Biological hydrogen cyanide emission globally impacts the physiology of both HCN-emitting and HCN-perceiving Pseudomonas.

IMPORTANCE: Bacteria communicate by exchanging chemical signals, some of which are volatile and can remotely reach other organisms. HCN was one of the first volatiles discovered to severely impact exposed organisms by inhibiting their respiration. Using HCN-deficient mutants in two Pseudomonas strains, we demonstrate that HCN's impact goes beyond the sole inhibition of respiration and affects both emitting and receiving bacteria in a global way, modulating their motility, biofilm formation, and production of antimicrobial compounds. Our data suggest that bacteria could use HCN not only to control their own cellular functions, but also to remotely influence the behavior of other bacteria sharing the same environment. Since HCN emission occurs in both clinically and environmentally relevant Pseudomonas, these findings are important to better understand or even modulate the expression of bacterial traits involved in both virulence of opportunistic pathogens and in biocontrol efficacy of plant-beneficial strains.

Downregulation of lncRNA PpL-T31511 and Pp-miRn182 Promotes Hydrogen Cyanamide-Induced Endodormancy Release through the PP2C-H2O2 Pathway in Pear (Pyrus pyrifolia).

Bud endodormancy is an important, complex process subject to both genetic and epigenetic control, the mechanism of which is still unclear. The endogenous hormone abscisic acid (ABA) and its signaling pathway play important roles in the endodormancy process, in which the type 2C protein phosphatases (PP2Cs) is key to the ABA signal pathway. Due to its excellent effect on endodormancy release, hydrogen cyanamide (HC) treatment is considered an effective measure to study the mechanism of endodormancy release. In this study, RNA-Seq analysis was conducted on endodormant floral buds of pear (Pyrus pyrifolia) with HC treatment, and the HC-induced PP2C gene PpPP2C1 was identified. Next, software prediction, expression tests and transient assays revealed that lncRNA PpL-T31511-derived Pp-miRn182 targets PpPP2C1. The expression analysis showed that HC treatment upregulated the expression of PpPP2C1 and downregulated the expression of PpL-T31511 and Pp-miRn182. Moreover, HC treatment inhibited the accumulation of ABA signaling pathway-related genes and hydrogen peroxide (H2O2). Furthermore, overexpression of Pp-miRn182 reduced the inhibitory effect of PpPP2C1 on the H2O2 content. In summary, our study suggests that downregulation of PpL-T31511-derived Pp-miRn182 promotes HC-induced endodormancy release in pear plants through the PP2C-H2O2 pathway.

Origami paper analytical assay based on metal complex sensor for rapid determination of blood cyanide concentration in fire survivors.

Cyanide-based blood poisoning can seriously damage fire victims and cause death if not detected quickly. Previous conventional methods require laboratory equipment, which are expensive and increase the duration of the analysis. Here, a simple origami based microfluidic device was introduced for point of need detection of blood cyanide concentration in people involved in fire. The device is made of four layers of paper. Each layer was in the size of 1 × 1 cm folded on each other. In this work, the blood sample was acidified by trichloroacetic acid to separate cyanide from methaemoglobin in the form of HCN gas. The produced gas released into borate buffer to recover free cyanide ions which interacted with the Pt complex ([Pt(p-MeC6H4)2(phen)]) used as a receptor in this study. Optimized conditions were applied to have a suitable interaction causing the color of the receptor to change from yellow to colorless. The color changes were recorded by a smartphone, and the sensor response was calculated by the routine image analysis software. The assay was capable of determining cyanide ions at different concentrations in the range of 1.0 to 100.0 µmol L-1. The detection limit of these determination was equal to 0.4 µmol L-1. The assay responses were not affected by the interfering species. As a practical analysis, the proposed sensor was applied to determine cyanide ions in the blood sample of 20 studied fire survivors and 10 controls with high accuracy.

Effect of silver nanoparticles and Bacillus cereus LPR2 on the growth of Zea mays.

