greenhouse studies. The treatments used in the greenhouse study were; unfertilized control (Blank), half recommended dose of chemical fertilizer (1/2CF), full recommended dose of chemical fertilizer (1CF), pig manure fertilizer (PMF), pig manure fertilizer + half recommended dose of chemical fertilizer (PMF + 1/2CF), and pig manure fertilizer + full recommended dose of chemical fertilizer (PMF + 1CF). All these treatments when inoculated with A. rugosum IMMIB AFH-6 inoculation were, respectively, In-Blank, In-1/2CF, In-1CF, In-PMF, In-PMF + 1/2CF, and In-PMF + 1CF. Significant increase in plant biomass and shoot N, P, Ca, and Fe was shown in the In-Blank treatment. Plant growth in soil amended with PMF and A. rugosum IMMIB AFH-6 was significantly lower than in soil treated with the chemical fertilizer, but inoculation combined with chemical fertilizer significantly elevated the plant biomass. The In-PMF + 1/2CF treatment showed the highest yield. A. rugosum IMMIB AFH-6 facilitated the accumulation of trace minerals in higher concentrations when PMF was combined with 1CF. To examine the benefits of inoculation by A. rugosum IMMIB AFH-6, we have proposed a new type of data analysis which considers both biomass and nutrient content of plants. This new type of analysis has shown the importance of the mineral content of plant.     

Spatial and temporal dynamics in genetic diversity in upland rice and late millet (<Emphasis Type="Italic">Pennisetum glaucum</Emphasis> (L.) R. Br.) in The Gambia

Ambiguity exists about the level of genetic diversity represented by farmer crop varieties, how it develops over time and how it relates to the diversity comprised by formal varieties. As part of an interdisciplinary technological/sociological study on farmer management of gene flow, upland rice (Oryza sativa L.) and late millet (Pennisetum glaucum (L.) R.Br.) from The Gambia were investigated for morphological and molecular variation. The goal of these analyses was to obtain insight into the level of crop genetic diversity of farmer’s materials planted in several case study villages in The Gambia. For both crops, samples were collected from villages and various research institutes. Based on variety names, different rice and millet varieties were expected to be used in different villages. In fact, there was a large overlap in genetic diversity for both crops, masked by the use of synonyms. The considerable similarity in rice genetic diversity between villages most likely results from the exchange of varieties between farmers. For millet this seems the result of development of varieties from the same gene pool. Some farmer varieties of rice, however, are apparent hybrid forms between the species O. sativa and O. glaberrima Steud., and farmer varieties in general displayed higher levels of genetic diversity than formal varieties. This indicates that, for rice, genetic diversity develops in farmers’ fields and may have potential use in formal breeding programs.     

Salt tolerance in <Emphasis Type="Italic">Zea mays</Emphasis> (L). following inoculation with <Emphasis Type="Italic">Rhizobium</Emphasis> and <Emphasis Type="Italic">Pseudomonas</Emphasis>

This study aimed to investigate the effect of inoculation with plant growth-promoting Rhizobium and Pseudomonas species on NaCl-affected maize. Two cultivars of maize (cv. Agaiti 2002 and cv. Av 4001) selected on the basis of their yield potential were grown in pots outdoors under natural conditions during July. Microorganisms were applied at seedling stage and salt stress was induced 21 days after sowing and maintained up to 50% flowering after 120 days of stress. The salt treatment caused a detrimental effect on growth and development of plants. Co-inoculation resulted in some positive adaptative responses of maize plants under salinity. The salt tolerance from inoculation was generally mediated by decreases in electrolyte leakage and in osmotic potential, an increase in osmoregulant (proline) production, maintenance of relative water content of leaves, and selective uptake of K ions. Generally, the microbial strain acted synergistically. However, under unstressed conditions, Rhizobium was more effective than Pseudomonas but under salt stress the favorable effect was observed even if some exceptions were also observed. The maize cv. Agaiti 2002 appeared to be more responsive to inoculation and was relatively less tolerant to salt compared to that of cv. Av 4001.     

