Optimization of soybean yield in ultisols through adaptive varieties screening and plant growth promoting Rhizobacter

This research was aimed at obtaining varieties of soybean adaptive to acid soils and to obtain Plant Growth-Promoting Rhizobacteria (PGPR) isolates that can improve the agronomic characteristics of soybean and increase the ultisols fertility. This research was conducted in two-stages research on Sampali Village, Percut Sei Tuan sub-District, Deli Serdang District, North Sumatra Province, Indonesia from August 2019 until March 2020. The first stage (adaptive varieties screening) using the non-factorial Randomized Block Design (RBD) with the varieties of Argomulyo, Wilis, Kaba, Dena-1, Devon-1, Dega-1, Demas-1, Burangrang, Detam-1, and Kipas Merah. The second stage (application of PGPR isolates singly and in combinations) using the factorial RBD, the first factor of applicative single and the combination of PGPR isolates, the second factor of adaptive varieties including Detam-1 and Wilis. Data were analyzed with ANOVA and followed by DMRT at P<0.05. The results showed that the Detam-1 and Wilis varieties had significantly higher yield per plant of 14.73 g and 14.54 g, respectively, than other varieties. The applications of a single and combination of PGPR isolates significantly increased the number of branches, stem diameter, plant height, yield per plant, soil pH, organic-C, available-P, and total-N and decreased the soil C/N. The Detam-1 variety had the higher in yield per plant compared to Wilis variety. The isolates combination of Rhizobium leguminosarum+Rhizobium sp2+Bacillus sp+Burkholderia sp for Detam-1 and Wilis varieties can be recommended to support the growth and yield of soybean on ultisols.

Isolation and Characterization of Plant Growth-Promotion Diazotrophic Endophytic Bacteria Associated to Sugarcane (Saccharum officinarum L.) Grown in Paraíba, Brazil

Abstract: Sugarcane is an important Brazilian commodity, being usually cultivated in soils with low natural fertility. This study aimed to isolate diazotrophic endophytes from sugarcane tissues and evaluate the morphological and physiological characteristics of their colonies as well as their plant growth-promoting (PGP) traits in select diazotrophic endophytic bacteria. Fifty-six bacterial isolates were identified in the sugarcane tissues, and these isolates presented distinct morphological and physiological traits. A total of thirty-five bacterial isolates were biochemically evaluated. Overall, Bacillus was the dominant genus. Isolates of Methylobacterium spp. and Brevibacillus agri were present only in leaves, while Herbaspirillum seropedicae occurred only in stems. Except to IPA-CF45A, all isolates were nitrogenase positive. All endophytes exhibit production of indol 3-acetic acid. Over 50% of endophytes solubilize phosphate, release N-acyl homoserine lactones, and present the activity of 1-aminocyclopropane-1-carboxylic acid deaminase, catalase, lipase and protease. The network analysis showed that isolates belonged to Burkholderia, Herbaspirillum, and Methylobacterium interact with Bacillus. Bacterial endophytes exhibited distinct morphological, physiological, and PGP traits that are useful for sustainable agriculture, highlighting the isolates IPA-CC33, IPA-CF65, IPA-CC9 and IPA-CF27. Further studies on the effects of these diazotrophic endophytes and their potential for providing microbial inoculants for improving sugarcane fields will provide valuable information to maintain the sustainability and environment quality.

Kinetics of Gluconacetobacter diazotrophicus Growth Using Cane Molasses and Sucrose: Assessment of Kinetic Models / Cinética de crecimiento de gluconacetobacter diazotrophicus usando melaza de caña y sacarosa: evaluación de modelos cinéticos

RESUMEN Gluconacetobacter diazotrophicus es una bacteria endófita promotora del crecimiento vegetal utilizada como inoculante microbiano en diferentes cultivos agrícolas. El objetivo del presente trabajo fue aplicar diferentes modelos matemáticos para representar su crecimiento en un cultivo sumergido por lotes empleando un biorreactor de 3 L y usando melazas de caña y sacarosa como fuente de energía. Se obtuvo el perfil temporal de pH, biomasa celular y azúcares totales. Se compararon los modelos estudiados por calidad de ajuste y complejidad y se realizó un análisis de sensibilidad paramétrica. Se consideraron modelos de cuatro y cinco parámetros con expresiones que incluyen efectos de inhibición por sustrato y por biomasa. El modelo con mayor calidad de ajuste fue el de Herbert-Pirt-Contois con coeficientes de determinación para biomasa y sustrato de 0,888 y 0,425 respectivamente. Estos valores indican una mayor correspondencia de los datos experimentales de biomasa con los datos calculados por el modelo, en comparación con los resultados obtenidos para azúcares totales para los que esta correspondencia fue menor. Este modelo generó la mejor combinación de calidad de ajuste y complejidad según el criterio de información de Akaike. El estudio cinético desarrollado permitió observar un comportamiento bifásico en la etapa de crecimiento de la bacteria cuando se cultiva en melaza y un efecto de limitación de su crecimiento por la biomasa. Los resultados obtenidos proporcionan una descripción matemática útil para el diseño, escalamiento y operación de un futuro proceso de producción de un inoculante microbiano a base de la bacteria G. diazotrophicus.

