Nature of the Interference Mechanism of Sugarcane (Saccharum officinarum L.) Straw

Sugarcane (Saccharum officinarum L.) straw left in the field after harvest interferes with the growth of winter and summer weeds. In the last years, there was a progressive move away from burning sugarcane straw to retaining it on the soil surface after harvest to prevent soil degradation and environmental pollution. Water-soluble phenolics leachated from straw into soil may suppress weed growth. A study was carried out to investigate (1) the effect of biotic (unautoclaved) soil treated with burned and unburned sugarcane straw leachates on seedling growth and foliar proline content of beggarticks (Bidens subalternans L.) and wild mustard (Brassica campestris L.), (2) the modification of sugarcane straw phytotoxicity in abiotic (autoclaved) soil and biotic (unautoclaved) soil plus activated charcoal, and (3) changes of inorganic ions and phenolic contents in biotic soil after treatment with burned and unburned sugarcane straw leachate. Unburned straw leachate significantly inhibited root elongation of 7-d-old beggarticks and wild mustard seedlings. Burned straw leachate did not affect seedling growth of the assayed weeds suggesting that organic straw phytotoxins were involved. Experiments with activated charcoal, however, did not provide clear evidence supporting the involve of organic molecules in straw phytotoxicity. Unburned straw leachate incorporated in biotic soil was more inhibitory than in abiotic soil on root growth suggesting that microbial activity is involved in sugarcane straw interference. There was no evidence of nutrient microbial immobilization. Unburned sugarcane straw leachate increased total phenolic content in biotic soil more than in abiotic soil or biotic soil plus charcoal. Burned sugarcane straw leachate did not increase phenolic compounds levels in biotic soil. Linear regression analysis indicated a strong correlation between levels of soil phenolic contents and root growth inhibition. Soil characteristics evaluated in soil treated with burned and unburned sugarcane straw leachate suggest that straw phytotoxicity is related with organic molecules, such as phenolic compounds, rather than to variations in inorganic nutrients. Unburned straw leachate induced proline accumulation in seedling leaves of both beggarticks and wild mustard. Proline foliar content was higher in seedlings grown in biotic soil than in seedlings grown in biotic soil plus charcoal suggesting that straw organic constituents induced proline accumulation. Proline foliar content of seedlings grown in biotic soil treated with burned straw leachate was not significantly different from water control. The present study showed that sugarcane straw leachate interferes with seedling growth of beggarticks and wild mustard and that water-soluble phenolics can play a role in the seedling growth inhibition of the assayed weeds.     

The phenolic, 3,4-dihydroxybenzoic acid,is an endogenous regulator of rooting in <Emphasis Type="Italic">Protea cynaroides</Emphasis>

Analysis of stem extracts identified large quantities of 3,4-dihydroxybenzoic acid and other similar phenolics. The exogenous application of 3,4-dihydroxybenzoic acid on Protea cynaroides explants in vitro significantly increased the root mass at 100 mg l⁻¹, but not at lower concentrations, while root inhibition was observed at 500 mg l⁻¹. HPLC analysis of cuttings during vegetative propagation showed a considerable increase in 3,4-dihydroxybenzoic acid levels from initial planting to when root formation took place, indicating for the first time that 3,4-dihydroxybenzoic acid may be an important phenolic compound in regulating root formation in P. cynaroides cuttings. HPLC analysis also identified caffeic, ferulic, gallic and salicylic acids in the cuttings.     

Towards a high-yield bioconversion of ferulic acid to vanillin

Natural vanillin is of high interest in the flavor market. Microbial routes to vanillin have so far not been economical as the medium concentrations achieved have been well below 1 g l⁻¹. We have now screened microbial isolates from nature and known strains for their ability to convert eugenol or ferulic acid into vanillin. Ferulic acid, in contrast to the rather toxic eugenol, was found to be an excellent precursor for the conversion to vanillin, as doses of several g l⁻¹ could be fed. One of the isolated microbes, later identified as Pseudomonas putida, very efficiently converted ferulic acid to vanillic acid. As vanillin was oxidized faster than ferulic acid, accumulation of vanillin as an intermediate was not observed. A completely different metabolic flux was observed with Streptomyces setonii. During the metabolism of ferulic acid, this strain accumulated vanillic acid only to a level of around 200 mg l⁻¹ and then started to accumulate vanillin as the principal metabolic overflow product. In shake-flask experiments, vanillin concentrations of up to 6.4 g l⁻¹ were achieved with a molar yield of 68%. This high level now forms the basis for an economical microbial production of vanillin that can be used for flavoring purposes. Received: 15 October 1998 / Received revision: 13 January 1999 / Accepted: 18 January 1999     

