Inhibitor Report for: Malathion

Malathion resistance in a strain of Culex tarsalis mosquitoes is due primarily to the activity of a malathion carboxylesterase (MCE). The resistant strain was 150 times more resistant to malathion than the susceptible strain and was weakly resistant to malaoxon and carbaryl, but not to any other insecticide tested. The phenotype could be reversed with the carboxylesterase inhibitor triphenylphosphate, but no synergism was observed with either the phosphatase or polysubstrate monooxygenase inhibitors, NaF and piperonyl butoxide. MCE is expressed throughout development and is most concentrated in the gut tissues of the larvae. Subcellular fractionation indicated that MCE was localized primarily in the mitochondria of resistant insects and the cytoplasm of susceptible insects. The enzyme was purified to homogeneity from both strains, and has a molecular weight of 59,000. However, chromatofocusing indicated that resistant insects have two MCEs with pIs of 6.8 and 6.2, while susceptible insects possessed only one MCE with a pI of 6.8. The MCE unique to the resistant strain hydrolysed malathion 18 times faster than the MCE common to both strains, suggesting that malathion resistance in C. tarsalis is due to the presence of a qualitatively different esterase in the resistant strain.

Resistance to the organophosphorus insecticide malathion in genetically related strains of the Australian sheep blowfly Lucilia cuprina was examined. Separate lines of blowflies were established by homozygosis of the fourth chromosome of the parental RM strain. Both the RM and the derived resistant (der-R) strains are approximately 100 times more resistant to malathion than the related susceptible der-S strain, resistance being correlated with a 45- to 50-fold increase in a malathion carboxylesterase (MCE) activity. MCE has a pH optimum ranging between 6.6 and 8.0 and is strongly inhibited by the carboxylesterase inhibitors triphenyl phosphate, paraoxon, and diisopropylfluorophosphate. Subcellular fractionation revealed that MCE was localized predominantly to the cytosol and mitochondria in both resistant and susceptible blowflies. A single MCE was purified to homogeneity from RM blowflies. It has a pI of 5.5, is a monomer of 60.5 kDa, and hydrolyzes malathion with a Vmax of 755 nmol/min/mg protein and a Km of 11.0 microM. L. cuprina have thus evolved a remarkable MCE which is faster and more efficient at hydrolyzing a specific insecticide than any other insect esterase vet described.

Ten day old chick sympathetic ganglia cultured in a microslide assembly were treated with a selected group of organophosphate pesticides to evaluate their cytotoxicity ranges, and the usefulness of such a model for screening pesticides. Examination by phase contrast and light microscopy for chemically-induced morphological alteration of nerve fibers, glial cells and neurons provided the criteria for quantitation and assessment of the toxic effects. Concentrations that produced half-maximal effects ranged from 1 x 10(-6)M (severely toxic) for methylparathian, diazinon, paraoxon, mevinphos, diisopropylfluorophosphate, tri-o-tolyl phosphate and its mixed isomers to a 1 x 10(-3)M (intermediate) for malathion, leptophos, coumaphos, mono- and dicrotophos. Some or no effects were evident at 1 x 10(2-)M for O'ethyl-O-p-nitrophenyl phenyl phosphonothioate, tri-m-tolylphosphate, chlorpyriphos and triphenyl phosphate. In all instances, nerve fibers were more sensitive than neurons or glial cells to insecticides. All cellular growth was inhibited at 1 x 10(-2)M (except triphenyl phosphate). Below 1 x 10(-7)M, no inhibitory effects were evident. The secondary abnormalities included decreased cellular migration, diffuse cellular growth pattern, increased vacuolization, nerve fiber swelling and cellular degeneration. The cytotoxic effects of these chemicals do not appear to be related to in vivo toxicity or cholinesterase inhibition potential.

To establish an insecticidal resistance surveillance program, Culex quinquefasciatus mosquitoes from So Paulo, Brazil, were colonized (PIN95 strain) and analyzed for levels of resistance. The PIN95 strain showed low levels of resistance to organophosphates [malathion (3.3-fold), fenitrothion (11.2-fold)] and a carbamate [propoxur (3.0-fold)]. We also observed an increase of 7.4 and 9.9 in alpha and beta esterase activities, respectively, when compared with the reference IAL strain. An alteration in the sensitivity of acetylcholinesterase to insecticide inhibition was also found in the PIN95 mosquitoes. The resistant allele (Ace.1R), however, was found at low frequencies (0.12) and does not play an important role in the described insecticide resistance. One year later, Cx. quinquefasciatus mosquitoes were collected (PIN96 strain) at the same site and compared to the PIN95 strain. The esterase activity patterns observed for the PIN96 strain were similar to those of the PIN95 mosquitoes. However the occurrence of the Ace.1R allele was statistically higher in the PIN96 strain. The results show that esterase-based insecticide resistance was established in the PIN95 Cx. quinquefasciatus population and that an acethylcholinesterase based resistant mechanism has been selected for. A continuous monitoring of this phenomenon is fundamental for rational mosquito control and insecticide application programs.

The urinary excretion rates of dimethyl-phosphate, -phosphorothioate and -phosphorodithioate were studied in six persons of whom four had ingested a concentrated solution of malathion and two of thiometon. The concentration decrease of single and total dimethylphosphorus metabolites was biphased, with a fast initial rate and a slow later rate. The excretion rate of total metabolites in the faster phase depended on the initial concentration in urine. At concentrations higher than 100 nmol/mg creatinine, the excretion half-times ranged from 7.5 to 15.4 h and at concentrations between 52 and 95 nmol/mg creatinine from 34.7 to 55.4 h. Non-metabolized malathion was detected only in one urine sample collected from one person immediately after hospitalization. Two persons poisoned with malathion were taken blood serum samples for the analysis of the parent pesticide and its metabolites on a daily basis after hospitalization. The parent pesticide was detectable in the serum only one day after the poisoning. The concentration of total malathion dimethylphosphorus metabolites in serum decreased very quickly within 1.5 days after hospitalization. The total metabolite elimination half-times were 4.1 and 4.7 h in the initial phase, and 53.3 and 69.3 days in the later slower elimination phase. There was no correlation between maximum concentrations of total metabolites measured in serum and/or urine on the day of admission to hospital and the initial depression of serum cholinesterase (BChE, EC 3.1.1.8) and erythrocyte acetylcholinesterase (AChE, EC 3.1.1.7).

Biomarker assays that provide measures of the toxic effects of chemicals on key organisms are of particular interest in ecotoxicology and environmental risk assessment. Typically, such assays provide measures of the molecular mechanisms that underlie toxicity (e.g., inhibition of brain acetylcholinesterase activity by organophosphorus insecticides and retardation of the vitamin K cycle by anticoagulant rodenticides). They are particularly valuable for detecting and quantifying toxicity where organisms are exposed to mixtures of compounds and for identifying cases of potentiation. In birds, inhibition of brain acetylcholinesterase activity can provide an index of potentiation of organophosphorus and carbamate insecticides by other pesticides. Inhibition of serum butyrylcholinesterase also is very useful as a nondestructive assay but is not simply related to inhibition of brain acetylcholinesterase. Assays for DNA damage can indicate where there is an increase in the rate of activation of carcinogens and mutagens due to induction of the cytochrome P450 system. Assays for blood levels of retinol (vitamin A) and thyroxine can establish thyroxine antagonism by metabolites of 3,3,4,4-tetrachlorobiphenyl. Assays for changes in levels of clotting protein in serum can give an indication of the effect of mixtures of anticoagulant rodenticides on the vitamin K cycle. The interactive effects of mixtures of pesticides in the field are starting to be investigated by this approach (e.g., a recent study of the combined action of malathion and prochloraz in the red-legged partridge).

We studied the effects of chronic exposure to malathion in the kidney of Octodon degus, a caviomorph whose habitat may be exposed to pesticides currently used in Chilean agriculture. A group of adult female animals received malathion (200 ppm) as sole drinking fluid for 90 days. Kidneys showed signs of histologic damage, marked by hyperplasia and hypertrophy of tubular cells. Exposed animals had unchanged glomerular filtration rates and renal handling of sodium and chloride, but a significant increase in fractional excretion of potassium resulted from this treatment. The activities of Na+/K(+)-ATPase and Mg(2+)-ATPase in renal cortex and outer medulla were not affected by malathion exposure. This study provides evidence of both morphologic and functional renal damage elicited by chronic exposure of O. degus to a low dose of malathion. Morphologic alterations in glomerulus were accompanied by either morphologic and functional impairments of the distal nephron.