The effect of Plant Growth Promoting Rhizobacteria (Bacillus sp.) and silver nanoparticles on Zea mays was evaluated. The silver nanoparticles were synthesized from Tagetes erecta (Marigold) leaf and flower extracts, whereas PGPR isolated from spinach rhizosphere. The silver nanoparticles (AgNPs) were purified using ultra centrifugation and were characterized using UV-Vis spectroscopy at gradient wavelength and also by High Resolution Transmission Electron microscopy (HRTEM). The average particles size of AgNPs was recorded approximately 60 nm. Almost all potential isolates were able to produce Indole Acetic Acid (IAA), ammonia and Hydrogen cyanide (HCN), solubilized tricalcium phosphate and inhibited the growth of Macrophomina phaseolina in vitro but the isolate LPR2 was found the best among all. On the basis of 16S rRNA gene sequence, the isolate LPR2 was characterized as Bacillus cereus LPR2. The maize seeds bacterized with LPR2 and AgNPs individually showed a significant increase in germination (87.5%) followed by LPR2 + AgNPs (75%). But the maximum growth of root and shoot of maize plant was observed in seeds coated with LPR2 followed by AgNPs and a combination of both. Bacillus cereus LPR2 and silver nanoparticles enhanced the plant growth and LPR2 strongly inhibited the growth of deleterious fungal pathogen. Therefore, LPR2 and AgNPs could be utilized as bioinoculant and growth stimulator, respectively for maize.

Exploring the mechanism of olfactory recognition in the initial stage by modeling the emission spectrum of electron transfer.

Olfactory sense remains elusive regarding the primary reception mechanism. Some studies suggest that olfaction is a spectral sense, the olfactory event is triggered by electron transfer (ET) across the odorants at the active sites of odorant receptors (ORs). Herein we present a Donor-Bridge-Acceptor model, proposing that the ET process can be viewed as an electron hopping from the donor molecule to the odorant molecule (Bridge), then hopping off to the acceptor molecule, making the electronic state of the odorant molecule change along with vibrations (vibronic transition). The odorant specific parameter, Huang-Rhys factor can be derived from ab initio calculations, which make the simulation of ET spectra achievable. In this study, we revealed that the emission spectra (after Gaussian convolution) can be acted as odor characteristic spectra. Using the emission spectrum of ET, we were able to reasonably interpret the similar bitter-almond odors among hydrogen cyanide, benzaldehyde and nitrobenzene. In terms of isotope effects, we succeeded in explaining why subjects can easily distinguish cyclopentadecanone from its fully deuterated analogue cyclopentadecanone-d28 but not distinguishing acetophenone from acetophenone-d8.

Functional characterization of potential PGPR exhibiting broad-spectrum antifungal activity.

This study describes the biocontrol potential of rhizobacteria against a range of fungal phytopathogens. Out of 227 bacteria isolated from the rhizosphere of maize, rice, wheat, potato, sunflower and soybean crops cultivated in different agro-ecological regions of Pakistan, 48 exhibited >60 % antifungal activity against Fusarium oxysporum, Fusarium moniliforme, Rhizoctonia solani, Colletotrichum gloeosporioides, Colletotrichum falcatum, Aspergillus niger, and Aspergillus flavus. The rhizobacteria inhibiting >65 % pathogen growth were selected for detailed molecular and in planta studies most of which were identified as Pseudomonas and Bacillus species based on 16S rRNA gene sequence analysis. Antifungal metabolites produced by these rhizobacteria analyzed through LCMS were identified as antibiotics (iturin, surfactins, fengycin, DAPG, Phenazine, etc.), cell wall degrading enzymes (protease, chitinase, and cellulase), plant growth promotion enzymes and hormones (indole-3-acetic acid, ACC-deaminase, phosphates, nitrogen fixation), N-acyl-homoserine lactones and siderophores. The growth room experiment validated the potential of these bacteria as biofertilizer and biopesticide agents. Of all, P. aeruginosa strain FB2 and B. subtilis strain RMB5 showed significantly higher potential as antagonistic plant-beneficial bacteria effective against a range of fungal phytopathogens. Both these bacteria can be used to develop a dual-purpose bacterial inoculum as biopesticide and biofertilizer. Rest of the antagonistic PGPR may be exploited for disease control in less-infested soils.

Redox metabolism and cell wall modifications as global and local targets respectively, of cyanide induced dormancy release of walnut kernels.

