Seed inoculation of desert-plant growth-promoting rhizobacteria induce biochemical alterations and develop resistance against water stress in wheat.

Water shortage limits agricultural productivity, so strategies to get higher yields in dry agricultural systems is vital to circumvent the effect of climate change and land-shortage. The plant rhizosphere harbors beneficial bacteria able to confer biotic/abiotic tolerance along with a positive impact on plant growth. Herein, three bacterial strains, Proteus mirabilis R2, Pseudomonas balearica RF-2 and Cronobacter sakazakii RF-4 (accessions: LS975374, LS975373, LS975370, respectively) isolated from native desert-weeds were investigated for their response to improve wheat growth under drought stress. The bacteria showed drought tolerance up to 20% polyethylene glycol (PEG; -0.6 MPa), and salt (65-97 g l-1 ), 1-aminocyclopropane-1-carboxylate (ACC)-deaminase activity, P/Zn/K-solubilization, calcite degradation, IAA, and siderophore production. The plant growth-promoting rhizobacteria (PGPR) were evaluated on wheat under water stress. The P. balearica strain RF-2 primed seeds showed a maximum promptness index and germination index under PEG-stress, that is, 68% and 100%, respectively. Inoculation significantly improved plant growth, leaf area, and biomass under water stress. P. mirabilis R2 inoculated plant leaves showed the highest water contents as compared to the plants inoculated with other strains. C. sakazakii RF-4 inoculated plants showed minimum cell injury, electrolyte leakage, and maximum cell membrane stability at PEG stress. After 13 days exposure to drought, C. sakazakii RF-4 treated plants showed an overall higher expression of cytosolic ascorbate peroxidase (cAPX) and ribulose-bisphosphate carboxylase (rbcL) genes. The activity of stress-induced catalase and polyphenol oxidase was reduced, while that of peroxidase and superoxide dismutase increased after inoculation but the response was temporal. Taken together, this data explains that different PGPR (especially C. sakazakii RF-4) modulate differential responses in wheat that eventually leads towards drought tolerance, hence, it has the potential to enhance crop production in arid regions.

Retrieved 16S rRNA and nifH sequences reveal co-dominance of Bradyrhizobium and Ensifer (Sinorhizobium) strains in field-collected root nodules of the promiscuous host Vigna radiata (L.) R. Wilczek.

In the present study, the relative distribution of endophytic rhizobia in field-collected root nodules of the promiscuous host mung bean was investigated by sequencing of 16S ribosomal RNA (rRNA) and nifH genes, amplified directly from the nodule DNA. Co-dominance of the genera Bradyrhizobium and Ensifer was indicated by 32.05 and 35.84% of the total retrieved 16S rRNA sequences, respectively, and the sequences of genera Mesorhizobium and Rhizobium comprised only 0.06 and 2.06% of the recovered sequences, respectively. Sequences amplified from rhizosphere soil DNA indicated that only a minor fraction originated from Bradyrhizobium and Ensifer strains, comprising about 0.46 and 0.67% of the total retrieved sequences, respectively. 16S rRNA gene sequencing has also identified the presence of several non-rhizobial endophytes from phyla Proteobacteria, Actinobacteria, Bacteroides, and Firmicutes. The nifH sequences obtained from nodules also confirmed the co-dominance of Bradyrhizobium (39.21%) and Ensifer (59.23%) strains. The nifH sequences of the genus Rhizobium were absent, and those of genus Mesorhizobium comprised only a minor fraction of the sequences recovered from the nodules and rhizosphere soil samples. Two bacterial isolates, identified by 16S rRNA gene sequence analysis as Bradyrhizobium strain Vr51 and Ensifer strain Vr38, successfully nodulated the original host (mung bean) plants. Co-dominance of Bradyrhizobium and Ensifer strains in the nodules of mung bean indicates the potential role of the host plant in selecting specific endophytic rhizobial populations. Furthermore, successful nodulation of mung bean by the isolates showed that strains of both the genera Bradyrhizobium and Ensifer can be used for production of inoculum.

Successful phytoremediation of crude-oil contaminated soil at an oil exploration and production company by plants-bacterial synergism.