A field population of Lutzomyia longipalpis from La Rinconada, Lara State, an endemic focus of visceral leishmaniasis in Venezuela, was tested for susceptibility to organochlorine (DDT 2%), carbamate (propoxur 0.01%), organophosphate (malathion 2%, fenitrothion 1%, and pirimiphos methyl 1%), and pyrethroid (deltamethrin 0.06%, lambdacyhalothrin 0.06%, and permethrin 0.2%) insecticides. Susceptibility to the insecticides tested was evaluated in the field population of L. longipalpis and compared with a laboratory reference strain. The (LT95) to propoxur and malathion insecticides for the field population was lower than the LT95 for the laboratory reference strain, demonstrating high susceptibility to these compounds. A low level of resistance at LT50 (< 3-fold) was found for fenitrothion, pirimiphos methyl, and permethrin insecticides, but no resistance was detected at LT95. No significant resistance at the LT50 and LT95 was detected for the pyrethroids deltamethrin and lambdacyhalothrin. The susceptibility levels of L. longipalpis to the insecticides tested are discussed in view of a future control program against endophilic vectors of leishmaniases based on the use of pesticides.

Previous studies have shown that acute, oral administration of malathion modulated the humoral immune response to T-cell-dependent antigen, mitogenic responses, macrophage function, and mast cell degranulation. While administration of malathion for 14 days did not affect the generation of an immune response to antigen, it was possible that macrophage and mast cell functions were affected. In this report, the effect of malathion administration for 14 days upon these parameters were assessed. This treatment regimen increased the respiratory burst capacity to a maximal level at a dose of 1 mg/kg/day or greater. The effect of oral administration of malathion for 14 days on the degranulation of mast cells in various organs (heart, skin, and small intestine) and peritoneal lavage fluid was also assessed. At doses of 1 mg/kg/day and above, the number of mast cells that was undegranulated decreased and the number that was severely degranulated increased. There was no change in mast cell integrity in biopsies from heart and skin, and in peritoneal fluid after 14-day administration of 0.1 mg/kg/day. However, the number of mast cells associated with the small intestine that had undergone degranulation was increased at this dose of malathion. These data indicate that repeated administration of malathion increased macrophage function at doses as low as 1 mg/kg/day and led to mast cell degranulation at doses as low as 0.1 mg/kg/day.

It was analyzed the behavior of the resistance of 3 organophosphated insecticides (malathion, clorpirifos and methyl-pyrimifos), 3 pyrethroids (deltamethrin, lambda-cyhalothrin and cypermethrin), and 1 carbamate (propuxur) in populations of Culex quinquefasciatus from 2 municipalities of the province of Santiago de Cuba. The values of the resistance factor proved that there is resistance to malathion and clorpirifos. However, in spite of the existence of a high frequency of the mechanisms of elevated esterases and altered acetylcholinesterase no resistance to methylpymirifos, was observed which demonstrated that this insecticide is not affected by these mechanisms selected in our populations of Culex quinquefasciatus. There was resistence to deltamethrin, and lambda-cyhalothrin in Santiago de Cuba, whereas it was moderate to cypermethrin in Santiago and San Luis. Resistance to deltamethrin was also found in San Luis, but it was mild to lambda-cyhalothrin. The results obtained from the use of the synergists S,S,S tributyl phosphotritiade (DBF) and piperonyl butoxide (PB) indicated that the mechanisms of resistance of unspecific esterases and oxidases of multiple function are involved in the resistance to pyrethroids in both strains from Santiago de Cuba and San Luis. It was determined by the biochemical tests that there existed a high frequency of the mechanisms of esterases and altered acetylcholinesterase. The results of the polyacrylamide gel electrophoresis (PAGE) showed that esterase B1 appears more frequently associated with esterases A6 and B6. It was inferred that this association could be connected with the resistance to pyrethroids.

A toxicity test for organophosphates (OP) and carbamates (C) was improved with the adult ragworm Nereis diversicolor. Animals were maintained in U-shaped glass tubes of 4-mm inner diameter fixed vertically on a plastic plate and placed in glass aquaria. Each tank was covered with glass in order to reduce evaporation and heat dissipation. Temperature varied between 15 and 16 degrees C and salinity was constant (34 per thousand) during the entire length of the experiment. Experiments were performed with a fixed day length of 12 h and seawater was gently aerated. The maintenance system allowed the administration of OP and C compounds via the seawater. An acclimatization period of 48 h was not sufficient to accomodate worms to their artificial burrows; accordingly, we chose to acclimate worms for a week before beginning the exposure. Choline acetyltransferase (ChAT) activity was very low and was not significantly modified by two OP compounds: malathion and parathion-ethyl. ChAT is not a target for these pesticides and should not be used for future studies about OP and C toxicity. On the other hand, inhibitory effects on acetylcholinesterase (AChE) activity were determined at concentrations of 10(-6) M for three OP compounds-malathion, parathion-ethyl, and phosalone-and a carbamate pesticide-carbaryl. We measured only short-term effects and no cumulative effect was determined, the maximum percentage of AChE activity inhibition being between 2 (carbaryl) and 7 (OP compounds) days after exposure and then remaining stable. Mortality occured only after a period of intoxication of 14 days. N diversicolor, which can be easily maintained at the laboratory, seems to be a good candidate for future laboratory studies to test the toxicity of other pollutants.

Approximately 200 citrus samples from markets of the Valencian Community (Spain) were analyzed to establish their residue levels in 12 organophosphorus pesticide residues during the 1994-1995 campaign. The organophosphorus pesticides carbophenothion, chlorpyriphos, chlorfenvinphos, diazinon, ethion, fenitrothion, malathion, methidation, methylparathion, phosmet, quinalphos, and tetradifon were simultaneously extracted by matrix solid-phase dispersion and determined by gas chromatography-mass spectrometry using selected ion monitoring mode. A total of 32.25% contained pesticide residues and 6.9% exceeded the European Union Maximum Residue Levels (MRLs). The pesticides found in the samples with residues above MRLs were carbophenothion, ethion, methidathion, and methyl parathion. Lower level residues of these and the other pesticides studied (except diazinon) were frequently found. The estimated daily intake of the 12 organophosphorus pesticide residues during the studied period was 4.87 x 10(-4) mg/kg body weight/day. This value is lower than the provisional tolerances dairy intakes proposed by the Food and Agriculture Organization and the World Health Organization.

We reviewed 130 admissions of organophosphate poisoning and analyzed the causes of death. Fenitrothion, malathion, dichlorvos, trichlorfon and fenitrothion/malathion were the most frequent chemicals involved. Mortality was 25% (32/130); delay in discovery and transport (18 cases), insufficient respiratory management (8 cases), and severe underlying or co-existing diseases (6 cases) were noted in the lethalities. Insufficient respiratory management consisted of delay in endotracheal intubation (5 cases) and failure in weaning (3 cases). About 3/4 of the severely serum cholinesterase-depressed cases needed ventilators. This suggests that better respiratory management would improve the outcome of organophosphate poisonings. Close observation of the clinical symptoms is essential, and detection of changes in serum cholinesterase may be helpful.

A rapid procedure has been developed that allows a single-step, selective extraction and cleanup of organophosphate (OP) pesticide residues from milk dispersed on solid-matrix diatomaceous material filled into disposable cartridges by means of light petroleum saturated with acetonitrile and ethanol. Recovery experiments were carried out on homogenized commercial milk (3.6% fat content) spiked with ethanolic solutions of 24 OP pesticides, viz., ethoprophos, diazinon, dimethoate, chlorpyrifos-methyl, parathion-methyl, chlorpyrifos-ethyl, malathion, isofenphos, quinalphos, ethion, pyrazophos, azinphosethyl, heptenophos, omethoate, fonofos, pirimiphos-methyl, fenitrothion, parathion, chlorfenvinphos, phenthoate, methidathion, triazophos, phosalone, azinphos-methyl, at levels ranging for the different OP pesticides from 0.02 mg/kg to 1.11 mg/kg. Average recoveries of four replicates were in the range 72-109% for the different OP pesticides, with relative standard deviations (R.S.D.) from ca. 1 to 19%, while dimethoate and omethoate were not recovered. Coextracted fatty material amounted to an average of about 4.0 mg/ml of milk. The extraction procedure requires about 30 min. The main advantages are that extraction and cleanup are carried out in a single step, emulsions do not occur, several samples can be run in parallel by a single operator, reusable glassware is not needed and simple operations are required.

