Altered pesticide use on transgenic crops and the associated general impact from an environmental perspective.

The large-scale commercial cultivation of transgenic crops has undergone a steady increase since their introduction 10 years ago. Most of these crops bear introduced traits that are of agronomic importance, such as herbicide or insect resistance. These traits are likely to impact upon the use of pesticides on these crops, as well as the pesticide market as a whole. Organizations like USDA-ERS and NCFAP monitor the changes in crop pest management associated with the adoption of transgenic crops. As part of an IUPAC project on this topic, recent data are reviewed regarding the alterations in pesticide use that have been observed in practice. Most results indicate a decrease in the amounts of active ingredients applied to transgenic crops compared with conventional crops. In addition, a generic environmental indicator -- the environmental impact quotient (EIQ) -- has been applied by these authors and others to estimate the environmental consequences of the altered pesticide use on transgenic crops. The results show that the predicted environmental impact decreases in transgenic crops. With the advent of new types of agronomic trait and crops that have been genetically modified, it is useful to take also their potential environmental impacts into account.

Basis for antagonism by sodium bentazon of tritosulfuron toxicity to white bean (Phaseolus vulgaris L.).

White bean (Phaseolus vulgaris L.) was used to study the antagonism caused by Na-bentazon on the phytotoxic action of the sulfonylurea (SU) herbicide tritosulfuron. After 168 h, uptake and translocation of [14C]tritosulfuron were reduced by 60 and 89%, respectively, when Na-bentazon was added to the mixture. Addition of (NH4)2SO4 or replacement of Na-bentazon with NH4-bentazon completely eliminated the negative effects on [14C]tritosulfuron uptake but not on its translocation. Scanning electron microscopy revealed that a mixture of Na-bentazon plus tritosulfuron plus DASH HC (0.156%) formed a rough layer of grain-like crystals on the leaf surface, whereas the addition of (NH4)2SO4 or replacement of Na-bentazon with NH4-bentazon resulted in amorphous deposits that may be more easily absorbed. The antagonism of tritosulfuron's phytotoxicity by Na-bentazon involves two separate processes, chemical (uptake effect) and biochemical (translocation effect).

2,4-D butoxyethyl ester kinetics in embryos of Xenopus laevis: the role of the embryonic jelly coat in reducing chemical absorption.

The role of the jelly coat in providing a protective barrier to chemical absorption was studied using the embryos of the amphibian, Xenopus laevis. Embryos with or without a jelly coat were water exposed to the butoxyethyl ester of 2,4-dichlorophenoxyacetic acid (2,4-D BEE) and the rates of uptake, metabolism, distribution, and excretion were determined. The water uptake clearance rates were slower for embryos with a jelly coat (1.5-4.5 ml(water).g (embryo) (-1).h(-1) or 0.040-0.022 ml(water).h(-1) per embryo) in comparison to dejellied embryos (14-21 ml(water).g (embryo) (-1).h(-1) 0.0066-0.021 ml(water).h(-1) per embryo). This accounted for the much lower residues in embryos with a jelly coat than in dejellied embryos during 8 h of exposure. Despite quantitative differences in uptake, once 2,4-D BEE had entered the embryos, metabolism and distribution were similar between the two test groups. 2,4-D BEE was metabolized to 2,4-dichlorophenoxyacetic acid (2,4-D) with half-lives ranging from 35 to 42 minutes. The radioactive residues, as determined by whole body autoradiography, appeared throughout the embryo with a slight accumulation in the blastocoel. Furthermore, 35% of the radioactive residues were located in the jelly coat and 65% in the developing embryo. Based on a slower 2,4-D elimination in embryos with a jelly coat, the diffusive properties that decreased 2,4-D BEE uptake appeared to similarly decrease elimination of its metabolite. The common practice of removing jelly coats prior to embryonic amphibian toxicity studies, as in the widely used Frog Embryo Teratogenesis Assay-Xenopus (FETAX), is discouraged based on the kinetic differences observed in this study.

MCPA (4-Chloro-2-ethylphenoxyacetate) resistance in hemp-nettle (Galeopsis tetrahit L.).