Phytoremediation is a promising approach for the cleanup of soil contaminated with petroleum hydrocarbons. This study aimed to develop plant-bacterial synergism for the successful remediation of crude oil-contaminated soil. A consortia of three endophytic bacteria was augmented to two grasses, Leptochloa fusca and Brachiaria mutica, grown in oil-contaminated soil (46.8 g oil kg-1 soil) in the vicinity of an oil exploration and production company. Endophytes augmentation improved plant growth, crude oil degradation, and soil health. Maximum oil degradation (80%) was achieved with B. mutica plants augmented with the endophytes and it was significantly (P < 0.05) higher than the use of plants or bacteria individually. Moreover, endophytes showed more persistence, the abundance and expression of alkB gene in the rhizosphere as well as in the endosphere of the tested plants than in unvegetated soil. A positive relationship (r = 0.70) observed between gene expression and crude oil reduction indicates that catabolic gene expression is important for hydrocarbon mineralization. This investigation showed that the use of endophytes with appropriate plant is an effective strategy for the cleanup of oil-contaminated soil under field conditions.

Bacterial rhizosphere and endosphere populations associated with grasses and trees to be used for phytoremediation of crude oil contaminated soil.

Different grasses and trees were tested for their growth in a crude oil contaminated soil. Three grasses, Lolium perenne, Leptochloa fusca, Brachiaria mutica, and two trees, Lecucaena leucocephala and Acacia ampliceps, were selected to investigate the diversity of hydrocarbon-degrading rhizospheric and endophytic bacteria. We found a higher number of hydrocarbon degrading bacteria associated with grasses than trees and that the endophytic bacteria were taxonomically different from rhizosphere associated bacteria showing their spatial distribution with reference to plant compartment as well as genotype. The rhizospheric soil yielded 22 (59.45 %), root interior yielded 9 (24.32 %) and shoot interior yielded 6 (16.21 %) hydrocarbon-degrading bacteria. These bacteria possessed genes encoding alkane hydroxylase and showed multiple plant growth-promoting activities. Bacillus (48.64 %) and Acinetobacter (18.91 %) were dominant genera found in this study. At 2 % crude oil concentration, all bacterial isolates exhibited 25 %-78 % oil degradation and Acinetobacter sp. strain BRSI56 degraded maximum. Our study suggests that for practical application, support of potential bacteria combined with the grasses is more effective approach than trees to remediate oil contaminated soils.

Genetic, physiological and biochemical characterization of Bacillus sp. strain RMB7 exhibiting plant growth promoting and broad spectrum antifungal activities.

BACKGROUND: Plant growth promoting rhizobacteria (PGPR) are functionally diverse group of bacteria having immense potential as biofertilizers and biopesticides. Depending upon their function, they may serve as partial replacements for chemical fertilizer or pesticides as an eco-friendly and cost-effective alternatives as compared to their synthetic counterparts. Therefore, isolation, characterization and practical evaluation of PGPRs having the aforementioned multifaceted beneficial characteristics, are essentially required. This study describes the detailed polyphasic characterization of Bacillus sp. strain RMB7 having profound broad spectrum antifungal activity and plant growth promoting potential. RESULTS: Based on 16S rRNA gene sequencing, strain RMB7 was identified as Bacillus specie. This strain exhibited the production of 8 mg. L(-1)of indole-3-acetic acid (IAA) in tryptophan-supplemented medium. It was able to solubilize 50.6 mg. L(-1) tri-calcium phosphate, reduced 601ηmol acetylene h(-1)/vial and inhibited >70% growth of nine fungal phytopathogens tested in vitro. Under natural pathogen pressure, inoculation with strain RMB7 and RMB7-supernatant conferred resistance by arugula plant against Pythium irregulare with a concurrent growth improvement over non-inoculated plants. The T-RFLP analysis based on 16S rRNA gene showed that inoculation with RMB7 or its supernatant have a major impact on the indigenous rhizosphere bacterial population. Mass spectrometric analysis revealed the production of lipopeptide surfactins as well as iturin A presence in crude extract of RMB7. PCR-amplification further confirmed the presence of genes involved in the biosynthesis of these two bioactive lipopeptide compounds. CONCLUSIONS: The data show that Bacillus sp. strain RMB7 has multifaceted beneficial characteristics. It may be an ideal plant growth promoting as well as biocontrol agent, for its integrated use in disease and nutrient management strategies.

RNA-Seq analysis of mung bean (Vigna radiata L.) roots shows differential gene expression and predicts regulatory pathways responding to taxonomically different rhizobia.

