Overexpression of citrate operon in Herbaspirillum seropedicae Z67 enhances organic acid secretion,mineral phosphate solubilization and growth promotion of Oryza sativa

Background and aims

Herbaspirillum seropedicae Z67, nitrogen fixing endophyte, significantly promotes the growth of cereals. Organic acid secreting nitrogen fixing rhizobacteria have better plant growth promotion abilities due to mineral phosphate solubilization.

Method

Plasmids pAB7, pJNK3 and pJNK4 containing Escherichia coli cs (gltA), NADH insensitive cs (gltA1), and citrate operon consisting of gltA1 gene along with Salmonella typhimurium Na⁺ dependent citrate transporter (citC) gene under constitutive lac promoter were constructed in broad host range plasmid pUCPM18-Km^r. The plasmid transformants of H. seropedicae Z67 were obtained by electroporation.

Results

Hs (pAB7) and Hs (pJNK3) had increased CS activity but citric acid secretion was not significant. Hs (pJNK3) secreted 45 mM acetic acid while Hs (pJNK4) secreted 2.7 mM citric and 51 mM acetic acids. Hs (pJNK3) and Hs (pJNK4) released 80 μM and 110 μM amount of P from rock phosphate, respectively, in buffered medium under both aerobic and micro aerobic conditions. These transformants showed better plant growth promoting factors. Upon inoculation to rice plants (Gujarat – 17), increase of Fresh weight, Dry weight N, P and K content was observed.

Conclusion

Thus the study demonstrates that artificial citrate operon in H. seropedicae Z67 enhances phosphate solubilization and plant growth promotion abilities.     

Impact of plant growth-promoting bacteria on grain yield,protein content,and urea-<Superscript>15</Superscript> N recovery by maize in a Cerrado Oxisol

Background and aims

The inoculation of cereal crops with plant growth-promoting bacteria (PGPB) is a potential strategy to improve fertilizer-N acquisition by crops in soils with low capacity to supply N. A study was conducted to assess the impact of three inoculants on grain yield, protein content, and urea-¹⁵ N recovery in maize (Zea mays L.) under Cerrado soil and climate conditions.

Methods

The main treatments included inoculants containing (i) Azospirillum brasilense strain Sp245, (ii) A. brasilense strains AbV5 + AbV6, (iii) Herbaspirillum seropedicae strain ZAE94, and (iv) a non-inoculated control. The subtreatments were (i) urea-N fertilization (100 kg N ha^?1) at 30 days after sowing and (ii) no N addition at the stage. To determine fertilizer-N recovery, ¹⁵N–labelled urea was applied in microplots.

Results

Inoculants carrying A. brasilense improved urea-¹⁵ N acquisition efficiency in maize and also improved grain yield compared to the non-inoculated control, while urea-N fertilization enhanced grain quality by providing higher protein content.

Conclusion

Our results suggest that the inoculation of maize grains with PGPB represents a strategy to improve fertilizer-N recovery and maize yield in Cerrado soil with a low capacity to supply N.

     

Survival of endophytic bacteria in polymer-based inoculants and efficiency of their application to sugarcane

Background & aims

Studies have been conducted to evaluate maintenance of cell viability and stability, as well as to select cheap carriers to extend the shelf life of plant beneficial bacterial inoculants for agricultural crops. The purpose of this study was to evaluate the shelf life and the colonization efficiency of novel liquid and gel-based inoculant formulations for sugarcane. The different inoculant formulations were all composed of a mixture of five strains of diazotrophic bacteria (Gluconacetobacter diazotrophicus, Herbaspirillum seropedicae, H. rubrisubalbicans, Azospirillum amazonense and Burkholderia tropica), which are recognized as sugarcane growth promoters.

