Corn microbial diversity and its relationship to yield.

This study aimed to identify possible relationships between corn (Zea mays L.) productivity and its endosphere microbial community. Any insights would be used to develop testable hypotheses at the farm level. Sap was collected from 14 fields in 2014 and 10 fields in 2017, with a yield range of 10.1 to 21.7 tonnes per hectare (t/ha). The microbial sap communities were analyzed using terminal restriction fragment length polymorphism (TRFLP) and identified using an internal pure culture reference database and BLAST. This technique is rapid and inexpensive and is suitable for use at the grower level. Diversity, richness, and normalized abundances of each bacterial population in corn sap samples were evaluated to link the microbiome of a specific field to its yield. A negative trend was observed (r = -0.60), with higher-yielding fields having lower terminal restriction fragment (TRF) richness. A partial least square regression analysis of TRF intensity and binary data from 2014 identified 10 TRFs (bacterial genera) that positively, or negatively, correlated with corn yields, when either absent or present at certain levels or ratios. Using these observations, a model was developed that accommodated criteria for each of the 10 microbes and assigned a score for each field out of 10. Data collected in 2014 showed that sites with higher model scores were highly correlated with larger yields (r = 0.83). This correlation was also seen when the 2017 data set was used (r = 0.87). We were able to conclude that a positive significant effect was seen with the model score and yield (adjusted R2 = 0.67, F[1,22] = 46.7, p < 0.001) when combining 2014 and 2017 data. The results of this study are being expanded to identify the key microbes in the corn sap community that potentially impact corn yield, regardless of corn variety, geographic factors, or edaphic factors.

Effect of Chemical Fertilization on the Impacts of Plant Growth-Promoting Rhizobacteria in Maize Crops.

The use of chemical fertilizers strongly promotes productivity in agricultural crops; therefore, large amounts of chemical fertilizers have been used. The use of plant growth-promoting bacteria may be a strategy to reduce the use of chemical fertilizers; however, little is known about the effect of chemical fertilization on the performance of these bacteria through plant-microbe interactions. The present study aimed to verify the performance of Bacillus subtilis, Azospirillum brasilense, B. pumilus, B. amyloliquefaciens, Herbaspirillum seropedicae, Gluconacetobacter diazotrophicus, and the mixtures A. brasilense + B. subtilis, B. pumilus + B. amyloliquefaciens, and H. seropedicae + G. diazotrophicus on parameters such as nitrogen and phosphorus extraction from soil, the concentrations of these nutrients in maize plants, and plant growth in both fertilized and unfertilized soil. The results showed that H. seropedica increased the nitrogen content by 6.6 g kg-1 in leaves and 2.2 g kg-1 in the root when comparing the unfertilized with the fertilized condition. G. diazotrophicus increased the nitrogen content by 3.7 g kg-1 in leaves and 2.4 g kg-1 in the root. B. pumilus increased the phosphorous content by 1.7 g kg-1 in leaves, and B. amyloliquefaciens increased the phosphorous content by 0.61 g kg-1. The present study showed that even though the bacteria presented good performance related to plant growth under fertilized conditions, H. seropedicae, G. diazotrophicus, B. pumilus, and B. amyloliquefaciens could be used in the maize crop with a reduced chemical fertilization dose.

λ-Carrageenan Suppresses Tomato Chlorotic Dwarf Viroid (TCDVd) Replication and Symptom Expression in Tomatoes.

The effect of carrageenans on tomato chlorotic dwarf viroid (TCDVd) replication and symptom expression was studied. Three-week-old tomato plants were spray-treated with iota(É©)-, lambda(λ)-, and kappa(κ)-carrageenan at 1 g·L-1 and inoculated with TCDVd after 48 h. The λ-carrageenan significantly suppressed viroid symptom expression after eight weeks of inoculation, only 28% plants showed distinctive bunchy-top symptoms as compared to the 82% in the control group. Viroid concentration was reduced in the infected shoot cuttings incubated in λ-carrageenan amended growth medium. Proteome analysis revealed that 16 tomato proteins were differentially expressed in the λ-carrageenan treated plants. Jasmonic acid related genes, allene oxide synthase (AOS) and lipoxygenase (LOX), were up-regulated in λ-carrageenan treatment during viroid infection. Taken together, our results suggest that λ-carrageenan induced tomato defense against TCDVd, which was partly jasmonic acid (JA) dependent, and that it could be explored in plant protection against viroid infection.

