Analysis of the Genome of the Heavy Metal Resistant and Hydrocarbon-Degrading Rhizospheric Pseudomonas qingdaonensis ZCR6 Strain and Assessment of Its Plant-Growth-Promoting Traits.

The Pseudomonas qingdaonensis ZCR6 strain, isolated from the rhizosphere of Zea mays growing in soil co-contaminated with hydrocarbons and heavy metals, was investigated for its plant growth promotion, hydrocarbon degradation, and heavy metal resistance. In vitro bioassays confirmed all of the abovementioned properties. ZCR6 was able to produce indole acetic acid (IAA), siderophores, and ammonia, solubilized Ca3(PO4)2, and showed surface active properties and activity of cellulase and very high activity of 1-aminocyclopropane-1-carboxylic acid deaminase (297 nmol α-ketobutyrate mg-1 h-1). The strain degraded petroleum hydrocarbons (76.52% of the initial hydrocarbon content was degraded) and was resistant to Cd, Zn, and Cu (minimal inhibitory concentrations reached 5, 15, and 10 mM metal, respectively). The genome of the ZCR6 strain consisted of 5,507,067 bp, and a total of 5055 genes were annotated, of which 4943 were protein-coding sequences. Annotation revealed the presence of genes associated with nitrogen fixation, phosphate solubilization, sulfur metabolism, siderophore biosynthesis and uptake, synthesis of IAA, ethylene modulation, heavy metal resistance, exopolysaccharide biosynthesis, and organic compound degradation. Complete characteristics of the ZCR6 strain showed its potential multiway properties for enhancing the phytoremediation of co-contaminated soils. To our knowledge, this is the first analysis of the biotechnological potential of the species P. qingdaonensis.

Increased Leaf Nicotine Content by Targeting Transcription Factor Gene Expression in Commercial Flue-Cured Tobacco (Nicotiana tabacum L.).

Nicotine, the most abundant pyridine alkaloid in cultivated tobacco (Nicotiana tabacum L.), is a potent inhibitor of insect and animal herbivory and a neurostimulator of human brain function. Nicotine biosynthesis is controlled developmentally and can be induced by abiotic and biotic stressors via a jasmonic acid (JA)-mediated signal transduction mechanism involving members of the APETALA 2/ethylene-responsive factor (AP2/ERF) and basic helix-loop-helix (bHLH) transcription factor (TF) families. AP2/ERF and bHLH TFs work combinatorically to control nicotine biosynthesis and its subsequent accumulation in tobacco leaves. Here, we demonstrate that overexpression of the tobacco NtERF32, NtERF221/ORC1, and NtMYC2a TFs leads to significant increases in nicotine accumulation in T2 transgenic K326 tobacco plants before topping. Up to 9-fold higher nicotine production was achieved in transgenics overexpressing NtERF221/ORC1 under the control of a constitutive GmUBI3 gene promoter compared to wild-type plants. The constitutive 2XCaMV35S promoter and a novel JA-inducible 4XGAG promoter were less effective in driving high-level nicotine formation. Methyljasmonic acid (MeJA) treatment further elevated nicotine production in all transgenic lines. Our results show that targeted manipulation of NtERF221/ORC1 is an effective strategy for elevating leaf nicotine levels in commercial tobacco for use in the preparation of reduced risk tobacco products for smoking replacement therapeutics.

Assessment of the Efficacy and Mode of Action of Benzo(1,2,3)-Thiadiazole-7-Carbothioic Acid S-Methyl Ester (BTH) and Its Derivatives in Plant Protection Against Viral Disease.

