Biochemical characteristics of insecticide resistance in the fall armyworm, Spodoptera frugiperda (J.E. Smith)

A strain of the fall armyworm, Spodoptera frugiperda (J.E. Smith), collected from corn in Citra, Florida, showed high resistance to carbaryl (562-fold) and methyl parathion (354-fold). Biochemical studies revealed that various detoxification enzyme activities were higher in the field strain than in the susceptible strain. In larval midguts, activities of microsomal oxidases (epoxidases, hydroxylase, sulfoxidase, N-demethylase, and O-demethylase) and hydrolases (general esterase, carboxylesterase, β-glucosidase) were 1.2- to 1.9-fold higher in the field strain than in the susceptible strain. In larval fat bodies, various activities of microsomal oxidases (epoxidases, hydroxylase, N-demethylase, O-demethylases, and S-demethylase), glutathione S-transferases (CDNB, DCNB, and p-nitrophenyl acetate conjugation), hydrolases (general esterase, carboxylesterase, β-glucosidase, and carboxylamidase) and reductases (juglone reductase and cytochrome c reductase) were 1.3- to 7.7-fold higher in the field strain than in the susceptible strain. Cytochrome P450 level was 2.5-fold higher in the field strain than in the susceptible strain. In adult abdomens, their detoxification enzyme activities were generally lower than those in larval midguts or fat bodies; this is especially true when microsomal oxidases are considered. However, activities of microsomal oxidases (S-demethylase), hydrolases (general esterase and permethrin esterase) and reductases (juglone reductase and cytochrome c reductase) were 1.5- to 3.0-fold higher in the field strain than in the susceptible strain. Levels of cytochrome P450 and cytochrome b₅ were 2.1 and 1.9-fold higher, respectively, in the field strain than in the susceptible strain. In addition, acetylcholinesterase from the field strain was 2- to 85-fold less sensitive than that from the susceptible strain to inhibition by carbamates (carbaryl, propoxur, carbofuran, bendiocarb, thiodicarb) and organophosphates (methyl paraoxon, paraoxon, dichlorvos), insensitivity being highest toward carbaryl. Kinetics studies showed that the apparent K_m value for acetylcholinesterase from the field strain was 56% of that from the susceptible strain. The results indicated that the insecticide resistance observed in the field strain was due to multiple resistance mechanisms, including increased detoxification of these insecticides by microsomal oxidases, glutathione S-transferases, hydrolases and reductases, and target site insensitivity such as insensitive acetylcholinesterase. Resistance appeared to be correlated better with detoxification enzyme activities in larval fat bodies than in larval midguts, suggesting that the larval fat body is an ideal tissue source for comparing detoxification capability between insecticide-susceptible and -resistant insects.     

Age-dependent variations in the response of several species of diptera to insecticidal chemicals

The toxicity of carbaryl to three species of fleshflies Sarcophaga bullata Parker, S. crassipalpis Macquart, and S. argyrostoma (Robineau-Desvoidy) varied considerably with age and sex. In contrast, the susceptibility of a blowfly, Phormia regina (Meigen) to carbaryl decreased with age and that of two muscid flies, Musca autumnalis DeGeer and Stomoxys calcitrans (L.), remained relatively constant. The synergistic activity of piperonyl butoxide varied inversely with the innate toxicity of carbaryl to each species suggesting that the observed age- and sex-dependent variations in carbaryl toxicity result mainly from differences in detoxifying capability. This was supported by in vitro measurements of oxidative microsomal metabolism. It was further established that differences in the rates of penetration and excretion of carbaryl and in the susceptibility of the head cholinesterase to carbaryl inhibition were of little importance in determining the susceptibility of the flies to this insecticide.     

Studies on the metabolism of vamidothion and its thioanalog in insecticide-resistant and susceptible house fly strains

