Impact of high temperature on sucrose translocation,sugar content and inulin yield in <Emphasis Type="Italic">Cichorium intybus</Emphasis> L. var. <Emphasis Type="Italic">sativum</Emphasis>

Aims

Dianthus caryophyllus is a commercially important ornamental flower. Plant growth promoting rhizobacteria are increasingly applied as bio-fertilisers and bio-fortifiers. We studied the effect of a rhizospheric isolate Klebsiella SGM 81 strain to promote D. caryophyllus growth under sterile and non-sterile conditions, to colonise its root system endophytically and its impact on the cultivatable microbial community. We identified the auxin indole-3-acetic acid (IAA) production of Klebsiella SGM 81 as major bacterial trait most likely to enhance growth of D. caryophyllus.

Methods

ipdC dependent IAA production of SGM 81 was quantified using LC-MS/MS and localised proximal to D. caryophyllus roots and correlated to root growth promotion and characteristic morphological changes. SGM 81 cells were localised on and within the plant root using 3D rendering confocal microscopy of gfp expressing SGM 81. Using Salkowski reagent IAA production was quantified and localised proximal to roots in situ. The effect of different bacterial titres on rhizosphere bacterial population was CFU enumerated on nutrient agar. The genome sequence of Klebsiella SGM 81 (accession number PRJEB21197) was determined to validate PGP traits and phylogenic relationships.

Results

Inoculation of D. caryophyllus roots with Klebsiella SGM 81 drastically promoted plant growth when grown in agar and soil, concomitant with a burst in root hair formation, suggesting an increase in root auxin activity. We sequenced the Klebsiella SGM 81 genome, identified the presence of a canonical ipdC gene in Klebsiella SGM 81, confirmed bacterial production and secretion of IAA in batch culture using LC-MS/MS and localised plant dependent IAA production by SGM 81 proximal to roots. We found Klebsiella SGM 81 to be a rhizoplane and endophytic coloniser of D. caryophyllus roots in a dose dependent manner. We found no adverse effects of SGM 81 on the overall rhizospheric microbial population unless supplied to soil in very high titres.

Conclusion

Klebsiella SGM 81 effectively improves root traits of D. caryophyllus in a dose dependent manner, likely through tryptophan dependent IAA production in the rhizoplane and potentially within the intercellular spaces of root tissue. Under optimal plant growth promoting conditions in non-sterile soil, the high total microbial titre in the rhizosphere supports a mutualistic relationship between Klebsiella SGM 81 and carnation that potentially extends to the wider rhizosphere microbiota.

     

Expression on roots and contribution to maize phytostimulation of 1-aminocyclopropane-1-decarboxylate deaminase gene <Emphasis Type="Italic">acdS</Emphasis> in <Emphasis Type="Italic">Pseudomonas fluorescens</Emphasis> F113

Aims

The plant-beneficial bacterium Pseudomonas fluorescens F113 harbours an acdS gene, which enables deamination of 1-aminocyclopropane-1-carboxylate. The impact of abiotic and biotic factors on the expression of this gene was assessed, as well as the plant-beneficial properties of F113 under different soil moistures.

Methods

An acdS-egfp biosensor was constructed in F113, validated in vitro and used to analyse, by microscopy, its expression on roots of Zea mays comparatively to Beta vulgaris. An acdS mutant was constructed and compared with the wild-type to characterize plant-beneficial effects of F113 on maize lines EP1 and FV2, under well-watered and water deficit conditions.

Results

Different patterns of root colonization and acdS expression were observed according to plant genotype. acdS rhizoplane expression was higher on Beta vulgaris, and on maize line FV2 and hybrid PR37Y15 than on maize line EP1 and teosinte. Strain F113 but not its acdS mutant promoted root growth of EP1 under well-watered conditions and germination of FV2 under water deficit conditions.

Conclusions

Maize lines differed in their ability to induce acdS expression and to respond to P. fluorescens F113. The maize line leading to higher acdS expression, FV2, was the one benefiting from inoculation under water deficit.