Systemic acquired resistance (SAR) induction is one of the primary defence mechanisms of plants against a broad range of pathogens. It can be induced by infectious agents or by synthetic molecules, such as benzo(1,2,3)-thiadiazole-7-carbothioic acid S-methyl ester (BTH). SAR induction is associated with increases in salicylic acid (SA) accumulation and expression of defence marker genes (e.g., phenylalanine ammonia-lyase (PAL), the pathogenesis-related (PR) protein family, and non-expressor of PR genes (NPR1)). Various types of pathogens and pests induce plant responses by activating signalling pathways associated with SA, jasmonic acid (JA) and ethylene (ET). This work presents an analysis of the influence of BTH and its derivatives as resistance inducers in healthy and virus-infected plants by determining the expression levels of selected resistance markers associated with the SA, JA, and ET pathways. The phytotoxic effects of these compounds and their influence on the course of viral infection were also studied. Based on the results obtained, the best-performing BTH derivatives and their optimal concentration for plant performance were selected, and their mode of action was suggested. It was shown that application of BTH and its derivatives induces increased expression of marker genes of both the SA- and JA-mediated pathways.

Chemical priming of immunity without costs to plant growth.

ß-Aminobutyric acid (BABA) induces broad-spectrum disease resistance, but also represses plant growth, which has limited its exploitation in crop protection. BABA perception relies on binding to the aspartyl-tRNA synthetase (AspRS) IBI1, which primes the enzyme for secondary defense activity. This study aimed to identify structural BABA analogues that induce resistance without stunting plant growth. Using site-directed mutagenesis, we demonstrate that the (l)-aspartic acid-binding domain of IBI1 is critical for BABA perception. Based on interaction models of this domain, we screened a small library of structural BABA analogues for growth repression and induced resistance against biotrophic Hyaloperonospora arabidopsidis (Hpa). A range of resistance-inducing compounds were identified, of which (R)-ß-homoserine (RBH) was the most effective. Surprisingly, RBH acted through different pathways than BABA. RBH-induced resistance (RBH-IR) against Hpa functioned independently of salicylic acid, partially relied on camalexin, and was associated with augmented cell wall defense. RBH-IR against necrotrophic Plectosphaerella cucumerina acted via priming of ethylene and jasmonic acid defenses. RBH-IR was also effective in tomato against Botrytis cinerea. Metabolic profiling revealed that RBH, unlike BABA, does not majorly affect plant metabolism. RBH primes distinct defense pathways against biotrophic and necrotrophic pathogens without stunting plant growth, signifying strong potential for exploitation in crop protection.

Investigation into the role of endogenous abscisic acid during ripening of imported avocado cv. Hass.

BACKGROUND: The importance of ethylene in avocado ripening has been extensively studied. In contrast, little is known about the possible role of abscisic acid (ABA). The present work studied the effect of 1-methylcyclopropene (1-MCP) (0.3 µL L-1 ), e+® Ethylene Remover and the combination thereof on the quality of imported avocado cv. Hass fruit stored for 7 days at 12 °C. Ethylene production, respiration, firmness, colour, heptose (C7) sugars and ABA concentrations in mesocarp tissue were measured throughout storage. RESULTS: Treatment with e+® Ethylene Remover reduced ethylene production, respiration rate and physiological ripening compared with controls. Fruit treated with 1-MCP + e+® Ethylene Remover and, to a lesser extent 1-MCP alone, had the lowest ethylene production and respiration rate and hence the best quality. Major sugars measured in mesocarp tissue were mannoheptulose and perseitol, and their content was not correlated with ripening parameters. Mesocarp ABA concentration, as determined by mass spectrometry, increased as fruit ripened and was negatively correlated with fruit firmness. CONCLUSIONS: Results suggest a relationship between ABA and ethylene metabolism since blocking ethylene, and to a larger extent blocking and removing ethylene, resulted in lower ABA concentrations. Whether ABA influences avocado fruit ripening needs to be determined in future research. © 2017 Society of Chemical Industry.

A Low Cost Compact Measurement System Constructed Using a Smart Electrochemical Sensor for the Real-Time Discrimination of Fruit Ripening.