Symbiotic interaction among legume and rhizobia is a complex phenomenon which results in the formation of nitrogen-fixing nodules. Mung bean is promiscuous host however expression profile of this important legume plant in response to rhizobial infection was particularly lacking and urgently needed. We have demonstrated the pattern of gene expression of mung bean roots inoculated with two symbionts Bradyrhizobium yuanmingense Vr50 and Sinorhizobium (Ensifer) aridi Vr33 and non-inoculated control (CK). The RNA-Seq data analyzed at two growth stages i.e., 1-3 h and 10-16 days post inoculation revealed significantly higher number of differentially expressed genes (DEGs) at nodulation stage. The DEGs encoding receptor kinases identified at early stage might be involved in perception of Nod factors produced by different rhizobia. At nodulation stage important genes involved in plant hormone signal transduction, nitrogen and sulfur metabolism were identified. KEGG pathway enrichment analysis showed that metabolic pathways were most prominent in both groups (Group 1: Vr33 vs CK; Group 2: Vr50 vs CK), followed by biosynthesis of secondary metabolites, plant hormone signal transduction and biosynthesis of amino acids. Furthermore, DEGs involved in cell communication and plant hormone signal transduction were found to be different among two symbiotic systems while DEGs involved in carbon, nitrogen and sulfur metabolism were similar but their expression varied in response to two rhizobial strains. This study provides the first insight into the mechanisms underlying interactions of mung bean host with two taxonomically different symbionts (Bradyrhizobium and Sinorhizobium) and the candidate genes for better understanding the mechanisms of symbiotic host-specificity.

Contribution of potassium solubilizing bacteria in improved potassium assimilation and cytosolic K+/Na+ ratio in rice (Oryza sativa L.) under saline-sodic conditions.

Sodium-induced potassium (K+) deficiency is more prevalent in salt-affected soils. Plants experience K+ starvation thus cytosolic K+/Na+ ratio is lowered, which is a prerequisite for their survival. K+ enrichment in crops can be acquired via K-solubilizing bacteria as a sustainable green agriculture approach. This study was conducted to explore potent K-solubilizing bacteria from the rhizosphere of wheat, rice, and native flora grown in salt-affected soils in two distinct regions of Pakistan. The aim of this work was to evaluate the contribution of microbial consortiums to the improvement of K+ assimilation and cytosolic K+/Na+ ratios in rice crops under saline-sodic conditions. Among 250 bacterial isolates, 9 were selected based on their salt (11% NaCl) and alkali (9) tolerance and K-solubilization indices (1.57-5.67). These bacterial strains were characterized for their plant growth-promoting traits and identified based on 16S rRNA gene sequencing. A consortium of five strains, namely, Enterobacter hormaechei, Citrobacter braakii, Pseudomonas putida, Erwinia iniecta, and Pantoea agglomerans, was used as a bio-inoculant to evaluate its role in K+ assimilation, cytosolic K+/Na+ ratio, and subsequent yield enhancement in rice grown under saline-sodic conditions. The impact of applied consortium on rice was assessed under variable salt levels (Control, 40, 80, and 120 mM) in a pot experiment and under natural saline-sodic conditions in the field. Plant agronomical parameters were significantly higher in the bacterial consortium-treated plants, with a concomitant increase in K+-uptake in root and shoot (0.56 and 0.35 mg g-1 dry wt.) of the salt-tolerant rice variety Shaheen. The root K+/Na+ ratio was significantly improved (200% in 40 mM and 126% in 80 mM NaCl) and in the shoot (99% in 40 mM and 131% in 80 mM) too. A similar significant increase was also observed in the salt-susceptible variety Kainat. Moreover, grain yield (30.39 g/1,000 grains wt.) and biomass (8.75 g) of the rice variety Shaheen, grown in field conditions, were also improved. It can be concluded that K-solubilizing bacteria can be used as bio-inoculants, contributing to growth and yield increment via enhanced K-assimilation and cytosolic K+/Na+ ratio in rice crops under salt stress.

Mitigation of water scarcity with sustained growth of Rice by plant growth promoting bacteria.