     

<Emphasis Type="Italic">Azospirillum</Emphasis> spp. from native forage grasses in Brazilian Pantanal floodplain: biodiversity and plant growth promotion potential

A sustainable alternative to improve yield and the nutritive value of forage is the use of plant growth-promoting bacteria (PGPB) that release nutrients, synthesize plant hormones and protect against phytopathogens (among other mechanisms). Azospirillum genus is considered an important PGPB, due to the beneficial effects observed when inoculated in several plants. The aim of this study was to evaluate the diversity of new Azospirillum isolates and select bacteria according to the plant growth promotion ability in three forage species from the Brazilian Pantanal floodplain: Axonopus purpusii, Hymenachne amplexicaulis and Mesosetum chaseae. The identification of bacterial isolates was performed using specific primers for Azospirillum in PCR reactions and partial sequencing of the 16S rRNA and nifH genes. The isolates were evaluated in vitro considering biological nitrogen fixation (BNF) and indole-3-acetic acid (IAA) production. Based on the results of BNF and IAA, selected isolates and two reference strains were tested by inoculation. At 31 days after planting the plant height, shoot dry matter, shoot protein content and root volume were evaluated. All isolates were able to fix nitrogen and produce IAA, with values ranging from 25.86 to 51.26 mg N mL^?1 and 107–1038 µmol L^?1, respectively. The inoculation of H. amplexicaulis and A. purpusii increased root volume and shoot dry matter. There were positive effects of Azospirillum inoculation on Mesosetum chaseae regarding plant height, shoot dry matter and root volume. Isolates MAY1, MAY3 and MAY12 were considered promising for subsequent inoculation studies in field conditions.     

The benefits of foliar inoculation with <Emphasis Type="Italic">Azospirillum brasilense</Emphasis> in soybean are explained by an auxin signaling model

Azospirillum sp. is one of the most studied genera of plant growth-promoting rhizobacteria (PGPR). The ability of Azospirillum sp. to promote plant growth has been associated with its ability to produce several phytohormones, such as auxins, gibberellins and cytokinins, but mainly indole-3-acetic acid (IAA). It has been propoosed that the production of IAA explains the positive effects of co-inoculation with Azospirillum sp. on the rhizobia-legume symbiosis. In this study, we constructed an IAA-deficient mutant of A. brasilense Az39 (ipdC ^? ) by using a restriction-free cloning method. We inoculated soybean seeds with 1·10⁶ cfu·seed^?1 of Bradyrhizobium japonicum E109 and co-inoculating leaves at the V3 stage with 1·10⁸ cfu.plant^?1 of A. brasilense Az39 wt or ipdC ^? or inoculated leaves with 20 μg.plant^?1 synthetic IAA. The results confirmed soybean growth promotion as there was increased total plant and root length, aerial and root dry weight, number of nodules on the primary root, and an increase in the symbiosis established with B. japonicum E109. Nodule weight also increased after foliar co-inoculation with the IAA- producer A. brasilense Az39. The exogenous application of IAA decreased aerial and root length, as well as the number of nodules on primary roots in comparison with the Az39 wt strain. These results allow us to propose a biological model of response to foliar co-inoculation of soybean with IAA-producing rhizobacteria. This model clearly shows that both the presence of microorganism as part of the colonization process and the production of IAA in situ are co-responsible, via plant signaling molecules, for the positive effects on plant growth and symbiosis establishment.     