Prebiotic effects of diet supplemented with the cultivated red seaweed Chondrus crispus or with fructo-oligo-saccharide on host immunity, colonic microbiota and gut microbial metabolites.

BACKGROUND: Gastrointestinal microbial communities are diverse and are composed of both beneficial and pathogenic groups. Prebiotics, such as digestion-resistant fibers, influence the composition of gut microbiota, and can contribute to the improvement of host health. The red seaweed Chondrus crispus is rich in dietary fiber and oligosaccharides, however its prebiotic potential has not been studied to date. METHODS: Prebiotic effects were investigated with weaning rats fed a cultivated C. crispus-supplemented diet. Comparison standards included a fructo-oligo-saccharide (FOS) diet and a basal diet. The colonic microbiome was profiled with a 16S rRNA sequencing-based Phylochip array. Concentrations of short chain fatty acids (SCFAs) in the feacal samples were determined by gas chromatography with a flame ionization detector (GC-FID) analysis. Immunoglobulin levels in the blood plasma were analyzed with an enzyme-linked immunosorbent assay (ELISA). Histo-morphological parameters of the proximal colon tissue were characterized by hematoxylin and eosin (H&E) staining. RESULTS: Phylochip array analysis indicated differing microbiome composition among the diet-supplemented and the control groups, with the C. crispus group (2.5% supplementation) showing larger separation from the control than other treatment groups. In the 2.5% C. crispus group, the population of beneficial bacteria such as Bifidobacterium breve increased (4.9-fold, p=0.001), and the abundance of pathogenic species such as Clostridium septicum and Streptococcus pneumonia decreased. Higher concentrations of short chain fatty acids (i.e., gut microbial metabolites), including acetic, propionic and butyric acids, were found in faecal samples of the C. crispus-fed rats. Furthermore, both C. crispus and FOS supplemented rats showed significant improvements in proximal colon histo-morphology. Higher faecal moisture was noted in the 2.5% C. crispus group, and elevated plasma immunoglobulin (IgA and IgG) levels were observed in the 0.5% C. crispus group, as compared to the basal feed group. CONCLUSIONS: The results suggest multiple prebiotic effects, such as influencing the composition of gut microbial communities, improvement of gut health and immune modulation in rats supplemented with cultivated C. crispus.

Transcriptional and metabolomic analysis of Ascophyllum nodosum mediated freezing tolerance in Arabidopsis thaliana.

BACKGROUND: We have previously shown that lipophilic components (LPC) of the brown seaweed Ascophyllum nodosum (ANE) improved freezing tolerance in Arabidopsis thaliana. However, the mechanism(s) of this induced freezing stress tolerance is largely unknown. Here, we investigated LPC induced changes in the transcriptome and metabolome of A. thaliana undergoing freezing stress. RESULTS: Gene expression studies revealed that the accumulation of proline was mediated by an increase in the expression of the proline synthesis genes P5CS1 and P5CS2 and a marginal reduction in the expression of the proline dehydrogenase (ProDH) gene. Moreover, LPC application significantly increased the concentration of total soluble sugars in the cytosol in response to freezing stress. Arabidopsis sfr4 mutant plants, defective in the accumulation of free sugars, treated with LPC, exhibited freezing sensitivity similar to that of untreated controls. The 1H NMR metabolite profile of LPC-treated Arabidopsis plants exposed to freezing stress revealed a spectrum dominated by chemical shifts (δ) representing soluble sugars, sugar alcohols, organic acids and lipophilic components like fatty acids, as compared to control plants. Additionally, 2D NMR spectra suggested an increase in the degree of unsaturation of fatty acids in LPC treated plants under freezing stress. These results were supported by global transcriptome analysis. Transcriptome analysis revealed that LPC treatment altered the expression of 1113 genes (5%) in comparison with untreated plants. A total of 463 genes (2%) were up regulated while 650 genes (3%) were down regulated. CONCLUSION: Taken together, the results of the experiments presented in this paper provide evidence to support LPC mediated freezing tolerance enhancement through a combination of the priming of plants for the increased accumulation of osmoprotectants and alteration of cellular fatty acid composition.

