Target-site basis for resistance to acetolactate synthase inhibitor in Water chickweed (Myosoton aquaticum L.)

Water chickweed is a widespread and competitive winter annual or biennial weed of wheat in China. One Water chickweed population (HN02) resistant to several acetolactate synthase (ALS) inhibitors was found in Henan province of China. Whole-plant bioassays showed that HN02 was high resistance to tribenuron (292.05-flod). In vitro ALS assays revealed that resistance was due to reduced sensitivity of the ALS enzyme to tribenuron. The I₅₀ value for HN02 was 85.53 times greater respectively than that of susceptible population (SD05). This altered ALS sensitivity in the resistant population was due to a mutation in the ALS gene resulting in a Pro197 to Ser substitution. Cross-resistance experiments indicated that HN02 exhibited various resistance patterns to pyrithiobac-sodium, florasulam and pyroxsulam, without resistance to imazethapyr. This is the first report of tribenuron-resistant Water chickweed in Henan province of China, target-site based resistance was established as being due to an insensitive form of ALS, resulting from a Pro to Ser substitution at amino acid position 197 in the ALS gene.     

山东省部分市县麦田杂草麦家公Lithospermum arvense对苯磺隆的抗药性

采用温室盆栽法和培养皿法测定了山东省部分市县冬小麦田杂草麦家公Lithospermum arvense L.对苯磺隆的抗药性水平,以及其抗药性生物型乙酰乳酸合成酶(ALS)对苯磺隆的敏感性。温室盆栽结果显示,供试杂草对苯磺隆产生了不同程度的抗药性,其中胶州麦家公生物型抗性水平最高,抗性倍数为12.8倍;培养皿法测定结果也显示胶州麦家公生物型抗性水平最高,但抗性倍数为3.89倍。交互抗性测定结果表明,胶州抗性麦家公生物型对其他ALS抑制剂噻吩磺隆和苄嘧磺隆已产生不同程度的交互抗性,其中对噻吩磺隆的抗性倍数达到3.11倍。离体条件下,与敏感生物型ALS活力的抑制中浓度(IC50)相比较,胶州抗性麦家公生物型的IC50值是敏感麦家公的 2.65倍。表明ALS敏感性降低可能是山东部分市县麦家公对苯磺隆产生抗药性的重要原因之一。     

Target-site mutation associated with glufosinate resistance in Italian ryegrass (Lolium perenne L. ssp. multiflorum)

BACKGROUND: Studies were conducted to elucidate the mechanism of glufosinate resistance in an Italian ryegrass population. RESULTS: Glufosinate rates required to reduce growth by 50% (GR₅₀) were 0.15 and 0.18 kg AI ha^?1 for two susceptible populations C1 and C2 respectively, and 0.45 kg AI ha^?1 for the resistant population MG, resulting in a resistance index of 2.8. Ammonia accumulation after glufosinate treatment was on average 1.5 times less for the resistant population than for the susceptible populations. The glufosinate concentrations (µM ) required to reduce the glutamine synthetase (GS) enzyme activity by 50% (I₅₀) were 31 and 137 for C1 and C2 respectively, and 2432 for the resistant population MG. One amino acid substitution in the plastidic GS2 gene, aspartic acid for asparagine at position 171, was identified in the resistant population. CONCLUSIONS: This is the first report of glufosinate resistance in a weed species that involves an altered target site. Copyright © 2012 Society of Chemical Industry     

A European biotype of Amaranthus retroflexus cross-resistant to ALS inhibitors and response to alternative herbicides

An acetolactate synthase (ALS)‐resistant Amaranthus retroflexus biotype was collected in a soyabean crop after repeated exposure to imazethapyr and thifensulfuron‐methyl in north‐eastern Italy. Studies were conducted to characterise the resistance status and determine alternative post‐emergence herbicides for controlling this biotype. Whole‐plant bioassay revealed that the GR₅₀ values were 1898‐ and 293‐fold higher than those observed for the biotype susceptible to imazethapyr and imazamox respectively. The biotype also displayed high cross‐resistance to sulfonylureas. Molecular analysis demonstrated that a single nucleotide substitution had occurred in domain B (TGG to TTG at position 574), conferring a change from the amino acid tryptophan to leucine in the resistant biotype. However, herbicides with other modes of action (PSII, 4‐HPPD and PPO inhibitors) provided excellent control. The GR₅₀ ratios for metribuzin, terbuthylazine and mesotrione were close to 1 and treatments with fomesafen gave 100% control of both susceptible and resistant biotypes at the recommended field dose. This study documents the first case of an imidazolinone and ALS‐resistant biotype in European crops and identifies the post‐emergence herbicide options available for managing this troublesome weed in soyabean crops. Alternative management strategies are also discussed.     