Isolation and characterization of nitrogen-fixing bacteria of the genus <Emphasis Type="Italic">Azospirillum</Emphasis> from the soil of a <Emphasis Type="Italic">Sphagnum</Emphasis> peat bog

he presence of nitrogen-fixing bacteria of the genus Azospirillum in the soils of acidic raised Sphagnum bogs is revealed for the first time. Three Azospirillum strains, B2, B21, and B22, were isolated as a component of methane-oxidizing enrichment cultures, whereas attempts to isolate them directly from peat samples have failed. The results of comparative analysis of the nucleotide sequences of 16S rRNA genes, DNA-DNA hybridization, and the analysis of the sequences of the functional genes encoding nitrogenase and ribulose-1, 5-bisphosphate carboxylase reveal that all the newly obtained strains can be classified as Azospirillum lipoferum. Yet, unlike A. lipoferum, the isolates do not require biotin and utilize sucrose, inositol, and glycerol for growth. The cell morphology of strain B2 differs from that of the type strain and strains B21 and B22. The results obtained indicate the variability of morphological, physiological, and biochemical properties in closely related Azospirillum strains and suggest the existence of metabolic relationships between methanotrophic bacteria and the representatives of the genus Azospirillum under peat bog conditions.     

Assessment of bacterial inoculant formulated with <Emphasis Type="Italic">Paraburkholderia tropica</Emphasis> to enhance wheat productivity

Paraburkholderia tropica is an endophytic nitrogen-fixing bacterium isolated from the rhizosphere, rhizoplane, and internal tissues of sugarcane and corn plants in different geographical regions. Other plant-growth-promoting abilities, such as phosphate solubilization and antifungal activity, have also been reported for this bacterium. With an aim at investigating the potential use of P. tropica as an inoculant for improving the performance of wheat crop, in this work we evaluated an experimental inoculant formulated with P. tropica MTo-293 with respect to root colonization, the practical aspects of its application, and the effects under field conditions when applied to wheat seeds. Bacterial colonization was monitored by culture dependent techniques and the wheat yield determined by quantifying the total grain production in two different seasons. Rhizoplane and endophytic colonization in wheat roots was achieved efficiently (on average, 8 and 4 log colony-forming units/g fresh weight, respectively) even at relatively low concentrations of viable bacteria in the inoculum under controlled conditions. P. tropica was compatible with a widely used fungicide, maintained viability for 48 h once applied to seeds, and was also able to colonize wheat roots efficiently. Furthermore, we were able to formulate an inoculant that maintained bacterial viability for relatively long time periods. Preliminary field assays were realized, and even though the average yields values for the inoculated treatments remained above the uninoculated ones, no significant effects of inoculation were detected with or without fertilization. The correct physiologic behavior of P. tropica suggests the necessity to continue with field experiments under different conditions.     

Evolution of <Emphasis Type="Italic">fixNOQP</Emphasis> genes encoding cytochrome oxidase with high affinity to oxygen in rhizobia and related bacteria

Many bacteria belonging to the order Rhizobiales have fixNOQP genes which encode cytochrome oxidase with high affinity to oxygen required for oxidative phosphorylation in microaerophilic conditions. There is one copy of the identified fixNOQP operon in ancestral forms of rhizobia (Bradyrhizobium), as well as in their putative evolutionary predecessors (bacteria related to Rhodopseudomonas). At the same time, forms deeply specialized in symbiosis (Rhizobium leguminosarum, Sinorhizobium meliloti) have multiple (2–3) copies, some of them have a high similarity (>90%) to fixNOQP genes of Bradyrhizobium and Rhodopseudomonas, and others have only 30–50% similarity. Two divergent copies fixNOQP are detected in Tardiphaga, which is a representative of the Bradyrhizobiaceae family, lacking the ability to fix N₂ (lack of nif genes encoding the synthesis of nitrogenase) and to induce the formation of nodules on legumes roots (lack of nod genes encoding the synthesis of signal Nod factors activating symbiosis development). The presence of Tardiphaga in nodule bacterial communities from a range of legumes, including Vavilovia formosa (relic representative of the tribe Fabeae, for which R. leguminosarum bv. viciae is the main microsymbiont), suggests that the ancestral gene duplication and subsequent divergence of fixNOQP operon in bacteria related to Tardiphaga opened the possibility of wide dissemination of functionally different copies of this cluster among symbiotically active forms of Rhizobiales. It is possible that the acquisition of fixNOQP genes determines adaptation of bacteria to microaerophilic niches not only in plants nodules but also in their environment (the rhizosphere, rhizoplane, internal portions of soil aggregates).     