The in vivo and in vitro metabolism of vamidothion [O,O-dimethyl S-[2-(1-methylcarbamoyl)-ethylthio] ethylphosphorothiolate] as well as the in vitro metabolism of thiovamidothion [O,O-dimethyl S-[2-(1-methylcarbamoyl)ethylthio] ethylphosphorodithioate] was investigated in insecticide-resistant and susceptible house fly strains. Vamidothion was converted in vivo to the sulfoxide, the principle metabolite, and subsequently to the sulfone at a slower rate. Vamidothion and vamidothion sulfoxide were hydrolyzed at the PS and SC bond. The resulting primary alcohol metabolite was further oxidized to a carboxylic acid followed by decarboxylation. No metabolism of vamidothion or thiovamidothion occurred in vitro without the addition of NADPH. The addition of NADPH resulted in rapid conversion of vamidothion to the sulfoxide, and thiovamidothion was oxidatively metabolized to six metabolic products. No qualitative differences were found between resistant and susceptible strains, but there were signficant quantitative differences. The metabolism was highest in the Rutgers strain followed by Cornell-R, Hirokawa, and then CSMA strain. The route of vamidothion and thiovamidothion metabolism was via the cytochrome P-450-dependent monooxygenase system, and none of the resistant strains showed glutathione S-transferase activity toward vamidothion or thiovamidothion. No further oxidation of vamidothion sulfoxide to the sulfone was observed and also no hydrolysis products were formed, in vitro.     

Induction of microsomal enzymes in the southern armyworm (Prodenia eridania)

Levels of microsomal epoxidation, N-demethylation, and cytochrome P-450 in the gut tissues of sixth instar southern armyworm larvae were considerably enhanced following oral in vivo treatment with a series of methylbenzenes. Induction increased with increasing methyl substitution and was maximal with pentamethylbenzene. The increase in microsomal activity occurred rapidly after initiation of treatment and the final levels of induction achieved were dependent on the concentration of the inducer in the diet and the time of exposure. Microsomal enzyme activity returned to control levels following termination of exposure and induction was blocked by puromycin and cycloheximide but not by actinomycin D. The in vivo tolerance of induced worms to orally administered carbaryl was increased in a manner reflecting the enhanced microsomal enzyme activity.     

Penetration of insecticides through the foregut of the honeybee (Apis mellifera L.)

The rates of penetration of ¹⁴C-labeled insecticides (parathion, carbaryl, and dieldrin) through the foregut of the honeybee (Apis mellifera L.) were measured in vitro and in vivo. Uptake of the insecticides from the lumen of the foregut into foregut tissue was directly proportional to insecticide lipophilicity, but penetration through the foregut was not. Of the three insecticides studied, parathion appeared to possess the optimal physicochemical characteristics required for penetration. The uptake of carbaryl and release of dieldrin by the foregut tissues may limit their respective penetration rates. Insecticide penetration was found to be inversely proportional to the sucrose concentration in the lumen of the foregut in both in vitro and in vivo studies. The oral toxicity of carbaryl showed a similar dependence on the sucrose concentration of the solution in which the insecticide was fed. The data presented indicate that the honeybee foregut is permeable to lipophilic compounds and strongly suggest that this permeability may contribute substantially to the toxicity of orally ingested insecticides in this insect.     

Pyrethroid-hydrolyzing esterases in southern armyworm larvae: Tissue distribution,kinetic properties,and selective inhibition

Esterase activity hydrolyzing both [1RS,trans]- and [1RS,cis]-permethrin was detected in crude homogenates of the following southern armyworm (Spodoptera eridania Cramer) larval tissues: cuticle, gut, fat body, head capsule, Malpighian tubules, and silk gland. Neither substrate was detectably hydrolyzed by hemolymph. The highest esterase activities per insect equivalent of tissue were found in cuticle, gut, and fat body for the trans isomer and in cuticle and gut for the cis isomer. Each preparation hydrolyzed the trans isomer more rapidly, but the degree of specificity varied greatly between tissues. Differences in apparent K_m and V_max values between the three most active tissues were threefold or less for trans isomer hydrolysis, but differences between tissues of up to 100-fold were found for K_m and V_max values for cis isomer hydrolysis. Hydrolysis of the trans isomer in cuticle, gut, and fat body homogenates was only partially inhibited by α-naphthyl N-propylcarbamate (NPC). Concentrations of NPC giving maximal inhibition of trans isomer hydrolysis had little effect on the hydrolysis of the cis isomer. These results demonstrate that pyrethroid-hydrolyzing activity is broadly distributed in insect tissues and results from the combined activity of several esterases with different properties. It is likely that the trans and cis isomers of permethrin are hydrolyzed by separate enzymes in this insect.     