苏云金杆菌对抗性及敏感小菜蛾的拒食活性

调查了苏云金杆菌对抗性和敏感小菜蛾的拒食活性。选择性和完全拒食试验表明,Bt对抗性和敏感小菜蛾种群都有一定的拒食作用,且拒食活性随Bt浓度的增加而显著增强。相对于抗性种群,小菜蛾敏感种群对Bt杀虫剂表现了更强的拒食趋势,其拒食中浓度AFC50(0.089 3 μg/mL)显著低于抗性种群(0.148 1 μg/mL)。试验结果表明,抗性种群对有毒叶片的行为避害能力弱于敏感种群,小菜蛾对Bt杀虫剂的抗性产生应与其行为机制无关。     

Resistance of dicot weeds to acetolactate synthase (ALS)-inhibiting herbicides in Australia

A biotype of Sonchus oleraceus L. and two bio types of Sisymbrium orientate Torn., SSO 3 and NSO 1, are the first dicot weeds in Australia to develop resistance to ALS-inhibiting herbicides. The resistant biotypes had been exposed to va rying periods of selection with sulfonylurea her bicides. All three biotypes are resistant to a range of sulfonylurea and imidazolinone herbicides. The S. orientale biotypes are also resistant to the triazolopyrimidine herbicide, flumetsulam. LD₅₀ ratios of resistant Sonchus oleraceus for sulfony lurea and imidazolinone herbicides are greater than 64-fold and 4.5-fold, respectively. GR₅₀ ratios are greater than 9 for sulfonylureas and 7.4 for imazapyr. The LD₅₀ ratios for both S. orien tale biotypes for chlorsulfuron, sulfometuron methyl, metsulfuron-methyl, flumetsulam and imazethapyr are greater than 110-, 15-, 7-, 24- and 29-fold, respectively. All resistant biotypes are susceptible to MCPA, diuron and diflufenican, herbicides which do not inhibit ALS.     

稻稗HJHL-715种群对五氟磺草胺的抗药性水平及抗性机理分析

为明确东北稻区稻稗Echinochloa oryzoides HJHL-715种群对五氟磺草胺的抗性水平及抗性机制,采用整株生物测定法测定稻稗种群对五氟磺草胺的敏感性,明确抗性种群的交互抗性和多抗性情况,研究3种细胞色素P450抑制剂对其敏感性的影响;并应用分子生物学方法进行稻稗的乙酰乳酸合酶(acetolactate synthase,ALS)离体活性测定、ALS基因序列分析及其表达量测定。结果表明:在东北稻区,五氟磺草胺对稻稗HJHL-715种群鲜重的抑制中剂量GR50为62.53 g/hm^2;稻稗HJHL-715的ALS基因序列中未发现氨基酸突变,其ALS离体活性与敏感种群的ALS离体活性无显著性差异,ALS基因表达量显著低于敏感种群。1-氨基苯并三唑(1-aminobenzotriazole,ABT)、胡椒基丁醚(piperomyl butoxide,PBO)、马拉硫磷3种P450抑制剂显著提高了稻稗HJHL-715种群对五氟磺草胺的敏感性,使其对五氟磺草胺的GR50由原来的62.53 g/hm^2分别下降到5.78、5.02、3.53 g/hm^2。表明东北稻区已经出现了对五氟磺草胺具有高水平抗性的稻稗种群,稻稗HJHL-715种群对五氟磺草胺的抗性很可能是由细胞色素P450介导的代谢增强所致。     