<Emphasis Type="Italic">Lactobacillus</Emphasis> spp. impair the ability of <Emphasis Type="Italic">Listeria monocytogenes</Emphasis> FBUNT to adhere to and invade Caco-2 cells

Objectives

The objective of this study was to evaluate the ability of Lactobacillus curvatus CRL705, CRL1532, and CRL1533 and Lactobacillus sakei CRL1613 to survive under simulated gastrointestinal conditions. Moreover, a microencapsulation approach was proposed to improve gastrointestinal survival. Finally, experiments were performed to demonstrate that Lactobacillus spp. can modulate the ability of Listeria monocytogenes FBUNT to adhere to and invade Caco-2 cells.

Results

Lactobacillus strains were encapsulated in alginate beads to enhance the survival of bacteria under in vitro gastrointestinal conditions. All strains hydrolyzed bile salts using chenodeoxycholic acid as a substrate and adhered to Caco-2 cells. Cell-free supernatants (CFSs) showed antimicrobial activity against L. monocytogenes as demonstrated by agar diffusion assays. The average percentages of L. monocytogenes adhesion decreased from 67.74 to 41.75 and 38.7% in the presence of 50 and 90% (v/v), respectively, for all CFSs tested. The highest concentrations of CFSs completely inhibited the L. monocytogenes invasion of Caco-2 cells.

Conclusions

The studied Lactobacillus strains have protective effects against the adhesion and invasion of L. monocytogenes FBUNT. Alginate encapsulation of these bacteria improved gastrointestinal tolerance such that they could be further studied as potential probiotics against intestinal pathogenic bacteria.

     

Use of a gnotobiotic plant assay for assessing root colonization and mineral phosphate solubilization by <Emphasis Type="Italic">Paraburkholderia bryophila</Emphasis> Ha185 in association with perennial ryegrass (<Emphasis Type="Italic">Lolium perenne</Emphasis> L.)

Aims

The mechanisms by which rhizosphere bacteria increase the availability of mineral P precipitates for plant use are understudied. However, Paraburkholderia bryophila Ha185 is known to solubilize inorganic phosphate in vitro via a novel process. Therefore, this study aimed to demonstrate P solubilization by Ha185 in association with roots of perennial ryegrass (Lolium perenne L.).

Methods

We developed a gnotobiotic plant assay to assess P solubilization by Ha185 on ryegrass roots under various nutrient conditions. A green fluorescent protein (GFP)-tagged derivative of Ha185 was used in conjunction with fluorescent microscopy and confocal microscopy to visualize colonization of ryegrass roots.

Results

Ha185 solubilized mineral P (hydroxyapatite) in association with ryegrass roots and increased ryegrass growth by 20% under P-limited conditions. The GFP-tagged Ha185 strain colonized the rhizoplane and penetrated the primary root of ryegrass, possibly through “crack entry” at the point of lateral root emergence, but also by entering the epidermal cells via root hairs.

Conclusions

Ha185 supported ryegrass growth under P-limited conditions, indicating this strain may improve availability of soil P for uptake by ryegrass. Tools developed in this study have broad application in the study of rhizobacteria-plant interactions.

     

Illumina-based analysis of the rhizosphere microbial communities associated with healthy and wilted Lanzhou lily (<Emphasis Type="Italic">Lilium davidii</Emphasis> var. <Emphasis Type="Italic">unicolor</Emphasis>) plants grown in the field