The effect of rhizobiophages on<Emphasis Type="Italic"> Sinorhizobium meliloti</Emphasis> -<Emphasis Type="Italic"> Medicago sativa</Emphasis> symbiosis

Using Tn5 mutagenesis of lysogenic Sinorhizobium meliloti strain L5–30 (L54), three neomycin-resistant transconjugants differing in phage-resistance profiles were isolated. Two of them increased the dry weight of plants and were capable of establishing root nodules, whereas the third one was ineffective. The bacterium-phage interaction did not have observable consequences in the Medicago sativa - S. meliloti symbiosis because it did not affect the number of nodules on M. sativa or plant dry weight.     

<Emphasis Type="Italic">Rhizobium</Emphasis>,<Emphasis Type="Italic"> Bradyrhizobium</Emphasis> and<Emphasis Type="Italic"> Agrobacterium</Emphasis> strains isolated from cultivated legumes

The present study was conducted to isolate and characterize rhizobial strains from root nodules of cultivated legumes, i.e. chickpea, mungbean, pea and siratro. Preliminary characterization of these isolates was done on the basis of plant infectivity test, acetylene reduction assay, C-source utilization, phosphate solubilization, phytohormones and polysaccharide production. The plant infectivity test and acetylene reduction assay showed effective root nodule formation by all the isolates on their respective hosts, except for chickpea isolate Ca-18 that failed to infect its original host. All strains showed homology to a typical Rhizobium strain on the basis of growth pattern, C-source utilization and polysaccharide production. The strain Ca-18 was characterized by its phosphate solubilization and indole acetic acid (IAA) production. The genetic relationship of the six rhizobial strains was carried out by random amplified polymorphic DNA (RAPD) including a reference strain of Bradyrhizobium japonicum TAL-102. Analysis conducted with 60 primers discriminated between the strains of Rhizobium and Bradyrhizobium in two different clusters. One of the primers, OPB-5, yielded a unique RAPD pattern for the six strains and well discriminated the non-nodulating chickpea isolate Ca-18 from all the other nodulating rhizobial strains. Isolate Ca-18 showed the least homology of 15% and 18% with Rhizobium and Bradyrhizobium, respectively, and was probably not a (Brady)rhizobium strain. Partial 16S rRNA gene sequence analysis for MN-S, TAL-102 and Ca-18 strains showed 97% homology between MN-S and TAL-102 strains, supporting the view that they were strains of B. japonicum species. The non-infective isolate Ca-18 was 67% different from the other two strains and probably was an Agrobacterium strain.     

Origin of rice (<Emphasis Type="Italic">Oryza sativa</Emphasis> L.) domestication genes

A number of genes that contribute to the domestication traits of cultivated rice have been identified. These include Sh4, Rc, PROG1 and LABA1, which are associated with non-shattering rachis, white pericarp, erect growth and barbless awns, respectively. The mutations giving rise to the “domestication alleles” of these genes are either invariable in cultivated rice, or have variability that is strictly associated with the phenotypic trait. This observation forms the basis to those current rice domestication models that envisage a single origin for the domesticated phenotype. Such models assume that the domestication alleles are absent or rare in wild rice, emerged under cultivation and spread across all rice groups by introgressive hybridization. We examined whole-genome sequencing datasets for wild and cultivated rice to test the former two assumptions. We found that the rc and laba1 alleles occur in wild rice with broad geographical distribution, and reach frequencies as high as 13 and 15%, respectively. These results are in agreement with previous observations of the prog1 and sh4 domestication alleles in wild populations. We also show that the diversity of the genomic regions surrounding the rc, laba1, prog1 and sh4 alleles in wild accessions is greater than that in cultivated rice, suggesting that these alleles emerged prior to domestication. Our findings indicate that the possibility that independent rice groups obtained identical domestication alleles directly from the wild population needs to be considered.     