中国北方部分地区麦田荠菜对苯磺隆的抗性水平

为明确北方地区冬小麦田荠菜对苯磺隆的抗性水平,运用培养皿法和温室盆栽法分别测定了山东、山西、河南、河北、陕西5省13个地区采集点麦田潜在抗药性生物型对苯磺隆的抗性水平,并分别测定了驻马店梁祝镇采集点荠菜潜在抗药性生物型和敏感生物型乙酰乳酸合成酶(ALS)对苯磺隆的敏感性。培养皿法测定结果表明:驻马店梁祝镇采集点荠菜抗药性生物型对苯磺隆的抗性水平最高,抗性倍数为6.17倍,其他采集点荠菜抗性倍数在0.94～2.04倍之间,仍处于较为敏感状态。温室盆栽法测定结果表明:驻马店梁祝镇采集点荠菜抗性倍数仍为最高,达到233倍, 其他地区采集点荠菜抗性倍数在1.23～3.73倍之间,尚未产生明显的抗药性。离体条件下,苯磺隆对荠菜抗药性和敏感生物型ALS的抑制中浓度(IC50)分别为0.664 μmol/L和0.053 3 μmol/L,抗药性生物型的抗性倍数达12.5倍。结果表明,驻马店梁祝镇采集点荠菜已对苯磺隆产生了较高水平的抗药性,而其体内ALS敏感性降低可能是抗药性产生的原因之一。     

Glyphosate resistance in common ragweed ( A mbrosia artemisiifolia L.) from Mississippi,USA

Glyphosate is one of the most commonly used broad‐spectrum herbicides over the last 40 years. Due to the widespread adoption of glyphosate‐resistant (GR) crop technology, especially corn, cotton and soybean, several weed species have evolved resistance to this herbicide. Research was conducted to confirm and characterize the magnitude and mechanism of glyphosate resistance in two GR common ragweed ( A mbrosia artemisiifolia L.) biotypes from Mississippi, USA. A glyphosate‐susceptible (GS) biotype was included for comparison. The effective glyphosate dose to reduce the growth of the treated plants by 50% for the GR1, GR2 and GS biotypes was 0.58, 0.46 and 0.11 kg ae ha^?1, respectively, indicating that the level of resistance was five and fourfold that of the GS biotype for GR1 and GR2, respectively. Studies using ¹⁴ C‐glyphosate have not indicated any difference in its absorption between the biotypes, but the GR1 and GR2 biotypes translocated more ¹⁴ C‐glyphosate, compared to the GS biotype. This difference in translocation within resistant biotypes is unique. There was no amino acid substitution at codon 106 that was detected by the 5‐enolpyruvylshikimate‐3‐phosphate synthase gene sequence analysis of the resistant and susceptible biotypes. Therefore, the mechanism of resistance to glyphosate in common ragweed biotypes from Mississippi is not related to a target site mutation or reduced absorption and/or translocation of glyphosate.     

我国南方稻区稗草对二氯喹啉酸的抗药性测定

用琼脂法对2000年和2001年采收的29个稗草生态型对二氯喹啉酸的抗性水平进行了测定。结果表明:广东花都稗草对二氯喹啉酸最为敏感,其抑制中浓度IC50是0.148 0 mg/L;湖南安乡稗草对二氯喹啉酸的抗性极为明显,2000年和2001年样本IC50值分别是7.458和13.80 mg/L,其相对抗性比分别为50.4和93.2;2000年采收的湖南常德稗草也对二氯喹啉酸表现出高抗,其相对抗性比值是4.47,黄梅、汉寿、常德(b)、高桥4地稗草也表现出抗性,其相对抗性比值分别为2.37、3.12、2.44和2.20;其他22个稗草生态型对二氯喹啉酸仍未表现出抗性。     

Possible involvement of herbicide sequestration in the resistance to diclofop-methyl in Italian biotypes of Lolium spp.

The mechanism of resistance to diclofop-methyl in three Italian populations of Lolium spp. (two resistant and one susceptible) was investigated. The major proportion of R-1 (Tuscania 1997) and R-2 (Roma 1994) plants (approximately 80%) survived after herbicide treatment by emitting new tillers from the crown. Both resistant (R-1 and R-2) and susceptible (Vetralla 1994) Lolium spp. populations were target-site sensitive. No difference in diclofop-methyl absorption by shoots of resistant and susceptible biotypes was observed. At the dose corresponding to 1× the recommended field rate, a relatively higher metabolism was found in R-2 biotype. In contrast, at the doses 2× and 10× the field rate no difference in herbicide metabolism between susceptible and resistant biotypes was observed. At all the three herbicide doses (1×, 2×, and 10× the field rate) 48 h after the treatment (HAT), the total amount of metabolites produced by wheat was more than three times higher than that produced by resistant and susceptible ryegrass biotypes. At the doses 1× and 2× the field rate, the herbicide translocation was different in the susceptible biotypes compared to resistant biotypes. The total amount of the radiolabel found 48 HAT in culm and root was approximately twice in susceptible biotype than in resistant biotypes. Susceptible and resistant ryegrass biotypes differed in the capability of their roots to acidify the external medium. Susceptible biotype acidified the external solution at approximately 6 times the rates of the resistant biotypes. In the present study, the mechanism responsible for resistance in the investigated resistant biotypes was not univocally identified. Indirect evidence supports the possible involvement of herbicide sequestration or immobilization.     