药剂对小菜蛾抗性及敏感品系乙酰胆碱酯酶抑制作用比较

采用浸叶法测定了云南通海、元谋和澜沧的小菜蛾plutella xylostella田间种群对常用杀虫剂的抗药性。结果表明,云南上述地区小菜蛾田间种群对各类杀虫剂均产生了不同程度的抗性。对有机磷类药剂的抗药性为1.74~31.1倍;对菊酯类药剂的抗药性为7.41~764倍;对阿维菌素类药剂则产生了 5.60~4.06×104倍的抗性。通过离体和活体试验测定了药剂对小菜蛾头部乙酰胆碱酯酶(AChE)的抑制作用。敌敌畏和灭多威对通海抗性品系AChE离体和活体内的抑制中浓度(I50)分别是敏感品系的209、26.5倍和2.21、2.16倍;敌敌畏对通海小菜蛾种群的离体和活体内抑制中时间(IT50)小于敏感品系,分别是敏感品系的0.32和0.17倍;而灭多威对通海小菜蛾种群的离体和活体内抑制中时间(IT50)则大于敏感品系,分别是敏感品系的1.37和1.74倍。     

Carboxylamidases inSpodoptera exigua: properties and distribution in the larval body

Resistance to many insecticides demonstrated by the beet armyworm,Spodoptera exigua (Hübner), can be caused by the action of carboxylamidases. A colorimetric method, based on the hydrolysis of 4-nitroacetanilide to 4-nitroaniline by carboxylamidases, was used for evaluating biochemical properties of these detoxifying enzymes in beet armyworm. The optimum pH and temperature were 7.5 and 38°C, respectively. K_m (Michaelis constant) and V_max (maximal velocity) at 28°C were 2.3 X 10^-4 M and 2.06 nmol min^-1 mg protein^-1, respectively. The enzyme activity was evaluated in several body parts and located mostly (66.2%) in the midgut. The soluble fraction (supernatant of 105,000g) contained the highest enzyme activity relative to the total (69.3%), and exhibited the highest specific activity. Carboxylamidase activity was totally inhibitedin vitro at a concentration of 10^-6 M methomyl. The analysis of thein vitro inhibition kinetics indicated the ability of methomyl and diflubenzuron to inhibit carboxylamidases noncompetitively. Over 95% inhibitionin vivo was obtained when the larvae were fed with castor bean leaves dipped in 250 mgl ^-1 of methomyl. Thein vivo enzyme activity could be reduced to half with a pretreatment of 15 mgl ^-1 diflubenzuron.     

Intestinal metabolism and absorption of carbaryl studied with a portal fistula

These data were obtained by use of a total and continuous portal vein fistula which virtually eliminated vascular redistribution of compounds absorbed from the gastrointestinal tract to nondigestive tissues (i.e., liver). The method allows direct measurement of the compounds absorbed, which is especially important in metabolism studies of ingested toxic compounds. These studies demonstrated that in vivo metabolism did occur within the intestine during the process of absorption of the pesticide carbaryl (naphthyl N-methylcarbamate) and naphthol, the hydrolysis product of the pesticide. Portal absorption of naphthol from a liquid diet (46 ± 4% of dose/120 min) was slower than from Ringer medium (75 ± 1%/120 min); portal absorption accounted for 82 ± 8 and 83 ± 4%, respectively, of the ¹⁴C absorbed from the intestine. The proportion of hydrophilic ¹⁴C-metabolites (water soluble) in portal blood varied from 6 to 89% and was a function of the substrate, dose vehicle (liquid diet vs Ringer), and time of portal fluid collection. Metabolism in the small intestine before absorption was confirmed for both substrates. The principal lipophilic constituent in portal fluid was the unmetabolized substrate for both carbaryl and naphthol; the principal ampholyte metabolite was naphthyl glucuronide. Although these in vivo data are qualitatively similar to evidence from previous in vitro studies, this in vivo evidence demonstrated that the extent of metabolism (and possibly detoxication) was considerably less than would be predicted from in vitro studies and indicates that the hazard of ingestion of carbaryl and other lipophilic toxic agents may be greater than realized.     

Pyrethroid synergists suppress esterase-mediated resistance in Indian strains of the cotton bollworm, Helicoverpa armigera (Hübner)

