Chromosomal location and expression of green fluorescent protein (gfp) gene in microspore derived transgenic barley (Hordeum vulgare L.).

Four doubled haploid barley lines (A, C, D, E) derived from gfp (green fluorescent protein) transformation and selection following particle bombardment of microspores were studied for gene expression pattern and the location of genome inserts. The integration sites were detected by fluorescence in situ hybridization (FISH) using the gfp plasmid DNA as a probe. Plants from events A, C, D and E all have a single insert site on chromosome 7L(5HL) at different locations while line E has a second insert site on chromosome 5S(7HS). All original transgenic plants were hemizygous for the transgenes and segregated in the T1 and T2 generations. Although line D had no GFP expression, FISH and PCR could detect gfp gene on its chromosome in transformed plants. Expression levels of GFP varied with lines and tissues examined. Plants from line C showed good expression in pollen and an intermediate level in root tips. Plants from A have intermediate expression of GFP in the pollen and light expression in the root tips. Line E showed strong expression in the root-tips and an intermediate level of GFP in the pollen. Lines A and C segregated as a single Mendelian locus while E segregated in a duplicate loci ratio (15:1) on seedling root tips but had low expression frequency in the pollen. PCR results were consistent with GFP expression on root tips in the three segregating lines. The expression of GFP for lines D and E was abnormal and may be related to the physical location of the transgene or the gene construct used.

Transgenic oat plants via visual selection of cells expressing green fluorescent protein.

New selectable markers and selection systems are needed to increase the efficiency and flexibility of plant transformation. The objective of this research was to determine if the green fluorescent protein (gfp) gene could be utilized as a visual selectable marker for transformation of oat (Avena sativa L.). A modified gfp gene was delivered into oat cells by microprojectile bombardment. Cell clusters expressing gfp were visually identified using fluorescence microscopy and physically isolated at each subculture. Eleven independent transgenic cell lines were obtained, and fertile plants regenerated from all lines. Transgene integration and expression were confirmed in transgenic plants and progeny. Transgene expression segregated in a 3 : 1 ratio in progeny of the majority of the transgenic lines.

Comparative sensitivity of Selenastrum capricornutum and Lemna minor to sixteen herbicides.

Aquatic plant toxicity tests are frequently conducted in environmental risk assessments to determine the potential impacts of contaminants on primary producers. An examination of published plant toxicity data demonstrates that wide differences in sensitivity can occur across phylogenetic groups of plants. Yet relatively few studies have been conducted with the specific intent to compare the relative sensitivity of various aquatic plant species to contaminants. We compared the relative sensitivity of the algae Selenastrum capricornutum and the floating vascular plant Lemna minor to 16 herbicides (atrazine, metribuzin, simazine, cyanazine, alachlor, metolachlor, chlorsulfuron, metsulfuron, triallate, EPTC, trifluralin, diquat, paraquat, dicamba, bromoxynil, and 2,4-D). The herbicides studied represented nine chemical classes and several modes of action and were chosen to represent major current uses in the United States. Both plant species were generally sensitive to the triazines (atrazine, metribuzin, simazine, and cyanazine), sulfonureas (metsulfuron and chlorsulfuron), pyridines (diquat and paraquat), dinitroaniline (trifluralin), and acetanilide (alachlor and metolachlor) herbicides. Neither plant species was uniformly more sensitive than the other across the broad range of herbicides tested. Lemna was more sensitive to the sulfonureas (metsulfuron and chlorsulfuron) and the pyridines (diquat and paraquat) than Selenastrum. However Selenastrum was more sensitive than Lemna to one of two thiocarbamates (triallate) and one of the triazines (cyanazine). Neither species was sensitive to selective broadleaf herbicides including bromoxynil, EPTC, dicamba, or 2,4-D. Results were not always predictable in spite of obvious differences in herbicide modes of action and plant phylogeny. Major departures in sensitivity ofSelenastrum occurred between chemicals within individual classes of the triazine, acetanilide, and thiocarbamate herbicides. Results indicate that neither species is predictively most sensitive, and that a number of species including a dicot species such as Myriophyllum are needed to perform accurate risk assessments of herbicides.

