Evaluation of acute and chronic toxicity of selected compounds in higher plants using cell culture

Summary The feasibility of using plant cell culture to measure toxicity was determined by investigating the toxicological effects of three chemical compounds, allyl alcohol, propargylglycine, and cadmium chloride, on cell cultures ofCatharanthus roseus G. Don (Madagascar periwinkle). Suspension cultures ofC. roseus were maintained in modified B5 medium and transferred every 5 d. Five-day-old cell cultures were exposed to various concentrations (10,3,1,0.3,0.1,0.03,0.01,0.003,0.001,0.0003,0.0001, 0.00003, and 0.0 mM) of the toxicants in both acute and chronic toxicity tests. In the acute test, cells were exposed to the toxicant for 24 h, washed three times with sterile medium, and plated in petri plates with an equal volume of 1.4% agar medium. Cells in the chronic test were plated with an equal volume of 1.4% agar medium containing various concentrations of the toxicant. Cells were incubated 28 d at 30°C in the dark. The colonies were counted and the results plotted as percent survival versus toxicant concentration. The results indicate, at the concentrations tested, thatC. roseus assay may be feasible in that it fulfills the criteria for a practical assay (e.g., rapid, simple, quantifiable, and reproducible). This work was submitted to the faculty of Miami University in partial fulfillment of the requirements for the degree of Master of Environmental Science, Institute of Environmental Sciences.     

How do elevated CO2 and O3 affect the interception and utilization of radiation by a soybean canopy?

Net productivity of vegetation is determined by the product of the efficiencies with which it intercepts light (?_i) and converts that intercepted energy into biomass (?_c). Elevated carbon dioxide (CO₂) increases photosynthesis and leaf area index (LAI) of soybeans and thus may increase ?_i and ?_c; elevated O₃ may have the opposite effect. Knowing if elevated CO₂ and O₃ differentially affect physiological more than structural components of the ecosystem may reveal how these elements of global change will ultimately alter productivity. The effects of elevated CO₂ and O₃ on an intact soybean ecosystem were examined with Soybean Free Air Concentration Enrichment (SoyFACE) technology where large field plots (20‐m diameter) were exposed to elevated CO₂ (～550 μmol mol^?1) and elevated O₃ (1.2 × ambient) in a factorial design. Aboveground biomass, LAI and light interception were measured during the growing seasons of 2002, 2003 and 2004 to calculate ?_i and ?_c. A 15% increase in yield (averaged over 3 years) under elevated CO₂ was caused primarily by a 12% stimulation in ?_c , as ?_i increased by only 3%. Though accelerated canopy senescence under elevated O₃ caused a 3% decrease in ?_i, the primary effect of O₃ on biomass was through an 11% reduction in ?_c. When CO₂ and O₃ were elevated in combination, CO₂ partially reduced the negative effects of elevated O₃. Knowing that changes in productivity in elevated CO₂ and O₃ were influenced strongly by the efficiency of conversion of light energy into energy in plant biomass will aid in optimizing soybean yields in the future. Future modeling efforts that rely on ?_c for calculating regional and global plant productivity will need to accommodate the effects of global change on this important ecosystem attribute.     

Leaf micromorphology inAgathis and its taxonomic implications

This study surveys the micromorphological surface structure of the leaves of the conifer genusAgathis (Araucariaceae) from throughout the range of the genus (Malaysia to New Zealand and Fiji) as seen with the scanning electron microscope. These data confirm that the vegetative parts ofAgathis are taxonomically highly conservative, but suggest the Melanesian and New Zealand elements to be distinctive from those of the rest of the genus, and from one another. Conclusions are compared with those that have been derived from studies based on other characters.     

Immunocytochemical localization of tubulin with colloidal gold in cilia of rabbit tracheal epithelial cultures

Ciliated tracheal epithelia cell cultures were investigated immunocytochemically with anti-tubulin and colloidal gold. When rabbit tracheal cultures were fixed in paraformaldehyde, treated with acetone, anti-tubulin and a second antibody coupled to FITC, fluorescence was associated with cytoskeletal and axonemal microtubules. Cilia covering the apical surface of the ciliated tracheal cells fluoresced very brightly thus facilitating identification of this cell type. Electron microscopy of tracheal cultures fixed as above, treated with Triton-X 100 and incubated in anti-tubulin and protein A coupled to colloidal gold resulted in the highly specific localization of tubulin in ciliary axonemes and basal bodies. Omission of primary or secondary antibody resulted in extremely low levels of fluorescence while no colloidal gold particles could be detected in cultures at the electron microscopy level when rabbit anti-tubulin was omitted.     

Haploid plant regeneration from unpollinated ovule cultures of niger (<Emphasis Type="Italic">Guizotia abyssinica</Emphasis> (L. f.) Cass.)