Lanzhou lily (Liliumdavidii var. unicolor) is the best edible lily as well as a traditional medicinal plant in China. The microbes associated with plant roots play crucial roles in plant growth and health. However, little is known about the differences of rhizosphere microbes between healthy and wilted Lanzhou lily (Lilium davidii var. unicolor) plants. The objective of this study was to compare the rhizosphere microbial community and functional diversity of healthy and wilted plants, and to identify potential biocontrol agents with significant effect. Paired end Illumina Mi-Seq sequencing of 16S rRNA and ITS gene amplicons was employed to study the bacterial and fungal communities in the rhizosphere soil of Lanzhou lily plants. BIOLOG technology was adopted to investigate the microbial functional diversity. Our results indicated that there were major differences in the rhizosphere microbial composition and functional diversity of wilted samples compared with healthy samples. Healthy Lanzhou lily plants exhibited lower rhizosphere-associated bacterial diversity than diseased plants, whereas fungi exhibited the opposite trend. The dominant phyla in both the healthy and wilted samples were Proteobacteria and Ascomycota, i.e., 34.45 and 64.01 %, respectively. The microbial functional diversity was suppressed in wilted soil samples. Besides Fusarium, the higher relative abundances of Rhizoctonia, Verticillium, Penicillium, and Ilyonectria (Neonectria) in the wilted samples suggest they may pathogenetic root rot fungi. The high relative abundances of Bacillus in Firmicutes in healthy samples may have significant roles as biological control agents against soilborne pathogens. This is the first study to find evidence of major differences between the microbial communities in the rhizospheric soil of healthy and wilted Lanzhou lily, which may be linked to the health status of plants.     

Ectopic expression of cucumber (<Emphasis Type="Italic">Cucumis sativus</Emphasis> L.) <Emphasis Type="Italic">CsTIR</Emphasis>/<Emphasis Type="Italic">AFB</Emphasis> genes enhance salt tolerance in transgenic <Emphasis Type="Italic">Arabidopsis</Emphasis>

Auxin receptors TIR1/AFBs play an essential role in a series of signaling network cascades. These F-box proteins have also been identified to participate in different stress responses via the auxin signaling pathway in Arabidopsis. Cucumber (Cucumis sativus L.) is one of the most important crops worldwide, which is also a model plant for research. In the study herein, two cucumber homologous auxin receptor F-box genes CsTIR and CsAFB were cloned and studied for the first time. The deduced amino acid sequences showed a 78% identity between CsTIR and AtTIR1 and 76% between CsAFB and AtAFB2. All these proteins share similar characteristics of an F-box domain near the N-terminus, and several Leucine-rich repeat regions in the middle. Arabidopsis plants ectopically overexpressing CsTIR or CsAFB were obtained and verified. Shorter primary roots and more lateral roots were found in these transgenic lines with auxin signaling amplified. Results showed that expression of CsTIR/AFB genes in Arabidopsis could lead to higher seeds germination rates and plant survival rates than wild-type under salt stress. The enhanced salt tolerance in transgenic plants is probably caused by maintaining root growth and controlling water loss in seedlings, and by stabilizing life-sustaining substances as well as accumulating endogenous osmoregulation substances. We proposed that CsTIR/AFB-involved auxin signal regulation might trigger auxin mediated stress adaptation response and enhance the plant salt stress resistance by osmoregulation.     

Regulation of root development in <Emphasis Type="BoldItalic">Arabidopsis thaliana</Emphasis> by phytohormone-secreting epiphytic methylobacteria

In numerous experimental studies, seedlings of the model dicot Arabidopsis thaliana have been raised on sterile mineral salt agar. However, under natural conditions, no plant has ever grown in an environment without bacteria. Here, we document that germ-free (gnotobiotic) seedlings, raised on mineral salt agar without sucrose, develop very short root hairs. In the presence of a soil extract that contains naturally occurring microbes, root hair elongation is promoted; this effect can be mimicked by the addition of methylobacteria to germ-free seedlings. Using five different bacterial species (Methylobacterium mesophilicum, Methylobacterium extorquens, Methylobacterium oryzae, Methylobacterium podarium, and Methylobacterium radiotolerans), we show that, over 9 days of seedling development in a light-dark cycle, root development (hair elongation, length of the primary root, branching patterns) is regulated by these epiphytic microbes that occur in the rhizosphere of field-grown plants. In a sterile liquid culture test system, auxin (IAA) inhibited root growth with little effect on hair elongation and significantly stimulated hypocotyl enlargement. Cytokinins (trans-zeatin, kinetin) and ethylene (application of the precursor ACC) likewise exerted an inhibitory effect on root growth but, in contrast to IAA, drastically stimulated root hair elongation. Methylobacteria are phytosymbionts that produce/secrete cytokinins. We conclude that, under real-world conditions (soil), the provision of these phytohormones by methylobacteria (and other epiphytic microbes) regulates root development during seedling establishment.     

