Rubens José Campo

Genome of Herbaspirillum seropedicae Strain SmR1, a Specialized Diazotrophic Endophyte of Tropical Grasses

The molecular mechanisms of plant recognition, colonization, and nutrient exchange between diazotrophic endophytes and plants are scarcely known. Herbaspirillum seropedicae is an endophytic bacterium capable of colonizing intercellular spaces of grasses such as rice and sugar cane. The genome of H. seropedicae strain SmR1 was sequenced and annotated by The Paraná State Genome Programme—GENOPAR. The genome is composed of a circular chromosome of 5,513,887 bp and contains a total of 4,804 genes. The genome sequence revealed that H. seropedicae is a highly versatile microorganism with capacity to metabolize a wide range of carbon and nitrogen sources and with possession of four distinct terminal oxidases. The genome contains a multitude of protein secretion systems, including type I, type II, type III, type V, and type VI secretion systems, and type IV pili, suggesting a high potential to interact with host plants. H. seropedicae is able to synthesize indole acetic acid as reflected by the four IAA biosynthetic pathways present. A gene coding for ACC deaminase, which may be involved in modulating the associated plant ethylene-signaling pathway, is also present. Genes for hemagglutinins/hemolysins/adhesins were found and may play a role in plant cell surface adhesion. These features may endow H. seropedicae with the ability to establish an endophytic life-style in a large number of plant species.

Nitrogen nutrition of soybean in Brazil: Contributions of biological N2 fixation and N fertilizer to grain yield

There has recently been concern in Brazil whether biological N2 fixation (BNF) is capable of meeting the increased N needs of newly released more productive cultivars, as well as doubts about the advantages of annual reinoculation of seeds. Forty experiments were performed over 3 yr in oxisols containing at least 103 cells of Bradyrhizobium g-1 in the State of Parana, southern Brazil to estimate the contributions of BNF and of N fertilizer. The experiments were performed at two sites, Londrina and Ponta Grossa, under conventional (CT) or no-tillage (NT) systems, with two cultivars [Embrapa 48 (early-maturing) or BRS 134 (medium-maturity group)]. Treatments included non-inoculated controls without or with 200 kg of N ha-1, and inoculation without or with N fertilizer applied at sowing (30 kg of N ha-1), or at the R2 or R4 stage (50 kg of N ha-1). Compared with the non-inoculated control, reinoculation significantly increased the contribution of BNF estimated by the N-ureide technique (on average from 79 to...

Genetic diversity of indigenous tropical fast-growing rhizobia isolated from soybean nodules

This study characterized genetically 30 fast-growing rhizobial strains isolated from nodules of Asian and modern soybean genotypes that had been inoculated with soils from disparate regions of Brazil. Analyses by rep-PCR (ERIC and REP) and RAPD indicated a high level of genetic diversity among the strains. The RFLP-PCR and sequencing analysis of the 16S rRNA genes indicated that none of the strains was related to Sinorhizobium (Ensifer) fredii, whereas most were related to Rhizobium tropici (although they were unable to nodulate Phaseolus vulgaris) and to Rhizobium genomic species Q. One strain was related to Rhizobium sp. OR 191, while two others were closely related to Agrobacterium (Rhizobium) spp.; furthermore, symbiotic effectiveness with soybean was maintained in those strains. Five strains were related to Bradyrhizobium japonicum and B. elkanii, with four of them being similar to strains carried in Brazilian inoculants, therefore modifications in physiological properties, as a shorter doubling time might have resulted from adaptation to local conditions. Phospholipid fatty acid analysis (PFLA) was less precise in delineating taxonomic relationships. The strains fit into eight Nod-factor profiles that were related to rhizobial species, but not to N2-fixation capacity or competitiveness. The data obtained highlight the diversity and promiscuity of rhizobia in the tropics, being capable of nodulating exotic legumes and might reflect ecological strategies to survive in N-poor soils; in addition, the diversity could also represent an important source of efficient and competitive rhizobial strains for the tropics. Putative new rhizobial species were detected only in undisturbed soils. Three species (R. tropici, B. japonicum and B. elkanii) were found under the more sustainable management system known as no-till, while the only species isolated from soils under conventional till was R. tropici. Those results emphasize that from the moment that agriculture was introduced into undisturbed soils rhizobial diversity has changed, being drastically reduced when a less sustainable soil management system was adopted.