Climate change augments the risk to food security by inducing drought stress and a drastic decline in global rice production. Plant growth-promoting bacteria (PGPB) have been known to improve plant growth under drought stress. Here in the present study, we isolated, identified, and well-characterized eight drought-tolerant bacteria from the rice rhizosphere that are tolerant to 20% PEG-8000. These strains exhibited multiple plant growth-promoting traits, i.e., 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase activity, exopolysaccharide production, phosphate (P)-solubilizing activity (51-356 µg ml-1), indole-3 acetic acid (IAA) production (14.3-46.2 µg ml-1), and production of organic acids (72-178 µg ml-1). Inoculation of bacterial consortium (Bacillus subtilis NM-2, Brucella haematophilum NM-4, and Bacillus cereus NM-6) significantly improved seedling growth and vigor index (1009.2-1100) as compared to non-inoculated stressed plants (630-957). Through rhizoscanning, efficiency of the consortium was validated by improved root parameters such as root length (17%), diameter, and surface area (18%) of all tested genotypes as compared with respective non-inoculated stressed treatments. Furthermore, the response of consortium inoculation on three rice genotypes was positively correlated with improved plant growth and drought stress ameliorating traits by the accumulation of osmoprotectant, i.e., proline (85.8%-122%), relative water content (51%), membrane stability index (64%), and production of antioxidant enzymes to reduce oxidative damage by reactive oxygen species. A decrease in temperature and improved chlorophyll content of inoculated plants were found using infrared thermal imaging and soil plant analyzer development (SPAD), respectively. The key supporting role of inoculation toward stress responses was validated using robust techniques like infrared thermal imaging and an infrared gas analyzer. Furthermore, principal component analysis depicts the contribution of inoculation on stress responses and yield of tested rice genotypes under water stress. The integration of drought-tolerant rice genotype (NIBGE-DT02) and potential bacterial strains, i.e., NM-2, NM-4, and NM-6, can serve as an effective bioinoculant to cope with water scarcity under current alarming issues related to food security in fluctuating climate.

Assessment of halotolerant bacterial and fungal consortia for augmentation of wheat in saline soils.

Adaptations of green technologies to counter abiotic stress, including salinity for crops like wheat by using halotolerant microbes, is a promising approach. The current study investigated 17 salt-affected agroecological zones from the Punjab and Sindh provinces of Pakistan to explore the potential of indigenous microbial flora, with their multiple biochemical characteristics in addition to plant growth promoting (PGP) traits, for enhanced wheat production in saline areas. Initially, 297 isolated pure bacterial colonies were screened for salt tolerance, biochemical, and PGP traits. Three bacterial strains belonging to Pantoea spp. and Erwinia rhaphontici with possession of multiple characteristics were selected for the development of the halotolerant bacterial consortium. Inoculation of two local wheat varieties, Faisalabad 2008 and Galaxy 2013, with the consortium for in vitro seed germination assay and sand microcosm experiments exhibited significant improvement of selected plant growth parameters like germination percentage and root structure. Two previously reported PGP fungal strains of Trichoderma harzianum and T. viridae were also used as fungal consortium separately for pot experiments and field trials. The pot experiments exhibited a positive correlation of consortia with metabolic viz. catalase, peroxidase, and proline and agronomical parameters including shoot length, dry weight, number of spikes, spike length, and 100 grain weight. To evaluate their performance under natural environmental conditions, field trials were conducted at three salt-affected sites. Agronomical attributes including days of flowering and maturity, flag leaf weight, length and width, shoot length, number of spikes, spike length, spike weight, number of seeds spike-1, 1,000 grain weight, and plot yield indicated the efficiency of these microbes to enhance wheat growth. Concisely, the bacterial consortium showed better performance and Faisalabad 2008 was a more resistant variety as compared to Galaxy 2013. Initial promising results indicate that further extensive research on indigenous microbes might lead to the development of Pakistan's first saline-specific biofertilizers and sustainable eco-friendly agriculture practices.

Root colonization and growth promotion of sunflower (Helianthus annuus L.) by phosphate solubilizing Enterobacter sp. Fs-11.

An Enterobacter sp. Fs-11 was isolated from sunflower rhizosphere, identified on the basis of 16S rRNA gene sequence analysis (GeneBank accession no. GQ179978) and studied for its root colonization and growth promotion ability in sunflower. Morphologically, it was rod shaped Gram-negative, motile bacterium, producing 4.5 µg mL(-1) indole acetic acid in tryptophan-supplemented medium. It utilized 27 out of 95 substrates in BIOLOG GN2 micro plate system. It was able to convert insoluble tri-calcium phosphate to soluble phosphorus up to 43.5 µg mL(-1) with decrease in pH of the medium up to 4.5 after 10 days incubation at 28 ± 2 °C in the Pikovskaya's broth. High performance liquid chromatography of cell free supernatant showed that Fs-11 produced malic acid and gluconic acid (2.43 and 16.64 µg mL(-1), respectively) in Pikovskaya's broth. Analysis of 900 bp fragment of pyrroloquinoline quinine pqqE gene sequence showed 98 % homology with that of E. cloacae pqqE gene. Confocal laser scanning microscope revealed strong colonization of fluorescently labeled Fs-11 with sunflower roots. Sunflower inoculation with Fs-11 and its rifampicin resistant derivative in sterile sand and natural soil showed that Fs-11 colonized sunflower roots up to 30 days after transplanting in both sterile sand as well as natural soil. Moreover, Fs-11 inoculation resulted in increased plant height, fresh weight, dry weight and total phosphorus contents as compared to un-inoculated plants. The data showed that Enterobacter sp. Fs-11 is an efficient phosphate solubilizing and plant growth promoting rhizobacterium and has great potential to be used as bio-inoculant for sunflower under phosphorus deficient conditions.