Acute and Sub-lethal Toxicity of Landfill Leachate Towards Two Aquatic Macro-invertebrates: Demonstrating the Remediation Potential of Aerobic Digestion

A specific landfill leachate that contained 1.036 mgl⁻¹of 2-chlorobiphenyl was used in the study (255 mg l⁻¹ COD and 133 mg l⁻¹ BOD₅). Three, 2-l semi-continuous batch reactors (SBRs) were used to simulate the treatment potential of this method on a small scale. Aerobic digestion effectively reduced the leachates COD concentration. Regardless of dilution, the leachates COD reached a <20 mg l⁻¹ equilibrium after 96 h exposure to aerobic digestion, however, increasing the level of dilution accelerated the process. In untreated leachate, the LC₅₀ for Asellus aquaticus was 57% v/v leachate in deionised water and 5% for Gammarus pulex (96 h, static LC₅₀ tests without nutrition and oxygen depleting conditions). After being exposed to aerobic digestion, these values rose to 95% and 40%, respectively. Prolonged exposure to a 1:20 sub-lethal dilution of the aforementioned leachate has been previously shown to affect the breeding colony size of Asellus aquaticus and a 1:66 dilution influenced the fecundity of a Gammarus pulex population. After remediation by aerobic digestion, however, the population dynamics of both test species remained unaltered.     

Role of sugarcane straw allelochemicals in the growth suppression of arrowleaf sida

Previous studies suggested that allelochemicals from sugarcane straw may suppress the growth of arrowleaf sida (Sida rhombifolia L.). A study was conducted to establish: (1) the direct or indirect role of the organic molecules from sugarcane straw leachate on the growth suppression of arrowleaf sida and (2) if leachate phytotoxins induce proline accumulation in arrowleaf sida tissues as an adaptative response to a water or an oxidative stress. Inhibition of root elongation was the primary effect of sugarcane straw leachate on arrowleaf sida grown in unsterile soil. Addition of activated charcoal to unsterile soil before incorporation of straw leachate reduced the inhibition in root growth suggesting a direct participation of organic molecules in leachate phytotoxicity on arrowleaf sida. Inorganic straw constituents did not inhibit root growth while microbial activity increased leachate phytotoxicity. Soil chemical analysis suggested a direct action of organic molecules in leachate phytotoxicity rather than variations in macro and micronutrients or nutrient microbial immobilization. Straw leachate induced proline accumulation in roots and cotyledons of arrowleaf sida. Proline increase was related with oxidative stress in the roots but not in the cotyledons. Our results indicate a direct action of organic compounds from sugarcane straw and/or their microbial transformation products on root growth of arrowleaf sida. These substances induced proline accumulation in roots mainly as consequence of an oxidative stress while water stress may be the main cause of high proline content in the cotyledons. Although the observed responses could be due to phenolic compounds, the involvement of organic molecules with other chemical nature could not be excluded.     

Effects of Rhamnolipids from Pseudomonas aeruginosa DS10-129 on Luminescent Bacteria: Toxicity and Modulation of Cadmium Bioavailability