Nitrogen fixation with the soybean crop in Brazil: Compatibility between seed treatment with fungicides and bradyrhizobial inoculants

Biological nitrogen fixation with the soybean crop can be improved by seed inoculation with superiorBradyrhizobium strains, but factors that reducethe population of inoculated bradyrhizobiaon the seedwill directly affect the efficiency of the process. Seed treatment with fungicides has been broadly practiced as cheap insurance against seed-and soil-borne pathogens, but toxicity of most fungicides to bradyrhizobia has often been underestimated. The compatibility between seed treatment with fungicides in single or mixed applications (including Benomyl, Captan, Carbendazin, Carboxin, Difenoconazole, Thiabendazole, Thiram, Tolylfluanid) and bradyrhizobial inoculants was examined in laboratory, greenhouse and field experiments during five crop seasons in Brazil. Bacterial survivalon the seeds was severely affected by all fungicides, resulting in mortalities of up to 62% after only 2 h and of 95% after 24 h. Fungicides also reduced nodule number, total N in grains and decreased yield by up to 17%. The toxic effects of fungicides were more drastic in sandy soils without soybean inoculation and cropping history, reducing nodulation by up to 87%, but were also important in areas with established populations of soybean bradyrhizobia. Therefore, fungicides should be used only when the seeds or soil are contaminated with pathogens, otherwise biological N2 fixation may be severely affected.

Conjunto mínimo de parâmetros para avaliação da microbiota do solo e da fixação biológica do nitrogênio pela soja

The objective of this work was to evaluate and validate a minimum set of parameters capable of monitoring, under field conditions, soil microbes and/or the biological N2 fixation process with the soybean crop. The field trials were performed in commercial fields (2002/2003) and experimental areas (2004/2004), with complete randomized blocks design. Microbial biomass C (MBC) and N (MBN) were adequate for the quantitative evaluation of soil microbes; the parameters were significantly correlated therefore the determination of only one of them is sufficient. Positive and significant correlations were also obtained between the parameters of shoot dry weight (SDW) and MBC and MBN. The analysis of total soil DNA by the denaturating gradient gel electrophoresis method (DGGE) detected qualitative changes in soil microbes, related to the homogeneity of the area, as well as to differences among treatments, areas and harvests. Nodule dry weight (NDW) was the most suitable parameter for the evaluation of nodulation. Significant correlations were obtained between the SDW and the total N accumulated in shoots (TNS). Similar results were obtained between SDW and total N-ureides (TNU). Monitoring of the symbiosis in soils traditionally cropped with soybean and poor on N may be achieved only by the determination of NDW and SDW.

Efeito de fontes e doses de molibdênio na sobrevivência do Bradyrhizobium e na fixação biológica de nitrogênio em soja

Molybdenum (Mo) is an essential nutrient for soybean because it is a component of enzyme nitrogenase, responsible for the biological nitrogen fixation (BNF). It has been recommended for application on soybean seeds along with soybean inoculants. However, several works have showed that Mo applied together with inoculant reduce Bradyrhizobium survival, nodulation and BNF efficiency. The aim of this study was to know if there are alternative ways to supply Mo to soybean without affecting Bradyrhizobium strains survival, nodulation and BNF efficiency. The alternative ways studied in vitro were to know the growth of strains in liquid medium with different sources and levels of Mo, and in glasshouse these sources of Mo were compared with the commercial ones on their effects on bacteria survival on the seeds, nodulation and BNF efficiency. The strains of Bradyrhizobium showed different growth rate for the Mo concentration and sources tested, being the strain SEMIA 5080 the best of them. Mo application on the seeds must be avoided, because Mo reduces bacteria survival, nodulation and, BNF efficiency. The best Mo sources to BNF were ammonium molybdate, Mo trioxide and the commercial sources Grap and Comol.