<Emphasis Type="Italic">Calamintha nepeta</Emphasis> (L.) Savi and <Emphasis Type="Italic">Micromeria thymifolia</Emphasis> (Scop.) Fritsch cultivated in Italy

Calamintha nepeta and Micromeria thymifolia have been traditionally used in the Mediterranean area as condiments and medicinal plants for a long time. Whereas in parts of Italy C. nepeta (special recipes have been developed in Lazio and Tuscany) is also an established garden plant showing different evolutionary products and their interaction among each other and the wild progenitor, M. thymifolia is being developed into a new crop plant. Both plants and their uses are described with regard to Italy. There is a marked tendency to broaden the use of condiments and spices which results in new crop plants which have to be documented and elaborated in further studies. Many species of Labiatae are predisposed to use by man and new items can be found even in areas which have to be considered as well studied.     

Nitrogen mineralization,N<Subscript>2</Subscript>O production and soil microbiological properties as affected by long-term applications of sewage sludge composts

A field study was conducted to investigate the long-term effect of surface application of sewage sludge composts vs chemical N fertilizer on total N, total C, soluble organic C, pH, EC, microbial biomass C and N, protease activity, deaminase activity, urease activity, gross and net rates of N mineralization and nitrification, CO₂ evolution, and N₂O production. Soil samples were taken from five depths (0–15, 15–20, 20–30, 30–40, and 40–50 cm) of a long-term experiment at the University of Tokyo, Japan. Three fields have been receiving sewage sludge composted with rice husk (RH), sawdust (SD), or mixed chemical fertilizer NPK (CF), applied at the rate of 240 kg N ha^–1 each in split applications in summer and autumn since 1978. Significantly higher amounts of total N and C and soluble organic C were found in the compost than in the CF treatments up to the 40-cm soil depth, indicating improved soil quality in the former. In the CF treatment, soil pH values were significantly lower and electrical conductivity values were significantly higher than those of compost-treated soils of up to 50 cm depth. Soil microbial biomass C and N, CO₂ evolution, protease, deaminase, and urease activities were significantly higher in the compost than in the CF treatments due to greater availability of organic substrates that stimulated microbial activity. Gross N mineralization rates determined by ¹⁵N dilution technique were eight and five times higher in the SD and RH treatments than in the CF treatment, respectively, probably due to high levels of microbial and enzyme activities. Net N mineralization rates were also significantly higher in the compost treatments and were negative in the CF treatment indicating immobilization. Net nitrification rates were higher in compost treatments and negative in the CF treatment. Nitrous oxide productions from compost treatments were higher than the CF treatment due to the greater availability of mineral N as a result of higher mineralization and nitrification rates and soluble organic C in the former. Most of the measured parameters were highest in the surface soil (0–15 cm) and were significantly higher in the SD treatment than in the RH treatment.     

<Emphasis Type="Italic">Ziziphus </Emphasis><Emphasis Type="Italic">spina-christi</Emphasis> (L.) Willd.: a multipurpose fruit tree

Neglected and underutilized species often play a vital role in securing food and livestock feed, income generation and energy needs of rural populations. In spite of their great potential little attention has been given to these species. This increases the possibility of genetic erosion which would further restrict the survival strategies of people in rural areas. Ziziphus spina-christi is a plant species that has edible fruits and a number of other beneficial applications that include the use of leaves as fodder, branches for fencing, wood as fuel, for construction and furniture making, and the utilization of different parts e.g. Fruits, leaves, roots and bark in folk medicine. Moreover, the plant is adapted to dry and hot climates which make it suitable for cultivation in an environment characterized by increasing degradation of land and water resources. Lack of research in Z. spina-christi hinders its successful improvement and promotion. Therefore, studies are needed to fully exploit this species. This article aims at summarizing information on different aspects of Z. spina-christi to stimulate interest in this crop which is of importance in Sudan and other countries of the semi-arid tropics.