What Did We Learn From Current Progress in Heat Stress Tolerance in Plants? Can Microbes Be a Solution?

Temperature is a significant parameter in agriculture since it controls seed germination and plant growth. Global warming has resulted in an irregular rise in temperature posing a serious threat to the agricultural production around the world. A slight increase in temperature acts as stress and exert an overall negative impact on different developmental stages including plant phenology, development, cellular activities, gene expression, anatomical features, the functional and structural orientation of leaves, twigs, roots, and shoots. These impacts ultimately decrease the biomass, affect reproductive process, decrease flowering and fruiting and significant yield losses. Plants have inherent mechanisms to cope with different stressors including heat which may vary depending upon the type of plant species, duration and degree of the heat stress. Plants initially adapt avoidance and then tolerance strategies to combat heat stress. The tolerance pathway involves ion transporter, osmoprotectants, antioxidants, heat shock protein which help the plants to survive under heat stress. To develop heat-tolerant plants using above-mentioned strategies requires a lot of time, expertise, and resources. On contrary, plant growth-promoting rhizobacteria (PGPRs) is a cost-effective, time-saving, and user-friendly approach to support and enhance agricultural production under a range of environmental conditions including stresses. PGPR produce and regulate various phytohormones, enzymes, and metabolites that help plant to maintain growth under heat stress. They form biofilm, decrease abscisic acid, stimulate root development, enhance heat shock proteins, deamination of ACC enzyme, and nutrient availability especially nitrogen and phosphorous. Despite extensive work done on plant heat stress tolerance in general, very few comprehensive reviews are available on the subject especially the role of microbes for plant heat tolerance. This article reviews the current studies on the retaliation, adaptation, and tolerance to heat stress at the cellular, organellar, and whole plant levels, explains different approaches, and sheds light on how microbes can help to induce heat stress tolerance in plants.

Diversity and functional traits of indigenous soil microbial flora associated with salinity and heavy metal concentrations in agricultural fields within the Indus Basin region, Pakistan.

Soil salinization and heavy metal (HM) contamination are major challenges facing agricultural systems worldwide. Determining how soil microbial communities respond to these stress factors and identifying individual phylotypes with potential to tolerate these conditions while promoting plant growth could help prevent negative impacts on crop productivity. This study used amplicon sequencing and several bioinformatic programs to characterize differences in the composition and potential functional capabilities of soil bacterial, fungal, and archaeal communities in five agricultural fields that varied in salinity and HM concentrations within the Indus basin region of Pakistan. The composition of bacteria with the potential to fix atmospheric nitrogen (N) and produce the enzyme 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase were also determined. Microbial communities were dominated by: Euryarchaeota (archaea), Actinobacteria, Proteobacteria, Planctomycetota, Firimicutes, Patescibacteria and Acidobacteria (bacteria), and Ascomycota (fungi), and all soils contained phylotypes capable of N-fixation and ACC-deaminase production. Salinity influenced bacterial, but not archaeal or fungal communities. Both salinity and HM altered the relative abundance of many phylotypes that could potentially promote or harm plant growth. These stress factors also appeared to influence the potential functional capabilities of the microbial communities, especially in their capacity to cycle phosphorous, produce siderophores, and act as symbiotrophs or pathotrophs. Results of this study confirm that farms in this region are at risk due to salinization and excessive levels of some toxic heavy metals, which could negatively impact crop and human health. Changes in soil microbial communities and their potential functional capabilities are also likely to affect several critical agroecosystem services related to nutrient cycling, pathogen suppression, and plant stress tolerance. Many potentially beneficial phylotypes were identified that appear to be salt and HM tolerant and could possibly be exploited to promote these services within this agroecosystem. Future efforts to isolate these phylotypes and determine whether they can indeed promote plant growth and/or carry out other important soil processes are recommended. At the same time, identifying ways to promote the abundance of these unique phylotypes either through modifying soil and crop management practices, or developing and applying them as inoculants, would be helpful for improving crop productivity in this region.

Exploring biocontrol and growth-promoting potential of multifaceted PGPR isolated from natural suppressive soil against the causal agent of chickpea wilt.