The HCN-induced seed dormancy release necessitates alterations in reactive oxygen species (ROS) metabolism and radicle cell wall loosening. Little is known about the interaction of ROS metabolism with cell wall hydrolytic enzymes during HCN-induced seed dormancy release. Thus dormant walnut (Juglans regia L.) kernels were exposed to HCN (4 h) and studied for redox metabolism and cell wall-modifying enzymes during 10 days of incubation (DI) i.e. before radicle emergence. HCN increased ROS especially in the embryonic axes (EAs) but decreased ROS-generating NADPH oxidase and ROS scavenging superoxide dismutase (SOD) and peroxidase (POX) with no effects on catalase (CAT), ascorbate peroxidase (APX) and cell wall-modifying enzymes activities in short term up to 2 DI. In long term roughly from 4 DI onwards, HCN-exposed EA displayed greater superoxide anions and enhanced activities of POX, APX, NADPH oxidase, cell wall peroxidase (CW-POX), ß- 1, 4-D glucanase, mannanase, polygacturonase and xylanase. Meanwhile HCN increased greater expression of POX and mannanase isoforms as revealed by in-gel activity assay. Except for higher activities of CAT, POX and APX, cotyledonary activities of CW-POX, mannanase and polygacturonase and to some extent ß- 1, 4-D glucanase remained unaffected by HCN. Thus short term ROS accumulation in HCN-treated EA is due to declined SOD and POX activities. In long term the enhanced activities of both NADPH oxidase: CW-POX couple and cell wall-modifying enzymes in EA bring about wall loosening in preparation for radicle emergence. Evidences for the simultaneous operation of both mechanisms are provided in walnut EAs during dormancy release.

Comparative transcriptome analysis reveals significant differences in the regulation of gene expression between hydrogen cyanide- and ethylene-treated Arabidopsis thaliana.

BACKGROUND: Hydrogen cyanide (HCN) is a small gaseous molecule that is predominantly produced as an equimolar co-product of ethylene (ET) biosynthesis in plants. The function of ET is of great concern and is well studied; however, the function of HCN is largely unknown. Similar to ET, HCN is a simple and diffusible molecule that has been shown to play a regulatory role in the control of some metabolic processes in plants. Nevertheless, it is still controversial whether HCN should be regarded as a signalling molecule, and the cross-talk between HCN and ET in gene expression regulation remains unclear. In this study, RNA sequencing (RNA-seq) was performed to compare the differentially expressed genes (DEGs) between HCN and ET in Arabidopsis. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses were subsequently performed to investigate the function and pathway enrichment of DEGs. Parts of key genes were confirmed by quantitative real-time PCR. RESULTS: The results showed that at least 1305 genes and 918 genes were significantly induced by HCN and ET, respectively. Interestingly, a total of 474 genes (|log2 FC| ≥1) were co-regulated by HCN and ET. GO and KEGG analyses indicated that the co-regulated genes by HCN and ET were enriched in plant responses to stress and plant hormone signal transduction pathways, indicating that HCN may cooperate with ET and participate in plant growth and development and stress responses. However, a total of 831 genes were significantly induced by HCN but not by ET, indicating that in addition to ET, HCN is in essence a key signalling molecule in plants. Importantly, our data showed that the possible regulatory role of a relatively low concentration of HCN does not depend on ET feedback induction, although there are some common downstream components were observed. CONCLUSION: Our findings provide a valuable resource for further exploration and understanding of the molecular regulatory mechanisms of HCN in plants and provide novel insight into HCN cross-talk with ET and other hormones in the regulation of plant growth and plant responses to environmental stresses.

Hydrogen cyanide produced by Pseudomonas chlororaphis O6 is a key aphicidal metabolite.

A biocontrol bacterium, Pseudomonas chlororaphis O6 promotes plant health through multifaceted mechanisms. In this study, we used P. chlororaphis O6 mutants to examine metabolites with aphicidal activity. Direct application of intact P. chlororaphis cells to the surface of second-instar nymphs of the green peach aphid resulted in no mortality. However, nymphs died when exposed only to the volatiles produced by the P. chlororaphis O6 wild-type strain grown on rich media. Mutants lacking the production of two antibiotics, phenazine and pyrrolnitrin, or the insect toxin FitD retained the aphicidal potential of the wild-type strain. However, the volatiles produced by mutants deficient in the production of hydrogen cyanide (HCN) or defective in the synthesis of the global regulator GacS, which regulates HCN synthesis, showed no aphicidal activity. Direct application of potassium cyanide caused mortality of green peach aphid nymphs. These results indicate that HCN production by a plant probiotic is involved in preventing insect growth.