Methods

Different inoculant formulations containing as carrier the polymers carboxymethylcellulose (CMC) and corn starch (60/40 ratio) at five different concentrations (named PIC, for Polymeric Inoculant Carrier) were supplemented, or not, with 2?% MgO, an interfacial stabilizing agent. Bacterial survival in the different formulations during storage was evaluated under controlled conditions, and two experiments with mini-cuttings of sugarcane variety RB72454 were carried out under greenhouse conditions.

Results

Laboratory tests showed that in the formulation composed of 0.8?g of the polymeric mixture per 100?g of the final product (PIC 0.8), survival of G. diazotrophicus and A. amazonense was around 10⁹?CFU?mL^?1 after 120?days of storage, regardless of the supplementation with MgO. The other formulation (2.2?g of polymeric mixture, PIC 2.2) presented survival levels of 10⁸?CFU?mL^?1 for up to 60?days of storage for all the individual strains. In the greenhouse, sugarcane seedlings showed a positive growth response 50?days after inoculation when inoculated with the mixture of five bacteria, with and without PIC 2.2.

Conclusions

The polymer carriers described here allowed for the long-term survival of the five different bacterial strains tested. In addition, short-term experiments in the greenhouse showed that their application as part of an inoculant on sugarcane cuttings was at least as effective in terms of bacterial colonization and the promotion of plant growth as that of the bacterial mixture without carriers.     

Utilizing pyrene-degrading endophytic bacteria to reduce the risk of plant pyrene contamination

Aims

Endophytic bacteria are ubiquitous in plants, but little information is available on the influence of endophytic bacteria on the uptake and metabolism of PAH by plants. Thus, we seek to investigate whether the colonization of a target plant by a PAH-degrading endophytic bacterium would improve the PAH metabolism of the plant and reduce the risk of plant PAH contamination.

Methods

A pyrene-degrading endophyte was isolated from PAH-contaminated plants using enrichment culture. After root inoculation with the isolated bacterium, greenhouse container experiments were conducted. Pyrene residues in soil and plant samples were analyzed by HPLC.

Results

A pyrene-degrading endophytic bacterium, Staphylococcus sp. BJ06, was isolated from Alopecurus aequalis and could degrade 56.0 % of pyrene (50 mg?·?L^?1) within 15 days. BJ06 grew and degraded pyrene efficiently under environmental conditions. The bacterium significantly promoted ryegrass growth and pyrene removal from contaminated soil in container experiments. The pyrene concentrations in ryegrass roots and shoots in endophyte-inoculated planted soil were reduced by 31.01 % and 44.22 %, respectively, compared with endophyte-free planted soil.

Conclusions

We have provided new perspectives on the regulation and control of plant uptake of organic contaminants with endophytic bacteria. The results of this study will be valuable to risk assessments of plant PAH contamination.     

Selection of phosphate-solubilizing diazotrophic Herbaspirillum and Burkholderia strains and their effect on rice crop yield and nutrient uptake

Background and Aims

Plant growth-promoting bacteria, mainly diazotrophs and phosphate solubilizers, can reduce the use of chemical fertilizers for rice crops. Here, diazotrophic bacteria isolated from rice were screened for their ability to solubilize inorganic P (P_i) in vitro and in association with rice plants cultivated in pots.

Methods

Forty-nine isolates were tested for the ability to solubilize P_i on NBRIP and GL agar plate media and seven selected strains were further evaluated in NBRIP liquid medium. Three of these strains were inoculated in rice plants grown in soil pots containing ¹⁵N-labeled fertilizer and two sources of P: tricalcium phosphate (TCP) or simple superphosphate (SSP). The dry matter, yield, N, P, and the ¹⁵N content accumulated in plant tissues were measured at 135 days after planting.

Results

Seven strains belonging to the genera Herbaspirillum and Burkholderia formed a halo of solubilized P_i on agar plates. The Burkholderia strains showed peak soluble P (around 200 mg P L^?1) on the fifth day when grown in NBRIP liquid medium for 14 days. Inoculation of Herbaspirillum strains (H18, ZA15) and a Burkholderia vietaminensis strain (AR114) increased rice grain yield from 33 to 47 % with TCP and 18 to 44 % with TSS, respectively. The bacterial inoculation led to enhanced N-use efficiency of the ¹⁵N-labeled fertilizer.