Chickpea is an important nutritive food crop both for humans and animals. Chickpea wilt caused by Fusarium oxysporum f.sp. ciceris (Foc) results in huge yield losses every year. Chickpea being a food crop requires the development of an eco-friendly bio-pesticide to effectively control the chickpea wilt disease. In this study, more than 50 bacterial stains isolated from the rhizosphere of healthy plants growing in wilt sick soil were examined for their Foc antagonist activities. Out of these, 17 strains showing > 90% growth inhibition of Foc were then characterized for their plant growth-promoting (PGP) and biocontrol traits. The biocontrol and PGP traits identified include amylase, hydrogen cyanide, protease, cellulase, chitinase activities, p-solubilization, nitrogen-fixing, and indole-3-acetic acid production. Two bacterial strains, IR-27 and IR-57, exhibiting the highest Foc proliferation inhibition and the PGP potential along with a consortium of four different strains (Serratia sp. IN-1, Serratia sp. IS-1, Enterobacter sp. IN-2, Enterobacter sp. IN-6) were used for controlling the chickpea wilt disease and growth promotion of the chickpea plants. Confocal laser scanning microscopy revealed their root colonization ability with partial or complete elimination of broken Foc mycelia and hyphae from roots. The bacterial inoculations particularly the consortium significantly suppressed the disease and improved the overall root morphology traits (root length, root surface area, root volume, forks, tips, and crossings), resulting in enhanced growth of the chickpea plants. Significant changes in growth (107% increase in root length, 23% increase in shoot length, and 54% increase in branches) in Foc-challenged plants were observed when inoculated with the consortium. Further investigations revealed that the chickpea plants inoculated with bacterial strains induced the expression of a number of key defence enzymes, including the phenylalanine ammonia lyase, peroxidase, polyphenol peroxidase, ß-1,3 glucanase, which might have helped the plants to thwart the pathogen attack. These findings indicate the potential of our identified bacterial strains to be used as a natural biopesticide for controlling the chickpea wilt disease.

Biodegradation and Subsequent Toxicity Reduction of Co-contaminants Tribenuron Methyl and Metsulfuron Methyl by a Bacterial Consortium B2R.

AllyMax is a widely used herbicide formulation in wheat-rice cropping areas of the world. The residues of its active ingredients, tribenuron methyl (TBM) and metsulfuron methyl (MET), persist in soil and water as co-contaminants, and cause serious threats to nontarget organisms. This study was performed to assess the potential of a bacterial consortium for the degradation and detoxification of TBM and MET individually and as co-contaminants. A bacterial consortium (B2R), comprising Bacillus cereus SU-1, Bacillus velezensis OS-2, and Rhodococcus rhodochrous AQ1, capable of degrading TBM and MET in liquid cultures was developed. Biodegradation of TBM and MET was optimized using the Taguchi design of experiment. Optimum degradation of both TBM and MET was obtained at pH 7 and 37 °C. Regarding media composition, optimum degradation of TBM and MET was obtained in minimal salt medium (MSM) supplemented with glucose, and MSM without glucose, respectively. The consortium simultaneously degraded TBM and MET (94.8 and 80.4%, respectively) in cultures containing the formulation AllyMax, where TBM and MET existed as co-contaminants at 2.5 mg/L each. Mass spectrometry analysis confirmed that during biodegradation, TBM and MET were metabolized into simpler compounds. Onion (Allium cepa) root inhibition and Comet assays revealed that the bacterial consortium B2R detoxified TBM and MET separately and as co-contaminants. The consortium B2R can potentially be used for the remediation of soil and water co-contaminated with TBM and MET.

Operational parameters optimization for remediation of crude oil-polluted water in floating treatment wetlands using response surface methodology.

The application of floating treatment wetlands (FTWs) is an innovative nature-based solution for the remediation of polluted water. The rational improvement of water treatment via FTWs is typically based on multifactorial experiments which are labor-intensive and time-consuming. Here, we used the response surface methodology (RSM) for the optimization of FTW's operational parameters for the remediation of water polluted by crude oil. The central composite design (CCD) of RSM was used to generate the experimental layout for testing the effect of the variables hydrocarbon, nutrient, and surfactant concentrations, aeration, and retention time on the hydrocarbon removal in 50 different FTW test systems planted with the common reed, Phragmites australis. The results from these FTW were used to formulate a mathematical model in which the computational data strongly correlated with the experimental results. The operational parameters were further optimized via modeling prediction plus experimental validation in test FTW systems. In the FTW with optimized parameters, there was a 95% attenuation of the hydrocarbon concentration, which was very close to the 98% attenuation predicted by the model. The cost-effectiveness ratio showed a reduction of the treatment cost up to $0.048/liter of wastewater. The approach showed that RSM is a useful strategy for designing FTW experiments and optimizing operational parameters.