<Emphasis Type="Italic">Piriformospora indica</Emphasis>-mediated salinity tolerance in <Emphasis Type="Italic">Aloe vera</Emphasis> plantlets

Piriformospora indica, a root endophytic fungus, has been reported to promote growth of many plants under normal condition and allow the plants to survive under stress conditions. However, its impact on an important medicinal plant Aloe vera L. has not been well studied. Therefore, this study was undertaken to investigate the effect of P. indica on salinity stress tolerance of A. vera plant. P. indica inoculated and non-inoculated A. vera plantlets were subjected to four levels of salinity treatment- 0, 100, 200 and 300 mM NaCl. The salinity stress decreased the ability of the fungus to colonize roots of A. vera but the interaction of A. vera with P. indica resulted in an overall increase in plant biomass and greater shoot and root length as well as number of shoots and roots. The photosynthetic pigment (Chl a, Chl b and total Chl) and gel content were significantly higher for the fungus inoculated A. vera plantlets, at respective salinity concentrations. Furthermore, the inoculated plantlets had higher phenol, flavonoid, flavonol, aloin contents and radical scavenging activity at all salinity concentrations. The higher phenolic and flavonoid content may help the plants ameliorate oxidative stress resulting from high salinity.     

In vitro investigation of <Emphasis Type="Italic">Debaryomyces hansenii</Emphasis> strains for potential probiotic properties

In this study, 23 Debaryomyces hansenii strains, isolated from cheese and fish gut, were investigated in vitro for potential probiotic properties i.e. (1) survival under in vitro GI (gastrointestinal) conditions with different oxygen levels, (2) adhesion to Caco-2 intestinal epithelial cells and mucin, and (3) modulation of pro- and anti-inflammatory cytokine secretion by human monocyte-derived dendritic cells. As references two commercially available probiotic Saccharomyces cerevisiae var. boulardii (S. boulardii) strains were included in the study. Our results demonstrate that the different D. hansenii yeast strains had very diverse properties which could potentially lead to different probiotic effects. One strain of D. hansenii (DI 09) was capable of surviving GI stress conditions, although not to the same degree as the S. boulardii strains. This DI 09 strain, however, adhered more strongly to Caco-2 cells and mucin than the S. boulardii strains. Additionally, two D. hansenii strains (DI 10 and DI 15) elicited a higher IL-10/IL-12 ratio than the S. boulardii strains, indicating a higher anti-inflammatory effects on human dendritic cells. Finally, one strain of D. hansenii (DI 02) was evaluated as the best probiotic candidate because of its outstanding ability to survive the GI stresses, to adhere to Caco-2 cells and mucin and to induce a high IL-10/IL-12 ratio. In conclusion, this study shows that strains of D. hansenii may offer promising probiotic traits relevant for further study.     

Usefulness of the Non-conventional <Emphasis Type="Italic">Caenorhabditis elegans</Emphasis> Model to Assess <Emphasis Type="Italic">Candida</Emphasis> Virulence