Conclusion

These results suggest that the selection and use of P-solubilizing diazotrophic bacteria are a good strategy to promote P solubilization and/or N use efficiency in rice plants.     

Nostoc, Microcoleus and Leptolyngbya inoculums are detrimental to the growth of wheat (Triticum aestivum L.) under salt stress

Background and aims

This study investigated the effect of cyanobacterial inoculants on salt tolerance in wheat.

Methods

Unicyanobacterial crusts of Nostoc, Leptolyngbya and Microcoleus were established in sand pots. Salt stress was targeted at 6 and 13 dS m^?1, corresponding to the wheat salt tolerance and 50 % yield reduction thresholds, respectively. Germinated wheat seeds were planted and grown for 14 (0 and 6 dS m^?1) and 21 (13 dS m^?1) days by which time seedlings had five emergent leaves. The effects of cyanobacterial inoculation and salinity on wheat growth were quantified using chlorophyll fluorescence, inductively coupled plasma-optical emission spectrometry and biomass measurements.

Results

Chlorophyll fluorescence was negatively affected by soil salinity and no change was observed in inoculated wheat. Effective photochemical efficiency correlated with a large range of plant nutrient concentrations primarily in plant roots. Inoculation negatively affected wheat biomass and nutrient concentrations at all salinities, though the effects were fewer as salinity increased.

Conclusions

The most likely explanation of these results is the sorption of nutrients to cyanobacterial extracellular polymeric substances, making them unavailable for plant uptake. These results suggest that cyanobacterial inoculation may not be appropriate for establishing wheat in saline soils but that cyanobacteria could be very useful for stabilising soils.     

Nitrogen dynamics following field application of biochar in a temperate North American maize-based production system

Background and aims

Biochar additions to tropical soils have been shown to reduce N leaching and increase N use efficiency. No studies exist verifying reduced N leaching in field experiments on temperate agricultural soils or identifying the mechanism for N retention.

Methods

Biochar derived from maize stover was applied to a maize cropping system in central New York State at rates of 0, 1, 3, 12, and 30 t?ha^-1 in 2007. Secondary N fertilizer was added at 100, 90, 70, and 50 % of the recommended rate (108 kg N ha^-1). Nitrogen fertilizer enriched with ¹⁵?N was applied in 2009 to the 0 and 12 t?ha^-1 of biochar at 100 and 50 % secondary N application.

Results

Maize yield and plant N uptake did not change with biochar additions (p?>?0.05; n?=?3). Less N (by 82 %; p?<?0.05) was lost after biochar application through leaching only at 100 %?N fertilization. The reason for an observed 140 % greater retention of applied ¹⁵?N in the topsoil may have been the incorporation of added ¹⁵?N into microbial biomass which increased approximately three-fold which warrants further research. The low leaching of applied fertilizer ¹⁵?N (0.42 % of applied N; p?<?0.05) and comparatively high recovery of applied ¹⁵?N in the soil (39 %) after biochar additions after one cropping season may also indicate greater overall N retention through lower gaseous or erosion N losses with biochar.

Conclusions

Addition of biochar to fertile soil in a temperate climate did not improve crop growth or N use efficiency, but increased retention of fertilizer N in the topsoil.     

Liebig’s law of the minimum applied to a greenhouse gas: alleviation of P-limitation reduces soil N2O emission

Background and aims

Emission of the greenhouse gas (GHG) nitrous oxide (N₂O) are strongly affected by nitrogen (N) fertilizer application rates. However, the role of other nutrients through stoichiometric relations with N has hardly been studied. We tested whether phosphorus (P) availability affects N₂O emission. We hypothesized that alleviation of plant P-limitation reduces N₂O emission through lowering soil mineral N concentrations.