Tasco®: a product of Ascophyllum nodosum enhances immune response of Caenorhabditis elegans against Pseudomonas aeruginosa infection.

The effects of Tasco®, a product made from the brown seaweed (Ascophyllum nodosum) were tested for the ability to protect Caenorhabditis elegans against Pseudomonas aeruginosa infection. A water extract of Tasco® (TWE) reduced P. aeruginosa inflicted mortality in the nematode. The TWE, at a concentration of 300 µg/mL, offered the maximum protection and induced the expression of innate immune response genes viz.; zk6.7 (Lypases), lys-1 (Lysozyme), spp-1 (Saponin like protein), f28d1.3 (Thaumatin like protein), t20g5.7 (Matridin SK domain protein), abf-1 (Antibacterial protein) and f38a1.5 (Lectin family protein). Further, TWE treatment also affected a number of virulence components of the P. aeuroginosa and reduced its secreted virulence factors such as lipase, proteases and toxic metabolites; hydrogen cyanide and pyocyanin. Decreased virulence factors were associated with a significant reduction in expression of regulatory genes involved in quorum sensing, lasI, lasR, rhlI and rhlR. In conclusion, the TWE-treatment protected the C. elegans against P. aeruginosa infection by a combination of effects on the innate immunity of the worms and direct effects on the bacterial quorum sensing and virulence factors.

Tasco(®), a product of Ascophyllum nodosum, imparts thermal stress tolerance in Caenorhabditis elegans.

Tasco(®), a commercial product manufactured from the brown alga Ascophyllum nodosum, has been shown to impart thermal stress tolerance in animals. We investigated the physiological, biochemical and molecular bases of this induced thermal stress tolerance using the invertebrate animal model, Caenorhabiditis elegans. Tasco(®) water extract (TWE) at 300 µg/mL significantly enhanced thermal stress tolerance as well as extended the life span of C. elegans. The mean survival rate of the model animals under thermal stress (35 °C) treated with 300 µg/mL and 600 µg/mL TWE, respectively, was 68% and 71% higher than the control animals. However, the TWE treatments did not affect the nematode body length, fertility or the cellular localization of daf-16. On the contrary, TWE under thermal stress significantly increased the pharyngeal pumping rate in treated animals compared to the control. Treatment with TWE also showed differential protein expression profiles over control following 2D gel-electrophoresis analysis. Furthermore, TWE significantly altered the expression of at least 40 proteins under thermal stress; among these proteins 34 were up-regulated while six were down-regulated. Mass spectroscopy analysis of the proteins altered by TWE treatment revealed that these proteins were related to heat stress tolerance, energy metabolism and a muscle structure related protein. Among them heat shock proteins, superoxide dismutase, glutathione peroxidase, aldehyde dehydrogenase, saposin-like proteins 20, myosin regulatory light chain 1, cytochrome c oxidase RAS-like, GTP-binding protein RHO A, OS were significantly up-regulated, while eukaryotic translation initiation factor 5A-1 OS, 60S ribosomal protein L18 OS, peroxiredoxin protein 2 were down regulated by TWE treatment. These results were further validated by gene expression and reporter gene expression analyses. Overall results indicate that the water soluble components of Tasco(®) imparted thermal stress tolerance in the C. elegans by altering stress related biochemical pathways.

Understanding the molecular basis of plant growth promotional effect of Pseudomonas fluorescens on rice through protein profiling.

BACKGROUND: Plant Growth Promoting Rhizobacteria (PGPR), Pseudomonas fluorescens strain KH-1 was found to exhibit plant growth promotional activity in rice under both in-vitro and in-vivo conditions. But the mechanism underlying such promotional activity of P. fluorescens is not yet understood clearly. In this study, efforts were made to elucidate the molecular responses of rice plants to P. fluorescens treatment through protein profiling. Two-dimensional polyacrylamide gel electrophoresis strategy was adopted to identify the PGPR responsive proteins and the differentially expressed proteins were analyzed by mass spectrometry. RESULTS: Priming of P. fluorescens, 23 different proteins found to be differentially expressed in rice leaf sheaths and MS analysis revealed the differential expression of some important proteins namely putative p23 co-chaperone, Thioredoxin h- rice, Ribulose-bisphosphate carboxylase large chain precursor, Nucleotide diPhosphate kinase, Proteosome sub unit protein and putative glutathione S-transferase protein. CONCLUSION: Functional analyses of the differential proteins were reported to be directly or indirectly involved in growth promotion in plants. Thus, this study confirms the primary role of PGPR strain KH-1 in rice plant growth promotion.