ABSTRACT Gluconacetobacter diazotrophicus is a plant-growth promoting endophytic bacterium used as a microbial inoculant for different crops. The objective of this work was to apply different mathematical models to represent its growth in a batch submerged culture employing a 3-L bioreactor and using sugarcane molasses and sucrose as energy sources. The time profile of pH, cell biomass, and total sugars was obtained. Models studied were compared considering their fit quality and complexity, and a parametric sensitivity analysis was performed. Four- and five-parameter models with expressions involving substrate and biomass inhibition effects were considered. The Herbert-Pirt-Contois model achieved the highest fit quality with determination coefficients of 0.888 and 0.425 for biomass and substrate, respectively. These values indicate a higher correspondence between the experimental data of biomass concentration and the data calculated by the model, compared to results obtained for total sugars for which this correspondence was lower. This model reached the best combination considering the fit quality and complexity according to the Akaike's information criterion. The kinetic study performed enabled to observe a bi-phasic behavior in the growth stage of the bacterium when grown on molasses, and a growth limitation effect due to biomass concentration. The outcomes obtained provide a mathematical description useful for design, scale-up, and operation of a future process for the production of a microbial inoculant based on G. diazotrophicus.

Azotobacter chroococcum as a potentially useful bacterial biofertilizer for cotton (Gossypium hirsutum): Effect in reducing N fertilization / Azotobacter chroococcum como biofertilizante bacteriano potencialmente útil para el algodón (Gossypium hirsutum L.): efecto en la reducción de la fertilización nitrogenada

The aim of this research was to evaluate whether the application of two plant growth-promoting (rhizo)bacteria might reduce nitrogen fertilization doses in cotton. We used strains Azotobacter chroococcum AC1 and AC10 for their proven ability to promote seed germination and cotton growth. These microorganisms were characterized by their plant growth-promoting activities. Then, we conducted a glasshouse study to evaluate the plant growth promoting ability of these strains with reduced doses of urea fertilization in cotton. Results revealed that both strains are capable of fixing nitrogen, solubilizing phosphorus, synthesizing indole compounds and producing hydrolytic enzymes. After 12 weeks, the glasshouse experiment showed that cotton growth was positively influenced due to bacterial inoculation with respect to chemical fertilization. Notably, we observed that microbial inoculation further influenced plant biomass (p<0.05) than nitrogen content. Co-inoculation, interestingly, exhibited a greater beneficial effect on plant growth parameters compared to single inoculation. Moreover, similar results without significant statistical differences were observed among bacterial co-inoculation plus 50% urea and 100% fertilization. These findings suggest that coinoculation of A. chroococcum strains allow to reduce nitrogen fertilization doses up to 50% on cotton growth. Our results showed that inoculation with AC1 and AC10 represents a viable alternative to improve cotton growth while decreasing the N fertilizer dose and allows to alleviate the environmental deterioration related to N pollution.

El objetivo de esta investigación fue evaluar si la aplicación de 2 (rizo)bacterias promotoras del crecimiento vegetal podría reducir la dosis de fertilizante nitrogenado en el cultivo de algodón. Se usaron las cepas Azotobacter chroococcum AC1 y AC10 por su habilidad para promover la germinación de semillas y el crecimiento del algodonero. Estos microorganismos fueron caracterizados sobre la base de sus actividades de promoción del crecimiento vegetal. Luego se realizó un estudio de invernadero con plantas de algodón para evaluar la capacidad de promoción del crecimiento vegetal de dichas cepas con dosis reducidas de urea. Los resultados revelaron que ambas cepas son capaces de fijar nitrógeno, solubilizar fósforo, sintetizar compuestos indólicos y producir enzimas hidrolíticas. Después de 12 semanas, el experimento de invernadero permitió observar que el crecimiento del algodón fue influido positivamente por la inoculación bacteriana con respecto a la fertilización química. En particular, se evidenció que la inoculación microbiana impactó más en la biomasa vegetal (p<0,05) que en el contenido de nitrógeno. Curiosamente, la coinoculación exhibió un mayor efecto positivo sobre los parámetros de crecimiento en comparación con la inoculación simple. Además, se observaron resultados similares, sin diferencias estadísticamente significativas, entre la coinoculación bacteriana más del 50% de urea y el 100% de fertilización. Estos hallazgos indican que la coinoculación de las cepas de A. chroococcum AC1 y AC10 permitiría reducir las dosis de fertilización nitrogenada del cultivo de arroz en hasta el 50% y aliviar, de esta manera, el deterioro ambiental relacionado con la contaminación por N.