Residue study was performed on several insecticides which could contaminate local Egyptian beans. The effect of storage periods and various processing steps on lowering the residues of malathion and pirimiphos methyl in treated seeds and their processed products were investigated. The data indicated that malathion and pirimiphos methyl persisted for more than 90 days on and in stored mature dry broad beans after postharvest treatment. However, stored broad beans could be safely used for human consumption after 90 days when the insecticide residues reached safe levels. Washing removed 69 and 75% of malathion and pirimiphos methyl residues of treated broad beans, respectively. Malathion residue was not detected in various processed products. More than 89 and 99% of malathion residues were absent in dehulled and heated dehulled broad beans. In addition, pirimiphos methyl residues were reduced to 92, 97, 87, 99, 99, and 95% from the initial levels in treated beans following dehulling, cooking of dehulled beans, germination, cooking of germinated beans and cooking of the beans by the common method and under pressure, respectively.

During Fiscal Years 1989-1994, the U.S. Food and Drug Administration (FDA) collected and analyzed 545 domestic surveillance samples of mixed feed rations (172 for cattle, 125 for poultry, 83 for swine, 61 for pets, 56 for fish, and 48 miscellaneous). All samples were analyzed by gas-liquid chromatography for organohalogen and organophosphorus pesticides. Of the 545 samples, 88 (16.1%) did not contain detectable pesticide residues. In the 457 samples with detectable pesticide levels, 804 residues (654 quantitable and 150 trace) were found. None of these 804 residues exceeded regulatory guidance. Malathion, chlorpyrifos-methyl, diazinon, chlorpyrifos, and pirimiphos-methyl were the most commonly detected pesticides. These 5 organophosphorus pesticides accounted for 93.4% of all pesticide residues detected (malathion, 52.9%; chlorpyrifos-methyl, 25.2%; diazinon, 7.7%; chlorpyrifos, 4.9%; and pirimiphos-methyl, 2.7%). Their median values in samples containing quantitable levels ranged from 0.014 to 0.098 ppm. The most commonly detected organohalogen compounds were methoxychlor, DDE, PCB, dieldrin, pentachloronitrobenzene, and lindane. These 6 compounds combined accounted for only 4.1% of all residues detected. FDA is continuing its pesticide surveillance of feeds to help ensure animal safety and prevent violative residues in food derived from animals.

Although the recent distribution of yellowfever mosquito, Aedes aegypti (L.), in Pakistan has been restricted to the port city of Karachi, adult and immature mosquitoes breeding in imported tires in warehouses at Landi Kotal (North-West Frontier Province) were identified as Ae. aegypti. The patterns of tire trade and the current disjunct distribution of Ae. aegypti indicated that the introduction into Landi Kotal may have been either from Karachi or India. Thermal fog application of pirimiphos-methyl and residual spray of malathion during 1993 reduced abundance in October-November. Living larvae or adults were not found during January 1994, apparently because of cold weather. However, Ae. aegypti reappeared during May-June 1994, most probably from eggs that overwintered. Population increased during late August when another round of spray using the same insecticides and fenthion as an additional larvicide again reduced abundance. Although this mosquito apparently has not spread into neighboring areas, its survival at Landi Kotal through all seasons despite control measures indicates its potential of becoming established in other areas of Pakistan.

A multiresidue extraction method based on matrix solid-phase dispersion (MSPD) is optimized for the extraction and gas chromatographic screening of eighteen insecticides (aldrin, carbophenothion, captafol, chlorpyriphos, chlorfenvinphos, diazinon, dicofol, alpha-endosulfan, beta-endosulfan, ethion, fenitrothion, folpet, methidathion, malathion, methyl-azinphos, methyl-parathion, phosmet, and tetradifon) from oranges. After optimization of different parameters, such as type of solid phase used and the amount of solid phase or eluent, recoveries ranged from 67 to 102% with relative standard deviations ranging from 2 to 10%. The limits of detection, calculated as 3 times the baseline noise ranged from 2 to 171 micrograms/kg. These limits of detection were about 10 times lower than the maximum residue levels established by the European Community. Compared with classical methods, the described procedure is simple, less labour intensive and does not require preparation and maintenance of equipment. Troublesome emulsions, such as those frequently observed in liquid-liquid partitioning did not occur.

Populations of Culex pipiens were sampled from 8 locations in Cyprus between 1987 and 1993. All population samples generally revealed organophosphate resistance to malathion, temephos, chlorpyrifos, fenthion, dichlorvos, and pirimiphos methyl, in decreasing order of magnitude. Of 7 populations assessed with the carbamate propoxur, all proved to be resistant to different degrees. Of the 6 populations tested with permethrin, 2 were resistant to permethrin. Resistance was associated with the presence of 5 different overproduced esterases (esterases A1, A2, A5, B2, and B5) as well as an insensitive form of acetylcholinesterase. These results are discussed in relation to the ongoing mosquito abatement program in Cyprus and to similar programs in other parts of the world.

A Culex quinquefasciatus Say 1823 strain with resistant genes to organophosphates was tested in the laboratory to know the reproductive potential after exposure, as larvae, at the LC30 and LC70 (mg/l) of three organophosphorus insecticides: malathion, chlorpyrifos and methyl-pirimiphos. Data showed that fecundity was decreased significantly by malathion at LC30 = 0.0025 and LC70 = 0.0075, whereas fertility has a no significant decrement by chlorpyrifos and methyl-pirimiphos at the LC70 (0.000016, 0.00043). The sexual index was affected by chlorpyrifos and methyl-pirimiphos showing a greater number of adult females.

During the experimental use of fenitrothion to replace malathion for the control of malaria in North West Frontier Province of Pakistan, serious intoxication of Afghan refugee spraymen occurred. A few weeks after commencement of the spraying operations, cholinesterase levels had fallen to 43.8% in personnel mixing the insecticide, and to 60.7% in spraymen, as measured by tintometry. Most of the personnel reported symptoms of overexposure and the spraying operations had to be discontinued. Intoxication of personnel resulted in poor coverage of the target area. High ambient temperatures during Pakistan's spray season discourage the use of full-protective clothing. Fenitrothion intoxication observed in the Afghan refugee programme, and similar experiences in Pakistan in the past, suggest that this insecticide is too toxic for routine use, when the compliance with safety precautions cannot be effectively supervised.

Biochemical biomarkers, such as inhibition of serum butyryl cholinesterase (BCHE) and brain acetyl cholinesterase (AChE), have been useful in studies of interactive effects of pesticides in birds. Examples of interactions due to increased activation or decreased detoxication are reviewed. Studies have shown that hybrid red-legged partridges (Alectoris rufa cross) pretreated with the inducing ergosterol biosynthesis inhibiting (EBI) fungicide, prochloraz, were more sensitive to the toxic effects of the organophosphorous (OP) insecticide, malathion, than controls. A dose of 90 mg/kg prochloraz produced greater inhibition at 1, 4 and 24 h following oral administration of 50 mg/kg malathion, compared to corn oil controls. Pigeons (Columba livia) given 180 or 90 mg/kg prochloraz showed greater inhibition of BCHE activity following malathion administration than did control birds. Starlings (Stumus vulgaris), however, appeared not to be induced by 180 or 300 mg/kg prochloraz, and no difference in BCHE activity following dosing with malathion was apparent in comparison with controls. Other EBIs and OP combinations have been investigated in the partridge. Birds pretreated with prochloraz showed a trend towards greater inhibition of serum BCHE activity at most time points following dosing with the OPs dimethoate and chlorpyriphos. Birds pretreated with the EBI penconazole showed significantly greater inhibition of serum BCHE activity at 1, 4 and 24 h after malathion administration than did controls. The mechanism of increased activation of malathion due to induction of cytochrome P-450 by prochloraz is reviewed. In the case of interactions due to inhibition of detoxication, inhibition of brain AChE activity was a useful biochemical biomarker

An outbreak of pediculosis at primary schools was recorded in the Czech Republic in 1992. Almost 20% of children in some schools were infested. This outbreak can be attributed to the resistance of head lice to permethrin, which has not been mentioned in literature yet. The resistance factors established in three towns range between 2 and 385 and between 5 and 557 for LC50 and LC90 values, respectively. This resistance has developed after exclusive use of pyrethroids lotion and shampoo in the Czech Republic since 1978, and it was accompanied by a cross-resistance to d-phenothrin and bioalethrin. But the susceptibility of head lice to malathion and pirimiphos-methyl in 1992 was very similar to that found in 1981. The lotion containing 0.3% of malathion (Diffusil H92 M) has been fully effective against the resistant lice. When introduced into the practice, it quickly reduced the infestation of children in primary schools. The other lotion and shampoo containing 0.3% and 0.7% of pirimiphos-methyl respectively were found to be effective as well.

