Effects of mixtures of herbicides on nutrient cycling and plant support considering current agriculture practices.

The use of mixtures of pesticides and consecutive pesticide applications challenge current regulations aimed at protecting ecosystem health due to unpredictable effects of complex and dynamic mixtures. In this study, we tested the ecotoxicological effects of mixtures of herbicides, applied following a real application scheme of soybean production on soil health in a mesocosm experiment. The experiment included two sequential applications; first, glyphosate + dicamba + clethodim, and 30 days later, flumioxazin + metolachlor. Commercial products were used at the recommended doses and at two other concentrations: half and double the recommended dose. Soybean plants were exposed to the herbicide-contaminated soil from the time of sowing to the beginning of pod formation. Half of the plants were harvested at the vegetative stage and the remaining plants at the reproductive stage to evaluate endpoints related to plant support and nutrient cycling. Plant biomass was significantly affected during the vegetative stage at the recommended and double the recommended dose, with the effects being mixture-dose dependent. Lower total and arbuscular colonization of mycorrhizas were also observed in double the recommended dose, and intermediate results were observed for the recommended dose. Nodule mass and phosphorous concentration in plants decreased with increasing herbicide doses. By the end of the experiment, nodule mass and total mycorrhizal colonization were low in the plants treated with double the recommended dose of herbicides. However, both endpoints reached similar values to the control at lower herbicide doses. Plant height and phenology were only lower at double the recommended dose during the experiment. The use of non-standard endpoints evidenced that important soil functions were transiently or permanently affected, while the realistic application scheme accounted for the impact of the management practice currently used. Pesticide risk assessment should therefore, incorporate both issues to effectively protect the ecosystems.

Effect of auxin on cytodifferentiation and production of quinoline alkaloids in compact globular structures of Cinchona ledgeriana.

Fine cell suspension cultures of Cinchona ledgeriana produce only very low amounts of quinoline alkaloids. These cultures formed self-propagating compact globular structures (CGS) on medium containing 2,4-D and BAP. These CGS could be induced to produce significant amounts of quinoline alkaloids by replacing 2,4-D by low amounts of 1-NAA, which was accompanied by histological changes of the CGS. A few high producing CGS clones could be selected. The stability of this trait was studied over a period of about one year of culture in maintenance medium.

Characterization of a Suspension Culture of Tabernaemontana elegans on Growth, Nutrient Uptake, and Accumulation of Indole Alkaloids*.

A suspension culture of TABERNAEMONTANA ELEGANS was characterized on growth, nutrient uptake (carbohydrate, nitrogen, and phosphate), and on accumulation of indole alkaloids. Besides tryptamine seven indole alkaloids were isolated and identified: vobasine, vobasinol, perivine, isositsirikine, apparicine, tubotaiwine, and O-acetylvallesamine. In spite of the presence of vobasinol as major product, only trace amounts of dimeric indole alkaloids could be detected. Alkaloid formation started at the early stationary phase.

Accumulation of indole alkaloids in a suspension culture of Tabernaemontana divaricata.

Cell suspension cultures of TABERNAEMONTANA DIVARICATA were found to produce relatively large amounts of indole alkaloids. For their isolation an ion-pair DCCC method was used in combination with preparative TLC. The alkaloids were identified as tabernaemontanine, perivine, vobasine, voaphylline, voaphylline-hydroxyindolenine, vallesamine, apparicine, 16-hydroxy-16,22-dihydroapparicine, pericyclivine, tubotaiwine, 19- S-heyneanine, and coronaridine. Voaphylline, the main alkaloid, was produced during growth and early stationary phase and reached a maximum of 23 mg/l at day 19 of the growth cycle. After this maximum voaphylline was rapidly metabolized. Apparicine, vobasine, and coronaridine reached their maximum levels at a later stage of the growth cycle. Tubotaiwine accumulation showed a similar profile as that of voaphylline. In light-grown cells the total production was about 2 times higher than in dark-grown cells, with respective main products voaphylline and apparicine.

Induction of triterpene biosynthesis by elicitors in suspension cultures of Tabernaemontana species.