Adubação nitrogenada suplementar tardia em soja cultivada em latossolos do Cerrado

The objective of this work was to evaluate the effects of nitrogen fertilization at the pre-flowering (R1) and seed filling (R5) stages, on the nodulation and yield of soybean grown in Cerrado Oxisols. The study was performed for seven years in a clayey Typic Haplustox, and for four years in a clayey Rhodic Haplustox, both with established populations of Bradyrhizobium. Fifteen experiments were carried out under conventional and no tillage management systems and the treatments were: standard inoculation (SI) with B. japonicum; SI + 200 kg ha-1 N (urea), split-applied 50% at sowing and 50% at the R1 stage; SI + 50 kg ha-1 N (ammonium nitrate) in R1; SI + 50 kg ha-1 N (ammonium sulfate) in R1; SI + 50 kg ha-1 N (ammonium nitrate) in R5; and SI + 50 kg ha-1 N (ammonium sulfate) in R5. The application of 200 kg ha-1 N harmed nodulation. In only two experiments, the yield responses to late season N applications were significant, with no net economical benefits for the producer. Late nitrogen fertilization on inoculated soybean, grown under Brazilian Cerrados Oxisols, has no economical advantages in any of the management systems evaluated, regardless of the N source applied.

Avaliação qualitativa e quantitativa da microbiota do solo e da fixação biológica do nitrogênio pela soja

The objective oh this work was to define, for edaphoclimatic conditions of Brazil, acceptable levels of dispersion of some biological parameters, used in studies of environmental impact due to the adoption of new technologies with the soybean crop. Two experiments were performed in eleven counties, in six states and in the Federal District, with conventional and transgenic soybean. Evaluated parameters were: carbon and nitrogen of the microbial biomass, microbial respiration, microbial metabolic quotient, electrophoresis of soil DNA in denaturating gradient gel (DGGE), biological nitrogen fixation, number of rhizobial cells, nodule number and nodule dry weight, nodule occupancy by Bradyrhizobium strains, shoot dry weight, total nitrogen in shoot and nitrogen as ureides in shoot. The temporal variability for carbon and nitrogen of microbial biomass, microbial respiration and microbial metabolic quotient was adequate, and the maximum coefficient of variation was estimated at 35%. The homogeneity between replicates, treatments and harvests was confirmed by the DGGE method. In N-poor soils, the parameters of nodules and shoot dry weight, with maximum coefficient of variation 33 and 18%, respectively, were adequate for evaluation of biological nitrogen fixation, which contributed from 72 to 88% of total nitrogen in shoot.

The Brazilian Experience with the Soybean (Glycine max) and Common Bean (Phaseolus vulgaris) Symbioses

Soybean and common bean are cultivated on more than 17 million hectares in Brazil. Soybean is the main cash crop and relies exclusively on BNF to supply the plant N demands. However, to maintain yields of more productive cultivars, a search for efficient, competitive and stress tolerant strains continues. Common bean is the most important legume used for food and inoculation with an efficient, competitive Rhizobium tropici strain increases in nodulation, nodule occupancy and yield. An adequate soil management is essential to guarantee the benefits of BNF with both crops.

Importance of Molybdenum and Cobalt to the Biological Nitrogen Fixation

Results indicated that: 5.1 Application of Mo + Co on seeds reduces nodule dry weight and BNF in glasshouse experiment; 5.2 There are no Mo + Co effects on nodulation in soil with established Bradyrhizobium population; 5.3 Application of Mo and especially Mo + Co, increase BNF; 5.4 Application of Mo + Co on leaves have the same effect on BNF as seed applications; 5.5 Soil application of Mo + Co does not supply the Mo and Co necessary to the plant and to the BNF; and 5.6 Seeds with high concentration of Mo show higher BNF than those with low Mo contents.