Biological nitrogen fixation in maize (<Emphasis Type="Italic">Zea mays</Emphasis> L.) by <Superscript>15</Superscript>N isotope-dilution and identification of associated culturable diazotrophs

The nitrogen-fixing capacity of a range of commercial cultivars of maize (Zea mays L.) was evaluated by the ¹⁵N isotope-dilution method. Biological nitrogen fixation (BNF) expressed as percent nitrogen derived from air (Ndfa) ranged from 12 to 33 regardless of nitrogen fertilization. BNF was not affected by mineral nitrogen fertilization except on cultivar Topacio and PAU-871 cultivars. Subsequently, culturable bacterial diazotrophs were isolated from endophytic tissue of maize: seed, root, stem, and leaf. All isolates were able to grow on N-free semisolid medium. Eleven bacteria isolates showed nitrogen-fixing capacity by the reduction of acetylene to ethylene and confirmed by PCR the presence of nifH gene in their genome. Identification of the 11 isolates was performed by bacteriological methods, 16S rRNA gene sequences, and phylogenetic analysis, which indicated that the bacteria isolated were closely related to Pantoea, Pseudomonas, Rhanella, Herbaspirillum, Azospirillum, Rhizobium (Agrobacterium), and Brevundimonas. This study demonstrated that maize cultivars obtain significant nitrogen from BNF, varying by maize cultivar and nitrogen fertilization level. The endophytic diazotrophic bacteria isolated from root, stem, and leaf tissues of maize cultivars may contribute to BNF in these plants.     

Comparative effects of applying leguminous and non-leguminous green manures and inorganic N on biomass yield and nitrogen uptake in flooded rice (<Emphasis Type="Italic">Oryza sativa</Emphasis> L.)

The beneficial role of green manures in rice production is generally ascribed to their potential of supplying plant nutrients, particularly nitrogen (N). However, the mechanisms through which green manures enhance the crop productivity are poorly understood. Pot experiments were conducted using a ¹⁵N-tracer technique: (1) to compare the biomass production potential of sesbania (Sesbania aculeata Pers.) and maize (Zea mays L.) as green manuring crops for lowland rice and (2) to compare the effect of the two types of green manure and inorganic N on the dry matter accumulation and N uptake by two rice (Oryza sativa L.) cultivars, viz. IR-6 and Bas-370. Although maize produced three times higher shoot biomass compared with sesbania, the latter showed higher N concentration; and thus the total N yield was similar in the two types of plants. Applying the shoot material of the two plants to flooded rice significantly enhanced the dry matter yield and N uptake by the two rice cultivars, the positive effects generally being more pronounced with sesbania than with maize amendment. The difference in the growth-promoting potential of the two plant residues was related more to an increased uptake of the native soil N rather than to their direct role as a source of plant-available N. A positive added nitrogen interaction (ANI) was observed due to both plant residues, the effect was much more pronounced with the application of sesbania than with maize residues. In both rice cultivars, inorganic N also caused a substantial ANI, particularly at higher application rate. Losses from the applied N were 2–3 times lower from sesbania, compared with maize treatment. Green manuring with sesbania also caused much lower N losses than the inorganic N applied at equivalent or higher rates. The overall benefit of green manuring to rice plants was higher than inorganic N applied at comparable rates. The two rice cultivars differed in their response to green manuring, IR-6 generally being more responsive than Bas-370.     

N<Subscript>2</Subscript> fixation by faba bean (<Emphasis Type="Italic">Vicia faba</Emphasis> L.) in a gypsum-amended sodic soil

The response of faba bean to the application of four rates of gypsum (0, 2.5, 5.0, 10.0 t ha⁻¹) to a non-saline, alkaline sodic soil was measured in terms of grain yield, dry matter (DM) production, N accumulation and the proportional dependence of the legume on symbiotic N₂ fixation (P _atm). A yield-independent, time-integrated ¹⁵N-dilution model was used to estimate symbiotic dependence. A significant decrease in the exchangeable sodium percentage and significant increases in exchangeable Ca⁺⁺ and the Ca⁺⁺:Mg⁺⁺ ratio in the 0–10-cm soil layer were measured 30 months after application of 10 t ha⁻¹ gypsum. Despite low and erratic rainfall during crop growth, faba bean DM and N uptake responded positively to gypsum application. The symbiotic dependence of the legume at physiological maturity was little affected by sodicity (P _atm = 0.74 at zero gypsum and 0.81–0.82 at 2.5–10 t ha⁻¹ gypsum). The increase in fixed N due to gypsum application was mainly due to increases in legume DM and total N uptake. At 10 t ha⁻¹ of gypsum, faba bean fixed more than 200 kg N ha⁻¹ in above-ground biomass.     