Molecular basis of diverse responses to acetolactate synthase-inhibiting herbicides in sulfonylurea-resistant biotypes of Schoenoplectus juncoides

Sulfonylurea-resistant biotypes of Schoenoplectus juncoides were collected from Nakafurano, Shiwa, Matsuyama, and Yurihonjyo in Japan. All of the four biotypes showed resistance to bensulfuron-methyl and thifensulfuron-methyl in whole-plant experiments. The growth of the Nakafurano, Shiwa, and Matsuyama biotypes was inhibited by imazaquin-ammonium and bispyribac-sodium, whereas the Yurihonjyo biotype grew normally after treatment with these herbicides. The herbicide concentration required to inhibit the acetolactate synthase (ALS) enzyme by 50% (I₅₀), obtained using in vivo ALS assays, indicated that the four biotypes were > 10-fold more resistant to thifensulfuron-methyl than a susceptible biotype. The Nakafurano, Shiwa, and Matsuyama biotypes exhibited no or little resistance to imazaquin-ammonium, whereas the Yurihonjyo biotype exhibited 6700-fold resistance to the herbicide. The Nakafurano and Shiwa biotypes exhibited no resistance to bispyribac-sodium, but the Matsuyama biotype exhibited 21-fold resistance and the Yurihonjyo biotype exhibited 260-fold resistance to the herbicide. Two S. juncoides ALS genes (ALS1 and ALS2) were isolated and each was found to contain one intron and to encode an ALS protein of 645 amino acids. Sequencing of the ALS genes revealed an amino acid substitution at Pro₁₉₇ in either encoded protein (ALS1 or ALS2) in the biotypes from Nakafurano (Pro₁₉₇ → Ser₁₉₇), Shiwa (Pro₁₉₇ → His₁₉₇), and Matsuyama (Pro₁₉₇ → Leu₁₉₇). The ALS2 of the biotype from Yurihonjyo was found to contain a Trp₅₇₄ → Leu₅₇₄ substitution. The relationships between the responses to ALS-inhibiting herbicides and the amino acid substitutions, which are consistent with previous reports in other plants, indicate that the substitutions at Pro₁₉₇ and Trp₅₇₄ are the basis of the resistance to sulfonylureas in these S. juncoides biotypes.     

Assessment of two biotypes of Solanum ptycanthum that differ in resistance levels to imazamox

Glasshouse and laboratory experiments were conducted on acetolactate synthase (ALS) homozygous resistant Solanum ptycanthum biotypes from Illinois (IL‐R) and Indiana (IN‐R), and homozygous susceptible biotypes from Illinois (IL‐S) and Indiana (IN‐S). Genetic similarity of biotypes was assessed by random amplified polymorphic DNA (RAPD) markers, which determined that the Illinois biotypes are more similar to each other than to the IN‐R biotype. ALS enzyme activity from the IL‐R and IN‐R biotypes had I₅₀ values of 362 and 352 μM imazamox respectively. Dose–response experiments using three‐ to four‐leaf‐stage plants of the IL‐R and IN‐R biotypes had GR₅₀ values of 242 and 69 g ae ha⁻¹ imazamox respectively. Whole‐plant and ALS enzyme results are different than previously reported values in the literature, which was attributed in the current study to the original IN‐R population having individuals that were segregating for ALS resistance. Metabolism studies showed no difference in percentage [¹⁴C]imazamox remaining between the IL‐R and IN‐R biotypes up to 72 h after treatment. The IL‐S biotype metabolised [¹⁴C]imazamox approximately two times faster than the IL‐R and IN‐R biotypes and this trait was heritable. Response of F₃ plants containing homozygous ALS‐resistant alleles from the IL‐R biotype in a genetic background of 50% Illinois and 50% Indiana biotypes suggests that genetic factors other than an altered target site or metabolism may also contribute to the magnitude of resistance at the whole‐plant level in resistant biotypes.     