The physiological basis for MCPA resistance in a hemp-nettle (Galeopsis tetrahit L.) biotype, obtained from a MCPA-resistant field population, was investigated. Dose-response studies revealed that the resistance factor for MCPA, based on GR50 comparisons of total dry weight of resistant (R) and susceptible (S) plants, was 3.3. Resistance factors for fluroxypyr, dicamba, 2,4-D, glyphosate, and chlorsulfuron were 8.2, 1.7, 1.6, 0.7, and 0.6, respectively. MCPA resistance was not due to differences in absorption, because both R and S biotypes absorbed 54% of applied [14C]MCPA 72 h after treatment. However, R plants exported less (45 vs 58% S) recovered 14C out of treated leaves to the apical meristem (6 vs 13% S) and root (32 vs 38% S). In both biotypes, approximately 20% of the 14C recovered in planta was detected as MCPA metabolites. However, less of the 14C recovered in the roots of R plants was MCPA. Therefore, two different mechanisms protect R hemp-nettle from MCPA phytotoxicity: a lower rate of MCPA translocation and a higher rate of MCPA metabolism in the roots. In support of these results, genetic studies indicated that the inheritance of MCPA resistance is governed by at least two nuclear genes with additive effects.

Effect of release herbicide on mortality, avoidance response, and growth of amphibian larvae in two forest wetlands.

Effects of Release herbicide (triclopyr butoxyethyl ester, [TBEE]) on mortality, avoidance response, and growth of larval amphibians (Rana clamitans, Rana pipiens) were investigated using in situ enclosures deployed in two forest wetlands in northern Ontario, Canada. Release was applied at nominal concentrations ranging from 0.26 to 7.68 mg TBEE acid equivalents (AE)/L. No significant deleterious effects of this herbicide on larval growth were detected. However, concentration-dependent mortality and abnormal avoidance response were observed. Most mortality occurred within 96 h following treatment. Median lethal concentration (LC50) values for each species and experimental site ranged from 2.79 to 3.29 mg AE/L, while median effective concentration (EC50) values (abnormal avoidance response) ranged from 1.67 to 3.84 mg AE/L. The LC10 and EC10 endpoints approximated aqueous concentrations (0.59 mg AE/L) expected under direct aerial overspray scenarios, indicating a potential risk of impacts for a small proportion of native amphibian larvae. However, given the low frequency and limited use of this herbicide formulation in Canadian forestry, these risks are considered negligible. Changes in usage patterns would require concurrent chemical and biological monitoring of operational spray programs to accurately quantify the probability and magnitude of real-world exposures and to relate these exposure levels to concentration-response relationships including those described in this study.

Physiological and biochemical characterization of quinclorac resistance in a false cleavers (Galium spurium L.) biotype.

The physiological and biochemical basis for quinclorac resistance in a false cleavers (Galium spurium L.) biotype was investigated. There was no difference between herbicide resistant (R) and susceptible (S) false cleavers biotypes in response to 2,4-D, clopyralid, glyphosate, glufosinate-ammonium, or bentazon. On the basis of GR(50) (growth reduction of 50%) or LD(50) (lethal dose to 50% of tested plants) values, the R biotype was highly resistant to the acetolactate synthase (ALS) inhibitor, thifensulfuron-methyl (GR(50) resistance ratio R/S = 57), and quinolinecarboxylic acids (quinclorac R/S = 46), resistant to MCPA (R/S = 12), and moderately resistant to the auxinic herbicides picloram (R/S = 3), dicamba (R/S = 3), fluroxypyr (R/S = 3), and triclopyr (R/S = 2). The mechanism of quinclorac resistance was not due to differences in [(14)C]quinclorac absorption, translocation, root exudation, or metabolism. Seventy-two hours after root application of quinclorac, ethylene increased ca. 3-fold in S but not R plants when compared to controls, while ABA increased ca. 14-fold in S as opposed to ca. 3-fold in R plants suggesting an alteration in the auxin signal transduction pathway, or altered target site causes resistance in false cleavers. The R false cleavers biotype may be an excellent model system to further examine the auxin signal transduction pathway and the mechanism of quinclorac and auxinic herbicide action.

Influence of herbicide-resistant canola on the environmental impact of weed management.

The growth of herbicide-resistant canola varieties increased from 10% of the canola area in Canada in 1996, when the technology was first introduced, to 80% in 2000. From 1995 to 2000, the amount of herbicide active ingredient applied per hectare of canola declined by 42.8% and the Environmental Impact (EI) per hectare, calculated using the Environmental Impact Quotient for individual herbicides and the amounts of active ingredients applied, declined 36.8%. The amount of herbicide active ingredient per hectare applied to conventional canola was consistently higher than that applied to herbicide-resistant canola each year between 1996 and 2000. Similarly, the EI of herbicide use per hectare in conventional canola was higher than that of herbicide-resistant canola during the same time period. Since 1996, herbicide use has shifted from broadcast applications of soil-active herbicides to post-emergence applications of herbicides with broad-spectrum foliar activity. The decline in herbicide use and EI since the introduction of herbicide-resistant varieties was due to increased use of chemicals with lower application rates, a reduced number of applications and a decreased need for herbicide combinations.