Ethylene as an indicator for evaluating fruit ripening can be measured by very sensitive electrochemical gas sensors based on a high-resolution current produced by a bias potential applied to the electrodes. For this purpose, a measurement system for monitoring ethylene gas concentrations to evaluate fruit ripening by using the electrochemical ethylene sensor was successfully developed. Before the electrochemical ethylene sensor was used to measure the ethylene gas concentrations released from fruits, a calibration curve was established by the standard ethylene gases at concentrations of 2.99 ppm, 4.99 ppm, 8.01 ppm and 10 ppm, respectively, with a flow rate of 0.4 L·min(-1). From the calibration curve, the linear relationship between the responses and concentrations of ethylene gas was obtained in the range of 0-10 ppm with the correlation coefficient R² of 0.9976. The micropump and a novel signal conditioning circuit were implemented in this measurement, resulting in a rapid response in detecting ethylene concentrations down to 0.1 ppm in air and in under 50 s. In this experiment, three kinds of fruits-apples, pears and kiwifruits-were studied at a low concentration (under 0.8 ppm) of trace ethylene content in the air exhaled by fruits. The experimental results showed that a low cost, compact measurement system constructed by using an electrochemical ethylene sensor has a high sensitivity of 0.3907 V·ppm(-1) with a theoretical detection limit of 0.413 ppm, and is non-invasive and highly portable.

Modelling microbial dechlorination of trichloroethene: investigating the trade-off between quality of fit and parameter reliability.

This work puts forth a heuristic approach for investigating compromises between quality of fit and parameter reliability for the Monod-type kinetics employed to model microbial reductive dechlorination of trichloroethene. The methodology is demonstrated with three models of increasing fidelity and complexity. Model parameters were estimated with a stochastic global optimization algorithm, using scarce and inherently noisy experimental data from a mixed anaerobic microbial culture, which dechlorinated trichloroethene to ethene completely. Parameter reliability of each model was assessed using a Monte Carlo technique. Finally, an alternate quantity of applied interest was evaluated in order to assist with model discrimination. Results from the application of our approach suggest that the modeler should examine the implementation of conceptually simple models, even if they are a crude abstraction of reality, as they can be computationally less demanding and adequately accurate when model performance is assessed with criteria of applied interest, such as chloroethene elimination time.

Assessment and modification of degenerate qPCR primers that amplify functional genes from etheneotrophs and vinyl chloride-assimilators.

AIMS: Degenerate qPCR primer sets that target the functional genes etnC and etnE in etheneotrophs and vinyl chloride-assimilating bacteria were assessed and modified in an effort to improve performance. METHODS AND RESULTS: Functional gene abundance in four pure cultures was estimated by qPCR using novel (MRTC and MRTE) and existing (RTC and RTE) degenerate primer sets and compared to abundances estimated with nondegenerate gene-specific primers (GSPs). Functional gene abundance in groundwater DNA extracted from several contaminated sites was also estimated with MRTC and MRTE primers. CONCLUSIONS: MRTC primers displayed significantly improved etnC quantification in both pure cultures and environmental samples. SIGNIFICANCE AND IMPACT OF THE STUDY: Application of MRTC and MRTE primer sets will enhance microbial ecology studies involving etheneotrophs and qPCR analyses that support vinyl chloride bioremediation strategies.

Assessment of ethylene removal with Pseudomonas strains.

This study investigated the biological removal of ethylene by Pseudomonas strains in a batch test and a biofilter column. In the batch test, no removal of ethylene was found in the absence of inoculated system, whereas more than 50% of the ethylene in the presence of inoculated system was degraded within 17 h, and completely removed after 25 h. The biofilter, packed with activated carbons, was capable of achieving ethylene removal efficiency as much as 100% at a residence time of 14 min and an inlet concentration of 331 mg m-3. Under the same conditions, carbon dioxide with a concentration of up to 1097 mg m-3 was produced. It was found that carbon dioxide was produced at a rate of 87 mg day-1, which corresponded to a volume of 0.05 L day-1. During operation with an inlet ethylene of 331 mg m-3, the maximum elimination capacity of the biofilter was 34 g C2H4 m-3 day-1. This biological system could reduce the ethylene concentration to levels below the threshold limit for the plant hormonal response (0.01 mg m-3), and provide an attractive treatment technology in horticultural storage facilities.

Assessment of inequality of root hair density in Arabidopsis thaliana using the Gini coefficient: a close look at the effect of phosphorus and its interaction with ethylene.