Anopheles culicifacies (probably species A) is the main vector of malaria in Baluchistan, southeastern Iran. Adult mosquitoes were collected during 1990-92 by five methods of sampling: knock-down pyrethrum space-spray indoors, human and animal bait (18.00-05.00 hours), pit shelters and CDC light traps, yielding 62%, 3%, 6%, 4% and 25% of specimens, respectively. Whereas spray-catches comprised c. 70% gravid and semi-gravid females, light trap catches were mostly (c. 60%) unfed females, while females from pit shelters comprised all abdominal stages more equally (13-36%). An.culicifacies populations peaked in April-May and rose again during August-November. Densities of indoor-resting mosquitoes were consistently greater in an unsprayed village than in villages subjected to residual house-spraying with propoxur, malathion or pirimiphos-methyl. Monthly malaria incidence generally followed fluctuations of An.culicifacies density, usually with a peak in May-June.

Susceptibility to six organophosphate (OP), two pyrethroid (PY), and one carbamate (C) insecticides was investigated in Culex pipiens quinquefasciatus Say, Aedes aegypti (L.), and Aedes polynesiensis Marks larvae from the island of Tahiti. Cx. p. quinquefasciatus and Ae. aegypti were compared with susceptible reference strains treated simultaneously. A low, but significant, resistance to bromophos (4.6x), chlorpyrifos (5.7x), fenthion (2.4x), fenitrothion (5.0x), temephos (4.3x) and permethrin (2.1x) was found in Cx. p. quinquefasciatus, and to malathion (1.5x), temephos (2.3x), permethrin (1.8x) and propoxur (1.7x) in Ae. aegypti. Cx. p. quinquefasciatus was shown to possess over-produced esterases A2 and B2, which are known to be involved in resistance to OPs in other countries. Ae. polynesiensis was less resistant than the Ae. aegypti reference strain to all insecticides except temephos (1.8x) and permethrin (6.7x). To determine whether Ae. polynesiensis had developed resistance to these insecticides in Tahiti, a geographical survey covering 12 islands of the Society, Tuamotu, Tubuai, Marquesas, and Gambier archipelagoes was undertaken with three insecticides (temephos, deltamethrin, and permethrin). Two- to threefold variations in LC50S were observed among collections. Results are discussed in relationship to the level of insecticide exposure on the different islands.

Malathion resistance in a strain of Culex tarsalis mosquitoes is due primarily to the activity of a malathion carboxylesterase (MCE). The resistant strain was 150 times more resistant to malathion than the susceptible strain and was weakly resistant to malaoxon and carbaryl, but not to any other insecticide tested. The phenotype could be reversed with the carboxylesterase inhibitor triphenylphosphate, but no synergism was observed with either the phosphatase or polysubstrate monooxygenase inhibitors, NaF and piperonyl butoxide. MCE is expressed throughout development and is most concentrated in the gut tissues of the larvae. Subcellular fractionation indicated that MCE was localized primarily in the mitochondria of resistant insects and the cytoplasm of susceptible insects. The enzyme was purified to homogeneity from both strains, and has a molecular weight of 59,000. However, chromatofocusing indicated that resistant insects have two MCEs with pIs of 6.8 and 6.2, while susceptible insects possessed only one MCE with a pI of 6.8. The MCE unique to the resistant strain hydrolysed malathion 18 times faster than the MCE common to both strains, suggesting that malathion resistance in C. tarsalis is due to the presence of a qualitatively different esterase in the resistant strain.

Resistance to the organophosphorus insecticide malathion in genetically related strains of the Australian sheep blowfly Lucilia cuprina was examined. Separate lines of blowflies were established by homozygosis of the fourth chromosome of the parental RM strain. Both the RM and the derived resistant (der-R) strains are approximately 100 times more resistant to malathion than the related susceptible der-S strain, resistance being correlated with a 45- to 50-fold increase in a malathion carboxylesterase (MCE) activity. MCE has a pH optimum ranging between 6.6 and 8.0 and is strongly inhibited by the carboxylesterase inhibitors triphenyl phosphate, paraoxon, and diisopropylfluorophosphate. Subcellular fractionation revealed that MCE was localized predominantly to the cytosol and mitochondria in both resistant and susceptible blowflies. A single MCE was purified to homogeneity from RM blowflies. It has a pI of 5.5, is a monomer of 60.5 kDa, and hydrolyzes malathion with a Vmax of 755 nmol/min/mg protein and a Km of 11.0 microM. L. cuprina have thus evolved a remarkable MCE which is faster and more efficient at hydrolyzing a specific insecticide than any other insect esterase vet described.

Syntheses of the enantiomers of malathion, malaoxon, and isomalathion are reported herein. Malathion enantiomers were prepared from (R)- or (S)-malic acid in three steps. Enantiomers of malathion were converted to the corresponding enantiomers of malaoxon in 52% yield by oxidation with monoperoxyphthalic acid, magnesium salt. The four isomalathion stereoisomers were prepared via two independent pathways using strychnine to resolve the asymmetric phosphorus moiety. The absolute configurations of the four stereoisomers of isomalathion were determined by X-ray crystallographic analysis of an alkaloid salt precursor. A high-performance liquid chromatography technique was developed to resolve the four stereoisomers of isomalathion, and to determine their stereoisomeric ratios.

Susceptibility status of five populations of Ae. albopictus mosquitoes from Maharashtra state, to DDT, malathion, fenitrothion, bromophos, propoxur and deltamethrin was studied and compared with the laboratory population. Four populations survived when adults were exposed to 4 per cent DDT impregnated paper for 2 h; though three of these populations had lower DDT-LC50 values at larval stages in comparison with the laboratory population. Results of topical application of DDT on these four populations supported these findings by showing comparatively higher LD50 values at adult stages in comparison with the laboratory population. All the populations were highly susceptible to other pesticides tested i.e., malathion, fenitrothion, bromophos, propoxur and deltamethrin. These populations were distinguished from each other by esterase isoenzyme patterns.