Hydrogen cyanide produced by the soil bacterium Chromobacterium sp. Panama contributes to mortality in Anopheles gambiae mosquito larvae.

Mosquito larvae continuously encounter microbes in their aquatic environment, which serve as food and play a critical role in successful development. In previous work, we isolated a Chromobacterium sp. (C.sp_P) with larvicidal activity from the midgut of dengue vector Aedes mosquitoes in Panama. In this study, we found a positive correlation between initial concentrations of C.sp_P and larval mortality rates, and that C.sp_P is more efficient at inducing larval mortality in a high nutrient environment. Multiple Chromobacterium species induce larval mortality with similar efficacy to C.sp_P except for C. subtsugae. We also found that a non-lethal dose of C.sp_P lengthens development time and increases mortality over multiple developmental stages, suggesting persistent effects of exposure. Additionally, we showed that larvicidal activity persists in the larval breeding water after removal of live bacteria, and that the larvicidal factor in C.sp_P-treated water is smaller than 3 kDa, heat resistant to 90 °C, and lost after vacuum centrifugation. We showed that C.sp_P produces hydrogen cyanide in culture and in larval water at concentrations sufficient to kill An. gambiae larvae, and treatment of the larval water with a cyanide antidote eliminated larvicidal activity. We conclude that a potential mechanism by which C.sp_P can induce larval mortality is via production of hydrogen cyanide.

Nematicidal effect of rhizobacteria on plant-parasitic nematodes associated with vineyards.

The action of metabolites and exoenzymes from rhizobacteria on different plant-parasitic nematodes has an influence on the nematicidal efficacy of the microbe. Seven rhizobacteria, divided into two bacterial groups, were evaluated in vitro for nematicidal activity on Meloidogyne ethiopica and Xiphinema index. The direct effect of their filtrates on egg hatching and juveniles of M. ethiopica as well as mobile stages of X. index was evaluated during a 72-h period. The production of four exoenzymes and two metabolites associated with nematode mortality was investigated. Molecular characterization of three isolates was performed, and the physiological profiles and lipase activity of all isolates were obtained using the BIOLOG EcoPlate system. While chitinase and collagenase were measured using the BIOLOG MT2 plate system, protease, hydrogen cyanide and hydrogen sulphide were directly determined in Petri dishes. Nematode mobile stages exposure to the bacterial filtrate revealed a nematicidal effect up to 93.7% on X. Index and up to 83.3% on M. ethiopica. The control of egg hatching varied between 35 and 85%. A positive correlation was found between the mortality of both nematode mobile stages and the concerted activities of the bacterial enzymes as well as the level of the volatile metabolites. The nematicidal effect of rhizobacteria strains varies by nematode genera and among the developmental stages evaluated.

Hydrogen cyanamide breaks grapevine bud dormancy in the summer through transient activation of gene expression and accumulation of reactive oxygen and nitrogen species.

BACKGROUND: Hydrogen cyanamide (HC) and pruning (P) have frequently been used to break dormancy in grapevine floral buds. However, the exact underlying mechanism remains elusive. This study aimed to address the early mode of action of these treatments on accumulation of reactive oxygen species (ROS) and reactive nitrogen species (RNS) and expression of related genes in the dormancy breaking buds of grapevine in the summer. RESULTS: The budbreak rates induced by pruning (P), hydrogen cyanamide (HC), pruning plus hydrogen cyanamide (PHC) and water (control) after 8 days were 33, 53, 95, and 0 %, respectively. Clearly, HC was more effective in stimulating grapevine budbreak and P further enhanced its potency. In situ staining of longitudinal bud sections after 12 h of treatments detected high levels of ROS and nitric oxide (NO) accumulated in the buds treated with PHC, compared with HC or P alone. The amounts of ROS and NO accumulated were highly correlated with the rates of budbreak among these treatments, highlighting the importance of a rapid, transient accumulation of sublethal levels of ROS and RNS in dormancy breaking. Microarray analysis revealed specific alterations in gene expression in dormancy breaking buds induced by P, HC and PHC after 24 h of treatment. Relative to control, PHC altered the expression of the largest number of genes, while P affected the expression of the least number of genes. PHC also exerted a greater intensity in transcriptional activation of these genes. Gene ontology (GO) analysis suggests that alteration in expression of ROS related genes is the major factor responsible for budbreak. qRT-PCR analysis revealed the transient expression dynamics of 12 specific genes related to ROS generation and scavenge during the 48 h treatment with PHC. CONCLUSION: Our results suggest that rapid accumulation of ROS and NO at early stage is important for dormancy release in grapevine in the summer, and the identification of the commonly expressed specific genes among the treatments allowed the construction of the signal transduction pathway related to ROS/RNS metabolism during dormancy release. The rapid accumulation of a sublethal level of ROS/RNS subsequently induces cell wall loosening and expansion for bud sprouting.