Buffer zone and windbreak effects on spray drift deposition in a simulated wetland.

The amount of agricultural spray that drifts into a wetland from an adjacent crop field is influenced by vegetation along the field boundary or any intentional setback distance (buffer zone) between the sprayer and the edge of the arable field. In this study, spray tracer drift deposits were measured in a simulated wetland area under different conditions of wind speed and buffer zone width. The effect of an artificial windbreak at the upwind edge of the simulated wetland was also evaluated. A level of tolerance of 0.1% of the in-swath spray deposition was established as a no-effect level for the response of aquatic plants to common herbicides. Our results indicate that a vegetated 10-m field margin (eg a fencerow) alone provides adequate protection from herbicide drift into a wetland area under wind conditions normally considered acceptable for spraying. For high winds (> 4m s(-1)) when field spraying would not normally be advised, adequate protection was afforded by the same 10-m margin plus a dense windbreak (25% porosity) or by the margin plus a 20-m buffer zone.

A comparison of two factorial designs, a complete 3 x 3 factorial and a central composite rotatable design, for use in binomial response experiments in aquatic toxicology.

Using an amphibian toxicity testing protocol, comparative studies were conducted to assess the predictive precision, degree of similarity of results and efficiency of a central composite rotatable design (CCRD) in relation to a conventional complete 3x3 factorial design. Data were treated with response surface analysis using generalized linear models (glm) and application of profile deviance to generate confidence intervals. Design comparisons were based on studies conducted using the Frog Embryo Teratogenesis Assay-Xenopus (FETAX) protocol to examine the interaction of three toxicants at pH levels ranging from 4.5 to 8.5. Test substances included two commercial herbicide formulations based on glyphosate ([N-phosphonomethyl]glycine) isopropylamine salt (Vision, Rodeo) as well as a polyethoxylated tallow amine surfactant blend (MON 0818), which is a key component of the Vision formulation. The generated models from both the CCRD and the factorial designs produced toxicity estimates similar to each other and to previously published results. Trends were also similar to published results in that the surfactant, MON 0818, was comparatively more toxic than Vision, which was more toxic than Rodeo. Further, all toxicants induced higher mortality under alkaline as opposed to acidic conditions. The CCRD was between 66 and 124% more efficient in the Vision and Rodeo experiments in comparison to the complete 3x3 factorial. Thus, the factorial experiment would have required at least 66% more observations to obtain the same precision. There appeared to be no efficiency gain with the use of the CCRD for MON 0818. The CCRD had tighter confidence intervals in 9 of 12 cases across all test substances. Computer simulations using the experimental data for Vision demonstrated that the LCalpha estimates generated from the 1,000 simulated data sets were very close to the "true" value for both designs. This was based on small bias and mean squared error calculations. Coverage probabilities in both designs were between 91.6 and 95.6%, close to the nominal coverage of 95%. Efficiency comparisons of the simulated Vision data sets demonstrated that the CCRD was more efficient in 93.3% of the comparisons. We suggest that a CCRD using glm and profile deviance for statistical analyses be considered an effective and efficient alternative in toxicity studies where interactive effects are of interest.

Comparative effects of pH and Vision herbicide on two life stages of four anuran amphibian species.

Vision, a glyphosate-based herbicide containing a 15% (weight:weight) polyethoxylated tallow amine surfactant blend, and the concurrent factor of pH were tested to determine their interactive effects on early life-stage anurans. Ninety-six-hour laboratory static renewal studies, using the embryonic and larval life stages (Gosner 25) of Rana clamitans, R. pipiens, Bufo americanus, and Xenopus laevis, were performed under a central composite rotatable design. Mortality and the prevalence of malformations were modeled using generalized linear models with a profile deviance approach for obtaining confidence intervals. There was a significant (p < 0.05) interaction of pH with Vision concentration in all eight models, such that the toxicity of Vision was amplified by elevated pH. The surfactant is the major toxic component of Vision and is hypothesized, in this study, to be the source of the pH interaction. Larvae of B. americanus and R. clamitans were 1.5 to 3.8 times more sensitive than their corresponding embryos, whereas X. laevis and R. pipiens larvae were 6.8 to 8.9 times more sensitive. At pH values above 7.5, the Vision concentrations expected to kill 50% of the test larvae in 96-h (96-h lethal concentration [LC50]) were predicted to be below the expected environmental concentration (EEC) as calculated by Canadian regulatory authorities. The EEC value represents a worst-case scenario for aerial Vision application and is calculated assuming an application of the maximum label rate (2.1 kg acid equivalents [a.e.]/ha) into a pond 15 cm in depth. The EEC of 1.4 mg a.e./L (4.5 mg/L Vision) was not exceeded by 96-h LC50 values for the embryo test. The larvae of the four species were comparable in sensitivity. Field studies should be completed using the more sensitive larval life stage to test for Vision toxicity at actual environmental concentrations.