In this study, the mixture of mono- and di-rhamnolipids produced by Pseudomonas aeruginosa DS10-129 was characterized for its toxicity and modulatory effects on Cd availability to different bacteria. Gram-negative naturally bioluminescent Vibrio fischeri and recombinant bioluminescent Pseudomonas fluorescens, P. aeruginosa, Escherichia coli, and Gram-positive Bacillus subtilis were used as model organisms. Rhamnolipids reduced the bioluminescence of these bacteria in less than a second of exposure even in relatively low concentrations (30-min EC₅₀ 45–167 mg l⁻¹). Toxicity of Cd to Gram-negative bacteria (30-min EC₅₀ values 0.16 mg l⁻¹ for E. coli, 0.96 mg l⁻¹ for P. fluorescens, and 4.4 mg l⁻¹ for V. fischeri) was remarkably (up to 10-fold) reduced in the presence of 50 mg l⁻¹ rhamnolipids. Interestingly, the toxicity of Cd to Gram-positive B. subtilis (30-min EC₅₀ value 0.49 mg l⁻¹) was not affected by rhamnolipids. Rhamnolipids had an effect on desorption of Cd from soil: 40 mg l⁻¹ rhamnolipids increased the water-extracted fraction of Cd twice compared with untreated control. However, this additionally desorbed fraction of Cd remained bound with rhamnolipids and was not available to bacteria. Hence, in carefully chosen concentrations (still effectively complexing heavy metals but not yet toxic to soil bacteria), rhamnolipids could be applied in remediation of polluted areas.     

In vitro callus induction and regeneration studies in Withania somnifera

Callus cultures were initiated from axillary leaves, axillary shoots, hypocotyls, and root segments on Murashige and Skoog (MS) (1962) medium supplemented with 2,4-D (2 mg l⁻¹) and KN (0.2 mg l⁻¹). Shoots differentiated best from axillary shoot base callus on MS medium containing BA (2 mg l⁻¹). Regenerated shoots rooted best on MS medium containing IBA (2 mg l⁻¹) alone, and IBA (2 mg l⁻¹) with IAA (2 mg l⁻¹). Plantlets were transferred to pots containing sand and soil mixture, acclimatized in a culture room and afterwards transferred to the glasshouse. This revised version was published online in June 2006 with corrections to the Cover Date.     

Optimization of biotin and thiamine requirements for somatic embryogenesis of date palm (Phoenix dactylifera L.)

Summary This study was conducted to examine the effect of biotin and thiamine concentrations on callus growth and somatic embryogenesis of date palm (Phoenix dactylifera L.). Embryogenic callus derived from offshoot tip explants was cultured on hormone-free MS medium containing biotin at 0, 0.1, 1, or 2 mg l⁻¹ combined with thiamine at 0.1, 0.5, 2, or 5 mg l⁻¹. Embryogenic callus weight, number of resultant embryos, and embryo length were significantly influenced by thiamine and biotin concentration. The optimum callus growth treatment consisted of 0.5 mg l⁻¹ thiamine and 2 mg l⁻¹ biotin. This treatment also gave the highest number of embryos. Embryo elongation was greatest at 0.5 or 2 mg l⁻¹ thiamine combined with 1 mg l⁻¹ biotin. Embryos from all treatments germinated and regenerants exhibited normal growth in soil. This study provides an insight into the importance of optimizing various culture medium components to overcome in vitro recalcitrace of date palm.     

Production of <Emphasis Type="SmallCaps">d</Emphasis>-lactic acid from sugarcane molasses,sugarcane juice and sugar beet juice by <Emphasis Type="Italic">Lactobacillus delbrueckii</Emphasis>

Lactobacillus delbrueckii was grown on sugarcane molasses, sugarcane juice and sugar beet juice in batch fermentation at pH 6 and at 40°C. After 72 h, the lactic acid from 13% (w/v) sugarcane molasses (119 g total sugar l⁻¹) and sugarcane juice (133 g total sugar l⁻¹) was 107 g l⁻¹ and 120 g l⁻¹, respectively. With 10% (w/v) sugar beet juice (105 g total sugar l⁻¹), 84 g lactic acid l⁻¹ was produced. The optical purities of d-lactic acid from the feedstocks ranged from 97.2 to 98.3%.     

Effects of hygromycin on cotton cultures and its application in <Emphasis Type="Italic">Agrobacterium</Emphasis>-mediated cotton transformation

We have evaluated the effects of the antibiotic hygromycin B on cotton (Gossypium hirsutum L.) callus induction, callus proliferation, and seed germination. Nontransgenic cotyledon and hypocotyl showed obvious variance in tolerance to hygromycin. Cotyledons were more sensitive to hygromycin than hypocotyls. Hygromycin at 7.5 and 20 mg l⁻¹ completely inhibited callus initiation from cotyledon and hypocotyl explants, respectively. Nontransformed calli did not grow on media supplemented with 10 mg l⁻¹ hygromycin and were killed at 15 mg l⁻¹. In seed germination assay, the presence of 20 mg l⁻¹ hygromycin significantly suppressed shoot and root elongation of seedlings. This hygromycin concentration was applied to select regenerated transgenic plantlets and their progenies. Based on these results, we developed an efficient hygromycin selection protocol for Agrobacterium-mediated cotton transformation and regeneration.     