Application of the Prunus spp. Cyanide Seed Defense System onto Wheat: Reduced Insect Feeding and Field Growth Tests.

Many crops are ill-protected against insect pests during storage. To protect cereal grains from herbivores during storage, pesticides are often applied. While pesticides have an undoubtable functionality, increasing concerns are arising about their application. In the present study, we investigated a bioinspired cyanogenic grain coating with amygdalin as cyanogenic precursor mimicking the feeding-triggered release of hydrogen cyanide (HCN) found for example in bitter almonds. The multilayer coating consisted of biodegradable polylactic acid with individual layers containing amygdalin or ß-glucosidase which is capable of degrading amygdalin to HCN. This reaction occurred only when the layers were ruptured, e.g., by a herbivore attack. Upon feeding coated cyanogenic wheat grains to Tenebrio molitor (mealworm beetle), Rhizopertha dominica (lesser grain borer), and Plodia interpunctella (Indianmeal moth), their reproduction as well as consumption rate were significantly reduced, whereas germination ability increased compared to noncoated grains. In field experiments, we observed an initial growth delay compared to uncoated grains which became negligible at later growth stages. The here shown strategy to artificially apply a naturally occurring defense mechanisms could be expanded to other crops than wheat and has the potential to replace certain pesticides with the benefit of complete biodegradability and increased safety during storage.

Pyrrolnitrin and Hydrogen Cyanide Production by Pseudomonas chlororaphis Strain PA23 Exhibits Nematicidal and Repellent Activity against Caenorhabditis elegans.

Pseudomonas chlororaphis strain PA23 is a biocontrol agent able to suppress growth of the fungal pathogen Sclerotinia sclerotiorum. This bacterium produces an arsenal of exometabolites including pyrrolnitrin (PRN), phenazine (PHZ), hydrogen cyanide (HCN), and degradative enzymes. Production of these compounds is controlled at both the transcriptional and posttranscriptional levels by the Gac-Rsm system, RpoS, PsrA, and the Phz quorum-sensing system. Beyond pathogen-suppression, the success of a biocontrol agent is dependent upon its ability to establish itself in the environment where predation by bacterivorous organisms, including nematodes, may threaten persistence. The focus of this study was to investigate whether PA23 is able to resist grazing by Caenorhabditis elegans and to define the role played by exoproducts in the bacterial-nematode interaction. We discovered that both PRN and HCN contribute to fast- and slow-killing of C. elegans. HCN is well-established as having lethal effects on C. elegans; however, PRN has not been reported to be nematicidal. Exposure of L4 stage nematodes to purified PRN reduced nematode viability in a dose-dependent fashion and led to reduced hatching of eggs laid by gravid adults. Because bacterial metabolites can act as chemoattractants or repellents, we analyzed whether PA23 exhibited attractant or repulsive properties towards C. elegans. Both PRN and HCN were found to be potent repellents. Next we investigated whether the presence of C. elegans would elicit changes in PA23 gene activity. Co-culturing the two organisms increased expression of a number of genes associated with biocontrol, including phzA, hcnA, phzR, phzI, rpoS and gacS. Exoproduct analysis showed that PHZ and autoinducer signals were upregulated, consistent with the gene expression profiles. Collectively, these findings indicate that PA23 is able to sense the presence of C. elegans and it is able to both repel and kill the nematodes, which should facilitate environmental persistence and ultimately biocontrol.