The role of esterase in pyrethroid resistance was studied in the final larval instar of different strains of the cotton bollworm, Helicoverpa armigera. The resistant strains viz., Nagpur strain and the Delhi strain were found to have elevated midgut esterase activity in comparison to the susceptible strain. Nagpur strain and Delhi strain have 2.24 and 1.73-fold higher esterase activity, respectively, than that of the susceptible strain. The Native PAGE displayed important differences in the midgut esterase isozyme pattern between the susceptible and the pyrethroid-resistant strains. Out of the 10 esterase isozyme observed, susceptible strain lacked three bands, E2, E6 and E10 that were found in the resistant strains. The potency of the synergists piperonyl butoxide (PBO) and dihydrodillapiole (DDA) as esterase inhibitor were also studied both in vitro and in vivo. The in vitro results clearly show that both PBO and DDA inhibited esterase activity in the two resistant strains, while there was almost no esterase inhibition in the homogenate of the susceptible strain. The in vivo inhibition studies (topical application of PBO and DDA followed by biochemical analysis) illustrated that PBO- and DDA-esterase binding is rather slow and non permanent process. Esterase inhibition did not occur immediately after the synergist treatment but at 4 and 8 h post treatment in case of PBO and DDA, respectively. Native PAGE revealed that the in vivo esterase inhibition caused by both PBO and DDA was due to the binding of the synergist with the E6 isozyme which was not present in the susceptible strain.     

Toxicological and biochemical characterizations of AChE in Liposcelis bostrychophila Badonnel (Psocoptera: Liposcelididae)

The toxicological and biochemical characteristics of acetylcholinesterases (AChE) in the resistant and susceptible strains (SS) of Liposcelis bostrychophila were investigated. The two resistant strains were the dichlorvos-resistant strain (DDVP-R) and the phosphine-resistant strain (PH₃-R) with resistance ratios of 22.36 and 4.51, respectively. Compared to their susceptible counterpart, the AChE activity per insect and the specific activity of AChE in DDVP-R and PH₃-R were significantly higher. There were also significant kinetic differences between DDVP-R and PH₃-R. The apparent Michaelis-Menten constant (K_m) for acetylthiocholine iodide (ATChI) was obviously lower in SS than that in PH₃-R, indicating a higher affinity to the substrate ATChI in the susceptible strains. The affinity for the substrate ATChI in DDVP-R and SS were not significantly different. The V_max value of the PH₃-R was significantly greater when compared to the V_max for the SS suggesting a possible over expression of AChE in this resistant strain. The inhibition of AChE to insecticide exposure in vitro revealed that all six insecticides were inhibitory for the extracted AChE’s. Based on the I₅₀ values, AChE of the SS were more sensitive to dichlorvos, paraoxon-ethyl, malaoxon and demeton-S-methyl than those of the two resistant strains. As for carbaryl and eserine, the PH₃-R suggested a significantly higher I₅₀s compared to the susceptible strain, while, no significant differences were found between SS and DDVP-R.     

Hydrolysis of permethrin by pyrethroid esterases from resistant and susceptible strains of Amblyseius fallacis (Acari: Phytoseiidae)

Pyrethroid-hydrolyzing activity in whole body homogenates of three insecticide-resistant and one susceptible strain of the predator mite, Amblyseius fallacis Garman has been examined in vitro. The highest esterase activity was found in the synthetic pyrethroid-resistant GH-1 strain body homogenate. All three pyrethroid-resistant strains had esterases that hydrolyzed trans-permethrin two times faster than cis-permethrin but isomer specificity was not observed in the susceptible strain. The pyrethroid esterases of the GH-1 strain were very sensitive to inhibition by dichlorovos, S,S,S-tributhylphosphorotrithioate, and 3-octylthio-1,1,1-trifluoro-2-propanone. Carbaryl and tetraethylpyrophosphate exhibited moderate inhibition in the GH-1 strain. Eserine sulfate and piperonyl butoxide only inhibited permethrin hydrolysis at higher concentrations. Fifteen esterase bands were resolved from body homogenates by gradient gel electrophoresis in the GH-1 strain, and were identified as carboxylesterases. The major pyrethroid-hydrolyzing activity was located in E5–E12 bands from GH-1 and composite susceptible strain esterases. Six esterase bands exhibiting low pyrethroid-hydroloyzing activity were also obtained from the two spotted spider mite, Tetranychus urticae (Koch).     

Enzyme activities and cytochrome P-450 levels of some Japanese quail tissues with respect to their metabolism of several pesticides