Invasive candidiasis is caused mainly by Candida albicans, but other Candida species have increasing etiologies. These species show different virulence and susceptibility levels to antifungal drugs. The aims of this study were to evaluate the usefulness of the non-conventional model Caenorhabditis elegans to assess the in vivo virulence of seven different Candida species and to compare the virulence in vivo with the in vitro production of proteinases and phospholipases, hemolytic activity and biofilm development capacity. One culture collection strain of each of seven Candida species (C. albicans, Candida dubliniensis, Candida glabrata, Candida krusei, Candida metapsilosis, Candida orthopsilosis and Candida parapsilosis) was studied. A double mutant C. elegans AU37 strain (glp-4;sek-1) was infected with Candida by ingestion, and the analysis of nematode survival was performed in liquid medium every 24 h until 120 h. Candida establishes a persistent lethal infection in the C. elegans intestinal tract. C. albicans and C. krusei were the most pathogenic species, whereas C. dubliniensis infection showed the lowest mortality. C. albicans was the only species with phospholipase activity, was the greatest producer of aspartyl proteinase and had a higher hemolytic activity. C. albicans and C. krusei caused higher mortality than the rest of the Candida species studied in the C. elegans model of candidiasis.     

Efficacy of bacterial seed treatments for the control of <Emphasis Type="Italic">Fusarium verticillioides</Emphasis> in maize

Maize colonization by the phytopathogenic fungi Fusarium verticillioides leads to economical and food quality losses and also implicates potential health risks. In order to control this fungal species different strategies are being considered. In the present work we investigated the in situ effects of the use of two in vitro proven bacterial biological control agents against Fusarium verticillioides, using maize seedlings grown in greenhouse conditions. The treatment of maize seeds with Fusarium verticillioides 10⁷ spores ml^?1 negatively affected the length of the stem and the weight of the root systems of resultant seedlings, and also reduced the numbers of non-rhizospheric organisms with ammonification and nitrification activities. The addition of Bacillus amyloliquefaciens or Microbacterium oleovorans, at a concentration of 10⁹ colony forming units ml^?1, to the seeds significantly reduced Fusarium verticillioides count at the root inner tissues of resultant seedlings. When testing the impact of bacterial treatments on soil populations, no alterations with respect to control numbers of organisms with nitrification, ammonification or cellulolytic potentials were observed. Culturable bacterial richness and diversity calculated at the rhizoplane and root inner tissues of maize seedlings neither changed in bacterized treatments when compared to control treatment. Our study showed that the Fusarium verticillioides in vitro proven antagonists, Bacillus amyloliquefaciens and Microbacterium oleovorans (at 10⁹ colony forming units ml^?1), were also effective at greenhouse conditions without causing major changes in culturable rhizospheric and endophytic microbial richness and diversity.     

Allelopathic Potential of <Emphasis Type="Italic">Koelreuteria bipinnata</Emphasis> var. <Emphasis Type="Italic">integrifoliola</Emphasis> on Germination of Three Turf Grasses

Allelopathy is very important for the scientific disposition of garden plants. To understand the allelopathic potential of Koelreuteria bipinnata Franch. var. integrifoliola, the germination of Agrostis tenuis Sibth., Festuca arundinacea Schreb. and Lolium perenne L. were determined under laboratory conditions. The results showed that root, stem and leaf aqueous extracts of K. bipinnata var. integrifoliola had allelopathic effects on all three turf grasses, and the allelopathic activity varied according to extract concentrations, test species, and extract sources. Lower extract concentrations did not affect or promoted the germination and initial seedling growth of turf grasses, but the highest concentrations almost had inhibitory effect. The order of allelopathic potentials of the three organs on germination of these receptors was root < stem < leaf. And at the highest concentration of leaf extract, the most strongly inhibition was found in A. tenuis, followed by F. arundinaces and then L. perenne. In addition, according to gas chromatography–mass spectrometry (GC–MS) analysis, the allelopathic potential compounds and their abundance in root, stem and leaf were obviously different. Therefore, the allelopathic compounds may responsible for allelopathy of K. bipinnata var. integrifoliola. These findings suggested that more attention should be paid to the leaf of K. bipinnata var. integrifoliola for the relative higher allelopathic effects.     