Dormancy alleviation by NO or HCN leading to decline of protein carbonylation levels in apple (Malus domestica Borkh.) embryos.

Deep dormancy of apple (Malus domestica Borkh.) embryos can be overcome by short-term pre-treatment with nitric oxide (NO) or hydrogen cyanide (HCN). Dormancy alleviation of embryos modulated by NO or HCN and the first step of germination depend on temporary increased production of reactive oxygen species (ROS). Direct oxidative attack on some amino acid residues or secondary reactions via reactive carbohydrates and lipids can lead to the formation of protein carbonyl derivatives. Protein carbonylation is a widely accepted covalent and irreversible modification resulting in inhibition or alteration of enzyme/protein activities. It also increases the susceptibility of proteins to proteolytic degradation. The aim of this work was to investigate protein carbonylation in germinating apple embryos, the dormancy of which was removed by pre-treatment with NO or HCN donors. It was performed using a quantitative spectrophotometric method, while patterns of carbonylated protein in embryo axes were analyzed by immunochemical techniques. The highest concentration of protein carbonyl groups was observed in dormant embryos. It declined in germinating embryos pre-treated with NO or HCN, suggesting elevated degradation of modified proteins during seedling formation. A decrease in the concentration of carbonylated proteins was accompanied by modification in proteolytic activity in germinating apple embryos. A strict correlation between the level of protein carbonyl groups and cotyledon growth and greening was detected. Moreover, direct in vitro carbonylation of BSA treated with NO or HCN donors was analyzed, showing action of both signaling molecules as protein oxidation agents.

The response of Caenorhabditis elegans to hydrogen sulfide and hydrogen cyanide.

Hydrogen sulfide (H2S), an endogenously produced small molecule, protects animals from various stresses. Recent studies demonstrate that animals exposed to H2S are long lived, resistant to hypoxia, and resistant to ischemia-reperfusion injury. We performed a forward genetic screen to gain insights into the molecular mechanisms Caenorhabditis elegans uses to appropriately respond to H2S. At least two distinct pathways appear to be important for this response, including the H2S-oxidation pathway and the hydrogen cyanide (HCN)-assimilation pathway. The H2S-oxidation pathway requires two distinct enzymes important for the oxidation of H2S: the sulfide:quinone reductase sqrd-1 and the dioxygenase ethe-1. The HCN-assimilation pathway requires the cysteine synthase homologs cysl-1 and cysl-2. A low dose of either H2S or HCN can activate hypoxia-inducible factor 1 (HIF-1), which is required for C. elegans to respond to either gas. sqrd-1 and cysl-2 represent the entry points in the H2S-oxidation and HCN-assimilation pathways, respectively, and expression of both of these enzymes is highly induced by HIF-1 in response to both H2S and HCN. In addition to their role in appropriately responding to H2S and HCN, we found that cysl-1 and cysl-2 are both essential mediators of innate immunity against fast paralytic killing by Pseudomonas. Furthermore, in agreement with these data, we showed that growing worms in the presence of H2S is sufficient to confer resistance to Pseudomonas fast paralytic killing. Our results suggest the hypoxia-independent hif-1 response in C. elegans evolved to respond to the naturally occurring small molecules H2S and HCN.

Volatile-mediated killing of Arabidopsis thaliana by bacteria is mainly due to hydrogen cyanide.

The volatile-mediated impact of bacteria on plant growth is well documented, and contrasting effects have been reported ranging from 6-fold plant promotion to plant killing. However, very little is known about the identity of the compounds responsible for these effects or the mechanisms involved in plant growth alteration. We hypothesized that hydrogen cyanide (HCN) is a major factor accounting for the observed volatile-mediated toxicity of some strains. Using a collection of environmental and clinical strains differing in cyanogenesis, as well as a defined HCN-negative mutant, we demonstrate that bacterial HCN accounts to a significant extent for the deleterious effects observed when growing Arabidopsis thaliana in the presence of certain bacterial volatiles. The environmental strain Pseudomonas aeruginosa PUPa3 was less cyanogenic and less plant growth inhibiting than the clinical strain P. aeruginosa PAO1. Quorum-sensing deficient mutants of C. violaceum CV0, P. aeruginosa PAO1, and P. aeruginosa PUPa3 showed not only diminished HCN production but also strongly reduced volatile-mediated phytotoxicity. The double treatment of providing plants with reactive oxygen species scavenging compounds and overexpressing the alternative oxidase AOX1a led to a significant reduction of volatile-mediated toxicity. This indicates that oxidative stress is a key process in the physiological changes leading to plant death upon exposure to toxic bacterial volatiles.