Quantitative analysis of phenolics in selected crop species and biological activity of these compounds evaluated by sensitivity of Echinochloa crus-galli

The purpose of this study was to determine the content of selected phenolic compounds in white mustard, buckwheat, spring barley, oat and rye grown under field conditions. Moreover, the allelopathic efficiency of these compounds was evaluated by sensitivity of Echinochloa crus-galli. The aromatic acids: trans-cinnamic, salicylic, ferulic, chlorogenic, p-hydroxybenzoic, protocatechuic, p-coumaric and vanillic were separated from crop plants by TLC and determined spectrophotometrically. Differences in concentrations of analysed compounds were observed for most of the examined plant species. The highest concentration was noticed for cinnamic acid and ranged from 360 μg·g⁻¹ DW in rye to 2770 μg·g⁻¹ DW in spring barley. The relatively high concentration was noticed for ferulic acid (from 73.8 μg·g⁻¹ DW in buckwheat to 1046 μg·g⁻¹ DW in spring barley) and p-coumaric acid (from 50 μg·g⁻¹ DW in oat to 1499 μg·g⁻¹ DW in buckwheat). The observed differences in the phenolics content between two successive vegetation seasons can reflect the effect of abiotic and biotic environmental factors on the phenolics level in studied plants. In the greenhouse experiment the effect of particular compounds on the growth of Echinochloa crus-galli was also studied. It has been found that the examined phenolics, and especially trans-cinnamic acid and mixture of phenolic compounds, significantly inhibit the growth of Echinochloa crus-galli. The obtained results may contribute to the explanation of the biological activity of some phenolic compounds.     

Simplified clonal multiplication of mulberry using liquid shake culture

Organogenesis was induced in callus derived from mature zygotic embryos of six families of loblolly pine (Pinus taeda L.) within 24 weeks of culture. Elongation of adventitious buds was achieved on TE medium supplemented with 0.5 mg l⁻¹ indole-3-butyric acid (IBA) and 2 mg l⁻¹ 6-benzyladenine (BA). The most suitable medium for root formation proved to be TE medium supplemented with 0.1 mg l⁻¹ IBA, 1 mg l⁻¹ BA, and 0.5 mg l⁻¹ gibberellic acid (GA₃). One hundred and sixty-nine regenerated plantlets were transferred to a perlite:peatmoss:vermiculite (1:1:1) soil mixture, and 98 plantlets survived in the field. This revised version was published online in June 2006 with corrections to the Cover Date.     

Large-scale cultivation of adventitious roots of Echinacea purpurea in airlift bioreactors for the production of chichoric acid, chlorogenic acid and caftaric acid

Adventitious roots of Echinacea purpurea were cultured in airlift bioreactors (20 l, 500 l balloon-type, bubble bioreactors and 1,000 l drum-type bubble bioreactor) using Murashige and Skoog (MS) medium with 2 mg indole butyric acid l⁻¹ and 50 g sucrose l⁻¹ for the production of chichoric acid, chlorogenic acid and caftaric acid. In the 20 l bioreactor (containing 14 l MS medium) a maximum yield of 11 g dry biomass l⁻¹ was achieved after 60 days. However, the amount of total phenolics (57 mg g⁻¹ DW), flavonoids (34 mg g⁻¹ DW) and caffeic acid derivatives (38 mg g⁻¹ DW) were highest after 50 days. Based on these studies, pilot-scale cultures were established and 3.6 kg and 5.1 kg dry biomass were achieved in the 500 l and 1,000 l bioreactors, respectively. The accumulation of 5 mg chlorogenic acid g⁻¹ DW, 22 mg chichoric acid g⁻¹ DW and 4 mg caftaric acids g⁻¹ DW were achieved with adventitious roots grown in 1,000 l bioreactors.     