Improved acid-stress tolerance of <Emphasis Type="Italic">Lactococcus lactis</Emphasis> NZ9000 and <Emphasis Type="Italic">Escherichia coli</Emphasis> BL21 by overexpression of the anti-acid component <Emphasis Type="Italic">recT</Emphasis>

Acid accumulation caused by carbon metabolism severely affects the fermentation performance of microbial cells. Here, different sources of the recT gene involved in homologous recombination were functionally overexpressed in Lactococcus lactis NZ9000 and Escherichia coli BL21, and their acid-stress tolerances were investigated. Our results showed that L. lactis NZ9000 (ERecT and LRecT) strains showed 1.4- and 10.4-fold higher survival rates against lactic acid (pH 4.0), respectively, and that E. coli BL21 (ERecT) showed 16.7- and 9.4-fold higher survival rates than the control strain against lactic acid (pH 3.8) for 40 and 60 min, respectively. Additionally, we found that recT overexpression in L. lactis NZ9000 improved their growth under acid-stress conditions, as well as increased salt- and ethanol-stress tolerance and intracellular ATP concentrations in L. lactis NZ9000. These findings demonstrated the efficacy of recT overexpression for enhancing acid-stress tolerance and provided a promising strategy for insertion of anti-acid components in different hosts.     

Interactions of the manganese hyperaccumulator <Emphasis Type="Italic">Phytolacca americana</Emphasis> L. with soil pH and phosphate

Hyperaccumulators are plants that store exceptionally high concentrations of heavy metals or metalloids in their leaves. Phytolacca americana is one of the few species known to hyperaccumulate manganese (Mn); however, it is a common weedy species and has no specific association with high-Mn soils. Neither the mechanism by which P. americana hyperaccumulates Mn nor the ecological significance of this trait are well understood. It has recently been suggested that P. americana secretes acids into the rhizosphere as a means of acquiring phosphate, which might coincidentally increase Mn uptake. To determine whether P. americana acidifies the surrounding soil, plants were grown in rhizoboxes providing access to living roots. A thin layer of agar containing bromocresol green pH indicator dye was placed on the roots to observe color changes indicating acidification. Comparative studies showed that P. americana acidifies the rhizosphere significantly more than the non-accumulating plant Acalypha rhomboidea. A second experiment studied whether adjustment of soil pH and phosphate affect foliar Mn concentrations of P. americana. Concentrations of Mn in leaves were highest when plants were grown in acidified soils but were significantly lower in soils that were alkaline and/or enriched with phosphate. These results suggest that Mn hyperaccumulation may be a side effect of rhizosphere acidification as a phosphorus-acquisition mechanism, rather than an adaptation in its own right. The findings provide fundamental information about hyperaccumulator physiology and evolution, and may be relevant to attempts to utilize P. americana for phytoremediation.     

Impact of soil salinity on the microbial structure of halophyte rhizosphere microbiome

The rhizosphere microbiome plays a significant role in the life of plants in promoting plant survival under adverse conditions. However, limited information is available about microbial diversity in saline environments. In the current study, we compared the composition of the rhizosphere microbiomes of the halophytes Urochloa, Kochia, Salsola, and Atriplex living in moderate and high salinity environments (Khewra salt mines; Pakistan) with that of the non-halophyte Triticum. Soil microbiomes analysis using pyrosequencing of 16S rRNA gene indicated that Actinobacteria were dominant in saline soil samples whereas Proteobacteria predominated in non-saline soil samples. Firmicutes, Acidobacteria, Bacteriodetes and Thaumarchaeota were predominant phyla in saline and non-saline soils, whereas Cyanobacteria, Verrucomicrobia, Gemmatimonadetes and the unclassified WPS-2 were less abundant. Sequences from Euryarchaeota, Ignavibacteriae, and Nanohaloarchaeota were identified only from the rhizosphere of halophytes. Dominant halophilic bacteria and archaea identified in this study included Agrococcus, Armatimonadetes gp4, Halalkalicoccus, Haloferula and Halobacterium. Our analysis showed that increases in soil salinity correlated with significant differences in the alpha and beta diversity of the microbial communities across saline and non-saline soil samples. Having a complete inventory of the soil bacteria from different saline environments in Pakistan will help in the discovery of potential inoculants for crops growing on salt-affected land.