Molecular basis of resistance to tribenuron-methyl in Descurainia Sophia (L.) populations from China

Two Descurainia Sophia populations, HB16 and HB08, were discovered in China and exhibit high resistance levels to tribenuron-methyl. Resistance ratio values of HB16 and HB08 were 153.47 and 651.20, respectively. The extractable acetohydroxy acid synthase (AHAS) activity was similar between resistant and susceptible populations. However, AHAS from HB16 and HB08 was less sensitive to inhibition of tribenuron-methyl comparing to susceptible population. The tribenuron-methyl I₅₀ values for HB16 and HB08 were 46.25 and 54.07 greater than that for susceptible population respectively. Pro-197-Ser and Pro-197-Leu mutations were identified in AHAS extracted from HB16 and HB08 plants respectively. AHAS insensitivity of resistant D. Sophia caused by Pro197 mutation could be responsible for high resistance to tribenuron-methyl.     

Molecular basis of resistance to acetolactate synthase-inhibiting herbicides in Sisymbrium orientale and Brassica tournefortii

Three Australian Sisymbrium orientale and one Brassica tournefortii biotypes are resistant to acetolactate synthase (ALS)-inhibiting herbicides due to their possession of an ALS enzyme with decreased sensitivity to these herbicides. Enzyme kinetic studies revealed no interbiotypic differences within species in K_m (pyruvate) (the substrate concentration at which the reaction rate is half maximal) but a greater V_max (the rate when the enzyme is fully saturated with substrate) for two of the resistant S orientale biotypes over susceptible levels. F₁ hybrids from reciprocal crosses between resistant and susceptible biotypes of S orientale showed an intermediate response to chlorsulfuron compared to the parental plants. ALS herbicide resistance in S orientale segregated in a 3:1 (resistant:susceptible) ratio in F₂ plants with a single rate of chlorsulfuron, indicating that resistance is inherited as a single, incompletely dominant nuclear gene. Two regions of the ALS structural gene known to vary in ALS-resistant biotypes were amplified and sequenced. Resistant S orientale biotypes NS01 and SS03 contained a single nucleotide substitution in Domain B, predicting a Trp (in susceptible) to Leu (in resistant) amino acid change. Two adjacent nucleotide substitutions (CC T to AT T) predicting a Pro (in susceptible) to Ile (in resistant) change in the primary amino acid sequence were identified in Domain A of resistant S orientale biotype SS01. Likewise, a single nucleotide substitution at the same site in the resistant B tournefortii biotype predicts a Pro (in susceptible) to Ala (in resistant) substitution. No other interbiotypic nucleotide differences predicted amino acid changes in the sequenced regions, suggesting that the amino acid substitutions reported above are responsible for resistance to ALS-inhibiting herbicides in the respective biotypes. © 1999 Society of Chemical Industry     

广东省水稻田稗对五氟磺草胺的抗性分析

为明确广东省水稻田杂草稗Echinochloa crus-galli对五氟磺草胺的抗性现状及其可能的抗性机理,采用整株剂量反应法测定不同地区稗种群对五氟磺草胺的抗性水平,对不同稗种群的乙酰乳酸合成酶（acetolactate synthase,ALS）基因片段进行扩增测序,分析细胞色素P450酶（cytochrome P450 monooxygenase,P450）和谷胱甘肽-S-转移酶（glutathione-S-transferase,GST）抑制剂胡椒基丁醚（piperonylbutoxide,PBO）和4-氯-7-硝基-2,1,3-苯并氧杂噁二唑（4-chloro-7-nitro-1,2,3-benzoxadiazole,NBD-Cl）对不同稗种群抗性水平的影响,并对替代药剂进行筛选。结果显示,广东省水稻田多数稗种群对五氟磺草胺仍表现敏感,但采自湛江市的1个种群BC-7对五氟磺草胺产生了抗性,抗性倍数达6.5倍。与敏感种群BC-2相比,BC-7种群并未发生已报道的ALS靶标抗性相关突变。PBO和NBD-Cl均可显著提高BC-7种群对五氟磺草胺的敏感性,其干重抑制中量GR₅₀由31.1 g/hm²分别降为11.0 g/hm²和24.7 g/hm²。BC-7种群对氰氟草酯和噁唑酰草胺仍较敏感,但对二氯喹啉酸和双草醚产生了抗性。表明P450和GST介导的代谢抗性是稗BC-7种群产生抗性的重要原因,氰氟草酯和噁唑酰草胺适用于治理该抗性种群。     

Transformation a mutant Monochoria vaginalis acetolactate synthase (ALS) gene renders Arabidopsis thaliana resistant to ALS inhibitors