The beneficial effect of small toxic molecules on dormancy alleviation and germination of apple embryos is due to NO formation.

Deep dormancy of apple (Malus domestica Borkh.) seeds is terminated by a 3-month-long cold stratification. It is expressed by rapid germination of seeds and undisturbed growth of seedlings. However, stimulation of germination of isolated apple embryos is also observed after applying inhibitors of cytochrome c oxidase: nitric oxide (NO) or hydrogen cyanide (HCN) during the first 3-6 h of imbibition of dormant embryos. The aim of this work was to compare the effect of yet another toxic gaseous molecule carbon monoxide (CO) with the effects of HCN and NO on germination of apple embryos and growth and development of young seedlings. We demonstrated that stimulation of germination after short-term pre-treatment with HCN, NO or CO was accompanied by enhanced NO emission from the embryo axes during their elongation. Moreover, similarly high NO production from non-dormant embryos, after cold stratification, was detected. Therefore, we propose that NO may act as signaling molecule in apple embryo dormancy break.

An FTIR study of the induction and release of kiwifruit buds from dormancy.

BACKGROUND: Many deciduous, perennial fruit crops require winter chilling for adequate budbreak and flowering. Recent research has shown that changes in sugar and amino acid profiles are associated with the release of buds from dormancy. This paper uses FTIR spectrometry to provide an alternative mechanism for tracking metabolic changes in the meristems of kiwifruit buds during winter dormancy. The results suggest that the application of multivariate analysis to FTIR spectra has the potential to be a reliable and fast method for detecting structural and compositional changes in fruit crops. RESULTS: Ten wave numbers of the FTIR spectra are used to calculate a bud development function. This function has been validated using data from two seasons and four orchards, and by monitoring the effects of hydrogen cyanamide application, sugar concentrations and soil temperatures on this function. These wave numbers appear to be associated with carbohydrate, pectin and cellulose levels in the meristems. CONCLUSION: It is expected that this FTIR signature can be used to advance our understanding of the influence of the various environmental and physiological factors on the breaking of bud dormancy and shoot outgrowth, including the optimum timing and concentrations of applications of budbreak regulators, such as hydrogen cyanamide.

The mechanisms involved in seed dormancy alleviation by hydrogen cyanide unravel the role of reactive oxygen species as key factors of cellular signaling during germination.

The physiological dormancy of sunflower (Helianthus annuus) embryos can be overcome during dry storage (after-ripening) or by applying exogenous ethylene or hydrogen cyanide (HCN) during imbibition. The aim of this work was to provide a comprehensive model, based on oxidative signaling by reactive oxygen species (ROS), for explaining the cellular mode of action of HCN in dormancy alleviation. Beneficial HCN effect on germination of dormant embryos is associated with a marked increase in hydrogen peroxide and superoxide anion generation in the embryonic axes. It is mimicked by the ROS-generating compounds methylviologen and menadione but suppressed by ROS scavengers. This increase results from an inhibition of catalase and superoxide dismutase activities and also involves activation of NADPH oxidase. However, it is not related to lipid reserve degradation or gluconeogenesis and not associated with marked changes in the cellular redox status controlled by the glutathione/glutathione disulfide couple. The expression of genes related to ROS production (NADPHox, POX, AO1, and AO2) and signaling (MAPK6, Ser/ThrPK, CaM, and PTP) is differentially affected by dormancy alleviation either during after-ripening or by HCN treatment, and the effect of cyanide on gene expression is likely to be mediated by ROS. It is also demonstrated that HCN and ROS both activate similarly ERF1, a component of the ethylene signaling pathway. We propose that ROS play a key role in the control of sunflower seed germination and are second messengers of cyanide in seed dormancy release.