Factors influencing efficiency of somatic embryogenesis of Gentiana kurroo (Royle) cell suspension

In this paper, we would like to show unexpected morphogenic potential of cell suspensions derived from seedling explants of Gentiana kurroo (Royle). Suspension cultures were established with the use of embryogenic callus derived from seedling explants (root, hypocotyl and cotyledons). Proembryogenic mass proliferated in liquid MS medium supplemented with 0.5 mg l⁻¹ 2,4-D and 1.0 mg l⁻¹ Kin. The highest growth coefficient was achieved for root derived cell suspensions. The microscopic analysis showed differences in aggregate structure depending on their size. To assess the embryogenic capability of the particular culture, 100 mg of cell aggregates was implanted on MS agar medium supplemented with Kin (0.0–2.0 mg l⁻¹), GA₃ (0.0–2.0 mg l⁻¹) and AS (80.0 mg l⁻¹). The highest number of somatic embryos was obtained for cotyledon-derived cell suspension on GA₃-free medium, but the best morphological quality of embryos was observed in the presence of 0.5–1.0 mg l⁻¹ Kin, 0.5 mg l⁻¹ GA₃ and 80.0 mg l⁻¹ AS. The morphogenic competence of cultures also depended on the size of the aggregate fraction and was lower when size of aggregates decreased. Flow cytometry analysis reveled luck of uniformity of regenerants derived from hypocotyl suspension and 100% of uniformity for cotyledon suspension.     

Direct organogenesis and plantlet regeneration from mature zygotic embryos of masson pine (Pinus massoniana L.)

Mature zygotic embryos of masson pine were cultured as initial explants to investigate the process of direct organogenesis. Adventitious buds were initiated on DCR medium (Douglas-fir cotyledon revised medium) supplemented with 0.5 mg l⁻¹ N⁶-benzyladenine (BA) and 0.05 mg l⁻¹ indolebutyric acid (IBA) or α-naphthaleneacetic acid (NAA). The highest induction frequency of adventitious buds was 99.3%. Subsequent transfer of buds to medium with lower concentrations of plant growth regulators in time was necessary for differentation of high quality adventitious buds. After culturing on elongating medium, in which the proportion of cytokinins to auxins was reduced, shoots higher than 2 cm were transferred for root induction to GD medium with half of the concentration of macro-salts (½ GD) and with 2 mg l⁻¹ IBA and 0.05 mg l⁻¹ BA. The average root frequency was over 70%. After adventitious roots had appeared, the shoots were transferred to ½ GD medium with a lower concentration of IBA (0.2 mg l⁻¹) for further root development.     

The effects of gap disturbance on nitrogen cycling and retention in late-successional northern hardwood–hemlock forests

Late-successional forests in the upper Great Lakes region are susceptible to nitrogen (N) saturation and subsequent nitrate (NO₃⁻) leaching loss. Endemic wind disturbances (i.e., treefall gaps) alter tree uptake and soil N dynamics; and, gaps are particular susceptible to NO₃⁻ leaching loss. Inorganic N was measured throughout two snow-free periods in throughfall, forest floor leachates, and mineral soil leachates in gaps (300–2,000 m², 6–9 years old), gap-edges, and closed forest plots in late-successional northern hardwood, hemlock, and northern hardwood–hemlock stands. Differences in forest water inorganic N among gaps, edges, and closed forest plots were consistent across these cover types: NO₃⁻ inputs in throughfall were significantly greater in undisturbed forest plots compared with gaps and edges; forest floor leachate NO₃⁻ was significantly greater in gaps compared to edges and closed forest plots; and soil leachate NO₃⁻ was significantly greater in gaps compared to the closed forest. Significant differences in forest water ammonium and pH were not detected. Compared to suspected N-saturated forests with high soil NO₃⁻ leaching, undisturbed forest plots in these late-successional forests are not losing NO₃⁻ (net annual gain of 2.8 kg ha⁻¹) and are likely not N-saturated. Net annual NO₃⁻ losses were observed in gaps (1.3 kg ha⁻¹) and gap-edges (0.2 kg ha⁻¹), but we suspect these N leaching losses are a result of decreased plant uptake and increased soil N mineralization associated with disturbance, and not N-saturation.     