Two field trials in the control of subperiodic brugian filariasis vectors, mainly Mansonia bonneae and Mansonia dives were carried out in Sarawak, East Malaysia. In the first trial, malathion ultra-low volume (ULV) spray was used to control the Mansonia mosquitos in two filariasis endemic villages. Six spray rounds were applied at biweekly intervals at Kampung Rasau and two spray rounds were applied at monthly intervals in Kampung Triboh. ULV malathion spray reduced biting Ma. bonneae population for 3 days after spraying. The biting density decreased to 50% of the pre-treatment level by the 12th - 13th day and reached the pre-treatment level by the 24th - 25th day. Contact bioassay tests on caged Mansonia mosquitos revealed considerable penetration of the malathion aerosol indoors and relatively adequate coverage outdoors. The estimated number of bites per case per day was 1.09 to 4 times less in the sprayed kampung than in an unsprayed control kampung. The parous and daily survival rates of Mansonia mosquitos were not significantly affected by the spraying. In a second trial, chemotherapy with diethylcarbamazine citrate (DEC) was combined with vector control through indoor residual spraying in Kampung Ampungan. The results were compared with the use of only DEC mass treatment in Kampung Sebangkoi and Kampung Sebamban. The combined control measures in Kampung Ampungan reduced the MfD-50 to 44% of the pre-treatment level over a period of 4 years. In the other two kampungs where only mass DEC therapy was applied, the microfilarial rate and MfD-50 declined significantly in the second blood survey but increased gradually in two subsequent follow-up blood surveys. The total insecticidal impact for Ma. bonneae was 3.9 to 1 indoors and 2.7 to 1 outdoors. These results indicated that quarterly pirimiphos-methyl indoor spraying used in integrated control could reduce indoor transmission by 3.9 times. The infective rate from the Ma. bonneae dissected in all three kampungs after the interventions, irrespective of DEC treatment alone or in combination with pirimiphos-methyl residual spraying were reduced by two fold. However the infection rate of brugian filarial larvae in Kampung Ampungan was significantly reduced after the use of DEC and insecticide. Annual Transmission Potential (ATP) showed a high significant reduction in Kampung Ampungan (p > 0.001) compared with Kampungs Sebangkoi and Schambam. In Ampungan, the ATP was reduced by 8.5 times indoors after the MDA and insecticidal application and 3 times outdoors. The reduction rate for Sebangkoi and Sebamban both indoors and outdoors were less than 2 fold.

Adult bedbugs, Cimex lectularius, were exposed for 24 h (25 degrees C) to filter paper treated with various dilutions of the technical grade of nine insecticides dissolved in acetone to determine the concentration-response relationships. The order of toxicity, from most to least based on the LC50's was: dichlorvos, pirimiphos methyl, lambda-cyhalothrin, bendiocarb, permethrin, malathion, carbaryl, tetrachlorvinphos, and fenvalerate. The residual toxicities of commercial formulations of six of the chemicals diluted with water and applied to wood, cardboard, cloth and galvanized metal, were determined by exposing adult bedbugs at 3, 7 and 12 weeks after treatment. The formulation of bendiocarb (FICAM 76% W) had little residual activity on all surfaces at 12 weeks after treatment. The formulation of carbaryl (SEVIN 21.5% L) was toxic to bedbugs on all surfaces at 12 weeks after treatment, but required high concentrations on wood, cardboard, and cloth. The formulation of pirimiphos methyl (ACTELLIC 57% EC) had no residual activity on any of the surfaces at 12 weeks after treatment. The formulation of tetrachlorovinphos (RABON 50% W) had residual activity for 12 weeks on all surfaces except metal. The formulation of permethrin (ATROBAN 11% EC) had residual activity on only metal and wood while the formulation of lambda-cyhalothrin (KARATE 13.1% EC) had residual activity 12 weeks on all surfaces.

Use of the organophosphorus insecticide malathion for mosquito control in Cuba, for 7 years up to 1986, selected elevated non-specific esterase and altered acetylcholinesterase (AChE) resistance mechanisms in Culex quinquefasciatus. In central Havana space-spraying of malathion was replaced by the pyrethroid cypermethrin in 1987: alternate cycles of malathion and cypermethrin were applied in some of the more rural areas of Havana district during 1987-91. Consequently, populations of Cx quinquefasciatus in the central area of Havana developed resistance to cypermethrin, but there is no evidence of pyrethroid resistance in the outlying areas. Malathion resistance levels declined significantly after 1986, measured both by bioassay and the frequency of the elevated esterase resistance mechanism, and then stabilized with no measurable decline during 1990 in any of the populations tested. These populations had less than 10% frequency of susceptible homozygotes for both the esterase and AChE resistance mechanisms, indicating that organophosphate resistance is still prevalent in Cuban Cx quinquefasciatus. These two mechanisms appear to be in linkage equilibrium, suggesting that current selection for double resistance is not strong. In the central Havana region, pirimiphos-methyl, an organophosphorus insecticide unaffected by the two common malathion resistance mechanisms, is now being used in a resistance management strategy designed to avoid pyrethroid resistance spreading.

Some compounds that inhibit acetylcholinesterase (AChE) activity compete directly with quinuclidinyl benzilate (QNB) binding, a muscarinic antagonist which binds to all subtypes equally, and with cis-methyldioxolane (CD), an agonist that binds with high affinity to the M2 subtype of muscarinic receptors. The relationship between inhibition of AChE activity and the capability to affect muscarinic receptors directly has not been systematically explored. The interaction of eight organophosphates with muscarinic receptors was compared to their ability to inhibit AChE activity in vitro in tissue homogenates from rat hippocampus and frontal cortex, two cholinergically enriched areas of the brain. Of the compounds tested only echothiophate competed for [3H]QNB binding and only at concentrations greater than 100 microM. The anticholinesterase compounds were also tested for their ability to compete with a muscarinic receptor agonist, [3H]CD, which binds with high affinity (approximate KD = 3.5 nM) to 10 and 3% of the muscarinic receptors in the frontal cortex and hippocampus, respectively. The anticholinesterase compounds inhibited high-affinity [3H]CD binding up to 80% and the effects were similar in both tissues. Echothiophate and DFP were potent inhibitors of [3H]CD binding, as were the active "oxon" forms of parathion, malathion, and disulfoton. The parent "thio" forms of these insecticides, however, were much less effective in competing for [3H]CD binding. A similar pattern of potency was observed for the inhibition of brain AChE activity. A strong correlation was found between the ability of a compound to inhibit AChE activity and the ability to compete with [3H]CD binding. These data suggest that the biological effects of cholinesterase-inhibiting compounds may be due to more than their ability to inhibit AChE.

Various organophosphorus compounds with low acute toxicity levels are widely used as insecticides. Human acute poisoning by organophosphates has often occurred accidentally. We determined the activity and isoenzyme patterns of serum cholinesterase (ChE) obtained from 13 human patients who attempted suicide with various organophosphates, i.e. Fenitrothion, Malathion, Isoxathion, Pyridaphenthion and Trichlorfon, and studied on the changes in the activity and isoenzyme patterns of serum ChE after ingestion. The following results were obtained. 1) Twenty ChE isoenzyme bands from normal human serum were detected by electrophoretic separation on polyacrylamide gradient gel. The main bands in the ChE isoenzyme pattern in normal serum were bands 4 and 5 which had the highest activity of acetylcholinesterase (AChE) with a molecular weight of 600,000-800,000, and bands 7, 12, 14, 17 and 18. 2) Inhibition of serum ChE activity was more severe as the amount ingested increased in patients who took Fenitrothion and Malathion. Reactivation of serum ChE activity was very slow in patients treated with PAM (2-pyridine aldoxime methiodide) in the late stage of ingestion or whose symptoms reappeared. 3) There were no differences in the patterns of serum ChE isoenzyme by organophosphorus compound. Band 7 disappeared in the serum ChE isoenzyme of almost every patient, and bands 12, 18, 14 and 17 of the serum ChE isoenzyme disappeared successively with the decline of serum ChE activity. Only band 5 of the isoenzyme remained in cases who had serum ChE activity lower than 5% of normal. 4) All 13 patients were treated with PAM and atropine immediately after being admitted to hospitals. We could not clearly determine the efficacy of PAM on reactivation of serum ChE activity and isoenzyme, because it was impossible in human poisoning to compare PAM efficacy with no treatment and with pre- and post-PAM treatment. 5) The activity and isoenzyme patterns of serum ChE recovered rapidly after combined hemoperfusion and hemodialysis treatment (HP-HD treatment) of the patients poisoned with Malathion. But HP-HD treatment had no effect on poisoning by Fenitrothion and Isoxathion. These findings demonstrated the changes in the activity and isoenzyme pattern of serum ChE in patients poisoned with several organophosphates after PAM and HP-HD treatment.