In the Japanese quail, cytochrome P-450, A- and B-esterase, amidase, and glutathione S-aryl transferase were assayed in postmitochondrial centrifugal fractions, in microsomes, and supernatant fractions of liver, lungs, kidneys, and testes. Liver microsomes contained the highest A-esterase activity and P-450 levels. B-esterase was more generally distributed and higher in the microsomal tissue fractions. Microsomal amidase activity was highest in quail lung and kidney, and lowest in the liver (per mg protein). Very little difference in glutathione S-aryl transferase activity was noted among the tissues assayed. In vitro metabolism of carbaryl, phosphamidon, and chlorotoluron by the various centrifugal fractions revealed that the production of 1-naphthyl-N-hydroxymethylcarbamate and 1-naphthol, the major metabolites, was greatest in the postmitochondrial fraction of the liver. The major carbaryl metabolite in all other quail tissue fractions was 1-naphthol. Phosphamidon metabolism in postmitochondrial preparations of quail liver was higher than in the supernatant and microsomes. Chlorotoluron metabolism occurred only in the postmitochondrial fractions of quail liver. The major products were the oxidative metabolites, N-(3-chloro-4-methylphenyl)-N′-methylurea and N-(3-chloro-4-hydroxymethylphenyl)-N′-methylurea.     

Comparative enzyme activities and cytochrome P-450 levels of some rat tissues with respect to their metabolism of several pesticides

Cytochrome P-450, A- and B-esterase, amidase, and glutathione S-aryl transferase were assayed in the postmitochondrial centrifugal fraction, microsomes, and supernatant of rat liver, lungs, kidneys, and testes. Liver microsomes contained the highest P-450 levels and A-esterase activity. B-esterase activity was more generally distributed and higher in the microsomal tissue fractions. Microsomal amidase activity was highest in rat lung and lowest in the liver (per mg protein). Glutathione S-aryl transferase activity was highest in the liver. The in vitro metabolism of carbaryl, phosphamidon, and chlorotoluron by the various centrifugal fractions revealed many differences. Carbaryl metabolism was greater in the liver microsomal fractions than in any other preparation. 1-Naphthol was the major metabolite in all tissue fractions. Although very little metabolism of phosphamidon occurred in the rat, metabolism in the rat liver postmitochondrial fraction was slightly higher with respect to the production of metabolites than in the supernatant and microsomes combined. Chlorotoluron was not metabolized by any of the tissue fractions of the rat. At least a low level of activity toward some compounds was observed in all tissues, but this study confirmed that the liver was the most active metabolizing tissue as well as having the highest levels of enzymatic activity usually associated with pesticide metabolism.     

Inhibition of permethrin-hydrolyzing esterases from Wiseana cervinata larvae

Inhibition of permethrin-hydrolyzing enzymes from larvae of the porina moth Wiseana cervinata has been examined in vivo and in vitro. Significant inhibition was shown by carbaryl and pirimiphos-methyl. 1-Dodecylimidazole substantially inhibited permethrin hydrolysis only in liver insects. The triphenylmethane dye tetrabromophenolphthalein was a moderate inhibitor only in vitro. TMDM (bis(N-dimethyl-4-aminophenyl)methane) had little effect on hydrolysis. These observations extend the range of species and substrates for which the triphenylmethane dyes and 1-dodecylimidazole are useful inhibitors of insecticide metabolism.     

Toxicity and metabolism of methomyl in the European corn borer

The contact and oral toxicity of methomyl (S-methyl N-[(methylcarbamoyl)oxy] thioacetimidate) was similar for two different strains of European corn borer, Ostrinia nubilalis (Hübner). In each case, third- and fourth-instar larvae were equally susceptible, but fifth-instar insects were considerably more difficult to kill. In vivo and in vitro studies revealed that borers from both strains metabolized methomyl via a mixed-function oxidase system to water-soluble products which could not be cleaved by acid or hydrolytic enzymes. By far, the greatest metabolic activity was localized in fat body tissues of last-instar larvae, and although both strains metabolized methomyl at a similar rate, a large difference was found in the rate of metabolism of methomyl oxime.     

Inhibition of Tribolium gut chitin synthetase

The chitin synthetase (CS) of Tribolium castaneum gut is inhibited 50% by 0.02 μM nikkomycin and 4 μM polyoxin D, two pyrimidine nucleoside fungicides, in in vitro assays with 10-min preincubation of enzyme and inhibitor prior to substrate addition. Tribolium CS is also sensitive to inhibition by the pyrimidine nucleotides uridine and cytidine di- and triphosphates. Captan, a known inhibitor of insect chitin synthesis, and the related fungicides captafol and dichlofluanid are highly potent inhibitors of Tribolium CS. Moderately active CS inhibitors are the acaricide oxythioquinox and the herbicide barban. One phenylcarbamate insect growth reatardant, H-24108, is weakly active in inhibiting Tribolium gut CS, as are three of its analogs but not 26 others. Many triazines are not inhibitory including several herbicides and an azido derivative, CGA 19255, which is active in blocking insect growth and chitin synthesis. Although the benzoylphenyl urea insecticides diflubenzuron and SIR 8514 are potent in vivo inhibitors of the polymerization step in insect chitin synthesis, they do not affect T. castaneum gut CS activity in vitro and greatly stimulate Tribolium brevicornis gut CS activity in vivo. These studies and preliminary findings on an integumental enzyme indicate that CS of these tissues is not sensitive to the direct action of benzoylphenyl ureas. This leads to speculation that the benzoylphenyl ureas act either as CS inhibitors via active metabolites formed in the integument or as blocking agents by direct binding to non-CS sites important in chitin polymerization and fibrillogenesis.     