Haploid plants were regenerated in vitro from unpollinated ovules of niger (Guizotia abyssinica (L. f.) (Cass.) on Murashige and Skoog nutrient medium (MS) supplemented with 10 μM naphthaleneacetic acid or 10 μM NAA + 1.5 μM kinetin and 30 g/l sucrose. Gamborg (B5) medium was the best for plant regeneration (in comparison with MS, Nitsch and Nitsch (NN), and Chu (N6) media) from cultured ovules, and 6.66 and 7.33 ovules of JNC-6 and Ootacamund cultivars were involved in direct plant regeneration on this medium. Matured ovules (ovules collected one day before anthesis or on the day of anthesis) only responded to cultural regimes and involved in direct plantlet development. Cytological preparation of root tips and chloroplast counts in the guard cells of leaf stomata of regenerated plants confirmed their haploid nature. This text was submitted by the authors in English.     

Effects of different sterols on the inhibition of cell culture growth caused by the growth retardant tetcyclacis

Cell division in cell suspension cultures can be completely blocked by the growth retardant tetcyclacis at a concentration of 10^-4 mol l^-1. In rice cells it has been demonstrated that the growth inhibition can be completely overcome by application of cholesterol independent of the duration of pretreatment with tetcyclacis. In suspension cultures of maize and soybean, too, the effect of tetcyclacis on cell division was neutralized by adding cholesterol. Other plant sterols, stigmasterol, campesterol and sitosterol were active in a decreasing order. Modifications in the cholesterol perhydro-cyclopentanophenanthrene-ring system indicate that the hydroxyl group at C-3 and the double bond between C-5 and C-6 in ring B are required for the activity. In contrast, gibberellic acid as well as ent-kaurenoic acid could not compensate retardant effects. Likewise, tetcyclasis did not change the level of gibberellins in rice cells as shown by radioimmunoassay. Thus, it is concluded that in cell suspension cultures sterols play a more important role in cell division than gibberellins.Abbreviation GA_x gibberelin A_x     

Diet of the Indochinese silvered langur (Trachypithecus germaini) in Kien Luong Karst area,Kien Giang Province

The Indochinese silvered langur (Trachypithecus germaini) is distributed to the west of Mekong River in Cambodia, Lao PDR, Thailand and Vietnam. During a two‐year study, from May 2014 to May 2016, we collected 320.44 hr of behavior, with 17,040 feeding bouts recorded (142 hr) for T. germaini on Chua Hang Karst Mountain, Kien Luong District, Kien Giang Province, Vietnam. Feeding accounted for 45% of the Indochinese silvered langurs’ activity budget. The plant diet of the Indochinese silvered langurs was principally composed of young leaves (58%), followed by mature leaves (9.5%), fruits (22.7%), flowers (4.7%), buds (3.3%), petioles (1.2%), and other (0.5%). A total of 58 plant species were fed on by the silvered langurs, and leaves of eight species (Phyllathus reticulatus, Ficus rumphii, Ficus tinctoria, Ficus microcarpa, Cayratia trifolia, Streblus ilicifolia, Combretum latifolium, and Streblus asper) were fed on throughout the year. P. reticulatus was most frequently eaten (13.9% feeding time, n = 1,733). Food selection differed significantly between months and seasons. The Indochinese silvered langurs ate 27 plant species in the wet season compared with 23 plant species in the dry season. Leaf chemical composition of two food categories, 16 eaten species (with 10 most frequently consumed species and six least consumed species), and four noneaten species, were analyzed. Feeding samples from eaten species in the Indochinese silvered langurs's diet contained lower amounts of condensed tannin, lignin, protein, ash, and lipids, but a higher amount of total sugar compared with samples from noneaten species. Furthermore, the most frequently consumed species contained lower amounts of lignin compared with the less frequently consumed species. Using a generalized linear model with five variables, including neutral detergent fiber (NDF), total sugar, lignin, lipid, and calcium (Ca) indicated that NDF positively correlated and lignin content negatively correlated with feeding records in the diet of these langur.     

Isolation and characterization of highly purified mosquito oostatic hormone

Oostatic hormone, the hormone that inhibits vitellogenesis in mosquitoes, was purified 7,000-fold with a recovery of 70% from the ovaries of the mosquito Aedes aegypti. The purification procedure included heat treatment and chromatography on ion exchange and gel filtration columns. The hormone is a small peptidelike molecule of molecular weight 2,200 at pH 4.5, which aggregates into larger molecular species of trimer and octamer at pH 7.0 as determined by gel filtration. The hormone is positively charged at pH 7.8 and has a low Rf at pH 9.4 on disc gel electrophoresis. Injection of purified oostatic hormone (9 ng) into female mosquitoes inhibited yolk deposition and vitellogenin synthesis. Activity of the oostatic hormone in the mosquito ovary increased rapidly following blood feeding and reached a maximum after 48 h. Oostatic hormone of A. aegypti injected into autogenous Aedes taeniorhynchus inhibited egg development. Repeated injections of dilute oostatic hormone at 24 h intervals partially arrested egg development, resulting in 60% reduction in the number of eggs laid. This hormone does not block release of egg development neurosecretory hormone (EDNH) from the mosquito brain but rather appears to act on the ovary.