Relative significance of soil and nitrogenous fertilizer in nitrogen nutrition and growth of wetland rice (Oryza sativa L.)

Summary A pot experiment was conducted to compare the yields from five commercially cultivated varieties (Bas-198, Bas-370, Bas-Pak, Bas-385, and IR-6) of rice (Oryza sativa L.) and to establish the relative significance of soil N and fertilizer N (¹⁵N-labelled ammonium sulphate) in affecting crop performance. Another aim was to study the interaction of fertilizer N and soil N as influenced by different rice varieties. Among the five varieties tested, Bas-Pak gave the maximum dry matter and N yield. The N-use efficiency (percentage of applied N taken up by the plants) of different varieties ranged between 33.7 and 43.7%, Bas-Pak being the most efficient. Significant losses of fertilizer N occurred from the soil-plant system. The maximum N loss (52.1% of applied N) was observed with IR-6 and the minimum loss (39.2%) with Bas-Pak. A substantial increase in the uptake of soil N following the application of fertilizer and an interaction between the two N sources were observed with all varieties except Bas-385. The interaction was attributed to greater root proliferation following the application of fertilizer. It was concluded that a varietey with greater potential to use soil N is likely to give a better yield and that, of the two N sources, the availability of soil N was more important in determining the yield performance of different varieties of rice.     

Exploring genetic diversity and potential novel disease resistance genes in a collection of rice (<Emphasis Type="Italic">Oryza</Emphasis> spp.) wild relatives

Wild relatives of rice (Oryza spp.) are an important source of novel resistance (R-)genes for rice improvement. Rice sheath blight, caused by Rhizotonia solani, and leaf blast, caused by Magnaportha oryzae, are major fungal diseases of rice worldwide. To identify novel R-genes, a group of Oryza spp. accessions represented by O. alta, O. australiensis, O. barthii, O. glaberrima, O. glumaepatula, O. latifolia, O. meridionolis, O. nivara, O. officinalis, and O. rufipogon, were evaluated for their reaction to leaf blast and sheath blight disease, and genotyped with 176 microsatellite (SSR) markers. Selected rice (O. sativa) accessions were included as reference. Cluster analysis performed with PowerMarker software using Rogers genetic distance and UPGMA, revealed most Oryza spp. accessions clustered with the same species or a closely related Oryza spp. Only a few Oryza spp. accessions grouped with the O. sativa accessions included as a reference. Analysis of this genotypic data in the software Structure revealed that the Oryza spp. accessions were assigned into eight different subpopulations and fit well into eight different backgrounds. Marker-trait associations between the SSR markers and disease reactions to blast and sheath blight were ascertained using the software TASSEL. Associations with blast disease were identified in ten different chromosomal regions and five of the ten were not located near known blast R-genes. Three associations were discovered with sheath blight disease and one was not near previously reported sheath blight QTL. These newly identified regions may represent novel R-genes that will be the basis future fine mapping studies. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Differential heterosis in a natural population of Asian wild rice (<Emphasis Type="Italic">Oryza rufipogon</Emphasis> Griff.) due to reproductive strategy and edge effect