Methods

We tested our hypothesis in a pot experiment with maize (Zea mays L.) growing on a P-limiting soil/sand mixture. Treatment factors included P and N fertilization and inoculation with Arbuscular Mycorrhizal Fungi (AMF; which can increase P uptake).

Results

Both N and P fertilization, as well as their interaction significantly (P?<?0.01) affected N₂O emission. Highest N₂O emissions (2.38 kg N₂O-N ha^?1) were measured at highest N application rates without P fertilization or AMF. At the highest N application rate, N₂O fluxes were lowest (0.71 kg N₂O-N ha^?1) with both P fertilization and AMF. The N₂O emission factors decreased with 50 % when P fertilization was applied.

Conclusions

Our results illustrate the importance of the judicious use of all nutrients to minimize N₂O emission, and thereby further underline the intimate link between sound agronomic practice and prudent soil GHG management.     

Assessment of nitrate leaching loss on a yield-scaled basis from maize and wheat cropping systems

Background and aims

It is so far a gap in knowledge to assess nitrate (NO₃ ^?) leaching loss linking with crop yield for a given cereal cropping system.

Methods

We conducted a meta-analysis on 32 published studies reporting both NO₃ ^? leaching losses and crop yields in the maize (N?=?20) and wheat (N?=?12) systems.

Results

On average, 22 % and 15 % of applied fertilizer N to wheat and maize systems worldwide are leached in the form of NO₃ ^?, respectively. The average area-scaled NO₃ ^- leaching loss for maize (57.4 kg N ha^?1) was approx. two times higher than for wheat (29.0 kg N ha^?1). While, if scaled to crop yields, the average yield-scaled NO₃ ^? losses were comparable between maize (5.40 kg N Mg^?1) and wheat (5.41 kg N Mg^?1) systems. Across all sites, the lowest yield-scaled NO₃ ^? leaching losses were observed at slightly suboptimal fertilization rates, corresponding to 90 % and 96 % of maximum maize or wheat yields, respectively.

Conclusions

Our findings suggest that small adjustments of agricultural N management practices can effectively reduce yield-scaled NO₃ ^? leaching losses. However, further targeted field experiments are still needed to identify at regional scale best agricultural management practices for reducing yield-scaled NO₃ ^? leaching losses in maize and wheat systems.     

No need to breed for enhanced colonization by arbuscular mycorrhizal fungi to improve low-P adaptation of West African sorghums

Background and aims

Herbaspirillum seropedicae (Hs) Z67 a diazotrophic endophyte was genetically engineered for secretion of 2-keto-D-gluconic acid by heterologous expression of genes for pqq synthesis and gluconate dehydrogenase to study its beneficial effect on plants.

Methods

Two plasmids, pJNK5, containing a 5.1 Kb pqq gene cluster of Acinetobacter calcoaceticus and pJNK6, carrying in addition the Pseudomonas putida KT2440 gluconate dehydrogenase (gad) operon were constructed in pUCPM18Gm^r under Plac promoter. H. seropedicae Z67 transformants were monitored for P and K solubilization, cadmium (Cd) tolerance and rice growth promotion.

Results

Hs (pJNK5) secreted 23.5 mM gluconic acid and Hs (pJNK6) secreted 3.79 mM gluconic acid and 15.8 mM 2-ketogluconic acid respectively. Under aerobic conditions, Hs (pJNK5) and Hs (pJNK6) solubilized 239.7 μM and 457.7 μM P on HEPES rock phosphate and, 76.7 μM and 222.7 μM K on HRPF (feldspar), respectively, in minimal medium containing 50 mM glucose. Under N free minimal medium, similar effects of P and K solubilization were obtained. Hs (pJNK5) and Hs (pJNK6) inoculation increased the biomass, N, P, K content of rice plants (Gujarat – 17). These plants also accumulated 0.73 ng/g PQQ, and had improved growth and tolerance to CdCl₂.