In Turkey, the mosquito Anopheles sacharovi has been under field selection pressure sequentially with DDT, dieldrin, malathion and pirimiphosmethyl over a period of 30 years for the purpose of malaria control. In 1984, the field population of An.sacharovi in the malarious Cukurova plain of Adana Province contained an altered acetylcholinesterase-based resistance gene giving broad spectrum resistance against organophosphorus and carbamate insecticides. The cross-resistance spectrum from this mechanism conferred resistance to malathion but not to the organophosphorus insecticide pirimiphos-methyl. Over the 6 years that pirimiphos-methyl has been applied for malaria vector control in this area, the frequency of the altered acetylcholinesterase resistance gene has declined, although in 1989 and 1990 it was still present at measurable frequencies in An.sacharovi from Cukurova. In addition to the acetylcholinesterase resistance mechanism there is evidence of an increased level of glutathione S-transferase in some of the An.sacharovi populations tested. This is known to be correlated with DDT resistance in other anophelines. In Turkish An.sacharovi, DDT resistance and elevated glutathione S-transferase occur in the same populations at similar frequencies. The continued prevalence of resistance to DDT and dieldrin, long after the 1971 cessation of DDT spraying for malaria control in Turkey, suggests that the DDT resistance gene has insufficient reduced fitness associated with it to have been lost from the field population during the past two decades. The implications of the slow decline in resistance gene frequencies in this field population are discussed in relation to mathematical models for managing resistance.

A flow injection system, incorporating an acetylcholinesterase (AChE) single bead string reactor (SBSR), for the determination of some organophosphorous (azinphos-ethyl, azinphos-methyl, bromophos-methyl, dichlorovos, fenitrothion, malathion, paraoxon, parathion-ethyl and parathion-methyl) and carbamate insecticides (carbofuran and carbaryl) is presented. The detector is a simple pH electrode with a wall-jet entry. Variations in enzyme activity due to inhibition are measured from pH changes when the substrate (acetylcholine) is injected before and after the passage of the solution containing the insecticide. The percentage inhibition of enzyme activity is correlated to the insecticide concentration. Several parameters influencing the performance of the system are studied and discussed. The detection limits of the insecticides ranged from 0.5 to 275 ppb. The determination of these compounds was conducted in Hepes buffer and a synthetic sea water preparation. The enzyme reactor can be regenerated after inhibition with a dilute solution of 2-PAM and be reused for analysis. The immobilized enzyme did not lose any activity up to 12 weeks when stored at 4 degrees C.

The effect of malathion on jugular plasma concentrations of follicle-stimulating hormone (FSH), estradiol (E2), progesterone (P4) and acetylcholinesterase (AchE) on conception in dairy cattle during a cloprostenol (prostaglandin F2 alpha analogue, PG)-induced estrus was studied. Malathion (1 mg/kg, intraruminally) given at the onset of estrus (48 h after PG) did not alter the plasma FSH or E2 concentrations but significantly (P less than 0.05) inhibited plasma P4 concentration. The mean P4 concentration in the malathion-treated group on days 8 and 12 were 0.8 +/- 0.4 and 1.0 +/- 0.5 ng/ml, as compared to 2.6 +/- 0.0 and 2.4 +/- 0.3 ng/ml in the control group. There was a nonsignificant (P greater than 0.05) inhibition of plasma AchE activity in malathion-treated cattle. Conception was 16.6% in malathion-treated cows and 50% in controls. Inhibition of progesterone secretion and poor conception occurred after the single intraruminal dose of malathion at the onset of estrus.

Use of malathion for mosquito control in Cuba for 7 years up to 1986 has selected for elevated non-specific esterase and altered acetylcholinesterase (AChE) resistance mechanisms in populations of the pest mosquito Culex quinquefasciatus Say. These mechanisms are still present in relatively high frequencies in the Havana area, despite the replacement of malathion by pyrethroid insecticides for the last 3 years in the mosquito control programme. Samples of Culex quinquefasciatus populations from within a 100 km radius of Havana had high levels of resistance to malathion and lower levels of resistance to propoxur, but there was little or no cross-resistance to the organophosphorus insecticide pirimiphos-methyl. Selection with malathion for twenty-two consecutive generations in the laboratory increased the level of malathion resistance to 1208-fold and propoxur level to 1002-fold, but the maximum level of pirimiphos-methyl resistance was only 11-fold. Pirimiphos-methyl is still operationally effective, despite the resistance mechanisms segregating, so this insecticide if used for control is unlikely to select either of the known resistance factors directly in the field population. Since 1986, pyrethroids have been used extensively, and low levels of pyrethroid resistance were detected in two of five field population samples tested. Malathion selection did not increase the level of pyrethroid resistance, which indicates that one or more distinct pyrethroid resistance factors are now being selected in the field populations of Culex quinquefasciatus.

The relative toxicities of ten acaricides to northern fowl mite, Ornithonyssus sylviarum (Canestrini and Fanzago), and the chicken mite, Dermanyssus gallinae (De Geer), were determined simultaneously by holding the mites inside disposable glass Pasteur pipettes previously immersed in acetone solutions of various concentrations (w/v) of technical grade acaricides. The LC90s (parts per million) of the acaricides after 24 h exposure for the northern fowl mite and the chicken mite, respectively, were: bendiocarb (13.1, 0.18), tetrachlorvinphos (14.5, 4.07), carbaryl (15.0, 0.83), pirimiphos methyl (18.3, 2.03), permethrin (23.1, 8.46), lambda cyhalothrin (80.7, 11.4), dichlorvos (252.8, 3.75), malathion (238.4, 6.59), amitraz (6741, 9430) and fenvalerate (greater than 10,000, 60.2). After 48 h exposure there were only slight increases in mortalities of both species except for increased mortalities for the northern fowl mite with lambda cyhalothrin, amitraz and fenvalerate, and for the chicken mite with amitraz.

The protective effects of atropine, diacetylmonoxime (DAM), and diazepam separately and in combination were investigated in rats exposed to malathion. Malathion (500 mg/kg, ip) inhibited acetylcholinesterase (AchE) activity in RBC and brain and produced hyperglycemia and hyperlactacidemia with depletion of glycogen in liver, triceps, and brain of animals 2 hr after its administration. Atropine (20 mg/kg, ip) given immediately after malathion abolished hyperglycemia and glycogenolytic effect but exhibited no effect on the recovery of inhibited AchE activity. DAM (100 mg/kg ip) given immediately after malathion significantly reactivated the inhibited AchE activity both in RBC and brain. It also partially modified hyperglycemia and glycogenolytic effect. Diazepam (50 mg/kg, ip) slightly modified AchE and abolished hyperglycemia, hyperlactacidemia, and glycogenolytic effects. A combination of these drugs protected the animals from the acute toxic effects of malathion.

A simple method for determination of organophosphorus pesticide residues at the parts per million level in milk was developed. Pesticide residues were extracted with acetonitrile added to aqueous milk, fat was removed by zinc acetate addition and dichloromethane partition, and analytes were concentrated and analyzed by wide-bore capillary column gas chromatography. Recoveries of 6 pesticides spiked in milk samples at levels of 0.1 and 1.0 micrograms/mL were 82.1-93.8% and 79.7-96.6%, respectively. Triplicate samples spiked with 6 pesticides at 1 microgram/mL were analyzed independently by 3 laboratories. Average recoveries were greater than 80%, and the mean coefficients of variation for the complete study were 2.9% for diazinon, 5.4% for dimethoate, 4.6% for malathion, 4.6% for parathion, 4.9% for EPN, and 6.1% for phosalone.

Organosphosphorus and carbamate insecticides are quite often used in greenhouses. They represent a group of active principles of toxicological relevance. Initial residues on the surface of cucumber, tomato and ornamental plants, and half-life periods for residue degradation are outlined for carbendazim, dimethoate, fenazox, malathion, methamidophos and pirimiphos-methyl. Residues on plants, concentration in the air, dermal exposition, and inhibition of serum choline esterase activity are shown for methamidophos and aldicarb, respective reentry times being discussed. On harvest and cultivation in greenhouses, dermal exposition as a rule is more relevant than inhalation.

An HPLC method for the simultaneous detection of six organophosphorus pesticides (Dimethoate, Ethion, Malathion, Phorate, Phosalone and Parathion) on a Zorbex ODS column using methanol + water (80:20) as solvent is described.