BACKGROUND AND AIMS: Root hair density (i.e. the number of root hairs per unit root length) in Arabidopsis thaliana varies among individual plants in response to different nutrient stresses. The degree of such variation, defined as inequality, serves as a unique indicator of the uniformity of response within a plant population to nutrient availability. METHODS: Using the Gini coefficient (G) as an inequality index, the inequality of root hair density in Arabidopsis thaliana 'Columbia' was evaluated under conditions of nutrient stresses; in particular the effect of phosphorus and its interaction with ethylene. KEY RESULTS: With decreasing phosphorus concentration, root hair density increased while inequality decreased logarithmically. The addition of the ethylene precursor 1-aminocyclopropane-1-carboxylate (ACC) under high phosphorus increased root hair density and decreased inequality by 7-fold. Inhibition of ethylene action with 1-methylcyclopropene (MCP) and silver thiosulphate (STS) under low phosphorus decreased root hair density, and increased inequality by 9-fold and 4-fold, respectively. The ethylene action inhibitors had little effect on root hair density under high phosphorus, but inequality increased 3-fold in the presence of MCP and decreased 2-fold in the presence of STS. Compared with the control, deficiencies in S, N and K increased inequality of root hair density, whereas deficiencies in P, Ca, B, Mn, Fe, Zn, Cu and Mg decreased inequality. In particular, the inequality of root hair density increased by over 2-fold under deficiencies of N or K, but decreased 14-fold under phosphorus deficiency. CONCLUSIONS: The inequality analysis indicates a strong correlation between prevalent signals from the environment (i.e. phosphorus stress) and the response of the plant, and the role of ethylene in this response. As the environmental signals become stronger, an increasing proportion of individuals respond, resulting in a decrease in variation in responsiveness among individual plants as indicated by reduced inequality.

Use of covalent binding in risk assessment.

Risk characterization comprises hazard identification describing the intrinsic toxic potential of a chemical, toxicokinetics, as well as the toxic mechanisms, information about dose response and exposure assessment. Compounds that induce reversible effects, which are repaired during and after exposure, are considered thresholded and allow definition of a NOEL. If damage is not repaired, the effect persists and accumulates upon repeated exposure. In such cases a NOEL cannot be determined. Biological reactive intermediates of chemicals have the potential to bind covalently to cellular macromolecules like proteins and DNA. Such interaction is not repaired completely and may persist. Thus, data on covalent binding (CB) are of qualitative and quantitative significance in the risk assessment process. Qualitatively, CB, especially with DNA and in correlation with this to proteins, is indicative for an irreversible and non-thresholded mutagenic and carcinogenic effect. Absence or presence of CB assists to differentiate between primarily genotoxic and thresholded non-genotoxic carcinogens. Quantitatively, CB is used to understand internal exposure and target dose, which is a prerequisite for species-species extrapolation, and to justify extrapolation from high dose to low dose. The reactive intermediates of ethylene, propylene and styrene have been determined in rodents and humans and modeled to predict dose responses of internal exposure. It is described in this communication that such information, together with other parameters like cell proliferation as a result of cytotoxicity, is the basis for quantitative risk assessment of human exposure to these compounds.

A thermodynamic assessment of possible substrates for sulphate-reducing bacteria.

A thermodynamic feasibility study was applied as a means of predicting suitable energy-yielding substrates for growth of sulphate-reducing microorganisms. The average free energy release per electron pair for a substrate-sulphate oxidoreduction may be more or less than the energy requirement for ATP synthesis from ADP and Pi. Substrates were divided into two groups on this thermodynamic basis and the division was shown to accord with previous experimental reports; those substrates which released an average of at least 8-4 kcal per electron pair (35-2 kJ per electron pair) were able to support growth whilst those releasing less than 8-4 kcal were unable to do so. It is proposed that the thermodynamic assessment could be applied to a wide range of possible substrates to predict the likelihood of their serving as sole substrates for growth of these organisms. The literature concerning the use of hydrocarbons by sulphate reducers is confused and indefinite, but inclines toward the idea that use of long-chain hydrocarbons by these organisms is possible. In contrast, however, thermodynamic analysis showed that the highest energy release is from the short-chain alkynes.