Conclusions

Incorporation of pqq and gad gene clusters in H. seropedicae Z67 imparted additional plant growth promoting traits of P and K solubilization and ability to alleviate Cd toxicity to the host plant.

     

Zinc fertilizer placement affects zinc content in maize plant

Background and aims

Adequate zinc (Zn) in maize (Zea mays L.) is required for obtaining Zn-enriched grain and optimum yield. This study investigated the impact of varying Zn fertilizer placements on Zn accumulation in maize plant.

Methods

Two pot experiments with same design were conducted to investigate the effect of soil Zn heterogeneity by mixing ZnSO₄·7H₂O (10 mg Zn kg^?1 soil on an average) in 10–15, 0–15, 25–30, 0–30, 30–60 and 0–60 cm soil layers on maize root growth and shoot Zn content at flowering stage in experiment-1, and assessing effects on grain Zn accumulation at mature stage in experiment-2.

Results

In experiment-1, Zn placements created a large variation in soil DTPA-Zn concentration (0.3–29.0 mg kg^?1), which induced a systemic and positive response of root growth within soil layers of 0–30 cm; and shoot Zn content was increased by 102 %–305 % depending on Zn placements. Supply capacity of Zn in soil, defined as sum of product of soil DTPA-Zn concentration and root surface area at different soil layers, was most related to shoot Zn content (r?=?0.82, P?<?0.001) via direct and indirect effects according to path analysis. In experiment-2, Zn placements increased grain Zn concentration by up to 51 %, but significantly reduced the grain Zn harvest index from 50 % by control to about 30 % in average.

Conclusion

Matching the distribution of soil applied Zn with root by Zn placement was helpful to maximize shoot Zn content and grain Zn concentration in maize.     

Real-Time PCR Quantification of the Plant Growth Promoting Bacteria Herbaspirillum seropedicae Strain SmR1 in Maize Roots

The plant growth promoting bacteria Herbaspirillum seropedicae SmR1 is an endophytic diazotroph found in several economically important crops. Considering that methods to monitor the plant–bacteria interaction are required, our objective was to develop a real-time PCR method for quantification of PGPB H. seropedicae in the rhizosphere of maize seedlings. Primer pairs were designed, and their specificity was verified using DNA from 12 different bacterial species. Ten standard curves of qPCR assay using HERBAS1 primers and tenfold serial dilutions of H. seropedicae SmR1 DNA were performed, and PCR efficiency of 91 % and correlation coefficient of 0.99 were obtained. H. seropedicae SmR1 limit of detection was 10¹ copies (corresponding to 60.3 fg of bacterial DNA). qPCR assay using HERBAS1 was used to detect and quantify H. seropedicae strain SmR1 in inoculated maize roots, cultivated in vitro and in pots, harvested 1, 4, 7, and 10 days after inoculation. The estimated bacterial DNA copy number per gram of root was in the range 10⁷–10⁹ for plants grown in vitro and it was around 10⁶ for plants grown in pots. Primer pair HERBAS1 was able to quantify H. seropedicae SmR1, and this assay can be useful for monitoring plant–bacteria interaction.     

Bioactivity of humic acids isolated from vermicomposts at different maturation stages

Background and aims

Vermicomposts are useful to improve environmental quality and sustainable agriculture. Moreover, it is enriched with highly bioactive humic acids (HAs)-like substances and can substitute no-renew source of humic substances to use as plant growth promoters in agriculture. The aim of this work was to evaluate the biological effects of HAs isolated at increasing vermicompost maturation stages.

Methods

Lateral root emergence, aqueous growth medium acidification and proton pumps of maize seedlings were used to monitor HAs bioactivity. Molecular conformation of the HAs was determined by size-exclusion and reverse-phase high performance liquid chromatography. We applied spectroscopy ¹³C-NMR on VC samples to follow the humification process.