Treatment of suspension cultures of some Tabernaemontana species (Apocynaceae) with elicitors (e.g. cellulase, Candida albicans) result in a rapid de novo production of antimicrobial active triterpenes. The triterpenes are identified as ursene carboxylic acid derivatives. These triterpenes are not produced by an elicited cell suspension culture of Catharanthus roseus, another Apocynaceae.

A method for the quantitative determination of anthraquinones and alkaloids in cell and tissue cultures of cinchona sp.

A procedure is described for the extraction of both anthraquinones and alkaloids from tissue culture material of CINCHONA species. Using this extraction method, it is possible to quantitatively determine the two groups of secondary metabolites produced in CINCHONA tissue cultures.

Correlation between the presence of membrane-bound auxin binding and root regeneration in cultured tobacco cells.

When cell-suspension cultures and callus tissue from Nicotiana tabacum are grown on medium containing α-naphthaleneacetic acid (NAA) and kinetin, three classes of auxin-binding proteins can be detected. When the herbicide 2,4-dichlorophenoxyacetic acid is used instead of both NAA and kinetin, one of these sites, which is membranebound, disappears. After retransferring cells to medium containing NAA and kinetin, this membrane-bound site reappears after four to eight weeks. This reappearance is correlated with the ability of the cells to regenerate roots.

Anthraquinones as phytoalexins in cell and tissue cultures of Cinchona spec.

The addition of autoclaved mycelia of Aspergillus niger and the known phytopathogenic fungus Phytophtora cinnamomi to cultured cells of Cinchona ledgeriana Moens. caused a marked increase in the anthraquinone content of the plant cells. This finding in combination with the antimicrobial activity of the anthraquinones isolated from calli of Cinchona pubescens Vahl. led to the conclusion that anthraquinones are phytoalexins.

Modulation of the number of membrane-bound auxin-binding sites during the growth of batch-cultured tobacco cells.

We studied the modulation of the number of membrane-bound naphthaleneacetic acid (NAA)-binding sites during the growth cycle of tobacco cells in batch culture. Both cell number and specific NAA-binding increased exponentially, but at different rates and for different periods. This caused a characteristic modulation of the number of binding sites per cell during the growth cycle: During the first day of the lag phase this number decreased; in the exponential phase it rose markedly, and in the stationary phase it was constant.

Auxin-binding by particulate fractions from tobacco leaf protoplasts.

In vitro binding of 1-naphthaleneacetic acid (NAA) to particulate fractions from tobacco leaf protoplasts was studied. In freshly isolated protoplasts no specific binding could be detected, whereas it was present in particulate fractions from tobacco leaves. It is concluded that the NAA-binding-sites are probably located at the external face of the plasma membrane; they are destroyed during protoplast isolation by proteolytic enzymes in the cellulase and macerozyme preparations. After culturing the protoplasts for 3-4 d, the first cell divisions were observed and at the same time specific NAA-binding became detectable. The affinity constant for NAA was approx. 2·10(6) mol(-1) and the number of binding sites increased during further culture.

Initial proliferation of cortical cells in the formation of root nodules in Pisum sativum L.

Root nodule initiation in Pisum sativum begins with cell divisions in the inner cortex at some distance from the advancing infection thread. After penetrating almost the entire cortex, the branches of the thread infiltrate the meristematic area previously initiated in the inner cortical cells. These cells are soon invaded by bacteria released from the infection thread and subsequently differentiate into non-dividing, bacteriod-containing cells. As the initial meristematic centre in the inner cortex is thus lost to bacteroid formation, new meristematic activity is initiated in neighbouring cortical cells. As development proceeds, more cortical layers contribute to the nodule, with the peripheral layer and apical meristem of the nodule not invaded by bacteria.Lateral root primordia are initiated in a region separate from that in which nodules are formed, with the lateral primordia being closer to the root apex. This is interpreted to indicate that the physiological basis for nodule initiation is distinct from that for initiation of lateral roots. The role of a single tetraploid cell in nodule initiation is refuted, as is the existence of incipient meristematic foci in the root. It is suggested that the tetraploid cells in nodule meristems arise from pre-existing endoreduplicated cells, or by the induction of endoreduplication in diploid cortical cells by Rhizobium.