The mycobiota of landfill leachates in the pretreatment process in a sequencing batch reactor

The paper discusses the dynamics of the accumulation of microscopic fungi, depending on the sludge load (Bx), in activated sludge used for landfill leachate pretreatment. The propagule washout from the sludge into pretreated leachates is determined, including genera and species that may threaten environmental health. An increased accumulation of microscopic fungi in sludge flocs occurred at Bx=0.23−0.45 mg chemical oxygen demand (COD) mg⁻¹ d⁻¹. Microscopic fungi were eluted at the maximal Bx value tested of 1.64 mg COD mg⁻¹ d⁻¹. Both the activated sludge and the leachate runoff from the sequencing batch reactor (SBR) pose health risks to the environment due to the occurrence of fungi such as Aspergillus fumigatus, Purpureocillium lilacinum, Cyberlindnera jadinii (C. utilis), Geotrichum candidum and G. fragrans. Their count is sufficient to cause multi-organ infections in homeothermal animals and in humans.     

Combined effects of phytohormone, indole-3-butyric acid, and methyl jasmonate on root growth and ginsenoside production in adventitious root cultures of Panax ginseng C.A. Meyer

Indole-3-butyric acid at 25 μM with methyl jasmonate (MJ) at 100 μM in Panax ginseng synergistically stimulated both root growth and ginsenoside accumulation compared with 100 μM MJ alone. Productivity of ginsenoside was 10 mg l⁻¹ d⁻¹ compared to 7.3 mg l⁻¹ d⁻¹ with MJ elicitation alone.     

Morphogenetic pathway in petiole derived callus of Amorphophallus albus in vitro

Two different morphogenetic pathways, adventitious bud and corm-like structure (CLS), were observed on organogenic calli derived from the petioles of Amorphophallus albus in vitro. The organogenic calli was established via culture of petiole segments on Murashige and Skoog (MS) medium supplemented with 1.0 mg l⁻¹ α-naphthaleneacetic acid (NAA) and 1.0 mg l⁻¹ 6-benzyladenine (BA) and subculture of the petiole-derived calli on MS medium with 0.5 mg l⁻¹ NAA and 0.5 mg l⁻¹ BA. These organogenic calli were used to induce morphogenesis via culture on MS medium with various concentrations of NAA and BA. BA alone favoured adventitious bud differentiation (57.0 ± 8.3% at maximum) from the organogenic calli but inhibited CLS formation. In the presence of NAA and BA, both adventitious bud and CLS were observed in a same culture system. The maximum CLS formation (71.2 ± 9.3%) were found on MS medium with 0.5 mg l⁻¹ NAA and 2.0 mg l⁻¹ BA, associated with 26.7 ± 8.6% adventitious bud differentiation. A small part of the adventitious buds developed into normal shoots which needed rooting culture phase to form complete plants. About 80% survival rate was obtained with these plants after transplantation to soil. More than 90% of the CLSs produced complete plants with shoots and root systems, regardless of the rooting media tested. Transplantation of the CLS-derived plants to soil gave 100% survival rate. Histological observations revealed both the two morphogenetic events originated from the meristematic cells located in superficial layers of callus tissue.     

Rapid in vitro multiplication and ex vitro rooting of Malus zumi (Matsumura) Rehd

Micropropagation system of Malus zumi was optimized by studying the influence of plant growth regulators and culture conditions. The axillary buds were used for mutiplication of in vitro shoot culture on agar Murashige and Skoog (1962) (MS) medium with combination of 1 mg l⁻¹ BAP, 0.5 mg l⁻¹ NAA or 0.5 mg l⁻¹ IAA or 0.5 mg l⁻¹ IBA under 16 h photoperiod. The shoot growth in culture was not significantly affected within a broad range (5.0–7.0) of initial medium pH. The highest shoot (13) was obtained on medium containing 1.0 mg l⁻¹ BAP and 0.5 mg l⁻¹ IAA. Well-developed shoots, 35–50 mm in length, were successfully rooted ex vitro at 86.3% by a 2-h-treatment with aqueous solution containing MS salts and 100 mg l⁻¹ IBA prior to their planting in growing substrate composed of soil and vermiculite (1:1 v/v). The survival rate of transplantation reached 88.0% when transferred to field condition.