Glutathione transferases and herbicide detoxification in suspension-cultured cells of giant foxtail (Setaria faberi)

Glutathione transferases (GSTs) catalysing the conjugation of 1-chloro-2,4-dinitrobenzene, the chloro-s-triazine herbicide atrazine, the chloroacetanilide herbicides metolachlor and alachlor and the diphenyl ether herbicide fluorodifen have been identified in suspension-cultured cells derived from the grass weed giant foxtail (Setaria faberi Herrm.). In contrast to suspension-cultured cells of maize, where atrazine-conjugating GSTs are lost during de-differentiation, the GSTs active toward this herbicide in S. faberi plants were also expressed in cultures, suggesting that these isoenzymes are subject to different regulation in the crop and weed. As a result, glutathione conjugation was the major route of atrazine metabolism in S. faberi cultures. Activities of these GSTs were maximal three days after sub-culturing when the cells were dividing most actively, when they were determined to be in the order CDNB>alachlor>metolachlor= fluorodifen>atrazine. This indicated that GSTs which are enhanced during cell division can metabolise herbicides. On the basis of activity per mg protein, GST activities in the cultures were between 20 and 60-fold higher than those determined in the foliage of S. faberi seedlings. The GSTs with activity towards CDNB were resolved into three peaks following anion-exchange chromatography at pH 7·8 using Q-Sepharose. Peak 1 GSTs were not retained, while peak 2 and peak 3 were sequentially resolved with an increasing concentration of salt. Peak 1 GSTs showed activity toward metolachlor and atrazine but showed little activity toward fluorodifen. Peak 2 and peak 3 GSTs were active toward atrazine and metolachlor, with peak 3 being particularly associated with activity toward fluorodifen. The GSTs in these peaks were then further purified using S-hexyl-glutathione-agarose affinity chromatography. In each case, the affinity-bound fraction of the GSTs consisted of 28 kDa and 26 kDa polypeptides, suggesting that the GST isoenzymes in S. faberi cultures are composed of related subunits. Our results demonstrate that the GST isoenzymes involved in herbicide metabolism in suspension cultures of a grass weed show a similar level of complexity to that determined in maize cell cultures. © 1998 SCI     

Glutathione Transferase Activities and Herbicide Selectivity in Maize and Associated Weed Species

The role of glutathione transferases (GSTs) in the selectivity of the herbicides alachlor, atrazine, fluorodifen and metolachlor, which are detoxified by glutathione conjugation in plants, was determined in seedlings of maize (Zeamays L.) and the associated weed species Abutilon theophrasti Medic., Digitariasanguinalis (L.) Scop. Echinochloa crus-galli (L.) Beauv., Panicum miliaceum (L.), Setaria faberi Herrm. and Sorghum bicolor (L.) Moench. The availability of glutathione was also determined in all species and tissue concentrations were found to be in the range 120–160 μm for all species except D. sanguinalis and S. bicolor, which contained half this amount. GST activities toward the herbicides were determined in crude protein extracts from the plants using HPLC to quantify the biosynthesis of the herbicide conjugates. The specific activities of the GSTs toward the substrates were in the order alachlor>fluorodifen> atrazine>metolachlor in all species except A. theophrasti, where fluorodifen was a better substrate than alachlor. In most cases there was a good correlation between GST activities and the selectivity of the herbicides applied pre-emergence. In the case of atrazine, GST activities were also related to the relative rates of herbicide conjugation in vivo. In contrast, there was no simple relationship between glutathione availability and the selectivity of the herbicides. However, with alachlor there was evidence that glutathione availability was limiting GST activity and influencing tolerance.     

Effects of an oil-based substrate (The Water Cleanser™) and bacterial additives on nitrogen and phosphorous dynamics in freshwater crayfish (Cherax cainii,Austin and Ryan 2002) aquaculture

Two experiments were conducted to evaluate the efficacy of an oil-based substrate, The Water Cleanser? (TWC). The first experiment studied the effects two substrates of different oil composition (TWC, TWC?+), and a commercial bacterial additive (Bio-Aid) on concentrations of nitrogen and phosphorous in indoor aquaria. The second experiment studied the effects of TWC, a bacterial additive, and a combination (TWC?+?B) on concentrations of nitrogen and phosphorous, phytoplankton abundance and diversity in outdoor freshwater crayfish (Cherax cainii, Austin and Ryan, Invertebr Syst 16:357–367, 2002) tanks. In the first experiment, the concentration of TAN decreased more rapidly with Bio-Aid, whilst the maximum concentrations of NO₂-N and NO₃-N were reduced with the substrates. The concentration of orthophosphate was reduced in aquaria with TWC?+?. In the second experiment, concentrations of TAN, NO₂-N, NO₃-N and total phosphate were not significantly affected by TWC. After addition of TWC?+?B, there was a significant decrease in the concentrations of nitrate and total phosphate, and a higher abundance of phytoplankton was maintained than with other treatments. Additionally, a population Bacillus sp. was found on the substrate surface. TWC had no adverse effects on phytoplankton abundance or diversity, or C. cainii physiology, weight gain or survival. TWC and TWC?+?were effective bioremediators of eutrophic water, whilst a combination of TWC?+?B was effective in short term bioremediation and in promoting phytoplankton abundance in C. cainii tank culture.