Effects of Vision herbicide on mortality, avoidance response, and growth of amphibian larvae in two forest wetlands.

The effects of Vision (glyphosate, 356 mg acid equivalents (a.e.)/L) on mortality, avoidance response, and growth of larval amphibians (Rana clamitans and Rana pipiens) were investigated using in situ enclosures deployed in two forest wetlands of northern Ontario, Canada. In addition to untreated controls, Vision was applied to yield initial concentrations ranging from 0.29 to 14.3 mg a.e./L (0.94-46.1 mg/L of Vision). Resultant 96-h median lethal concentration (LC50) values ranged from 2.70 to 11.5 mg a.e./L (8.71-37.1 mg/L of Vision) depending on the species or site involved. Substantial mortality and incidences of abnormal avoidance response occurred only at concentrations exceeding the expected environmental concentrations (EEC) (1.43 mg a.e./L, or 4.61 mg/L of Vision) as calculated by Canadian regulatory authorities. The concentration dependence of larval growth rate and maximum size varied depending on site and species. Mean growth rates and maximum sizes exposed to 1.43 mg a.e./L (EEC) treatments were the same or greater than controls. Experimental site and biotic/abiotic factors therein, such as pH and suspended sediments, substantially affected the expression of Vision herbicide toxicity in the amphibian larvae tested. Overall, results suggest that the silvicultural use of Vision herbicide in accordance with the product label and standard Canadian environmental regulations should have negligible adverse effects on sensitive larval life stages of native amphibians.

Chemical and biomonitoring to assess potential acute effects of Vision herbicide on native amphibian larvae in forest wetlands.

In conjunction with operational forest herbicide spray programs in Ontario, Canada, chemical and biological monitoring studies were conducted in 51 different wetlands to quantify the probability and magnitude of contamination by a glyphosate herbicide formulation (Vision). Wetlands were classified as oversprayed, adjacent, or buffered in relation to the operational target spray blocks. Results show that vegetated buffers significantly mitigated against exposure and thus potential for acute effects. Aqueous concentrations of glyphosate in buffered wetlands were below analytical limits of quantitation (0.02 mg acid equivalent [a.e.]/L) in 14 of 16 cases, with mean concentration (0.03 +/- 0.02 mg a.e./L) significantly (p < 0.05) less than that of either adjacent (0.18 +/- 0.06 mg a.e./L) or oversprayed wetlands (0.33 +/- 0.11 mg a.e./L). Biomonitoring with caged amphibian larvae showed no significant differences among mean mortality (48 h) of either Rana pipiens (p = 0.194) or Rana clamitans larvae (p = 0.129) exposed in situ to Vision under these various wetland conditions. Percent mortality was not significantly (p = 0.05) correlated with exposure concentrations for either amphibian species tested. Results suggest that exposures typically occurring in forest wetlands are insufficient to induce significant acute mortality in native amphibian larvae.

Effect of pH and Release on two life stages of four anuran amphibians.

Using three native Canadian and one exotic anuran species, the interactive toxicity of pH and the forestry used-herbicide Release (triclopyr [3,5,6-trichloro-2-pyridl-oxyacetic acid]) was assessed. Embryonic and larval (Gosner 25) stages of Rana pipiens, Rana clamitans, Bufo americanus, and Xenopus laevis were exposed to treatments for at least 96 h in a static-renewal system using a central composite rotatable design. Mortality and the prevalence of malformations were modeled using generalized linear models with a profile deviance approach to obtain confidence intervals. Consistent trends of greater toxicity with lower pH were observed, with the majority of models (five of seven models) showing significant (p < 0.05) inverse relations. Larval lethal concentration estimates were eight to twenty-three times less than those observed for embryos, indicating that the larval stages were more sensitive to treatments. Further, the median lethal concentration (LC50) values for the larvae were below the expected environmental concentration (EEC) as calculated by Canadian regulatory authorities for Release. Species sensitivity was similar, with an average larval 96-h LC50 of 0.89 mg acid equivalents (AE)/L at pH 5.5 and 1.6 mg AE/L at pH 7, suggesting that X. laevis is a reasonable surrogate for native amphibians in laboratory toxicity testing. For the embryo tests, R. pipiens were slightly less sensitive in comparison with the other three species. Based on a hazard quotient analysis (EEC/LC50 > 1) for the most sensitive larval life stages, higher tier ecotoxicological testing under more realistic environmental conditions is strongly recommended.