Use of knowledge bases and QSARs to estimate the relative ecological risk of agrichemicals: a problem formulation exercise.

Ecological risk assessments can be used to establish the likelihood that an adverse effect will result from exposure to one or more chemicals. When evaluating contaminated sites with many chemicals present, risk assessors must grapple with the problem of quickly identifying the chemicals that are most likely to be of concern, based on effect and exposure assessment information. Many times data gaps exist and the risk assessor is left with decisions on which models to use to estimate the parameter of concern. In the present paper, a procedure is presented for ranking agrichemicals, utilizing the ASTER (ASsessment Tools for the Evaluation of Risk) system. The procedure was employed to rank the relative ecological risk of forty-nine pesticides historically used in agricultural sites in the Walnut Creek watershed near Ames, lowa, USA. Empirical data from the ASTER system were used when available in the associated databases, and quantitative structure-activity relationships and expert systems were invoked when data were lacking. Separate rankings were conducted based on major species taxonomic groupings. Resulting toxic effects thresholds were compared to surface water concentrations.

Effect of management practices on the Streptococcus suis carrier rate in nursery swine.

Management practices on swine farms were analyzed to determine factor(s) associated with high prevalence of pigs that were carriers of Streptococcus suis. Samples were obtained for bacteriologic culture via direct swabbing of palatine tonsils of healthy nursery pigs on 35 farms throughout the United States. Overall, 36.7% of the pigs were determined to be carriers. Isolates of S suis were serotyped, and antimicrobial susceptibility tests were performed by use of Kirby-Bauer techniques. Streptococcus suis types 1 and 2 were most commonly isolated. All isolates were susceptible to enrofloxacin, 97% of the isolates were susceptible to ceftiofur, and 94% were susceptible to ampicillin. However, only 80% of the isolates were susceptible to penicillin, and only 18% were susceptible to tetracycline. Environmental, managerial, nutritional, and health factors were measured on each farm. Excessive temperature fluctuation, high relative humidity, crowding, and an age spread of > 2 weeks between pigs in the same room were the 4 most commonly encountered problems on farms with higher-than-average percentages of carrier pigs. Continuous flow facilities were found on 50% of these farms, and various disease problems, vitamin E/selenium deficiency, inadequate vaccination programs (attributable to the presence of atypical serotypes), and penicillin-resistant strains were found on 6 to 28% of these farms. Overall, 83% (15/18) of farms with higher-than-average percentages of carrier pigs also had a history of clinical S suis disease.

Integrated assessment of contaminated sediments in the lower Fox River and Green Bay, Wisconsin.

Samples of sediment and biota were collected from sites in the lower Fox River and southern Green Bay to determine existing or potential impacts of sediment-associated contaminants on different ecosystem components of this Great Lakes area of concern. Evaluation of benthos revealed a relatively depauperate community, particularly at the lower Fox River sites. Sediment pore water and bulk sediments from several lower Fox River sites were toxic to a number of test species including Pimephales promelas, Ceriodaphnia dubia, Hexagenia limbata, Selenastrum capricornutum, and Photobacterium phosphorum. An important component of the observed toxicity appeared to be due to ammonia. Evaluation of three bullhead (Ictalurus) species from the lower Fox River revealed an absence of preneoplastic or neoplastic liver lesions, and the Salmonella typhimurium bioassay indicated relatively little mutagenicity in sediment extracts. Apparent adverse reproductive effects were noted in two species of birds nesting along the lower Fox River and on a confined disposal facility for sediments near the mouth of the river, and there were measurable concentrations of potentially toxic 2,3,7,8-substituted polychlorinated dibenzo-p-dioxins (PCDDs) and dibenzofurans (PCDFs), and planar polychlorinated biphenyls (PCBs) both in the birds and in sediments from several of the study sites. Based on toxic equivalency factors and the results of an in vitro bioassay with H4IIE rat hepatoma cells, it appeared that the majority of potential toxicity of the PCB/PCDF/PCDD mixture in biota from the lower Fox River/Green Bay system was due to the planar PCBs. The results of these studies are discussed in terms of an integrated assessment focused on providing data for remedial action planning.