Herbicidal interference with the photochemical activities of chloroplasts (in vivo and in vitro) of certain crop and weed species

The independent modes of action of diuron and atrazine on the photochemical activities of chloroplasts (In vivo and in vitro) from the leaves of crop plants Pisum sativum and Pennisetum typhoides and the weeds Amaranthus viridis and Cyperus rotundus were investigated. Hill reaction activity (DCPIP photoreduction) of in vivo chloroplasts (chloroplasts isolated from herbicide-sprayed plants) was unaffected by treatment at sublethal or intermediate levels of diuron or atrazine while that of in vitro chloroplasts (chloroplasts incubated in the required herbicidal concentration) was severely inhibited. The ferricyanide catalyzed noncyclic photophosphorylation was markedly reduced in both the in vivo and in vitro chloroplast systems. N-Methyl phenozonium sulfate (PMS)-mediated cyclic photophosphorylation was inhibited in the in vivo system while a pronounced enhancement of activity was noticed in the in vitro chloroplasts. The rate of NADP⁺ photoreduction was severely inhibited in the in vitro chloroplasts. The unaffected in the in vivo system. The herbicidal effects on the photoreactions of isolated chloroplasts were compared with chloroplasts isolated from herbicide-sprayed plants.     

Insensitivity of acetylcholinesterase in a field strain of the fall armyworm, Spodoptera frugiperda (J.E. Smith)

Acetylcholinesterase (AChE) was purified from adult heads of the fall armyworm (Spodoptera frugiperda) by using a two-step procedure involving gel filtration on a Sephadex G-200 column and affinity chromatography on a procainamide-ECH Sephadex 4B column. Both susceptible and field strains possessed two AChE isozymes, namely, AChE-1 and AChE-2, with subunit molecular weights of 63.7 and 66.1 kDa. The purified AChE had an apparent K_m value of 33.5 μM and a V_max of 7.07 μmol/min/mg protein in the susceptible strain. The apparent K_m and the V_max were 2.2- and 2.0-fold higher, respectively, in the field strain than in the susceptible strain. The purified AChE from the field strain was 17- to 345-fold less sensitive than that from the susceptible strain to inhibition by carbamates (carbaryl, eserine, methomyl, and bendiocarb) and organophosphates (methyl paraoxon and paraoxon), insensitivity being highest toward carbaryl. The results further support the notion that insensitive AChE played an important role in the insecticide resistance observed in the field strain.     

Rapid in vitro screening assay for immunotoxic effects of organophosphorus and carbamate insecticides on the generation of cytotoxic T-lymphocyte responses

There has been an increasing need for rapid and easily interpreted techniques for the screening of possible immunotoxicants. Besides the obvious detrimental effects of exposure to immunosuppressive agents, the modulation of the immune system which results from exposure to these toxicants may be a sensitive index to the toxicologic effects of such agents. Other researchers have proposed assays to screen the effect of in vitro treatment with immunotoxicants on mitogenic and humoral immune responses. In this report, we have described an in vitro technique for screening the effect of immunotoxicants, in the presence and absence of a NADPH fortified liver postmitochondrial supernatant (S-9) from Arochlor 1254-treated rat, on another aspect of the mammalian immune system, the generation of a T-cell-mediated cytolytic (CTL) response. This enzyme system altered the effect of organophosphorus compounds on the generation of a CTL response. Malathion and fenitrothion were no longer suppressive following this pretreatment; however, ethyl and methyl parathion and fenthion were only partially detoxified. In contrast, the S-9 enzyme system did not alter the effect of carbamate pesticides, carbaryl and carbofuran, on the generation of CTL responses. This report describes the effects of these seven organophosphorus and carbamate pesticides on the generation of the CTL response. In addition, some of the in vivo data published on the immunomodulatory effects of these compounds were collated from the literature and a correlation between in vitro and in vivo studies was discussed.