Designing Synergistic Biostimulants Formulation Containing Autochthonous Phosphate-Solubilizing Bacteria for Sustainable Wheat Production.

Applying phosphate-solubilizing bacteria (PSB) as biofertilizers has enormous potential for sustainable agriculture. Despite this, there is still a lack of information regarding the expression of key genes related to phosphate-solubilization (PS) and efficient formulation strategies. In this study, we investigated rock PS by Ochrobactrum sp. SSR (DSM 109610) by relating it to bacterial gene expression and searching for an efficient formulation. The quantitative PCR (qPCR) primers were designed for PS marker genes glucose dehydrogenase (gcd), pyrroloquinoline quinone biosynthesis protein C (pqqC), and phosphatase (pho). The SSR-inoculated soil supplemented with rock phosphate (RP) showed a 6-fold higher expression of pqqC and pho compared to inoculated soil without RP. Additionally, an increase in plant phosphorous (P) (2%), available soil P (4.7%), and alkaline phosphatase (6%) activity was observed in PSB-inoculated plants supplemented with RP. The root architecture improved by SSR, with higher root length, diameter, and volume. Ochrobactrum sp. SSR was further used to design bioformulations with two well-characterized PS, Enterobacter spp. DSM 109592 and DSM 109593, using the four organic amendments, biochar, compost, filter mud (FM), and humic acid. All four carrier materials maintained adequate survival and inoculum shelf life of the bacterium, as indicated by the field emission scanning electron microscopy analysis. The FM-based bioformulation was most efficacious and enhanced not only wheat grain yield (4-9%) but also seed P (9%). Moreover, FM-based bioformulation enhanced soil available P (8.5-11%) and phosphatase activity (4-5%). Positive correlations were observed between the PSB solubilization in the presence of different insoluble P sources, and soil available P, soil phosphatase activity, seed P content, and grain yield of the field grown inoculated wheat variety Faisalabad-2008, when di-ammonium phosphate fertilizer application was reduced by 20%. This study reports for the first time the marker gene expression of an inoculated PSB strain and provides a valuable groundwork to design field scale formulations that can maintain inoculum dynamics and increase its shelf life. This may constitute a step-change in the sustainable cultivation of wheat under the P-deficient soil conditions.

Inoculation With Azospirillum spp. Acts as the Liming Source for Improving Growth and Nitrogen Use Efficiency of Potato.

Nitrogen (N) is one of the limiting factors for plant growth, and it is mainly supplied exogenously by fertilizer application. It is well documented that diazotrophic rhizobacteria improve plant growth by fixing atmospheric N in the soil. The present study investigates the nitrogen-fixing potential of two Azospirillum spp. strains using the 15N isotope-dilution method. The two diazotrophic strains (TN03 and TN09) native to the rhizosphere of potato belong to the genus Azospirillum (16S rRNA gene accession numbers LN833443 and LN833448, respectively). Both strains were able to grow on an N-free medium with N-fixation potential (138-143 nmol mg-1 protein h-1) and contained the nifH gene. Strain TN03 showed highest indole acetic acid (IAA) production (30.43 µg/mL), while TN09 showed highest phosphate solubilization activity (249.38 µg/mL) while both diazotrophs showed the production of organic acids. A 15N dilution experiment was conducted with different fertilizer inputs to evaluate the N-fixing potential of both diazotrophs in pots. The results showed that plant growth parameters and N contents increased significantly by the inoculations. Moreover, reduced 15N enrichment was found compared to uninoculated controls that received similar N fertilizer levels. This validates the occurrence of N-fixation through isotopic dilution. Strain TN09 showed higher N-fixing potential than TN03 and the uninoculated controls. Inoculation with either strain also showed a remarkable increase in plant growth under field conditions. Thus, there were remarkable increases in N use efficiency, N uptake and N utilization levels. Confocal laser scanning and transmission electron microscopy showed that TN03 is an ectophyte, i.e., present outside root cells or within the grooves of root hairs, while TN09 is an endophyte, i.e., present within root cells, forming a strong association withroot it. This study confirms that diazotrophic Azospirillum spp. added to potato systems can improve plant growth and N use efficiency, opening avenues for improvement of potato crop growth with reduced input of N fertilizer.