Characterization of phosphate solubilizing bacteria and nitrogen fixing bacteria from limestone mining region

Aims: Phosphate and nitrogen are major macronutrients needed by plants. Phosphates in the soil are present in the organic and inorganic form. The amounts of phosphate and nitrogen in marginal soil can be increased by plant growth promoting rhizobacteria (PGPR). The aim of this study was to isolate and characterize phosphate solubilizing bacteria which has ability to fix nitrogen from the soil around limestone mining area. Methodology and results: There were 22 isolates that could solubilize phosphate and fix nitrogen. There were 9 isolates that could not cause hypersensitivity symptoms (necrotic) on tobacco leaf. Quantitative assay of phosphate solubilizing was done by colorimetric method. Quantitative assay of phosphate showed that isolate GPC1.7 had the highest phosphate solubilizing activity on Pikovskaya broth (450 mg/L) on the 6th and 7th day of incubation whereas isolate GPA2.2 had the highest nitrogen fixing activity (0.162 ppm/h), measured with Acetylene Reduction Assay whereas nitrogenase activity of GPC1.7 was unidentified. Isolate GPA2.1 and GPA2.2 were Gram negative bacteria whereas isolate GPC1.7 was Gram positive bacteria. Identification based on 16S rRNA gene showed that GPA2.1 was closely related to Pseudomonas psychrotolerans, GPA2.2 was closely related to Stenotrophomonas maltophilia, GPC1.7 was closely related to Bacillus megaterium and B. aryabhattai. Conclusion, significance and impact of study: Phosphate solubilizing bacteria and nitrogen fixing bacteria isolate could be further used for revegetation process of the ex-limestone mining area.

Accumulation of ammonia in culture broth by wild-type nitrogen-fixing bacterium, stenotrophomonas maltophilia.

Stenotrophomonas maltophilia was isolated from paddy field of Kyaukse District, Mandalay Division, Myanmar. Nitrogen fixing activity of S. maltophilia was detected in nitrogen free mineral medium supplemented with different carbon sources. S. maltophilia accumulated highest amounts of ammonia in glucose supplemented medium. But their nitrogen fixing activity was best at 0.7% glucose concentration. When 0.5609 ppm of glucose still remained in the medium after 24 hours incubation, no ammonia was detected. After incubation of 32 hours, ammonia began to accumulate and the amount of 0.5 ppm of ammonia was detected but no sugar remained in the medium. Highest amount of ammonia (2 ppm) was accumulated in the medium after 48 hour incubation. So we found that ammonium accumulation began when no sugar remained in the medium, supposing that sugar plays a key role in ammonia accumulation.

Distribución de bacterias potencialmente fijadoras de nitrógeno y su relación con parámetros fisicoquímicos en suelos con tres coberturas vegetales en el sur de la Amazonia colombiana

Distribution of potentially nitrogen-fixing bacteria and its relationship with physicochemical parameters in soils with three vegetation types in the southern Colombian Amazon region. Potentially nitrogen-fixing microaerobic and aerobic bacteria were isolated from several Colombian Amazon soils (forest, pastures and chagras) and two landscapes (floodable and non floodable areas). The abundance and distribution of bacteria were evaluated, as well as their relationship with soil physical and chemical characteristics. Landscape had a direct influence on the abundance of the microaerobic bacteria, with higher numbers in forest and pasture soils in non- floodable zones. The aerobic isolates (N=51) were grouped into 19 morphologies, with the highest numbers found in forest soil in floodable zones. A higher number of aerobic morphologies was shared among forest sites (Nonmetric Multidimensional Scaling and Analysis of Similarity p<0.05), and 40% of the distribution was explained by lime percentage and Al concentration. Rev. Biol. Trop. 57 (4): 915- 927. Epub 2009 December 01.

Se evaluó la abundancia y distribución de bacterias microaerófilas y aerobias potencialmente fijadoras de nitrógeno aisladas a partir de suelos bajo coberturas de bosque, pastizal y chagra en dos paisajes, terraza y llanura inundable, en el sur de trapecio amazónico. Se relacionó el recuento en placa en el medio Ashby de estas bacterias aerobias con las características físicas y químicas del suelo. Se encontró mayor abundancia de bacterias aerobias y microaerofilas en suelos bajo cobertura de pastizal. Se observó influencia directa del paisaje sobre la abundancia de bacterias microaerófilas registrando mayores recuentos los suelos bajo terraza en las coberturas de bosque y pastizal. Los aislamientos aerobios obtenidos (51) se agruparon en 19 morfologías, de las cuales se obtuvo mayor número en suelos bajo bosque y en el paisaje de llanura inundable. A través del análisis multidimensional no métrico (NMDS) y análisis de similaridades (ANOSIM) (p<0.05) se comprobó que entre los sitios de muestreo bajo la cobertura de bosque se comparten mayor número de morfologías de bacterias aerobias que bajo las demás coberturas. El programa BIOENV indicó que esta distribución fue explicada en un 40% por las variables % limo y Al (meq/100 g).