Introducing key microbes from high productive soil transforms native soil microbial community of low productive soil.

This study aimed to understand the changes in rhizosphere microbial structure and diversity of an average corn yielding field site soil with the introduced microbial candidates from a high-yielding site. Soils used in this study were from two growers' fields located in Dunnville, Ontario, Canada, where one of the farms has an exceptional high corn yield (G-site soil; ca 20 tons/acre) and the other yields an average crop (H-site soil; 12 tons/acre) (8 years of unpublished A & L data). In growth room experiments using wheat as the indicator crop, calcium alginate beads with microbes composed of Azospirillum lipoferum, Rhizobium leguminosarum, Burkholderia ambifaria, Burkholderia graminis, Burkholderia vietnamiensis, Pseudomonas lurida, Exiguobacterium acetylicum, Kosakonia cowanii, and Paenibacillus polymyxa was introduced into the soil at planting to the average-yielding soil. These bacteria had been isolated from the high-yielding farm soil. Among the nine microbial candidates tested, three (P. polymyxa, E. acetylicum and K. cowanii) significantly impacted the plant health and biometrics in addition to microbial richness and diversity, where the microbial profile became very similar to the high productive G-site soil. One hundred and forty-two bacterial terminal restriction fragments (TRFs) were involved in the community shift and 48 of them showed significant correlation to several interacting soil factors. This study indicates the potential of shifting microbial profiles of average-yielding soils by introducing key candidates from highly productive soils to increase biological soil health.

Sugarcane growth and nutrition levels are differentially affected by the application of PGPR and cane waste.

Mineral and organic fertilization can be optimized by using rhizobacteria which increases dry matter, yield, and nutrients in the soil and plant, among the other biological inputs. However, the discovery of single microbes or a consortium that can benefit plants has been a challenge. In this context, this study aimed to evaluate the effects of Bacillus subtilis and Bacillus pumilus combined with mineral fertilization and sugar and alcohol industry by-products in presprouted and the initial growth phase of sugar cane seedlings. The study was carried out in two phases. Phase 1 included presprouted seedlings with T1 =  untreated control, T2 =  B. subtilis, T3 =  B. pumilus, and T4 =  B. subtilis + B. pumilus treatments. Phase 2 included the same treatments with four types of fertilization: F1 =  mineral fertilization, F2 =  mineral fertilization + vinasse, F3 =  mineral fertilization + filter cake, and F4 =  mineral fertilization + filter cake compost. Of the phase 1 treatments, T2 (B. subtilis) was the best promoter of root growth and the total dry matter compared to the control with an increase of 23.0% compared to the control. In phase 2, B. pumilus application, increased the total dry matter by 13%, the number of tillers by 37%, and the diameter of the tillers by 48% when combined with mineral fertilization. The combined application of B. subtilis and B. pumilus increased the phosphorus content by 13% in soil treated with mineral fertilization and filter cake compost. The results of the this study strongly suggest that the use of B. subtilis and B. pumilus together with these by-products can improve soil fertility parameters and decrease adverse effects associated with vinasse fertilization, in addition to providing shoot and root growth and providing collective synergy for a high yield of sugarcane production with environmental benefits.

Gene expression during imidacloprid-induced hormesis in green peach aphid.

Imidacloprid-induced hormesis in the form of stimulated reproduction has previously been reported in green peach aphid, Myzus persicae. Changes in gene expression accompanying this hormetic response have not been previously investigated. In this study, expression of stress response (Hsp60), dispersal (OSD, TOL and ANT), and developmental (FPPS I) genes were examined for two generations during imidacloprid-induced reproductive stimulation in M. persicae. Global DNA methylation was also measured to test the hypothesis that changes in gene expression are heritable. At hormetic concentrations, down-regulation of Hsp60 was followed by up-regulation of this gene in the subsequent generation. Likewise, expression of dispersal-related genes and FPPS I varied with concentration, life stage, and generation. These results indicate that reproductive hormesis in M. persicae is accompanied by a complex transgenerational pattern of up- and down-regulation of genes that likely reflects trade-offs in gene expression and related physiological processes during the phenotypic dose-response. Moreover, DNA methylation in second generation M. persicae occurred at higher doses than in first-generation aphids, suggesting that heritable adaptability to low doses of the stressor might have occurred.