Asian common wild rice (Oryza rufipogon) is a polycarpic perennial plant with a mixed inbreeder/outbreeder strategy. Seeds and clones are both reproductive components of the population. However, when collected in the flowering stage, the two differ substantially as to whether they have experienced natural selection directly or not. This study is aimed at evaluating mechanisms for the survival of a population by comparing genetic structure among subpopulations that are classified in terms of: (1) reproductive systems (clones or seeds); and also (2) location (fringe or inside). First, the genotypes determined at seven SSR (Simple Sequence Repeat) loci showed that parameters of observed heterozygosity (H_O) and outcrossing rate (t) were clearly higher in clones (H_O = 0.609, t = 82.4%) than in seeds (H_O = 0.346, t = 35.5%), although the two had approximately the same values for the number of alleles (A) and expected heterozygosity (H_E)(A = 4.43 in both clones and seeds, H_E = 0.709, 0.692 in clones and seeds, respectively). This result indicates that the individuals showing heterosis, with high numbers of heterozygous loci and outcrossing rates, are more likely to survive in the natural habitat. Secondly, parameters of observed heterozygosity and outcrossing rates were lower for seeds from the fringe area (H_O = 0.238, t = 24.6%) compared to seeds from the inside area (H_O = 0.443, t = 60.7%), although values were similar in clones of both the fringe and inside areas. This result suggests that the edge effect might be due mainly to the restriction of wind strength in fringe area of the forested swamp.     

The Genetic Resources of European Sweet Chestnut (<Emphasis Type="Italic">Castanea sativa</Emphasis> Miller) in Andalusia,Spain

Sweet chestnut (Castanea sativa Miller) is the only native species of the genus Castanea in Europe. A collecting mission has been carried out in Andalusia (South of Spain) with the purpose to evaluate of the genetic variability of this species in this region. During the field trips, two different types of chestnut trees have been detected: one of clonal nature (varieties obtained by graft) and other of sexual nature (trees derived from nut). For the former, 156 trees were located in plantations in the Genal river Valley (Malaga) and the Natural Park of `Sierra de Aracena y Picos de Aroche' (Huelva), where 43 varietal denominations were recorded. Thirty out of them had never been mentioned. For the latter, 29 stands were located in Almería, Córdoba, Granada, Jaén, and Málaga. This work reveal that this species present a high degree of genetic variability. Nevertheless, their preservation is seriously threatened in the most zones due to the varietal substitution, the low profit or the abandonment, which determinate the urgent necessity to safeguard these genetic resources.     

Effect of methods of application on the efficiency of urea broadcast onto lowland rice (Oryza sativa L.)

Summary We evaluated the effect of different methods of application on the efficiency of urea broadcast at a rate of 100 kg N ha^-1 onto lowland rice (Oryza sativa L. var. SPR 60) in a field experiment conducted on a Phimai soil (Fluvic Tropaquepts) during the dry season of 1989. Analysis of the floodwater on the first day after the fertilizer application showed a high initial concentration of urea-N. Addition of the urease inhibitor N-(n-butyl) thiophosphoric triamide (nBTPT), broadcast with the urea into the floodwater, caused an apparent reduction in the rate of urea disappearance and a subsequent accumulation of NH₃–N in the floodwater; this ureas inhibitor also suppressed the rise in floodwater pH, with a resultant reduction in the partial pressure of ammonia (pNH₃) compared with the unamended urea application. The use of nBTPT did not decrease the N loss from broadcast urea not did it increase the grain yield. Among the different methods of applying broadcast urea that we tested, the broadcast application of granular urea onto drained soil shortly after removing floodwater followed by flooding 2 days later appeared to be a good N management practice, offering considerable potential for improving the efficiency of urea applied to lowland rice crops.     

A bradyrhizobial-<Emphasis Type="Italic">Penicillium</Emphasis> spp. biofilm with nitrogenase activity improves N<Subscript>2</Subscript> fixing symbiosis of soybean

A bradyrhizobial-fungal biofilm (i.e. Bradyrhizobium elkanii SEMIA 5019-Penicillium spp.) developed in vitro was assayed for its nitrogenase activity and was evaluated for N₂-fixing symbiosis with soybean under greenhouse conditions. The biofilm showed nitrogenase activity, but the bradyrhizobial strain alone did not. Shoot and root growth, nodulation and N accumulation of soybean increased significantly with an inoculum developed from the biofilm. This study concludes that such biofilmed inoculants can improve N₂-fixing symbiosis in legumes, and can also directly contribute to soil N fertility in the long term. Further studies should be conducted to investigate the performance of these inoculants under field conditions.