Responses of Paramecium caudatum, a ciliated protozoan, to acute exposures of certain organic solvents and organophosphorus insecticides (OPI) were studied by determining their lethal concentration (10 min-LC100) and median lethal concentration (4 h-LC50). The solvents and OPI evoked a distinct sequence of responses. Among the five solvents tested, acetone proved most toxic [LC-2.9% and LC50-0.68% (v/v)], while dimethyl sulphoxide (DMSO) showed least toxicity [LC-11.0% and LC50-3.16% (v/v)]. The order of toxicity of solvents was: acetone greater than ethanol greater than methanol greater than N, N-dimethylformamide greater than dimethylsulphoxide. The LC values of six OPI dissolved in either acetone or DMSO indicated that they were more toxic when dissolved in acetone and least toxic in DMSO. Among the OPI, bromophos proved most toxic (LC-10 ppm) while malathion showed least toxicity (LC-200 ppm) in DMSO. The order of toxicity of OPI was: bromophos greater than pirimiphos-methyl greater than parathion methyl greater than dichlorvos greater than fenitrothion greater than malathion. The 4 h-LC50 values computed for bromophos and malathion (dissolved in DMSO) were 575 ppb and 19.9 ppm, respectively, indicating the high susceptibility of P. caudatum to bromophos. The results indicate that the Paramecium toxicity assay could be used as a complementary system to rapidly elucidate the cytotoxic potential of compounds.

In developing countries, extensive use of pesticides to meet with increased agricultural needs is inevitable and the indiscriminate use has led to several toxicological implications in humans. The toxic effects, however, to a large extent depend on the nutritional status of the individuals. In the present work, the hepatic susceptibility to pesticide toxicity has been studied in experimental animals maintained on diets containing different levels of protein for a period of 3 weeks. Along with protein deprivation, the rats were also exposed to three organophosphorus pesticides, viz., parathion, malathion, and phosalone, individually at various doses for the same time period. Phosphatases, the functionally important enzymes of the liver, were estimated and the results obtained indicated that protein deprivation further aggravated the pesticide-induced effects on the hepatic phosphatases.

An enzyme test for determining isomalathion (O,S-dimethyl-S-(1,2-dicarbethoxyethyl) phosphorodithioate) impurities in water-dispersible powders of malathion (WDP malathion) is described. The test is based on inhibition of acetylcholinesterase (EC 3.1.1.7) by isomalathion extracted from WDP malathion. The lower limit of detection of the test is 0.01% (w/w) isomalathion. For 18 samples of WDP malathion there was good correlation between the levels of isomalathion found using the enzyme test and those obtained by thin-layer chromatography.

The purpose of this study was to determine if 4 major organophosphate impurities of malathion were active as alkylators of nitrobenzylpyridine (NBP) or as mutagens in the Salmonella typhimurium bioassay. Malathion, isomalathion, O,O,O-trimethyl phosphorothioate, O,O,S-trimethyl phosphorothioate, and O,S,S-trimethyl phosphorodithioate produced alkylated NBP at varying rates. In order of increasing NBP reactivity, the compounds ranked: O,O,O-trimethyl phosphorothioate = O,O,S-trimethyl phosphorothioate less than O,S,S-trimethyl phosphorodithioate less than isomalathion = malathion. At 37 degrees C, the most reactive compounds produced an NBP alkylation rate equal to approximately 25% of the rate produced by methyl methanesulfonate, a potent Salmonella mutagen. However, none of the organophosphates were mutagenic in S. typhimurium TA97, TA98 and TA100 when tested by the standard plate-incorporation method or by the preincubation modification of the plate-incorporation method. The possible relationships between NBP reactivity and the biological activities of these organophosphates are discussed.

Impurities such as O,S,S-trimethyl phosphorodithioate (TMPD) and the S-methyl isomer of malathion (isomalathion) strongly potentiated the mammalian toxicity of malathion. In contrast, impurities present in the phosphoramidothioate insecticide acephate had an antagonizing effect on its mammalian toxicity. The potentiation of the toxicity of malathion was attributed to inhibition of mammalian liver and serum carboxylesterase. O,O,S-Trimethyl phosphorothioate (TMP), another impurity present in technical malathion and in other organophosphorus insecticides, proved to be highly toxic. Rats given a single oral dose of TMP at a level as low as 20 mg/kg died over a period of three weeks, with death occurring with non-cholinergic signs of poisoning. TMPD also caused similar delayed death in rats. O,O,O-Trimethyl phosphorothioate (TMP=S), also another impurity in technical malathion and a structural isomer of TMP, was a potent antagonist to the delayed toxicity of TMP. Examination of a number of related trialkyl phosphorothioate and dialkyl alkylphosphonothioate esters revealed several of these compounds to be highly toxic to rats.

The emergence of strains of malaria vectors resistant to malathion in an area of Pakistan, and the continuing search for improved methods of control, necessitated the examination of alternative safe insecticides, with improved residual effects, for future use in the Malaria Control Programme in Pakistan. For these reasons, the effectiveness of pirimiphos-methyl, as Actellic 25 WP, was evaluated on a large scale in one sub-sector of Sheikhupura district of Punjab Province near Lahore. Entomological and parasitological evaluations demonstrated that 1 g of pirimiphos-methyl/m2 was as effective as 2 g/m2. Vector mosquito densities were reduced to zero, or almost so, in all areas sprayed with pirimiphos-methyl, and only began to approach vector levels in unsprayed areas after 9-10 months. No new cases of malaria were detected in those areas sprayed with pirimiphos-methyl. Blood cholinesterase determinations after the application of the pirimiphos-methyl spray confirmed the absence of any toxic effect on the spray operators, nor were there any toxic symptoms in the house occupants.

Details of cases involving the inadvertent exposure of birds to eight toxic substances are recorded. The organophosphate insecticides dichlorvos, diazinon and malathion produced respiratory symptoms which in the former and latter cases were initially thought to be caused by infectious disease. Birds which consumed feed containing fenitrothion showed nervous signs before death. On three separate occasions feral starlings (Sternus vulgaris) were found dead and their gizzard contents contained mevinphos. The rodenticide warfarin was associated with petechial haemorrhages in the skeletal muscles and on the serosal surfaces of one hen. Cyanogenic glycosides from Eucalyptus cladocalyx were responsible for the sudden deaths of ducks and guinea fowl. 'Ornamental dough' containing sodium chloride was fed to birds which were deprived of water and they showed diarrhoea and nervous disorders before death.

A single oral dose of DDT (60.0 mg/kg), malathion (687.5 mg/kg), phosalone (60.0 mg/kg) and elsan (175.0 mg/kg) to adult male albino rats produced significant impairment in acid and alkaline phosphatase activities within 1 h of dosing. Renal and hepatic alkaline phosphatase activity was significantly inhibited in all cases, while treatment with malathion and phosalone resulted in an increase in intestinal alkaline phosphatase activity. Elsan induced an increase in acid phosphatase activity in liver, kidney and intestine whereas the response to DDT, malathion and phosalone was variable.

Evidence is given that the organo-phosphates mevinphos and malathion perturb the retinal functioning of the white mouse by acting 1. directly on the photoreceptors; 2. by a synaptolytic effect due to the inhibition of cholinesterases; 3. likely by a damage of the bipolar and/or the ganglion neurones.

Two malathion carboxylesterase fractions, designated as esterase fraction A and esterase fraction B, that hydrolyze malathion were purified 13- and 18-fold, respectively, from rat liver microsomes. The two enzymes could not be distinguished kinetically, but fraction A contained at least one electrophoretic species not present in fraction B. The molecular weight of fraction A was estimated as 50,000-60,000; the molecular weight of fraction B was about twice this value. Incubation of [methoxy-14C]malathion with fraction A or fraction B resulted in a mixture of malathion alpha and beta monoacids, but the composition of the mixture produced by fraction A (alpha/beta = 1.5) differed from that produced by fraction B (alpha/beta = 0.2), indicating the presence of multiple species of carboxylesterases in mammalian liver microsomes. Isomalathion was substantially more potent as an inhibitor of both rat liver and rat serum malathion carboxylesterases than O,S,S-trimethyl phosphorodithioate. Isomalathion appeared to be equipotent in inhibiting the rat liver carboxylesterase-catalyzed reactions leading to either alpha or beta-monoacid. O,S,S-Trimethyl phosphorodithioate, on the other hand, preferentially diminished the reactions leading to alpha monoacid. In contrast, the rat serum carboxylesterase-catalyzed reactions leading to either alpha or beta monoacid were inhibited to approximately on equal degree by isomalathion and O,S,S-trimethyl phosphorodithioate.