Efficacy of alginate- and clay-encapsulated microorganisms on the growth of Araçá-Boi seedlings (Eugenia stipitata)

The present work aimed to evaluate the effects of encapsulated microorganisms on seedlings of Eugenia stipitata, popularly known as araçá-boi, to evaluate the interaction between the inoculum and encapsulating agents such as clay and alginate. The experiment was carried out in a completely randomized design using a 3×2 factorial scheme. The treatments were control, inoculum, clay without microbial inoculum, clay with microbial inoculum, alginate without microbial inoculum, and alginate with microbial inoculum. The seedlings were grown under nursery conditions over a period of 3 months. No treatment increased the height, stem diameter, shoot dry matter or root dry matter of the araçá-boi seedlings. The use of alginate increased the ammonium content compared to the clay and control treatments. Alginate and clay increased the nitrate content in relation to the control. Alginate increased the total number of bacteria in relation to the clay and control treatments. The application of inoculum combined with alginate increased the nitrate content only in relation to the clay and control treatments. Although the application of inoculum promoted an increase in the nitrate content compared to the uninoculated treatments, there was no effect for the other parameters analyzed. The results suggest that clay and alginate encapsulating agents with the presence or absence of microorganisms may improve some soil parameters.

Efficacy of alginate- and clay-encapsulated microorganisms on the growth of Araçá-Boi seedlings (Eugenia stipitata)

The present work aimed to evaluate the effects of encapsulated microorganisms on seedlings of Eugenia stipitata, popularly known as araçá-boi, to evaluate the interaction between the inoculum and encapsulating agents such as clay and alginate. The experiment was carried out in a completely randomized design using a 3×2 factorial scheme. The treatments were control, inoculum, clay without microbial inoculum, clay with microbial inoculum, alginate without microbial inoculum, and alginate with microbial inoculum. The seedlings were grown under nursery conditions over a period of 3 months. No treatment increased the height, stem diameter, shoot dry matter or root dry matter of the araçá-boi seedlings. The use of alginate increased the ammonium content compared to the clay and control treatments. Alginate and clay increased the nitrate content in relation to the control. Alginate increased the total number of bacteria in relation to the clay and control treatments. The application of inoculum combined with alginate increased the nitrate content only in relation to the clay and control treatments. Although the application of inoculum promoted an increase in the nitrate content compared to the uninoculated treatments, there was no effect for the other parameters analyzed. The results suggest that clay and alginate encapsulating agents with the presence or absence of microorganisms may improve some soil parameters.(AU)

A fucose containing polymer-rich fraction from the brown alga Ascophyllum nodosum mediates lifespan increase and thermal-tolerance in Caenorhabditis elegans, by differential effects on gene and protein expression.

The extracts of the brown alga, Ascophyllum nodosum, which contains several bioactive compounds, have been shown to impart biotic and abiotic stress tolerance properties when consumed by animals. However, the physiological, biochemical and molecular mechanism underlying such effects remain elusive. We investigated the effect of A. nodosum fucose-containing polymer (FCP) on tolerance to thermally induced stress using the invertebrate animal model, Caenorhabditis elegans. FCP at a concentration of 150 µg mL(-1) significantly improved the life span and tolerance against thermally induced stress in C. elegans. The treatment increased the C. elegans survival by approximately 24%, when the animals were under severe thermally induced stress (i.e. 35 °C) and 27% under mild stress (i.e. 30 °C) conditions. The FCP induced differential expression of genes and proteins is associated with stress response pathways. Under thermal stress, FCP treatment significantly altered the expression of 65 proteins (54 up-regulated & 11 down-regulated). Putative functional analysis of FCP-induced differential proteins signified an association of altered proteins in stress-related molecular and biochemical pathways of the model worm.