Results

We observed a decrease of carbohydrate content and selective preservation of hydrophobic alkyl and aryl C components by ¹³C-NMR during vermicompost maturation. Apparent molecular weight distribution of HAs did not change with vermicompost maturation, but was possible observed increase on hydrophobic moieties.

Conclusion

After 60 days of vermicomposting, all HAs promotes lateral root emergence, acidification of growth aqueous medium and induction of proton pumps without changes on apparent molecular weight but with enhance on hydrophobic domains.     

Nitrogen contributions from faba bean (Vicia faba L.) reliant on soil rhizobia or inoculation

Background and Aims

Understanding the impact of soil rhizobial populations and inoculant rhizobia in supplying sufficient nodulation is crucial to optimising N₂ fixation by legume crops. This study explored the impact of different rates of inoculant rhizobia and contrasting soil rhizobia on nodulation and N₂ fixation in faba bean (Vicia faba L.).

Methods

Faba beans were inoculated with one of seven rates of rhizobial inoculation, from no inoculant to 100 times the normal rate of inoculation, sown at two field sites, with or without soil rhizobia present, and their nodulation and N₂ fixation assessed.

Results

At the site without soil rhizobia, inoculation increased nodule number and increased N₂ fixation from 21 to 129 kg shoot N ha^?1, while N₂ fixation increased from 132 to 218 kg shoot N ha^?1 at the site with high background soil rhizobia. At the site without soil rhizobia, inoculation increased concentrations of shoot N from 14 to 24 mg g^?1, grain N from 32 to 45 mg g^?1, and grain yields by 1.0 Mg (metric tonne) ha^?1. Differences in nodulation influenced the contributions of fixed N to the system, which varied from the net removal of 20 kg N ha^?1 from the system in the absence of rhizobia, to a net maximum input of 199 kg N ha^?1 from legume shoot and root residues, after accounting for removal of N in grain harvest.

Conclusions

The impact of inoculation and soil rhizobia strongly influenced grain yield, grain N concentration and the potential contributions of legume cropping to soil N fertility. In soil with resident rhizobia, N₂ fixation was improved only with the highest inoculation rate.     

Inoculation of seed-borne fungus in the rhizosphere of Festuca arundinacea promotes hydrocarbon removal and pyrene accumulation in roots

Background and aims

The selective inoculation of specific hydrocarbon-degrading microbes into the plant rhizosphere offers a useful means for remediating hydrocarbon-contaminated soils. The effect of inoculating a seed-borne filamentous fungus (Lewia sp.) on hydrocarbon removal by Festuca arundinacea and its growth was studied on perlite (model soil) and soil, both spiked with hydrocarbons.

Methods

A hydrocarbon mixture (1,500 mg kg^?1) of two polycyclic aromatic hydrocarbons (PAH), phenanthrene and pyrene, blended with hexadecane (1.0:0.5:0.5 weight) was used. Greenhouse experiments were carried out for 45 days. Inoculated and non-inoculated plants were grown in dark cylindrical glass pots containing perlite or soil.

Results

Inoculation with Lewia sp. stimulated (100 %) root growth in spiked perlite. Inoculated plants showed higher phenanthrene removal (100 %) compared to non-inoculated plants in perlite and soil. Pyrene removal by inoculated plants was 37-fold higher than that by non-inoculated plants in perlite; in soil, pyrene removal by inoculated plants (97.9 %) differed significantly from that of non-inoculated plants (91.4 %). Accumulation of pyrene in roots (530.9 mg kg^?1 of dry roots) was promoted in perlite.

Conclusions

Our results demonstrate that Lewia sp. (endophytic fungus) improved the efficiency of PAH removal by F. arundinacea, on both perlite and soil, stimulating pyrene accumulation in roots.     