Acetolactate synthase (ALS) genes from Monochoria vaginalis resistant (R) and susceptible (S) biotypes against ALS inhibitors found in Korea revealed a single amino acid substitution of Proline (CCT), at 169th position based on the M. vaginalis ALS sequence numbering, to serine (TCT) in conserved domain A of the gene (equal to the proline 197 in Arabidopsis thaliana ALS gene sequence). A. thaliana plants transformed with the single mutated (Pro₁₆₉ to Ser) M. vaginalis ALS gene (including transit signal peptide) showed cross-resistance patterns to ALS-inhibiting herbicides, like as sulfonylurea-herbicide bensulfuron methyl (R/S factor of 9.5), imidazolinone-herbicide imazapyr (R/S factor of 5.1), and triazolopyrimidine-herbicide flumetsulam (R/S factor of 17.6) when measuring hypocotyls’ length of A. thaliana. The ALS activity from the transgenic A. thaliana plants confirmed the cross-resistance pattern to these herbicides like as R/S factor of 8.3 to bensulfuron methyl, 2.3 to imazapyr, and 13.2 to flumetsulam.     

Triazophos resistance mechanisms in the rice stem borer (Chilo suppressalis Walker)

A field population of the rice stem borer (Chilo suppressalis Walker) with 203.3-fold resistance to triazophos was collected. After 8-generation of continuous selection with triazophos in laboratory, resistance increased to 787.2-fold, and at the same time, the resistance to isocarbophos and methamidophos was also enhanced by 1.9- and 1.4-fold, respectively, implying some cross-resistance between triazophos and these two organophosphate insecticides. Resistance to abamectin was slightly enhanced by triazophos selection, and fipronil and methomyl decreased. Synergism experiments in vivo with TPP, PBO, and DEM were performed to gain a potential indication of roles of detoxicating enzymes in triazophos resistance. The synergism results revealed that TPP (SR, 1.92) and PBO (SR 1.63) had significant synergistic effects on triazophos in resistant rice borers. While DEM (SR 0.83) showed no effects. Assays of enzyme activity in vitro demonstrated that the resistant strain had higher activity of esterase and microsomal O-demethylase than the susceptible strain (1.20- and 1.30-fold, respectively). For glutathione S-transferase activity, no difference was found between the resistant and the susceptible strain when DCNB was used as substrate. However, 1.28-fold higher activity was observed in the resistant strain when CDNB was used. These results showed that esterase and microsomal-O-demethylase play some roles in the resistance. Some iso-enzyme of glutathione S-transferase may involve in the resistance to other insecticides, for this resistant strain was selected from a field population with multiple resistance background. Acetylcholinesterase as the triazophos target was also compared. The results revealed significant differences between the resistant and susceptible strain. The V_max and K_m of the enzyme in resistant strain was only 32 and 65% that in the susceptible strain, respectively. Inhibition tests in vitro showed that I₅₀ of triazophos on AChE of the resistant strain was 2.52-fold higher. Therefore, insensitive AChE may also involved in triazophos resistance mechanism of rice stem borer.     

Purification and biochemical characterization of glutathione S-transferases from three strains of Liposcelis bostrychophila Badonnel (Psocoptera: Liposcelididae): Implication of insecticide resistance

Glutathione S-transferases (GSTs) catalyzing the conjugation of reduced glutathione (GSH) to a vast range of xenobiotics including insecticides were investigated in the psocid Liposcelis bostrychophila Badonnel. GSTs from susceptible and two resistant strains (DDVP-R for dichlorvos-resistant strain and PH₃-R for phosphine-resistant strain) of L. bostrychophila were purified by glutathione-agarose affinity chromatography and characterized by their Michaelis-Menten kinetics towards artificial substrates, i.e., 1-chloro-2,4-dinitrobenzene (CDNB), in a photometric microplate assay. The specific activities of GSTs purified from two resistant strains were significantly higher than their susceptible counterpart. For the resistant strains, GSTs both showed a significantly higher affinity to the substrate GSH while a declined affinity to CDNB than those of susceptible strain. The inhibitory potential of ethacrynic acid was very effective with highest I₅₀ value (the concentration required to inhibit 50% of GSTs activity) of 1.21 μM recorded in DDVP-R. Carbosulfan also exhibited excellent inhibitory effects on purified GSTs. The N-terminus of the purified enzyme was sequenced by Edman degradation, and the alignment of first 13 amino acids of the N-terminal sequence with other insect GSTs suggested the purified protein was similar to those of Sigma class GSTs.