Phylogenetic diversity analysis reveals Bradyrhizobium yuanmingense and Ensifer aridi as major symbionts of mung bean (Vigna radiata L.) in Pakistan.

The present study was carried out to evaluate the diversity of rhizobia associated with nodules of mung bean in Pakistan, because this information is necessary for inoculum development. Based on sequence analysis of 16S rRNA gene of thirty-one bacteria, 11 were assigned to genus Bradyrhizobium, 17 to Ensifer, and 3 to Rhizobium. Phylogenetic analyses on the basis of 16S-23S ITS region, atpD, recA, nifH, and nodA of representative strains revealed that B. yuanmingense is the predominant species distributed throughout different mung bean-growing areas. Among the fast-growing rhizobia, Ensifer aridi was predominant in Faisalabad, Layyah, and Rawalpindi, while E. meliloti in Thal desert. Sequence variations and phylogeny of nifH and nodA genes suggested that these genes might have been co-evolved with the housekeeping genes and maintained by vertical gene transfer in rhizobia detected in the present study. Host infectivity assay revealed the successful nodulation of host by rhizobia related to genera Bradyrhizobium, Ensifer and Rhizobium. Among all, Bradyrhizobium and Ensifer spp. inoculation exhibited a significantly higher number of nodules (11-34 nodules plant-1) and nitrogenase activity (nodule ARA 60-110 µmol g-1 h-1). Contrary to the previous studies, our data reveal that B. yuanmingense and E. aridi are predominant species forming effective nodules in mung bean in Pakistan. Furthermore, to the best of our knowledge, this is the first report showing the effective symbiosis of E. aridi, E. meliloti, and Rhizobium pusense with mung bean. The diversity of rhizobia in different habitats revealed in the present study will contribute towards designing site-specific inocula for mung bean.

Bioactive Nutrient Fortified Fertilizer: A Novel Hybrid Approach for the Enrichment of Wheat Grains With Zinc.

Zinc (Zn) is a critical micronutrient that synergizes nutrient use efficiency, and improves plant growth and human health. Low Zn bioavailability in soils affects produce quality and agricultural productivity worldwide ultimately inducing deficiency in humans and animals. Zn deficiency is a leading cause of malnutrition in underdeveloped countries where a widespread population depends upon staple cereals for daily intake of calories. Modern cereal cultivars are inherently low in Zn, eventually, plants need to be enriched with soil application of ZnSO4, but due to higher fixation losses, it becomes an inefficient source. Rhizosphere microbiome contains Zn-solubilizing bacteria (ZSB) that improve Zn bioavailability, thus increase the root function, Zn uptake, and plant growth. Niha Corp developed a hybrid process of bioactive nutrient fortified fertilizer (BNFF), which has been used to formulate Zabardast Urea (ZU) by coating bioactive Zn (BAZ) and ZSB on urea. Data obtained for 15 wheat varieties from 119 farmer field demonstration plots and eight replicated trials on 42 locations across multi-environment conditions conclude that ZU significantly improved the plant biomass and yield by 12% over non-Zn control and produced grains with 57 µg/g Zn contents, which can meet a major part of the recommended dietary allowance (RDA) of humans. The study recommends that this microbe-mediated hybrid invention (ZU) is a feasible approach to boost Zn bioavailability and Zn use efficiency, with enhanced yield and quality that may contribute to improve human health. To the best of our knowledge, this is the first wide-scale field testing of Zn enrichment in the grains of bread wheat using an innovative BNFF Urea Z technology.

Impact of economic capabilities and population agglomeration on PM2.5 emission: empirical evidence from sub-Saharan African countries.

The utilization of economic capabilities to raise production in the economy enhances the industrial activities and use of transportation. These activities deteriorate the quality of the environment and raise the level of particulate matter (PM2.5). The objective of this study is to investigate the impact of economic capabilities and population agglomeration on PM2.5 emissions for the sample of 23 sub-Saharan African countries-a highly polluted region of the world. The study used panel regression from 2007 to 2015 and found that production capabilities are highly significant and directly effecting PM2.5 emissions. The Kuznets hypothesis does not hold in this region, and the concentration of population in urban areas is also significant. To avoid the problem of possible endogeneity in the empirical model and for robustness check, the study also utilized system GMM and found consistent results. The findings of the study are highly valuable to reduce PM2.5 and propose that the policymakers in Sub-Saharan African countries should opt for industrial specialization rather than economic complexity and ensure that the proper measures are taken to control PM2.5. Moreover, population concentrations should not be taken place in a few main urban cities.