Alfalfa plant-derived biostimulant stimulate short-term growth of salt stressed Zea mays L. plants

Background & Aims

The effects of an alfalfa plant (Medicago sativa L.) hydrolysate-based biostimulant (EM) containing triacontanol (TRIA) and indole-3-acetic acid (IAA) were tested in salt-stressed maize plants.

Methods

Plants were grown for 2 weeks in the absence of NaCl or in the presence (25, 75 and 150 mM). On the 12th day, plants were supplied for 48 h with 1.0 mg L^?1 EM or 11.2 μM TRIA.

Results

EM and TRIA stimulated the growth and nitrogen assimilation of control plants to a similar degree, while NaCl reduced plant growth, SPAD index and protein content. EM or TRIA increased plant biomass under salinity conditions. Furthermore, EM induced the activity of enzymes functioning in nitrogen metabolism. The activity of antioxidant enzymes and the synthesis of phenolics were induced by salinity, but decreased after EM treatment. The enhancement of phenylalanine ammonia-lyase (PAL) activity and gene expression by EM was consistent with the increase of flavonoids.

Conclusion

The present study proves that the EM increases plant biomass even when plants are grown under salinity conditions. This was likely because EM stimulated plant nitrogen metabolism and antioxidant systems. Therefore, EM may be proposed as bioactive product in agriculture to help plants overcome stress situations.     

Increased plant density of winter wheat can enhance nitrogen–uptake from deep soil

Background and Aims

Increased plant density improves grain yield and nitrogen (N)–use efficiency in winter wheat (Triticum aestivum L.) by increasing the root length density (RLD) in the soil and aboveground N–uptake (AGN) at maturity. However, how the root distribution and N–uptake at different soil depths is affected by plant density is largely unknown.

Methods

A 2–year field study using the winter wheat cultivar Tainong 18 was conducted by injecting ¹⁵?N–labeled urea into soil at depths of 0.2, 0.6, and 1.0 m under four plant densities of 135 m^?2, 270 m^?2,405 m^?2, and 540 m^?2.

Results

We observed significant RLD and ¹⁵?N–uptake increases at each soil depth as the plant density increased from 135 to 405 m^?2. ¹⁵?N–uptake increased with plant density as the soil depth increased, although the corresponding RLD value fell with depth. The ¹⁵?N–uptake at each soil depth was positively related to the RLD at the same depth. The total AGN was positively related to RLD in deep soil, especially at 0.8–1.2 m.

Conclusions

Increasing the plant density from 135 m^?2 to the optimum increases AGN primarily by increasing the RLD in deep soil and therefore increasing the plant density of winter wheat can be used to efficiently recover N leached to deep soil. Moreover, the total root numbers per unit area and RLD still increased at supraoptimal density while shoot number and N uptake stagnated.     

Maintaining elevated Fe2+ concentration in solution culture for the development of a rapid and repeatable screening technique for iron toxicity tolerance in rice (Oryza sativa L.)

Background and aims

Iron toxicity decreases rice (Oryza sativa) grain yield especially in acid soils after flooding. Our aim was to establish a high-throughput screening technique using nutrient solution culture for identifying Fe-toxicity-tolerant genotypes.

Methods

Varying levels of Fe, pH, and chelators in Yoshida nutrient solution culture were tested to maintain sufficient Fe²⁺ concentration over time to optimize the severity of Fe toxicity stress for distinguishing between a tolerant (Azucena) and sensitive (IR64) genotype. The optimized solution was tested on 20 diverse genotypes in the greenhouse, with measurement of leaf bronzing scores and plant growth characteristics at the seedling stage. The same 20 genotypes were grown to maturity in a field with natural Fe toxicity stress, with measurement of seedling-stage leaf bronzing scores and grain yield to determine their inter-relationship.