Physiological traits of the symbiotic bacterium Teredinibacter turnerae isolated from the mangrove shipworm Neoteredo reynei

Nutrition in the Teredinidae family of wood-boring mollusks is sustained by cellulolytic/nitrogen fixing symbiotic bacteria of the Teredinibacter clade. The mangrove Teredinidae Neoteredo reynei is popularly used in the treatment of infectious diseases in the north of Brazil. In the present work, the symbionts of N. reynei, which are strictly confined to the host's gills, were conclusively identified as Teredinibacter turnerae. Symbiont variants obtained in vitro were able to grow using casein as the sole carbon/nitrogen source and under reduced concentrations of NaCl. Furthermore, cellulose consumption in T. turnerae was clearly reduced under low salt concentrations. As a point of interest, we hereby report first hand that T. turnerae in fact exerts antibiotic activity. Furthermore, this activity was also affected by NaCl concentration. Finally, T. turnerae was able to inhibit the growth of Gram-negative and Gram-positive bacteria, this including strains of Sphingomonas sp., Stenotrophomonas maltophilia, Bacillus cereus and Staphylococcus sciuri. Our findings introduce new points of view on the ecology of T. turnerae, and suggest new biotechnological applications for this marine bacterium.

Associative diazotrophic bacteria in grass roots and soils from heavy metal contaminated sites

This work aimed to evaluate density of associative diazotrophic bacteria populations in soil and grass root samples from heavy metal contaminated sites, and to characterize isolates from these populations, both, phenotypically (Zinc, Cadmium and NaCl tolerance in vitro, and protein profiles) and genotypically (16S rDNA sequencing), as compared to type strains of known diazotrophic species. Densities were evaluated by using NFb, Fam and JNFb media, commonly used for enrichment cultures of diazotrophic bacteria. Bacterial densities found in soil and grass root samples from contaminated sites were similar to those reported for agricultural soils. Azospirillum spp. isolates from contaminated sites and type strains from non-contaminated sites varied substantially in their in vitro tolerance to Zn+2 and Cd+2, being Cd+2 more toxic than Zn+2. Among the most tolerant isolates (UFLA 1S, 1R, S181, S34 and S22), some (1R, S34 and S22) were more tolerant to heavy metals than rhizobia from tropical and temperate soils. The majority of the isolates tolerant to heavy metals were also tolerant to salt stress as indicated by their ability to grow in solid medium supplemented with 30 g L-1 NaCl. Five isolates exhibited high dissimilarity in protein profiles, and the 16S rDNA sequence analysis of two of them revealed new sequences for Azospirillum.

Objetivou-se avaliar a densidade de populações de bactérias diazotróficas associativas em amostras de solos e de raízes de gramíneas oriundas de sítios contaminados com metais pesados, e caracterizar isolados destas populações através da análise fenotípica (tolerância aos metais pesados zinco e cádmio e à NaCl in vitro, perfis protéicos), e genotípica (seqüenciamento de 16S rDNA), comparados às estirpes tipo das mesmas espécies. As densidades foram avaliadas nos meios NFb, Fam e LGI, comumente utilizados para culturas de enriquecimento de populações de bactérias diazotróficas associativas. As densidades encontradas em amostras de solo e raiz de sítios contaminados foram semelhantes àquelas relatadas na literatura para solos agrícolas. Isolados de Azospirillum spp. de solos contaminados e estirpes tipo oriundas de solos não contaminados variaram substancialmente com relação à tolerância a Zn+2 e Cd+2, sendo que Cd+2 mais tóxico que Zn+2. Dentre os isolados mais tolerantes (UFLA 1S, 1R, S181, S34, e S22), alguns(1R, S34 e S22) foram mais tolerantes a metais pesados que rizóbios isolados de solos de áreas tropicais e temperadas. A maioria dos isolados mais tolerantes a metais pesados também foi tolerante ao estresse salino, o que foi indicado por seu crescimento em meio sólido suplementado com 30 g L-1 de NaCl in vitro. Cinco isolados apresentaram alta dissimilaridade em perfis protéicos e o seqüenciamento do gene 16S rDNA em dois deles revelou que apresentam novas seqüências de Azospirillum.