Common house mosquito Culex pipiens molestus Forskal was used to test biologically the residual toxicity of 15 organophosphorus, 5 carbamate and 5 pyrethroid insecticide preparations sprayed on whitewashed or limewashed wall surfaces. The doses of 0.1 g and 1.0 g of active ingredient per 1 m2 of wall surface were used in this experiment. At the dose of 1 g/m2, organophosphates chlorpyriphos, diazinon, fenitrothion, malathion, pirimiphos-methyl and propetamphos, cambamates bendiocarb, dioxacarb, propoxur and promecarb, and pyrothroids bioresmethrin, decamethrin-permethrin and tetramethrin produced on whitewashed wall surfaces the residual toxicity persisting for at least four months. At the dose of 0.1 g/m2, a long-lasting residual toxicity persisting on whitewashed wall surfaces for at least two months was observed after bendiocarb, decamethrin, fenitrothion, permethrin, pirimiphos-methyl and propoxur application. The residual toxicity of organophosphates, carbamates except for bendiocarb and pyrethroids except for permethrin sprayed on limewashed wall surfaces was considerably shorter than on whitewashed surface.

During a malaria eradication programme in Pakistan in 1976, out of 7,500 spraymen, 2,800 became poisoned and 5 died. The major determinant of this poisoning has been identified as isomalathion present as an impurity in the malathion. It seems almost certain that the isomalathion was produced during storage of the formulated malathion. The quantitative correlation found between isomalathion content and toxicity of many field samples of malathion has been confirmed by an examination of mixtures of pure compounds. Addition of known amounts of isomalathion to technical malathion indicates that other active substances are present. These impurities have been identified (trimethyl phosphorothioates) and have been shown to behave like isomalathion in potentiating the toxicity of malathion. Some preliminary work on their toxicological properties is reported. The mechanisms involved in the potentiation of the toxicity of malathion are discussed.

A screening method has been developed for determining organophosphorus pesticides at ng/L levels in drinking water. Sixteen organophosphorus pesticides, diazinon, diazinon-oxon, dimethoate, ronnel, beta-phosphamidon, methyl parathion, ethyl parathion, malathion, chlorpyrifos, fenitrothion, ruelene, methidathion, ethion, EPN, phosalone, and phosmet, were extracted by Amberlite XAD-2 resin from 100 and 200 L drinking water previously spiked with these pesticides. The pesticides were eluted from the XAD-2 resin with acetone-hexane (15+85). The concentrated extract was analyzed by gas chromatography using a nitrogen-phosphorus selective detector and by gas chromatography-mass spectrometry using selected ion monitoring. Recoveries at the 10 and 100 ng/L spiking levels were greater than 90%, except recoveries for dimethoate and phosphamidon were 37 and 42%, respectively. The analysis of 300 L Ottawa tap water showed no detectable amounts (less than 1 ng/L) of any of the 16 organophosphorus pesticides.

In 1976, epidemic organophosphate insecticide poisoning due to malathion occurred among 7500 field workers in the Pakistan malaria control programme. In July, the peak month of the epidemic, it is estimated that there were about 2800 cases. In field studies low red-cell cholinesterase activities were associated with the signs and symptoms of organophosphate insecticide intoxication. Toxicity was seen with 3 different formulations of the insecticide and was greatest with the products containing increased amounts of isomalathion, a toxic malathion degradation product. Poor work practices, which had developed when D.D.T. was the primary insecticide for malaria control, resulted in excessive skin contact with and percutaneous absorption of the pesticide. Airborne malathion concentrations were very low. Implementation of good work practices and proscription of use of the 2 pesticide formulations most contaminated with isomalathion halted the epidemic in September. An extensive training programme and surveillance system for pesticide toxicity preceded 1977 spraying operations.

Ten day old chick sympathetic ganglia cultured in a microslide assembly were treated with a selected group of organophosphate pesticides to evaluate their cytotoxicity ranges, and the usefulness of such a model for screening pesticides. Examination by phase contrast and light microscopy for chemically-induced morphological alteration of nerve fibers, glial cells and neurons provided the criteria for quantitation and assessment of the toxic effects. Concentrations that produced half-maximal effects ranged from 1 x 10(-6)M (severely toxic) for methylparathian, diazinon, paraoxon, mevinphos, diisopropylfluorophosphate, tri-o-tolyl phosphate and its mixed isomers to a 1 x 10(-3)M (intermediate) for malathion, leptophos, coumaphos, mono- and dicrotophos. Some or no effects were evident at 1 x 10(2-)M for O'ethyl-O-p-nitrophenyl phenyl phosphonothioate, tri-m-tolylphosphate, chlorpyriphos and triphenyl phosphate. In all instances, nerve fibers were more sensitive than neurons or glial cells to insecticides. All cellular growth was inhibited at 1 x 10(-2)M (except triphenyl phosphate). Below 1 x 10(-7)M, no inhibitory effects were evident. The secondary abnormalities included decreased cellular migration, diffuse cellular growth pattern, increased vacuolization, nerve fiber swelling and cellular degeneration. The cytotoxic effects of these chemicals do not appear to be related to in vivo toxicity or cholinesterase inhibition potential.

Organophosphate poisoning with malathion caused large increases (up to 125 and 440%, respectively) in the level of cyclic GMP in larvae of Mamestra configurata Wlk. and in the fly Sarcophaga bullata Parker. Cyclic AMP was little affected. The malathion-induced increase in cyclic GMP was time and dose dependent. Time-course studies with the head and thorax of S. bullata demonstrated that the increase in cyclic GMP level occurred precipitously after a lag period of about 1 h, during which time the activity of acetylcholinesterase (EC 3.1.1.7) was progressively inhibited. The abrupt increase in cyclic GMP began when acetylcholinesterase activity had been inhibited to a sufficient extent to permit accumulation of acetylcholine. It is suggested that the accumulation of acetylcholine in the malathion-poisoned insects caused cyclic GMP levels to rise. Cyclic GMP may have a role in cholinergic transmission in normally functioning insect neural tissue. Increased levels of cyclic GMP induced by organophosphate and organocholorine (Bodnaryk, R. P. (1976) Can. J. Biochem. 54, 957-962) insecticides appear to be a vital and previously unrecognized biochemical lesion in insects poisoned by these compounds.

Two major classes of enzymes, i.e., hydrolases and transferases, comprise all the nonoxidative enzymes, and together these enzymes catalyze a wide variety of biotransformations of insecticides. The hydrolytic enzymes involved in insecticide metabolism are carboxylesterase (EC 3.1.1.1), arylesterase (EC 3,1.1.2), alkylamidase, and DFPase (EC 3.8.2.1). Recent experimental evidence suggests that carboxylesterase enzymes(s), formerly known to hydrolyze malathion-type insecticides, can also catalyze hydrolysis of a variety of diversified insecticidal esters such as benzilic acid derivatives, carbanilate compounds, and pyrethroids. These organo-phosphate-sensitive esterases, with the exception of the enzyme which hydrolyzes malathion, are all present in microsomes. Similarly, the action of amidases now extends to those insecticidal compounds or their intermediates which contain an aminoformyl (N--CHO) moiety. Arylesterase and DFPase catalyze the P--anhydride bond cleavage of the leaving group, a major hydrolytic pathway for organophosphate insecticides. Transferal enzymes which are presently known to metabolize insecticidal organophosphates are GSH-S-alkyltransferase (EC 2.5.1.12) and GSH-S-aryltransferase (EC 2.5.1.13). These enzymes cleave P--O--R (R = alkyl) or P--O--X (X = aromatic), with subsequent transfer of the R or X group to glutathione. Regarding the other conjugating enzymes, UDP-glucuronyltransferase (EC 2.4.1.17), UDP-glucosyltransferase (EC 2.4.1.35), and arylamine acetyltransferase (EC 2.3.1.5), much work is needed to understand their interactions with insecticidal compounds. There is some evidence that arylsulfotransferase (EC 2.8.2.1) may play a prominent role in the conjugative mechanisms of insects.