Results

Optimized nutrient solution conditions were 300 mg L^?1 Fe supplied as Fe²⁺ at pH 4.0 with a 1:2 molar ratio of Fe:EDTA, which maintained sufficient Fe²⁺ stress over 5 days. The highest correlation of nutrient solution phenotypic data with field grain yield was found with leaf bronzing scores at 4 weeks, with a Pearson r of 0.628 for simple association and a Spearman corrected r of 0.610 for rank association (P?<?0.01) using 20 diverse rice genotypes with proven Fe toxicity tolerance reaction. The Leaf bronzing scores at 4 weeks in nutrient culture solution were also found highly correlated with LBS under natural field stress after 8 weeks that had highest correlation with grain yield under stress.

Conclusion

This culture solution-based standardized screening technique can be used in plant breeding programs as a high-throughput technique to identify genotypes tolerant to Fe toxicity.     

Interaction between root growth allocation and mycorrhizal fungi in soil with patchy P distribution

Aims and Background

Many plants preferentially grow roots into P-enriched soil patches, but little is known about how the presence of arbuscular mycorrhizal fungi (AMF) affects this response.

Methods

Lotus japonicus (L.) was grown in a low-P soil with (a) no additional P, (b) homogeneous P (28 mg pot^?1), (c) low heterogeneous P (9.3 mg pot^?1), and (d) high heterogeneous P (28 mg pot^?1). Each P treatment was combined with one of three mycorrhiza treatments: no mycorrhizae, Glomus intraradices, indigenous AMF. Real-time PCR was used to assess the abundance of G. intraradices and the indigeneous AMF G. mosseae and G. claroideum.

Results

Mycorrhization and P fertilization strongly increased plant growth. Homogeneous P supply enhanced growth in both mycorrhizal treatments, while heterogeneous P fertilization increased biomass production only in treatments with indigenous AMF inoculation. Preferential root allocation into P-enriched soil was significant only in absence of AMF. The abundance of AMF species was similar in P-enriched and unfertilized soil patches.

Conclusion

Mycorrhization may completely override preferential root growth responses of plants to P- patchiness in soil. The advantage of this effect for the plants is to give roots more freedom to forage for other resources in demand for growth and to adapt to variable soil conditions.     

Blue-light regulation of ZmPHOT1 and ZmPHOT2 gene expression and the possible involvement of Zmphot1 in phototropism in maize coleoptiles

Main conclusion

ZmPHOT1 and ZmPHOT2 are expressed differentially in maize coleoptiles and leaves, with Zmphot1 possibly involved in first-positive phototropic curvature of red-light-adapted maize coleoptiles exposed to pulsed low-fluence blue light. Unilateral blue-light perception by phototropin(s) is the first event of phototropism, with the subsequent signal causing lateral transport of auxin at the coleoptile tip region of monocots. In this study, we analyzed the behavior of two maize phototropin genes: ZmPHOT1 and ZmPHOT2, the latter identified from the maize genome database and newly characterized. Quantitative real-time PCR analysis demonstrated that ZmPHOT1 was abundantly expressed in etiolated coleoptiles, while lower expressions of both ZmPHOT1 and ZmPHOT2 were observed in young leaves. Interestingly, these genes were not specifically expressed in the coleoptile tip region, a key position for photoperception in phototropism. Exposure to pulsed low-fluence blue light (LBL) (0.33 µmol m^?2 s^?1 × 8 s) and continuous high-fluence blue light (HBL) (10 µmol m^?2 s^?1) rapidly decreased ZmPHOT1 gene expression in coleoptiles, with levels of ZmPHOT2 not significantly altered in that tissue. In young leaves, no drastic expression changes were induced in either ZmPHOT1 or ZmPHOT2 by LBL or HBL irradiation. The Zmphot1 protein was investigated by Western blot analysis with anti-Osphot1 antibodies. Zmphot1 was detected in microsomal fractions, with higher levels in coleoptiles than in leaves. HBL caused rapid phosphorylation of the protein, whereas no phot1 phosphorylation was induced by LBL. The involvement of Zmphot1 in LBL-induced phototropic curvature of maize coleoptiles is discussed.